Microbial Metabolism

Metabolism: Catabolic vs. Anabolic Processes

Metabolism encompasses all chemical reactions occurring within a living cell, essential for maintaining life. These reactions are categorized as catabolic or anabolic:

• Catabolic Reactions: Break down complex molecules into simpler ones, releasing energy (exergonic). Example: Breakdown of glucose into CO2 and H2O. Often involve hydrolytic reactions.

• Anabolic Reactions: Build complex molecules from simpler ones, requiring energy (endergonic). Example: Synthesis of proteins from amino acids. Often involve dehydration synthesis reactions.

• Energy Connection: Energy released from catabolic reactions is used to drive anabolic reactions, mediated by ATP.

ATP: Structure and Role in Metabolism

Adenosine triphosphate (ATP) is the cell's energy currency, composed of adenine, ribose, and three phosphate groups. Hydrolysis of ATP to ADP and inorganic phosphate releases energy for cellular activities.

• Catabolism: Generates ATP by converting ADP and inorganic phosphate using energy released from breakdown of molecules.

• Anabolism: Utilizes ATP to drive biosynthetic reactions.

• ATP Coupling: Ensures balanced energy flow between catabolic and anabolic pathways.

Enzymes: Function and Classification

Enzymes are globular proteins that catalyze biochemical reactions by lowering activation energy, enabling reactions at physiological temperatures. Their specificity is determined by their active sites.

• Naming: Enzymes typically end with "-ase" and are classified by the reaction they catalyze:

Factors Influencing Enzyme Activity

• Substrate Concentration: Reaction rate increases with substrate concentration until saturation is reached.

• Temperature: Activity increases with temperature up to an optimum (35–40°C), then declines due to denaturation.

• pH: Each enzyme has an optimal pH; deviations reduce activity and may cause denaturation.

Graphical Representation: Enzyme activity vs. temperature and pH shows a peak at optimal values; activity vs. substrate concentration plateaus at saturation.

Oxidation and Reduction in Glucose Metabolism

• Oxidation: Loss of electrons, often addition of oxygen or removal of hydrogen.

• Reduction: Gain of electrons, often addition of hydrogen or removal of oxygen.

• Glucose Metabolism: Glucose is oxidized to CO2 and H2O, releasing energy captured as ATP. Electrons are transferred to oxygen, which is reduced to water.

ATP Production: Substrate-Level vs. Oxidative Phosphorylation

• Substrate-Level Phosphorylation: Direct transfer of phosphate from substrate to ADP. Occurs in glycolysis and Krebs cycle.

• Oxidative Phosphorylation: Electrons from NADH/FADH2 pass through electron transport chain, creating a proton gradient used by ATP synthase to generate ATP (chemiosmosis).

Electron Transport Chain and Chemiosmosis

• NADH: Donates electrons to ETC.

• H+: Pumped across membrane, creating gradient.

• Oxygen: Final electron acceptor, forms water.

• ATP Synthase: Uses proton gradient to convert ADP + Pi to ATP.

Equation:

Respiration vs. Fermentation

• Aerobic and Anaerobic Respiration: Both use glycolysis, Krebs cycle, and ETC.

• Fermentation: Only glycolysis, followed by conversion of pyruvate to lactic acid or ethanol.

Microbial Identification Tests

• Amino Acid Catabolism: Detects enzymes for amino acid breakdown; pH indicator changes color based on products.

• Fermentation Test: Measures acid and gas production from carbohydrate metabolism.

• Cytochrome c Oxidase Test: Identifies bacteria with cytochrome c oxidase; distinguishes species.

• Urease Test: Detects urease activity; pH indicator changes with ammonia production.

Metabolic Categories

Humans: Chemoheterotrophs; obtain energy and carbon from organic compounds.

Microbial Growth

Optimal pH for Bacteria and Fungi

• Bacteria: Optimal pH 6.5–7.5 (near neutral).

• Fungi: Optimal pH 5–6 (acidic).

Halophiles: Types and Adaptations

• Obligate Halophiles: Require high salt (up to 30%) for growth.

• Facultative Halophiles: Tolerate high salt but do not require it; grow up to 15% salt.

Essential Elements for Cellular Growth

• Nitrogen: Protein and nucleic acid synthesis.

