Microbial Metabolism, Growth, and Control: Study Notes for Health Sciences

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Microbial Metabolism

Overview of Metabolism

Metabolism encompasses all chemical reactions within a cell, divided into energy-releasing (catabolic) and energy-consuming (anabolic) processes. Enzymes, which are highly specific proteins, regulate these reactions.

Catabolism: Exergonic reactions that break down complex molecules, releasing energy (often via oxidation).
Anabolism: Endergonic reactions that build complex molecules, requiring energy input (often via reduction).
Enzymes: Biological catalysts that lower activation energy, speeding up reactions without being consumed. Types include hydrolases, lyases (catabolic), ligases, synthases (anabolic), isomerases, and transferases.
Optimal Conditions: Enzyme activity depends on pH and temperature; inhibitors can reduce activity.

Reaction Coupling

Cells couple exergonic and endergonic reactions to drive essential processes.

Exergonic Reactions: ; spontaneous, release energy.
Endergonic Reactions: ; require energy input.

Catabolism of Macromolecules

Microbes degrade macromolecules externally using secreted enzymes before absorbing monomers for energy extraction.

Carbohydrates: Cellulose, chitin, agar, lignin, and pectin are hydrolyzed into monomers.
Proteins: Proteolytic enzymes cleave proteins into amino acids; some enzymes (e.g., hyaluronidase, collagenase) aid in tissue invasion.
Lipids: Fatty acids are separated from glycerol and transported into the cell.
Nucleic Acids: DNases degrade DNA, aiding in tissue invasion (e.g., Staphylococcus aureus).

Catabolism of Monomers

Monosaccharides: Enter glycolysis, producing ATP and NADH; pyruvate proceeds to the Krebs cycle or fermentation.
Amino Acids: Enter glycolysis, pyruvate oxidation, or Krebs cycle after deamination.
Fats: Broken into acetyl groups for the Krebs cycle; glycerol enters glycolysis.

Glycolysis Pathways

Glycolysis splits glucose into two three-carbon molecules, generating ATP and NADH. Pathways include:

Embden-Meyerhof Pathway: Used by most organisms.
Entner-Doudoroff Pathway: Used by some Gram-negative bacteria.
Phosphoketolase Pathway: Found in certain prokaryotes and eukaryotes.

Electron Carriers: NAD+ and NADP+ shuttle electrons; NADH for energy, NADPH for biosynthesis.

Fermentation

Fermentation regenerates NAD+ by transferring electrons to organic molecules, producing various end products. Types include:

Homolactic Acid: Only lactic acid produced.
Heterolactic Acid: Lactic acid, ethanol, and CO2 produced.
Alcoholic: Ethanol and CO2 produced.
Propionic, Butyric, Butanediol, Mixed Acid: Various acids and gases produced; important for bacterial identification (e.g., methyl red and Voges-Proskauer tests).

Krebs Cycle (Citric Acid Cycle)

A series of enzyme-catalyzed reactions extracting high-energy electrons from acetyl-CoA, producing ATP, NADH, and FADH2.

Electron Transport Chain and Chemiosmosis

High-energy electrons from NADH and FADH2 are transferred through membrane-bound carriers, pumping H+ to create a gradient.
ATP synthase uses this gradient to generate ATP (oxidative phosphorylation).
Oxygen is the final electron acceptor in aerobic respiration.
Facultative anaerobes can switch between aerobic respiration and fermentation.

Anaerobic Respiration

Uses electron acceptors other than oxygen (e.g., nitrate, sulfate).
Denitrifying bacteria reduce nitrate to nitrogen gas, important in the nitrogen cycle.
Sulfate-reducing bacteria produce H2S, contributing to environmental processes.

Metabolic Diversity

Phototrophs: Use light for energy.
Chemotrophs: Use chemicals for energy (chemoorganotrophs: organic; chemolithotrophs: inorganic).
Autotrophs: Use CO2 as carbon source.
Heterotrophs: Use organic compounds as carbon source.
Combined terms: photoautotroph, chemoheterotroph, etc.

