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Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Bacterial cell structures and function a) b) c) d) The structure of bacterial cells A typical bacterial cell is composed of: Capsule Cell wall Cell membrane Cytoplasm containing nuclear material and appendages such as flagella and pili (fimbriae). Certain species of bacteria can produce forms termed endospores or spores, which are resistant to environmental influences. Some of the structural features of pathogenic bacteria which are important in the production of disease or may be useful for the laboratory diagnosis of infection. Capsule Bacteria can synthesize extracellular polymeric material which is usually described as glycocalyx. In some bacterial species this polymeric material forms a capsule, a well defined, organized, and not easily removed structure closely adherent to the cell wall. A slime layer is unorganized material that easily removed formed when the polymeric material is present as a loose meshwork of fibrils around the cell. Most capsules are composed of polysaccharides; Bacillus species such as B. anthracis produce polypeptide capsules. Defined capsules can be visualized by light microscopy using negative staining techniques. Bacteria with well defined capsular material produce mucoid colonies on agar media. However, the capsules of most species of bacteria can be demonstrated only by electron microscopy or by immunological methods using antisera specific for the capsular (K) antigens. The main functions of capsules a -Protection of the bacterium from adverse environmental conditions such as desiccation. b- In the body, capsules of pathogenic bacteria may facilitate adherence to surfaces and interfere with phagocytosis. 1 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Cell wall The tough, rigid cell walls of bacteria protect them from mechanical damage and osmotic lysis. As cell walls are non-selectively permeable, they exclude only very large molecules. Differences in the structure and chemical composition of the cell walls of bacterial species account for variation in their pathogenicity and influence other characteristics including staining properties. Peptidoglycan, a polymer unique to prokaryotic cells, imparts rigidity to the cell wall. This polymer is composed of chains of alternating subunits of N-acetyl glucosamine and N-acetylmuramic acid cross-linked by short tetrapeptide side chains and peptide cross-bridges. Based on their color when stained by the Gram method, bacteria can be divided into two major groups, Gram-positive and Gram-negative. Gram-positive bacteria have a relatively thick uniform cell wall which is composed mainly of peptidoglycan and teichoic acids. In contrast, Gram- negative bacteria have cell walls with a more complex structure, consisting of an outer membrane and a periplasmic space containing a comparatively small amount of peptidoglycan Similarity of Gram positive and Gram negative cell walls Characteristic Number of major layers Chemical composition Overall thickness Outer membrane Periplasmic space Permeability to molecules Gram positive One Peptidoglycan Teichoic acid Lipoteichoic acid Mycolic acid and polysaccharides in some cells Thicker (20-80 nm) No No More penetrable Gram negative Two Lipopolysaccharide (LPS) Lipoprotein Peptidoglycan Porin proteins Thinner (8-11 nm) Yes Yes Less penetrable Outer membrane description of Gram negative bacteria: The outer membrane of Gram-negative bacteria is a protein-containing asymmetrical lipid bilayer. The structure of the inner surface of the membrane resembles that of the cytoplasmic membrane, whereas that of the outer surface is composed of lipopolysaccharide (LPS) molecules. 2 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Low molecular weight substances such as sugars and amino acids enter through specialized protein channels, known as porins, in the outer membrane. The outer membrane LPS, the endotoxin of Gram-negative bacteria, is released only after cell lysis. The major components of LPS molecules are core polysaccharides bound to lipid A and long external polysaccharide side chains. The polysaccharide side chains of the LPS molecules stimulate antibody production. Lipid A is the molecular component in which endotoxic activity resides. The outer membrane excludes hydrophobic molecules and renders Gram-negative bacteria resistant to some detergents which are lethal to most Gram-positive bacteria. Gram staining This color reaction is determined by the composition of the cell wall. Gram positive bacteria, stain blue, whereas Gram negative bacteria stain red. The Gram-staining procedure consists of a primary stain (crystal violet), a mordant (Gram’s iodine), alcohol as a decolorizer and a counterstain (carbol fuchsin). The crystal violet penetrates the cell wall of both Gram-positive and Gramnegative bacteria. Following addition of the iodine, a complex with the primary dye is formed. The alcohol dehydrates the peptidoglycan of Gram-positive organisms making them less permeable to the crystal violet complex than Gram-negative bacteria. Thus the dye–iodine complex is retained in Gram-positive organisms whereas it is removed in Gram-negative organisms. Both Gram-positive and Gram-negative organisms absorb the counterstain but its pink color is visible only in Gram-negative organisms as the deep purple color of the crystal violet masks the pink color in Gram-positive organisms. The Mycoplasma comprise an important group of bacteria without cell walls. Conventional bacteria, exposed to the action of antibiotics such as penicillin, or other substances which interfere with the synthesis of peptidoglycan, cannot produce cell walls and are termed L forms. 