Download 06EnterobacII2012 - Cal State LA

Enterobacteriaceae II:
Intestinal Pathogens
Salmonella, Shigella, Yersinia
Salmonella
• Animals main reservoir of human disease:
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Gastroenteritis
Typhoid (Enteric) fever
Septicemia
Asymptomatic carriage
• Transmission:
– Ingest contaminated food (poultry, eggs, dairy
products)
– Direct fecal-oral spread in children
Salmonella Genera
• Describe >2500 serotypes
• DNA homology studies:
– One Genus-species - Salmonella enterica
– Seven subspecies (1, 2 ,3a ,3b ,4 ,5, and 6)
– Subgroup 1 causes most human infections
• Clinically Salmonella isolates often still
reported as serotypes by Kauffman-White
classification:
– Based on O (cell) and H (flagella) antigens
– H antigens occur in two phases (antigenic
variation)
– Polyvalent antisera used, followed by group
specific antisera (A, B, C1, C2, D, and E)
– Salmonella Typhi (capitalize, not italics to
designate Genus-Serotype) - also has capsular
Vi antigen
Salmonella: Biochemical
Test ID
• TSI = K/A + gas, H2S
– Salmonella Typhi small amount of H2S, no gas
– Salmonella Paratyphi no H2S
• LIA = K/K (+)lysine DC, +H2S
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– Salmonella Paratyphi = K/A
Urea(–)
PA(-)
Citrate(±)
Indole(-)
Salmonella: Infection
• MO found in all animals – poultry, reptiles,
livestock, rodents, domestic animals, birds,
and humans
• Highly adapted strict human pathogen:
– Salmonella Typhi
– Salmonella Paratyphi
• Peak incidence of infections in warm
months via ingesting contaminated food at
outdoor social gatherings
• ~50,000 cases reported annually in U.S
Salmonella: Infection of GI
Tract
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Ingest contaminated food
Infect GI cells
Blood (systemic infection, target organs)
RES (Reticuloendothelial System) –
macrophages, liver, spleen, bone marrow
Infection of GI Tract
• MO adheres to intestinal mucosa epithelium
• Invasion of enterocytes, M cells (microfold cells
of Peyer patches)
• Induced endocytosis by bacteria effector
proteins
• Salmonella multiplies within phagosome,
released; may be phagocytosed by macrophages
Salmonella: Gastroenteritis
• Salmonellosis most common form of
gastroenteritis
• Due to Salmonella Enteritidis in U.S.
• Symptoms 6-48 hours after consumption
contaminated food:
– Nausea, vomiting, non-bloody diarrhea
– Also common with elevated temperature,
abdominal cramps, myalgia, headache
• Persist 2-7 days, spontaneous resolution
• Supportive care, no antibiotics required
• Incidence greatest children <5 yrs. and
adults >60 yrs.
Gastroenteritis
• MOs multiply, induce strong inflammatory
response
• Causes disease symptoms (fever, diarrhea,
abdominal cramps)
• Inflammatory response prevents spread
beyond GI tract, eventually kills MOs
Salmonella: Typhoid ( Enteric)
Fever
• Typhoid fever - Salmonella Typhi
• Paratyphoid fever (milder) - Salmonella
Paratyphi
• 10-14 day incubation period following
ingestion of contaminated food
• Fever, headache, myalgia, malaise, anorexia
• Persist 1 week, followed by GI symptoms
(GI→blood→GI)
• Mortality ~20% if untreated
Typhoid Fever
• MO disseminate before high enough levels
to stimulate inflammatory response
• Initial symptoms low fever, constipation
• MO move via lymphatics & bloodstream to
liver & spleen; phagocytosis &
multiplication occurs
• MO re-enter bloodstream, disseminate to
all organs; fever, headaches, myalgia, GI
problems
• Rose spots (erythema, maculopapular
lesions) on abdomen
• Osteomyelitis, cystitis, gall bladder
infections may occur
Salmonella: Septicemia
• Most frequently:
– Salmonella Typhimurium
– Salmonella Choleraesuis
– Salmonella Paratyphi
• At risk: children, elderly, AIDS patients
