BMS1-1.b. Bacterial morphology and cell structure-Dr. Dr. Emine Atasayar.pdf

1 BACTERIAL MORPHOLOGY AND CELL STRUCTURE Assist. Prof. Dr. Güner EKİZ DİNÇMAN Faculty of Pharmacy Dept. of Pharmaceutical Microbiology  2 Bacterial Cellular Morphology Basiller §...

1 BACTERIAL MORPHOLOGY AND CELL STRUCTURE Assist. Prof. Dr. Güner EKİZ DİNÇMAN Faculty of Pharmacy Dept. of Pharmaceutical Microbiology  2 Bacterial Cellular Morphology Basiller § Çomak şekilli bakteriler § Silindir benzeri § 1-8 µm uzunlukta § Çok kısa basil→ kokobasil (coccobacilli) § 2’li basil→ diplobasil (diplobacilli) § Zincir şekilli basil→ streptobacilli  3 Bacterial Cellular Morphology Cocci § Round, spherical § 0.6-1.5 µm in size § Cell division in a single axis; streptococci § In pairs; diplococci § Clusters of four cocci arranged within the same plane; tetrads § Irregular, grape-like clusters of cocci; staphylococci  4 Bacterial Cellular Morphology Spirilla Divided into 3 groups: § Vibrios; rigid, curved, comma-shaped and capable of movement due to flagella § Spirilla; spiral-shaped, curved § Spirochetes; spiral (helical)-shaped, tightly coiled, flexible body  5 Bacterial Cellular Morphology Other shapes and arrangements of bacteria: Filamentous → very long thin filament-shaped bacteria, Actinomyces Pleomorphic → do not have any characteristic shape, Mycoplasma  6 Bacterial Cell Structure Nucleoid (nuclear material) § No nuclear membrane → nucleoid § 10% of cell content § Circular, double-stranded DNA § Bacterial chromosome length is 1 mm § Invagination of plasma membrane towards cytoplasm: mesosome  7 Bacterial Cell Structure Cytoplasm and cytoplasmic structures § Bacterial cytoplasm; transparent, colloidal system § Cytoplasm contains water, ions, metabolites for biosynthesis, metabolite products of catabolism and macromolecules § Proteins, nucleic acids and granules § Ribosomes (70s) § Plasmids and transposons § No mitochondria and chlorophlast therefore electron transport enzymes are found in the cytoplasmic membrane  8 Bacterial Cell Structure Granules § Concentrated deposits of certain substances located in the cytoplasm of certain bacteria are known as cytoplasmic granules or inclusion bodies. § They serve as storage areas for complex polymers used for nutrients and energy § If the nitrogen, sulphur, phosphate resources are insuffient: § Some bacteria convert carbon into poly-(3-hydroxybutyric acid polymers (PHB granules) § Some bacteria convert carbon into polysaccharides and glycogen (glycogen and polysaccharide granules) § Granules are used as energy source for the synthesis of proteins and nucleic acid § Most bacteria store polyphosphate granules during ATP synthesis § Volutin granules and metachromatic granules (eg. Corynebacterium)  9 Bacterial Cell Structure Ribosomes § Main structure: RNA, function: protein sythesis § 80% of cellular RNA is found in ribosomes § Bacterial ribosomes: 50S and 30S subunits make 70S § Eukaryotic ribosomes: 60S and 40S subunits make 80S IMPORTANT: The differences between rRNA of bacteria and eukaryotes allows the use of antibiotics that inhibit bacterial protein synthesis without having an effect on the human rRNA  10 Bacterial Cell Structure Plasmids § Plasmids are extrachromosal DNA molecules which are distinct from a cell’s chromosomal DNA § Can be transferred from one bacterium to another § Present in Gram positive and Gram negative bacteria § Resistance to antibiotics in bacteria, to heavy metals such as silver and mercury and UV § Provide bacteria with the genetic advantage of having fimbriae, hemolysins and exotoxins  11 Bacterial Cell Structure Transposons § DNA sequence that can change its position within a genome, can move between plasmids and bacteriophages § Called jumping genes § Difference from plasmids, cannot independently multiply § Responsible for bacteria problematic in hospital infections – most resistance genes are acquired by transposons  12 Cytoplasmic membrane § Surrounds the cell cytoplasm in all bacteria § Contains phospholipids, proteins and carhohydrates in small amounts § In Gram negative bacteria, is also called inner membrane § Cytoplasmic membrane; phopholipid bilayer with embedded proteins – mosaic apperance § Prokaryotic cells cannot synthesize sterol (only mycoplasma membranes contain sterol eg:cholesterol)  13 Functions of cytoplasmic membrane 1. Selective permeability and transport of various molecules into the cell 2. Electron transport and oxidative phosphorylation 3. Production of hydrolytic enzymes 4. Biosynthesis function 5. Chemotactic system  14 Functions of cytoplasmic membrane Mesosome § Invaginations in the plasma membrane towards the cytoplasm 2 types: 1) Septal mesosomes: in contact with the bacterial nucleoid, helps to form the plasma membrane during bacterial cell division 1) Lateral mesosome: located in the periphery, and not associated with nucleus. Involved in plasmid multiplication and spore formation  15 Bacterial Cell Structure Cell wall § Rigid layer lying outside the cell membrane, gives bacteria their shape § Multilayered, complex and resistant – protects bacterial integrity § Different layers contain antigenic determinants Peptidoglycan layer § Fundamental structure in bacterial cell wall is peptidoglycan (murein, mucopeptide) § Present in all prokaryotes except mycoplasma and archeabacteria § Not present in humans, therefore is the target of antibacterial drugs (eg. penicillin and cephalosporins) § Lysozyme enzyme (muramidase) present in animal secretions including tears, saliva and other body fluids; destroys peptidoglycan (protects against bacterial infections)  16 Bacterial Cell Structure Peptidoglycan layer The glycan chain acts as a backbone to peptidoglycan, and is composed of alternating residues of N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG). They are connected by inter-peptide bridges.  17 Bacterial Cell Structure § Gram (+) bacteria: crosslinked polymers, around 25 layers § Makes up more than 50-90 % of the cell wall § Gram (-) bacteria: only a thin layer of peptidoglycan § Makes up more than %5-10 of the cell wall  18 Bacterial Cell Structure Penicillin-binding proteins (PBPs) § Bacterial proteins found in the cell wall and essential for cell wall synthesis § PBPs bind to penicillin and other β-lactam antibiotics § Member of group of enzymes called transpeptidases § 4 PBPs in bacteria § Inhibition of PBPs by β-lactam antibiotics causes cell lysis 19  20 Bacterial Cell Structure Teichoic acid § Lipoteichoic/teichoic and teichouronic acid are found in the peptidoglycan layer § Anionic glycopolymers (gives rigidity) § Made up of ribitol phosphate and glycerol phosphate § Lipoteichoic acid is a major surface antigenic determinant and adhesin in Gram positive bacteria § Not found in Gram negative bacteria § Protects bacteria from the effects of lysozyme § Teichoic acid also acts as a receptor molecule for Gram positive bacteriophages 21  22 Bacterial Cell Structure  23 Gram negative bacteria - cell wall § Thinner peptidoglycan layer compared to Gram positive bacteria § Complex structure above the peptidoglycan layer made up of lipoprotein and lipopolysaccharides § No teichoic acid (different to Gram positive bacteria) § Periplasmic space between the lipoprotein layer and the cytoplasmic membrane (various enzymes and proteins exist in this space)  24 Gram negative bacteria - cell wall Components of Gram negative bacteria cell wall and their properties: (inside to outside) § Peptidoglycan layer: Thinner than Gram positive and not on the exterior surface § Lipoprotein layer: Connects exterior membrane to peptidoglycan (stabilizes the exterior membrane) § Exterior (outer) membrane: Different to other membranes, made up of two layers § Inner layer made up of phospholipids, external layer made up of lipopolysaccharides  25 Gram negative bacteria - cell wall Lipopolysaccharide (LPS): Gram negative cell wall LPS is made up of 3 parts § Lipid A: Glucosamine disaccharides and C14 fatty acids (3-hydroxyl-myristic acid) § Core: Polysaccharide structure, similar in all Gram-negatives § O-Polysaccharide: Linear trisaccharide, branched tetrasaccharides or pentasaccharides § connects to core § makes up bacterial specific O antigen (somatic antigen)  26 Gram negative bacteria - cell wall § LPS is toxic to humans and animals and called endotoxin in Gram negative bacteria § Endotoxins may cause fever, shock and death § Lipid A domain is responsible for the toxicity  27 Gram negative bacteria - cell wall O Polysaccharide: § Major surface antigen called the O antigen § Multiple variants of O antigens (Salmonella genus has more than 1000 O antigens identified)  28 Protoplasts, Spheroplasts and L forms § Lysozyme or antibiotics such as penicillin destroy the cell wall § In Gram (+) forms that do not contain cell wall are called protoplasts, in Gram (-) spheroplast § If the cells without cell wall has the property of multiplication and division, they are called L forms § L forms are difficult to culture § L forms hidden in the body may revert back to normal forms and may cause relapse (re-infection)  29 Bacterial Surface Structures Capsule or slime layer is used to describe glycocalyx which is a thin, high molecular weight secretory substance present in many bacteria external to cell wall It is composed of polysaccharide, polypeptide, or both. If glycocalyx is rigid and organized in a tight matrix, it is called capsule and most capsules consist of polysaccharides.  