Major Historical Developments in Microbiology

Germ Theory and Koch’s Postulates

The Germ Theory established that microorganisms are the cause of many diseases. Koch’s Postulates are a set of criteria used to prove the causative relationship between a microbe and a disease.

• Koch’s Postulates:

  1. The microorganism must be found in all organisms suffering from the disease, but not in healthy organisms.

  2. The microorganism must be isolated from a diseased organism and grown in pure culture.

  3. The cultured microorganism should cause disease when introduced into a healthy organism.

  4. The microorganism must be re-isolated from the experimentally infected host and identified as identical to the original.

• Spontaneous Generation: The disproven theory that life arises spontaneously from non-living matter. Pasteur’s experiment used swan-neck flasks to show that microbes come from the environment, not spontaneously.

• Cultivation Techniques: Methods such as isolation and enrichment cultures allow scientists to grow and study specific microbes.

• Characteristics of Life: Cellular organization, metabolism, growth, reproduction, response to stimuli, and evolution.

• The Three Domains of Life: Bacteria, Archaea, and Eukarya, based on molecular and cellular differences.

• Beneficial Microbes: Microbes play roles in nutrient cycling, food production, and health.

• Key Figures: Louis Pasteur (disproved spontaneous generation, vaccines), Robert Koch (germ theory, postulates), Sergei Winogradsky (soil microbiology), Martinus Beijerinck (enrichment culture, viruses).

Example: Pasteur’s swan-neck flask experiment demonstrated that sterilized broth remained free of microbes unless exposed to air, disproving spontaneous generation.

Cell Structure, Function and Behavior

Cytoplasmic Membranes and Cell Walls

The cytoplasmic membrane is a selectively permeable barrier, anchors proteins, and is involved in energy conservation.

• Membrane Functions:

  • Selective permeability barrier

  • Protein anchor

  • Energy conservation (generation of proton motive force)

• Differences Among Domains:

  • Bacteria: Phospholipid bilayer with ester linkages

  • Archaea: Unique lipids with ether linkages, sometimes monolayer

  • Eukarya: Phospholipid bilayer, sterols present

• Prokaryotic Cell Morphologies: Cocci (spherical), bacilli (rod-shaped), spirilla (spiral), etc.

• Osmotrophy: Small unicellular cells have high surface-to-volume ratio, facilitating nutrient uptake by active transport.

• Cell Wall: Provides shape and protection; Gram-positive (thick peptidoglycan), Gram-negative (thin peptidoglycan, outer membrane).

• Outer Membrane: Present in Gram-negative bacteria, contains lipopolysaccharides.

• DNA Arrangement: Bacteria and Archaea: single, circular chromosome; Eukarya: multiple, linear chromosomes.

• Flagella: Motility structures; Bacterial flagella rotate, Archaeal flagella are structurally different, Eukaryotic flagella use microtubules and whip-like motion.

• Chemotaxis: Movement toward or away from chemical stimuli.

• Cell Surface Structures: Pili (attachment, conjugation), capsules (protection, adherence).

• Cell Inclusions: Endospores (resistance), gas vesicles (buoyancy), storage polymers (energy reserves).

Example: Gram staining differentiates bacteria based on cell wall structure.

Microbial Metabolism

Elemental Composition and Nutrient Acquisition

Microbes require macronutrients (C, N, P, S, K, Mg, Ca, Na) and micronutrients (trace elements) for growth.

• Growth Factors: Organic compounds required by some organisms (e.g., vitamins, amino acids).

• Nutrient Acquisition: Osmotrophs use active transport to assimilate dissolved compounds.

• Permeases:

  • Simple transport (driven by proton motive force)

  • Group translocation (chemical modification during transport)

  • ABC systems (ATP-binding cassette, use ATP)

• Lab Media: Defined, complex, selective, differential media.

• Energy Classes: Phototrophs (light), chemotrophs (chemical), organotrophs (organic), lithotrophs (inorganic).

Bioenergetics and Metabolic Pathways

Microbial metabolism involves energy conservation and transfer through chemical reactions.

• Free Energy: indicates spontaneity; negative values mean reactions are energetically favorable.

• Catalysis: Enzymes lower activation energy, speeding up reactions; enzyme structure determines specificity.

• Redox Reactions: Transfer of electrons; important for energy generation.

• Redox Couples: Pairs of oxidized and reduced forms; electron tower ranks by reduction potential.

• Electron Carriers: NADH, FADH2, cytochromes.

• High Energy Compounds: ATP, phosphoenolpyruvate; energy storage products include glycogen, polyhydroxybutyrate.

• Energy Conservation: Substrate-level phosphorylation and oxidative phosphorylation.

• Respiratory Pathways:

  • Aerobic respiration (uses O2 as terminal electron acceptor)

  • Anaerobic respiration (uses other acceptors)

  • Fermentation (no external electron acceptor)

• Electron Transport Chain (ETC): Series of membrane-bound carriers; generates proton motive force (PMF).

• Proton Motive Force: Drives ATP synthesis, solute transport, and motility.

