Medical Bacteriology I and II PDF

1 Introduction to Medical Bacteriology Dr. Özge YILMAZLI 2 Medical Bacteriology  Bacteria are relatively simple in structure. They are prokaryotic organisms — simple unicellular organisms with no nuclear membrane, mitochondria, Golgi bodies, or endoplasmic reticulum— that reproduce by asexual...

1 Introduction to Medical Bacteriology Dr. Özge YILMAZLI 2 Medical Bacteriology  Bacteria are relatively simple in structure. They are prokaryotic organisms — simple unicellular organisms with no nuclear membrane, mitochondria, Golgi bodies, or endoplasmic reticulum— that reproduce by asexual division  Bacterial cells range from 0,1(μm) to 5 μm in length and thus are not visible to the human eye without help of a microscope  The cell wall is the outermost component common to all bacteria (except Mycoplasma species) 3 Shapes of Bacteria Coccus: spherical or round shape Spirillum: a thick, rigid spiral shaped Bacillus: rod shaped, Vibrio: comma shaped Spirochete: a thin, flexible spiral shaped Cocobacillus: oval shaped 4 Shapes of Bacteria Diplococcus: division in one plane coccus arranged in pairs Streptococcus: division in one plane coccus arranged in chains Tetrad: division in two planes coccus arranged in squares of 4 Sarcina: division in three planes regularly coccus arenged in cubes of 8 Staphylococcus: division in three planes irregularly coccus arenged grape-like clusters 5 Bacterial Structure  Cytoplasmic Structures;  The cytoplasm of the bacterial cell contains the DNA chromosome, ribosomes, proteins, and metabolites  In contrast eukaryotes, most bacterial chromosomes are a single, double-stranded circle that is contained not in a nucleus  Bacteria have 70S ribosome  The cytoplasmic membrane has a lipid bilayer structure similar to the structure of the eukaryotic membranes, but it contains no steroids (e.g., cholesterol); (except Mycoplasma species) 6 7 Bacterial Structure  Cell Wall;  Rigid peptidoglycan (murein) layers surround the cytoplasmic membranes of most bacteria (except Mycoplasma species which have no peptidoglycan)  It is responsible for the shape of the cell  It protects cell from osmotic lysis and also provides a barrier against certain toxic chemical and biologic agents  Two different types of cell wall in bacteria, called Gram-positive and Gram-negative. The names originate from the reaction of cells to the Gram stain  Gram stain is a common technique used to differentiate two large groups of bacteria based on their different cell wall constituents. Developed in 1884 -Hans Christian Gram-, it’s been in use ever since 8 Bacterial Structure  Cell Wall;   Gram-positive bacteria have a thick, multilayered cell wall consisting mainly of peptidoglycan surrounding the cytoplasmic membrane and also includes other components such as proteins, teichoic and lipoteichoic acids, and complex polysaccharides Gram-negative bacteria have a thinner layer of peptidoglycan (10% of the cell wall) and external to the peptidoglycan layer is the outer membrane which contains lipopolysaccharide (LPS) 9 Gram Stain GRAM STAIN  Heat Fixation: A thin layer of the biological sample is spread on the slide and slide with biological sample is passed through a flame to paste the sample.  Crystal-violet dye(Primary dye): Cover the smear with crystal-violet dye (a purple colour dye) for 1 minute. Gently rinse off the stain with water  Gram's iodine (Mordant): Cover the smear with Gram’s iodine, the mordant, for 1 minute. Gently rinse off the stain with water 10 Gram Stain  Decolorisation solution: Run the acetone/alcohol decolorizer over the smear until the solution appears clear. Dye will flow from slide. Gently rinse with water  Safranin (counter stain): Cover the smear with safranin, the counterstain, for 30 seconds. Gently rinse the stain with water 11 Gram Stain alcohol 12 13 Bacterial Structure  External Structures;  Some bacteria have surface features external to the cell wall, such as capsule, flagella, and pili  Capsule; Some bacteria (gram-positive or gram-negative) are closely surrounded by loose polysaccharide or protein layers called capsules  The capsule can protect bacteria from phagocytosis by immune system,  The capsule can also act as a barrier to toxic hydrophobic molecules, such as detergents,  The capsule can promote adherence to other bacteria or host tissue surfaces 14 Bacterial Structure 15 Bacterial Structure  External Structures;  Flagella; Flagella are long, slender, thin hair like appendages which are responsible for the motility of bacteria. Flagella are made up of several thousand molecules of a protein subunit called flagellin  Flagella provide motility for bacteria, allowing the cell to swim (chemotaxis) toward food and away from poisons  Flagella are attached to cells in different places. As the number and location of flagella are distinctive for each genus, it can be used in the classification of bacteria.  