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IID EXAM I Immunogenicity—requires foreignness, high MW & chemical complexity; proteins are most immunogenic Antigen—any substance that reacts specifically w/ Ab and/or TcR Protein Ag—polyvalent (multiple epitopes) induces polyclonal response; linear or discontinuous (conformational) Bacterial Ag—outermost component of bacterium induces greatest immune response; can alter polysaccharides to create multiple strains Gram(+)—cell wall peptidoglycan w/ peptide & oligosaccharide epitopes Gram(-)—outer membrane lipopolysaccharide (LPS) w/ oligosaccharide & lipid epitopes Encapsulated—capsular polysaccharides w/ oligosaccharide epitopes Epitope (antigenic determinant)—site on angtigen that interacts w/ Ab or TcR; linear or conformational (discontinuous) Hapten—compound, usually of low MW, that can act as an epitope but not as an immunogen Immunogen—substance that can elicit an immune response 4 Classes of pathogens—bacteria, parasites, fungi (extracellular); bacteria, parasites (intracellular); viruses; parasitic worms Genetic variation—allows some antigens to evade long-term immunity Antigenic drift—point mutations create new epitopes or strains (serotypes); causing epidemics Antigenic shift—recombination creates entirely new strain of disease, e.g. when human & avian strains present in same host; causing pandemics Gene conversion—variation, by e.g. African trypanosomes, that allows parasite to create new antigens by utilizing inactive genes (variable surface glycoproteins) Ab/Ag interaction—noncovalent (electrostatic, H-bonds, van der Waals, hydrophobic interactsion, aromatic stacking) Reversible interaction, but effectively unreversible b/c high ionic strength & pH changes needed to break bonds do not occur naturally Heterogeneous—multiple antibody clones may bind any given epitope Dependence on temperature (little effect), ionic strength (reverses binding at high concentrations), & pH (reverses binding at either high or low pH) Specificity—potential of Ab to bind Ag; high affinity Ab are highly specific, else crossreactive Precipitin reaction—demonstrates potential of Ab/Ag binding w/ precipitation of multi-determinant Ag up to equivalence point Excess Ag prevents crosslinking b/w Ab & Ag that normally forms precipitate, remains in solution & prevents proper clearance by macrophages Ab/Ag testing: Immunodiffusion—assays that detects precipitation of Ab:Ag complexes in gel Radial diffusion—quantitatively determine concentration of Ag in particular well based on diameter of precipitate in Ab gel Double diffusion (Ouchterlony)—qualitative assay to determine presence of Ag based on # of bands emanating from center Ag well towards surrounding Ab wells Immunoelectrophoresis—qualitative assay in which serum components are first separated by electrophoresis followed by double diffusion analysis Radioimmunoassay/enzyme immunoassay (ELISA)—sensitive, quantitative assays that use Ab labeled w/ radioactive or enzymatic marker Ag or purified Ab bound to sold phase substrate → Ab or Ag binds substrate if present in serum wash → secondary labeled-Ab (anti-human Ig) binds Immunohistochemistry—used to identify cells w/in tissue w/ labeled-Ab Western blot—identifies Ag w/in complex mixture following separation by electrophoresis & binding of Ab; confirms ELISA test Flow cytometry—immunofluorescent assay to rapidly quantitate immune cells in blood; laser detects cells & charged deflection plates separate cell into tubes Monitors AIDS patients (stain cells w/ anti-CD4 Ab), immunophenotyping, organ rejection, RBC count Monoclonal Ab—homogeneous Ab produced from hybridoma cells (fusion of B-cell & myeloma tumor cell) to provide single clone of B-cells for above mentioned tests Myeloma tumor cells lack Ab secretion & enzyme HGPRT; hybridoma cells selected for b/c cell dies if not fused w/ B-cell to obtain HGPRT for purine synthesis Ab humanization—Ag-binding site of mouse monoclonal Ab transferred to human Ab to avoid immune response when injected into human host for treatment 1° (central) lymphoid tissue—bone marrow & thymus; site of maturation of lymphocytes Bone marrow—cells migrate towards central sinus as mature; marrow of ribs, sternum, iliac crest, & vertebrae remain cellular throughout life Endothelial & reticular cells lining central sinus are source of GF & cytokines, reticular fibers provide microenvironment & cell contact signals for developing cells Thymus—bilobed structure under sternum, develops from 3rd & 4th pharyngeal pouches; site of T-cell maturation; atrophies almost completely by 30yo Lobules—outer cortex & inner medulla; cells migrate towards medulla as mature & enter venules at corticomedullary junction TcR added → T-cells proliferate → recognize self/non-self → CD4/8 differentiation → mature naïve T cell Cortical epithelial cells—provide support network & release cell contact signals (GF & cytokines) for developing thymocytes Nurse cells—may help eliminate dying thymocytes in cortex 2° (peripheral) lymphoid tissue—adenoids, tonsils, lymph nodes, appendix, spleen, Peyer’s patches; site of initiation of immune response Spleen—respond to Ag w/in blood w/in white pulp & removal of damaged RBCs Circulation—Ag & lymphocytes follow same pathway thru spleen Trabecular artery → central artery → marginal zone sinuses → red pulp sinuses → pulp veins → trabecular veins M line sinusoids to remove dying RBCs or Ag that escaped white pulp & then migrate back to marginal zone Follicle—central arteriole surrounded by PALS (T cells); germinal center (B cells) surrounded by B-cell corona; marginal sinus & marginal zone surrounds all 1° lymphoid follicle—all resting B cells, no Ag stimulation 2° lymphoid follicle—outer resting B cells w/ central germinal center of proliferating B cells Lymph nodes—respond to tissue-derived Ag (antigens that have penetrated skin or mucous membranes) Ag circulation—afferent lymphatic → subcapsular sinus → lymphoid follicle (B-cells w/ germinal center) → paracortical area (T-cells) → medullary cords (macrophages & plasma cells) → medullary sinus → efferent lymphatic vessel Lymphocyte circulation—postcapillary high endothelial venule (HEV) → subcapsular sinus → lymphocytes migrate out of blood at follicle → activated T-cells migrate to infection site thru efferent lymphatic & activated B-cells lodge in lymph node medulla or bone marrow DCs (paracortical) & follicular dendritic cells bind Ag & activate Ag-specific T-cells (paracortical) & B-cells (follicular) Mucosal-associated lymphoid tissue—respond to inhaled & ingested Ag; tonsils & adenoids, Peyer’s patches, appendix, lamina propria (diffuse) Lymph node-like accumulations of lymphocytes in subepithelia of all mucosal tissue; diffuse b/c no fibrous capsule M-cells—transport luminal contents from gut lumen into follicle for sampling for Ag Hematopoietic stem cell (HSC)—pluripotent CD34 cells in bone marrow (1:100,000 cells) or blood (only when mobilized by cytokines) Regulation: Colony stimulating factor (CSF)—drives HSC to differentiation Spatial regulation—HSC must physically interact w/ other cells in bone marrow & thymus to become responsive to cytokines Temporal regulation—stages defined by unique CD expression & responsiveness to certain cytokines Common lymphoid progenitor (bone marrow) → B- & T-cell (blood) → plasma cell & effector T-cell (w/ antigen contact) OR NK cell Myeloid progenitor (bone marrow) → basophil, eosinophil, neutrophil (polymorphonuclear PMN leukocytes, blood) OR dendritic cell (tissue) Erythroid progenitor (bone marrow) → megakaryocyte & erythroblast (bone marrow) → platelets & erythrocyte (blood) 2 1st Innate (natural) immunity—rapidly activated & acting line of defense; non-specific response; constitutive Inducible innate immunity—thru alternative pathway of complement or acute inflammatory response (promoting opsonization) Epithelium—skin & mucosal barriers provide first defense against infection Mechanical—epithelial cells joined by TJs; stratification; mucus secretion & removal Chemical—fatty acids (skin), hydrolytic enzymes that target Gram(+) cell walls, low pH (stomach) Antibacterial peptides (cationic peptides that destroy bacterial membranes), defensins (skin, gut), cryptidines (Paneth cells in base of ileal crypts) Microbiological—normal flora compete for attachment & nutrients & can produce antibacterial substances Monocyte—2-10% of WBC in blood (t1/2 = 1-4 days); peanut-shaped nucleus w/ numerous vesicles; phagocytic, produces cytokines, Ag-presenting, microbicidal Differentiates into macrophages in tissue (t1/2 = 4-12 days), Kupffer cells in liver, microglia in brain Expresses MHCII, FcR (binds effector portion of Ab-Ag complex), C3R (binds complement component C3b), CD14, receptors for many bacterial constituents Binding to effector T-cell stimulates release of cytokines (IL-1, IL-6, IL-8, IL-12, TNF-) that initiate inflammation Dendritic cell—<1% blood & tissue; pseudopods (↑SA to sample environment); Ag-presenting (MHCII), NOT microbicidal & loses phagocytic capacity once presents Ag Interdigitating dendritic cell (DC)—bone marrow-derived; constitutively express MHCII Immature DC—found in interstitium of most organs (e.g. Langerhans cells throughout epidermis); present degraded Ag on MHCII & migrate to LN Mature DC—found in T-cell-rich areas of LN & spleen; NOT phagocytic, high expression of co-stimulators Principal APC for activating naïve T-cells, attract to LN & spleen w/ chemokine DC-CK Follicular DC (FDC)—not bone marrow-derived; found in germinal centers of lymphoid follicles; not constitutively express MCHII Express complement & FcR; NOT phagocytic (“trap” Ag on surface complexed w/ Ab or complement proteins); present immune complexes to B-cells Granulocytes—50-70% bone marrow, 65% blood; multilobed nucleus, unique cytoplasmic granules Neutrophil (polymorphonuclear or PMN)—most abundant WBC; phagocytic, microbicidal; FcR (binds IgG) & complement receptors Normally NOT in tissue, but functions best in anaerobic conditions seen in damaged tissue; unable to reform granules, short-lived, forms pus Azurephilic granules—light color, lysosomes containing myeloperoxidase (H2O2 + Cl- → HOCl or bleach), microbicidals Secondary specific granules—dark color, containing lysozyme (degrade bacterial cell wall), lactoferrin, alkaline phosphatases & defensins Eosinophil—1-6% WBC; eosinophilic (red) granules containing “major basic protein” or crystalline bands that degranulate mast cells & basophils Cytotoxic to worms; phagocytic; synthesized w/ IL-5 stimulation & migrates into tissue w/ chemokine eotaxin Basophil/mast cell—<1% WBC; large purple cytoplasmic granules containing histamine, heparin & proteases; FcR (binds IgE) Mast cells found in ALL CT, especially near epithelial surfaces; allergic reactions & parasitic resistance Natural killer (NK) cells—or large granular lymphocytes (LGL); express CD16 & CD56 & FcR; NOT phagocytic, NOT Ag-specific Fight infection until CD8 cells develop → IL-10 produced by CD8 cells deactivates NK cells Selectively kills virus-infected cells (viral infection → ↑IFN- & - → ↑MHCI) or cells lacking MHCI; tumor cells w/ granzyme; maturation completed in marrow IFN- & IFN- cause ↑cytotoxicity & proliferation of NK cells M release TNF- & IL-12 → NK cells activated to produce IFN- → activates M (positive feedback) Inhibitory receptor—normally binds MHCI on healthy cells to deactivate NK cell; CD94/NKG2 or killer cell Ig-like receptors Antigen-presenting cells (APCs)—DCs (viruses), B-cells (soluble Ag) & M (extracellular peptides); degrade proteins into fragments; must express MHCI/II & costimulator (B7) Ag binding promotes DC migration & differentiation; microbial products enhance presenting function of M & DC; specific Ag enhance presenting of B-cells LPS-binding protein—serum protein that binds bacterial LPS & delivers bacteria to CD14 on M & neutrophils Bactericidals—agents produced or released by phagocytes after ingestion of microorganisms Acids, ROS, NO, antimicrobials (defensins & cationic peptides), lysozyme & acid hydrolases, lactoferrin & vitamin B12-binding protein Pathogen-associated molecular patterns (PAMPs)—molecular structures not shared w/ host; relatively invariant; often polysaccharides or polynucleotides Pattern recognition receptors (PRR)--M & DC receptors that bind PAMPs & promote Ag-presentation, release CKs & induce expression of costimulators Disorders of innate immunity: Neutropenia—congenital or acquired deficiency in phagocytes resulting in no inflammation response (<500 PMN/mm3 at risk for infection, <200 PMN/mm3) Chronic idiopathic neutropenia (300-1500 PMN/mm3)—usually presents in childhood w/ recurrent infections, fever & malaise, also Schwachman syndrome Schwachman syndrome—pancreatic insufficiency, abnormal bone development & FTT Cyclic neutropenia—AD disorder w/ marked cyclic decrease in phagocytes at 15-35 day intervals; treated w/ G-CSF Depressed BM production—idiopathic, drug induced (cyclophosphamide, phenothiazides, mercurial diuretics), tumor invasion Peripheral destruction—Ab against neutrophils (e.g. RA or SLE), splenic trapping Felty’s syndrome—RA w/ splenomegaly Splenectomy—treatment for neutropenia to reduce rate of clearance of opsonized WBCs Chronic granulomatous disease—inherited defect in NADPH oxidase, renders phagocytes unable to generate ROS; ↓Ag degradation leading to chronic T-cell stimulation Treatment—IFN- therapy, prophylactic antibiotic & granulocyte transfusions for infections, gene therapy Diagnosis—nitroblue tetrazolium + O2 → formazan + O2 (blue/black precipitate); recurrent infections & enlarging granulomas (obstruct GU or GI tracts) G6PD deficiency—defect of first enzyme in HMPS that results in deficiency of NADPH → nonsphereocytic anemia Hereditary myeloperoxidase (MPO) deficiency—most common (1:2000) inherited (AR) disorder of phagocytes MPO—component of primary (azurophilic) neutrophil granules that catalyze H2O2 + HCl → HOCl/OCl (halogenates bacterial proteins) Adaptive (acquired) immunity—slower, specific, diverse, accentuates effects of innate response, memory, self-limited to prevent collateral damage Clonal selection—pool of mature naïve lymphocytes exposed to antigen → proliferation & differentiation to form clone of effector cells Clonal deletion—removal of self-reactive immature lymphocytes after recognizing self-Ag in thymus (T cell) or bone marrow (B cell) to establish tolerance Anergy—no response to subsequent Ag exposure b/c immature B-cell binds soluble self-Ag; IgM no longer expressed, die in T-cell area of 1° follicle OR naïve T-cell binds MHC:Ag w/o costimulatory B7 signal → T-cell unable to produce IL-2 Lymphocytes—20-45% WBC (75% T cells, 20% B cells, 5% NK cells); little cytoplasm, most resting & naïve Humoral immunity—most effective against extracellular Ag; B-cells express BcR, Ig, & MHCII; maturation completed in bone marrow Recognize ALL chemical classes of Ag w/ or w/o MHC, usually entire intact Ag w/ surface epitopes, conformational & hydrophilic 2 biochemical signals required for proliferation—BcR + costimulator (CD4 cell interaction & CK release) Memory—repeated exposure → ↑# of Ag-specific B-cells, ↑affinity for Ag & ↑MHC expression Iccosomes—Ag that remains bound to follicular DCs as immune complexes, bud off to present to & maintain memory of B- & T-cells Original antigenic sin—memory cells suppress activation of naïve B- & T-cells on subsequent exposure, only new epitopes stimulate naïve response Antibody—2 (divalent) variable regions (Ag-binding sites) + constant region (effector function); composed of 2 heavy (VH + CH1-3 domains) & 2 light chains (VL + CL domains) Hinge—b/w variable & constant regions allows flexibility of Ag-binding arms to move to reach epitopes; covalently linked w/ disulfide bonds Papain—protease that cleaves (unhinges) Ig into 2 fragment Ag-binding (Fab) & separate effector fragment crystallizable (Fc) Pepsin—cleaves Fc, but leaves Ag-binding arms coupled Ab repertoire—>1 billion; over 3 million combinations of heavy & light chains w/ additional junctional diversity & somatic hypermutation 3 Somatic recombination: Germline DNA—unrearranged Ig gene found in germ cells & all non-B-cells (vs. somatic DNA in each cloned B-cell); ALL B cells begin producing IgM Heavy chain locus—encodes ALL heavy chain types on chromosome 14 w/ “diversity segment” (D) & “joining segment” (J) between V & 3C domains Light chain loci-- (chromosome 22) & (chromosome 2) w/ J segment b/w V & C domains; no functional difference b/w chains, but human have 2/3 chains “Leader” peptide (L)—membrane secretion signal Recombination signal sequence (RSS)—flank V,D, & J gene segments; cleavage sites that “pull” segments together during recombination Recombination activating gene (RAG-1 & RAG-2)—expressed ONLY at early stages of B- & T-cell development, shut down when functional protein made Junctional diversity—variations in sequence at V(D)J junctions due to varied length of repair joint in HV region Non-productive rearrangement—most (2/3) rearrangements disrupt triplet codon reading frame of mRNA, no functional protein made & die in marrow N-region addition—“non-templated” addition of nucleotides at junctions by Tdt to create novel triplet codons for new AA P-nucleotides—palindromic sequences added to ends of DNA cleavage at RSS Allelic exclusion—only 1 heavy chain & 1 light chain permitted to complete rearrangement, ensuring that each clone makes only 1 type Hypervariable (HV) or complementary determining region (CDR)—loops that form Ag-binding sites, 3 HV regions on both light & heavy chains Affinity—strength of sum total of all noncovalent interactions b/w single Ag-binding site & single epitope Equilibrium dialysis—used to determine affinity of Ab by binding of free Ag w/ known amount of Ab, also Scatchard analysis Avidity—strength of binding entire Ab to multi-determinant antigen; always greater than affinity; less likely to diffuse away from each other w/ multiple bonds B-cell development—proceeds thru stages defined by somatic recombination & expression of Ig genes Ag-independent stage—generation of B-cells in bone marrow & elimination of self-reactive B-cells; dependent on marrow stromal cells for cell contact & GFs Double VDJ rearrangement in heavy chain locus → single VJ rearrangement in light chain loci ( first & if rearrangments fail, cell dies if both fail) Stem cell binds stromal cell via cell adhesion molecules (CAMs) & VLA4 (dimer of CD49d/CD29) on stem cell to VCAM1 (CD106) on stromal cell → early pro-B cell binds via Kit (SCF receptor, CD117) to stem cell factor (SCF) → stromal cell releases IL-7 to bind IL-7R Immature B-cell expresses IgM on surface & functional protein product on B-cell surface signal termination of gene rearrangment Mature naïve B cell expresses IgD & IgM from alternatively spliced heavy chain mRNA, signals B-cell to leave marrow & migrate to periphery Ag-dependent stage—activation of B cells by Ag in 2° lymphoid tissue & differentiation to plasma & memory cells Somatic hypermutation—point mutations in V-region of B-cells ONLY that lead to increased affinity of Ab for Ag, preferential selection (affinity maturation) Secretion—switch from membrane-bound to secreted Ig thru alternative RNA processing of heavy chain C-terminus; loss of hydrophobic anchor Isotype (class) switching—regulated by CD4s; deletion (looping out) of DNA in constant region w/ interaction of switch regions preceeding C regions Differentiation—plasma cells lose ability to grow, somatic hypermutate, isotype switch, or express surface Ig & MHCII B-1 cells—alternate pathway of development that occurs embryologically in pleural & peritoneal cavities; self-renewing cells (thru IL-10) that express CD5 Secrete mainly IgM; low somatic hypermutation (polyspecificity leads to binding of many different Ag) Common source of chronic lymphocytic leukemia (CLL); protect body cavity during initial innate immune response B-cell activation—mature naïve B-cells enter lymph nodes via HEV & exit via efferent lymphatics; contact follicular DC in light zone then migrate to outer light zone to contact CD4 cells Naïve B-cells encounter CD4 cells trapped in T-cell zone of 2° lymphoid tissue to form germinal centers (t1/2 = 3-8 weeks vs. 3 days if not form follicle) Differentiate into memory & plasma cells (remain in LN medullary cords, red pulp or migrate to marrow or lamina propria); no longer express surface Ig or MHC BcR—membrane Ig associated w/ 2 other signaling proteins Ig (CD79a) & Ig (CD79b); activates TFs leading to cell proliferation & differentiation & secretion of Ig Crosslinking of BcR w/ Ag activates Src kinases (Lyn, Blk, Fyn) → phosphorylates ITAM (associated w/ Ig & Ig on cytoplasmic domain) → Syk binds phosphorylated Ig → Syk activates PI3 kinase → PI3 kinase recruits Btk to cell membrane → Btk phosphorylated by Syk → signaling cascade Bruton’s disease (X-linked agammaglobulinemia)—mutation in Bruton’s tyrosine kinase (Btk) resulting in no circulating Ab B-cell co-receptor—pulls 2 receptors together; activation amplified when Ag also contains bound fragments of complement proteins, still requires additional CD4 cell signals CR2 (CD21)—receptor for iC3b, C3dg, C3d complement proteins CD19—signaling molecule w/ ITAM, phosphorylated by Ig-bound tyrosine kinase Lyn CD81—“tetraspan” family (spans membrane 4X), promotes interactions w/ integrins Neutralization—Ab covers up sites on Ag that are necessary for growth or replication Opsonization—coating of pathogen w/ Ab to promote phagocytosis & destruction by complement or M w/ specific FcR Ab-dependent cell-mediated cytotoxicity (ADCC)—preformed Ab binds Ag on target cells → FcR (CD16) on NK binds Ab → FcR crosslinking activates NK → apoptosis IgG acts through FcRIII receptor (CD16) & IgA acts through FcRI receptor (CD89) T-cell independent (TI Ag) B-cell Ag—polysaccharides, LPS & proteoglycans from bacterial cell walls & capsules TI-1 antigen—bacterial LPS (B-cell mitogen on Gram- bacteria) in low concentration, activates B-cells only in combination w/ BCR but w/o CD4 cell assistance Higher concentrations of LPS causes polyclonal B-cell activation for severe toxicity Mimic T-cell dependent Ab response—Ag directly stimulate CK production causing isotype switching, e.g. pneumococcal capsular polysaccharides Macrophages in marginal zone of spleen follicles may “trap” polysaccharides for long-lived Ab response w/o true memory TI-2 antigen—high density repetitive carbohydrate or protein epitopes on surface of microorganisms Activate only mature B1-cells (induces IgM & IgG w/o isotype switching, hypermutation or memory); causes strong crosslinking of BcR & co-receptors T-cell dependent (TD Ag) B-cell Ag—proteins Ag-binding to BcR delivers first signal; TH2 cell delivers 2nd signal via CD40L (CD154) & CKs → B-cell isotype switching, affinity maturation, memory generation CD27 binds CD70 to cause differentiation of B-cells into plasma cells CD134L (OX40L) binds CD134 (OX40) to stimulates Ig production Hyper-IgM syndrome—congenital deficiency of CD40L, characterized by loss of isotype switching, lack of germinal centers, affinity maturation & memory cell generation B-cell early response—entry of resting cells into G1 phase of cell cycle; ↑expression of MHCII & co-stimulators; ↑expression of CK receptors Ab isotypes—variants of Ab w/ different heavy chains; separated into 5 classes by function & localization (IgA, IgD, IgE, IgG, IgM); varied # constant domains & amount of glycosylation Allotypes—different alleles of same constant gene in different individuals due to genetic polymorphisms Idiotype—different arrangements of VH & VL gene to form B cell clones IgA—most synthesized daily but lost in secretions; transports across epithelium; 2 subclasses (monomeric from plasma cells & dimeric from mucosal 2° lymphoid tissue) IgE—crosslinking on mast cell, basophil & eosinophil surface leads to rapid release of granules; allergies when activated against harmless Ag; always bound to FcR IgG—most abundant in blood; opsonizes; transports across placenta & into extravascular spaces; 4 subclasses #ed by relative abundance Brambell receptor (FcRB)—receptor that carries IgG from blood vessel lumen across endothelium into extracellular space Transient low IgG levels from birth to 1yr as synthesized IgG catching up after drop in passively transferred maternal IgG IgM—pentamer activates complement; 10 binding sites means strongly binds repetitive epitopes but large size means can’t leave blood Polymers—IgM & IgA form polymers w/ greater avidity for multivalent antigens; may immobilize bacteria by wrapping around flagellae J chain—junction at which IgM forms pentamers & IgA forms dimers, trimers & tetramers 4 Cell-mediated immunity (CMI)—thru T-cells acting directly on infected cells T-cells—all express T-cell receptor (TcR), CD3, & coreceptors CD4/8; maturation begins in bone marrow & completed in thymus Recognize only peptide Ag fragments on MHC of APC w/ epitopes internal, linear & hydrophobic Helper T-cells—express CD4, most abundant lymphocyte in blood that differentiates twice into immature effector T cell (TH0) TH1—activates macrophages thru IFN-, T-cell proliferation thru IL-2, isotype switching & opsonizing thru TNF-; travel to site of infection TH2—activates B cells to produce Ab thru IL-4, IL-5 & IL-6, inhibit TH1 cells thru IL-10 & TNF-; remain in 2° lymphoid tissue (humoral response) Cytotoxic T-cells—express CD8; release granzyme to induce apoptosis 2 biochemical signals required for proliferation—TcR (binds to Ag:MHCII) + costimulator (CD28 on T-cell w/ B7 on APC) Tolerance—discrimination of self-Ag during different stages of development; Ag-specific May result from persistent Ag exposure, Ag recognition w/o inflammation, APC binding to soluble non-bacterial Ag w/o costimulator signal (anergy) Central tolerance—selection of immature (double positive) T-cells in thymus OR binding of multivalent Ag to newly formed IgM causing apoptosis in marrow Positive selection—double positive T-cells capable of binding self-MHCI/II isoforms (able to respond to Ag presentation) signaled to mature (1-2% of T-cells) Occurs in cortex of thymus w/in 3-4 days of formation; signals end of -chain rearrangement Negative selection—deletion by DC & M of T-cells that bind too strongly to self-MHC Bare lymphocyte syndrome—lack of CD4/8 cells due to defect in TAP protein that prevents peptides from entering ER & joining MHCI/II Peripheral tolerance—absence of costimulator signal, abundance of self-Ag, rarity of self-Ag causing ignorance, or regulatory T-cells suppressing self-reactive mature T-cells Binding to self Ag arrests B-cell migration; downregulation of surface IgM in periphery increases tolerance; CD4 tolerance may induce B-cell tolerance T-cell development—T-cell receptor (TcR) always membrane-bound; two mutually exclusive isotypes ( & ); multiple TcR bind multiple MHC:Ag complexes T-cells—only 1-5% of T-cells, protect body surface (epithelium) during initial innate response TcR complex—CD3 ( chains & chains) required for cell surface expression & signal transduction & chain for signal transduction TcR gene recombination— & genes (similar to Ig light chains), & genes (similar to Ig heavy chains) 1. Germline gene configuration (immature CD4-8- thymocyte) 2. DJ rearrangement of -chain (CD25+CD44 thymocyte) 3. V-DJ rearrangement of -chain (CD25+CD44 thymocyte w/ expression of intracellular β chain) 4. Surface expression of β → β rearrangement stops → cell proliferates → CD4/8 induction → transcription (CD4+8+ “double positive”) 5. VJ rearrangement of -chain → surface expression of ::CD3 (CD4+8+ : thymocyte) Requires RAG-1 & RAG-2, allelic exclusion, productive & non-productive rearrangements, junctional diversity, & N-region addition Fewer V segments than Ig but greater junctional diversity, NO somatic mutations, NO isotype switching Defective V(D)J recombination: Severe combined immunodeficiency (SCID)—AR, mutations in RAG1 or RAG2 resulting in lack of both B- & T-cells Omenn syndrome (OM)—AR, mutations in RAG1 or RAG2 w/ partial function resulting in lack of B cells & oligoclonal T cells Characterized by T-cell infiltration of skin, gut, liver & spleen → erythroderma, diarrhea, FTT, hepatosplenomegaly RS-SCID—SCID w/ associated cellular radiosensitivity; AR, mutation in Artemis (DNA repair protein) resulting in lack of both B- & T-cells & defective DNA repair T-cell activation: Cell adhesion molecules—non-specific receptors that aid naïve T-cell transport into 2° lymphoid tissue, contact w/ APC, & interaction of effector T-cell w/ target cell Leukocyte adhesion deficiency (LAD) type I—AR defect in CD18; characterized by recurrent or progressive infection, poor wound healing, NO pus formation LAD type II—deficient sialyl-Lewis X (glycosylation) on neutrophils causing defective binding to endothelial E- & P-selectins Migration—T-cell enters HEV → L-selectin binds vascular addressins on HEV (GlyCAM-11 & CD34) → T-cell rolls → integrin LFA-1 activated by chemokines from ECM → LFA-1 binds tightly to intercellular adhesion molecule (ICAM-1) → T-cell migrates across endothelium into lymph tissue Recognition—Ag recognition & binding of TcR → activates LFA-1 causing conformational change → ↑affinity of LFA-1:ICAM-1 → prolonged cell contact Proliferation—2nd signal from co-stimulator (B7) on APC binding to CD28 on naïve T-cell (low-affinity IL-2R) → express IL-2R (high affinity IL-2R) → secrete IL-2 (thru NFAT activation) → IL-2 binds self IL-2R → signals T-cell proliferation Cyclosporin—inhibits IL-2 production & prevents transplant rejection by preventing T-cell proliferation Activation—binding of MHC & TcR (receptor clustering) → phosphorylation of ITAMs on CD3 tail by receptor-associated kinases → CD4/8 coreceptor binding activates Lck → phosphorylates ZAP-70 → ZAP-70 binds phosphorylated chain of TcR complex → activates PLC- → cleaves PIP2 into DAG & IP3 → activates PKC (by DAG & Ca2+) & calcineurin → activates NFB (by PKC) & NFAT (by calcineurin) → proliferation & differentiation CD4 T-cell—IL-2 signals prolifeation → immature effector T-cell (TH0) → TH1 (w/ IL-12) or TH2 (w/ IL-4) CD8 T-cell—activated thru DC expressing high levels of B7 → T-cells to make IL-2 OR CD4 releases CKs that activate APC to express B7 → stimulates naïve CD8 T-cell OR simultaneous activation such that APC activates CD4 & CD8 cell to express IL-2R → IL-2 secreted by activated CD4 binds CD8 cell T-cell effector function—no longer needs costimulator signal; CD8 cell moves on after dumping contents (~5min) TH2 cells activate B-cells thru CD40L:CD40 interaction → IL-4 causes proliferation & clonal expansion → IL-5 & IL-6 drives differentiation into plasma cells TH2 cells inhibit TH1 cells & M thru TGF-, IL-4, IL-10, IL-13 TH1 & CD8 T-cells migrate to site of infection via activated endothelium (VCAM expressed w/ CK stimulation from infection site) → binds integrin VLA-4 on lymphocyte (VLA-4 switch from LFA-1 prevents from exiting blood at HEV) → TH1 cells activated by Ag:MHCII → bind M thru CD40:CD40L → activate M w/ IFN- (↑MHCII, B7, CD40, IL-1/6, TNF-, oxidative burst, lysosomes) TH1 cells secrete IL-3 & GM-CSF → production of M & neutrophils in marrow → TNF- activates vascular CAMs → MCP directs M to site of infection IFN---inhibits viral replication & activate M to remove dying cells, synergizes w/ TNF- to further activate M CD8+ binds Ag:MHCI → redistributes cytoskeleton (MTOC) & cytoplasmic components → release of lytic granules at site of contact → perforin creates pore on surface of target → granzymes enter target cell thru pore → activates nucleases Fas ligand on CD8 cell also binds Fas on target cell to induce apoptosis NK cells—activating receptor binds ligand on healthy cell, but inhibitory receptor binds MHCI to prevent killing Virus-infected cells lose expression of MHCI (mechanism to evade CD8 T-cells) or anti-MHCI Ab covers MHCI → no longer inhibition of NK cells MHC—membrane-associated molecules that bind & transport peptides to cell surface for recognition by T-cells (antigen presentation); present self-peptides in absence of infection Polymorphisms—multiple alleles of each MHC gene w/in population forming various allotypes MHC complex or human leukocyte Ag (HLA)—cluster of closely linked genes on chr6; 2 classes w/ similar structure; each bind different length peptides via “anchor residues” Anchor residues—residues in MHC-binding peptides that interact w/ pockets of MHC-binding groove forming peptide-binding motif MHCI (HLA-A, -B, or -C)—transmembrane heavy chain & small protein 2-microglobulin (chr15); present intracellular (endogenous) Ag (8-10AA length) to CD8 cells Expressed on ALL nucleated cells (NOT RBCs → susceptible to persistent malarial infection) Proteins degraded by proteasomes in cytoplasm → peptides transported into ER by transporter associated w/ Ag processing (TAP) (requires ATP) 5 → chaperones aid folding of MHC in ER → MHCI- chain binds calnexin → 2-microglobulin binds MHCI- → releases from calnexin → binds chaperonin complex (calreticulin & tapasin) → tapasin binds TAP-1 & positions MHC w/ peptide entering ER → MHC completes folding around peptide → releases from TAP1 → transports to cell surface via vesicle Viruses often interfere w/ transport of viral peptides into ER by inhibiting TAP or binding MHCI to prevent from leaving ER for cell surface MHCII (HLA-DR, -DQ, -DP)—2 transmembrane domains ( & ); expressed on APCs (B cells, M, DCs); present processed extracellular Ag (13-25AA) to CD4 cells Ag phaogytosed into vesicles → acidification of vesicles by proton pumps → vesicles fuse w/ lysosomes to form phagolysosomes → acid activates proteases & hydrolases → vesicles w/ peptides fuse w/ MHCII-containing vesicles → bound peptide transported to cell surface Invariant chain—identical (chr5) in all humans; prevents binding of peptides to MHCII in ER & directs to peptide-containing vesicles (separates MHCI/II peptides) CLIP—prevents binding of peptides to MHCII in vesicles, but HLA-DM facilitates release of CLIP & peptides bind MHCII Mycobacterium—some, e.g. leprosy & TB, exploit vesicular system to avoid attack by CD8 cells, preventing fusion of phagosome w/ lysosome MHC restriction—T-cell receptor must recognize both specific Ag & MHC molecule to bind to APC CDR3 loops of TcR & chains form central part of binding site to peptide; CDR1 & CDR2 loops form periphery of binding site to helices of MHC HLA typing—serological or molecular DNA techniques used to identify allelic forms of individual’s HLA (haplotype) Public specificity—determinant found on MHC molecules from several alleles (vs. private specificity unique to 1 allele) Microcytotoxicity test—use of anti-HLA Ab to bind particular HLA molecules, then lyse cells w/ complement PCR—amplify cellular DNA sequences unique for different HLA alleles, then analyze by electrophoresis to identify HLA alleles by distinct banding Interallelic conversion (segmental exchange)—genetic recombination b/w 2 alleles of locus to generate new MHCI/II alleles Gene conversion—recombination b/w 2 homologous genes on different MHC genes Mucosal immune system—distinct lymphoid tissue w/ preferential “homing” of activated lymphocytes towards mucous membranes; mainly secrete IgA Induction sites—Peyer’s patches (w/in subepithelial dome) & mesenteric lymph nodes (MLN); afferent lymphatics drain Peyer’s patches & villus lamina propria into MLN Effector sites—scattered lymphoid cells throughout lamina propria & epithelium of mucosa Follicle-associated epithelium (FAE)—cover Peyer’s patches; contains columnar epithelial cells, microfold (M) cells, APCs Lymphocyte activation—Ag passively transported into dome region by M cells to local interdigitating DCs, B/T cells & M Intraepithelial lymphocytes (IEL)—mainly -T-cells that suppress -T-cells & antagonize TH1 cells; also lamina propria lymphocytes (LPL) B-cells—divide in germinal center, isotype switch to IgA, affinity maturation, differentiate into memory & plasma cells; upregulate mucosal homing receptors Plasma cells secrete dimeric IgA, transported across epithelial cells into external secretions T-cells—divide in parafollicular region, differentiate into memory & effector cells; upregulate mucosal homing receptor Mucosal homing receptor (47 integrin)—along w/ CK receptor CCR9, binds endothelial cell addressin MAdCAM-1 Directs CD4+ cells out of efferent lymphatics or HEV into mucosa or peripheral blood IgA—transcytosed across epithelial cells via poly-Ig receptor into lumen along w/ secretory component of poly-Ig receptor Polymeric Ig receptor—can transport both dimeric IgA & pentameric IgM in humans; IgM can participate in mucosal immunity & compensate for IgA-deficiency Passive protection—neutralization (also IgG & IgE); forms immune complexes for clearance of Ag; prevents adherence of pathogens to epithelial cells & teeth Passive transfer of immunity to newborns; Does NOT enhance inflammatory response; NOT activate classical complement pathway; poor opsonin Mucosal tolerance—lack of systemic immune response against mucosal Ag (e.g. dietary Ag or resident microflora); local immune homeostasis Food proteins & commensal bacteria taken up by DC → maturation of DC (w/ PGE2 from M & mesenchymal cells; TGF-, IL-10 from epithelial cells) → Ag presented to naïve CD4+ cells in MLN or Peyer’s patch → T-cells differentiate → local IgA production (thru TGF- from TH2 cells) Pathogen receptors: Toll-like receptors (TLR)—family that recognize microbial components (LPS, techoic acid, bacterial DNA, viral RNA & proteins); expressed on APCs & intestinal epithelia TLR1, 2, 6 (Gram+ bacteria), TLR3 (dsRNA), TLR4 (LPS, Gram- bacteria), TLR5 (flagellin), TLR7 (imidazoquinoline), TLR9 (CpG DNA) LPS bound by LPS-binding protein → complex binds CD14 on M surface → interacts w/ TLR-4 to activate NFB → TNF- & IL-12 synthesis → activates NK cells to release IFN- → activates M to phagocytose bacteria & local IgA production (thru TGF-, IL-4 & IL-5) Acute phase proteins—soluble plasma proteins secreted from liver that selectively bind pathogen C-reactive protein—pentroxin protein that