• Infection presents like Gram(-)
bacteremia
• 10% develop osteomyelitis, endocarditis,
arthritis
Salmonella: Asymptomatic
Infection
• Salmonella Typhi
• Salmonella Paratyphi
• MO’s from enteric fever maintained by
human carriage
• ~1-5% infected patients chronic carrier >1
year
• Reservoir in gall bladder
Lab Diagnosis: Typhoid
Fever
• Blood cultures positive during first week,
after second week
• Stool culture, sometimes urine culture
positive after second week
• Widal Test (serology):
– Antibodies against Salmonella Typhi
– Look for 4-fold rise in titer between acute and
convalescent stage (~one month)
Salmonella: Prevention
• Gastroenteritis:
– Public Health education
– Improved hygiene, especially food handlers
– Safe preparation of poultry & eggs,
refrigerate food
– Antibiotics not recommended, may prolong
disease
• Enteric fevers:
– Treat with fluoroquinolone (ciprofloxacin),
chloramphenicol ( decrease mortality to <2%)
– Live oral vaccine (attenuated Salmonella Typhi)
for travelers to endeminc countries
– Clean water supply
– ID, treat chronic carriers
Shigella: Gastroenteritis
• Shigellosis (Bacillary dysentery)
• ~150 million cases annually worldwide;
450,000 in U.S.
• Humans only reservoir, transmitted
person-to-person by fecal-oral route
• Outbreaks in communities where sanitary
standards, level of hygiene low
• Common in daycare centers, nurseries,
custodial institutions
Shigella Genera
• Four species by O antigen serotype:
– S. dysenteriae (Group A) – most severe; Shiga
toxin
– S. flexneri (Group B) – developing countries
– S. boydii (Group C) – not commonly isolated
– S. sonnei (Group D) – developed countries; U.S.
• By DNA analysis, now shown to be biogroup
within E. coli
Shigella: Biochemical Test ID
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TSI = K/A with NO gas
LIA = K/A (-)lysine DC
Urea (–)
Motility(–)
S. sonnei may show delayed lactose
fermentation
Shigella: Virulence Factors
- Enterotoxin
• Shiga toxin - S. dysenteriae
• Smaller amounts by S. flexneri, S.
sonnei
• Inhibit protein synthesis by
inactivating 60S ribosomal subunit
• Role in ulceration intestinal mucosa
Shigella: Virulence Factors Cell Wall Proteins
• Expressed at body temperature
• Upon contact with intestinal
mucosa M cells (microfold cells,
Peyer’s patches), induce
phagocytosis of bacteria into
vacuoles
• Shigella destroys vacuoles,
escape into cytoplasm
• Spread laterally (actin filaments
propel MO through cytoplasm) to
epithelial cells; multiply
• Do not usually disseminate
beyond epithelium
Shigella: M Cell
Attachment
Shigella: M Cell
Penetration
Shigella: Epithelial Cell
(CCEC) Invasion
Shigellosis
• Fecal-oral route, primarily contaminated
hands
• Infective dose very low (100-200 MO)
• Incubation 1-7 days
• Fever, cramping, abdominal pain, watery
diarrhea for 1-3 days (due to exotoxin)
• Followed by frequent, scant stools with
blood, mucous, pus (invasion of intestinal
mucosa)
• Rare for MO to disseminate, generally selflimited but may lead to death
• Severity of disease depends upon species:
– S. dysenteriae most pathogenic
– Followed by S. flexneri, S. sonnei, S. boydii
Shigella: Treatment and
Prevention
• Trimethoprim-sulfamethoxazole (SXT) or
fluoroquinolone (ciprofloxacin) - shortens
course of disease & fecal shedding;
recommended to reduce spread to
contacts
• Resistant strains becoming increasingly
common, so antibiotic sensitivity testing
required
• Infection control by proper hand washing
and disposal of soiled diapers/linens
• Public Health monitoring for clean water
supply
Comparison of Invasion:
Salmonella versus Shigella
Yersinia Genera
• Infections are zoonotic, humans accidental
host
• Three species important pathogens in