• Phosphorus: Nucleic acids, phospholipids, ATP.

• Sulfur: Sulfur-containing amino acids, vitamins.

• Magnesium: Enzyme cofactor, nucleic acid stabilization.

• Potassium: Enzyme activation, osmotic balance.

Oxygen Requirements: Growth Patterns

Biofilms: Characteristics and Clinical Impact

• Resistance: Highly resistant to antibiotics and disinfectants.

• Complexity: Multi-species, coordinated community.

• Communication: Quorum sensing regulates activities.

• Clinical Problems: Cause persistent infections on medical devices; difficult to treat.

Types of Culture Media

• Chemically Defined: Exact composition known; used for specific bacteria.

• Complex: Contains extracts; supports heterotrophs.

• Selective: Suppresses unwanted microbes, encourages desired ones.

• Differential: Distinguishes colonies based on reactions (e.g., color change).

• Reducing: Removes oxygen for anaerobes.

Capnophiles: Growth Requirements

• Capnophiles: Require elevated CO2 (above 3%). Found in intestinal and respiratory tracts.

• Aerobes: Require O2; do not thrive in high CO2.

• Facultative Anaerobes: Grow with or without O2; do not require high CO2.

Bacterial Growth Curve

• Lag Phase: Metabolic activity, no increase in cell number.

• Log Phase: Rapid, exponential cell division.

• Stationary Phase: Cell division equals cell death; population stabilizes.

• Death Phase: Cell death exceeds division; population declines.

Binary Fission and Generation Time

• Binary Fission: One cell divides into two; exponential growth.

• Generation Time: Time for population to double; varies by species and conditions.

Equation:

Where is final cell number, is initial cell number, is number of generations.

Cell Counting Methods

Control of Microbial Growth

Definitions: Microbial Control Agents

• Bacteriostatic: Inhibits bacterial growth; reversible.

• Bacteriocidal: Kills bacteria.

• Germicide: Kills microorganisms (not endospores).

• Virucide: Inactivates viruses.

• Sterilant: Destroys all forms of microbial life, including endospores.

• Fungicide: Kills fungi.

Thermal Death Time (TDT)

• Definition: Minimum time to kill all bacteria at a given temperature.

• Graph: Logarithmic plot of surviving cells vs. time; TDT is where line meets x-axis.

Wet Heat vs. Dry Heat Sterilization

Bacterial Cell Death Graphs

• Bactericidal: Downward slope (log scale).

• Bacteriostatic: Horizontal line; cell number unchanged.

Food Preservation Mechanisms

• Heat Treatment: Denatures proteins; moist and dry heat.

• Cold Preservation: Slows growth; bacteriostatic.

• High Pressure: Alters molecular structures.

• Desiccation/Osmotic Pressure: Removes water; causes plasmolysis.

• Radiation: Destroys DNA.

• Aseptic Packaging: Maintains sterility.

• Industrial Canning: Steam under pressure; targets spores.

Physical and Chemical Sterilants

• Physical: Heat, filtration, cold, radiation.

• Chemical: Phenolics, halogens, alcohols, aldehydes, gaseous sterilants.

Disk Diffusion Test Interpretation

• Zones of Inhibition: Clear areas around antibiotic disks indicate effectiveness.

• Larger Zones: Higher sensitivity.

• Smaller Zones: Resistance.

• Compare to standards: Classify as sensitive, intermediate, or resistant.

Applications of Chemical Agents

Use Dilution Test Interpretation

• Bacteriostatic: Inhibits growth; bacteria resume growth after removal.

• Bactericidal: Kills bacteria; no growth after removal.

• Effective Dilution: Lowest concentration with no growth is bactericidal.

Chemical Sterilants: Comparison

• Aldehydes: Cross-link proteins; formaldehyde (carcinogenic), glutaraldehyde (hospital instruments).

• Gaseous Chemosterilants: Ethylene oxide; heat-sensitive materials.

• OPA: More effective, less irritating than glutaraldehyde.

Hospital-Acquired Infections (HAIs)

• Microorganisms: Hospitals harbor pathogens, including resistant strains.

• Compromised Host: Patients with weakened immunity are susceptible.

• Chain of Transmission: Spread via direct/indirect contact, medical devices, airborne routes.

Example: Catheter-associated infections, surgical site infections.