binds phosphorylcholine on bacteria & fungi; opsonizes thru C1q Mannose-binding protein (MBP)—binds multiple sites on bacteria & activates MASP proteolytic enzyme complex to cleave C4 & C2 initiating complement, Also opsonizes; similar to pulmonary surfactants A & D, & LPS-binding protein Interferons (IFN- & IFN-)—induce formation of oligoadenylate synthetase → polymerizes ATP into oligomers → activates endonucleases to degrade viral DNA P1 kinase → phosphorylates synthesis initiation factor (eIF-2) → inhibits translation of viral proteins Pattern recognition molecules—recognize patterns of repeating structural motifs Collectins (MBL, C1q)—bind pathogen & initiates C3b Mannose binding protein (lectin)—binds mannose on pathogen surface & initiates phagocytosis Scavengar receptors—recognize anionic polymers & acetylated LDLs Complement system—>30 blood proteins found in ALL vertebrates that act rapidly during infection; results in C3b coating pathogen surface Promote inflammatory response (↑vascular permeability), enhance phagocytosis thru opsonization, directly kill microorganisms Classical pathway—late (adaptive); thru Ag-Ab complexes activated whenever IgM or IgG binds Ag; coats pathogen w/ C3b for opsonization or directly lyses Recognition phase—minimum of 2Ab bind Ag (IgM most efficient b/c only need to bind one Ag, or 2IgG) → C1 complex (C1q, C1r, C1s) binds 2Ab Fc region → C1r proteolytic zymogen cleaves other C1r (conformational change) → C1r then cleaves C1s Activation phase—C1s cleaves C4 & C2 → C4b covalently binds membrane thru newly exposed thioester bond to OH or NH2 → C2b binds C4b to form activated enzyme C4b2b (C3 convertase) → C3 convertase cleaves C3 → C3b mostly inactivated by water, or binds membrane next to C4b2b → C4b2b3b loses ability to cleave C3 (becomes C5 convertase) Lytic phase—C5 convertase cleaves C5 → C5b randomly non-covalently binds membrane → C6 & C7 associate w/ C5b non-covalently → C8 binds C5-7 scaffold & membrane begins to leak → 10-16 C9 molecules binds to form membrane attack complex (MAC) → pore allows imbalance of intracellular ions (Na+ in & K+ out) → intracellular water accumulation → cell lysis Alternative pathway—in absence of infection, continuously synthesizes Ab-independent “sensing system” of C3b looking for membrane w/ which to bind C3 spontaneously hydrolyzed by H2O to C3(H2O) → binds Factor B to form C3B → Factor D binds C3B & cleaves to release C3Ba → C3Bb (C3 convertase) cleaves C3 → C3b inactivated by H2O or binds membrane → C3b binds factor B w/ infection → bound factor B is cleaved by protease factor D → Ba is released → C3bBb complex (C3 convertase) cleaves C3 → Factor P (properdin) stabilizes C3 convertase against Factor H → C3 convertase cleaves C3 until another C3b binds C3bBb (becomes C5 convertase) → enters classical pathway to assemble MAC C3b molecules cause positive amplification & restart alternative pathway Lectin pathway—rapid (innate); thru mannose-binding protein (lectin, MASP-1 & 2 or C1r & C1s) binding to mannose residues on bacteria, normally buried in humans Opsonization—complement C3b binds to surface; C3bR & FcR on surface of phagocytes bind Ab & C3b on bacterial surface to signal internalization & destruction 6 Clearance—C3bR on RBC carry immune complex to spleen & liver for removal of immune complexes & recirculation of RBC CR2—B-cell coreceptor & CR1 on follicular DCs binds iC3b, C3d, or C3dg (components of C3b); Epstein-Barr virus uses CR2 to attach & infect host cells Anaphylatoxins—C3a, C4a, C5a; cause vascular leakage via direct interaction w/ endothelial cells or indirectly by activating mast cells Histamine released quickly, leukotrienes released slowly as made → SM contraction, degranulation, ↑blood flow & permeability, ↑CR1 & CR3 expression Chemotaxin—C5a gradient from site of infection causes direct migration of neutrophils & M to infected tissue Regulation—regulatory proteins focus damage & prevent damage of own cell membranes; normally complement proteins inactive w/ short half-life C1INH (C1 inhibitor)—dissociates C1s & C1r & blocks enzymatic site to prevent uncontrolled C3b depletion C4BP—binds C4b & displaces C2b → allowing Factor I to inactivate C4b by enzymatic cleavage (dissociate C3 convertase) Factor H—binds C3b & allows Factor I to inactivate C3b (dissociate C5 convertase) “Protective proteins”—membrane-bound proteins that prevent accidental damage to self tissue by complement activation Decay-accelerating factor (DAF)—binds C3b & C4b, dissociating C3 convertase & preventing assembly of new C3 convertase Membrane cofactor protein (MCP)—binds C3b & C4b, making them susceptible to proteolytic cleavage by Factor I C59 (protectin)—inhibits assembly of membrane attack complex by preventing C9 from binding, especially on RBCs Primary infection—elicits temporally related immune response involving coordinated effort of both innate & adaptive systems Innate immunity (0-4hrs)—recognition by pre-formed, non-specific effectors (DCs, M, complement proteins) Early induced response (4-96hrs)—recruitment of effector cells (neutrophils, M), recognition & activation Late adaptive response (4-5days)—transport of Ag to lymphoid organs, recognition by naïve B & T cells, clonal expansion & differentiation of effector cells Secondary infections—elicity more rapid adaptive immune response w/ less lag, greater response & slower decline Cell injury—biochemical changes start immediately & morphologic changes occur as a result Adaptation—cell changes to cope w/ excess stress, escapes injury Injury—reversible (cell reverts back to normal state if injurious agent removed) or irreversible (persistent or severe injury, death is inevitable); dependent on length & dose Death—end stage of irreversible cell injury, either by necrosis or apoptosis Necrosis—no apoptotic bodies & elicit inflammatory response Cytoplasmic changes—eosinophilia (deep pink staining cytoplasm w/ H&E binding to denatured cytoplasmic proteins) Hyalinization (glossy & homogenous vs. granular cytoplasm due to glycogen loss); vacuolization; calcification Nuclear changes—2 days after necrosis onset, nucleus simply disappears Karyolysis—pale blue nucleus w/ H&E due to DNA degradation by DNAases Pyknosis—nucleus shrinks & stains dark blue due to DNA condensation Karyorrhexis—fragmentation of nucleus Coagulative necrosis—cell outline still recognizable (not autolyze); denaturation of cellular proteins; characteristic of hypoxia/ischemic death Liquefaction necrosis—cell “liquefy” due to lysis & digestion from own lysosomes (autolysis) or WBC (heterolysis); characteristic of focal bacterial infections Caseous necrosis—tissue appears “cheesy,” central coagulated cells surrounded by granulomatous inflammation; characteristic of TB Fat necrosis—fat cells undergo liquefaction & coagulation due to lipases from necrotic pancreatic cells; characteristic of acute pancreatitis Apoptosis—programmed cell death, e.g. embryogenesis & development, or hormonal dependent involution of adult cells (menstruation) Affects single or few cells; chromatin condenses & aggregates peripherally against nuclear membrane; not elicit inflammatory response Apoptotic bodies—cytoplasmic blebbing then ingested by macrophages or parenchymal cells Calcium homeostasis—injury interferes w/ Ca2+ ATPases → influx of Ca2+ from organelles & extracellular → ↑cytosolic Ca2+ → activation of enzymes → cell damage Phospholipases (destroy membrane phospholipids), proteases (destroy proteins of membrane & cytoskeleton), ATPases (deplete ATP) Ischemia/hypoxia—lack of oxygen → ↓ATP production → loss of membrane integrity Reversible injury—↓oxidative phosphorylation → ↓ATP → ↑anaerobic glycolysis (↑lactic acid → ↓pH) & failure of Na + pumps (Na+ & H2O influx) ↑cell & organelle size, clumping of chromatin (due to ↓pH), detachment of ribosomes from ER (due to ↓ATP); cytoplasmic blebs & autolysis Irreversible injury—inability to reverse mitochondrial dysfunction (permanently lose ability to generate ATP) Membrane damage—progressive loss of phospholipids (degraded by Ca2+-activated phospholipases & ↓synthesis from ↓ATP) Cytoskeleton abnormalities (from Ca2+-activated proteases), reperfusion injury (from ROS), lipid breakdown products (like detergent) ↓intracellular AA, especially glycine (normally protect against hypoxic damage) Morphologic changes—defects in cell membrane, abnormal nucleus, lysosome rupture & ER lysis Myelin figures (fragments of damaged membrane in cytoplasm) Free radical injury—primarily superoxide (O2-.), hydrogen peroxide (H2O2), & hydroxyl ion (OH.) Removal—spontaneously decays into non-reactive elements; antioxidants (vit. A, C, E & glutathione) block formation or inactivate O2 + NADPH → O2-. + NADP + H+ (by NADPH oxidase) → H2O2 + O2 (by SOD) → (OH.)(O2.) Storage & transport proteins (e.g. ceruloplasmin & transferrin)—bind iron & copper, needed for formation of ROS Intracellular enzymes—(superoxide dismutase, catalase & glutathione peroxidase) covert radicals into inert substances Injury—lipid peroxidation of membranes (chain reaction), oxidative modification of proteins, DNA damage Inflammation—reaction of vascularized tissue to injury; characterized by heat, swelling & pain Edema—excessive fluid w/in tissues or body cavity, consistent feature of inflammation Exudate (inflammatory edema)—high protein content (specific gravity >1.020) w/ cellular debris; results from ↑vascular permeability Transudate (plasma filtrate)—low protein content (only water & albumin); results from ↑hydrostatic pressure or ↓plasma oncotic pressure Vascular changes—arterioles w/in injured tissue vasoconstrict (for few seconds), then vasodilate (by PGs) & recruitment of capillaries to increase blood flow ↑Permeability—leaves concentrated (viscous) blood remaining in vessels & slowing of flow (stasis) Endothelial cell contraction (“immediate, transient response”)—venules only; cells shorten & cell junctions widen; lasts 15-30min, reversible Mediated by histamine, BK & leukotrienes Endothelial cell retraction—venules & capillaries; reorganization of endothelial cytoskeleton causing membrane to “wrinkle” Reversible, begins 4-6hrs. after injury & lasts >24hrs.; mediated by CKs Direct endothelial cell injury (“immediate, sustained response”)—all vessels; necrosis & detachment caused by burns, chemicals/toxins, infections Begins immediately & lasts till tissue healed Leukocyte mediated endothelial injury—mostly venules; activated leukocytes release proteolytic enzymes & ROS to damage endothelial cells Increased transcytosis—venules only; ↑# & size of endothelial cytoplasmic channels; mediated by vascular endothelial growth factor (VEGF) 7 Lymphocyte emigration—extravasation of leukocytes from blood to site of injury in tissue as enter lymph nodes thru HEV & peripheral tissue thru post-capillary venules 1. Adhesion—leukocytes move from center to periphery of blood vessels (margination) w/ stasis; vascular endothelium activation produces MHCII & adhesion molecules Pavementing—selectins mediate weak bond that allows rolling of leukocytes along endothelial surface, studding endothelium w/ leukocytes Adhesion molecules—present of endothelium w/ complements on leukocyte membranes; 4 families (selectins, Ig family, integrins, & mucin-like glycoproteins) Selectins—L-selectin expressed on nearly ALL leukocytes; E- & P-selectin inducible on vascular epithelium & expressed on platelets (P-selectin) Integrins (LFA-1, Mac-1 or VLA-4)—bind ECM proteins of BM or Ig-superfamily Ig superfamily (ICAM-1 or VCAM-1)—contain at least one Ig domain Modulation—promotes binding of adhesion molecule & leukocyte Redistribution—migration of adhesion molecules from cytoplasm to cell surface e.g. P-selectin migrates from Weibel-Palade bodies to membrane w/ chemical mediators (histamine, thrombin, PAF) Induction—delayed synthesis of adhesion molecules, e.g. synthesis of E-selectin w/ inflammation mediator (IL-1 & TNF) Avidity—increase binding of adhesion molecules to leukocytes e.g. ↑LFA1/ICAM1 & CR3 interaction by leukocyte activation (by chemoattractants, IL-8) causing conformational changes 2. Transmigration (diapedesis)—leukocyte squeezes through endothelial gaps in venules; involves adhesion molecule PECAM1 & CD31, break thru BM w/ proteases Integrins bind Ig-superfamily to activate lymphocyte → cell stops rolling (arrest & adhesion) & flattens shape 3. Chemotaxis—leukocyte migrates along chemoattractant gradient (IL-8) towards site of injured tissue (“ground zero”) Chemoattractants—bind specific receptor on leukocyte membrane → activates phospholipase C → ↑intracellular Ca 2+ → actin crosslinking in cytoskeleton → pseudopod formation → cell movement → pseudopod degrades Exogenous—originate from source of inflammation, e.g. bacterial products Endogenous—produced by injured host, e.g. complement components, leukotrienes, cytokines, PAF, fibrinopeptides 4. Activation—stimulation by chemoattractant causes production of arachidonic acid metabolites, release of lysosomal enzymes, oxidative burst, ↑avidity 5. Phagocytosis—ingestion & destruction of inflammation-causing material by leukocyte Recognition & attachment—requires opsonization (Fc fragment of IgG & complement C3b) Engulfment—via pseudopods to form phagosome; lysosomes fuse to form phagolysosome Killing & degradement—acid hydrolases degrade bacteria Oxygen-dependent (oxidative burst)—hydrogen peroxide-myeloperoxidase-halide system uses ROS to insert halide covalently into bacteria Chronic granulomous disease (CGD)—no oxidase to initiate oxidative burst NADPH → NADP+ & O2 → O2-. (by membrane oxidase) → H2O2 → OCl. (+ Cl-) hypochlorous acid (bleach) Oxygen-independent—lysosomal contents, e.g. bactericidal permeability increasing protein (BPI), lysozyme, lactoferrin Release of leukocyte products—“regurgitation during feeding,” lysosomal contents released & damage tissue; e.g. RA damage to joints Lymphocyte recirculation—naïve lymphocytes tend to remain in blood & 2° lymphoid tissue, searching for Ag for activation Effector & memory lymphocytes that migrate into peripheral non-lymphoid tissues reenter blood thru thoracic duct Chemical mediators of inflammation—originate from plasma (precursors must be enzymatically activated) or cells (preformed or synthesized de novo) May bind other mediator-containing cells causing 2° release (amplification); act on single or variety of cell types; short-lived; most have harmful effects Vasoactive amines—histamine & serotonin Histamine—preformed; primarily w/in mast cells; cause vasodilation & ↑permeability via endothelial cell contraction; notorious for tissue damage Serotonin—preformed; found w/in platelet granules; cause vasodilation & ↑permeability Plasma proteases—all plasma derived Complement system—lyses microorganisms thru membrane attack complex; C3a-5a cause vasodilation & ↑permeability; C5a chemoattractant; C3b opsonin Kinin system—factor XII (Hageman factor) activated by injured tissue → activates kallikrein (thru factor XIIa) → activates Hageman factor (amplification) → activates bradykinin → pain of inflammation & ↑vascular permeability thru endothelial contraction Clotting system—Hageman factor activates: Fibrinopeptides—chemoattractants & ↑vascular permeability Thrombin—modulation of adhesion molecules (redistribution of P-selectin) Arachidonic acid metabolites (eicosanoids)—normally bound to membrane phospholipids, released & metabolized when activated by inflammation Steroids—anti-inflammatory b/c inhibit phospholipases that release these metabolites Cyclooxygenase pathway—occurs in leukocytes, platelets & endothelial cells; produces prostaglandins → pain & fever PGI2 (prostacyclin)—causes vasodilation & inhibits platelet aggregation Tthromboxane A2—causes vasodilation & promotes platelet aggregation PGD2, PGE2 & PGF2—cause vasodilation & potentiates edema by ↑permeability effects of other mediators Aspirin & NSAIDS—anti-inflammatory b/c inhibit cyclooxygenase Lipoxygenase pathway—occurs in neutrophils; produces leukotrienes Leukotriene B4—chemoattractant C4,D4 & E4—cause vasoconstriction, bronchospasm (maybe fatal) & ↑permeability thru endothelial cell contraction Platelet activating factor (PAF)—synthesized in leukocytes & endothelial cells; w/ TNF- triggers clotting to prevent pathogen from entering blood ↑Vascular permeability, enhances leukocyte adhesion (redistribution), chemoattractant, promotes synthesis of other mediators (e.g. leukotrienes) Blood-borne pathogens (sepsis) induce systemic TNF- release from liver M → systemic vasodilation & clotting (septic shock) Cytokines—synthesized by activated leukocytes; act thru cell-surface receptors to induce changes in gene expression Chemokines—2 families, either c-c or c-x-c (2 adjacent cysteins or separated by another AA) IL-1 & TNF---leukocyte adhesion (↑synthesis of adhesion molecules, activates endothelium), ↑synthesis of PG & other CKs; fever, loss of appetite, ↑sleep IL-2—growth of T-cells IL-3—GF for HSC IL-4—stimulates isotype switching to IgG, IgE, ↑MHCII & growth of T-cells IL-6—activates lymphocytes to ↑Ab production IL-8—activates neutrophils & chemotactic factor that recruits neutrophils, basophils & T-cells to site of infection along gradient IL-12—activates NK cells & induces differentiation of CD4 cells into TH1 cells IFN---↑MHCI/II TNF---activates M CD40L—growth, differentiation & isotype switching of B-cells TNF-, IL-1 & IL-6—together initiate innate immune respone → fever (endogenous pyrogens) to potentiate Ag-processing & slow replication of pathogens f-MLP (N-formylated methionine)—chemoattractant produced by bacterial proteins Nitric oxide—soluble free radical gas produced by endothelial cells & macrophages; potent vasodilator (relaxes SM); combines w/ O 2 to form free radicals w/in macrophages 8 Acute inflammation—resolution, healing, abscess formation, or progression to chronic inflammation Resolution—tissue returns to normal in morphology & function; only when injury is short-lived, lacks major tissue destruction & tissue capable of regeneration Healing—CT replacement (fibrosis or scarring) if tissue consists of permanent cells, no surviving cells remain, or CT framework (scaffolding) is disrupted/destroyed Abscess formation—supperative (purulent) inflammation caused by pyogenic (pus-producing) organisms Pus cavity (exudate of neutrophils & necrotic cell debris) w/ band of surrounding neutrophils secreting enzymes that digest tissue (liquefactive necrosis) Cavity either collapses, followed by healing & scar formation; or microorganisms break out & disseminate via bloodsteam (sepsis) Chronic inflammation—progressive acute inflammation or low-grade, asymptomatic progressive process due to: Persistent infections—certain microorganisms (e.g. mycobacterium tuberculosis, treponema pallidum) & some fungi; low toxicity does not elicit acute response Prolonged exposure—e.g. coal miner’s lung or atherosclerosis from persistent hypercholesterolemia Autoimmune disease—e.g. RA or systemic lupus erythematosus (SLE) Characteristics—tissue infiltration by mononuclear inflammatory cells (not neutrophils), tissue destruction, healing by fibrosis Official start of chronic inflammation when monocyte replaces neutrophil as primarily cell type extravasating into tissue by 48hrs. Activated by CKs from T cells & non-immunologic stimuli (e.g. bacterial endotoxins) to form macrophages, epithelioid or Giant cells Lymphocytes—secrete lymphokines that activate M, which secrete monokines to activate lymphocytes (cycling) Granulomatous inflammation—chronic inflammation characterized by focal aggregates (granulomas) of M Immune granuloma—M phagocytose substance unable to digest (usually microorganism, e.g. TB or syphilis) Epithelioid cells—M that phagocytosed immunogenic substance that induces T-cell response & transforms M Langhans-type Giant cells—fusion of epithelioid M, up to 50 M nuclei arranged around periphery of cell Surrounding epithelioid & giant cells is collar of lymphocytes, plasma cells & fibroblasts (may produce collagen); may have caseous necrotic center TB (tubercle granuloma), leprosy, histoplasmosis, coccidioidomycosis, Q fever, brucellosis, syphilis (gumma tubercle), sarcoidosis, Crohn’s disease Foreign body granuloma—non-immunogenic indigestible substances (e.g. talc, asbestos, silica, sutures); foreign substance usually visible Giant cell nuclei arranged randomly (not peripherally), may or may not have collar of other cells Hypersensitivity (allergic) reactions: Type I (anaphylactic type)—rapid response w/in min; combination of Ag & IgE bound to mast cells (tissue) or basophils (blood) in individuals previously sensitized to Ag Nature of Ag, genetic factors & CK profile → TH2 CD4 cells secrete IL-4 → activates IgE-producing B-cells → IgE attaches to IgE FcR on mast cells & basophils → Ag binding directly to & crosslinking of IgE causes release of 1 & 2 mediators from granules → tissue reaction & lymphocyte infiltration 1 mediators—e.g. histamine & TNF-; rapid & short-acting vs. 2 mediators—e.g. leukotrienes C4, D4, E4; begin w/in hours & long-lasting Local reactions (local anaphylaxis)—allergic rhinitis, urticaria, angioedema, asthma Systemic anaphylaxis—systemic release of mediators if Ag enters vascular bed; general itchng (hives) to respiratory distress, vascular collapse (shock) & death Epi—stimulates reformation of endotheilial TJs (↓edema & ↑BP); relaxes bronchial SM & stimulates HR Type II—mediated by Ab directed towards Ag present on surface of cells or other tissue components Complement dependent reactions—e.g. transfusion reactions, erythroblastosis fetalis, autoimmune hemolytic anemia, pemphigus vulgaris, drug reactions IgG or IgM binds cell surface & activates complement cascade → MAC causes cell lysis & C3b binds M receptors to enhance phagocytosis Ab-dependent cell-mediated cytotoxicity (ADCC)—e.g. tumor cell destruction, parasite destruction, allograft rejection IgG coats target cell & FcR on nonsensitived effector cells (monocytes, neutrophils, eosinophils, NK cells) bind to cause cellular injury Ab-mediated cellular dysfunction—e.g. myasthenia gravis, Ab directed against AChR on MEP of skeletal muscle resulting in muscle weakness e.g. Grave’s disease, anti-thyroid hormone receptor Ab stimulate secretion of TH resulting in hyperthyroidism Ab cause dysfunction w/o cell injury or inflammation Type III (immune complex mediated)—soluble immune complexes produce tissue damage as a result of complement activation thru deposition at filter barriers Complex size—small or intermediate size complexes formed in slight Ag excess most pathogenic b/c tend to survive longer in circulation w/o phagocytosis Mononuclear phagocytic system—dysfunction of these cells or their overload by extensive complex formation reduces elimination Also affected by charge of complexes, valence of Ag, avidity of Ab, affinity of Ag, 3D structure of complex & hemodynamic factors Systemic immune complex disease—e.g. acute & chronic serum sickness, SLE, farmer’s lung 1. Formation of Ag:Ab complexes in circulation, typically 5 days after Ag introduction 2. Deposition in various tissue, facilitated by ↑vascular permeability w/ CK release after activation of inflammatory cells FcR or C3bR 3. Inflammation reaction thru complement & inflammatory cell activation, coagulation & platelet aggregation; typically 10 days after Ag intro Local immune complex disease (Arthus reaction)—localized area of tissue necrosis resulting from acute immune complex vasculitis Begins few hours & peaks 4-10hrs after Ag introduction Type IV (cell mediated)—initiated by Ag-specific T-cells; delayed-type hypersensitivity (intracellular pathogens) & CD8 cytotoxicity (virus infection & allograft rejection) Delayed-type hypersensitivity—prior sensitization to Ag → memory TH1 cells activated → secrete CKs (IL-2, IL-12, IFN-, TNF-) → M activation e.g. (+) TB skin test—previously exposed individual shows reddening & induration of injection site w/in 8-12hrs & peaking at 24-72hrs Epitheloid cells—morphologic transformation of M following persistent Ag; surrounded by lymphocytes to form granuloma (granulomatous inflammation) T-cell-mediated cytotoxicity—sensitized CD8 cells kill Ag-bound target cells (virus infected cells & acute cellular allograft rejection) Perforin-granzyme-dependent killing—perforin pokes holes in target cell membrane facilitating osmotic lysis & granzyme induces apoptosis Fas-Fas ligand-dependent killing—Fag ligand on surface of activated CD8 cells interacts w/ Fas on some target cells to induce apoptosis Cytokine-mediated injury—“CK storm” or “capillary leak syndrome,” directly injures tissue (particularly vascular endothelium) & may cause multisystem organ failure Healing—always involves cell growth, either own tissue cells (regeneration) or fibroblasts & endothelial cells (fibrosis) Labile cells—continuously dividing throughout life; short G0; e.g. epithelial cells & hematopoietic cells Stable (quiescent) cells—low rate of cell division; prolonged G0; reenter cell cycle only under certain conditions; e.g. endothelial, parenchymal & mesenchymal cells Permanent cells—non-dividing cells; e.g. neurons of CNS, cardiac & skeletal muscle cells Surgical incision—blade injures limited # epidermal cells; space immediately fills w/ clotted blood & dehydrated scab on surface 24hrs—cellular evidence of acute inflammation (neutrophils) appear 3 days—acute inflammation replaced by chronic inflammation; granulation tissue begins to form; GFs induce regeneration of basal cells of epidermis 5 days—granulation tissue fills incisional space 2 weeks—collagen accumulates (dense fibrous union) & leukocytes, edema, vascularity disappear 1 month—CT scar completely formed & epidermis fully regenerated Growth factors—promote growth by recruiting G0 cells back into cell cycle; also influence differentiation & locomotion Competence factors—some render cells in G0 or G1 ready to undergo DNA synthesis Progression factors—stimulate DNA synthesis in cells already rendered competent Epidermal GF family: EGF—progression factor produced by several cells including platelets; mitogenic for epithelial cells & fibroblasts; binds EGF receptor (c-erb B1) Transforming GF (TGF-)—identical in structure & function to EGF; also binds c-erb B1 receptor Platelet-derived GF (PDGF)—competence factor produced by M, endothelial, SM & some some tumor cells; stored in granules of platelets & released when activated 9 Fibroblast GF (FGF)—acidic & basic versions; produced by M, neural tissue; stimulates growth of fibroblasts & induces angiogenesis TGF---produced by platelets, endothelial cells, T cells & M; inhibits (epithelial) or stimulates cell growth (collagen & fibronectin) & fibroblast chemotaxis Fibrogenic cytokines (IL-1 & TNF-)—mitogenic & chemotactic for fibroblasts, stimulates collagen production by fibroblasts (scar formation) Fibrosis—formation of new blood vessels (angiogenesis or neovascularization), migration & proliferation of fibroblasts, deposition of ECM 1. 24hrs—fibroblasts & vascular endothelial cells proliferate 2. 3-5 days—granulation tissue forms; pink, soft granular appearance Angiogenesis—“budding” off pre-existing vessels; immature & leaky (granulation tissue is edematous); induced by FGF & vascular endothelial GF Fibroblast proliferation—triggered by EGF, TGF- (stimulates new fibroblasts to synthesize & secrete collagen), PDGF, FGF 3. Weeks to months—granulation tissue transformed (remodeled) into mature scar; ↑collagen, ↓fibroblasts & vascularity Immunosurveillance—theory that primary reason for cellular immunity is for defense of body against neoplastic (tumor) cells Neoplastic cells immunogenic—b/c spontaneous regression of tumors, regression of metastases after excision of primary tumor, infiltration of solid tumor w/ immune cells Tumor-specific Ab in circulation, higher incidence of certain tumors among immunodeficient patients Tumor-specific antigens (TSA)—found only in tumor cells; endogenously processed Ag; point mutations in normal cytosolic proteins Tumor-associated antigens (TAA)—found on both tumor & normal cells; often reexpressed embryonic antigens or overexpression Onco-fetal antigens—TAA normally expressed during fetal development, e.g. alpha-fetoprotein & carcinoembryonic antigen Evade immune response—low immunogenicity (no peptide:MHC, no adhesion molecules, no co-stimulators) Antigenic modulation—Ab against tumor antigen induces endocytosis & degradiation leading to selection of unrecognizeable antigen variants Tumor-induced immune suppression—factors (e.g. TGF-) secreted by tumor cells actually inhibit TH1 cells & cell-mediated immunity directly Tumor cell destruction—complement-mediated lysis; Ab-dependent cell cytotoxicity (ADCC); non-specific direct cytotoxicity by NK cells & M Immunotherapeutics—generally no more effective than conventional cancer treatment protocols Active immunization (tumor vaccines) or passive immunity (w/ injection of anti-tumor Ab to activate cell-mediated immunity or directly w/ immunotoxins) Expression of transfected genes in tumor cells—transfect tumor cell w/ B7 → activates TSA CD8 cells OR tranfect tumor cells w/ GM-CSF → recruits DCs that present tumor antigens to T-cells Transplant immunology—universal rule of transplantation (engrafted tissue will be rejected by immune system if any MHC antigens on graft are absent from recipient) Syngeneic—always accepted by recipient Autograft—graft from one part of body to another on same individual Isograft—graft b/w genetically identical individuals Allogeneic—always rejected by recipient; causes alloreaction due to alloantibodies reaction against alloantigen on foreign MHCs Allograft—graft from genetically dissimilar donor to recipient of same species Xenograft—graft from donor of different species HLA (MHC) matching—improves survival of transplanted organs by ↓incidence & severity of allograft rejection; only kidney & marrow routinely typed Mismatches of MHCII often more rapid & severe rejecting than mismatches of MHCI Minor histocompatibility antigens—allelic (different AA sequence b/w individuals) forms of non-MHC molecules Responsible for transplant rejection when MHC matched; generated during normal protein turnover & peptides presented on surface of cell A,B,O blood groups—Ag differences in CHO on glycolipids of RBCs; cross-match test recipient’s Ab to donor serum Tissue typing—detection of HLA antigens on surface of lymphocytes Serologically (microcytotoxicity test)—test all HLA loci DNA or molecular techniques (PCR)—best for MHCII, all HLA loci Mixed lymphocyte culture (MLC)—for MHCII; in vitro culture containing blood leukocytes from both donor & recipient Culure of irradiated donor blood (not able to proliferation but still present Ag) & recipient labeled (3H-thymidine) T-cells T-cell proliferation depends on differences in MHCII alleles Ab screening—detection of Ab in recipient that could lead to graft rejection, panel of 40-60 cells of known tissue type to cover all recognized HLA specificities Crossmatch of recipient serum w/ potential donor lymphocytes; panel reactive antibody (PRA) Graft rejection—exhibits immunological memory & Ag specificity; either hyperacute, acute, or chronic Alloreactivity—T-cell recognition of peptide Ag on incorrect HLA type Graft survival—depends on establishment of blood supply, passenger leukocytes in graft, grafting site, & type of tissue grafted (e.g. cornea, liver) Hyperacute—type III hypersensitivity of immune complex deposition & complement activation in vessels walls; due to ABO mismatch Preformed MHCI Ab can arise from pregnancy, blood transfusions, or previous transplants Acute—type IV hypersensitivity reaction w/in days of transplantation Chronic—months or years after transplantation; thickening of vascular endothelium leading to ischemia & graft death Graft vs. host disease (GVHD)—HLA differences b/w bone marrow graft & recipient; presence of immunocompetent cells in graft; immunodeficiency in patient Acute—rash, diarrhea, hepatosplenomegaly Chronic—desquamation, hepatosplenmegaly, lymphadenopathy, diarrhea, wasting syndrome, immunodeficiency Immunosuppressive drugs: Cytotoxic agents: Alkylating agents (cyclophosphamide)—converted to phosphoramide mustard (chemical weapon in WWI) → alkylates & crosslinks DNA Anti-metabolites (azathioprine)—converted to 6-mercacptopurine → 6-thioinosinic acid Inhibits production of inosinic acid essential for DNA replication (targets only actively dividing cells) Methotrexate—prevents DNA replication by inhibiting dihydrofolate reductase (synthesis of thymidine); prevents marrow GVHD Corticosteroids (prednisone)—pro-drug synthetic derivative of hydrocortisone → prednisolone; used as acute immunosuppressant b/c adverse side effects ↑synthesis of IB that prevents NFB from entering nucleus → ↓CK production → ↓inflammation Antibiotics (cyclosporin)—derived from soil fungus; inhibits activation of T-cells w/o targeting dividing cells Binds calcineurin w/ cyclophilins → inhibits phophatase activity & prevents NFAT activation → no synthesis of IL-2 (T-cell activation) Tacrolimus (FK506)—binds calcineurin w/ FK-binding proteins to prevent NFAT activation Rapamycin (sirolimus)—binds FK-binding proteins but blocks T-cell activation by preventing signal transduction from IL-2R; more toxic Antibodies (anti-T-cell reagents)—injected at 5-15day courses during episodes of acute rejection; used only once per patient b/c immune complex formation later Anti-T-cell Ab made in sheep or goats (antithymocyte globulin) or from hybridoma cells 10 Autoimmunity—immune response specific against one or more self tissues (systemic or organ-specific), usually resulting in chronic tissue destruction; specific Ag generally unknown 3 Requirements for autoimmunity: 1. Presence of autoimmune reaction 2. Evidence that reaction not secondary to tissue damage 3. Absence of another well-defined cause of disease Peripheral tolerance—failure of T-cell anergy & failure of activation-induced cell death, failure of regulatory T-cells, polyclonal activation (superantigen) IFN- induces MHCII expression on cells normally not APC; loss of anatomical barriers to T-cell infiltration (e.g. CNS) Molecular mimicry—crossreactivity of immune response to infection damages host collaterally, e.g. strep M protein & cardiac myosin Genetic factors—HLA associations (particular alleles more likely to bind self peptide) & non-HLA associations Endocrine factors—tissue-specific hormone production & well vascularized; higher frequency in females (due to higher levels of P & E, fluctuate during month) Environmental factors—drugs & toxins; stress, infectious agents; e.g. smoking & immune complex deposition in alveolar BM of Goodpasture’s syndrome Organ-specific—thru type II (Ab response) & IV (cell-mediated response) hypersensitivity mechanisms Grave’s disease—Ab against TSH receptor stimulating TH secretion (TH2 response) Hashimoto’s disease (chronic thyroiditis)—Ab against thyroglobulin, thyroid peroxidase & TSH receptor (TH1 response) → hypothyroidism Pernicious anemia—Ab against gastric parietal cells & intrinsic factor Autoimmune adrenalitis (Addison’s disease)—Ab against adrenal cortex Primary biliary cirrhosis—Ab against liver mitochondria Antiglomerular basement membrane disease—Ab against BM (Goodpasture syndrome if no crossreactivity against pulmonary alveolar BM) Myasthenia gravis—Ab against ACh receptor in muscle Autoimmune thrombocytopenia (ITP)—Ab against platelets Autoimmune hemolytic anemia (AIHA)—Ab against RBCs Type I diabetes mellitus—Ab against pancreatic islet cells Systemic—thru type III (immune-complex mediated) hypersensitivity mechanisms Systemic lupus erythematosus—1:500 African & Asian women; remitting & relapsing, often febrile illness characterized by injury to skin, joints, kidney & serosa Butterfly-shaped skin rash on face giving appearance of wolf’s head (Latin lupus means wolf) Sjogren syndrome—characterized by dry eyes (keratoconjuctivitis sicca) & dry mouth (xerostomia) from destruction of lacrimal & salivary glands Systemic sclerosis (scleroderma)—characterized by excessive fibrosis throughout body Inflammatory myopathies—characterized by injury & inflammation of mainly skeletal muscle Mixed CT disease—coexistence of featuers suggestive of SLE, polymyositis, & systemic sclerosis; high titers of Ab to RNP particles containing U1 RNP Rheumatoid arthritis—affecting mainly joints thru nonsuppurative proliferative synovitis progressing to destruction of articular cartilage & ankylosis of joints Superantigens—microbial proteins capable of activating both polyclonal CD4 response & M; can activate autoreactive lymphocytes or exacerbate autoimmune disease Forms bridge b/w CD4 cell receptors & MHCII on APC (not need Ag-processing, binds directly to TcR -chain & MHCII -chain) → T-cell & M activation → large amounts of CKs from activated CD4 cells & M (systemic toxicity & suppression of adaptive immune response) Toxic shock syndrome (TSS)—suddent onset of fever (103-106°) lasting 2-5days, cough persisting up to 4wks; higher exposure may lead to septic shock & death Associated w/ use of tampons (rayon) since 1978; strains of staphylococcus aureus that produce bacterial toxin TSST-1 or enterotoxins A, B, & C Streptococcal TSS (STSS)—strains of streptococcus that produce toxin exotoxin A or B; highly invasive group A named “flesh-eating bacteria” Mouse mammary tumor virus (MMTV)—milk-born retrovirus passed on to suckling pups, integrated into host genome & code integral membrane proteins MMTV superantigen stimulates CD4 cells → activate B-cells thru IL-4 & CD40L → virus replicates well in activated B-cells MMTV expression in fetal thymus causes V-chain clonal deletion Intracellular bacteria—intracellular habitat (mainly monocytes & M); delayed type hypersensitivity; granulomatous tissue reaction (containment) T-cell mediated response—turn M into effector (CD8) cells to kill intracellular bacteria; B-cells rarely interact w/ Ag b/c intracellular location Listeria—primarily acute infection that causes listeriosis usually associated w/ food-borne illness, presents as meningoencephalitis Internalins A & B—bind to epithelial E-cadherin