3 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Cytoplasmic membrane The cytoplasmic membranes of bacterial cells are flexible structures composed of phospholipids and proteins. They can be observed only by electron microscopy and are structurally similar to the plasma membranes of eukaryotic cells. However, bacterial cytoplasmic membranes, with the exception of those present in mycoplasmas, do not contain sterols. The inner and outer faces of cytoplasmic membranes are hydrophilic while the interior is hydrophobic, forming a barrier to most hydrophilic molecules. Only a limited range of small molecules such as water, oxygen, carbon dioxide and some lipid-soluble compounds can enter bacterial cells by passive diffusion. Functions of the cytoplasmic membrane a) Active transport of nutrients into the cell. b) Elimination of waste metabolites. c) The cytoplasmic membrane is also the site of electron transport for bacterial respiration, and of enzymes and carrier molecules that function in the biosynthesis of DNA, cell wall polymers and membrane lipids. Cytoplasm The cytoplasm, which is enclosed by the cytoplasmic membrane, is essentially an aqueous fluid containing the nuclear material, ribosomes, nutrients and the enzymes and other molecules involved in synthesis, cell maintenance and metabolism. Storage granules may be present under certain environmental conditions, usually those unfavorable for bacterial growth. These granules, which may be composed of starch, glycogen, polyphosphate or other compounds, can often be identified using particular dyes. Ribosomes All protein synthesis takes place on ribosomes. These structures are composed of ribonucleoproteins and are up to 25 nm in size. They consist of two subunits, a larger 50S subunit and a smaller 30S subunit. The Svedberg (S) unit is a measure of sedimentation rate, which is dependent on both the size and shape of a particle. 4 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Ribosomal ribonucleic acid (rRNA) is complexed with many different proteins and accounts for about 80% of the RNA of the cell. Smaller amounts of transfer RNA (tRNA) and messenger RNA (mRNA) account for the remaining cellular RNA. Ribosomes may be present either in the cytoplasm or associated with the inner surface of the cytoplasmic membrane. During active bacterial growth and rapid protein synthesis, individual ribosomes are joined by mRNA into long chains known as polysomes. Nuclear material Chromosomes: The bacterial genome is usually composed of a single haploid circular chromosome containing double- stranded DNA. However, some bacteria have two circular chromosomes, and others, have a linear chromosome. The genes in the bacterial chromosome encode all the vital functions of the cell. Because of its length, the bacterial chromosome is extensively folded to form a dense body which can be seen by electron microscopy. Plasmids Small circular pieces of DNA which are separate from the genome, are capable of autonomous replication. Plasmid DNA may code for characteristics such as antibiotic resistance and exotoxin production. Flagella Bacteria that possess flagella are motile. Flagella are usually several times longer than the bacterial cell and are composed of a protein called flagellin. They consist of a filament, hook and basal body. The hook functions as a universal joint between the filament and the basal body. The basal body is anchored to the cell wall and to the cytoplasmic membrane. The positions at which flagella are inserted into the bacterial cell vary and may be characteristic of a genus or family. Motile bacteria can move into suitable microenvironments in response to physical or chemical stimuli. 5 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Flagella can be demonstrated by electron microscopy, Motility can be confirmed in young broth cultures using the hanging drop technique or in a semisolid motility medium containing tetrazolium salts. Pili Fine, straight, hair-like appendages called pili or fimbriae, composed of the protein pilin, are attached to the cell wall of many bacteria. The number of pili on each bacterial cell varies widely. They are most common on Gram-negative bacteria and they may have different functions. They frequently function in adhesion to host tissues with the adhesion located at the tips of fimbriae. Pili are also found in many Gram-positive genera such as Corynebacterium, Actinomyces, Enterococcus and Streptococcus species. As in Gram-negative bacteria, these pili function in adhesion to host tissues. A unique type of pilus, the F (sex or conjugation) pilus, functions in male or donor cells of Gram negative bacteria as a conduit for the transfer of DNA to female or recipient cells during conjugation process. Endospores Dormant highly resistant bodies termed endospores are formed by some bacteria to ensure survival during adverse environmental conditions. The only genera of pathogenic bacteria that contain endospore-forming species are Bacillus and Clostridium. Endospores, which are produced inside the bacterial cell, show species variation in shape, size and position within the mother cell. Because of the resistance and impermeability of the spore coat, special staining procedures which employ heat are required to demonstrate endospores. The resistance of endospores is attributed to a) Their layered structure. b) Their dehydrated state 6 Dr.A.K.AL-Yassari c) d) e) 2016-2017 Microbiology Year:third Their negligible metabolic activity. Their high levels of small acid- soluble proteins (SASPs). Their high content of dipicolinic acid. Because spores are thermo stable, they can be destroyed with certainty only by moist heat at 121°C for 15 min. Endospores germination When an endospore is reactivated, germination occurs in three stages, namely activation, initiation and outgrowth. Activation may occur in response to factors such as brief exposure to heat, abrasion of the spore coat or environmental acidity. If other environmental conditions including the presence of adequate nutrients are favorable, initiation of germination will occur. The spore cortex and coats are degraded, water is absorbed, calcium dipicolinate is released and outgrowth develops. Outgrowth is a period of active biosynthesis and terminates with division of the new vegetative cell. Structural features of a mature bacterial endospore. 7 Dr.A.K.AL-Yassari 2016-2017 Microbiology Year:third Comparative features of the cell walls of Gram-positive and Gram-negative bacteria. Bacterial flagella. A, Monotrichous (polar) flagellum; B, amphitrichous flagella; C, lophotrichous flagella; D, peritrichous flagella. 8