1. Microbial metabolism (energy production and storage mechanisms)
   1. Catabolic reactions (breakdown of larger molecules into smaller molecules, usually releasing energy which is trapped in chemical bonds of ATP)
      1. Glycolysis
         1. Conversion of one glucose to two pyruvate in series of enzyme catalyzed reactions
         2. Energy stored in chemical bonds attaching phosphate groups to adenosine to make ATP (esp. in the third phosphate bond)
         3. Products: net gain of 2 ATP molecules, 2 pyruvate molecules from each glucose
      2. Fermentative breakdown of pyruvate
         1. No oxygen required
         2. Specific enzymes required for each kind of fermentation end product
         3. No additional ATP’s made
         4. Examples: Lactic acid fermentation (Streptococcus mutans, involved in dental caries), alcoholic fermentation (yeast), mixed acid fermentation (E. coli, and many other bacteria), propionic fermentation (Propionibacterium), butanediol fermentation (Enterobacter aerogenes), butyric/butylic fermentation (anaerobes)
      3. Oxidative breakdown of pyruvate (usually in aerobic respiration)
         1. Oxygen required (terminal electron acceptor for electron transport system)
         2. Enzymes required at each step
         3. Lots more ATP’s made means that organisms can grow faster
         4. Krebs cycle (or TCA cycle)
            1. Starting materials: pyruvate
            2. Products: carbon dioxide, high energy electrons
         5. Electron transport system
            1. Starting materials: high energy electrons, oxygen
            2. Products: ATP, water
         6. Net gain of 38 ATP’s from one glucose molecule
      4. Other macromolecules (fats, proteins, carbohydrates) can be broken down to enter catabolic pathways at points beyond glucose or can be converted to glucose
   2. Energy currency: ATP (energy released during catabolic reactions trapped in chemical bonds in ATP, an energy storage molecule)
   3. Enzyme-catalyzed reactions (enzyme/substrate/product)
      1. Example: invertase converts sucrose to fructose and glucose
      2. Example: amylase converts starch to sugars
      3. Enzyme lowers energy of activation to increase efficiency of reaction
   4. Anabolic reactions (biosynthesis of larger molecules from smaller molecules, usually requiring consumption of energy in the form of ATP)
      1. Amino acids used to make proteins
      2. Nucleotides used to make nucleic acids
      3. Glycerol and fatty acids used to make lipids
      4. Simple sugars used to make complex sugars and starches (carbohydrates)
      5. Macromolecules used to make new cell components
2. Microbial genetics
   1. Maintenance and use of genetic information
      1. One circular chromosome (genome, DNA) contains one copy of each bacterial gene
      2. Genetic changes easy to observe in bacteria because effect of change is not masked by other genes and because bacteria grow rapidly
      3. DNA structure (A, T, C, G bases attached to sugar-phosphate backbone, double-stranded helix)
      4. Order of bases in DNA contains genetic information encoded in base sequence
      5. Base sequence is read in three-base units (triplets, codons)--any change in one unit affects the way the others are read, change in first base in triplet more important than change in third
      6. DNA replication (copying DNA to make new DNA, requires DNA polymerase and energy)
      7. Transcription (using DNA to make messenger RNA, requires RNA polymerase and energy, ribose instead of deoxyribose, uracil instead of thymine)
      8. Translation (using mRNA as template, with ribosome and tRNA-amino acids, to make proteins)
      9. Order of bases in mRNA determines order of amino acids in protein and therefore the structure and function of the protein
   2. Genetic change in bacteria
      1. Mutation
         1. Change in order of bases in existing DNA
         2. Spontaneous (rare) or induced (more common)
         3. Lethal (bacteria are killed) or nonlethal (sublethal, change in bacterial characteristic such as amino acid synthesis, nutrient utilization, pigment production, resistance to antibiotics or virus infection)
         4. Achieved by inserting a base, deleting a base, or changing a base
         5. Radiation
            1. Nonionizing--ultraviolet light (formation of pyrimidine dimers that lead to errors when DNA is used, nonpenetrating)
            2. Ionizing--X rays, gamma rays (chromosome breakage)
            3. Example: inducing a sublethal pigment production mutation in Serratia marcescens with UV light
         6. Chemical mutagens
            1. Induce mutations by many mechanisms
            2. Example: 5-bromuracil (acts as base analog for thymine, but may bind to guanine instead of adenine, causing a complete base pair change in three generations)
         7. Ames test
            1. Screening test to detemine whether chemical is mutagenic for bacteria and to infer from that the probability that the chemical may be carcinogenic for human cells)