to gain access to host cell Listeriolysin—pore activated at low pH that allows evasion of listeria from endosome into cytoplasm of M Infection of hepatocytes—listeria forms assembly of actin filaments that push pathogen to surface forming pseudopod that invaginates into adjacent cell NK cells produce IFN- → M activation → Kuppfer cell phagocytosis → recruitment of M & neutrophils → microabscess formation → T-cell recruitment → granuloma formation Leprosy—spectral disease ranging from tuberculoid pole (T-cell mediated immunity) to lepromatous pole (deficient TH2 immunity & multibacillary lesions) Tuberculoid—few anesthetic plagues, close to skin lesions, granulomas Lepromatous—multiple plagues & nodules, diffuse inflammatory involvement of nerves, no granulomas Tuberculosis—grows w/in M vacuole; most individuals contain primary infection & develop CD4 cell response, 5-10% develop progressive disease Activates M or DCs thru TLR-2 & TLR-4 → IL-12 drives TH1 response → CD4/8 & NK cells produce IFN- → TNF- synergizes w/ IFN- to induce NO & form granulomas Chlamydia trachomatis—obligate intracellular pathogen that grows w/in phagosome Causes trachoma, inclusion conjunctivitis, neonatal pneumonia, genital tract infection, lymphogranuloma venereum Rickettsioses—grow in cytoplasm of vascular endothelial cells & M & SM (Rocky Mountain spotted fever) Immunodeficiencies: Primary—due to genetic defect that affects development of immune system; usually affects children (95% <6yo), life-long; only 1:10,000 births Ab deficiencies (50%)—susceptibility to pyogenic bacteria & lytic viruses Cellular deficiencies (10%)—viruses & intracellular bacteria, fungi & protozoa Phagocyte deficiencies (18%)—susceptibility to both intra- & extracellular organisms b/c cannot phagocytose Complement deficiencies (2%)—pyogenic bacteria b/c loss of opsonization, Neisseria infections w/ loss of membrane attack complex Combined (20%)—severe b/c affects both arms of immunity Secondary—acquired from external factors that affect immune system; more common Symptoms—more severe, persistent, recurrent infections w/ unusual (opportunistic) organisms; FTT or MR, thrush, paucity of LN (no immune response) Hematologic abnormalities, evidence of autoimmunity, diarrhea & malabsorption, skin lesions HSC differentiation defects: Severe combined immunodeficiency—mutations in RAG-1 or RAG-2 recombination that impairs HSC Adenosine deaminase (ADA) or purine nucleoside phosphorylase (PNP)—requires for conversion of adenosine & guanine into uric acid Defects in enzymes results in buildup of deoxy-ATP & -GTP w/ toxic effects that cause destruction of B- & T-cells 11 Reticular dysgenesis—↓T- & B-cells due to defect in HSC maturation Wiskott-Aldrich syndrome—progressive ↓T-cells due to defect in membrane glycosylation & HSC maturation Thymic epithelium—defect affecting structure for T-cell maturation DiGeorge syndrome—defect in thymic epithelium development resulting in ↓T-cells, often from transient perinatal exposure or FAS B-cell progenitor defect—before or after pre-B-cell formation in marrow X-linked agammaglobulinemia—↓B-cells (↓all Ig isotypes) due to defect in Btk (Bruton’s tyrosine kinase) Ig production defect—defect in activation of B-cell into plasma cell, Ig synthesis, or catabolism of Ig (too rapid turnover) Common variable immunodeficiency—most frequent; defect of differentiation of B-cells into plasma cells affecting various isotypes Selective Ig isotype deficiencies—defect in Ig synthesis (isotype switching) that most commonly affects IgA T-cell activation defect—either ↓CD4 cells or ↑supressor cells X-linked SCID—mutation in common c chain from IL-2R, IL-4R & IL-7R resulting in defect in differentiation of B- & T-cells Bare lymphocyte syndrome—defective transcription of MHCI/II genes resulting in deficient CD4/8 cells AIDS—HIV virus binds CD4 & chemokine receptor (CCR5) on host cell; some individuals immune b/c defective CCR5 receptor prevents its expression on cell surface RNA virus w/ RNA nucleocapsid surrounded by lipid envelope derived from host-cell membrane; contains protease, reverse transcriptase & integrase Lentiviruses—(Latin lentus meaning slow) group of retroviruses that cause slowly progressing disease Acute period—often asymptomatic or flu-like symptoms; appears w/in 2 weeks & lasts 2-6wks Targets CD4 cells, M & DC (all express CD4) via coreceptor CCR5 resulting in sudden dramatic drop in CD4 cell count Clinical latency—no clinical symptoms w/ low detectable viremia as immune response removes virus from blood (seroconversion) Virus trapped on follicular DCs w/ 2° lymphoid tissues; lasts 2-15yrs AIDS—characterized by <200/mm3 CD4 cell count; ↑opportunistic infections, lymphoid cancers & Kaposi’s sarcoma; dementia; death w/in 2yrs Switch to lymphocyte-tropic from macrophage-tropic type late in infection aggressively infecting CD4 cells thru CXCR4 co-receptor HIV vaccine—enormous genetic diversity w/ recombination among clades (unique genotypes) & even subtypes (9) common Different subtypes affecting world (HIV-1 primarily in developed countries & less-virulent HIV-2 in Africa & Asia) Whole or modified virus vaccine: Live, attenuated HIV—assessed in primates; most closely mimics HIV (excellent protection in animal models); could potentially cause disease Whole, killed HIV—risk that prep might include some active virus & difficult to produce in large amounts Pseudovirions (non-replicating HIV-like particles)—close to phase I trials; difficult to produce Protein-based vaccines: Viral surface proteins (gp120)—in phase II/III trials; safe & simple to prepare but not always able to elicit immune response to HIV HIV peptides—in phase I trials; simple & cheap but poor immunogenic & unstable Genetically-engineered vaccines: Naked DNA—in phase II trials; simple & cheap, but possibly harmful & elicit only modest immune response Live viral vectors (non-HIV viruses w/ HIV proteins)—in phase II trials; complicated to prepare, modest response DCs infected w/ Canarypox/HIV vector—in phase I trials; strong T-cell response b/c HIV proteins introduced into Ag-processing Combination vaccines—in phase I/II trials; stimulates both arms of immune response Vaccine development—providing protective immunity w/o causing disease (safe & effective); Latin vaccus meaning cow from Jenner’s cowpox vaccine for smallpox Toxoids—toxins denatured for vaccination w/ formalin, neutralizing Ab made against toxoid in host (e.g. diphtheria & tetanus) Passive immunity—transferring of protective Ab to new host (e.g. anti-venom) States of vaccine development: 1. Study characteristics of natural infection & protective immune response 2. Develop animal model for disease 3. Develop vaccine concept/design 4. Test vaccine in animal model 5. Clinical trial in humans—phase I for safety; phase II/III for safety & efficacy Types of vaccines: Live attenuated microorganisms—e.g. polio (oral), influenza virus (nasal, e.g. trivalent strains in FluMist), MMR (injection); more potent b/c limited replication Inactivated whole microorganisms—poliovirus, hepatitis A virus, influenza virus; treated w/ formalin, heat or irradiation Subunit vaccines—composed of individual Ag viral components; e.g. diphtheria & tetanus toxoids Acellular pertussis—3 purified Ag (inactived pertussis toxin (PT), filamentous hemagglutin (FHA) & pertactin) Pneumococcal polysaccharide (PPV)—purified from 23 strains of streptococcus pneumoniae Meningococcal polysaccharide—purified from serogroups A, C, Y, & W-155 Hepatitis B virus—recombinant surface Ag Glycoconjugates—polysaccharides coupled to protein carriers; CD4 cells respond to protein carrier → stimulate specific B-cell response to polysaccharide Haemophilus influenzae type B (Hib) conjugate vaccine—capsular oligosaccharides from serotype B conjugated to tetanus toxoid Pneumococcal conjugate vaccine (PCV)—capsular oligosaccharides from 7 serotypes conjugated to mutant diphtheria toxin Adjuvants (immune enhancers)—↑size & ↓solubility of Ag → induce inflammation & ↑helper immune response Aluminum salts, CKs, strong helper T-cell Ag (combination vaccines), Bacterial cell wall constituents (TLR ligands)—muramyl dipeptide, monophosphoryl lipid A, bacillus calmette-guerin (BCG) Bacterial exotoxins (cholera toxin & heat-labile toxin), biopolymers (immune-stimulating complexes ISCOMs & biodegradable microspheres) DTP—pertussis bacteria serves as adjuvant to diphtheria & tetanus toxoids; or DTaP w/ acellular pertussis toxoid instead New approaches—including using transgenic plants as vaccine production systems DNA recombination—production of subunit vaccines, removing genes, “naked” DNA, live viral/pseudoviral/bacterial vectors “Naked” DNA vaccine—plasmid infects myocytes → Ag released & phagocytosed by APCs → T-cell activation & B-cell Ab production Peptide epitopes—protein fragments & peptides conjugated to protein carriers 12 IID EX AM 2 History of virology: Hippocrates (400-500BC)—epidemic of apparent parotitis (mumps) Democratus (500BC)—rabies Aristotle (322BC)—behavior of rabid dogs & transmission thru bites Celsus (100AD)—related hydrophobia in man to animal rabies; Described “virus” as disease producing vapor vs. venom Jenner (1798)—cowpox & smallpox Pasteur (1880)—rabies Buist of Edenburgh (1887)—first to microscopically view vaccinia Ivanovsky & Beijerinck (1892)—filterable agent that transmits disease Loffler & Frosch (1899)—agent of foot-and-mouth disease Ellerman & Bang (1908)—transmission of fowl leucosis Rous (1911)—transmission of avian sarcoma Twort & D’Herelle (1917)—bacteriophages Enders & Robbins (1945)—cell culture Virus definition—genetic material that utilizes (parasitizes) biochemical machinery of host cell (intracellular) w/ transmission of infectious stage (extracellular) Size—some bacteriophages & parvoviruses (smallest) to vaccinia (largest) Virion—infectious viral particle Structure—protomer (protein) → capsomer (5-6 protomers) → capsid (protomers & capsomers) → nucleocapsid (nucleid acid core + capsid) Icosahedral—regular polyhedron nucleocapsid of 20 triangular faces w/ 12 corners Helical—protomers/capsomers of capsid follow helical structure of interior nucleic acid core Enveloped—pleomorphic glycoprotein membrane surrounding nucleocapsid & tegument (area b/w envelope & nucleocapsid) Complex—lack clear definition of capsid & envelope (e.g. poxvirus & influenza) OR inherent complexity of capsid (e.g. bacteriophage T2) Central dogma: Transcription—RNA polymerase synthesizes (+) strand mRNA in 5-3’ direction & antiparallel orientation Translation—binding of ribosomes w/ charged tRNAs to mRNA to produce polypeptide RNA transcriptase—rare in eukaryotes; copies mRNA into dsRNA w/ antisense or (-) strand mRNA (negative to synthesis of proteins) Reverse transcriptase—forms DNA complement from mRNA, resulting in RNA paired to cDNA (violation of central dogma) Viral classes: Class I—continuous (+) strand RNA genome; infectious; NO transcriptase; produces single large mRNA primary transcript (e.g. picornviruses) Class II—(-) strand RNA; contains transcriptase; NOT infectious; produces multiple small distinct mRNAs (e.g. paramyxoviruses) Class III—several small (-) strand RNAs; contains transcriptase; NOT infectious; produces multiple small distinct mRNAs (e.g. orthomyxoviruses) Class IV—several small dsRNA; contains transcriptase; NOT infectious; produces multiple small distinct mRNAs (e.g. reoviruses) Class V—two large (+) strand RNAs; contains reverse transcriptase; NOT infectious; produces several different size polypeptides (e.g. retroviruses) Viral lifecycle—productive or lytic cycle; all processes not provided by host cells must be encoded by virus & competition for supplies & machinery determine disease manifestation Permissive cells—allow production of viral progeny following infection vs. non-permissive cells that prevent initiation of cycle OR abortive infections that fails to finish cycle 1. Attachment (adsorption)—protein or CHO attachment molecules bind to cellular receptors (mutations may create new variants with ↑host range or pathogenicity Virus receptors—complement receptor CR2 for EBV; CD4 for HIV; heparin sulfate for HSV Hemagglutination—aggregation of RBCs coated w/ viruses thru attachment proteins; used to titer H-containing viruses like influenza 2. Penetration—usually controlled by virion structure: Non-enveloped—enter by receptor-mediated endocytosis (viropexis) or direct penetration of membrane (e.g. papovaviruses) Enveloped—fuse w/ plasma membrane & deliver nucleocapsid directly into cytoplasm (↓pH controlled w/ endosomes to facilitate optimal fusion) 3. Uncoating—delivery of nucleocapsid to site of replication; initiated by receptor, promoted by acidic environment or proteases w/in endosomes OR partial uncoating may release or allow synthesis of viral proteins to complete uncoating (e.g. reovirus or poxvirus) OR nucleocapsids may be released upon fusion, travel to nuclear membrane & release contents directly into nucleoplasm (e.g. herpesviruses) Eclipse period—stage after virus entry into host cell that causes virus to loose infectivity & identifiable structure; determines length of latency before exiting cell 4. Maturation—assembly of DNA viruses (except poxvirus) occurs in nucleus OR cytoplasm for RNA viruses (except influenza & retroviruses) Empty capsids formed first & then filled w/ genome (e.g. picornviruses & herpes) OR capsids assemble around genome (e.g. retroviruses, togaviruses, (-) strand) Envelopes—acquired by budding from plasma membrane (most RNA viruses) & modified (glycoproteins added) by viral proteins OR budding into ER & Golgi (e.g. flavi-, corona- & bunyaviruses) to remain highly cell-associated Early-late regulation—stages of protein synthesis prior to replication of viral genome are early (proteins need for replication); late proteins for virion formation 5. Release—exocytosis, lysis (naked viruses released w/ cell death), or budding (budding produces most progeny, membrane coated w/ viral M protein) Burst size—yield of virus per cell at end of cycle Differences from eukaryotes—transcription occurs in nucleus & translation in cytoplasm (uncoupled); mRNAs transported across nuclear membrane; monosistronic mRNAs RNA processing (addition of 5’ methyl cap & 3’ poly(A) tail & splicing) reduces volume 30% Adenovirus-5 lifecycle—naked icosahedral DNA virion penetrates cell or phagocytosed → breaks down w/ cellular enzymes (loses penton fiber spikes) to form UVI → loses polygonal shape of capsid to form nucleoprotein core UV2 → penetrates nuclear membrane thru nuclei pocket (UV3) or nuclear pore (UV4) → host RNA polymerase II starts semi-conservative transcription of early genes (5 early mRNAs, initiated at both ends w/ asymmetric displacement) → terminally bound protein initiates replication of displace DNA strand (form “pandhandle” structure due to terminal inverted repeats) → late transcription begins (13 late mRNAs) → mRNAs in cytoplasm translated into structure proteins needed for procapsid & virion formation → release of completed virions (only 1% reach extracelullar fluid, most remain in nucleus) Viral gene product (VA RNAs)—produced early during infection to bind active component of interferon system & prevent antiviral action on dsRNA Poxvirus lifecycle—enveloped DNA virus phagocytosed → host enzymes remove outer membrane → transcription of early genes → uncoatase removes core membrane → release of DNA to newly synthesized DNA polymerase → replication w/in cytoplasmic inclusion bodies (factories, virosomes, or Guarnieri’s inclusion bodies) → transcription of late genes for structural & virion proteins → first membrane assembles around factories forming intracellular naked virus (INV) → second membrane (envelope) surrounds INV at golgi to from extracelullar enveloped virus (EEV) → lysis releases 10,000 virions from several factories ONLY ribosomes missing from self-machinery Poliovirus lifecycle—naked icosahedral RNA virus enters cell by endocytosis → release (+) strand RNA to polyribosomes → cleavage of polypeptide generates RNA polymerase → transcription of (-) strand & replication of genome → viral protein (VPg) attaches to 5’ end of progeny RNA to facilitate translation (like primer) → procapsid & capsid proteins assemble into empty heads → heads filled w/ newly synthesized (+) strand progeny w/ VPg attached → lysis to release virus 13 Precursor polyprotein (NCVPoo)—produced by Class I viruses Rapidly cleaved into NCVP1 (non-capsid viral protein) → cleaved into functional viral proteins (VP0, VP1 & VP3) used in formation of procapsid → processed into structural proteins (VP4, VP2, VP3 & VP1) & non-structural proteins (NCVP-1, -2, & -4) including RNA polymerase Retrovirus (HIV) lifecycle—gp120 viral protein binds CD40 on host cells → enters by fusion → (+) strand RNA released into cytoplasm → reverse transcribed into dsDNA (provirus) → transported into nucleus & integrated into host DNA → mRNAs produced & translated into proteins to modify membrane, form capsids & package viral progeny → virus buds from plasma membrane Viral genetics—genetic drift occurs thru mutations, either spontaneous (thru replication errors) or induced (thru physical or chemical agent) Results of mutation—plaque morphology (associated w/ more rapidly replicating viruses); host range (# or type cells infected); drug resistance Temperature sensitivity (affects protein conformation which affects function) Viral polymerase—NO proofreading mechanism, so RNA viruses, large DNA viruses & retroviruses (2 polymerases) particularly prone to mutation Defective interfering (DI) particles—cause decline in infectious virus thru deletion mutations that compete w/ nondefective viruses for replication machinery Complementation—defective gene supplied by co-infecting (helper) virus; each virus unable to replicated independently Recombination—breakage (DNA physically broken & reformed w/ DNA from another virus) Reassortment—surviving combinations of RNA segments (antigenic shift) or strand switching during transcription (retroviruses Reactivation—defective virus regain capacity to replicate as normal wild-types; regain original material or functionally equivalent Genomic insertion—integration of viral DNA into host genome Viral oncology—all DNA viruses (except parvoviruses) capable of transforming normal cells into cancer cells by continually synthesizing viral proteins Malignant tumor cells—fail to respond to signals controlling growth & location of normal cells Characterized by altered morphology; failure to grow in organized pattern; overgrowth; growth w/ less nutrients; immortality; anchorage-independent growth Focus formation? Neoplastic transformation—converting normal cells into tumor cells (excess growth) Malignant transformation—converting tumor cells into malignant tumor cells due to persistent association of viral genes w/ cells & expression of viral transforming proteins Lysogenic conversion—viral genes incorporated into host (bacterial) genome (nonspecific recombination) for continual expression & transformation w/ time Proto-oncogenes—normally functioning gene w/ capacity to change & cause cancer Oncogene—cancer-causing genes that arise from nearly all types of cells due to one or more mutations Mutational activation—virus encodes protein that interferes w/ cell signaling or disrupts normal genes (e.g. retroviruses inserting into tumor suppressor gene) e.g. RAS GTP-binding signal transduction enzyme, mutation causes to be continually active; found in 20-30% of all cancers Overexpression—chromosomal translocation places active promoter next to proto-oncogene (e.g. Burkitt’s lymphoma & CML & RAS in bladder, breast cancers) Viral oncogenes—retrovirus codes for protein (transactivating factor) to maximize transcription of proviral DNA & cellular genes (acute transforming viruses) Insertional activation—viral promoter or enhancer causes inappropriate expression of cellular genes e.g. long-term repeats (LTRs)—gene promoters encoded by RNA viruses to regulate own transcription Cancer-causing viruses: Epstein-Barr virus—Burkitt’s lymphoma (Sub-Saharan Africa), nasopharyngeal carcinoma (Asians); infectious mononucleosis Hepatitis (B, C & D)—hepatocelllular carcinoma; HBV vaccine helping to prevent cancer HHV8—Karposi’s sarcoma HPV—cervical, anal & oral carcinomas Human T-cell leukemia virus (HTLV)—leukemia Viral immunology—cellular-mediated immunity aids in recovery from intracellular viral infections, Ab determine how virus replicates & spreads Viral spread—extracellular, intracellular to adjacent cells, from parent to progeny Ab—neutralization (binds attachment proteins & prevents binding to cell receptor); opsonization (digestion by M) Complement-mediated lysis (enveloped viruses & virus-infected cells); Ab-dependent cellular cytotoxicity (virus-infected cells) Cell-mediated—CD8+ cells lyse infected cells (Ag-specific); NK cells lyse infected cells (nonspecific); activated M are more resistant to infection IL-2 T-cell proliferation & activation of M APC release IL-1 to stimulate T-cell differentiation TH1 cells—produce IL-2 & IL-3 to stimulate M & TH2 cells TH2 cells—produce IL-3, IL-4, IL-5, & IL-6 to activate B-cells Interferon (IFN)—early non-specific immune response to infection; secreted by various cells to cause noninfected cells to become resistant (antiviral protein production) Subtypes—classified by cells of origin —derived from leukocytes; Tx of herpes zoster & CMV (particularly in immunocompromised), chronic HBV, HCV, & HIV —derived from fibroblasts —product of activated T-cells; induce activation & proliferation of NK cells & CD8+ cells, activates M, induces MHC expression Function—IFN binds IFNR induces oligoadenylate synthetase & protein kinase enzymes activated by presence of viral dsRNA (induction) hydrolysis of ATP by enzymes causes activation of RNase & phosphorylation of elongation initiation factor (eIF-2) degrades mRNAs & inhibits formation of protein synthesis initiation complex NO viral replication or protein synthesis w/in cytoplasm Induction—viruses (dsRNA); bacterial endotoxins; metabolic activators Negative effects—inadvertently enhance inflammation thru NK & CD8+ cells OR transiently suppress cell-mediated immunity by inhibiting protein synthesis Viral evasion of immune system—intracellular spread; antigenic drift; induce non-neutralizing Ab; induces immunosuppression; latency Antiviral therapy—chemical (drugs) & immunological (vaccines) that inhibit various steps unique to viral replication Adsorption—neutralizating Ab prevents attachment to cell receptor Penetration & uncoating—synthetic amines (amantadine & rimantadine) prevent uncoating of influenza viruses; resistance develops w/ single AA change Replication—ideally act on virus-specific polymerases (interfere w/ synthesis or terminate chain upon incorporation) & avoid host nucleic acid synthesis Deoxyribonucleotide analogs: Adenosine dideoxyinosine & adenosine arabinoside Guanosine acyclovir ( ganciclovir) & ribavarin (ribonucleotide) Cytidine dideoxycytidine Thymidine azidothymidine & iododeoxyuridine ( trifluridine) DNA antivirals: Acyclovir (acycloguanosine)—phosphorylated by thymidine kinase, again by cellular kinases & triphosphate form selectively binds & inhibits DNA polymerase Little host toxicity; CMV resistant b/c does not encode thymidine kinase OR resistance develops w/ mutation of thymidine kinase Ganciclovir—acyclovir analog that inhibits ALL herpes viruses including CMV; significant host toxicity Adenine arabinoside (vidarabine)—purine that inhibits DNA polymerase (particular herpes viruses) w/ less systemic toxicity Idoxouridine & trifluorothymidine—halogenated pyrimidines that block synthesis when incorporated in place of thymidine; significant host toxicity 14 Foscarnet—pyrophosphate analog that directly inhibits DNA polymerases of ALL herpes viruses, RNA polymerase of influenza & reverse transcriptase NOT require activation by thymidine kinase (highly active against CMV & acyclovir-resistant herpes); nephrotoxic Antiretrovirals: Azidothymidine (AZT)—thymidine analog phosphorylated to inhibit reverse transcriptase & terminate DNA elongation Reduces opportunistic infections, transient CD4+ cells, delays onset of AIDS; significant toxicity for bone marrow (severe anemia in 80% patients) Dideoxyinosine & dideoxycytidine—inhibits reverse transcriptase; CD4+ cell count, HIV p24 levels, delays onset of AIDS; peripheral neuropathy & pancreatitis RNA antivirals: Ribavarin—synthetic triazole that inhibits synthesis of guanosine 5’-P necessary for viral nucleic acid synthesis; aerosol Tx of some respiratory infections & HIV Vaccines—ideally cause significant illness; exist as only one serotype; stimulates Ab to block infections; have NO oncogenic potential; heat stable Active immunization: Live, attenuated (e.g. Sabin oral polio, measles)—limited host range, temperature-sensitive, cold-adapted, genetically manipulated Whole live (e.g. adenovirus) Killed whole (e.g. Salk polio) or extracts (e.g. pneumococcal capsular polysaccharide OR HBV)—subunits, peptides, toxoids, or whole Passive immunization—specific Ig selected from fractionation of pools of donor plasma w/ high Ab titer to specific virus Inactivated vs. live vaccines—CONS require high doses (high cost); multiple doses; requires adjuvant; shorter duration of immunity; poor cell-mediated immune response PROS—heat stable, NO interference; NO reversion to virulence, fewer side effects Prion diseases (transmissible spongiform encephalopathies (TSEs))—inherited AND infectious, neurodegenerative diseases of CNS Prions—small infectious proteins that lack nucleus acids Species barrier—ease w/ which prions from one species can infect another species Strains—characterized by incubation times & neuropathological profile (i.e. pattern of PrPSc deposition in CNS); properties all retained during transmission Prion protein (PrP)—aberrantly metabolized in prion diseases PrPc—normal form of prion protein, expressed on surface of neurons via glycophosphatidyl inositol (GPI) anchor Primarily -helical 2° structure; sensitive to proteases & soluble in detergents PrPSc—disease-associated isoform, found only in infected brains; partially resistant to protease & insoluble in detergents; -sheets that form aggregates Prion propagation—PrPSc coerces PrPc to adopt disease-associated conformation; disease properties depend on PrPSc conformation Scrapie—prototypic prion disease; recognized in sheep & goats 250yo; up to 1% of sheep population infected in UK Bovine spongiform encephalopathy (BSE)—discovered in 1985; spread in contaminated offal used in manufacture of meat & bone meal fed to cattle BSE-contaminated foodstuffs then led to feline spongiform encephalopathy (FSE) in domestic & exotic cats; transmissible mink encephalopathy (TSE) in US Chronic wasting disease (CWD)—lateral transmission among Rocky Mountain elk & deer Human prion diseases: Creutzfeldt Jakob disease (CJD)—80% of all human prion diseases; affects 1:million persons (usually 60-65yo); 70% die w/in 6m Kuru—spread w/ cannibalism in Eastern Highland of Papua New Guinea; disease nearly gone except for previously exposed older tribe members; death w/in 9m New variant CJD (vCJD)—>120 teens & young adults (usually 20-25yo) in UK; homozygous for methionine at codon 129; death w/in 14m Transmission—corneal implants, contaminated surgical instruments, dura mater grafts, cadaveric pituitary-derived GH for CJD Cannibalism of brain of dead tribal member by women & children for Kuru Pathology—mutation of PrP gene (PRNP) leading to spontaneous conversion to PrPSc; not linked to infectious source ~10-20% human prion diseases inherited (AD) thru 20 different missense & insertion PRNP mutations (homozyogsity at codon 129) Deposition of amyloid plaques composed of insoluble aggregates of PrP (characteristic of Kuru & vCJD) Characteristic triad—neuronal vacuolation (“spongiform” appearance to grey matter), astroglial proliferation, & lack of inflammatory response Presentation—survival times short after symptoms appear; incubation period depends on inoculation site distance from CNS Scrapie—anxiousness & hypersensitivity, followed by intense pruritis & unsteady gait progressing to severe ataxia & recumbency in terminal phase CJD—rapidly progressive dementia accompanied by myoclonus; cerebellar ataxia & cortical blindness, periodic sharp wave complexes (PSWCs) on EEG Kuru—progressive cerebellar ataxia w/ dementia vCJD—resembles Kuru more than CJD Diagnosis—detection of PrPSc in brain tissue (posterior thalamic region or “pulvinar sign”) & tonsil biopsy (infection w/ lymphoreticular tissue) Presymptomatic testing—DNA sequencing of mutation in PNRP for affected families Prevention—recombinant GH; surgical instrument incineration Treatment—anti-PrP Ab inhibit prion replication & delay development of disease (especially w/in lymphoid tissue); branched polyamines; tricyclic derivatives Cysteine protease inhibitors (calpains); tetrapyrrole compounds; inhibitors of PrP structure Hepatitis—inflammation of liver; generally present w/ jaundice (icteric infection) & signs of hepatocyte damage (elevated serum transaminases) HAV—formerly known as infectious or short incubation hepatitis; ~40% acute hepatitis; 50% US adults show evidence of prior infection Structure—naked ssRNA; resistant to detergents, acid & high temperatures Transmission—fecal-oral (high concentration in feces 2w before onset of jaundice); food (shellfish concentrate virus from contaminated water) Pathology—enteric mucosa (enterovirus type 72) viremia spreads to liver lymphoid cell infiltration, parenchymal necrosis & Kupffer cell proliferation Presentation—immune-mediated liver damage; milder (anicteric) in children; 99% complete recovery (no chronic carriers); fulminant hepatitis rare (80% mortality) Symptoms (fever, anorexia, pain) occur after ~25d incubation period & increase 4-6d before onset of jaundice (iceteric phase) Diagnosis—presence of anti-HAV IgM in serum by ELISA or RIA Prevention—chlorine treatment of drinking water; passive prophylaxis w/ immune serum globulin (ISG) before or early in incubation (80-90% effective) HAV vaccine—2 formalin-inactivated viruses approved & recommended for high-risk groups Treatment—NONE, only supportive measures HBV—0.1-0.5% chronic carriers in US Structure—enveloped partially dsDNA w/ short ssDNA; HBcAg & HBeAg in core, HBsAg on envelope surface (aggregates in serum) Transmission—HBsAg found in most body fluids (only 0.0001mL blood needed to cause infection) & sex; mother-to-child results in chronic carrier state in child Replication—HBV enters hepatocyte uncoating of nucleocapsid completion of dsDNA genome by core enzymes delivery to nucleus transcription produces 4mRNAs transported into cytoplasm mRNAs translated core proteins assemble around largest mRNA DNA synthesized by reverse transcriptase w/in core RNA degraded as DNA synthesized core enveloped before (+) strand DNA completed → exocytosis Presentation—more severe & pronounced symptoms vs. HAV infection; cell-mediated immune lysis of infected hepatocytes Rash & arthritis—early symptoms after ~10w incubation period due to immune complex deposition (complement activation) Chronic HBV—presence of HBsAg in serum for >6-12m & absence of anti-HBsAg; HBsAg w/o HBeAg in serum signals minimal liver damage Survivors of fulminant infection rarely become chronically infected Hepatocellular carcinoma—10-300X risk for malignancy w/ chronic HBV infection Diagnosis—HBsAg, HBeAg & HBV in serum during acute infection 15 Anti-HBsAg & anti-HBeAg—protective & sign of disease resolution Anti-HBcAg—appears during acute infection (diagnostic for acute infection); NOT protective; associated w/ chronic carrier state Prevention—passive hepatitis B immune globulin (HBIG) prophylaxis & active recombinant HBsAg vaccine Newborns—combined passive HBIG in delivery room w/ active HBsAg vaccine in 3 doses over first 24m HBsAg vaccine—initially restricted to live or killed virus or HBsAg isolated from plasma of chronically infected individuals Heptavax—liscensed in 1981; purify HBsAg from plasma 3 steps to inactivate any remaining virus test for residual HBV infectivity Recombinant—HBsAg expressed in yeast (Recombivas HB, Engerix-B) MS controversy—NO evidence linking HBV vaccine to demyelinating disease; more studies currently underway Treatment—IFN- effective in minority of chronic infections; 3TC (lamivudine) inhibits reverse transcriptase; OR liver transplantation HCV—occurs in 5-10% transfusion recipients, 50% develop chronic infections Transmssion—blood; less thru sex or mother-to-child Presentation—milder, slower disease but usually chronic infection; viremia develops w/in 1-3w lasting 4-6m Chronic infections range from asymptomatic to cirrhosis w/ risk of hepatocellular carcinoma; less frequent fulminant infections Chronic HCV also associated w/ development of autoimmune disease; rate of progression varies (depending on coinfections or EtOH use) Diagnosis—anti-HCV Ab by ELISA (Ab often delayed or absent from serum); virion RNA in serum by PCR (susceptible to contamination) Preventation—NO vaccine available b/c no effective viral Ag identified & genetic diversity of HCV (quasispecies variants of HCV w/in individual) Treatment—IFN- + ribavirin for minority of patients; pegylated-IFN- slightly more effective; OR liver transplantation HDV—small circular ssRNA w/ coat of HBsAg Transmission—blood Replication—only in HBV-infected cells; binds & enters hepatocytes in same manner as HBV Co-infection—HBV & delta agent infections simultaneously; more self-limiting than HBV infection alone but causes 40% of fulminant hepatitis Superinfection—infection of delta agent in person w/ chronic HBV; more severe & rapid progression (fulminant infection & chronic HDV infection) HEV—15-25% fulminant HEV infections that are fatal in pregnant women Transmission—fecal-oral, NOT person-to-person Presentation—similar to HAV infections; NO chronic infections Non-A thru E HV—serological & epidemiological evidence for at least 1 other virus w/ blood-related transmission & 1 enteric transmission HFV, HGV & TT viruses isolated from individuals w/ hepatitis, but no evidence that viruses actually cause hepatitis Viral gastroenteritis—viral infection of surface epithelium of small intestine (stomach usually NOT affected, acid resistant) resulting in vomiting & diarrhea Rotavirus—leading cause of childhood diarrhea (mortality in developing countries); 3.5million cases annually in US Structure—reovirus (respiratory enteric orphan virus)—wheel-like appearance of 11 dsRNA segements, nonenveloped Outer capsid proteins—VP4 (hemaglutinin) & VP7 (glycoprotein) induce neutralizing Ab & determine serotype A-G (A clinically relevant) Reassortment—RNA segments can combine during coinfection to produce novel viral variants Transmission—fecal-oral Presentation—1-3d incubation period followed by vomiting (early in infection) & diarrhea Prevention—breast feeding provides passive protection; intestinal immune response PARTIALLY protective Vaccine—RotaShield rhesus rotavirus tetravalent vaccine approved in 1998 & withdrawn following year due to complications (bowel obstruction) Treatment—supportive (fluid & electrolyte replacement) Enteric adenovirus—adenovirus types 40 & 41 (members of subgenus F); naked dsDNA Presentation—7-10d incubation period followed by moderate symptoms like rotavirus (vomiting & fever, but protracted diarrhea ~10d) Norovirus & Sapovirus—Caliciviridae family; naked (+) ssRNA; most common cause of nonbacterial gastroenteritis in adults Transmission—contaminated food/water (1°) & person-to-person (2°) Presentation—1-2d incubation followed by vomiting (early) & diarrhea (~5d) Low infectious dose, prolonged asymptomatic shedding, environmental stability, strain diversity, & no lasting immunity aid endemic spread Astrovirus—star-shaped appearance; naked (+) ssRNA Transmissions—contaminated food Presentation—3-4d incubation period; most common in young children Enteroviruses—new enteroviruses numbered rather than named; humans only natural host Structure—picornviruses; naked (+) ssRNA; acid resistant & optimum T of 37°C; 4 groups based on serotyping & disease characteristics Polioviruses (3 serotypes); coxsackieviruses (groups A & B); echoviruses (enteric cytopathic human orphan viruses); enteroviruses Transmission—fecal-oral (virus stable at room T for several days & prolonged period of shedding in stool) Pathology—replication w/in cytoplasm w/ synthesis of large precursor polyprotein → multiple proteolytic cleavages to produce viral proteins Initial URI & GI tract infection → viremia → replication in virus-specific tissues Presentation—often asymptomatic; 2-10d incubation followed by symptoms due to lysis of infected cells & immune response Nonspecific febrile illness—fever, malaise, anorexia, headache, sore throat, myalgia, GI symptoms Aseptic meningitis or encephalitis; acute myocarditis or pericarditis; exanthems; muscle weakness & paralysis; conjuctivitis Disseminated infection of neonate—acquired perinatally from infected mother; encephalitis, myocarditis, hepatitis & shock; possibly fatal Poliovirus—infection of CNS (particularly motor neurons); 90-95% asymptomatic Abortive poliomyelitis (4-8%)—minor illness (URI, GI disturbances & influenza-like symptoms) w/o evidence of CNS invasion; complete recovery w/in week Nonparalytic poliomyelitis (1-2%)—initial symptoms of minor illness, aseptic meningitis (stiffness of neck, back or legs for 2-10d) followed by complete recovery Paralytic poliomyelitis (1-2%)—initial minor symptoms followed by paralysis 1-10d later (NOT always permanent, ↑risk w/ age, progression ends w/ fever ends) Spinal form—most common; often asymmetric; usually affecting legs Bulbar form—paralysis of muscle groups innervated by CN (affects breathing); high mortality rate Bulbospinal form—combination spinal & bulbar forms Post-polio syndrome—neuromuscular symptoms seen ~35y after recovery from acute paralytic poliomyelitis; present w/ fatigue, muscle weakness & pain Prevention—Advisory Committee on Immunization Practices recommended in July 1999 to