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10-14-09 Structure/Classification/Replication 1. Identify the structural components of a bacterial cell.

The following characteristics are unique to bacteria (are not found in eukaryotes): Peptidoglycan** Cell wall* Single supercoiled circular chromosome Multiple plasmids 70S ribosomes instead of 80S Cellular respiration located on cell membrane rather than in mitochondria Flagella Does not contain sterols *Fungi, which are eukaryotes, do have cell walls. **Peptidoglycan is composed of prefabricated polysaccharide polymers crosslinked by peptide bonds. Such crosslinking is performed by transpeptidases, which are the target of penicillin. 2. Become familiar with the growth conditions, survival and death of bacteria. All bacteria grow asexually via binary fission. Pathogenic bacteria derive their energy by metabolizing sugars, fats, and proteins. These macronutrients are broken down into pyruvate, which is then fermented into different end products in different bacteria. The fermentation pathways and end products of pyruvate breakdown are used to distinguish different bacteria. However, different bacteria have different growth requirements. For example, E. coli can synthesize all AAs, nucleotides, lipids, and carbohydrates necessary for growth and division. On the other hand, T. pallidum has complex growth requirements. As noted below, some gram positive bacteria can enter a vegetative or dormant state under harsh environmental conditions. These are called spores. 3. Become familiar with the criteria for classification and identification of bacteria. Classification by phenotypic characteristics Morphological characteristics (size, shape*, colony formation, pigmentation) Biotyping (biochemical tests for proteases, lipases, nucleases, carbon sources) Serotyping (identification of antigens) Antibiogram patterns (susceptibility to antibiotics) Phage typing (susceptibility to bacteriophages) Classification by analytic characteristics cell wall fatty acid analysis whole-cell lipid analysis

whole-cell protein analysis (proteomics via mass spec) enzyme typing Classification by genotypic characteristics G:C ratio (an old method) DNA hybridization Nucleic acid sequence analysis Plasmid analysis Ribotyping Chromosomal DNA fragment analysis

*bacterial shapes include: coccus, bacillus, coccobacillus, fusiform bacillus, virbio, spirillum, and spirochete A very basic form of bacterial classification is gram positive VS gram negative bacteria. Gram + bacteria: have a high amount of peptidoglycan in their cell walls, allowing for retention of the dark blue/violet Gram stain. The key components of gram positive bacteria include: Thick peptidoglycan layer (thick cell wall) No outer membrane Lysozyme sensitive Teichoic acid: polymer of chemically modified ribitol or glycerol phosphate that adds rigidity to cell wall The nature of the modification can define the serotype of the bacteria Lipotechoic acid: lipid linked techoic acid Sporulation (inactive form during harsh conditions) Capsules* Biofilms** Gram bacteria: have a very thin peptidoglycan layer that takes up the counterstain (safranin red or fuschin) resulting in a red or pink color. Thin peptidoglycan layer (thin cell wall) Outer membrane Lysozyme resistant Periplasmic space: contains enzymes involved in transport, synthesis and degradation LPS: Gram negative bacteria are associated with greater pathogenicity because of the presence of LPS, which consists of Lipid A, core polysaccharide, and O antigen Lipid A is responsible for the endotoxin activity of LPS and is therefore essential for bacterial viability. It has been implicated with: fever Activating complement Cytokine release Leukocytosis (high leukocyte count) Thrombocytopenia Disseminated intravascular coagulation (DIC)

Decreased peripheral circulation Shock death The O antigen can be used to distinguish serotypes of a bacterial species Capsules* Biofilms**

*Capsule, also known as a glycocalyx, can be found on both forms of bacteria. It is a layer of high molecular weight polysaccharides and proteins loosely adherent to the cell wall. A loosely adherent capsule of non-uniform density is called a slime layer. Capsules are poorly antigenic (meaning that they arent easily recognized by the immune system) as well as antiphagocytic (so they make the bacteria difficult to eat). Consequently, they are considered a major virulence factor! Ex: capsulated Streptococcus pneumoneiae causes fatal infection, while the uncapsulated form doesnt cause disease at all. dextran and levan capsules allow bacteria to attach to tooth enamel **Biofilm is composed of polysaccharides that establish a bacterial community, protecting them from antibiotics and host defenses.