            2. Test for revertants from auxotrophy (mutants unable to synthesize histidine, unable to grow on a medium without histidine) to prototrophy (wild type, can synthesize histidine and grow on medium without histidine) after exposure to chemical being tested
            3. Growth on medium without histidine indicates that the chemical has induced mutation
            4. 90% of chemicals that are mutagenic in Ames test are carcinogenic for human cells
      2. Recombination
         1. Genetic change in which new section of DNA is incorporated into existing bacterial chromosome
         2. Occurs after transfer of DNA from donor cell to recipient cell
         3. First predicted after experiment done by Griffith with mice injected Streptococcus pneumoniae (see Figure 8.1 in text)
         4. Types of genetic transfer in bacteria
            1. Transformation (naked DNA transferred, no cell contact required, plasmid transfer)
            2. Conjugation (direct cell-to-cell transfer involving F pili, cell contact required)
               1. May result in change in mating type, but no genetic change (F+ x F- mating)
               2. May result in genetic change by recombination, but rarely change in mating type (Hfr x F- mating)
            3. Transduction (bacteriophage-mediated transfer, no contact required, very small amount of DNA transferred)
         5. Experimental recombination (genetic engineering)
            1. Isolation of human gene (cut out with restriction endonuclease)
            2. Isolation of bacterial plasmid (cut with same restriction endonuclease)
            3. Insertion of human gene into bacterial plasmid (complementary sticky ends of DNA)
            4. Recombinant plasmid sealed with ligase
            5. Insertion of recombinant plasmid into recipient bacterial cell (transformation)
            6. Growth of bacteria to produce gene product (human growth hormone, insulin)
            7. Growth of bacteria to produce copies of human gene (gene cloning)
3. Antimicrobial chemotherapy
   1. Antibiotics are antimicrobial drugs that are produced by or derived from substances produced by microorganisms--many antimicrobial drugs are not antibiotics.
   2. Properties of an ideal antibiotic (broad spectrum, stable--long shelf life, soluble in body fluids, stable toxicity, nonallergenic, reasonable cost, selectively toxic, not likely to induce bacterial resistance)
   3. Selective toxicity (toxic to bacteria or fungi, nontoxic to human cells)
   4. Major genera that produce clinically useful antibiotics (Bacillus, Streptomyces, Cephalosporium, Penicillium)
   5. Measurement of bacterial susceptibility to antibiotics: zones of inhibited growth on Kirby-Bauer disk diffusion plates
   6. Antibiotics are not useful for treating virual disease (viruses grow in human cells; hard to make drug that affects virus without damaging human cell also
   7. Major targets of antimicrobial activity
      1. Cell wall synthesis: penicillins, cephalosporins (beta-lactamase producing bacteria resistant to both, require active cell wall synthesis in actively growing cultures), bacitracin
      2. Cell membrane function: amphotericin B (no growth requirement, changes membrane permeability by binding to sterols in fungal membranes, more side effects since membranes similar in all cells)
      3. DNA synthesis: mitomycin C (antitumor, not clinically useful)
      4. Protein synthesis
         1. Transcription (DNA --> mRNA): rifampin (TB), actinomycin D
         2. Translation (mRNA--> protein):
            1. Block movement of ribosome along mRNA: streptomycin, tetracycline
            2. Prevent peptide bond formation by binding to ribosome: chloramphenicol, erythromycin
      5. Antimetabolites (structural analogs of natural substances important in metabolism): PASA, sulfa drugs, INH
         1. PASA very similar in structure to PABA, required by bacteria (but not human cells) for synthesis of folic acid
         2. When PASA is used in synthesis of folic acid, results in nonfuctional folic acid analog and bacterial cell dies
   8. Antibiotic Resistance
      1. Transfer of resistance plasmids from resistant strains (beta-lactamase producing bacteria)
      2. Variation in resistant populations during the course of antibiotic treatment (Figure 14.11)
      3. Double drug combinations (Fig. 14.16.: combination of quinolone with beta-lactam ring antibiotic, resistant bacteria may break lactam ring, but would still be susceptible to released quinolone)

Host-Pathogen Relationships: Mechanisms of Pathogenesis

1. Introduction
   1. Vocabulary: host, pathogen, infection, disease, opportunistic pathogen, normal flora, virulence
   2. Opportunistic pathogen: normally harmless, but can take advantage of weakened host defenses or can acquire more virulence factors
      1. Weakened host: very young, very old, malnourished, cancer pt. or treatment, HIV disease, chronic disease
      2. Enhanced virulence: normal flora E. coli can become pathogenic if they acquire capacity to synthesize certain toxins (E. coli O157:H7)