shift all vaccines to IPV due to slight risk of OPV & worldwide decline in disease Poliovirus vaccines—contain all 3 serotypes; inactivated vaccine (Salk or IPV, 98% effective) & oral live, attenuated vaccine (Sabin or OPV, 95% effective) Vaccine-associated paralytic poliomyelitis—low incidence (1:2.4million doses); most common after 1st immunization in normal individuals w/ OPV Influenza—kills 20,000 annually; ~80% respiratory disease is viral Structure—orthomyxoviruses; A, B, C subtypes (A & B clinically relevant); enveloped, segmented (-) RNA 15 hemagglutination (H) antigen (3 in humans)—aid virus attachment 9 neuraminidase (N) antigens (2 in humans)—aid attachment & release of virus from cell Antigens that determine serotype & infectivity, reassort to create new strains 16 Transmission—respiratory droplets; antigenic shift & drift w/ type A, only drift for type B b/c no type B in animal populations (simultaneous infection unlikely) Pathology—only RNA virus to replicate in nucleus by stealing methyl caps of cellular mRNA & transported into nucleus Replication in ciliated respiratory epithelial cells → desquamation of mucous-secreting & ciliated cells → IFN & cell-mediated reponse Presentation—short incubation period of 2d followed by abrupt onset of URI symptoms (particularly dry nonproductive cough); LRI may cause pneumonia Systemic symptoms due to IFN & lymphokine response, followed by cell-mediate response; local symptoms due to epithelial cell damage Bacterial superinfection—due to suppression of IFN & cell-mediate response & induction of bacterial adhesion to epithelium; acute worsening of symptoms Prevention—monitor viruses in birds (migration b/w Australia & China), pigs & humans; anti-H Ab prevents infection (neutralizes virus) & anti-N Ab prevents viral spread Vaccine—2 type A & 1 type B strains in killed vaccine (70-85% effective) for high risk patients Treatment—amantadine or rimantadine administered early for type A ONLY; CNS side effects Neuraminidase inhibitors—prevent viral infection & spread if given early for types A & B; less side effects Parainfluenza—15-20% nonbacterial LRI in children; transient immunity ONLY Structure—paramyxoviruses; enveloped (-) ssRNA w/ N & H antigens (stable serotypes w/o drift or shift); 4 serotypes: HPIV-1—major cause of croup (laryngotracheitis) in children & mild URI in adults HPIV-2—less significant but associated w/ croup or mild URI/LRI HPIV-3—major cause of severe LRI in children; ~50% children exposed by 1yo HPIV-4—least common & associated w/ mild URI only Prevention—washing w/ soap & water; NO vaccine available Respiratory syncytial virus (RSV)—named derived from ability to cause cell fusion; most important viral agent in infant respiratory infection Structure—paramyxovirus; (-) ssRNA w/ no H or N antigens; G (attachment protein) & fusion (F) glycoproteins (initial fusion & syncytium formation); A & B subgroups Pathology—confined to respiratory epithelium w/ occasional viremia; necrosis of epithelial cells, mononuclear cell infiltration & plugging of small airways Presentation—short incubation of 1-4d followed by rhinitis progressing to bronchiolitis & pneumonitis (lung hyperexpansion, hypoxemia & hypercapnia); lasting up to 2w Rhinoviruses—common cold; replicates best at 33C (T in nasopharynx, suitable for URIs) Structure—picornviruses; naked (+) ssRNA; not acid resistant, optimum T of 33°C (nasopharynx); >100 serotypes; binds to ICAM-1 adhesion molecule Adenoviruses—100 serotypes, 49 affect humans (95% adults seropositive to types 2 & 5) Structure—naked icosahedral dsDNA Transmission—fecal-oral & respiratory droplets; stable in environment & affects all cell types (vector for gene therapy) Presentation—common cold or gastroenteritis; URIs & rare severe LRIs or hemorrhagic cystitis, conjunctivitis (swimming pools); acute respiratory disease in military recruits Type 7—inhaled deeply causing severe pneumonia; depending on dose & route of infection Persist in tonsils & adenoids w/ shedding for years after initial infection Prevention—live coated vaccine of serotypes 4 & 7 for military recruits Coronaviruses—5-10% common colds Structure—enveloped (+) ssRNA w/ club-shaped spikes projecting around envelop to give crown appearance SARS—most develop pneumonia (10-20% require mechanical ventilation); 813 deaths from 8,437 confirmed cases Structure—coronavirus; (+) ssRNA Presentation—2-7d incubation; high fever (>100.4°F), chills, headache, body ache, diarrhea (10-20%) progressing to dry nonproductive cough, hypoxia & pneumonia Diagnosis—RT-PCR or Ab detection Herpes—over 50 different herpesviruses affecting all species (transmission across species usually severe); 8 herpesviruses in humans Structure—enveloped, dsDNA; 3 subfamilies: —broad host range; short reproductive cycle; latent in neurons (e.g. HSV & HZV) —restricted host range; long reproductive cycle; enlarged infected cells (e.g. CMV, HHV-6 & HHV-7) —lymphotropic; long reproductive cycle; associated w/ cell proliferation (e.g. rhadinoviruses (HHV-8) & lymphocryptovirsuses (EBV)) Pathology—coordinate regulation of protein synthesis (, regulatory or immediate early proteins , replicative , structural), NOT early-late control Replication by early Cairns mode (immediate early) shifting to rolling circle (producing multiunit length concatemers) Virus fuses w/ plasma membrane or phagocytosed capsid enters cytoplasm DNA released into nucleus thru nucleopores DNA immediately transcribed by RNA polymerase (10% of mRNA reaches functional polyribsosomes for synthesis of protein) transcription of mRNAs proteins turn off protein synthesis & start mRNA synthesis (turn off protein synthesis) → mRNA processing → genome packaged into head mechanism → enveloped as released from nucleus (most remain highly cell associated) Presentation—after primary infection, virus not eliminated & may replicate at any time to cause recurrent disease (latency) Reactivation stimuli—sunlight, menstruation, emotional stress & trauma; often small, less severe, shorter duration or asymptomatic HSV-1 & HSV-2—>50% middle socioeconomic class have circulating Ab by 21yo; HSV-1 primarily oral (99%) & HSV-2 primarily genital (50-92%) Pathology—entry via lesion in mucous membrane or skin multiplication at inoculation site spreads to regional lymph node (symptoms develop) → infected cells enlarge (intranuclear inclusions & margination of chromatin) → lesions develop papular to vesicular → inflammatory cells invade underlying dermis (vesicular fluid becomes cloudy) → lesions crust over (↓infectivity) or ulcerate (in mouth & genitals) Presentation—primary illness more severe; worse in children, malnourished & immunocomprised (only 1-2% infected present w/ recognizable illness) Diagnosis—restriction endonuclease analysis of DNA purified from isolate for distinct fingerprint of HSV-1 or -2; PAGE to examine proteins made in infected cells CMV—70% adults seropositive; latency w/in leukocytes or endothelial cells Transmission—shed in saliva, semen, vaginal secretions & breast milk w/o clinical symptoms; blood transfusion (1-5%) & transplantation (60-80%) Presentation—most commonly asymptomatic mononucleosis Congenital symptomatic at birth—ONLY 1% of fetal infections; 20% die during infancy or suffer brain damage Congenital asymptomatic at birth—may develop hearing defects or impaired intelligence Transplant recipients (leukopenia & hepatitis); bone marrow recipients (pneumonitis); AIDS (25% developed CMV infection before HAART) Diagnosis—PCR, culture, serology or Ag detection w/ IFA Prevention—blood & transplant screening; prophylactic therapy for bone marrow & solid organ transplant patients Treatment—NONE in normal persons; antivirals drug therapy in immunocompromised Varicella-zoster virus (VZV or HHV-3)—varicella (chicken pox) & zoster (shingles) Transmission—probably respiratory droplets early in varicella; slight risk for vertical transmission (especially in 1 st trimester) Pathology—latency w/in sensory dorsal root ganglia; often reactivated after 50yo or immunocompromised to cause zoster Infection of conjunctivae or URI replication in lymph nodes primary viremia (by 4-6d) replication in liver & spleen secondary viremia infection of skin (vesicular rash, by 14d) Presentation—fever & pruritic rash (maculopapules vesicles pustules scabs in varying stages, localized to head & trunk, self-limiting) Complications—bacterial superinfection (group A strep); pneumonia; encephalitis or meningitis; Reye syndrome (aspirin in children) Congenital varicella—vertical transmission thru maternal viremia causing scarring, eye abnormalities & abnormal limbs Zoster—unilateral rash; complications of post-herpatic neuralgia (elderly), eye damage from CNV infection; encephalitis Ramsay-Hunt syndrome—reactiviation of zoster in geniculate ganglion of ear canal resulting in facial palsy, loss of taste Diagnosis—characteristic rash; immunofluorescent staining of Ag; culture; known exposure; serology; PCR to detect viral DNA 17 Prevention—passive immuniziation w/ varicella-zoster Ig (VZIG) before lesions appear; active vaccine Treatment—symptomatic therapy for varicella; antivirals for adult varicella, eldery zoster, immunocompromised or severe disease (IV acyclovir) Epstein-Barr virus (EBV)—2 strains that differ in genes that maintain latency (EBNAs); 90-95% worldwide population infected Transmission—intimate contact; saliva (low titer detectable for life w/ sporadic replication in oropharyngeal epithelium) Pathology—infection of epithelial cells of oropharynx spreads to B-cells (infects ~10%) in adjacent lymph nodes spread thru lymphoreticular system → cell-mediated response targets virus-infected B-cells → 1:million B-cells remain infected (latent) Presentation—most 1° infections asymptomatic; most severe during 2nd decade of life; children often asymptomatic (may develop IM or heterophile Ab) Infectious mononucleosis (IM)—usually self-limiting fever, sore throat, lymphadenopathy, malaise, splenomegaly Atypical lymphocytes in peripheral blood; heterophile Ab (IgM from polyclonal activation of EBV-infected B-cells that target RBCs) Complications—rash when treated w/ ampicillin; splenic rupture; encephalis & meningitis & myelitis Burkitt’s lymphoma—common childhood malignancy in malaria belt of Africa Post-transplant lymphoproliferative disorder—ranges from benign polyclonal B-cell activation (most common) to malignant B-cell lymphoma HIV-associated—non-Hodgkin’s lymphoma & oral hairy leukoplakia (wart-like disease of lingual squamous epithelium) Hodgkin’s disease—50% US cases contain EBV Nasopharyngeal carcinoma—common in southern China T-cell lymphoma—chronic EBV infections Multiple sclerosis—anti-EBV Ab titer high in individuals who later develop MS Diagnosis—presence of atypical lymphocytes; anti-EBV Ab in serum; heterophile Ab Treatment—supportive therapy ONLY HHV-6 (B lymphotropic virus, HBLV)—causes roseola infantum (exanthem subitum); 70% acquire HHV-6 during 1st year of life Structure—HHV-6A (often isolated from lymphoproliferative or AIDS patients) & HHV-6B (often isolated from patients w/ roseola infantum) Presentation—latency w/in CD4 cells, epithelial cells & possibly moncytes Roseola infantum—benign disease of children; fever that subsides w/ appearance of non-pruritic, slightly elevated, blanchable rash Liver dysfunction during acute phase; febrile seizures possible Presents as mild infectious mononucleosis in adults Transplant recipients—7-80% asymptomatic OR febrile syndrome, bone marrow suppression, encephalitis, GVHD? Diagnosis—HHV-6 detected in peripheral blood during febrile phase (PCR, culture, IFA, Western blot w/ HHV-6 Ab) Treatment—antivirals (gancyclovir & foscarnet) HHV-7—isolated from saliva w/ PCR on peripheral blood mononuclear cells, culture or IFA; unknown whether causes disease HHV-8—10-20% seropositive in US, up to 100% in endemic areas (Central Africa) Transmission—sex, needle sharing (not as efficient as w/ HIV or HBV), horizontal spread in endemic areas; organ transplantation Presentation—viremia strongly associated w/ Kaposi’s sarcoma (KS) development (HHV-8 detectable long before appearance of KS) KS—characterized by proliferation of spindle-shaped cells & irregular slit-like vascular channels; lymphocyte & M infiltration Non-AIDS-related KS—classic (elderly Mediterranean men); immunosuppression; endemic areas (mostly males) AIDS-related KS—very aggressive spread from skin to viscera Primary effusion lymphoma (PEL)—non-Hodgkin’s; no solid tumor but effusion into viscera Multicentric Castleman’s disease—atypical lymphoproliferative disorder w/ generalized lymphoid hyperplasia Diagnosis—PCR or Southern blot detection of viral DNA; ELISA or IFA on serum Treatment—radiation & chemotherapy; excision of localized tumors; IFN therapy; antivirals & HAART (lowers risk for KS & remission) Cercopithecene herpesvirus-1 (Simian herpesvirus B)—macaques are natural host w/ severe & fatal disease in humans Transmission—monkey bites or needle sticks; 1 documented person-to-person spread; airborne transmission among macaques Presentation—localized vesicular lesion at bite complete CNS involvement & encephalitis Diagnosis—contact w/ monkeys; PCR for viral DNA; culture Treatment—high dose IV acyclovir & high dose ganciclovir if neurological symptoms present Human retroviruses: HTLV-1—adult T-cell leukemia HTLV-2—variant of hairy T-cell leukemia HTLV-3—HIV-1 & HIV-2 HTLV-4—related to simian retrovirus HIV presentation—CDC classification: Stage I (initial infection)—mononucleosis-like illness (fever, adenopathy, rash, pharyngitis, arthralgia & myalgia, thrombocytopenia) Stage II (chronic asymptomatic infection)—lab abnormalities in CD4, CD8 & WBC counts Stage III (persistent generalized lymphadenopathy)—palpable lymph nodes w/o 2° conditions Stage IV (AIDS)—constitutional disease, neurological disorders, 2° infectious disease & cancers Recurrent pneumonia, progressive multifocal leukoencephalopathy, Wasting syndrome, cervical cancer, retinitis, lymphomas; KS; sinusitis; dementia Sore mouth & trouble swallowing (Candida); shortness of breath & nonproductive cough (pneumocystis carinii pneumonia); Headache & confusion (CD4 + <200) CNS toxoplasmosis or cryptococcosis; fever (CD4+ <100); mycobacterium infection; rash; viral infections Lipodystrophy—centripetal fat accumulation & buffalo hump, sunken cheeks & temporal wasting; ↑breast size; prominent veins in extremities Psychiatric problems—memory & concentration impairment, confusion, depression, anxiety, motor abnormalities; psychoses or hallucinations or mania Prevention—HIV testing of persons in risk group, STD, recurrent TB or pneumonia, abnormal pap, squamous cell carcinoma, recurrent Candida infection, PID, pregnant May take weeks to months to be positive following exposure; 2 positive ELISA tests w/ confirmatory Western blot (3 weeks) HIV vaccine—difficulty to develop b/c state of protective immunity unknown in humans; high mutation rate; virus latency; lack of animal model for testing Treatment—failure due to nonadherence, suppressive drugs regimens, high baseline viral load, syncytium-inducing phenotype, poor drug penetration Disease monitoring—viral load (measure of HIV RNA in plasma) & CD4+ cell count (measure of immune status) Adherence—education, simplification of drug regimens, medication reminders, reinforcement of success, coping w/ side effects, social support Persistence—slow release of viral progency w/o cell death or destruction (permissive & non-lytic); associated w/ defective host immunity Portal of entry—NOT always same as site of persistence (e.g. VZV entry into respiratory tract & persistence in dorsal root ganglia) 1. Latent infection—persists in non-infectious form following acute disease episode w/ recurrence similar to acute infection (e.g. herpesviruses) 2. Persistent & slow progressive infection that follows acute & recovery phase (e.g. measles) 3. Typical persistent infection never goes through early acute episode & recovery (e.g. CJD) Persistent diseases—can be reactivated; associated w/ chronic disease & cancers; keep virus persistent in host community Latency—viral genomes usually resident as DNA forms in extrachromosomal (e.g. herpesvirus) or integrated w/ host chromosome (e.g. retroviruses) & replicate w/ cell duplication Immune modulation model—ganglion cells permissive for HSV replication; Ab production produces non-lytic latent infection after primary site cleared of virus 18 Immune elimination model—two populations of ganglion cells, non-permissive cells establish latent infection, permissive cells establish productive infection Non-permissive model—all ganglion cells non-permissive; replication (directly) or inflammation (indirectly) provides signal that converts ganglion cells to permissive state Host immune response eventually shuts down signal Reactivation—resumption of viral activity (phase A) & replication & spread of virus (phase B); travels down sensory axon to epithelium & mucosa Sensory prodrome—phase A happens more frequently than phase B, so some fraction of reactivations are non-lesional May indicate replication in sensory neurons (appearance of EBV & CMV in saliva & blood) Human papillomavirus (HPV)—most common viral STD (20-60% adult women seropostive in US) Structure—unenveloped dsDNA; 50 genotypes (12 causing genital warts) Presentation—epidermal papillomas & warts; associated w/ cervical & anal cancers Treatment—destroy & remove infected epithelium (recurrence common); IFN therapy Rabies—recognized 3000yo; Aristotle noted that could be spread by rabid dog; up to 50,000 deaths in India each year, <5 cases annually in US Structure—bullet-shaped, enveloped (-) ssRNA 2 forms—urban (from unimmunized dogs & cats) & sylvatic (from wild animals, especially bats) Transmission—animal bite or inhalation of contaminated material (e.g. bat droppings) Pathology—incubation period from 10d-1y depending on dose, host immunity, & distance from site of inoculation to CNS Entry thru dermis replication in local striated muscle enters PNS at NMJ spreads up axons to CNS replicates exclusively w/in gray matter spreads back thru ANS to other tissues (e.g. salivary glands) Presentation—initially fever, headache, malaise & GI upset progressing to encephalitis, coma & respiratory paralysis; death w/in 4d (ONLY 6 human survivors) Excessive motor activity & spasms; hallucinations; combativeness; meningeal irritation; seizures; focal or ascending paralysis Salivation—infection w/in salivary glands via ANS & difficulty swallowing via CN involvement (also facial palsy & double vision) Hydrophobia—painful, violent involuntary diaphragm contracts when swallowing liquid; seen in 50% cases Diagnosis—characteristic brain lesions w/ Negri bodies (eosinophilic cytoplasmic inclusions) Prevention—preexposure prophylaxis for risk groups (14,000 per year in US); postexposure before appearance of symptoms (40,000 per year) Postexposure—immediate washing w/ soap & water, passive hyperimmune globulin (half w/in wound site) & active antirabies vaccine (5 doses) Treatment—NONE (mortality >90%) Arenaviruses—enveloped, ssRNA (-) sense segmented; contain host ribosomes inside (granular appear or arenosus “sandy”); e.g. hemorrhagic fevers Filoviruses—enveloped, ssRNA (-) sense; e.g. Marburg & Ebola (30-80% mortality) Arbovirus—togaviruses, flaviviruses, bunyaviruses & reoviruses Transmission—reservoirs in non-humans w/ spread following extrinisic incubation period (virus multiplication in vector) Viremia—in humans aids transmission by reinfecting vectors; birds often hosts b/c longer viremia Overwintering—survival due to sustained viremia in lower vertebrates; hibernation of infected vectors; transovarial transmission among vectors 3 types of viral transmission—depending of cycle of hosts Urban—large # humans in close proximity to arthropod allows direct transmission to humans & reinfection of vector Sylvatic—single non-human reservoir w/ humans usually dead-end host Arthropod-sustained—transovarial transmission w/in arthropod Pathology—replication w/in reticuloendothelial system & vascular endothelium causing endothelial cell swelling & edema w/ mononuclear cell infiltration Severe infection causes perivascular edema & serosal effusions; virus-Ab complexes activate complement & vascular permeability leads to hemorrhage Togaviruses & flaviviruses—most arthropod-borne viruses; enveloped (+) ssRNA Eastern equine encephalitis—ranges from mild febrile illness to meningitis & severe encephalitis (20% mortality) Dengue—hemorrhagic fever thru destruction of endothelium & thrombocytopenia; erythematous rash, severe pain in back, head, muscles & joints Bunyaviruses—enveloped (-) ssRNA; causing fulminant respiratory disease (>60% mortality) Hantavirus—or Korean hemorrhagic fever (KHF); 3 segments resorted to create new strains (>50% mortality) Transmission—thru rodents (deer mice in SW US), inhalation of excrement, conjuctiva or direct contact Presentation—initially no rash or edema, progressing to bilateral diffuse interstitial pulmonary edema & pneumonia Treatment—supportive ONLY w/ aggressive respiratory support, ribavarin (Dengue virus); prevention of bites & vector control Mumps—affects primarily 5-15yo; 30-40% asymptomatic Structure—paramyxovirus; enveloped (-) ssRNA w/ H & N antigens Pathology—entry in respiratory tract → local replication → viremia & spread to salivary glands, CNS, & kidneys (viruria & renal impairment common) Presentation—2-3w incubation period followed by fever & swelling of salivary glands (especially parotids) Complications—meningitis (10%), encephalitis, myelitis, pancreatitis, orchitis (10-20% infected men), severe CNS infection (hearing loss) Diagnosis—culture from saliva, throat swab or CSF; ELISA for IgM & IgG Prevention—1968 live, attenuated vaccine (95% effective) given in first year of life Treatment—NO specific therapy Measles—5 –day measles; passive immunity in <6mo prevents infection; 1millions death annually from pneumonia Structure—paramyxovirus; enveloped (-) ssRNA; glycoproteins M (viral assembly), H (attachment) & F (fusion), no N antigen Pathology—entry in respiratory tract → replication in respiratory mucosal epithelium (disrupts cytoskeleton) → viremia w/ infection of lymphocytes & monocytes Vasculitis, giant cell formation w/ inclusions, damage to neurons & cells thru CD8+ cells Infection of lymphocytes causes susceptibility to other bacterial infections (↓NK cells, ↓ROS, ↓Ig synthesis) Presentation—7-18d incubation followed by cough, coryza & conjunctivitis, fever; rash (papular & semiconfluent) starts on head & spreads down trunk to extremities Koplik’s spots—white spots w/ surrounding erythema on mucous membranes, 1-3d after onset of symptoms Bacterial superinfection (5-15%); pneumonia & sepsis; encephalitis (15% mortality); thrombocytopenic purpura; abdominal pain & acute appendicitis Subacute sclerosing panencephalitis—progressive neurologic disease from chronic measles infection in children (1:100,000); presents 2-10y after infection Prevention—live, attenuated vaccine highly immunogenic, not recommended for pregnant women & immunocompromised; passive ISG w/in 6d of exposure Treatment—supportive therapy ONLY Rubella—German or 3-day measles; mild exanthem of children, profound BD in fetus Structure—togavirus; enveloped (+) ssRNA Pathology—URI w/ viremic spread; cell-mediated immunity mediates arthritis & rash; fetal infection chronic in first trimester (20-30% risk) due to chromosomal breaks Presentation—only 30-60% develop symptoms; 16d incubation followed by mild fever, URI & lymphadenopathy, macular rash (faint over head, neck & trunk), arthralgia Congenital rubella syndrome (CRS)—cardiac defects, eye defects, hepatosplenomegaly, thrombocytopenia, microcephaly, MR Prevention—1969 live, attenuated vaccine (95% effective); not recommended for pregnant women Parvovirus B19—commonly presents as erythema infectiosum (fifth disease or academy rash) Structure—small naked ssDNA; only replicates in dividing cells Pathology—replication w/in RBC precursor nuclei 19 Presentation—mild fever, malaise, headache, myalgia & confluent rash on fash (“slapped cheeks”) spreading to exposed surfaces; arthralgia Transient aplastic crisis—acute, severe anemia in patients w/ chronic hemolytic disease IID EX AM 3 Bacteria (prokaryotes)—genus (first of two names) always capitalized & species never capitalized 2 major subdivisions—archaebacteria (thrive in extreme environmental conditions) & eubacteria (“true bacteria,” most human pathogens) Differences vs. eukaryotes—heterotrophs (rely on preformed organic compounds as nutrients) Nucleus—nucleoid lacks nuclear membrane; single (haploid), covalently-closed, supercoiled dsDNA chromosome; NO histones, NO introns Cytoplasm—NO organelles; ribosomes smaller Ribosomes—70S composed of 30S & 50S (16S, 23S & 5S rRNAs); assembled into polysomes Plasmids—extrachromosomal, self-replicating, circular dsDNA; usually multicopy; transmissible or nontransmissible Cytoplasmic membrane—NO sterols (except Mycoplasma) Mesosomes—invaginations of membrane that form divisional walls & associates w/ nucleoid during replication Cell wall—stress bearing & protects from osmotic lysis (except Mycoplasma); essential for growth & division; nonselectively permeable; site of Abx action Unicellular—free living (except Chlamydia & Rickettsia) Cell size—10-100X smaller (average 1m length); high SA:V ratio (diffusion of nutrients) Division—asexually by binary fission, simpler & exponential every 15-20min (vs. 1/day); transcription/translation coupled; polycistronic mRNAs Capsule—hydrophilic gel surrounding cell wall; also slime layer (glycocalyx); role in adherence, protection against clearance, identification thru Ag properties Flagella—motility & chemotaxis; long spiral shaft composed of flagellin forms major surface antigen (H-Ag, located at poles (peritrichously) Hook (universal joint) & basal body (associated w/ cell envelope)—anchors flagellum & produces rotational force driven by proton-motive force Pili—hair-like projections from cell membrane w/ no apparent anchor; pilin protein arranged helically to form cylinder that mediates attachment Common pili (fimbriae)—cover all of bacterial surface for adherence Sex pili—thicker & only 1-6/cell for attachment b/w mating bacteria & transfer of DNA from male bacteria to female (conjugation) Endospores—metabolically-inactive bacterial (Bacillus & Clostridium) cells, highly resistant to extreme environmental conditions, dormant in soil Formed w/in mother cell w/ complete genome in core, thick wall & thick keratin-like coat; liberated when mother cell autolyses Biofilms—heterogenous mixture of microorganisms & extracelullar material; provides nutrients & protection; difficult to eradicate (persistent & resistant) Dental plaques—tooth covered by saliva provides film for organic polymers (e.g. Strep. mutans); 2° colonizers mostly anaerobes Classification—according to cell wall, morphology, or specific characteristics: Cell wall—Gram-staining (+) purple vs. (-) red Poor-staining—acid fast (e.g. Mycobacterium); no cell wall (e.g. Mycoplasma, L-forms); obligate intracellular (e.g. Chlamydia, Rickettsia); spirochetes Morphology—cocci (spherical), bacilli (rods), spiral OR pleomorphic Cocci—diplococci (e.g. S. pneumoniae, Neisseria), chains (e.g. Strep), & clusters (e.g. Staph) Bacilli—round ends (e.g. E. coli), square end chains (e.g. Bacillus), shorter (e.g. coccobacilli), curved (e.g. Vibrio) Club-shaped (e.g. Corynebacterium), tapered ends (e.g. Fusobacterium) Spiral—rigid wall (e.g. Spirilla) Specifics—physiologic (endospore formation, O2 requirements, motility); colony morphology (pigment, hemolytic activity); biochemical (catalase or oxidase) Cell wall—b/w cytoplasmic membrane & capsule (except Mycoplasma); activates alternative complement pathway, especially Gram(+) w/ adjuvant properties Murein—universal peptidoglycan of cell wall; repeating, alternating N-acetylglucosamine (NAG) & N-acetylmuramic acid (NAM) NAM not present in humans; chains crosslinked by tetrapeptides of alternating D & L amino acids attached to NAM & crosslinked to each other D-AAs confer resistance to human peptidases Gram(+)—thick w/ 40 layers PG (up to 50% teichoic acids) linked to NAM Membrane teichoic acid (lipoteichoic acid, LTA)—covalently linked to membrane glycolipid; forming major surface Ag & taxonomic markers Functions—binds Mg2+, impermeable to hydrophobic compounds, anchors cell wall, regulates autolysin of complement Teichuronic acids—repeating sugar acids vs. phosphoric acids (teichoic acids); synthesized when phosphate or Mg2+ limited Gram(-)—characteristic outer membrane; ONLY 1-3 layers PG covalently linked to outer membrane by lipoproteins (Braun lipoprotein) Periplasmic space—PG layer & gel-like proteinaceous solution Outer membrane—bilayered; inner leaflet of phospholipids & outer leaflet of LPS; forms impermeable barrier to all compounds LPS—composed of lipid A + core polysaccharide + O Ag polysaccharide Lipid A—phosphorylated glucosamine dissacharide attached to long-chain FA; endotoxin that causes fever & septic shock Polysaccharide tail—repeating linear trisaccharides; forms major surface Ag (O-Ag) Porins—passive diffusion of low-MW compounds; large Abx penetrate slowly (high resistance) VS. transport proteins—energy-requiring coupled translocation Bayer junctions—adhesions b/w cytoplasmic membrane & outer membrane Acid-fast—thinner PG layer w/ high lipid & wax content Mycolic acids—complex branched, long FA chains bound to PG by arabinogalactan; forms lipoarabinomannan (LAM) Cord factor—glycolipid of trehalose & mycolic acid; that forms “serpentine cords” Impermeable to hydrophilic compounds; resistant to dehydration, acids, alkali, many disinfectants & antibiotics; multiply intracellular Bacterial growth—heterotrophs Nutrients—energy (ATP & proton-motive force); C, N, S, P; inorganic elements; GFs Oxygen—grouped into 5 classes: Obligate aerobes—requires O2 (e.g. Mycobacterium, Neisseria, Pseudomonas); SOD & catalse(+) Obligate (strict) anaerobes—only in absence of O2 (e.g. Clostridium); NO enzymes to detoxify ROS Facultative anaerobes—presence OR absence (e.g. most medically important bacteria); SOD & catalase(+) Aerotolerant anaerobes—w/ or w/o O2, but exclusively anaerobic (fermentation) metabolism (e.g. many Strep); SOD & peroxidase(+) Microaerophiles—grow best at low O2 (e.g. Helicobacter); small amounts of SOD & catalase(+) ROS—cellular enzymes detoxify, e.g. SOD (2O2. + 2H+ → H2O2 + O2) & catalase/peroxidase (H2O2 + AH2 → 2H2O + A) Temperature—psychrophiles grow best at low T (10-20°C); mesophiles at room T (20-40°C); thermophiles at higher T (55-75°C) pH—acidophiles grow best at pH<6.0; neutrophiles between 6.0-8.0, best at physiological pH (7.2-7.6); alkaliphiles at pH>8.0 Growth cycle: 1. Lag phase—metabolite-depleted cells adapt to new environment; little or NO cell division 2. Exponential phase—balanced growth; binary fission at constant rate; most Abx effective against metabolically active, rapidly growing cells Generation (doubling) time (t1/2)—time required for bacterial population to double in # or mass; depends on strain, nutrient supply & environment 20 3. Stationary phase—nutrients limiting & toxic products accumulate; NO net growth (balance b/w growth & death) metabolic activity & cell size, resistance Sporulation—Bacillus & Clostridium 4. Death phase—NOT always observed; metabolic activity not meeting cellular maintenance activation of autolytic enzymes Bacterial identification: Culturing—liquid or solid agar VS. complex (undefined) tissue digest & yeast extract VS. synthetic medium; most grown at 35°C & 5% CO 2 or anaerobic conditions Turbidity—growth (viable & nonviable cells) in liquid culture determined by optical density (OD) Cell concentration—# viable cells or colony forming units (CFUs) per volume of culture w/ serial dilutions of liquid culture & cultivation on solid medium Supportive—ALL bacteria except most fastidious (e.g. blood agar) Enriched supportive—including most fastidious (e.g. heated whole blood or “chocolate agar”; additional nutrients for Haemophilus & detoxifies for Neisseria) Selective—restricts growth of some bacteria (e.g. MacConkey agar selects for Gram(-)) Differential (indicator)—reveals specific characteristics of bacteria (e.g. MacConkey agar determines lactose fermentation properties of Gram(-)) Staining—Gram stain (most bacteria, sputum smears); acid fast (Mycobacterium); methylene blue (Corynebacterium) Fluorescent stains (Mycobacterium, Chlamydia, Legionella & Treponema) Microscopy—light (brightfield) for Gram or AF; fluorescent; darkfield for spirochetes (not very sensitive) Immunologic—detect antigen material w/in specimen (e.g. Group A Strep from throat swabs) Serodiagnostic—detect Ab w/in serum; Ag on plastic substrate washed w/ patient serum w/ some visible label DNA probe—detect specific bacteria or non-culturable bacteria (PCR & sequencing) Antibiotic susceptibility tests: Diffusion method—measures zone of inhibition around disc containing Abx; compare zone size to standard test organism Dilution method—automated process of serial dilution of antibiotic; endpoint is amount of drug needed to inhibit or kill bacteria (minimal inhibitory concentration) E-test—combination of disc diffusion & dilution method w/ strips of serial Abx concentration placed on inoculated agar Metabolism—NAD+ & ATP major carriers of energy CHO catabolism—1st step of both fermentation & respiration Emden-Meyerhof-Parnas (EMP) or glycolytic pathway—oxidation of glucose to pyruvate HMS—provides NADPH & pentoses Entner-Douderoff (ED) pathway—exclusive to bacteria, alternative pathway for glucose oxidation w/ lower energy yield Fermentation—incomplete oxidation; terminal electron acceptor is organic compound (organic acids, alcohols, ketones, gases) NAD+ reduced to NADH reoxidized to NAD+ by reducing pyruvate; substrate level phosphorylation forms 1-3ATP Respiration—complete oxidation, inorganic compound terminal electron acceptor (O2 for aerobic VS. nitrate, sulfate, carbonate, fumarate for anaerobic respiration) Electron transport chain generates proton-motive force for ATP synthesis, transport & motility; oxidative phosphorylation forms 38ATP Complete oxidation to CO2 (or NO2- & N2) & H2O Chemotaxis—find nutrients & avoid toxic compounds thru proton-motive force; net movement based on regulation of tumbling (longer swimming towards chemoattractant) “Biased random walk”—swimming (counterclockwise) & tumbling (clockwise) toward chemoattract (AA or sugar) or away from chemorepellent (EtOH) Exoenzymes—degrade complex CHO or nucleic acids & proteins for transport across membranes Iron acquisition—usually active transport systems synthesized in response to low Fe conditions to retrieve from host Direct uptake from host iron complexes, cell bound Fe-uptake proteins (e.g. OM receptors), extracellular chelated-mediated uptake (e.g. siderophores) Active transport—mediated by membrane carrier proteins (permeases); some Abx actively transported (e.g. tetracycline) Group translocation—unique to bacteria; nutrient chemically modified as crosses membrane & energy released drives own transport Phosphotransferase (PTS) systems—obligate & facultative anaerobes; transport & phosphorylate sugars coupled w/ fermentation metabolism Normal flora—acquired by normal contact w/ environment; composition depends on body site, age, diet & secretions; potential pathogens; some transient Highest concentration on moist skin, colon, pharynx, vagina; some in small intestine, distal cm of urethra, conjunctivae; sterile blood, CSF, urine, internal organs, synovium Benefits—prime immune system (CK production, phagocytes, maturation of GALT & IgA plasma cells), maintains integrity of GI mucosa; control inflammation Digestion/absorption; suppression of food allergies; exclusionary effect (competes w/ other bacteria for nutrients); produces vitamins B & K Probiotics—normal flora used to treat GI disease in 109-1010 doses; must have beneficial effect, human strain, safe, stable in acid & bile, adhere to intestinal mucosa Types—most common Lactobacillus acidophilus (yogurt culture), Bifidobacterium bifidum, Strep thermophilus Uses—diarrhea (kids rotavirus, traveler’s diarrhea, enteritis, post- or long-term antibiotic use); lactose intolerance; food allergies GI inflammation (disruption of mucosal barrier w/ Crohn’s disease), atopic eczema, food allergies Potentially for inflammatory bowel disease, cancer prevention, immunomodulations (arthritis), respiratory disease (e.g. pneumonia w/ CF) Negative effects—imbalance affects mucosal barrier, immunocompromised, disease in other body sites, quorum sensing molecules; gingivitis/caries Halitosis (volatile sulfur compounds, HS, methyl marcaptan, dimethyl sulfide) Parasites—one-sided benefit (disease-causing) VS. commensal—neutral, non-damaging relationship VS. mutualistic—reciprocally beneficial Primary pathogen—seriousness of disease depends on first-time infection, survival/transmission aided by symptoms, immunocompromised, virulence, resistance D = nV/R (seriousness of disease proportional to # invading bacteria & virulence, inversely proportional to host resistance Infectious dose—# bacteria needed to cause infection Facultative intracellular—grow both inside & outside host cell (e.g. Mycobacterium, Legionella) Obligate intracellular—grow ONLY inside host cells (e.g. Chlamydia, Rickettsia) Pathogenesis—gain access to host → find/create unique niche → evade host immunity → multiply → disease (toxin production & tissue damage) → transmission Attachment—counteracts mechanical clearance mechanisms Loose attachment to mucus (common or type 1 pili) VS. specific, tight attachment (special pili, OMP, surface proteins, glycoprotein receptors) Establishing niche: Extracellular parasites—contact-dependent injection of bacterial toxins to block phagocytosis Intracellular parasites—parasite-induced uptake through contact-dependent injection of toxins to induce phagocytosis Toxins—often proteins, usually secreted on surface or into medium or into cytoplasm (contact-dependent) Local tissue damage