Biochemical Tests for Bacterial Identification

Phenol Red Test: Detects acid production from fermentation (yellow at low pH).
Methyl Red Test: Detects strong acid production (red at pH < 4.4).
Citrate Test: Identifies bacteria that use citrate as sole carbon source (color change from green to blue).
Catalase Test: Detects catalase enzyme (breaks down H2O2).
Oxidase Test: Detects cytochrome c oxidase.
IMViC Tests: Indole, Methyl Red, Voges-Proskauer, Citrate; differentiates Enterobacteriaceae.

Organism	Indole	Methyl Red	Voges-Proskauer	Citrate
Escherichia coli	+	+	-	-
Enterobacter, Klebsiella	-	-	+	+
Citrobacter freundii	-	+	-	+

Growth and Culturing of Bacteria

Temperature Requirements

Cardinal Temperatures: Minimum, optimum, and maximum for growth.
Psychrophiles: -10°C to 25°C (optimum 10–20°C).
Mesophiles: 10°C to 45°C (optimum 20–40°C); includes most human pathogens.
Thermophiles: 30°C to 80°C (optimum 45–80°C).
Hyperthermophiles: 80°C and above; many Archaea.

pH and Moisture Requirements

Acidophiles: Grow at low pH (e.g., Helicobacter pylori).
Alkalophiles: Grow at high pH (8.5–11.5).
Water Activity (aw): Most bacteria require aw > 0.90; halotolerant and halophilic microbes tolerate or require low aw.
Osmotolerant/Xerophiles: Survive high solute or low water environments.

Inorganic and Organic Nutritional Needs

Nitrogen: Needed for amino acids, nucleic acids; sources include ammonia, nitrate, and nitrogen gas.
Sulfur: For amino acids, coenzymes.
Phosphorus: For nucleic acids, ATP.
Trace Elements: Iron, calcium, sodium, etc.
Vitamins: Often required as growth factors.

Oxygen Requirements

Aerobes: Require oxygen.
Microaerophiles: Require lower oxygen levels.
Facultative Anaerobes: Grow with or without oxygen.
Aerotolerant Anaerobes: Tolerate oxygen but do not use it.
Obligate Anaerobes: Cannot tolerate oxygen.

Culture Media Types

Nondefined (Complex) Media: Contains extracts (e.g., beef, yeast); composition varies.
Defined (Synthetic) Media: Exact chemical composition known.
Broth, Agar Plates, Deeps, Slants: Used for different culturing purposes.
Enrichment Media: Enhances growth of specific microbes.
Selective Media: Inhibits unwanted microbes, promotes desired ones.
Differential Media: Distinguishes microbes by colony appearance or metabolic products.

Media Type	Purpose	Example
Enrichment	Enhance growth of specific microbes	Selenium for Salmonella
Selective	Suppress unwanted, encourage desired	Sabouraud dextrose with antibiotics
Differential	Distinguish by appearance/metabolism	Blood agar, MacConkey agar

Measuring Bacterial Growth

Binary Fission: Main method of bacterial division (not mitosis).
Generation (Doubling) Time: Time required for population to double.
Population Growth Curve: Four phases: lag, exponential, stationary, decline.

Phase	Description
Lag	Adaptation, little division
Exponential	Rapid cell division
Stationary	Growth rate = death rate
Decline	Death rate > growth rate

Counting Methods: Direct microscope count, viable plate count (colony forming units), turbidimetric (optical density), dry weight.

Control of Microbial Growth

Key Terms

Sterilization: Complete destruction of all life forms, including spores and viruses.
Disinfection: Destruction of most life forms (not necessarily spores); disinfectants for surfaces, antiseptics for living tissue.
Sanitizing/Degerming: Milder forms of disinfection.
“-cide” Suffix: Kills microbes (e.g., bactericide).
“-stat” Suffix: Inhibits growth (e.g., bacteriostat).