There are four types of flagellar arrangement. Monotrichous, Lophotrichous, Amphitrichous, Peritrichous. MLAP 16 Bacterial Structure 17 Bacterial Structure  External Structures;  Fimbria (Pili); Fimbriae (pili) are hairlike structures on the outside of bacteria; they are composed of protein subunits (pilin)  Fimbriae can be morphologically distinguished from flagella because they are smaller and usually are not coiled in structure than flagella  Two classes can be distinguished:  Ordinary pili, which play a role in the adherence of bacteria to other bacteria or to the host cells  F pili (sex pili), which are responsible for the attachment pili (sex pili) bind to other bacteria and are a tube for transfer of large segments of bacterial chromosomes between bacteria 18 Bacterial Structure 19 Bacterial Structure  Special structure; Spore; Under poor environmental conditions, such as loss of a nutritional requirement, these bacteria can convert from a vegetative state to a dormant state, or spore  Some Gram positive—but never Gram negative—bacteria, such as members of the genera Bacillus and Clostridium are spore formers  The spore is a dehydrated multishelled structure that protects and allows the bacteria to exist in “suspended animation”  It contains a complete copy of the chromosome, the bare minimum concentrations of essential proteins and ribosomes, and a high concentration of calcium bound to dipicolinic acid 20 Bacterial Structure  Special structure; Spore;  The spore has an inner membrane, two peptidoglycan layers, and an outer keratin-like protein coat  Depletion of specific nutrients from the growth medium triggers a cascade of genetic events leading to the production of a spore (6 to 8 hours)  Germination of spores into the vegetative state is stimulated by disruption of the outer coat by mechanical stress, pH, heat, or another stressor and requires water and a triggering nutrient (90 min) 21 Bacterial Structure 22 Bacterial Metabolism 23 Bacterial Metabolism Bacteria must obtain or synthesize the amino acids, carbohydrates, and lipids used for building blocks of the cell. The minimum requirements for growth are;  Energy source,  Water,  Essential elements (C, O, H, N, S, P),  Important ions (K, Na, Mg, Ca, Cl),  Components of enzymes (Fe, Zn, Mn, Mo, Se, Co, Cu, Ni) 24 Bacterial Metabolism Oxygen (O2 gas), although essential for the human host, is actually a poison for many bacteria Microorganisms can be classified according to their oxygen requirements necessary for growth and survival;     Obligate aerobes require the presence of molecular oxygen for metabolism and growth Microaerophiles sit in the middle requiring 5% to 10% oxygen for optimal growth Facultative anaerobes grow in either the presence or the absence of oxygen Obligate anaerobes can’t grow in the presence of oxygen 25 Bacterial Metabolism Obligate anaerobes Obligate aerobes Microaerophiles Facultative anaerobes 26 Bacterial Metabolism Bacteria multiply by binary fission;  Cell elongates,  DNA is replicates,  Cell wall and plasma membrane begin to constrict,  Cross-wall forms, completely separating the two DNA copies  Cell separate 27 28 Bacterial Metabolism  Media; are an artificially prepared mixture of various nutrients in appropriate concentration and are prepared for considering the biochemical requirements of microbes  Media are used in the laboratory for the cultivation of bacteria  Culture; Microbes that grow and multiply in or on a culture media  The growth rate of a bacterial culture depends on three factors: the species of bacterium, the chemical composition of the medium, and the temperature.  