allows for new sites of attachment & further spread of infection Evading host immunity: Serum resistance—evade complement Bind or inactivate Ab—block opsonization (complement-mediated lysis) OR cleavage of IgA1 Degradation of chemical defenses—catalase & SOD Antiphagocytosis—surface structures (capsules) inhibit phagocytosis OR contact-dependent export of anti-phagocytotic effects into host cell EPEC induce formation of pedestals on intestinal epithelial cells that prevent uptake of bacteria into cell; extracelullar parasites Kill phagocytes—OR escape from phagosome into cytoplasm OR inhibit maturation of phagocytes Contact-dependent secretion of toxin into phagocyte, pore-forming toxins or exotoxins Intracellular—silent entry (w/ complement receptor & NO oxidative burst) Evasion of lysosomes (fusion w/ phagosome), tolerate lysosome environment, escape into cytoplasm 21 Antigenic disguise—bind host compounds to bacterial surface to appear as nonforeign Antigenic mimicry—resemble components in tissue; e.g. sialic acid or gangliosides on bacterial surfaces; extracelullar parasites Antigenic variation & phase variation (turning on/off protein synthesis) Immunosuppression—down-regulation of immune response, inhibition of CK expression, induced apotosis Multiplication—toxins alter cells or release nutrients; iron-acquisition (siderophores, cell-bound systems); exoenzymes (nutrition & infection spread) Damage—toxins, metabolic byproducts, host response (inflammation, autoimmunity, deposition of immune complexes) Transmission—reservoirs; adaptation to insect vectors Hardiness—resistance to desiccation, UV, pH extremes; promotes transmission by fomites & dust Toxins or metabolic byproducts produce symptoms that aids spread (e.g. respiratory droplets, exudates, diarrhea) Exploitation of human habits—rubbing face, handshaking, sex Antibiotics—bactericidal (kills bacterial cells) VS. bacteriostatic (inhibits growth); classified according to target area: Cell membrane function: Polymyxin B—cationic detergents that damages membrane containing phosphatidylethanolamine, mainly Gram(-) cell leakage Growing OR resting cells; toxic to kidneys & CNS (used topically) Gram cell wall synthesis: PG synthesis—synthesis of UDP-NAM-pentapeptide from UDP-NAM in cytoplasm +AA to form UDP-NAM-tripeptide +D-Ala-D-Ala (2L-Ala → 2D-Ala (by racemase) → D-Ala-D-Ala (by dipeptide synthetase)) to form UDP-NAM-pentpeptide translocation of PG to exterior transglycosylation (enzymatic -1,4 linkage of NAM to NAG & release of bactroprenol-PP) on membrane bactoprenol-PP converted to bactoprenol-P (by membrane-bound phosphatase) & cycled back to cytoplasmic side of membrane transpeptidation (enzymatic cross-linking of peptides & release of terminal D-Ala) to form NAM-NAG-tetrapeptide Species-specific modifications—addition of teichoic acid (TA) or Gly5 bridge Lysozyme—autolysin that hydrolyzes -1,4 linkages b/w NAM & NAG; found in phagocytes of innate immune response Fosfomycin—broad spectrum & widely distributed; analog of PEP that inhibits production of UDP-NAM from UDP-NAG; single dose Tx for UTIs Cycloserine—analog of D-Ala that inhibits formation of D-Ala-D-Ala; widely distributed; 2nd line Tx for TB Vancomycin & teicoplanin—block transglycosylation by binding D-Ala-D-Ala; most effective against Gram(+) Bacitracin—inhibits dephosphorylation of bactropenol-PP to bactoprenol-P; can’t penetrate OM of Gram(-), but effective against Gram(+); toxic (topical use) -lactam antibiotics—narrow to broad; low toxicity; effective against growing, walled-bacteria ONLY Structural analog of D-Ala-D-Ala that binds penicillin-binding proteins (PBPs) & inhibit transpeptidation & activates autolysin Clavulanic acid (clavam)—weak OR “suicidal” -lactamase inhibitor Wall-less (L forms)—able to grow & remake PGs forming persistent or chronic infections (difficult to clear) Lysozyme or PCN—produce Gram(+) protoplasts OR Gram(-) spheroplasts Mycobacterium cell wall synthesis—most drugs specific to M. tuberculosis, used in combination therapies Isoniazid—inhibits mycolic acid synthesis; even penetrates w/in M Ethambutol—bacteriostatic; inhibits synthesis of arabinogalactan & LAM Pyrazinamide—inhibits lipid metabolism prevents mycolic acid synthesis Folic acid synthesis—from PABA, glutamate & pteridine; PABA converted to DHF (by dihydropteroate synthesis) reduced to THF (by dihydrofolate reductase) Humans must obtain DHF or THF from diet (NO DHF-synthetase)—synthetic PABA derivates against dihydropteroate synthetase NOT toxic to humans Trimethoprim—selectively inhibits bacterial enzyme that reduces DHF to THF; Tx for UTIs b/c highly concentrated in urine Sulfonamides (sulfa drugs)—analogs of PABA that competively inhibit dihydropteroate synthetase; concentration must be > PABA Bacteriostatic & broad spectrum; well distributed; Tx for meningitis, otitis media & conjunctivitis, concentrated in urine p-aminosalicylic acid (PAS)—combination Tx for Mycobacterium TB Dapson—combination Tx for M. leprae Trimethoprin—bacteriostatic & broad spectrum; synthetic pyrimidine analog that competitively inhibits dihydrofolate reductase; concentrated in urine DNA replication & repair: Replication—begins at origin of replication (oriC) & proceeds bidirectionally (5’→3’) till fork meets terminus DNA gyrase (topoisomerase II)—tetramer of A2B2; Topo I relaxes coils, Topo 2 maintains supercoils, Topo 3 & 4 unlink daughter chromosomes Quinolones (nalidixic acid) & fluoroquinolones—binds (cross-links gyrase & DNA) & inhibits gyrase subunit A after double-stranded break in DNA Narrow spectrum (1° generation) to broad (3-4° generations); well distributed; restricted for UTIs Fluoroquinolones—broader spectrum; binds several targets on A subunit (slower resistance development); anaerobes generally resistant Novobiocin—inhibits B subunit, bacteriostatic w/ little clinical use Nitroimidazoles (metronidazole)—reduction of nitro group in DNA itself causing strand breakage; effective against anaerobes ONLY b/c requires reduction RNA synthesis: Transcription—RNAP holoenzyme ( subunit) binds promoter to form closed complex → DNA unwinds to form transcription bubble (open complex) → polymerization 5’-3’ → DNA rewound behind RNAP → subunit released after nascent transcript 8-9 nucleotides long Bacterial mRNA—NOT transported through nuclear membrane, NO poly(A) tail, NO introns, coupled transcription/translation, short half-life Rifamycins—bind subunit of RNAP (RNA polymerase) to inhibit transcription initiation Rifampin—well distributed; against most Gram(+) & front-line TB drug, in combination to prevent resistance Rifabutin—against M. avium-intracellular complex (MAC) & TB in immunocompromised Protein synthesis: Translation—16S rRNA of 30S subunit binds Shine Dalgarno sequence of mRNA → initiator tRNA (fMet-tRNA in bacteria) binds start codon → 50S subunit binds to form initiation complex → peptidyl transferase (part of 50S) makes peptide bond → EF-G translocates ribosome by 1 codon w/ GTP peptide shifts from A site (aminoacyl) to P (peptidyl) → dissociates when reach stop codon Tetracycline—binds reversibly to 30S to prevent initiation; inhibits human ribomomes also at high concentrations; bacteriostatic Some bacteria have active transport mechanisms; discolors teeth in <8yo; well distributed; broad spectrum for wall-less & some obligate intracellular Aminoglycosides (e.g. streptomycin)—binds irreversibly to 30S to form “dead” or inactive 70S ribosome (NOT human 80S) Gentamicin, amikacin, & tobramycin—bind 50S & 30S subunits, increasing spectrum & decreasing development of resistance Actively transported by bacteria w/ oxidative phosophorylation; effective against aerobic Gram(-) bacilli, NOT strict anaerobes Streptomycin is front-line TB & plague; neomycin very toxic (oral & topical use ONLY) Linezolid (oxazolidinones)—bind 50S; effect against most Gram(+) cocci & methicillin-resistant staph; well-distributed Tx for VRE infections, community & nosocomial acquired pneumonia, complicated skin infections Chloramphenicol—binds reversibly to peptidyltransferase of 50S; broad spectrum, well distributed Bone marrow toxicity (gray baby syndrome w/ aplastic anemia b/c immature liver cannot clear drug); Tx for serious infections ONLY Macrolides (e.g. erythromycin)—blocks translocation of peptide chain; bacteriostatic, broad spectrum & well-distributed; Tx for Legionnaire’s & Mycoplasmas Lincosamides (e.g. clindamycin)—inhibits peptidyl transferase; bacteriostatic; Tx serious anaerobic Gram(-) infections; side effect of diarrhea 22 Streptogramins (e.g. Synercid)—mixture of group A & B antibiotics; binds irreversibly to 50S Combination strongly bacteriostatic; synergy due to conformational change in ribosome; effective against Gram(+) cocci; well-distributed except CSF Mupirocin—analog of isoleucine that inhibits isoleucyl-tRNA synthetase; topical Tx for Strep & Staph Fusidic acid—binds EF-G but inhibits 80S ribosome also (toxic); Tx ONLY in severe & drug-resistant Staph infections Antibiotic resistance—mutation of intrinsic genes or acquisition of novel genes; over 53% of ALL pneumococci now resistant to macrolides Excess use—broad-spectrum, prophylactic for simple surgeries, inadequate dosage, broad use → selects for resistant bacteria when given for viral infections Altered permeability or uptake—intrinsic barriers Gram(+)—porous PG is poor barrier to drugs Gram(-)—OM & LPS more effective barrier, but drugs enter thru pores Mycobacteria—additional barrier makes intrinsically resistant Reduced uptake—e.g. mar mutations in E. coli caused ↓expression of OmpF porin & loss of OprD porin in Pseudomonas results in imipenem resistance Active efflux—pump Abx out of cell w/ ATP; found in both Gram(+) & Gram(-); encoded by transposable elements Altered cellular targets: Mutant proteins that do NOT bind Abx—e.g. Streptomycin resistance (mutant 30S ribosomal subunit) Rifampin resistance (-subunit of RNA polymerase) OR Quinolone resistance (GyrB subunit of DNA gyrase) Methicllin resistance S. aureus (MRSA)--MRSA acquire gene encoding new PBP w/ low affinity for -lactams Vancomycin resistant enterococci (VRE)—normally inhibits cell wall synthesis by binding D-Ala-D-Ala, resistant bacteria add D-Ala-D-Lactate instead VanH converts pyruvate to D-latate → VanA ligates D-Ala & D-lactate together → VanX cleaves any D-Ala-D-Ala formed by native enzymes Drug-modifying enzymes—inactivates drug; e.g. -lactamase, chloramphenicol acetyltransferase, aminoglycoside phosphatase Medical solution—co-administer -lactam w/ -lactamase inhibitor, but bacteria produce more -lactamase to bind added competition M. tuberculosis—~10million new cases of TB each year, ~3million deaths Primary Tx—Rifampin, Streptomycin, some Fluoroquinolones TB-specific antibiotics—Isoniazid (inhibits mycolic acid synthesis), Ethambutol (inhibits cell wall formation), Pyrazinamide (unknown function) Abx resistant mutations: rpoB (b-subunit of RNA polymerase)—resistant to Rifampin rpsL (S12 ribosomal subunit)—resistant to Streptomycin pncA (nicotinamidase cleaves PZA)—resistant to pyrazinamide embB (polyermizes cell wall saccharides)—resistant to Ethambutol katG (catalase-peroxidase, activates INH)—resistant to Isoniazid inhA (mycolid acid synthesis)—low-level resistance to INH Sterilization—destruction of ALL microbes, including spores Disinfection—reduction of microbes from inanimate objects; 3 levels: High—effective against ALL microbes except high levels of endospores Glutaraldehyde—alkylates sulfhydryl, hydroxyl & carboxyl & amino groups Formaldehyde—alkylates amino & sulfhydryl groups of proteins & ring nitrogens of purines; toxic fumes Peracetic acid—disrupts sulfhydryl & sulfur bonds; versatile; broad spectrum for critical/semi-critical; combined w/ H2O2 H2O2—free radicals that destroy membranes & DNA; versatile; residues may cause hemolysis & enteritis Intermediate—kills M. tuberculosis & most resistant viruses, NOT endospores Low—kills vegetative bacteria & fungi Chlorine compounds—oxidize thiol groups & halogenate amino groups; most common hypochlorites & organic chloride Release hypochlorous acid (HOCl); low cost, broad spectrum; corrosive & irritating if not diluted; for environmental surfaces Alcohol—denatures proteins at 70% concentration; NOT sporicidal; NO residue; semicritical/non-critical Phenol compounds—destroys cell wall & precipitates cellular protein; residue may cause irritation & hyperbilirubinemia; non-critical/environmental surfaces Iodophors—iodine + carrier (continous iodine release) that disrupts protein & nucleic acid structure & synthesis; Betadine; antiseptics; corrosive & discoloring Quaternary ammonium compounds (Quats)—disrupts cell membranes & denatures proteins; reduced by hard water, soap & protein; environmental surfaces Pasteurization—disinfection using heat to kill microbes in liquids; 65°C for 30m or 71°C for 15s Sanitization (decontamination)—reduction of microbes to “safe” levels Antisepsis—elimination of microbes from living tissue w/ non-chemotherapeutic agents (antiseptics) Critical items—introduced into normally sterile body regions, MUST be sterilized Semi-critical items—onto mucosa or broken skin, MUST be sterilized or high disinfection Non-critical items—onto contact skin, low disinfection or sanitization Efficacy—tested against battery of resistant microbes Bacillus subtilis spores—test dry heat, ethylene oxide, gas plasma B. stearothermophilus spores—tests steam, chemi-clave T & pressure gauges unreliable; indicator (Bowie-Davies) tape doesn’t measure time Chemical & physical agents: Nucleic acids—alkylating agents (pyrimidine dimer formation), acridine dyes (intercalation into DNA helix), UV radiation (dsDNA breaks) Proteins—organic solvents (denaturation), crosslinkage of free sulfhydryl groups, altered enzymatic activity Cell membrane—altered permeability thru release of ions & small molecules, ↓active transport & energy generation Cell wall—disruption causing osmotic fragility Heat—macromolecular denaturation Moist heat (autoclave)—steam under pressure most efficient Dry heat (ovens)—require higher T & longer time Chemiclave—formaldehyde, ethanol, water; often for dental applications Chemicals—sterilize items that can be damaged by autoclave or dry heat Ethylene oxide—causes pyrimidine dimer formation; extremely toxic Peracetic acid—MUST be immersed; causes denaturation & ↑membrane permeability Gas plasma—for T & moisture sensitive items; electric field applied to H2O2 vapor → hydroxyl & hydroperoxy free radicals (NO toxic endproducts) Vapor-phase H2O2, ClO2 or O3—strong oxidizers Radiation—used in OR to control airborne microbes UV radiation (~260nm)—interferes w/ DNA replication; bactericidal but w/ poor penetration Ionizing radiation—x-rays, -rays, cathode rays; produce oxidizing & reducing agents; spores resistant Filtration—size, electrostatic & adsorptive means; porous discs of inert cellulose esters Intrinsic resistance—spores (spore coats) > mycobacterium (waxy cell wall) > Gram(-) w/ OM > Gram(+) (glycocalyx ONLY) 23 Acquired resistance—mutation or acquisition of plasmids/transposons; surface alteration, inactivation, efflux; correlates w/ Abx resistance Bacterial genetics: Genomics—sequence genome → examine for ORFs (usually >100AAs) → compare ORF sequence w/ previously identified proteins of known function → predict function → predict capabilities of organism (but at least half ORFs encode novel proteins) Operon—basic unit of transcription, includes gene, promoters & operators; organized into regulons (multiple operons) Genetic code—essentially same as humans w/ ATG (methionine) start codon, except stop codons (TAA, TGA, TAG) b/c no corresponding AA Opening reading frame (ORF)—region of DNA b/w start & stop codon that encodes protein Gene (cistron)—usually ORF encoding for protein but also structural RNAs Operator—affects interaction b/w RNA polymerase & promoter thus controlling level of transcription RNA polymerase—4 main proteins (, ’, 2) --guides RNA polymerase to promoter to begin transcription; --aids termination of transcription NusA, NusB, NusC—subunits that affect efficiency of transcription Gene regulation—adapt to environment & host (nutrients & surface protein interaction); efficiency; development (depending on stage, e.g. spores) Inducer—↑transcription (e.g. lactose stimulates transcription of lac operon & arabinose induces ara operon) VS. repressor—↓transcription (e.g. lacI) Activator—↑transcription (e.g. AraC) Negative regulation—e.g. E. coli lac operon that transports lactose into bacterium & degrades LacZ (-galactosidase), lacY (lactose permease), lacA (galactose acetylase), lacI (or lacR, lac repressor not found w/in lac operon) Absence of lactose—lacI binds lac operator & prevents transcription VS. presence—lacI binds lactose transcription of lac operon Positive regulation—e.g. E. coli araBAD operon for degradation of sugar arabinose araA, araB, araD—structural genes for arabinose degradation; araC—ara activator NOT found w/in araBAD operator Absence of arabinose—promoter does NOT bind RNA polymerase & NO transcription VS. presence—araC binds arabinose binds ara operator Regulon—multiple opersons; e.g. cCMP & CRP (cAMP-receptor protein); cAMP levels inversely proportional to cellular glucose CRP-cAMP complex binds DNA to stimulate transcription of operons & catabolize “alternative” C sources e.g. ppGpp (AA starvation response) & SOS system (senses DNA damage) Multiple effectors—e.g. E. coli lac operon sensitive to both lactose & glucose levels (RNA polymerase requires CRP-cAMP bound to operator also) Multple ezymes—bacteria may encode several versions of same enzyme; e.g. E. coli encodes 3 genes for enzyme in synthesis of isoleucine, leucine & valine (ilvB, ilvG, ilvH) ilvB—transcription stimulated by ppGpp & CRP-cAMP, sensitive to levels of leucine & isoleucine, inhibited by valine ilvG—transcription stimulated by ppGpp, sensitive to levels of leucine, isoleucine, & valine ilvH—regulated by activator protein that cannot bind operator in presence of leucine; inhibited by valine Environmental sensing—sense T of human host, C source, AA, N, P, nucleic acids; 2-component regulator systems: Sensor protein—usually in membrane; stimulus causes transfer of phosphate from ATP onto histidine residue on sensor protein (autophoshorylation) Response regulator (transducer)—usually soluble cytoplasmic protein; activated by phosphorylation on aspartate residue by sensor protein Quorum sensing—cells produce signal molecules (peptides or homoserine lactones) that adjacent cells detect Mutations—classified as wild-type, silent, missense, nonsense, frameshift, or inversion Transition—purine to purine (A to G) or pyrimidine to pyrimidine (T to C); usually deleterious Transversion—purine to pyrimidine & vice versa; usually deleterious Insertion—usually deleterious unless large DNA segment added w/ useful function (e.g. antibiotic resistance); often w/ long repeated DNA sequences Deletion—almost always deleterious; often w/ long repeated DNA sequences Duplication—deleterious OR beneficial addition of multiple copies of a DNA sequence w/o transfer from another organism Inversion—usually deleterious reversal of DNA sequence Spontaneous mutations—occur in normal cells NOT exposed to mutagenic factors Uncorrected replication errors—transitions, transversions, small deletions/additions Spontaneous deamination—conversion of C to U Recombination b/w homologous segments—deletions, inversions or duplications e.g. Neisseria gonorrhoeae—11 different opa genes (encoding antigenic surface proteins); lack of opa permits survival from opa-directed Ab Spontaneous insertions or deletions of repeated 5 base sequence (CTCTT) w/in protein coding region Induced mutations: DNA reactive agents—alter base pairing or cause cross-linking b/w bases (e.g. UV radiation induces thymine dimerization) Base analogs—incorporated into DNA w/ frequent mispairing errors during replication Planar polycytic molecules—slide b/w bases of helix (intercalate) & cause frameshifts Base substitution—e.g. single base change in rpsL encodes a S12 protein resistant to streptomycin rpsL codon 42—wild-type (AAA) encodes lysine (antibiotic sensitive) VS. mutant (ATA) encodes arginine (resistant) Recombination: Homologous—requires extensive base pairing b/w homologous regions (double crossover event); mediated by RecA protein Site-specific—b/w specific short segments of homologous DNA; mediated by specific enzymes; used by many bacteriophages for genomic integration into host Non-homologous (“illegitimate”)—NO homology, unknown mechanism e.g. E. coli—synthesis of fimbriae by site-specific recombination; transcription of fimA (gene encoding fimbrial subunit) dependent on orientation of promoter Catalyzed by products of fimB & fimE genes e.g. Borrelia hermsii—synthesis of different VMPs (variable membrane proteins) to evade recognition by Ab; causes relapsing fever Transformation—some species naturally competent to take up & recombine extracelullar DNA (e.g. H. influenzae, S. pneumoniae, N. gonorrhoeae) S. pneumoniae—can take up any segment of DNA; develop competence in response to activating signals (quorum sensing) N. & H.—take up DNA segments containing specific DNA-uptake sequences (DUS) found frequently in bacterial genomes E. coli—NOT naturally competent but can be induced by chemical treatment Natural competence—spread of Ab resistance, ↑genetic variation, ↑virulence, acquisition of beneficial genes, ↑host range DNA restriction systems—maintain integrity DNA methylase—recognizes specific DNA sequence & modifies base by adding methyl group Restriction endonucleases—recognizes some specific DNA sequence, does NOT cut if sequence methylated (“self”), cuts if not methylated (“non-self”) Plasmids—circular or linear, extrachromosomal DNA (episomes); metabolic drain on bacteria but some selective advantages Provide essential proteins, Abx resistance (R-factors), produce Abx (kill competing bacteria), catabolic enzymes (use of certain nutrients), virulence determinants Integration to form high-frequency recombination (Hfr) chromosome, exact excision to re-form F plasmid or inexact excision to form F’ plasmid Host range—determined by mechanism of plasmid replication Limited host range—replicate ONLY in single bacterial species or closely related; uses host DNA replication machinery 24 Broad host range—replicate in wide variety of species; generally encode own replication machinery Conjugation—plasmid transferred from donor (“male”) to recipient (“female”) thru conjugation bridge Mobilization—plasmid parasitizes conjugation machinery of another plasmid to facilitate transfer b/w bacteria Bacteriophages—bacterial viruses that recognize appropriate hosts by interacting w/ specific surface components Lytic phage—produces multiple copies of phage particle then lyses bacterial cell Lysogenic phage—prophage replicates DNA along w/ host cell, does NOT kill cell; e.g. integrant or episome (plasmid-like) Virulence properties—often carried by phages; e.g. cholera toxin of Vibrio cholerae, Shiga-like toxins of E. coli, erythrogenic toxin of group A Strep Transduction—phage w/ some bacterial DNA injects genome into recipient cell, recombination forms transduced recipient Transposable elements—defined DNA segments that catalyze own transfer b/w or to other species; NOT dependent on homologous or site-specific recombination Defined ends w/ inverted terminal repeats, encode specific transposase, cause mutations, generate direct repeat in target DNA Transposase makes staggered cut at target site → TE rejoins ends of new transposon into space → gaps filled by host enzymes Insertion sequence (IS) elements—contain inverted terminal repeats & transposase; central region encodes proteins required for transposition Transposons—contain inverted terminal repeats, transposase, & additional DNA sequences (e.g. antibiotic resistance genes) e.g. IS10/Tn10 confers tetracycline resistance Bacteriophage Mu—both phage & transposon that infects E. coli & injects DNA; lytic or lysogenic (random transposition integration) Conjugative transposons—inverted repeats, encode transposase, carry antibiotic resistance genes Do NOT generate direct repeat in target DNA, can transfer themselves b/w cells, different mechanism of transposition Intracellular transposition—excision & integration back into chromosome or plasmid w/in same cell Intercellular transposition—excision, transfer & replication to dsDNA, integration back into genome Chlamydiae: Bacteriology—small (15% size of E. coli), round Gram(-), NO PG, non-motile, obligate intracellular, metabolically deficient, dimorphic Elementary body (EB)—small, infectious, extracelullar, metabolically inactive, spore-like form w/ highly cross-linked OMP by disulfide bridges Reticulate body (RB)—larger, replicative, intracellular, metabolically active, fragile form Pathology—adhesion of EB (heparin-sulfate-like ligand) to receptors on epithelial cell surface → endocytosis → inhibition of phagosome fusion → fusion w/ sphingomyelin-containing vesicles near golgi → differentiation of EB to RB (9-10h) → replication of RB by binary fission forming intracytoplasmic inclusions (glycogen-filled) → attach to inclusion membrane → differentiation back into EB (16-20h) → detach from inclusion membrane → lysis of host cell & spread C. pneumoniae—first isolated in 1965 from conjunctiva of Taiwanese child; major respiratory pathogen by 1983 Transmission—respiratory droplets w/ possible reservoir in amoebae, throughout year; infection & reinfection universal Presentation—~10% pneumonia & 5% bronchitis, pharyngitis & sinusitis in adults Initial pharyngitis → LRI 1-3w later → cough may persist for weeks Complications—possibly associated w/ atherosclerosis (binds LDL → endothelial injury → foam cell formation → atherosclerosis) & late-onset AD Diagnosis & Treatment—serological testing NOT routine; tetracycline & erythromycin (recovery slow) C. psittaci—originally discovered in psittacines (parrots & parakeets); 100-200 cases/y in US Transmission—respiratory droplets or droppings from infected birds; occupational hazard of poultry workers & bird owners Presentation—acute bilateral interstitial pneumonia w/ fever, malaise, dry cough, aches; hepatosplenomegaly common Complications—mycoarditis, encephalitis & hepatitis Diagnosis—serological fluorescence or complement fixation Treatment—tetracycline & erythromycin C. trachomatis: Transmission—sexually transmitted Virulence factors—LPS 100X less inflammatory; heat shock proteins (HSP) causes intense inflammation → scarring Presentation: Trachoma—serotypes A-C; developing countries, cause 9million blindness 15-20y after infection; conjunctivitis → scarring of cornea Urogenital infections—serotypes D-K; most common bacterial STD; 10-15% sexually active teens infected; 45% co-infections w/ gonnorrheae Urethritis, dysuria & discharge, epidiymitis, or cervicitis (70% asymptomatic) 5-30% ascending infection → salpengitis & PID → scarring, infertility, ectopic pregnancy (10% patients) Lymphogranuloma venereum (LGV)—L1, L2, L3 serotypes; 200-400 cases/year in US Suppurative inflammation of inguinal LNs; hemorrhagic ulcerative proctitis in homosexuals Inclusion conjunctivitis (IC)—2-6% newborns in US; serotypes D-K Acute, self-limiting swelling of eyelids w/ purulent discharge 2-25d after birth → may progress to pneumonia Neonatal pneumonia—1-6m after birth; afebrile, pertussis-like cough w/ dyspnea; rarely fatal Diagnosis—culture is expensive & time-consuming; glycogen detected by iodine; fluorescence, immunoassays of LPS or DNA probes Treatment—erythromycin & tetracycline; problems w/ noncompliance Legionella pneumophila—at least 10% of community-acquired pneumonia; 15-50% mortality Bacteriology—Gram(-) bacilli (stains poorly), motile, aerobic, NOT spore-forming; serogroup 1 accounts for ~80% of Legionnaire’s dz Requires complex medium (buffered charcoal yeast extract agar, BCYE-) supplemented w/ -ketogluterate, cytsteine, & Fe Transmission—fresh water & soil; aerosols (faucets, shower heads, ACs, whirlpools), NOT person-to-person; parasite of protozoa (amoebae) Pathology—binds Gal/GalNac lectin protozoan receptor → endocytosis → coiling phagocytosis by alveolar M → inhibits lysosome fusion (NO acidification) → phagosome surrounded by ER (replicative phagosome) Virulence factors—pore-forming activity causes lysis & induces apoptosis in M & epithelial cells Type IV pili—attachment dot/icm loci—inhibition of lysosome fusion & assembly of contact-dependent (type IV) pili Characteristic intracellular replication w/in monocytes & M in alveolar spaces (NOT bronchi) → intense acute inflammation → CK release → extensive necrosis of alveolar epithelium & lysis of inflammatory cells → microabscesses → coalesce to form cavities Presentation—fever, cough, dyspnea, chest pain, NOT purulent sputum; possibly watery diarrhea, low blood O2, abnormal liver function Pontiac fever—infection or hypersensitivity reaction to Legionella or protozoan Ag; NOT produce pneumonia & NEVER fatal Transmission—air-borne Presentation—flu-like illness, self-limiting in 2-5d Diagnosis—Gram stain of sputum usually suggest atypical pneumonia w/ abundance of PMNs & monocytes w/o presence of bacteria; culture MOST diagnostic Treatment—erythromycin (DOC), Rifampin, Tetracycline Streptococci—generally opportunistic pathogens 25 Bacteriology—normal flora, Gram(+) round, diplo- or chains, extracelullar, aerotolerant, catalase(-) & oxidase(-) Classification—based on , , hemolysis on BAP Lancefield classification—differentiates -hemolytics based on antigenic cell wall CHO (except group D lipoteichoid acid Ag); most human isolates groups A-E S. pyogenes—Group A Strep primary pathogen Presentation—impetigo, pharyngitis (“strep sore throat”), toxic shock-like syndrome (TSLS), necrotizing fascitis → non-suppurative sequelae (ARF & AGN) S. agalactiae—Group B Strep (GBS); leading cause of neonatal sepsis & meningitis (1-3/1000 births); mortality 30-60% Bacteriology—-hemolytic; sialic acid capsule Transmission—up to 40% pregnant women carriers & 60% transmission to newborns; aspiration of contaminated amniotic fluid OR infected birth canal Virulence factors—sialic acid capsule (↓C3 deposition on surface), peptidase (inactivates C5a), absence of maternal IgG Presentation—UTI in females, neonatal sepsis & meningitis Early onset disease—w/in ~20h; vertical transmission; primary focus in lung → respiratory distress Late onset disease—w/in ~24d; vertical OR horizontal transmission; bacteremia → meningitis → neurological sequelae → death Treatment—aggressive combination of PCN & aminoglycoside to prevent permanent neurological damage Prophylaxis—intrapartum w/ PCN/ampicillin Vaccine—under development; protein carriers (tetanus toxoid & Strep c Ag) S. equisimilis—Group C; causes UTIs, wound infections, endocarditis, brain abscesses, sepsis Enterococci—Group D Strep, cause UTIs, endocarditis, wound infections, nosocomial infections Groups E-V—NO Linnaean designations; Group G causes pharyngitis, cellulitis, bone & joint infections Viridans streptococci---hemolytic; normal flora of pharynx, GI & female GU tracts; causes caries, wound infections, endocarditis S. pneumoniae---hemolytic; pneumonia & acute purulent meningitis Peptostreptococci—anaerobic, -hemolytic; cause brain abscesses & gingivitis Infective endocarditis—10,000-20,000 cases/year; >50% patients >50yo Pathology—altered valve surface → deposition of platelets & fibrin (non-bacterial thrombotic endocarditis, NBTE) → attachment & colonization of NBTE Virulence factors: Fibronectin—common host receptor; fibrinogen-binding proteins of S. aureus (coagulase) S. mutans surface protein—binds ECM & endothelium (enhanced by dextran synthesis) FimA—adherence to platelet-fibrin matrix; platelet aggregation enhances NBTE colonization (S. sanguis) Poor M penetration, bacterial population grows to 1010/g tissue, reduced metabolic activity (↓antibiotic susceptibility), occlude valves → emboli & infarcts Acute bacterial endocarditis (ABE)—abrupt onset → fulminant course → death w/in 6w (40% mortality) Caused by nosocomial-acquired Streptococci, S. aureus, N. gonorrhoeae, & Candida Subacute bacterial endocarditis (SBE)—slow progressing → death w/in >6w Caused by Viridans streptococci & Enterococci; opportunistic normal flora (oral strep often caused by dental induced bacteremia) w/ underlying HD Treatment—valve replacement (~25% cases); relapse common; extended dosage, combination Abx Group A Streptococci (GAS)—localized & systemic infections, non-suppurative sequelae Virulence factors: M-protein—major cell surface Ag (“fuzzy coat”) anchored to PG & binds ECM & Fc of IgG (adhesin); determines >90 serotypes Binds Factor H → ↓C3b deposition on surface Also M-related proteins, fibronectin binding proteins (F1 & F2) & collage binding proteins (Epa) Hemolysins—demonstrated by hemolytic reaction on BAP; streptolysin O (anti-streptolysin O (ASO) test determines recent Strep infection) & streptolysin S Streptokinase—plasminogen-plasmin system activator that spreads of infection Hyaluronic acid capsule—↓C3b binding, camouflage & cross-react Ab (w/in synovial tissue & cartilage) C5a peptidase—↓neutrophil migration Streptococcal inhibitor of complement (Sic)—disrupts MAC Streptococcal pyogenic exotoxins (Spe)—superantigen carried by bacteriophage; SpeA most important (TSLS) & SpeB (cysteine protease for spread) Lipoteichoic acid—polyglycerol-P attached to lipid, complexed w/ M-protein to ↑colonization → septic shock Presentation—pharyngitis & impetigo most common (also w/ Groups C & G) w/ non-suppurative sequelae (ARF & AGN) Pharyngitis—most often in 5-15yo w/ 15-20% carrier rate in kids; transmitted by respiratory droplets; diagnosed by throat culture Treatment—PCN start <9d after symptoms to prevent sequelae Pyoderma (impetigo)—localized, purulent skin infection; most often in 2-5yo Presentation—entry & colonization of broken skin; few systemic symptoms w/ complications of septicemia & AGN Treatment—PCNs & cephalosporins SpeA-producing GAS: Toxic shock-like syndrome (TSLS) or Streptococcal TSS (STSS)—sudden onset of shock & organ failure (mortality 10X greater than TSS) Associated w/ skin wound, RTIs → bacteremias (SpeA-producing GAS w/ M1 or M3 serotype) Treatment—quick & aggressive surgical exploration & debridement, support, Abx Necrotizing fascitis (Streptococcal gangrene)—deep subcutaneous tissue & fascial infection Erysipelas—acute inflammation of skin & cutaneous lymph vessels Streptococcal cellulitis—acute, spreading inflammation of skin & subcutaneous tissue Acute rheumatic fever (ARF)—characterized by non-suppurative inflammation of heart, joints, subcutaneous tissue & CNS; usually in 6-15yo Presentation—<3% untreated pharyngitis followed by ~18d latent period & acute, febrile, self-limiting; may cause valve damage (autoimmune cross-reactivity) Complication—long-term rheumatic HD Treatment—Abx, corticosteroids; prophylaxis until early 20s for rheumatic patients; vaccine development slow (fear of cross-reactivity) Acute glomerulonephritis (AGN)—delayed, non-suppurative acute inflammation (immune complex deposition) of renal glomeruli Presentation—edema (fluid overload), HTN, ↓output of rusty-colored urine More common after skin infection (M49) w/ 21d latent period, OR after pharyngitis (M12) w/ 10d latent period <10-15% cases & recurrence rare; >90% kids recover BUT adults often develop subacute/chronic infection Treatment—salt & fluid restriction, diuretics, benzathine PCN G Diagnosis—culture on BAP; bacitracin sensitive; latex agglutination (differentiate from other -hemolytics), catalase(-) (differentiate from Staph) Staphylococcus—ubiquitous causing ~80% suppurative infections Bacteriology—Gram(+) cocci, clusters, facultative aerobes S. aureus—primary human pathogen in “healthy” hosts; coagulase(+) & heat resistant endonuclease, mannitol fermentation S. epidermidis—opportunistic pathogen; coagulase(-) S. saprophyticus—opportunistic pathogen; coagulase(-) 26 Transmission—colonization of body surface begins at birth (normal flora), breach of skin or mucosal barriers exposes bacteria to tissues S. epidermidis & saprophyticus—~11% nosocomial dz w/ high affinity for foreign materials Transmission—S. epidermidis—65-90% of all normal human surface Staph, especially axilla, inguinal & perineal regions S. saprophyticus—colonized inguinal & perineal regions; cause young female UTIs (tropism for UT epithelium oligosaccharide receptors) Virulence factors: S. epidermidis—fimbrial protein for attachment, polysaccharide intracellular adhesion (PIA) binds other bacteria & helps establish biofilms Transferrin binding protein for Fe acquisition S. saprophyticus—hemagglutinin & fibrillar surface proteins bind UT epithelium; urease causes invasion Presentation—bacteremias mostly catheter-related w/ 50-70% due to S. epidermidis CSF shunt infections—50-70% S. epidermidis Prosthetic cardiac valve infections—almost always S. epidermidis; native valve endocarditis rare but high mortality UTIs—S. saprophyticus cause “community-acquired” UTIs in 16-35yo females w/ sex, outdoor swimming, occupational meat packing Treatment—methicillin (80% resistant); oxacillin (DOC), cephalosporins S. aureus—~37% carrier in general population Structure—Gram(+) w/ 40-60% PG, mannitol(+), catalase(+), -hemolytic; carotenoid pigments (“aureus” Latin for “gold”) Transmission—reservoir in anterior nares Virulence factors: PNSG (surface polysaccharide)—prevent phagocytosis & ↑colonization Ribitol teichoic acid—phage receptor & regulates daughter cell separation Capsule—serotypes 5 & 8 most significant; inhibits alternative complement & phagocytosis, ↑adherence to hydrophobic surfaces Protein A—covalently bound to PG & released into environment; binds Fc of IgG (except IgG3) Toxic shock syndrome toxin & Staph enterotoxin—superantigens Exfolative toxins (exfoliants)—separate epidermis from underlying tissue; from bacteriophage group II; ETA (chromosomal) & ETB (plasmid) Staphylokinase—dissolves clots by activating plasminogen Hyaluronidase—causes dissolution of ECM -toxin (-hemolysin)—forms pores in host