Physical Methods of Control

Heat: Denatures proteins, oxidizes cell contents, breaks DNA.
Thermal Death Point: Lowest temperature to kill all microbes in 10 min.
Thermal Death Time: Minimum time to kill all microbes at a set temperature.
Autoclaving: Moist heat under pressure (121°C).
Pasteurization: Mild heat (e.g., 65°C for 30 min).
Dry Heat: Flaming, hot air (higher temps needed).
Filtration: Removes microbes from liquids/air (0.22 µm filters).
Low Temperature: Slows growth, can eventually kill.
High Pressure, Desiccation, Osmotic Pressure, Radiation, Ultrasound: Various effects on microbial viability.

Chemical Methods of Control

Phenols/Phenolics: Disrupt membranes, denature proteins (e.g., Lysol™).
Bisbiguanides: e.g., chlorhexidine in surgical soap.
Halogens: Disrupt protein function (e.g., chlorine, iodine).
Alcohols: Disrupt proteins and lipids (e.g., ethanol, isopropanol).
Heavy Metals: Bind sulfhydryl groups, denature proteins.
Surface Active Agents: Soaps, detergents, quaternary ammonium compounds (quats).
Aldehydes: Disrupt proteins (e.g., formaldehyde).
Peroxygens/Oxidizing Agents: e.g., hydrogen peroxide.
Plasmas, Supercritical Fluids: Advanced sterilization methods.

Antimicrobial Drugs

History and Key Terms

Paul Ehrlich: Developed Salvarsan, first chemotherapeutic agent.
Alexander Fleming: Discovered penicillin.
Gerhard Domagk: Developed sulfa drugs.
Antibiotic: Natural antimicrobial produced by organisms.
Antimicrobial Drug: Any agent (natural, synthetic, semisynthetic) for microbial diseases.

Attributes of Effective Antimicrobials

Soluble in body fluids, non-toxic, selective toxicity, non-allergenic, bacteriostatic or bactericidal, minimal disruption of normal flora, resistance not easily acquired, long shelf-life, reasonable cost, appropriate spectrum (broad or narrow).

Mechanisms of Action

Inhibit Cell Wall Synthesis: β-lactams (penicillins, cephalosporins), carbapenems, vancomycin, isoniazid.
Inhibit Protein Synthesis: Chloramphenicol, aminoglycosides, tetracyclines, macrolides.
Injure Plasma Membrane: Daptomycin (Gram+), polymyxin B (Gram-), bacitracin.
Inhibit Nucleic Acid Synthesis: Rifamycins, quinolones/fluoroquinolones.
Inhibit Synthesis of Metabolites: Sulfonamides (folic acid synthesis inhibitors).

Antifungal, Antiviral, Antiprotozoan, and Antihelminthic Drugs

Antifungals: Target ergosterol (nystatin, amphotericin), β-glucan (echinocandins), nucleic acid synthesis (flucytosine).
Antivirals: Block entry, uncoating, nucleic acid synthesis, assembly, or exit; include interferons and antiretrovirals.
Antiprotozoan/Antihelminthic: Various mechanisms (not detailed here).

Assessment of Antimicrobial Susceptibility

Disc Diffusion (Kirby-Bauer): Measures zone of inhibition around antibiotic discs.
MIC (Minimal Inhibitory Concentration): Lowest concentration preventing growth.
Broth Dilution: Serial dilutions to determine MIC.
Serum Killing Power: Tests patient blood for antimicrobial activity.
β-lactamase Testing: Detects resistance enzymes.

Antimicrobial Resistance

Mechanisms: Enzymatic inactivation (e.g., β-lactamases), decreased entry, altered targets, alternative pathways, drug efflux.
Types: Intrinsic, mutation-based, or acquired.
Notable Resistant Strains: MRSA, MDR tuberculosis, artemisinin-resistant malaria, cephalosporin-resistant gonorrhea.

Laboratory Skills (Summary)

Aseptic technique for inoculation.
Colony morphology and motility testing (e.g., SIM agar).
Staining methods: simple, negative, Gram, spore, capsule, acid-fast.
Microscopy, including oil immersion.

Additional info: Where details were brief, standard microbiology context was added for completeness (e.g., definitions, test principles, and examples).