31 Functions of the capsule § Capsule increases the virulence of many bacteria § Capsule protects bacteria from phagocytosis § Capsule protects bacteria from antibiotics and other environmental factors eg. detergents, dessication § Organisms produce protective antigens when encounter encapsulated bacteria § This property is used for the preparation of various vaccines – capsule polysaccharides can be used ans antigens (H. influenzae type b)  32 Bacterial surface formations Slime layer § If glycocalyx is more easily deformed and loosely attached to cell wall it is called slime layer. § a protein-linked (glycoproteins) and lipid-linked carbohydrate fuzz-like structure that spans the plasma membrane (also called biofilm layer) § contributes to cell-cell recognition, communication, and intercellular adhesion (bacterial adherence to host cell surface – skin, catheters)  33 Flagella § The locomotion organelle in bacteria, proteinaceous appendages which protrude from the cell body § Flagella size is 2-3 times the lenth of bacterial cell, diameter 12-30 nm § Flagella is found in more than half of bacilli, in all of spirilla bacteria, but not commonly seen in cocci § Flagella structure contains helical shaped proteins called flagellin which have antigenic properties § Flagellin protein is termed H antigen and varies amoing different bacteria types  34 Flagella § Single polar flagellum – monotrichous § Single flagellum on both sides – amphitrichous § Tufts of flagella at one or both sides – lophotrichous § Numerous flagella all over the bacterial body – peritrichous § No bacterial flagella - atrichious  35 Fimbria (Pili) § Most Gram negative bacteria contain filamentous projections § Fimbria is not a locomotion organelle § Thinner, shorter than flagella, without folding § Made up of simple protein units called pilin § Fimbria length, thickness, protein sequence and antigenic proteins vary in different bacteria types § Fibria divided into two types – normal and sex fimbria  36 Fimbria (Pili) § Normal fimbria allow bacterial colonization on a surface via adhesion – important in pathogenesis § Sex pili play a role in conjugation § Sex pilus have an empty cylindrical central space and transfer genetic material § Normal fimbria do not have a central canal § Sex pilus is longer and fewer than normal fimbria  37 Spores (Endospores) § Some bacteria form resistant spores when the environmental nurtients are scarce § Spores; are resistant to physical (heat, dryness, radiation) and chemical (toxic materials) environmental factors for extended periods of time § The role of spores is to protect bacteria in non-ideal conditions § Formation of spores – sporulation § Loss of spore-specific properties and conversion to vegetative cells - germination  38 Spore location  39 Bacterial staining techniques § Biological molucules in microorganisms do not absorb light § Therefore microorganisms are observed via staining § Staining molecules are absorbed differently and allow bacterial visualization § In generall, basic staining is used for bacteria § Acidic staining is generally used for contract in the background  40 Gram staining technique Gram Staining is the common, important, and most used differential staining techniques in microbiology, which was introduced by Danish Bacteriologist Hans Christian Gram in 1884. This test differentiate the bacteria into Gram Positive and Gram Negative Bacteria, which helps in the classification and differentiations of microorganisms.  41 Gram staining technique § This technique uses the basic stain crystal violet to stain bacteria. § Under the light microscope, bacteria that retain the stain appear purple (Gram positive), whereas bacteria which do not retain the dye appear pink (Gram negative).  42 Gram staining procedure 1. Bacterial sample is fixed on the a glass slide and stained with crystal violet 2. The dye is fixed into the bacterial cell wall using iodine 3. The sample is washed with absolute alcohol (%95 ethanol) 4. Non lipid-containing Gram positive bacterial cell wall is not affected by alcohol treatment and keeps the purple color 5. Gram negative cell wall is thinner and has a lipid structure therefore loses integrity with alcohol treatment and the dye diffuses out 6. Safranin is used to stain the transparent bacteria which appear pink in Gram positives  43 Gram staining procedure