cells Leukocidin—cytotoxic for PMNs & M, changes cellular phospholipid metabolism G3PDH—transferrin receptor for Fe acquisition Presentation—most common cutaneous infection w/ characteristic abscess formation Cutaneous Staph infections—most common human bacterial infections Pathology—Langerhan’s cells produce IL-1 & TNF; endothelium produces IL-8 (chemoattractant for PMNs) → PMNs engulf bacteria w/ C3R Impetigo—encrusted pustules → folliculitis—subcutaneous extension (boil, furuncle OR stye) → carbuncles—multiple, interconnected abscesses Staph scalded skin syndrome (SSSS)—Ritter’s dz or bullous impetigo; skin layers separate & outer layers slough off; neonates-4yo & immunocompromised Toxic shock syndrome (TSS)—associated w/ high absorbency tampons → inhibit normal flora → Staph overgrowth; possible genetic predisposition Food poisoning—ingestion of pre-formed toxin (intoxication) Pneumonia—>10% bacterial pneumonias; aerosol from URIs; up to 50% mortality; <1yo or imczd; associated w/ influenza epidemics Bone & joint infections—septic arthritis & osteomyelitis Bacteremia—50% hospital-acquired; associated w/ underlying dz or foreign bodies; up to 38% develop ABE Treatment—Vancomycin (DOC) w/ MIC susceptible (<4g/mL), intermediate (8-16), resistant (>32) Methicillin resistant S. aureus (MRSA)—bacteria acquire mecA gene that encodes modified PBPs Corynebacterium diphtheriae: Transmission—normal flora of skin, oropharynx & distal urethra; opportunistic; aerosol inhalation → colonization of pharynx → forms pseudomembrane → toxin production Virulence factors—K & O antigens; diphtheria toxin (DT), encoded by corynephage & carry toxin structural gene (tox+) DT—non-toxic pro-enzyme; Part A (active) linked by disulfide bridge to Part B (binds target cell membrane “R domain”) Part B binds heparin binding epidermal growth factor (HB-EGF) receptor → triggers endocytosis → ↓pH in phagosome → alters DT structure → Part B forms channel & Part A translocates into cytoplasm → completely disrupts target cell protein synthesis (inhibits EF-2) Bacteriology—Gram(+) pleomorphic rods; aerobic; non-spore forming; club-shaped; may appear beaded (internal Babes-Ernst granules); catalase(+) & oxidase(+) Cultured on Loeffler’s or tellurite medium—inhibits competing bacteria & high lipid concentration Presentation—superficial inflammation & necrosis of respiratory tract mucosa & skin; pharyngitis, tonsillitis, inflamed cervical LNs Bacteremia → myocarditis & arrhythmias; paralysis, neuritis, nerve dysfunction → death due to asphyxiation, cardiac arrest (mortality up to 30%) Diagnosis—presence of pseudomembrane Treatment—early horse diphtheria antitoxin therapy (DAT) & erythromycin eliminates carrier state Vaccine—treat toxin w/ formalin (toxoid); given during first 6-8w, booster entering kindergarten & every 10y Enterobacteriaceae—largest, most diverse group of Gram(-) bacilli; mostly opportunists Bacteriology—small Gram(-), non-spore forming rods (coccobacilli); facultative anaerobes, lactose(+) & oxidase(-) Virulence factors—O antigen (LPS capsule), H antigen (flagella), fimbriae & K antigen (capsule, slime layer) Classification—based on patterns of CHO use, variations in endproducts, serotyping (antigens) E. coli—opportunistic & primary pathogen Transmission—fecal contamination; colonizes large intestine Presentation—>85% community-acquired UTIs, uropathogenic strains (UPEC); females <10yo & 20-40yo (anatomy & sex); primarily ascending infections Hospital-acquired UTIs—flow obstruction, ~50% of patients catheterized >5d; ascending or descending Bacteremia—Gram(-) sepsis & endotoxic shock Virulence factors: Adhesins—type I (common) pili bind mannose-containing receptors of bladder & stimulate uptake by uroepithelial cells P fimbriae—bind P blood group Ag (globobiose) of bladder & kidney cells LPS (O antigen)—endotoxin, ↓complement-mediated lysis & ↓opsonization Hemolysin—lyses host cells thru pores, ↓phagocytosis, ↑CKs Siderophores (aerobactin)—chelate Fe Capsule (K antigen)—polysaccharide (some protein), ↓phagocytosis & ↓alternative complement activation Flagella (H antigen)—motility Klebsiella—3 species (pneumoniae, rhinoscleromatis, oxytoca); common GI inhabitants Structure—lactose(+), non-motile K. pneumoniae—most common; burn infections, UTIs (9%), bacteremia (14%), meningitis Community-acquired lobar pneumonia—opportunistic; predisposed w/ alcoholism, DM, COPD; 25-50% mortality 27 Virulence factor—capsule (K antigen) determines serotypes; endotoxin (O antigen) → ↓lysis by MAC; pili; heat stable & labile toxins Treatment—3rd generation cephalosporins; naturally resistant to CAN & carbenicillin; R-plasmid provides -lactamase Enterobacter—most common aerogenes, cloacae, agglomerans Structure—lactose(+), motile, capsule E. cloacae—most common hospital isolate (burn, wounds, respiratory, UTIs) Virulence factors—O antigens, capsule (K antigens), flagella (H antigens), heat labile & stable toxins Treatment—3rd generation cephalosporins b/c inducible -lactamase Serratia—most common marcescens Structure—lactose(+ or -); differentiated by extracelullar DNAse, resistance to colistin & cephalothin, prodigiosin (red pigment) Presentation—75-90% nosocomial infections (catheters, foreign bodies → UTIs, wound infections, pneumonia, septicemia) Virulence factors—O & H antigens Treatment—amikacin; -lactamase producing Citrobacter—amalonaticus, diversus, freundii (most common) Structure—uses citrate as sole C & energy source; forms H2S that cleaves disulfide bonds of CKs Presentation—UTIs & respiratory infections in hospitalized patients; neonatal meningitis & brain abscesses (C. diversus) Virulence factors—O, H & K antigens; OMP in more virulent strains; heat stable toxins Treatment—plasmid-mediated -lactamase; resistance greater in C. freundii Proteus, Providencia & Morganella—historically classified as Proteus; P. mirabilis most common Structure—lactose(-), motile, phenylalanine deaminase(+), indole(+) except P. mirabilis Presentation—up to 15% of all nosocomial infections; community-acquired UTIs (upper) Virulence factors—fimbriae colonize uroepithelium (mannose-resistant Proteus-like, MR/P & P. mirabilis fimbriae, PMF) Flagella for spread, hemolysins, urease (stone formation w/ metalloprotease) P. stuartii—common in catheterized nursing home patients Treatment—senstive to most common Abx; resistant to PCN & aminoglycosides (indole+); inducible -lactamases Pseudomonas—naturally inhabit soil; P. aeruginosa most common Structure—mucoid growth (pyanocyanin pigment & fruity odor), biofilm formation, oxidase(+), motile; versatile Presentation—2nd most common cause of hospital-acquired pneumonia (particularly in CF, neoplastic dz & burn patients) CF—viscous pulmonary secretions, less NO, bacterial biofilm→ cannot eliminate bacteria 80-90% develop chronic pulmonary infection w/ selective pressure due to ABC use Virulence factors—adhesin pili bind ganglosides, non-pilus adhesions bind epithelial cells; LPS binds CFTR Exoenzyme S (ExoS)—disrupts signal transduction, ↓phagocytosis Elastase (LasA & B) Phospholipase C (rhamnolipid)—lung tissue destruction Alginate capsule (mucoid exopolysaccharide, MEP)—↓opsonization, ↓ABC access, biofilm formation & adhesion Pyocyanin—endothelial cell damage thru OH- synthesis Pyochelin (pyoverdin)—siderophore Endotoxin & exotoxin A—hypotension & shock, local inflammation Treatment—resistant to most Abx due to restricted access, efflux pumps, biofilm formation & -lactamases; 4th cephalosporins & broad fluoroquinolones Salmonella—enteritidis, typhimurium (most common), typhi, colerasuis (swine pathogen) Bacteriology—enterobactereaceae; lactose(-), produce H2S from AA; bile resistant, motile; O, K, H antigens (at least 2200 serotypes); acid tolerant S. typhimurium—ubiquitous; dz of industrial society, 15% food-borne infections of US; 1/3 of epidemics occur in nursing homes Transmission—poultry & infected eggs (animals fed high-protein diets of other animal remnants) Asymptomatic carriers shed bacteria in stool; antacids increase risk of transmission (smaller inocula) Virulence factors—5 pathogenicity islands w/ 2 contact-dependent secretion systems SPI-1—invasion of epithelial cells SPI-2—intracellular survival w/in phagosome, blocks NADPH oxidase-dependent killing Attachment to intestinal epithelium → induce signal transduction cascade → cytoskeletal rearrangments → ruffles surround bacteria → internalization w/in vacuole → acidification but NO maturation into phagosome Regulatory loci (phoP/phoQ)—genes for intracellular survival (induce M apoptosis), induced by ↓[Mg2+] Presentation—nausea, abdominal cramps, diarrhea (watery-bloody, NO WBCs) & vomiting w/in 8-48h lasting 3-4d; fever in 50% Treatment—self-limiting; fluid & electrolyte replacement; Abx for systemic infections; NO vaccines (except for chickens) S. typhi & S. paratyphi—enteric fever; invades intestinal mucosa thru CFTR (may explain high prevalence of CF in Europeans) Virulence factors—invasion & killing of M cells → invasion of M w/in Peyer’s patches → replication w/in M vacuole → spread to RES, liver & spleen Presentation—incubation >1w followed by fever & headache → faint rash on abdomen & head → bacteremia (multi-organ involvement) Prolonged infection (>2w)—perforation & hemorrhage of intestinal wall Chronic infection—w/in gallbladder, particularly stones; produces carrier state (Typhoid Mary Mallon) Yersinia enterocolitica: Bacteriology—enterobactereaceae; urease(+) at 37°C & urease(-) at 25°C; motile at 25°C Transmission—ubiquitous, particularly in Scandinavian countries; fecal-oral (contaminated food, water, milk); mostly in children Virulence factors—adhesins; YopM inhibits platelet aggregation; heat-stable enterotoxin Contact-dependent secretion system—effectors block phagocytosis (disrupts cytoskeleton), induces caspase-1-mediated apoptosis, ↓CKs Pathology—invasion of M cells → replicate locally in mesenteric LNs Presentation—usually self-limiting; severe sx in older children (acute mesenteric lymphadenitis) Autoimmune reactive arthritis or thyroiditis w/in 2-6w—associated w/ HLA-B27 Y. pseudotuberculosis—less common than Y. enterocolitica; more invasive bacteremia; NO diarrhea; mimics appendicitis; self-limiting Vibrio cholerae: Bacteriology—curved, Gram(-) rods, motile, oxidase(+); mostly type O:1 (El Tor); fragile & acid sensitive (large infective dose required) Transmission—contaminated water; concentrated in shellfish Virulence factors—toxin co-regulated pilus (TCP)—major colonization factor on bacteriophage (VPI) Cholera toxin (CT)—encoded on bacteriophage (CTX); 5 binding (B) subunits & A1, A2 subunits Binds GM1-ganglioside receptor → A1 subunit translocates into cell → ADP-ribosylates G protein → prevents dissociation from active adenylate cyclase → ↑cAMP → hypersecretion of Cl-, K+, HCO3-, H2O ToxR regulatory system—response to environmental stimuli 28 Zonula occludens toxin (Zot)—disrupts TJs b/w mucosal cells → ↑permeability Accessory colonization factor (Acf) & accessory cholera enterotoxin (Ace) Vac cytotoxin—vacuolation of epithelial cells Metalloprotease—degrades occludin in TJs b/w epithelial cells Presentation—colonization of small intestine; rapid onset of vomiting & diarrhea (rice-watery w/ mucous flecks, NO blood or WBCs); NO fever; NO invasion of epithelium Treatment—fluid & electrolyte replacement; tetracycline shortens duration & magnitude of fluid loss; NO vaccine Vibrio vulnificus—halophilic (sea water) & contaminated raw oysters Presentation—invasion of intestinal mucosa → septicemia → death w/in 24h (50% mortality) Wound infections in seashore bathers or fisherman → local tissue necrosis V. parahemolyticus—coastal waters & raw shellfish (Japan) Presentation—incubation of 24-48h; severe cramping, vomiting, diarrhea (watery but less than cholera); self-limiting Campylobactor jejuni—4-30% of ALL cases of diarrhea; C. fetus causes infant gastroenteritis Bacteriology—Gram(-) curved rods; microaerophilic Transmission—poultry & milk Virulence factors—cytolethal distending toxins (Cdt) block cell cycle of epithelial cells in G2 Presentation—incubation of 1-7d; colonizes intestinal mucosa; lower abdominal pain & diarrhea (blood & WBCs); fever common; self limiting w/in 3-14d Guilian-Barre’s syndrome (GBS)—acute inflammatory polyneuropathy → progressive paralysis (10% mortality) Treatment—PCN resistant Helicobacter pylori—90% of gastritis, 20-30% develop ulcers; 20-30% higher rate in males; 50% world population chronically infected Bacteriology—Gram(-) curved-to-spiral rods (motile) OR coccoid (nonmotile); microaerophilic; urease(+) Transmission—fecal-oral (contaminated water); colonizes stomach (acquired in childhood) Virulence factors—adhesins (BabA binds Lewis B blood group antigen on gastric epithelium) Neutrophil activating protein (NAP)—iron-binding protein Contact-dependent secretion system—effacement of microvilli, pedestal formation, activation of MAP kinases Vacuolating toxin (Vac) Presentation—type I (most clinically isolates causing ulcers or adenocarcinoma, induce pedestal formation & tyrosine phosphorylation), ↑IL-8 (neutrophil chemoattractant) Type II (asymptomatic, NO pedestal formation or Tyr phosphoryation) Diagnosis—Giemsa stain & urea breath test highly sensitive Treatment—2w course of 2 Abx & antiacid Tuberculosis—>2million cases in India Transmission—human primary reservoir; respiratory droplets (5m); bacteria viable & virulent, many in sputum, cause mechanisms to transmit (cough, hoarseness) Presentation—persistent cough (>3w) + hemoptysis, night sweats, weight loss (>10% over 6m), fever (>101°F over past 3w) Pulmonary—77% Primary—lower lobe infiltrate w/ hilar lymphadenopathy; few bacteria; may resolve spontaneously (become latent) Reactivation—apical infilatrates (↑O2 tension or ↓lymphatic clearance?); often cavitary; very contagious Extrapulmonary—36% lymphatic; 20% pleural; 11% skeletal; 7% GU… Latent TB infection (LTBI)—(+) PPD; NOT symptomatic, NOT contagious; needs preventative therapy Active TB—symptomatic, potentially contagious; requires multiple agent therapy Tuberculin reaction—10% chance of developing TB during lifetime w/ (+)PPD; 5% w/in first 1-2y; 10% chance each year for HIV patients 5mm—HIV, recent contacts of TB, Ghon complex on chest x-ray, imczd 10mm—recent foreign arrivals, IV drug users, high-risk employees, mycobacteriology lab personnel, at-risk dz, children <4yo or exposed to TB 15mm—no known risk factors Diagnosis—isolate bacteria & sensitivity testing; sputum specimens X3 (spontaneous, induced, bronchoscopy or gastric aspirate in children) Treatment—INH for 9m + Rifampin for 4m + 2 drugs for sensitivities (shortest time, never add 1 drug to failing regimen, ensure compliance w directly observed therapy) Multi-drug resistant TB (MDR-TB)—resistant to INH & RMP, only 56% success of therapy VS. >95% for drug-sensitive TB Mycobacteria—high lipid content of cell wall; acid fast; strict aerobes; slow growing; ~40 cases/year in US M. leprae—obligate, intracellular parasite; infects nerve tissue; grows best in cool T (skin, ears, URT, testis); humans main reservoir; chronic granulomatous dz Classification—based on progression of dz & host immune response Tuberculoid (TT) → borderline tuberculoid (BT) → mid-borderline (BB) → borderline lepromatous (BL) → lepromatous (LL) Tuberculoid—CD4 response (IL-2 & IFN- secretion), delayed hypersensitivity, granuloma formation, macular lesions w/ few bacilli (pauci-bacillary, PB); 75% lung Lepromatous—impaired cell-mediated response (NO CD4 cells), large nodular lesions w/ many bacilli (5 or more positive patches, multi-bacillary, MB); skin & nerve Up grade—transition to tuberculoid, type I, delayed hypersensitivity reaction of pre-existing lesions; treatment slows taper of steroids Down grade—transition to lepromatous, type 2, erythema nodosum leprosum (ENL), increased TNF- in tissues, multi-drug therapy w/ steroids Pathology—bacteria colonize skin & mucosa → spread to regional LNs → parasitization of Schwann cells Transmission—inhalation of aerosol Presentation—skin lesions (macular areas of hypopigmentation); nerve involvement—claw hand (ulnar/median), foot drop (peroneal), claw foot (posterior tibial) Loss of eyebrows & eyelashes; perforated nasal septum; hoarseness Diagnosis—Hx of travel to endemic area; skin biopsy & smear Nontuberculous Mycobacteria—progressive pulmonary dz (MAC & M. kansasi), superficial lymphadenitis (cervical in children, MAC, scrofulaceum, malmoense) Disseminated dz (imczd); skin & soft tissue infection Mycobacterium avium complex (MAC)—M. avium + M. intracellulare; pulmonary dz; acquired from environment Older white men w/ COPD & EtOH or tobacco use; prior Hx of TB or CF Lady Windermere syndrome—older non-smoking female w/ interstitial lung infiltrate, multiple <5mm nodules in association Shigella—bacillary dysentery; 4 species (sonnei, flexneri, boydii, dysenteriae), mostly S. sonnei in kids <5yo; S. flexneri in sexually-active gay males Structure—Gram(-); nonmotile; lactase(-); acid resistant Transmission—fecal-oral Virulence factors: Pathogenecity island—encodes contact-dependent secretion system → actin polymerization & formation membrane ruffles → bacteria uptake into vacuole Degrades phagosome membrane → actin polymerization on IcsA protein causes unidirectional movement thru cytoplasm Transcytosis to subepithelium → invasion of adjacent cells → lysis of double layer membrane of phagosome 29 IpaB—activates IL-1 converting enzyme (ICE) w/in subepithelial M → secretion of IL-1 & apoptosis of M (thru caspase-1) Shiga toxin—leads to hemolytic uremic syndrome (HUS) Presentation—acute inflammatory colitis, blood diarrhea (WBCs & mucoid), cramps, fever Treatment—3rd-generation cephalosporins & quinolones; reinfection possible Escherichia coli—5 types depending on virulence w/ common plasmid-mediated adherence to epithelial cells Enterotoxigenic (ETEC)—traveler’s diarrhea Transmission—fecal-oral Virulence factors—type 1 pili; long thin pili (colonization factors) I & II (adhesins) for attachment to microvilli; BFP Heat-labile toxin (LT)—5 B subunits & 1 active (A) subunit; leaks out in presence of bile salt, trypsin & low Fe Heat-stable toxin (ST)—binds & activates membrane-bound guanylate cyclase Presentation—watery diarrhea & vomiting Enteropathogenic (EPEC)—20% diarrhea in bottle-fed <1yo in developing countries Virulence factors—NO toxins; BFP; locus of enterocyte effacement (LEE) pathogenicity island for attachement (intimin) & contact-dependent secretion system Attachment to BFP → contact-dependent secretion → activation of PLC-1, ↑IP3 & Ca2+, IL-8 (PMN chemoattractant) → loss of microvilli → pedestal formation Intimin binds translocated intimin receptor (Tir) translocated from bacteria to host cell membrane for intimated attachment Presentation—occasional fever; watery diarrhea; self-limiting; chronic sometimes in infants Enteroaggregative (EAEC)—mucous gel on intestinal mucosa, damage & watery diarrhea; chronic in kids Virulence factors—thin pili (bacterial clumping & attachment), EAST (heat-stable toxin), hemolysin Plasmic-encoded toxin (Pet)—serine protease that alters epithelial cell cytoskeleton Enterhemorrhagic (EHEC)—Shiga toxin-producing E. coli (STEC); O157:H7 causes ~70% cases; ~52 deaths/year in US (absent from third world) Transmission—contaminated ground beef & milk (reservoir in intestine of asymptomatic cows); USDA estimates 90% ground beef contaminated Virulence factors—LEE, intimin & Tir, NO BFP Shiga-like toxins (SLT)—5 B & 1 A subunits, internalized into vacuole, A crosses vacuolar membrane into cytoplasm Removes adenine from host cell 28S rRNA → inhibits protein synthesis → cell death Presentation—lower small & large intestine; watery for 4d followed by blood diarrhea (NO mucous or WBCs) w/in 1-2d (hemorrhagic colitis); resolves in 3-10d Hemolytic uremic syndrome (HUS)—8-11% children <5yo, elderly & imczd; thrombosis of glomerular capillaries → acute renal failure Treatment—feed cows hay prior to slaugher (corn-fed cows shed 100X more acid resistant E. coli) Enteroinvasive (EIEC)—similar to Shigella but less severe; uncommon in US Bordetella pertussis—whooping cough; 4000 cases/year in US (only 10% reported); B. bronchisepticum—kennel cough in dogs; causes 1:1000 cases of whooping cough Bacteriology—small Gram(-) coccobacilli; strict aerobe (5% CO2); small, round, moist colonies on Bordet-Gengou plate Transmission—respiratory droplets Virulence factors: Filamentous hemagglutinin—like pili binds to cilia of respiratory epithelium Pertussis toxin—5 B & 1 A subunit; binds CHO on target cells & prevents normal internal regulation of cyclase Invasive adenylcyclase—interferes w/ chemotaxis & superoxide production Tracheal cytotoxin—invades & kills epithelial cells, induces paroxysms at CNS sites, stimulates bone marrow to produce lymphocytosis Presentation—tropism for ciliated epithelium of trachea & bronchi; 6-20d incubation followed by 3 clinical phases: Catarrhal phase—rhinorrhea for several days Paroxysmal phase—sudden paroxysms of coughing (5-20X in 15s) followed by inspiratory “whoop,” w/ drooling & vomiting for ~2w Convalescent phase—↓paroxysms; “hundred day cough” in Chinese Complications—conjunctival hemorrhage; bronchopneumonia; convulsions → encephalitis (1:9500); meningitis Diagnosis—marked lymphocytosis Treatment—usually NO Abx except w/ severe 2° bacterial infections; recurrence possible but mild Vaccine—formalin-killed whole cell DPT shots for kids <6yo; possible neurological complications (convulsions & blindness) for <6yo Purified acellular vaccine—pertussis toxin & filamentous hemagglutinin, NOT as effective, only for boosters Mycoplasma pneumoniae—20% of all pneumonias usually in 5-15yo Bacteriology—smallest, free-living, self-reproducing organism; dsDNA 1/5 of E.coli (limited metabolic versatility); slow growing & prefer aerobic conditions NO cell wall; triple layer plasma membrane → PCN resistant, poor staining, pleomorphic, fragile, plastic (filterable) Transmission—respiratory droplet; peaks during school season, 4-6y cycle of epidemics Virulence factors—cytadhesion protein binds bronchial epithelial cells & impairs ciliary action; induces desquamation of respiratory epithelium (paroxysmal coughing) Presentation—2-3w incubation; gradual onset of low fever, headache, coryza & pharyngitis, cough (often paroxysmal w/ acute substernal pain), little sputum “Walking pneumonia”—only 10% develop pneumonia, few rales; unexpected, extensive consolidation on chest x-ray; lasts 2-3w Diagnosis—slight neutropenia; cold agglutinins (IgM that agglutinate human type O RBCs) Treatment—tetracycline in adults, erythromycin in kids Mycoplasma hominis—common in GU tract; isolated from blood of 10% post-abortion & postpartum fever women; resistant to erythromycin Ureaplasma urealyticum—inhabits oropharynx & GU tract; isolated in 50-80% non-gonococcal urethritis; urease(+) → alkaline urine (distinguish from M. pneumoniae) Hemophilus influenzae & Neisseria meningitides—along w/ S. pneumoniae cause bacterial meningitis in kids <2yo; extracelullar parasites that avoid innate & humoral immunity H. influenzae—spectrum of pediatric respiratory infections, chronic pulmonary dz in elderly & imczd; often 2 to viral URI Bacteriology—small, pleomorphic Gram(-) rods/coccobacilli, some encapsulated; catalase(+), aerobic & anaerobic; autolyze Grown on chocolate/Levinthal-Fildes agar w/ NAD (Factor V) & protoporphyrin IX (Factor X) & elevated CO2 Major surface Ag—capsular polysaccharide, LOS (shorter sugar chain than LPS) & OMPs Transmission—aerosol w/ seasonal variation; 60-90% healthy kids carry NTHi in nasopharynx Presentation—respiratory tract infection; historically major cause of meningitis in kids 2m-5y but dramatically reduced w/ vaccine since 1987 NTHi—nonencapsulated; usually cause localized infections (e.g. URIs, LRISs, gut) Typeable—cause localized AND systemic dz; URIs, cellulitis, bacteremia, meningitis; characteristic fever due to CK induced by LOS Virulence factors—CHO capsule that reduces complement activation & phagocytosis; determines 6 major serotypes (a-f) Type b—phosphoribosylribitol-P (PRP); major cause of meningitis in US LOS endotoxin—adhesin that paralyzes ciliated respiratory epithelium & induces inflammation w/ IL-1 & TNF Other adhesions—fimbriae & OMPs that bind to specific host cell receptors IgA1 protease—hydrolyzes hinge region of Ab Treatment—Cephalosporins; local infections (NTHi) w/ oral antimicrobials 30 Type b vaccine—type b capsular polysaccharide (PRP) conjugated to protein carrier (diphtheria/tetanus toxoids); eliminates carrier state & prevents invasive dz N. meningitides (meningiococcinia)—exclusively human pathogen that colonizes URT & other mucosal surfaces; 300,000 cases/year (10% mortality) Bacteriology—Gram(-) diplococci; non-motile, non-spore forming; aerobic, catalase(+) & oxidase(+) Virulence factors—polysaccharide capsule; determines 13 serotypes (largest epidemics w/ group A) Types B, C, Y—primary causes of endemic dz in US Type B sialic acid—resembles host (molecular mimicry) host response (poor Ag that doesn’t activate complement) Pili—mediate attachment to non-ciliated pharyngeal mucosal cells IgA1 protease OMPs—mediate adherence & invasion; important in dz spread & prevent Ig-mediated opsonizatoin LOS—stimulates inflammation that may cause fulminating dz, necrosis & shock Transferrin binding proteins—acquiring Fe Transmission—large respiratory droplets; carried in pharynx of 5-15% healthy kids & adults Presentation—highest in kids <1yo & >50yo w/ underlying disease Flu-like illness; failure to clear causes continued bacterial growth endotoxin release causes overt dz (blebbing & CK induction) Colonization of naso-pharyngeal mucosa thru adhesins, pili & OMPs (bacteria modulate adhesin expression to evade immune system) Binding stimulates engulfment & transcytosis; modulate mucosal cell metabolism to ensure survival Meningitis—antecedent URT common Bacteria crosses blood-brain barrier at choroids plexus thru endothelial cell vacuoles, endothelial cell junctions & inflammation due to endotoxin Proliferates in subarachnoid space endotoxin mediates CK activation TNF & IL-1 permeability of BBB neutrophil infiltration → inflammation → complement & coagulation, intracranial pressure cerebral vein thrombosis Presentation—mild URI headache, fever, GI upset, nuchal rigidity, impaired consciousness, Brudzinksi’s sign Fulminant meningococcal sepsis (FMS)—systemic manifestation of meningitis; mortality 20-80% w/ 50% dead w/in 24h of first symptoms Presentation—malaise, weakness, headache, hypotension, prostration shock & disseminated intravascular coagulation (DIC) Release of elastase acute respiratory distress syndrome (ARDS) & other proteinases from neutrophils endothelial cell damage CK induction (TNF, IL-1, IL-6 & IL-8) induces NO vasodilation Skin hemorrhages characteristic of invasive meningococcal dz Effects ALL organs, especially adrenals (Waterhouse-Friderichsen syndrome) Endotoxin & CK induces vasculitis & activation of Factor XII (Hageman factor) Diagnosis—history & presentation, lab tests confirm Treatment—high dose IV PCN for N. meningtitidis Vaccine—effective in adults, control epidemics in military (NOT group B); do NOT reduce carrier state, NOT recommended for routine use Streptococcus pneumoniae: Structure—Gram(+) diplococci in chains; catalase(-), -hemolytic, bile soluble (autolysin), optochin sensitive, latex agglutination Transmission—aerosol w/ mid-winter peak; colonization in nasopharynx Virulence factors—capsule—elicits strong, type-specific Ab response (quelling reaction); induces C-reactive protein; change capsules by expressing different genes Adhesin NOT characterized, possibly PsaA/choline binding protein GlcNAc 1-3 Gal—adherence to oral epithelium & type II pneumocytes Platelet-activating factor (PAF)—upregulated by inflammation Pneumolysin (Ply)—cytotoxin released w/ cell death for growth & spread; covers Ab or C3b opsonizers to prevent phagocytosis & prevents C3bBb formation Binds cholesterol creates pores in PMNs & respiratory epithelial cells activates classical complement induces IL-1 & TNF- synthesis inhibits PMN oxidative burst reduces mucociliary escalator Presentation—usually asymptomatic & transient; major cause of URIs (sinusitis, otitis media), pneumonia (15-25% of community-acquired, 5-7% mortality) Bacteremia (19% mortality), meningitis (40% mortality), & occassionally infects heart valves, conjunctivae, joints, & peritoneal cavity Otitis media—children 4m-6y; also NTHi & Moraxella catarrhalis; viral URI predisposes Sinusitis—persons w/ allergies or viral URIs; mucosal congestions creates obstruction that prevents bacterial clearance Pneumonia—aspirated into lungs (requires additional viral RTI, allergies or compromised innate immunity) Teichoic acid & PG stimulate endothelial cells to synthesize CKs (IL-1 & TNF-) & ↑PAG for adherence; w/ pneumolysin activate complement Proliferation in alveoli (alveolar M ineffective) influx of fluid & PMN inflammation w/o bacterial clearance Meningitis—most common cause; crosses BBB via choroids plexus → inflammation in subarachnoid space (C5a, TNF, IL-1 & IL-6) Hearing loss, blindness, drowsiness, seizures & coma Treatment—PCN standard (25% resistant in 1998) w/ altered penicillin binding proteins (PBPs) from other strep strains Pneumonia—PCN, macrolides, quinolones; meningitis—vacomycin + -lactam Prevention—CDC recommends vaccination for at-risk adults Vaccine—capsular polysaccharide from 23 most common serotypes (Pneumovax or Pnu-Imune); infants don’t repond & less in elderly Conjugate vaccine—for infants & kids (Prevnar); 7 serotypes w/ mutant diphtheria toxoid carrier; prevents invasive dz & reduces otitis media; four dose regimen Subunit vaccines—surface protein Ag, under development Neisseria gonorrhoeae—1million cases/year in US; 30-50% asymptomatic females & 5-10% males Bacteriology—Gram(-); “kidney bean” shaped diplococci; aerobic; nonmotile; fragile; require 5-10% CO2 for growth; obligate human pathogen N. meningitidis—oxidase(+), glucose(+) & maltose(+) N. gonorrhoeae—maltose(-) Transmission—sexually transmitted Virulence factors—attachment to nonciliated cells & damage to ciliated cells that slough off from mucosa OMP—attachment to lutropin (hCG/LH) receptor on endometrial cells; induces NF-B & CK expression Outer membrane porin (OMP1)—1A on most invasive strains & IB strains that cause localized infections Opa proteins (PII)—adherence b/w gonococcal cells, attachment & invasion of epithelial cells & PMNs; phase & antigenic variation Binds pyruvate kinase; source of energy & induces expression of sialyltransferase PIII (Rmp)—invasion of epithelial cells, NOT PMNs Pili—attachment to CD46R on epithelial cells; phase & antigenic variation LOS—major toxin; undergoes sialation by pyruvate → serum resistance & dissemination (genetic defect in MAC); attachment & invasion; antigenic variation PG—inflammation & tissue damage IgA1 protease—cleaves IgA1 & LAMP-1 (lysosomal associated membrane protein) → prevents vacuole maturation Presentation—oropharyngeal infection (pharyngitis, cervical adenitis) OR anal infection (rectal discharge, bleeding) Male genital infection—urethritis, discharge, dysuria → epididymitis or prostatitis 31 Female genital infection—cervicitis, discharge, dysuria, abdominal pain, menstrual abnormalities → endometriosis, PID Scarring & stricture of fallopian tubes → ectopic pregnancy & infertility Dissemination—endocytosis by nonciliated cells → thru BM → dissemination by serum resistant strains (genetic defect in MAC) → bacteremia Meningitis, endocarditis, most frequently purulent arthritis (elbows & knees) Gonococcal conjunctivitis—possible blindness w/ characteristic influx of PMNs LOS activates complement & induce TNF-, IL-8, IL-1, GM-CSF by M & PMNs Secrete IL-6 & IL-8 in urine Diagnosis—Gram stain of discharge (diplococci w/in PMNs) & culture ASAP Treament—most PCN-resistant; 3rd-generation cephalosporins; NO vaccine b/c extensive phase & antigenic variation w/ molecular mimicry Spirochetes—highly motile (thru viscous fluids) w/ flagella b/w inner & outer membranes, highly antigenic & highly conserved; white Oral treponemes—normal flora of mouth associated w/ periodontal dz; T. denticola, vincentii, socranskii; Ab to oral treponemes give false(+) for other spirochetes T. vincentii—causes Vincent’s angina (“trench mouth”) Pathogen-related oral spirochetes (PROS)—never been cultivated Virulence factor—protease dentilysin Syphilis—Treponema pallidum Bacteriology—thin spirochetes; humans only natural host; NOT cultured, only transiently in rabbit testicles Transmission—primarily thru sex; OR contact w/ skin lesions of 2 syphilis, congenital infection, or blood transfusion Virulence factors—very few proteins on surface once establish infection OMP—bind ECM, laminin, fibronectin, collagen T. pallidum rare OMP (Tromp1) Tpr proteins—function unknown, but Ab to Tpr correlate w/ resistance to reinfection Presentation—symptoms due to immune response, NOT toxins Primary syphilis—bacteria penetrate mucous membranes or thru skin abrasions disseminate w/in hours (slow replicating) Primary lesion (chancre)—appears at site of inoculation; painless; begins as papule; heals spontaneously after 2-6w Oral chancres mistaken for other types of sores; vaginal & anal chancres often missed Local lymphadenopathy w/in 1w (hard, nonsuppurative, painless) Secondary sphylis—lesions on skin & mucous membranes (syphilides); 5-10mm diameter; macular, papular or pustular; w/ constitutional symptoms ~25% unaware; may enlarge & erode (moist, pink condylomata lata) in warm, moist areas (highly infectious) Complications—iritis (pain, photophobia), hepatitis, neuropathy, GI upset, arthritis Latent syphilis—asymptomatic but infectious (congenital); w/in 1y of primary infection (~90% relapses w/in first year) or late latent (w/in 4y) Tertiary syphilis—neurosyphilis → optic lesions Meningeal syphilis—may resemble stroke Paresis—changes in personality, affect, reflexes, eyes (loss of pupil reflexes), speech, intellect, & sensorium Cardiovascular tertiary syphilis—rare w/ ABCs; 10-40y after infection Late benign syphilis—formation of gumma; may heal spontaneously w/ scarring; very destructive; painful in bone & deep tissue Congenital syphilis—infection any time during pregnancy; 75-95% transmission w/ early untreated syphilis (~35% w/ later latent) Lesions appear around 4mo; may result in spontaneous abortion, pulmonary hemmorhage, hepatitis, or secondary infection Diagnosis—dark-field microscopy of lesion exudates; Reagin serologic test for nontreponemal Abs induced during infection (VDRL, TRUST) specific Ag Treatment—traditionally massive doses of mercury & arsenic; now PCN is DOC Jarisch-Herxheimer reaction—sudden release of endotoxins as bacteria killed; possibly fatal; accompany Tx w/ hydrocortisone & acetaminophen Endemic treponematoses—rare in US Endemic syphilis (betel)—T. pallidum subspecies epidemicum; focused in Eastern tropics, transmitted thru casual contact Presentation—oral lesions common, skin lesions rare; few late complications Yaws—subspecies pertenue; focused in Caribbean, South America & Africa; transmitted skin-to-skin → nondestructive skin lesions Pinta—T. ca;rateum; focused in West; transmitted skin-to-skin → skin lesions (pintides) initially red & turning darker w/ sun exposure Leptospirosis—Leptospira interrogans Transmission—many animals carry asymptomatically & shed in urine; enters human thru skin or mucous membranes Rat urine in inner cities; coal mines; contaminated lakes; rainy season in tropical areas Presentaiton—many asymptomatic; infection primarily of kidneys & liver Anicteric phase (90%)—flu-like symptoms; occasional meningitis; resolves w/in 1-3w Weil’s syndrome (10%)—severe leptospirosis w/ jaundice, renal dysfuntion, spontaneous bleeding; high mortality Treatment—oral -lactams or tetracyclines; IV Abx in severe cases Lyme disease—Borrelia burgdorferi; 18,000 cases/y in US, mostly in Northeast & around Great Lakes Transmission—ticks (lxodes scapularis (dammini) in eastern US (deer tick) OR l. pacificus in western US (black-legged tick); also transmit Anaplasma (Ehrlichia) & Babesia No transovarial transmission; tick larvae feed on infected mice molt into nymphs transmit to other mice (deer serve only as food source, no role in cycle) Non-endemic areas—ticks feed on non-competent animals (e.g. lizards) VS. endemic areas—nymphal feedings precedes larval feeding Bacteria enter midgut migrate to salivary glands w/in 2d (tick removed w/in 48h unlikely to transmit infection) Virulence factors—surface proteins synthesized during mammalian infection: DbpA & B—decorin binding proteins for adherence p47—fibronectin binding protein for adherence Erp protein family—bind complement Factor H & inhibit alternative pathway H/FHL-1 binding surface proteins (CRASPs) BmpA (p39)—unknown function, but highly conserved & diagnostic Ag OspC (p24)—transmission VlsE—varied expression during infection Presentation—due to immune system response; initial flu-like illness → rarely fatal due to heart block (<12 confirmed deaths since first recognized in 1970s) Erythema migrans (EM)—“bulls eye” rash >5cm around bite; diagnostic for Lyme dz 2 symptoms—weeks or years later; NO evidence that causes AD, MS, ALS, SLE or other chronic dz Inflammatory arthritis—knee or elbow most common, normally unilaterally Neuroborreliosis—Bell’s palsy (highly suggestive of Lyme dz), meningitis, encephalitis Cardiac problems—AV nodal block, myocarditis, pericarditis Acrodermatitis chronicum atrophicans (ACA)—patches of scaly, flaky skin; more common in Europe Diagnosis—presence of EM; Hx of tickbite associated w/ characterisitic manifestations Serologic testing—only as part of dx of probable cases; many false(+) b/c cross-reactivity due to oral treponemes & conserved heat-shock proteins 32 ELISAs—simple & cheap, but high false(+); must follow w/ confirmatory Western blot Skin biopsies—from edge of EM; sometimes cultured; difficult & expensive PCR—of tissue biopsies; sensitive but inaccurate Treatment—very sensitive to -lactams & tetracyclines; 3-4w oral therapy or >1m IV Abx for persistent infections Naturally resistant to rifampin, sulfonamides, phosophomycin Vaccine—taken off market in 2002; recombinant form of OspA OspA Ab do NOT develop in animals infected, ONLY in late-stage human infections; OspA also shut off expression during transmission High titers must constantly be maintained (frequent boosters) & only 1 bacterium needs to survive & transmit to cause infection Southern tick-associated rash (STAR or Master’s dz)—outside endemic Lyme dz areas Presentation—erythema migrants-like rash surrounding bite Diagnosis—NO serologic evidence of B. burgdorferi infection (speculated non-Lyme dz Borrelia); unculturable; possibly B. lonestarii Tick-borne relapsing fever (endemic RF)—primarily in southwest US; several Borrelia species Transmission—Ornithodoros tick (fast feeders, feed multiple times during life) Pathology—bacteria migrate to tick salivary glands w/in days transmitted as soon as begins feeding replicate to high levels in host blood cycles Presentation—characterized by periods of fever & normalcy (like malaria) w/ different surface protein (variable membrane protein, VMP) each relapse Louse-borne relapsing fever (epidemic RF)—B. recurrentis; endemic in Ethiopia & Peru; more fatal than TBRF Transmission—human body louse crushed & fluids enter wound in skin Treatment—tetracycline preferred over -lactams b/c quicker clearance of bacteria Rickettsiaes—obligate intracellular pathogens Rocky Mountain spotted fever—Rickettsia rickettsii; 500-1000 cases/y in US, mostly eastern & southern Transmission—transovarially in ticks but gradually lose viability, need vertebrate host to restore viability Tick saliva dissemination thru blood attachment to epithelial cells via OmpA induce phagocytosis escape phagosome & replicate in cytoplasm spread to other cells via actin tail filament propulsion & cell lysis local vascular permeability localized edema & hemmorhage organ failure & death Virulence factors—OmpA (adherence) & capsule (infectivity) Presentation—initial flu-like illness; progresses rapidly (20-25% mortality before Abx, now ~5%) Rash—appears 50% by 3d (10% never have rash) first on wrists & ankles & spreading over whole body; macules to petechiae → gangrene Diagnosis—serologic simple but (-) for week or more; latex agglutination; immunofluorescence (most accurate & sensitive); Weil-Felix test obsolete & inaccurate Culture NOT done b/c risk to lab workers; immunohistochemistry; RMSF triad (exposure to ticks, flu-like illness, rash) Endemic typhus (flea-borne typhus)—R. typhi & R. felis; found in southern Texas & California Transmission—flea or rat bites, or inhalation of flea feces Presentation—initial flu-like illness; rash in 50% → nonproductive cough, pulmonary densities (ventilation in 10%) Epidemic typhus (louse-borne typhus)—R. prowazekii Transmission—human body louse, poor hygiene; epidemics often w/ war, refugee camps; flying squirrel fleas in eastern US Presentation—rash begins on upper trunk by 5d & spreads to rest of body (except face, palms & soles), may progress to petechial CNS abnormalities (confusion & stupor), cough (40% untreated mortality) Brill-Zinsser disease—milder form of epidemic typhus that occurs years after primary infection, often in elderly, generally NOT fatal; NO rash or headache Rickettsialpox—R. akari; ~100 cases/y in US; transmitted by mouse bite; flu-like symptoms w/ macular rash lasting 6-10d; NOT fatal Scrub typhus—Orietia tsutsugamushi; transmitted by chiggers in Asia; maculopapular rash that fades rapidly Ehrlichiosis: Human monocytic ehrlichiosis (HME)—Ehrlichia chaffiensis; spread by Lonestar ticks Human granulocytic ehrlichiosis or anaplasmosis (HGE)—Anaplasma phagocytophila; spread by lxodes ticks Presentation—intial flu-like illness; rashes rare; leukopenia & thrombocytopenia common; 2-5% mortality Morulae—clusters of cells w/in vacuole of respective leukocyte sometimes seen Q fever—Coxiella burnetti; spread by ticks, contaminated meat, hides or wool; spores resist desiccation & persist in soil Presentation—initial flu-like illness → common pulmonary involvement & thrombocytopenia → death rare Tick paralysis—reaction to substance in tick saliva w/ recovery after tick removal; paralysis may involve entire body death Anaerobes—first organisms (cyanobacteria?) used little or no water; produced O2 to allow facultative & aerobic organisms to grow; outnumber aerobes 1000:1 in GI, 5-10:1 on skin Pasteur (1860)—revealed essential role of anaerobes in producing wine & beer; fermentation (anaerobiosis) Bacteriology—Gram(+) bacilli—Clostridium & Bifidobacterium Gram(-) bacilli—Bacteroides (95% of normal human fecal flora), Prevotella (commensal in GU & GI tracts), Porphyromonas, & Fusobacterium Function—source of vitamins & CHO; keep pathogenic bacteria in check (“wellness bacteria”) Before WWII, most knowledge limited to dz produced by spore-forming, toxin-producing anaerobes Classification—based on response to oxygen; obligate & strict anaerobes do NOT produce enzymes (SOD, catalase, peroxidase) that degrade ROS Obligate aerobes—require O2 (e.g. M. tuberculosis) Facultative anaerobes—fermentation OR respiration (e.g. Staph, E. coli) Aerotolerant anaerobes—fermentation ONLY in presence or absence of O2 (e.g. Actinomyces) Microaerophilic—inhibited by atmospheric concentration of O2 (e.g. Campylobacter) Obligate anaerobes—ONLY in absence of O2 (e.g. Clostridium tetani) Strict anaerobes—killed by 10min exposure to O2, found mainly in cow rumen Percentage of O2 present—crude system of measurement <10% (Campylobacter), <6% (Bacteroides), <3 (Prevotella), <1% (Clostridium), <0.1% (Treponema microdentum, spirochete found in dental plaques) Oxygen-reduction potential (eH)—most precise method of measurement; eH inversely related to distance form oxygen-supplying RBCs 100% oxygen = +800mV & 0% oxygen = -400mV; well vascularized tissue +125 to +250mV (anaerobes do NOT grow at these O2 levels) Dental plaques, gingival pockets, tonsillar crypts, nasal sinuses, colon & vagina—obligate & strict anaerobes (<-300mV) Diagnosis: Thioglycolate medium—tube of semisolid agar w/ reducing agent for growing C. perfringens (+100eH) GasPak—air-tight sealed glass jar w/ internally generated H2 (relative anaerobic atmosphere), -50 to -150eH Glove box & roll tubes—steady stream of N2 flushes out O2; standard lab method today Culture—C. perfringens produce double zone of hemolysis on BAP; Prevotella melaninogenica produce darkly pigmented (black or “melanino”) colonies Gas chromatography—unique collection (fingerprint) of volatile gases, soluble (CO2, ammonia, H2S) & insoluble (H2, CH4, SO2) Anaerobic infections—normally commensal; cause tissue necrosis that produce foul-smelling (fetid) gases Conditions for infection—escape from normal habitat, oral Abx suppress other enteric bacteria, reduced O2 in tissue (metabolism of local facultative organisms) 33 Depressed immunity, impaired local blood supply Virulence factors—enzymes (e.g. lecithinase), capsule, endotoxins (LPS), short chain FAs Intra-abdominal infections—contamination of peritoneal cavity w/ feces (surgery, wounds, inflammatory bowel dz), salpingitis Bacteroides fragilis (95% of normal fecal flora), Clostridium, Fusobacterium, Peptococcus & Peptostreptococcus OB/Gyn infections—septic abortions, salpingitis, surgery, endometriosis; Prevotella Pulmonary infections—aspiration, pneumonia, lung cancer, PE, underlying dental infections; Bacteroides fragilis (20%), Porphyromonas, Fusobacterium Head & neck infections—periodontal & peritonsillar abscesses, chronic sinusitis, otitis media, mastoiditis → brain abscesses; Actinomyces Soft tissue infections—bites (especially human), pressure ulcers, pilonidal cysts, Ludwig’s angina (infection of floor of mouth) Septicemia—most common after abdominal or pelvic infections (25-35% mortality); endocarditis of artificial heart valves (Propionibacterium acnes); Strep + Staph Mixed infections (polymicrobic)—bacterial synergy; use of local O2 by facultative, secrete mutual GFs as byproducts, resistance to PCN b/c local -lactamase production Aspiration pneumonia → lung abscesses Vincent’s angina—acute necrotizing ulcerative gingivitis (ANUG)—Fusobacterium + Furospirochete Ludwig’s angina—anaerobe + GAS Treatment—Metronidazole for intra-abdominal infections & high dose PCN G for oral & pulmonary infections Clostridia—C. difficile (pseudomembranous enterocolitis), C. perfringens (myonecrosis & gastroenteritis), B. cereus (mild food poisoning from old rice) Bacteriology—obligate anaerobic, spore-forming (C. perfrigens aerotolerant & NOT spore-forming); ferment sugars; produce acid & gases (blebs → crepitation) Myonecrosis (gas gangrene)—C. perfringens (60-90%), C. novyi, C. septicum; contaminated wounds or surgery w/ impaired blood supply Pathology—tissue destruction by -toxin (lecithinase C) that degrades lecithin in cell membranes → lysis Presentation—local edema & pain, discolored skin, crepitation, fetid odor Treatment—extensive debridement; Abx rarely cure infection; possibly hyperbaric oxygen Anaerobic cellulitis—bacteria spread thru subcutaneous tissue along fascial planes b/w muscles VS. necrotizing fascitis—Strep or Staph infection, NO crepitation, characteristic anaesthesia; often rapidly fatal, requires aggressive debridement Food poisoning—Abx-associated colitis due to overgrowth of C. difficile Pathology—enterotoxins A & B cause granulocytes to release CKs → intestinal mucosa damage → excess fluid loss Clostridial gastroenteritis—C. perfringens; 3rd most common cause of food poisoning Transmission—spores heat-resistant, ingestion of meat containing preformed toxins (heat-labile) Toxin C—more severe illness, occasionally produces necrotizing jejunitis Presentation—8-24h incubation followed by nausea, abdominal pain & diarrhea (rarely vomiting), fever & headache; resolves in 12-18h C. botulinum—mortality 32% for type A, 17% for type B, 40% for type E Transmission—most heat-resistant but heat-labile spores; intoxication of preformed toxin in home-canned foods; Japan from sushi; or wound infection Botulinum toxin—neurotoxin that blocks ACh release; types A, B, E most common in US (B in KY) Presentation—12-36h incubation followed by severe nausea & vomiting (NO diarrhea), constipation, weakness, rarely fever CN palsies—diplopia, dilated or fixed pupils, dysphagia & dysphonia Symmetrical descending paralysis of large muscle groups → sudden respiratory paralysis → death Treatment—polyvalent botulinum antitoxin; supportive care; Abx of some use if toxin still being produced Infant botulism—“floppy baby syndrome” in 1-2mo Presentation—initial weakness of neck, descending → paralyzed tongue → suffocation or paralysis of diaphragm → death Botox—toxin lasts several months; for correcting strabismus or smoothing out facial wrinkles C. tetani—tetanus Transmission—spores ubiquitous in soil; contaminated puncture wounds, burns, compound fractures, infected umbilical cords Tetanospasmin—neurotoxin prevents release of inhibitory (GABA) transmitters, allows contraction of more powerful opposing muscle Presentation—4-8d incubation followed by muscle stiffness → spasm of masseters (lockjaw) → repeated brief tetanospasms (arching of back or opisthotonos) Flexion of arms & extension of legs → aspiration pneumonia or respiratory failure → death Treatment—immune human globulin administered locally & debridement; large dose PCN, muscle relaxants; supportive care Prophylaxis—tetanus toxoid, 4 injections in infancy & boosters every 10y; transient passive immunity to newborn from immunized mother Zoonoses—30 transmitted by domesticated animals Bacillus anthracis—1st dz-causing bacterium cultured & isolated by Koch from sheep spleen fragments Bacteriology—box car shaped Gram(+) bacillus, chains; facultative; forms spores (ONLY aerobically) Transmission—endemic in cattle & sheep; farmers, vets & slaughter house workers; spores brought to surface by worms → contaminate plants Virulence factors—capsule polypeptide anti-phagocytic (Ab to capsular Ag NOT protective); exotoxin of 3 components: Protective Ag (PA)—barrel-like structure binds LF & EF Edema factor (EF)—reacts w/ calmodulin to generate excess cAMP → prevents CK release from M Lethal factors (LF)—impairs CK production & release from M Presentation—death from sudden release of massive amounts of CKs Cutaneous anthrax—malignant pustule Woolsorter’s disease (inhalation anthrax)—up to 58d incubation period followed by initial flu-like illness Progressing to fatal hemorrhagic mediastinitis, lymphangitis & meningitis Enteritis—fatal enteritis & peritonitis IV—contaminated heroin in N. Europe Treatment—Ciprofoxacin, Doxycycline, PCN; permanent immunity w/ Ab to protective Ag of exotoxin Human vaccine—precipitated part of exotoxin produces short-lived immunity; mandate for all US military Listeria monocytogenes—450 deaths/year US (300X in AIDS patients) Bacteriology—small Gram(+) coccobacilli; aerobic to microaeorophilic; grows 3-45°C, in high salt solution, b/w pH 4.1-9.6; motile 20-25°C; -hemolytic Catalase(+), slow growing; facultative intracellular (liver M & hepatocytes) w/ 90% killed in phagolysosomes; forms palisades Transmission—infects >27 animal species; food-borne (milk, coleslaw, soft cheese, ice cream, hot dogs), placental, OR sexual Enter M cells → lyse primary vacuole → assemble actin tail → migrate w/in cell → infect adjacent cells → lysis of double membrane Virulence factors—complement receptors mediates phagocytosis by M Adhesins (internalins)—InlA binds E-cadherin receptor on GI epithelium, InlB binds hepatocytes Listerliolysin O (LLO) & PI-PLC—lyses primary vacuole, activated by acidification PC-PLC—lysis of double-membrane vacuole after cell-to-cell spread Metalloprotease—Zn-dependent activation of PC-PLC Act-A—surface protein that forms actin tail PrfA—positive regulator of other virulence factors; regulated by T Presentation—tropism for placenta & CNS → neonatal meningitis (10% of neonatal meningitis), sepsis, encephalitis 34 Imczd (50%)—opportunistic infections causing meningitis & endocarditis Typhoidal listeriosis (25%)—generalized infection w/ high fever Angina (8%)—sore throat w/ monocytosis Oculo-glandular form (7%)—purulent conjunctivitis Pregnant women—crosses placenta in 3rd trimester; asymptomatic to severe, usually self-limiting flu-like Neonates—3-50% mortality Early onset—infected in utero → stillborn or premature w/ septicemia → granuloma infantisepticum (focal liver necrosis) Late onset—meningitis d-w after birth, lower mortality Diagnosis—slow-growing culture & elevated agglutinin titer Treatment—clearance by 8d → long-lasting immunity; PCN; 30% mortality despite ampicillin + aminoglycoside (90% if untreated) Erysipelothrix rhusiopathiae—erythros (red) + pella (skin) + thrix (hair) + rusio (seize) Bacteriology—Gram(+) bacillus; survives long periods in dry state Transmission—decaying organic matter; punctures w/ infected fish bones OR wood splinters Presentation—cutaneous infection (purple-red lesions) → deep red advancing edge w/ local burning & pain → rare septicemia Treatment—PCN Yersinia pestis—Bubonic plague; 3 world pandemics (542, 1347, & 1894) Bacteriology—short, plump Gram(-) bacillus; bipolar staining (safety pin appearance); slow growing Transmission—rat flea vector; reservoir in black house rat (roof rat) & gray sewer rat; Sylvatic plague first noted in W. US (prairie dogs, rabbits & squirrels) Blocks intestine of flea → regurgitate during feeding → bacteria (ONLY need single bacterium) reach local LN → multiply in monocytes Virulence factor—capsular Ag anti-phagocytic; endotoxin (LPS) Protein Ag—acquired (phenotypic conversion) at 37°C in mammal (NOT 28°C of normal flea) Presentation: Bubonic plague—2-8d incubation following flea bite → regional LN swelling (bubo) → death w/in 10d Pneumonic plague—inhalation of aerosol → 1-6d incubation period → sudden high fever → septicemia → pneumonia Characteristic hemoptysis VS. inhalation anthrax Disseminated intravascular coagulation (DIC)—intravascular thrombi, purpura, gangrene of distal digits Treatment—Streptomycin or Tetracycline Francisella tularensis—tularemia or rabbit fever Bacteriology—small Gram(-) cocci or bacillus (older cultures); facultative intracellular; slow growing Transmission—endemic in US wild rabbits Presentation: Ulceroglandular form (75%)—tick, deerfly, louse or mosquito bite OR broken skin → regional bubo Typhoid tularemia (25%)—undercooked rabbit meat Oculoglandular form (5%)—infection of conjunctiva Inhalation tularemia—inhalation of aerosol, septicemia, spread from local LN → 3-4d incubation → acute febrile illness → pneumonia Diagnosis—serologic testing Treatment—Streptomycin; prophylaxis w/ Doxycycline or Cipro; live-attenuated vaccine Brucella: Bacteriology—small Gram(-) cocci; facultative intracellular; slow growing Transmission—goats & sheep (B. melitensis), cows (B. abortus), pigs (B. suis); broken skin (90% US cases), milk, conjunctiva OR inhalation Tropism for cattle mammary glands & uterus b/c high concentration of erythritol (NOT in human tissue) Presentation—weeks-months incubation followed by remittent low-grade fever (undulant fever) → weakness & fatigue → vague chronic course Occasionally jaundice, splenomegaly & lymphadenopathy Treatment—Tetracycline for 6w; brucellergen nucleoprotein used as skin test for prior exposure Rat bite fever—normal flora of rat nasopharynx Streptobacillus moniliformis—more common; rat bite → 3-10d incubation → painful & red site → adenitis & fever → acute arthritis (1 or more joints) Bacteriology—pleomorphic Gram(-) w/ yeast-like swellings Spirillum minus—Soduko fever in Japan; 1-3w incubation after rat bite → lesion swells (purple & painful) → chancre-like ulcer → local adenitis & relapsing fever Bacteriology—spiral-shaped Gram(-) bacillus, flagellated Cat scratch fever—Bartonella henselae Presentation—papule at scratch site → local lymphadenopathy → malaise & low-grade fever → rare encephalitis & follicular conjunctivitis Bacillary angiomatosis—skin condition characterized by proliferation of vascular endothelial cells; imczd patients ONLY Cat bite fever—Pasteurella multocida (multi killing) → hemorrhagic septicemia (dogs & cats), shipping fever (cattle), fowl cholera Bacteriology—small Gram(-) coccobacilli, non-hemolytic Presentation—cat bite → lymphadenitis → septicemia → rare meningitis, endocarditis, osteomyelitis Treatment—suturing of bites NOT recommended b/c creates good environment for growth of anaerobes Biological warfare—1930-40s Japanese army doctors testing Chinese prisoners; 1750s French giving smallpox-infected blankets & clothes to Indians 1347—Tartar troops catapulted plague-infected corpses into besieged Kafka 1940s—CA coastal cities sprayed w/ S. marcescens; 1966—S. marcescens dropped onto subway tracks of NYC to test dissemination of aerosol CDC’s 7 most dangerous—inhalation anthrax, smallpox, pneumonic plague, botulism toxin, pneumonic tularemia, filoviruses (e.g. Ebola), & Arenaviruses (e.g. Lassa fever) BW checklist—obtain virulent strain → cultivate in sufficient quantity → disperse effectively (particle size 1-5) → maintain stability → achieve infectious dose per person → facilitate person-to-person spread → overcome prophylaxis & therapy (Abx-resistance) 35 IID EX AM 4 Fungi—NO chlorophyll, usually non-motile, spore-forming, multicellular, discrete nucleus (nonseptate are multinucleated) Hyphae—mycelium (network of hyphae) or thallus (colony of hyphae); vegetative (hyphae growing into substrate) VS. aerial; septate (pores allow exchange) VS. nonseptate Cell wall—thatched network of microfibrils, usually multi-layered; composed of glucans (-1,3), mannans, & chitin (N-AGA, -1,4 unbranched) complexes Plasmalemma—2-layered; composed of lipoprotein & sterols (ergosterol) w/ differential permeability Mitochondria—concentrated in growing tip of hyphae Ribosomes—80S (60S + 40S) Yeast—single-celled w/ budding of daughter cells; some exhibit pseudohyphae (daughter cells fails to separate, forming chain of yeast cells) Dimorphic—ability to grow in either mold (mycelium) or yeast forms Requires ↑CO2, ↑T (35-37°C), organic N source, SH compounds (cysteine), abundance of simple sugars 5 most common—Blastomyces dermatidis, Histoplasma capsulatum, Sporothrix schenckii, Coccidioides immitis, Paracoccidioides brasiliensis Metabolism—molds aerobic & yeasts facultative; heterotrophic & utilize almost any C source (simple sugar, plastic, jet fuel), NOT solely CO2 Nitrate, nitrites or ammonia sole N source Exoenzymes—break down complex molecules for transport across cell wall & plasmalemma Growth factors: Temperature—psychorophilic (0-20°C), mesophilic (30-40°C, most fungi), thermophilic (30-50°C) pH—some yeast grow at 2.0 & molds at 8.5; most fungi b/w 6.5-7.0 optimal Moisture—most fungi require 60-95% relative humidity Light—most molds photosensitive (especially growing tips) Asexual reproduction: Thallospores—spores produced directly from thallus Arthroconidium—spore formed by septation of hyphae & separation at septum Blastoconidium—spore formed by budding (yeasts) Chlamydospore—spore formed by thick-walled intercalary or terminal cell w/in hyphae Exogenous spores—simple to complex conidium, supported by conidiophore; found on septate hyphae Deuteromycetes—NO sexual spore & asexual conidia; septate; e.g. Aspergillus, Trichophyton, Candida Endogenous spores—sporagiospore containing sporangium, supported on sporangiophore; found on non-septate hyphae Sexual reproduction—haploid gametes → plasmogamy → karyogamy (diploid) → meiosis (haploid) → mitosis (haploid) Zygospores—fusion of hyphae to form sexual spore Zygomycetes—sexual zygospore & asexual sporangiospore; non-septate; e.g. Rhizopus, Absidia, Mucor Ascus—contains ascospores w/in fruiting body ascocarp (cleistothecium most common, apothecium, perithecium, OR naked ascus) Ascomycetes—sexual ascospore & asexual conidium; septate; e.g. Pseudoallescheria boydii, Piedraia hortae, Saccharomyces Basidium—contains basidiospores Basidiomyctes—sexual basidiospore & asexual conidium (rare); septate; e.g. Coprinus, Amanita (mushrooms), Filobasidiella neoformans Fungal dz—depends on interaction b/w virulence factors (inoculum size, structure, adherence, enzymes, toxins) & host immunity; typically opportunistic infections Superficial—e.g. Tinea versicolor (skin), Tinea nigra (skin), Black & White Piedra (hair) Tinea versicolor (pityriasis versicolor)—Malassezia (pityrosporium) furfur; Ep—worldwide distribution; all ages; equal b/w sexes Sx—mild, dry skin lesions w/ NO inflammation Dx—mycology (wet prep “meatballs & spaghetti” appearance); culture NOT necessary but requires FAs/olive oil; histopathology Cutaneous—Tinea capitis, corporis, cruris, unguium, barbae, & pedis; Epidermophyton floccosum, Microsporum, Trichophyton Transmission—direct contact (ONLY truly communicable fungal dz), either anthrophophilic (man-to-man), zoophilic, or geophilic (soil-to-man) Microsporosis—Microsporum M. audouinii—anthrophophilic; primarily in kids; Sx—infects hair & skin; Dx—salmon color colony, RARE conidia, NOT grow on rice medium M. canis—zoophilic; usually in kids; Sx—infects hair & skin; Dx—septate hyphae; large, thick, rough-walled, pointed macroconidia; FEW microM. gypseum—geophilic; often in adults; Sx—infects hair & skin; Dx—tan colony; septate hyphae; large, thick, rough-walled macroconidia Trichophytosis—abundant microconidia & few thin, smooth-walled macroconidia T. mentagrophytes—most common cause of athletes foot in US; worldwide distribution; Sx—infects hair & skin, often inflammatory Dx—floccose to granular, abundant microconidia in grape-like clusters; septate hyphae T. rubrum—worldwide distribution, in kids & adults; Sx—infects skin, hair & nails; causes chronic dz Dx—floccose colony w/ non-diffusing red pigment; tear-shaped microconidia T. tonsurans—worldwide distribution; Sx—infects hair (primarily, destroys & causes “black dots”), skin, nails Dx—white felt-like colony; club-shaped “balloon form” microconidia T. verrucosum—zoophilic; in kids & adults; Sx—infects hair, skin, & nails; often inflammatory Dx—white waxy colony; usually NO conidia; grows well at 37°C Epidermophytosis—Epidermophyton floccosum; Ep—anthrophophilic; usually in adults Sx—associated w/ jock itch; infects primarily skin, may be inflammatory Dx—tan to olive-drab (OD), felt-like colony; septate hyphae; club-shaped segmented macroconidia, NO microconidia Subcutaneous—common in soil; transmitted into wounds w/ trauma Chromoblastomycosis (Cladosporium, Fonsecaea, Phialophora)—primarily in tropics Sx—chronic, granulomatous verroucus lesions; often 2° bacterial infections Dx—slow growing, dermatiaceous; histopathology (round, dark-rimmed sclerotic bodies w/in granuloma) Phaeohyphomycosis—numerous genera of dermatiaceous (darkly-pigmented) fungi; worldwide distribution Sx—abscess formation Dx—darkly-pigmented hyphae & colonies Sporotrichosis (Sporothrix schenckii)—worldwide distribution (especially in peat moss); all ages; higher in males (occupational exposure) Sx—ulceration at site of inoculation w/ spread along lymphatics Dx—dimorphic; budding, cigar-shaped yeast, gray/black mold w/ delicate septate hyphae & tapered conidiophores, conidia in “daisy flowers” Histopathology (granulomatous or suppurative); serologic (agglutination); fungus may NOT be in exudates Systemic—primarily pulmonary dz (inhalation of conidia) w/ predictable, self-limiting dz course in immunocompetent Histoplasmosis (Histoplasma capsulatum)—Mississippi-Ohio river valleys; predominately in adult, white, males; grows in alkaline N waste of bird droppings Sx—pulmonary (indistinguishable from other mycoses, TB or other pulmonary dz) Acute (94%)—benign (70%) asymptomatic or mild flu-like (25%), severe epidemic (5%) OR chronic (5%) Extra-pulmonary (1%)—disseminated, always starts w/ pulmonary infection; characteristic oral lesions; acute OR chronic 36 Dx—serologic w/ agglutination (quantitative in acute cases ONLY); ID precipitation (qualitative, detects H&M bands) Complement fixation (CF)—quantitative, titer of >1:8, possible cross-reactivity; immunofluorescence (Ag in urine w/ dissemination) Mycology—dimorphic; direct exam of specimen NOT diagnostic b/c yeast not unique (small, single bud w/ narrow septum) Slow growth (8-21d); large, tuberculated conidia (diagnostic) & small, round, smooth-walled microconidia; septate hyphae Blastomycosis (Blastomyces dermatitidis)—Mississippi-Missouri river valleys; predominately adult, white, males; in soil & decaying organic matter Sx—pulmonary (acute OR chonic) VS. disseminated (ALL organs may be infected) Dx—serology (CF or ID but high false(-), cross-reactivity w/ H. capsulatum); histopathology (suppurative to granulomatous) Mycology—dimorphic; septate hyphae, slender conidiophores; thick-walled, wide bud base yeast Coccidioidomycosis (Coccidioides immitis)—Valley fever; desert Southwest US; adults, ALL races, higher dz in males; found in desert soil Sx—pulmonary & disseminated Dx—serology (Ab in 10-21d, CF, ID, agglutination in acute cases); histopathology (suppurative to granulomatous) Mycology—dimorphic; septate hyphae w/ barrel-shaped arthroconidia; large spherical spherules containing numerous endospores Opportunistic systemic: Cryptococcosis (Cryptococcus neoformans OR Filobasidiella neoformans in sexual state)—also C. terreus, albidus, diffluens, laurentii Ep—found in soil w/ bird droppings Sx—pulmonary (asymptomatic, pneumonia, cavitary, effusion), CNS involvement OR disseminated (meningitis, cryptococcoma) Dx—budding yeast w/ polysaccharide capsule w/ narrow bud base; Mucicarmine stain (capsule appears red) Latex agglutination—detects capsular Ag; highly sensitive for meningitis Candidiasis (C. albicans)—3rd most common nosocomial infection w/ Abx therapy, IV catherization, UT catherization (indwelling), neutropenia Sx—wide spectrum of dz Dx—pseudohyphae in fresh specimens (except C. glabrata); latex agglutination for Candida Ag (NOT sensitive) Aspergillosis (Aspergillus fumigatus, flavus, niger)—grows in 2-3d on most media Sx—allergic, pulmonary, fungus ball, invasive, rhinofacial, cerebral Dx—septate, branched hyphae in specimen (diagnostic of invasion); ID, CF (high OR rising titer), immunofluorescence (galactomannans of cell wall) Zygomycosis (Mucor, Rhizopus, Absidia)—predisposed by DM, hematologic malignancies, severe burns, malnutrition, leucopenia, transplantation, IV drug use Sx—rhinocerebral (most common), pulmonary, GI, disseminated Dx—non-septate, branching, ribbon-like hyphae in fresh specimen; NO serologic; culture NOT grow w/ cyclohexamide, presence of sporangiophores Toxicoses—aflatoxin (hepatotoxin, cancer-causing); trichothenes (dermal necrosis & hemorrhage); ochratoxins (nephrotoxin); vomitotoxins; mycotoxins (mushrooms) Antifungals: Polyene (Amphotericin B, Nystatin)—binds ergosterol & disrupts plasmalemma; systemic & topical Azoles—inhibits ergosterol synthesis; systemic Pyrimidine (5-FC)—inhibits DNA & RNA synthesis; systemic Echinocandins (Cancidas)—inhibits -1,3 glucan synthetase; systemic Grisans (Griseofulvin)—inhibits microtubule assembly; topical Host immunity—mechanical barriers (skin, cilia, mucosa), phagocytosis (PMNs & M), cell-mediated response (delayed hypersensitivity), Ab-mediated response 1. Alveolar M ingest some inhaled conidia (NOT killed effectively) & growth proceeds in alveolar spaces 2. Invasion of intestinal tissue activates alternative pathway of complement → C3b deposition (opsonization) → phagocytosis by M (killing still ineffective) 3. Acute inflammatory response—influx of neutrophils (phagocytose & kill yeast forms) → hyphal forms incompletely phagocytosed → release of hydrolytic enzymes 4. Adaptive immune response—Ab (opsonization of neutrophils) & cellular immunity (TH1 response leads to recovery, TH2 leads to uncontrolled infections) 5. TH1 cells release IFN- → activates M (effectively kills fungi, restrict intracellular growth) 6. Granuloma formation walls off fungi (also hyperplasia, inflammation & suppuration in tissue) >90% develop specific Ab to fungus (NOT effective in clearing), Ab response greater in disseminated infection Cutaneous hypersensitivity—epidimeologic testing for exposure thru DTH skin reaction Strong reaction (asymptomatic infections), weak DTH reaction (active pulmonary infection), NO reaction (anergy, during disseminated infection) Parasites—“one who dines at table of another”; EVERYONE is infected w/ parasites Protozoa—most free living & NOT parasites; classified by morphology & motility (amoeba, flagellates, ciliates, sporozoa, & reclassified organisms) Metazoa—classified by structure; transmitted by direct ingestion (adult worms, larvae, eggs), insect vectors, OR penetration thru skin Nematoda (ingested OR vector); Cestoda (ingested); Trematoda (ingested OR direct penetration); Ectoparasite Vectors—Arthropods often lay eggs into wounds (Insecta, Arachnida, Crustacea) Intermediate hosts—often reservoirs in Mollusca Survival in host—minimize immunity, establish dormancy, adapt to environment (cyst forms); balance b/w damage to host & opportunity for growth & dissemination Vaccine—absence of single virulence determinant; absence of well-defined metabolic pathways; difficulty growing & studying; lack of economic incentive Stealth (low immunogenicity of surface Ag OR intracellular growth); antigenic variation; manipulation of immune system (skew TH1 vs. TH2) Entamoeba, Giardia, Trichomonas—earliest eukaryotes Structure—facultative anaerobes; NO mitochondria; extracelullar (adhere to plasmalemma) w/ contact-dependent cytotoxicity & mechanical damage; binary fission Amebiasis (Entamoeba histolytica)—amoebic dysentery; Ep—predisposed w/ poverty, alcoholism, malnutrition (changes to GI flora); anal sex Life cycle—ingest cysts in feces → 4 nuclei emerge from cyst wall → division (1 round) → progeny divide into trophozoites in intestine (binary fission) → reconversion to 4 nuclei cysts → breach gut wall & travel along portal vein to liver (extraintestinal amoebiasis) Trophozoites adhere to intestinal wall at sites of mucus depletion → spread into LP & submucosa Lyse tissue (hence the name) & neutrophils → further tissue damage (absence of neutrophils around lesions) Sx—2-21d incubation; localized infections (abscess formation, characteristic flask-shaped ulcers) in liver, lungs & brain Intestinal amoebiasis—acute (abdominal pain, bloody mucoid diarrhea, flatulence) OR chronic (MOST asymptomatic; diarrhea, weight loss, fatigue) Extraintestinal—URQ pain, weight loss, fever, hepatomegaly → liver dysfunction Dx—trophozoites (usually laden w/ RBCs) OR 4 nuclei cysts in stool; glycogen granules by PAS stain; relative leucopenia Liver abscesses (brown semiliquid) visualized by CAT or MRI → invasion of lungs thru diaphragm → trophozoites in sputum Tx—Metronidazole (Flagyl, selective for anaerobes); Parmomycin (Humatin & Iodoquinol for asymptomatic carriers) Other intestinal amoebae—classified by morphology of trophozoites & cysts; e.g. Entamoeba, Endolimax nana, Lodamoeba buetschlii, Diantaoeba fragilis Acanthamoeba castellanii—free living; found in soil & fresh water; generally opportunistic; asymptomatic in nasopharynx Sx—1d-2w incubation; targets eye (contact lens of health persons, conjunctivitis, keratitis, uveitis) & CNS (seizures, granulomatous encephalitis or GAE) Dx—presence of large trophozoites w/ smaller cyst forms in axenic culture Tx—Ketoconazole & surgical excision of granulomas; topical Azoles for ocular infections Naeglaria fowleri—found in fresh water & pools; asymptomatic in nasopharynx 37 Sx—1-3d incubation; rapidly fatal primary amoebic meningoencephalitis (PAM) → CSF thick w/ parasite burden Tx—Amphotericin B (limited efficacy & high toxicity) Blastocystis hominis—diarrheal dz; transmission by cysts; Dx—detection of cysts in feces; usually after Tx for bacterial & viral diarrheas; NO curative Tx Balantidium coli—found in pig feces & resident in lumen of colon; Sx—usually asymptomatic, diarrhea, perforation (rare); Dx—detection of cysts in feces; NO curative Tx Giardiasis (Giardia lamblia)—Beaver Fever; most common cause of non-bacterial diarrhea in US (nearly half asymptomatic) Ep—bi-nucleate, pear-shaped flagellate; found in duodenum & upper ileum; predisposed by deficient mucosal immunity & anal sex; cysts removed by filtration Life cycle—excystation in duodenum → release of trophozoites → attach to epithelium via ventral disk → localized effacement of microvilli → cyst formation Express different major surface glycoproteins → failure of mucosal immunity → leukocyte infiltration into LP → chronic dz Sx—3-12d incubation; followed by abdominal cramps, foul-smelling diarrhea (NOT bloody), malabsorption (excessive fat in stool), NO fever Dx—characteristic bi-nucleate “monkey face” trophozoites or 4 nuclei cysts Tx—Metronidazole (Flagyl), Quinacrinehydrochloride (if unresponsive to Flagyl, potentially toxic) Trichomonas vaginalis—adapted almost exclusively to damp, warm environment of human vagina & prostate Ep—highly motile (4 flagella w/ undulating membrane); NO cyst form (relies entirely on human-to-human transmission); strict anaerobes; NO mitochondria Hydrogenosome—unique single-membrane organelle required for energy production; catabolism of pyruvate to acetate, CO2 & H2 Predisposed w/ changes in acidity (resident microflora) of vagina during menstrual cycle, pregnancy, or Abx therapy Pathology—trophozoites attach to vaginal epithelium → desquamation → inflammatory response → leukocyte infiltration Sx—intense vaginal itching & burning; foul-smelling whitish vaginal discharge (leukocytes, metabolic byproducts & H2); asymptomatic in males Dx—presence of trophozoites in vaginal or prostatic secretions; axenic culture Tx—treat ALL partners to avoid ping-pong infections; Flagyl (possible teratogen) Opportunistic parasites—Toxoplasmosis & PCP in homo populations early in epidemic, now indicators of underlying AIDS infection Tachzoite—rapidly growing parasite form associated w/ acute infection VS. bradyzoite—slow-growing tissue cyst form Sporozoite—state of development found in oocyst following sexual reproduction VS. bradyzoites—contained in tissue cyst from asexual reproduction Endodyogeny—mother cell “consumed” by 2 daughter progeny Schizogeny— divisions produce trophozoite (schizonts or meronts) w/ multiple nuclei at periphery as internal membranes separate into daughter cells (merozoites) Endopolygeny—multiple nuclear & organelle divisions w/o cytoplasmic division; progeny form all over cytoplasm Intracellular parasitism: Pros—protected from humoral immunity, nutrient rich & stable environment, dispersed by circulation, usually NO competition Cons—must overcome lysosomes, neutralize ROS & RNS, cell-mediated immunity, need strategy for dissemination Toxoplasmosis (Toxoplasma gondii)—nearly 1/3 world population exposed to parasite, up to 90% in Western Europe Ep—obligate intracellular; warm-blooded animals (intermediate hosts) & felines (definitive hosts); gliding motility (NOT flagella or cilia) Sporozoites found on vegetables, water, sandboxes, grazing animals, steak tartare (raw beef); survive high heat & freezing Sexual cycle in feline GI tract, asexual cycle in other animals producing tachyzoites (acute phase) & bradyzoites (chronic phase) Host immune system (IFN-) limits growth & cues conversion to bradyzoites (NO signals in imczd → more severe disseminated dz) Life cycle—ingest bradyzoites in meat & brain → cysts rupture in feline GI tract → invade tissue → differentiate into tachyzoites → differentiate into male & female gameteocytes → mate & form oocyst → released in feces (up to million/day w/o symptoms) → maturation in soil & water to form 4 sporozoites → humans ingest sporozoites → sporozoa invade target cell (tropism for brain & muscle) → establish in parasite-modified parasitophorous vacuole (PV) surrounded by PV membrane (PVM) → replicate by endodyogeny → trophozoites develop w/in vacuole → infect adjacent cells (including M) Sx—generally asymptomatic; else flu-like symptoms w/ possible lymphadenopathy, slenomegaly, low-grade