humans:
– Yersinia pestis – plague (bubonic or pneumonic)
– Yersinia enterocolitica – gastroenteritis,
transfusion-related sepsis
– Yersinia pseudotuberculosis – mainly disease of
animals, uncommon human gastroenteritis
Yersinia: Infection
• Y. pestis:
– Mammalian reservoir: rats, squirrels,
rabbits, domestic animals
– Humans infected by:
• Fleas
• Direct contact with infected animal tissue
• Aerosols from patient with pulmonary
disease
• Y. enterocolitica:
– Ingest contaminated food products
– Infusion of contaminated blood products
(growth in blood stored at 4°C)
Yersinia pestis: Lab ID
• Non-motile - 37°C &
25°C
• Bipolar staining
• Slow growth, small
colonies at 37°C (grows
better 25°C)
• TSI = K/A
• Urea(-)
• ODC(-)
• Guinea pig LD50<10
• Direct fluorescent
antibody test
• New DNA probe test
Y. pestis: Bubonic Plague
• By flea bite infected animal (rat,
squirrel, rabbit) to human
• Endemic in local So Cal mountains
• MO travel to nearest lymph node,
engulfed by macrophage
• High fever, buboe (enlarged lymph
node), MO proliferate, stimulate
inflammatory response
• MO multiply in lymph node, leak into
bloodstream
• Lysis of bacteria releases LPS causing
septic shock, DIC
• Subcutaneous hemorrhage, disease
named Black Death in Middle Ages
• High mortality rate (30-40%) if
untreated
Pneumonic Plague
• Eventually bacteria reach lungs, engulfed
by lung macrophages, cause pneumonic
plague
• Transmit directly to others via aerosol
• Direct inhalation produces disease that
progress more rapidly
• Mortality rate close to 100%
Y. pestis: Treatment and
Prevention
• Streptomycin, tetracycline, SXT
• Control by reducing rodent population
• Vaccine (formalin killed bacteria) for
individuals at risk; no longer available
Yersinia enterocolitica: Lab ID
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Non-motile at 37°C
Motile at 25°C
Bipolar staining
Slow growth, small colonies at 37°C (grows
better at 25°C)
• TSI = A/A (sucrose fermentation)
• Urea(+)
• ODC(+)
CIN Agar
• Isolation of Y. enterocolitica
from stool specimen (also
Aeromonas, Plesiomonas)
• Selective - antimicrobials
(Cefsulodin-Irgasan-Novobiocin),
bile, crystal violet inhibit NF
• Differential - mannitol,
neutral red
• Incubate room temperature
• After 48 hours at RT, Y.
enterocolitica produce typical
pink colonies (mannitol
fermentation) with bulls-eye
appearance
Y. enterocolitica:
Gastroenteritis
• Ingestion of contaminated food or water
• Common cause of human disease (mostly in
children) involving fever, abdominal pain,
watery diarrhea
• Intestinal epithelium invasion of M cells,
transcytosed through M cells, released at
basal surface
• Bacteria penetrate into underlying
lymphoid tissue, multiply both inside and
outside host cells
Gastroenteritis
• Inflammatory response to MO
responsible for extreme pain (~acute
appendicitis)
• Fever due to LPS endotoxin
• Sometimes drain into adjacent
mesenteric lymph nodes, causing
lymphadenitis
• Reactive arthritis may occur in some
people; thought cross reacting T cells or
antibodies that attack joint
• Usually self-limited
• If treatment indicated, susceptible to
broad-spectrum cephalosporins
Summary: Yersinia Infections
Lab: Enterobacteriaceae
II Group Unknowns
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Salmonella
Proteus
Citrobacter
Yersinia
TSI
K/A, H2S
K/A, H2S
K/A, H2S
K/A
LIA
K/K, H2S
R/A, H2S
K/A, H2S
K/A
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Lecture Exam I
Thursday, Feb. 2, 2012
Introduction thru Enterobacteriaceae
Lecture
Reading (Chap. 14, 15, 17, 19)
Key Terms, Learning assessment
Questions
• Case Study 1,2,3
• Exam Format:
– Multiple Choice
– Terms
– True/False Statements
– Short Essay
• Review, Review, Review!