   3. Highly virulent pathogen: Many successful strategies for invading and colonizing host, few cells/viruses may be required for successful colonization
   4. Less virulent pathogen: Larger population required for successful colonization of host, more susceptible to host defenses, strategies for colonization less effective
   5. If pathogen strategies outweigh host defenses, interaction may result in acute disease, which may result in death of host, depending on treatment and severity of disease
   6. If host defenses outweigh pathogen strategies, host may successful resist invasion by pathogen and interaction may result in elimination of pathogen from host or control of pathogen spread in host
   7. If interaction is balanced, may result in chronic disease
2. Transmission between hosts
   1. GI tract route tract
      1. Fecal contamination of food or water (portal of exit: anus/feces; portal of entry: mouth, oropharynx)
      2. Feces, flies, fingers, food sequence
      3. Examples: cholera, bacillary dysentery, typhoid fever, salmonellosis and other food poisoning
   2. Respiratory route
      1. Pathogens are usually transferred in aerosols produced by coughing or sneezing, resulting in droplets that can be inhaled
      2. Pathogens can also be transferred when hands are contaminated with mucous secretions from nose or mouth
      3. Upper respiratory examples: diphtheria, influenza, rhinoviruses, adenoviruses, coronaviruses
      4. Lower respiratory examples: pneumonias, tuberculosis, influenza
   3. Direct contact route (transfer between infected tissues)
      1. Contact may be sexual or nonsexual
      2. Skin to skin, skin to mucous membrane, or mucous membrane to mucous membrane contact may be route of transmission
      3. Contact may deliver organisms to new host in fluid from lesions or with skin particles
      4. Examples: impetigo, fungal skin infections, syphilis, gonorrhea, chlamydia, herpes, HIV, HPV
      5. Vector-bourne route (living organism acts as agent for transmission between hosts)
         1. Bubonic plague (rat, rat flea, human cycle)
         2. Malaria (mosquitos)
         3. Rocky Mountain spotted fever (ticks)
         4. Lyme disease (deer ticks)
   4. Invasion and colonization of host by pathogen
      1. Invasion and colonization of host by pathogen
         1. Resistance to elimination by host defenses
            1. Resistance to phagocytosis (due to encapsulation)
            2. Resistance to outward flow of mucous secretions (due to pili and adhesins)
         2. Penetration from body surface to deeper tissues (Vibrio cholerae, remains on surface of intestinal lining; Salmonella typhi, invades lining and then moves out to other organs)
         3. Ability to survive inside host cells (intracellular growth)
            1. Salmonella typhi, typhoid fever, in macrophages
            2. Mycobacterium tuberculosis, tuberculosis, in macrophages
         4. Nutritional requirements/tissue specificity
            1. Brucella abortus, erythritol in placenta of cows
            2. Mycobacterium tuberculosis, oxygen level in tissues
      2. Virulence factors
         1. Nontoxic virulence factors
            1. Capsule (prevents phagocytosis, Streptococcus pneumoniae, pneumococcal pneumonia)
            2. Pili (attaches bacteria to linings of genital tract--Neisseria gonorrhoeae, gonorrhea, or to the linings of the respiratory tract--Bordetella pertussis, whooping cough)
            3. Adhesins (enteropathic E. coli, many other GI tract pathogens)
            4. Enzymes (aggressins)
               1. Proteases, lipases, amylases: release nutrients
               2. DNase, hyaluronidase: promote spread of bacteria through tissue
         2. Toxic virulence factors
            1. Exotoxins (protein, extremely potent, specific action unique to each type of exotoxin, relatively heat labile, can be used to prepare a toxoid, usually released from living cell) (cholera toxin, hemolysins, tetanus toxin, diphtheria toxin)
            2. Examples of exotoxins
               1. Diphtheria, Corynebacterium diphtheriae--toxin prevents protein synthesis by host cells
               2. Botulism, Clostridium botulinum--toxin inhibits nerve signal transmission to muscles, resulting in flaccid paralysis
               3. Tetanus, Clostridium tetani--toxin causes uncontrolled nerve signal transmission, resulting in violent uncontrolled muscle spasms
               4. Cholera, Vibrio cholerae--toxin causes massive fluid loss from intestinal lining cells
            3. Endotoxins
               1. Lipopolysaccharide, lipid portion toxic, part of Gram negative cell wall, only released when cell dies
               2. Relatively heat stable, cannot be used to prepare a toxoid
               3. Similar structure and effects in all organisms
               4. Symptoms include fever, circulatory system disturbance, intravascular clots, inflammation
               5. Less potent than exotoxins, but if released in large amounts (i.e., due to massisve die-off of gram negative bacteria in peritonitis) can cause circulatory system collapse and endotoxic shock (irreversible)