fever & malaise Cerebral toxoplasmosis—reactivation of brain cysts (cycles of reinfection) → cerebral abscesses → multifocal neurologic symptoms Encephalitis—common in organ transplant patients w/ cysts in organ donation Congenital toxoplasmosis—severity depends on gestational age w/ highest risk b/w 10-24w; most in 3rd trimester unaffected Symptomatic (10-20%)—spontaneous abortion, hepatitis, splenomegaly, pneumonia, rash, anemia, FTT Sabin tetrad—hydrocephalus, retinochorditis, intracerebral calcifications, psychomotor retardation Subclinical (75%); mild w/ retinochorditis (15%); severe w/ brain damage, hydrocephalus, death (10%) TORCH (toxoplasma, rubella, CMV, HSV)—standard prenatal testing Anti-Toxoplasma IgM (recent or active infection) & IgG (resolved or reactivated if rising) Ocular toxoplasmosis—retinochorditis (destruction of retina mediated by parasite growth & inflammatory response) → blindness; maybe 20y later Dx—serologic (immune response against surface Ag (SAG-1 or p30); amniocentesis (parasites in amniotic fluid); PCR of CSF; histopathology of brain Tx—chemotherapy inhibits tachyzoite (limited effect on bradyzoites) growth during reactivation Pyrimethamine+Sulfadiazine—10X more effective w/ synergy; contraindicated in pregnancy (Spiramicin instead) Macrolides—disrupt small genome of the apicoplast, essential organelle in the apicomplexa; NOT widespread clinical use Cryptosporidiosis (Cryptosporidium parvum)—outbreaks due to failure in water purification (fecal contamination), e.g. 1993 Milwaukee that affected 400,000 & killed 50 imczd Life cycle—ingest oocyts (imcpt must consume up to 300) → sporozoites released → attach to intestinal microvilli → invade & form schizont w/in vacuole → schizogeny to form 8 merozoites → infect adjacent cells → some differentiate into gametocytes (multinucleate macrogamont & microgametes) → fertilized gametes develop into oocysts → sporulate to contain 4 sporozoites → released into feces OR autoinfection Sx—abdominal pain, diarrhea, fever for 3-10d (mistaken as bacterial food poisoning); imczd—profuse diarrhea (months), malabsorption, severe weight loss Tx—ALL limited efficacy; support & anti-diarrheals instead PCP (Pneumocystis carinii)—traditionally classified as protozoa, now fungus; most common & rapid death in AIDS patients; >75% US population seropositive by 4yo Ep—until AIDS, only sporadic in malnourished & imczd; inhibits O2 exchange in lungs; diffuse infiltrate on x-ray; CD4 counts predictive of infection in imczd Life cycle—trophozoites attach to non-surfactant secreting lung epithelium → pre-cysts (sporocytes) → cyst (thicker-walled) → spore formation → trophozoites Dx—Type I alveolar cells colonized & Type II (surfactant & mucous producing) cells proliferate PAS stain detect parasites attached to alveolar epithelium; GMS gold standard; pre-cyst & cyst forms stain intensely w/ toluidine blue & PAS Tx—Trimethoprim+Sulfamethoxyazole (TMC-SMX or Trim/sulfa); prophylactic in imczd; Pentamidine IV in acute cases Microsporidia—obligate intracellular; Enterocytozoon bienusi (enteric dz), E. cuniculi & E. hellem (disseminated infections) Sx—enteric, neurologic, ocular, pulmonary infections; Dx—detection of spores in feces; PCR; NO curative Tx Malaria (Plasmodium falciparum, vivax, ovale, malariae)—association w/ swamps (mal aria, “bad air”); >300 million infected worldwide w/ >1 million deaths annually, mostly kids <5yo 1897—Sir Ronald Ross completed life cycle of avian malaria w/ sexual cycle in mosquitos (Nobel Prize in 1902) 90% in sub-Saharan Africa; GDP estimated 32% greater if malaria eliminated 35 years ago, an extra $100 billion; slows Africa’s economic growth by 1.3% annually Ep—transmitted by Anopheles mosquito (other mosquitos cannot complete sexual cycle); evolved in Africa & migrated to Mediterranean; Sporozoan (Apicomplexan) Female mosquitos use blood for production & nourishment of eggs laid in stagnant water; live 7d-several months, laying 4-5 clutches Proboscis injecting saliva prevents clotting & local anesthetic; best defense remains bed nets & repellants applied to clothing (many now DDT resistant) 38 Life cycle—bite releases sporozoites into blood → invades hepatocytes (extraerythrocytic stages) → replication & release of merozoites into blood → invades RBCs (ring stage to trophozoite stage) → shizogeny & release of merozoites → attack new RBCs → some convert to gametes → mosquitos take up gametocytes w/in RBCs → male gamete exflagellates → fertilizes female gamete forming ookinete → transverses midgut wall → develops into oocyst → maturation (10-14d) & release of sporozoites into hemolymph → travel to salivary glands RBC cycle—digest Hgb (polymerization into hemozoin or “malarial pigment”) for anabolic activity Modify cytoplasm (Maurers & vesicular clefts) & plasmolemma (surface knobs w/ adhesive proteins) interact w/ endothelium (cerebral malaria) Sx—classic shivering followed by high fever that coincides w/ synchronous lysis & release of merozoites from RBCs Tertian malaria—48h periodicity of fever; P. vivax, ovale, falciparum Quartan malaria—72h periodicity of fever; P. malariae Dx—blood smear w/ Wrights-Giemsa stain & parasites w/in RBCs (active infection), scored based on staging; species identified based on staining pattern; or PCR P. falciparum—severe malaria, Malaria tropica, malignant tertian malaria Sx—8-24d incubation in hepatocytes; 5-12d to symptoms (anemia); continuous fever, headache, GI upset → rapid-onset coma (cerebral malaria) → death Cerebral malaria—knobs aid binding to endothelium → promote aggregation of infection RBCs → capillary occlusion → coma → death PfEMP1 (P. falciparum erythrocyte membrane protein 1)—family of var proteins that aid binding to endothelium; antigenic variation Pregnancy—premature, spontaneous abortion (after 3m) or LBW; higher w/ 1st pregnancy; targets maternal sinuses of placenta, accumulation of infected RBCs Hepatic—early stages, liver damage → jaundice, characteristic reddish-brown color & hepatomegaly w/ destroyed RBCs in liver & spleen Renal—presence of Hgb in urine → dark colored urine (Blackwater fever) P. vivax—Malaria tertiana, benign tertian malaria; most common form of malaria; Ep—endemic in parts of US in late 19th century Sx—12-18d incubation; 8-17d symptomatic; 5y or more reactivation (more frequent than other malarias); rarely fatal; can cause LBW in pregnancy Distinct tertian patter of high fever preceded by hour of shivering (physical exhaustion); more predictable & shorter duration fever spikes Targets immature RBCs & relatively few reticulocytes (self-limiting) Dx—enlarged RBCs, Schauffners dots (stippled appearance to cytoplasm), mature rings forms appear coarse, distinctive appearance of gametocytes P. ovale—Malaria tertiana; rarest malaria; restricted to western Africa & Pacific; causes tertian fever pattern; 10-17d incubation, 5-7 patent period P. malariae—Malaria quartana; mildest form of malaria; targets older RBCs; 18-42d incubation, 30y patent period (longest) Tx—must know species, site of infection (drug resistant patterns), prior prophylaxis, age & pregnancy status, cost, compliance Quinine & Chloroquine—1600 Peruvian Indians extracted from Cincona tree bark; British introduced for malaria in India Compliance difficult b/c extreme bitterness, gin & tonic created as prophylactic delivery method of quinine DOC for >300y; targets digestive vacuole & blocks inactivation of malarial pigment hemozoin (toxic to parasite); ONLY RBC stages Some Chloroquine-resistant Plasmodia show sensitivity to Quinine; inexpensive; prophylaxis for travelers Chloroquine & Mefloquine—more effective w/ chemical synthesis, but increasing resistance Mefloquine—highly effective; used ONLY in regions resistant to other meds; targets RBC stages Pyrimethamine + Sulfonamides (Fansidar, anti-folates)—combination Tx to avoid resistance to Pyrimethamine thru DHFR mutations Targets RBC stages; NO longer effective in Southeast Asia & South America Proquanil (anti-folates, bi-guanides)—combination Tx to avoid resistance Doxycycline & Macrolides—adjuncts to Quinine for multiple drug resistant P. falciparum; targets apicoplast Arteminisin/Qinghaosu (Sesuuiterpene lactone)—traditional anti-malarial in Chinese medicine; extracted from wormwood tree; very rapid acting Last resort for cerebral malaria; NOT prevent reactivation Atovaquone—targets RBC stages by interfering w/ mitochondrial electron transport & respiration; combination Tx Primaquine—DOC for clearance of P. vivax; prevents reactivation by targeting dormant liver stages Kinetoplastids—possess mastigone (flagellum) w/ life cycle stages defined by flagellar properties; Ag surface proteins for host interaction & evasion of immunity Flagellar pocket (FP)—highly specialized structure in which endocytic & exocytic activities occur Kinetoplast (KI)—associated w/ mitochondria; DNA organized in concatemers (intertwining rings) of maxi & mini circles Trypanosomiasis—African sleeping sickness T. brucei rhodesiense (central & eastern sub-Saharan) & T. brucei gambiense (western & central sub-Saharan), T. brucei brucei (Nagana in cattle) Ep—both fly sexes feed ONLY on blood, during day; attracted to moving targets, scent of sweat & bodily secretions Variant surface glycoprotein (VSG)—main target of Ab response, antigenic variation causing “relapsing parasitemia”; repertoire encoded in genes Life cycle—trypomastigotes (NO surface coat) ingested by Tsetse fly → migrate to midgut → convert to epimastigotes (smooth surface coat) → enter salivary glands → form coat for survival in blood (25-50d cycle) → fly bites human → slender forms alter organization of mitochondria & kinetoplast in blood → rapidly proliferation (binary fission) → conversion to “stumpy” forms Sx—1-21d incubation, progression of symptoms in 3m (slower in T. brucei gambiense, over 9m-2y); chronic infections for years 1st stage—chancre (Winterbottoms sign) or papular to ulcerating lesion at bite site; lasts 2w 2nd stage—bloodstream → replicate by binary fission → immune response (specific Ab) → fever (lasts 1-3w) 3rd stage—lymphatic system (detect parasites in cervical LNs) 4th stage—CNS (headaches, apathy, atrophy, emaciation, apraxia, somnolence, paralysis, coma, death) Dx—detection of parasites in blood smears; serologic Tx—Suramin—effective against both strains; ineffective against CNS involvement (can’t cross BBB); clears bloodstream; side effects (shock, proteinuria) Pentamidine—effective against both (especially gambiense); early hemolymphatic dz (NOT CNS); side effects (pancreatitis, BP drop, renal) Melasoprol (MelB)—for CNS involvement; highly toxic (reactive encephalopathy) requires hospital supervision; pretreat to clear blood stages Eflornithine (DFMO)—inhibitor of parasite ornithine decarboxylase; clinical trials (“awaken” comatose) & available ONLY from WHO Effective ONLY against gambiense; expensive; active ingredient in cosmetic creams to retard facial hair growth Chagas disease (Trypanosoma cruzi)—named after Brazilian parasitologist Oswaldo Cruz Ep—transmitted by Riduviid bug (Triatome or kissing bug) by defecating parasites onto skin of host, scratching promotes infection Life cycle—infection w/ metacyclic trypomastigotes → parasite enters vacuole → lyses into cytoplasm & converts to amastigotes (NO flagella) → phagocytosed & replication repeated OR differentiate into trypomastigotes (flagellated, protein coat) → blood → ingested by Riduviid bug → differentiate into epimastigotes → differentiate into metacyclic trypomastigotes in bug rectum Sx—5-20d incubation; initial infection at bite site, adjacent muscle & fat cells in dermis forming inflammatory nodule chagoma (Romanas sign), persists for weeks Spreads to lymphatics → cardiac mycoytes, esophagus, colon; Symptomatic period 1-2m; asymptomatic parasitemia 20-40y Cardiac—infection of pericardium & destruction by intracellular amastigotes → microabscesses → healing by fibrosis Profound hypertrophy, dilation, weaking, damage to nerves (arrythmias), hypotension → cardiac arrest → death GI—destruction of autonomic ganglia → disrupt peristaltic function → megaesophagus & megacolon Encephalitis—most often in kids → death in 1-2m Pregnancy—abortion or death w/in few days Chronic infection—minor skin & adipose lesions → lipochagoma or lipo granuloma (painful) on face Dx—serologic; blood smears; xenodiagnosis using uninfected Riduviid bugs Tx—Nitrofuran derivatives w/ IFN---reduces acute phase, targets amastigotes & blood trypomastigotes; side effects common 39 Benznidazole—DOC for patients in Brazil; targets both amastigotes & trypomastigotes; side effects common Allopurinol—purine analog (hypoxanthine) w/ some effectiveness Leishmaniasis (Leishmania donovani)—named after William Leishman & Charles Donovan who first identified amastigotes in M Ep—primarily zoonotic dz; transmitted by phlebotomous sandflies (Phelbotomus & Lutzomyia) Life cycle—amastigoes ingested by sandfly → differentiate into promastigotes → bind midgut & proliferate → blockage of gut forces parasite to mouth → injected into human → infects M → differentiate into amastigotes → repeated division → lysis & infection of adjacent cells → repeated amastigote cycle in reticuloendothelial system & internal organs Visceral Leishmaniasis—Kala Azar; old world (L. donovani & L. infantum) & new world (L. chagasi); wide distribution Sx—10d-1y incubation; acute dz 1-3w; hepatosplenomegaly (laden w/ amastigotes) → erythrophagocytosis → anemia, hyperglobulinemia, & hypoalbuminia Renal complications w/ Ig deposition; leucopenia w/ impaired hematopoesis in bone marrow Characterized by fever peaking 24h after first symptoms; cachexia; may resolve spontaneously after chemotherapy Post-Kala Azar dermal Leishmaniasis (PKDL)—aggressive dermal lesions follow resolution of visceral symptoms in 10-20% of “cured” patients 120d Tx of daily injections of Pentavalent antimonials OR 90d Amphoterecin B infusions Dx—serologic; biopsy or bone marrow smears Tx—cell mediate immunity plays critical role in control, often combine Tx w/ IFN- Pentavalent antimonials (Sb5+)—first line Tx for 50y; injections into cutaneous lesion OR IM for systemic Tx, daily for 20-28d; toxic; inexpensive Amphotericin B—visceral & mucocutaneous Leishmaniasis; liposome encapsulation reduces toxicity; expensive, DOC in US; 7-10d regimen Aminosidine—Abx injected over 21d; up to 97% long-term cure rate Miltefosine—chemotherapy synthetic phospholipids derivative; delivered orally over 21d; 97% long-term cure rate Nematodes—1.5 billion infected w/ Ascaris (30,000 deaths annually); 1 billion infected w/ Hookworms; 500 million infected w/ Enterobius Structure—roundworms, NOT segmented; 0.3mm to 8.5m long; inhabit soil, fresh & salt water; some hermaphrodites Generalized life cycle—ingestion of embryonated eggs, larval stages OR penetrate skin Adult worms copulate to produce eggs → viviparous (“give birth”) to larval stages or deposit eggs → eggs “mature” thru 3 larval stages (L1-L3) Intestinal nematodes: Pinworms (Enterobius vermiculus)—most common parasitic infection in US Ep—humans definitive host; males 2-5mm & females 8-13mm long; transmited by autoinfection, person-to-person, eggs on fomites, OR eggs in dust/air Life cycle—L1 hatch in duodenum → sexually mature as enter colon → copulate & gravid females migrate towards anus (at night) → deposit up to 10,000 eggs Sx—1-4w incubation; intense itching (pruritis ani) in anal region, often seen in kids scratching; also diarrhea, sleep disturbances Dx—coffee-bean shaped eggs detectable 5-10w post infection; scotch tape test (tape on tongue depressor at night or morning prior to bathing or BM) Tx—Mebendazole in single dose, repeated after 2w; must treat ALL family w/ thorough cleaning Whipworms (Trichuris trichiura)—estimated 2 million cases in US; 3-5cm long Life cycle—ingested embryonated eggs (L2) → hatch in small intestine & burrow into epithelium → mature w/ 4 molts → mature worms return to lumen → carried to colon → anchor anterior portion into mucosa → mate & release unembryonated eggs → mature in soil for 18d-1m Sx—2-3m incubation; embeds in epithelium of colon; localized inflammation → blood (0.005mL per worm per day) & mucous in stool (colitis) → anemia Rectal prolapse—complication of rectum forced externally in kids due to tenesmus (ineffectual attempts at deffication) Dx—detection of lemon-shaped eggs in stool Tx—Mebendazole for larval stages; Oxantel for adult worms in colon; rectal prolapse must be treated surgically w/ Abx for 2° bacterial infection Ascariasis (Ascaris lumbricoides)—nearly 1/3 world population infected; prevalent in rural South US Ep—largest nematodes in humans w/ males 15-30cm & females 20-35cm long; worm burden can obstruct gut & compete for nutrition Life cycle—eggs shed in feces (240,000 per day), fertilized eggs ovoid/spherical w/ thick knobby shell → mature in soil for 10-40d → L2 embryonated eggs ingested → hatch & release larvae → cross intestine into liver, heart, lungs → L3 forms in lungs → emerges from alveoli up trachea → worms swallowed & mature w/ final molt Sx—7d incubation (lungs) OR 3w (GI); 85% asymptomatic, else abdominal pain, vomiting, eosinophilia, fever (high worm burden) Worms aggregate & obstruct gut → worms enter bile duct, esophagus, mouth, pancreatic duct, & liver → perforation & peritonitis Dx—worms often passed as bolus w/ Tx OR coughing up L3, emergence in oral & nasal compartments; detection of eggs in stool Tx—Mebendazole & Albendazole effective against both adult & larval forms (can cause aggregation in heavy infections) Piperazine or Levamisol—renders worms unable to swim against perstalsis & passed; NOT as effective in mixed worm infections Visceral larva migrans (Toxocara canis or cati)—humans (ONLY intermediate host) & canines or felines (definitive hosts); 10-12cm long Life cycle—ingested embryonate eggs → hatch in intestine & release L2 worms → develop into L3 & burrow into circulation → visceral organs → attacked by eosinophils & form granulomas Sx—usually asymptomatic, else granuloma formation in heart, brain, or eyes Ocular infections—damage to retina & immune response, usually unilateral, older kids & teens → blindness & retinal detachment Mistaken for retinoblastoma w/ unnecessary enucleation of eye CNS infections—seizure, personality changes, encephalopathy Dx—serologic (Ab against L3 by ELISA); elevated eosinophil levels Tx—Piperazine derivaties for L2; Albendazole for L3; corticosteroids to reduce inflammatory response; laser ablated larvae in eye Extraintestinal nematodes: Trichinosis (Trichinella spiralis)—infects 50 million worldwide Ep—reservoir in pigs & bears; transmitted thru undercooked game meat; very small w/ males 1.5mm & females 3mm long, larval stages microscopic Life cycle—ingest larval cysts in meat → penetrate epithelium & mature (5-7d) → mate & female releases >2000 live larvae into gut (viviparous) → larvae penetrate intestine into portal circulation → target skeletal muscle & form cysts → calcification of cyst by immune response (protects cyst for 20y) → eosinophil infiltration Sx—diarrhea, vomiting, abdominal pain; eosinophil infiltration → muscle pain, edema, eosinophilia Tx—Hx of consumption of undercooked game meat sufficient reason to give corticosteroids; Benzimidazole some effectiveness Guinea worm (Dracunculus medinensis)—Fiery serpent, Medina worm Ep—transmitted by ingestion of cepapods (Cyclops water fleas) infected w/ larval worms; mature females 50-120cm long Life cycle—ingested water fleas → larvae penetrate intestine & migrate thru CT (3m) → mate in CT & male dies → female migrate to subQ tissue (7m) → worm emerges thru boil or blister in extremities (foot & lower leg) by 10-14m → gravid female releases 1000s of live larvae into water Sx—3-4m incubation; intense burning at site of worm exit due to intense allergic reaction; potential for 2° bacterial infection Traditionally spool worm on stick as it emerges; surgical excision if worm breaks & causes intense skin reaction Tx—Metronidazole kills bacteria & facilitate removal of worm; Benzimidazoles kill worm but then require surgical removal Anisakisis—Herring worm dz, Codworm Ep—infect marine mammals; transmitted thru raw fish; prevalent in Japan, Netherlands, Scandinavia 40 Life cycle—worm migrates from GI tract to muscle as fish is caught; freezing for 2-3d destroys larvae; worms passed, vomited, or surgically removed Sx—acute dz w/in hours; acute epigastric pain as larvae invade mucosa → bloody vomiting, heartburn, diarrhea Ectopic anisakiasis—worms penetrate intestinal mucosa & cause localized infection of visceral organs Dx—endoscopic exam & excision of worms; maybe detectable on x-rays as coiled defects in GI tract; serologic (Ag by ELISA) Skin-penetrating nematodes: Strongyloidiasis (Strongyloides stercoralis)—Cochin China diarrhea; 40y patent period Ep—humans, dogs & cats; larval forms in soil penetrate skin from soil or auto-infection; 1-3mm long w/ 2 alternative life cycle stages Life cycle—larvae penetrate skin → migrate thru circulation to lungs → migrate up trachea & swallowed → adult females establish in intestine → reproduce by parthenogenesis → deposit eggs in mucosa of jejunum → filariform larvae hatch & emerge in feces → some larvae invade mucosa or anal skin (chronic infection) → develop into male & females in soil → mate & produce eggs (rhabditiform larvae) → convert to filariform larvae Sx—12-18h incubation (dermatitis), 1w (lungs), 2w (GI); allergic dermatitis at entry site → bronchitis & allergic bronchopneumonia → diarrhea & malabsorption Dx—detection of larvae in feces or duodenal aspirates Tx—Ivermectin against female worms & larvae; Thiobenzamidazoles for disseminated infection Hookworms—2nd to Ascaris in human infections worldwide; Necator americanus & Anclyostoma duodenale & animal hookworms occasionally Ep—small 7-10mm; distinctive mouthparts that anchor into mucosa & copulatory organs; secrete immuno-modulatory molecules to depress host immunity Life cycle—penetration of skin by filariform larvae in soil → migrate thru circulation to lungs → migrate up trachea & swallowed → establish in intestine (patent infection) OR L3 migrate to other tissues (dormant infection) → develop into male & females → anchor in intestinal epithelium using hooks → mate & shed eggs → partially embryonates (8 cell stage) as passed thru gut → form rhabditiform larvae in soil → differentiate into infectious filariform larvae (L3) Sx—4h incubation (dermatitis), 2w (GI); 20y patent; most subclinical, else intense itching → papular & vesicular rash → focal hemorrhage & allergic pneumonia GI—abdominal pain, blood loss (stool appears black w/ up to 100mL/day), eosinophilic enteritis → anemia, cechexia Pregnancy—iron deficient anemia, LBW Immune response—potent TH2 infection mediated by IL-4; allergic response w/ elevated IgG1, IgG4 & IgE (directly correlate w/ worm burden) Dx—detection of eggs in feces Treatment—Benzimidazoles against adult forms; Tetrahydropyrimidines (Pyrantel) against larval forms; nutrition & supplemental Fe for anemia Cutaneous larva migrans—creeping eruptions; Anclyostoma caninum (dogs) & A. brasiliense (cats) Ep—humans NOT definitive host, NO eggs shed or detected; restricted to skin Sx—radiating serpiginous lesions under skin, persist for months until larvae die Tx—topical or systemically w/ Thiabendazole Vector-borne nematodes: Mosquitos—Lymphatic filarisasis (night feeding Anopheles & Culex), Brugia malayi, Wuchereria bancrofti, canine heartworm occasionally Flies—ocular filariasis (Loa Loa transmitted by Crysops biting fly), Onchocerca volvulus (River blindness transmitted by black flies) Lymphatic filariasis—Wuchereria bancrofti (wide distribution) & Brugia malayi (restricted to southeast Asia) Ep—worms migrate to proboscis surface from flight muscle & contaminate bite wound; males 4cm & females 5-10cm long (5-10y), microfilariae 0.3mm (18m) Target lymphatic system & accumulate in LN, remain for decades mating & releasing microfilaria (larvae) into blood w/ nocturnal circadian rhythm Sx—maturation in LN of lower extremities, occasionally arm & breasts (6m-1y) → remain paired & mate (5-40y) → viviparous release of microfilariae Immune granulomatous response → occlude lymphatics → lymphoedema → fibrosis Initial fever, lymphadenopathy, chyleuria (milky appearance of urine b/c chyle, lymph + chylomicrons, blocked from lymphatics) Elephantiasis—giant swelling of organs; acute orchitis w/ hydrocele; verrucous skin growth w/ hyperproliferation of new tissue Grossly deforming; killing adult worms in lymphatics exacerbates condition Tx—Piperizine derivatives used for >50y to immobilize microfilariae → exit circulation at liver & cleared Side effects—trigger potentially fatal allergic reactions due to large-scale destruction of microfilariae; corticosteroids or antihistamine also Ivermectin & Mebendazole—very effective against microfilariae, NOT adults Loa Loa (Ocular filariasis)—central & western Africa, Central America; males 40mm & females 70mm long Life cycle—larvae enter fly bite & develop slowly into adults (1y) → mate & migrate thru tissue shedding microfilariae → localized inflammation → subQ (Calabar swelling) & subconjunctival spaces of eye → dead worms & microfilariae trigger microabscess formation Sx—2-12m incubation; subQ swellings (Calabar swellings), visual impairments → retinal damage Tx—DEC kills both microfilariae & immature stages, NOT adults; adult worms surgically removed during transit thru eye River blindness (Onchocerca volvulus)—follows major rivers in Africa; ONLY female fly draws blood Life cycle—transmit L3 or microfilariae → migrate thru skin & mature into adults (1y) → nodule formation under skin (contain multiple worms) mate & produce microfilariae Sx—papular dermatitis (craw craw) → thickening of skin (worse by scratching) & changes in pigmentation; scrotal elephantitis in lymphatic system Ocular—primarily by dead microfilariae w/ repeated cycles of infection → keratitis → scarring & damage to optic nerve → blindness Dx—skin snips of nodules detect motile worms or larvae; serologic Tx—prophylaxis & vector control; Ivermectin targets microfilariae Nematocidal drugs: Ivermectin—targets Helminth nervous system causing paralysis Benzimamidazoles—prevents tubulin polymerization; Albendazole inhibits glucose uptake & Mebendazole inhibits tubulin polymerization Levamisol—inhibits succinate dehydrogenase (energy production) Piperazine derivatives & Tetrahydropyrimidine (Pyrantel)—paralysis Cestodes—tapeworms (flatworms or Platyhelminths); hermaphroditic; NO internal digestive tract, absorb nutrients thru tegument; live in intestinal tract of host Segmented—anterior (contains scolex or holdfast organ that anchors worm to intestinal wall w/ suckers, grooves or hooks), neck (connects scolex to body) Proglottids—immature, mature, or gravid (laden w/ eggs); contains both male & female organs; chronic infections compete w/ host for nutrients; up to 25m long General life cycle—ingestion of embryonated eggs or cysts in muscle → development of scolex → attachment to intestinal wall → production of proglottid segments → maturation of proglottids & egg formation → release of gravid proglottids in feces → eggs released in thin hyaline membrane (egg shell) that disintegrates → thick striated embryophore encloses oncosphere (infectious form) that develops into larvae Hydatidosis (Echinococcus granulosus)—Hydatid dz; Ep—resides in intestinal tract of dogs (definitive host); small 2.5-6mm long; small neck & ONLY 3-4 proglottids Life cycle—ingestion of embryonated eggs → larvae released & migrate to liver, lungs, brain & eye → form hydatid cysts filled w/ asexually dividing larvae (protoscolex) → protoscolex attaches to intestine & matures to adult Hydatid dz—rare in humans; hydatid cysts in herbivores (sheep) metastasize by rupture & seepage, release larvae (hydatid sand) & trigger anaphylactic shock 41 E. multilocularis—significant metastatic dz Sx—liver dysfunction, lung problems, ascities, abdominal pain, neurologic & eye problems Dx—ultrasound, CAT scan of organs; calcified cysts seen by x-ray; serologic Tx—hydatid cysts removed surgically; Albendazole Taeniasis & Cysticercosis—beef (Taenia saginata) & pork (T. solinum) tapeworms Ep—humans (definitive host) w/ sexual cycle in GI tract; T. solinum (2-7m) & T. saginata (6-15m) w/ 1000s proglottids; distinctive scolices & proglottids for ID Life cycles—adult worms live exclusively in GI tract of humans → shed gravid proglottids in feces → eggs embryonate → ingested by cattle or pigs → larval oncospheres enter circulation → cyst formation in muscle → ingestion of undercooked meat → release of larvae & maturation Sx—8-10w incubation; 25y patent period (intestinal) & 2y (cysts); weight loss (competition for nutrients), malnutrition, abdominal pain, anal itching Cyst formation in muscle (Cysticercus bovis or Cysticercus intermis w/ beef) & muscle/other organs (Cysticercus cellulose w/ pork) T. solinum—forms cysts in brain, eye, & other tissues; traditionally fed “night soil” or refuse contaminated w/ human feces Dx—detect proglottids in stool; cysts detected by CAT scan, MRI; serologic Tx—Niclosamide (single dose), Praziquantel (single dose), Mebeldazole (3d) for GI worms; Albendazole & Praziquantel for larval worms w/ surgical excision Diphyllobothrium latum—fish tapeworm Ep—fish-eating mammals (definitive hosts); freshwater crustacean (copepods) & freshwater fish (trout, whitefish, pike) intermediate hosts Larvae in undercooked, picked or smoked fish; often Gefuellte fish (Jewish) & gravlax (Scandinavia) Life cycle—embryonated eggs produce ciliated coracidium larvae → released from egg & infects cepapods → forms procercoids → fish eat infected cepapods → larvae migrate to muscle Sx—3w incubation; vague symptoms w/ neurologic problems (weakness, ataxia) from vitamin B12 deficiency (competition) Tx—Praziquantel & vitamin B12 Rodent tapeworms (Hymenolepis nana, H. dimunata) & Canine tapeworm (Diphylidium caninum) Trematodes—flukes (Platyhelminths) Structure—small, leaf-shaped; simple, blind alimentary canal (mouth but NO anus); suckers or hooks; locomotion by muscles (some ciliated); most hermaphroditic Schistosoma dioecious (separate sexes), adult male & females worms mate & remain paired for life; vertebrate (definitive hosts) & snails (intermediate hosts) Life cycle—fertilized eggs released in urine OR feces → eggs embryonate forming pear-shaped larvae (miracdidium) → miracidia break thru egg shell or operculum → target freshwater snails w/ highly motile miracidium → miracidia attach & invade snail body forming sporocyst → spawns daughter sporocysts → produce cercariae → exit snail into water via forked tail → penetrate skin of human → induces maturation of adults → mate & produce eggs Schistosoma—Schistosomiasis, Bilharziasis; infects 200 million annually, 120 symptomatic, 200,000 deaths Ep—blood flukes; grow anchored in venous circulation; feed on blood Life cycle—fresh water cercariae penetrate skin OR swallowed → convert to schistosomules (lose forked tail) → mature in liver → swim against flow to mesenteric veins around colon (GI—S. mansoni & S. japaniocum) OR pelvic veins (GU—S. heamotobium) → monogamous males & females pair for life → release eggs that are shed in feces or urine → eggs embryonate & release miracidiae Acute Sx—cercarial infection maculopapular rash in few hours; NOT severe GI (Katayama syndrome)—14-90d incubation; symptoms coincide w/ egg deposition (fever, headache, myalgia, URQ pain, bloody diarrhea) GU—10-12w incubation; hematouria first symptom Chronic Sx—granuloma formation destroys eggs but deposits fibrotic lesions; forces eggs from blood across intestinal or bladder into feces or urine GI—1-3w incubation; 25y patent; granulomas in intestine & liver; severe worm burden misdiagnosed as colon cancer Liver—hepatomegaly, portal HTN, splenomegaly → anemia (particularly in kids) & dysfunction (late stages) Co-infection w/ HBV or HCV promotes liver degeneration & risk for hepatocellular carcinoma GU—4-7w incubation; 25w patient; dysuria & hematouria → proteouria, calcification of bladder, ureter obstruction, renal colic & renal failure, 2° UTI Correlation b/w S. heamotobium infection & bladder cancer Female genital schistosomiasis (FGS)—vulval & perineal hypertrophic & ulcerative lesions; misdiagnosed as genital warts Pulmonary—ecotpic granulomas → pulmonary HTN CNS—most often w. S. japoniocum; granuloma formation → seizures Childhood—anemia, MR & growth retardation Dx—detect eggs in feces or urine Tx—Praziquantel & Tetrahydroquinolones for suppression of egg production, up to 100% cure rate, against adult parasites, NOT larvae Microbilharzaria variglandis—swimmers itch Ep—infects migratory waterfowl that deposit eggs in freshwater → eggs embryonate & release miracidia → intermediate host in snails → cercariae production → infect swimmers → cercariae die & trigger intense dermal reaction Sx—cercarial dermatitis caused by avian schistosomes; rash, papular or nodular, localized edema, itching Tx—topical steroid & anti-histamines Ingested flukes—ALL treated w/ Praziquantel Fasciola hepatica—liver fluke; parasite of sheep w/ snails as intermediate host (found on watercress) Life cycle—ingest encysted cercariae (metacercariae) on watercress → excysts in duodenum → migrates thru peritoneum to liver → develop in bile ducts → eggs produced that enter intestine w/ bile → excreted in feces → eggs embryonate & miracidia infect snails → rapid growth of cercariae → sporocyst aggregate into redia → free-swimming cercariae form cysts on watercress Sx—3-12w incubation; 1-20y patent; hepatomegaly, fever, dyspepsia, ascities, eosinophilia, occlusion of bile duct Tx—reinfections common, respond poorly to Tx; Bisphenols & Praziquantel against adults; Albendazole against larvae Clonorchis sinesinsis—liver fluke; endemic in southeast Asia & US veterans; definitive hosts in humans, cats & dogs; intermediate hosts in snails or freshwater fish Life cycle—ingest metacercariae from raw fish → adult worms migrate to liver → develop in bile ducts Sx—2w incubation; 20y patient period; inflammation & intermittent obstruction of biliary ducts; long-term infections (cholangitis, pancreatitis, cholangiocarcinoma) Paragonimus westermani—liver fluke; Asia & Africa; intermediate hosts in snails & crabs Life cycle—worms penetrate intestine & diaphragm to reach lung → pairs of copulating worms form fibrotic lesions → eggs expectorated in sputum → eggs swallowed & transmitted in feces → miracidia develop from embryonated eggs → infect snails → infect crabs or crayfish Sx—9-12w incubation; 20y patent; infiltrate surrounds worms w/ fibrous capsule; hemorrhage of capsule releases eggs → bronchitis, thoracic & abdominal pain Fasciolopsis buski—intestinal fluke; southeast Asia; intermediate host in snails (water chestnut & bamboo) Life cycle—fluke attaches to duodenal wall rather than penetrate to liver Sx—most asymptomatic; else diarrhea, abdominal pain, fever, ascities, intestinal obstruction Ectoparasites—feed on outside of body (suck blood, eat skin, sting/bite as defense mechanism); true ectoparasites remain intimately associated w/ host for prolonged time Pediculosis—lice infestations of head, body or pubic hair; anoplura (bloodsucking lice) VS. mallophaga (feed on skin & secretions) Head lice (Pediculus humana capita)—infect eyebrows & eyelashes; prefer straight hair NITS—lice eggs, cement to individual hairs near scalp for warmth; hatch after 1w, series of molts in adults; feed daily on blood; lay up to 8 nits/day 42 Sx—usually asymptomatic; else itches & tickling Dx—adult lice attached to scalp; nits w/in 6mm of scalp indicates active infection Tx—Pediculicide & washing; ALL family treated; hair combed w/ NIT comb to dislodge nits from hair shaft (nit-picking) Body lice (Pediculus humana corporis)—transmit typhus, trench fever, relapsing fever Pubic lice (Phthirus pubis)—Crabs; tropism for pubic hairs; transmitted by sex (condoms NOT useful), NOT surfaces; cause intense itching Tx—topical Pediculocides; must treat ALL partners Mites (Sarcoptes scabiei)—Scabies in humans, Mange in dogs Ep—wide range of arthropods; feed on dust; some bloodsucking & transmit dz; dig into epidermidis Sx—allergies to dust mites by contact or inhalation; lay eggs in burrows & metabolic products trigger inflammation & thickening of skin Lesions first on hands & spread to feet, genitals & axilla, scratching promotes 2° infections Epidermal scaling—Norwegian scabies, seen in imczd Dx—skin scrapings; Tx—topical Acarizides Myasis—maggot infestations; over 80 species Obligatory myasis—infest living hosts VS. facultative or accidental myasis—infect necrotic tissue, deposit eggs directly Human bot flies—Dermatoba hominis Life cycle—flies lay eggs on legs or body of bloodsucking insects or ticks → eggs hatch & larvae burrow into wounds, nose, ear, or scalp → inflamed furuncle (microabcess) → mature larvae escape lesion & pupate into adult flies in soil Sx—intestinal, GU, naso-pharyngeal, dermal (creeping eruptions) Tx—larvae must be surgically removed (anchoring hooks) OR encouraged to emerge w/ pork fat or meat applied to lesion Screw worms—lay eggs in open wounds, grow on necrotic tissue ONLY; Wohlfaritia (both necrotic & healthy tissue) & Dermatobia hominis Lucilia serricata—lab-raised flies used to debride necrotic tissue; do NOT attack healthy tissue Ep—often in rural, indigent, poor hygiene, alcoholics, poor wound healing (DM), overcrowding Sand fleas—Tunga penetrans (Tungiasis); female fleas invade skin on soles of feet; up to 1cm; vectors of dz Life cycle—inseminated female penetrates skin & suck blood → tail oriented out w/ hole for breathing & shed eggs into epidermis Sx—small erythymatous spot, swells into itchy nodule w/ dark central spot → painful (impedes walking) w/ 2° infection common Tx—extrication of flea w/ sterile needle; topical antiseptics Leeches—parasitic b/c suck blood, 2° bacterial infections common Medicinal leeches—Hirudo medicinalis; bloodletting; still used in plastic surgery to reattach digits b/c promote circulation & prevent localized coagulation