Some species can double in 20 minutes (Escherichia coli)  Some take almost 20 hours (Mycobacterium tuberculosis) 29 Bacterial Metabolism When bacteria are added to a medium that provides all of the nutrients that are necessary for their growth, the population exhibits four phases of growth that are representative of a typical bacterial growth curve;     Lag Phase; When bacteria are added to a new medium, they require time to adapt to the new environment before they begin dividing Log Phase; During this phase, the bacteria will grow and divide with a doubling time characteristic of the strain and determined by the conditions. Period of most rapid growth. Stationary Phase; The culture eventually runs out of metabolites, or a toxic substance builds up in the medium; the bacteria then stop growing and enter the stationary phase, Death phase; Population size begins to decrease. Cell number decreases at a logarithmic rate and cells lose their ability to divide. Number of cells dying > Number of cells produced 30 Bacterial Metabolism 31 Bacterial Metabolism Culture media can be subdivided into four general categories:  Enriched Nonselective Media  Selective Media  Differential Media  Specialized Media 32 Bacterial Metabolism  Enriched Nonselective Media;These media are designed to support the growth of most organisms without fastidious growth requirements (e.g., Blood Agar)  Selective Media; has added inhibitors that discourage the growth of certain organisms without inhibiting growth of the organism being sought.   Differential Media ; A differential medium permits the differentiation of organisms that grow on the medium.   For example, MacConkey Agar inhibits growth of Gram-positive bacteria and thus is selective for Gram-negative bacteria For example, MacConkey agar differentiates between lactose-fermenting and non-lactose fermenting Gram (-) bacteria. Gram-negative bacteria capable of fermenting lactose produce pink colonies, whereas those that are unable to ferment lactose produce colorless colonies Specialized Media; A large variety of specialized media have been created for the detection of specific organisms that may be fastidious. (e.g., BCYE Agar) 33 Bacterial Metabolism 34 Bacterial Metabolism 35 Bacterial Metabolism  Colony; A colony is defined as a visible mass of microorganisms all originating from a single mother cell, therefore a colony constitutes a clone of bacteria all genetically alike  Bacteria grow in colonies; each colony is like a city of as many as a million or more organisms. The sum of their characteristics provides the colony with distinguishing characteristics such as color, size, shape, and smell. 36 Bacterial Metabolism 37 Bacterial Genetics  4. Bakteri Genetiği 38 Bacterial Genetics  Bacterial chromosomes are a single, double-stranded circle that is contained not in a nucleus  Bacteria may also contain extrachromosomal genetic elements such as plasmids or bacteriophages.  These elements are independent of the bacterial chromosome and in most cases can be transmitted from one cell to another 39 Bacterial Genetics 40 Bacterial Genetics  Genetic material can be transferred between different strains of bacteria  The exchange of DNA between cells allows exchange of genes and characteristics between cells, thus producing new strains of bacteria.  This exchange may be advantageous for the recipient, especially if the exchanged DNA encodes antibiotic resistance.  The exchange of genetic material between bacterial cells may occur by one of three mechanisms;     Transformation Transduction Conjugation Transposition 41 Bacterial Genetics  Transformation Transformation is the process by which bacteria take up fragments of naked DNA and incorporate them into their genomes 42 Bacterial Genetics  Transduction Transduction is the transfer of genetic information from donor to recipient cell by viruses of bacteria called bacteriophages 43 Bacterial Genetics  Conjugation Conjugation results in one-way transfer of DNA from a donor (or male) cell to a recipient (or female) cell through the sex pilus The mating type (sex) of the cell depends on the presence (male) or absence (female) of a conjugative plasmid, such as the F plasmid 44 Bakterial Genetics 45  Transposition Mobile DNA segments that move in genom, between plasmids and plasmid and genom. 46