• Citric Acid Cycle: Central pathway for energy and precursor production.

• Catabolic Diversity: Prokaryotes have greater metabolic diversity than eukaryotes.

• Key Intermediates: 12 central metabolites produced by catabolic processes, used in biosynthesis.

Example: ATP synthesis via oxidative phosphorylation uses PMF generated by ETC.

Microbial Growth

Cellular and Population Growth

Microbial growth can refer to increase in cell size or population size. Population growth is often measured mathematically.

• Cell Division: Controlled by processes such as binary fission, budding, and division of stalked organisms.

• Population Growth Equations:

  • (discrete generations)

  • (continuous growth)

  • (growth rate constant)

• Growth Phases in Batch Culture:

  • Lag phase (adaptation)

  • Exponential phase (rapid growth)

  • Stationary phase (nutrient depletion)

  • Death phase (decline)

• Continuous Cultures (Chemostats): Maintain cells in exponential phase; control nutrient supply and waste removal.

• Measuring Growth: Direct counts, turbidity, viable plate counts.

• Environmental Factors: Temperature, O2, pH, water activity.

• Cardinal Temperatures: Minimum, optimum, and maximum temperatures for growth.

• Oxygen Requirements:

  • Aerobes (require O2)

  • Microaerophiles (low O2)

  • Aerotolerant (ignore O2)

  • Facultative aerobes (can use O2 or not)

  • Obligate anaerobes (cannot tolerate O2)

• Coping with Superoxides: Enzymes like superoxide dismutase, catalase, peroxidase detoxify reactive oxygen species.

• Molecular Adaptations: Proteins, membranes, and genomes adapt to extreme conditions.

• Sterilization and Disinfection: Methods include heat, filtration, chemicals.

• Disinfectant Types:

  • Bacteriostatic (inhibit growth)

  • Bacteriocidal (kill cells)

  • Bacteriolytic (lyse cells)

Example: Chemostats are used in industrial microbiology to maintain cultures in optimal growth phase.

Viruses and Virology

Virus Structure and Replication

Viruses are acellular entities that require host cells for replication.

• Types of Viruses: DNA, RNA, double-stranded (ds), single-stranded (ss), retroviruses.

• Structures: Simple (naked), complex, enveloped.

• Transmission and Infection: Viruses attach to host via specific receptor sites; host specificity depends on receptor compatibility.

• Replication Cycles:

  • Lytic (acute): Virus replicates and lyses host cell.

  • Lysogenic (temperate): Virus integrates into host genome, replicates with cell.

  • Other cycles: Chronic, latent, persistent infections.

• Impact on Microbial Food Webs: Viruses can lyse cells, releasing nutrients.

• Viral Titer: Concentration of virus particles; assayed by plaque assays.

• Key Terms:

  • Burst size: Number of virions released per cell

  • Assembly: Formation of new virions

  • Latent period: Time between infection and lysis

  • Capsomere: Protein subunit of capsid

  • Nucleocapsid: Capsid plus nucleic acid

  • Icosahedral symmetry: Common viral shape

• Viral Enzymes: Polymerases, proteases, reverse transcriptase (in retroviruses).

• Immunity: Populations develop resistance via CRISPR, restriction enzymes, or adaptive immunity.

• Bacterial vs Animal Viruses: Bacterial viruses (phages) often inject DNA; animal viruses enter by endocytosis or fusion.

• Retrovirus: RNA virus that uses reverse transcriptase to integrate into host genome.

• Viroids: Infectious RNA molecules without protein coat.

• Prions: Infectious proteins causing neurodegenerative diseases.

Example: HIV is a retrovirus that integrates its genome into host DNA.

Metabolic Regulation

Gene Expression and Regulation

Microbial cells regulate gene expression at multiple levels to adapt to environmental changes.

• DNA-Binding Proteins: Regulate transcription by interacting with promoters and operators.

• Promoters and Operators: Promoters are DNA sequences where RNA polymerase binds; operators are regulatory sites.

• Transcription: Carried out by RNA polymerase.

• Negative Control:

  • Repression: Inhibits transcription when product is present.

  • Induction: Activates transcription in response to substrate.

• Positive Control: Activator proteins enhance transcription.

• Operon: Cluster of genes under control of a single promoter.

• Regulon: Multiple operons regulated by the same control mechanism.

• Global Control Systems: Coordinate responses to environmental changes.

• Diauxic Growth: Sequential use of two carbon sources.

• Two-Component Regulatory Systems: Sensor kinase and response regulator; regulate processes like chemotaxis.

• Quorum Sensing: Cell-to-cell communication to coordinate group behaviors.

• Feedback Inhibition: End product inhibits enzyme activity.

• Post-Translational Modification: Chemical changes to proteins after synthesis.

Example: The lac operon is induced in the presence of lactose.

Genetics of Bacteria and Archaea

Mutations and Genetic Exchange

Genetic variation in microbes arises from mutations and horizontal gene transfer.

• Mutation: Change in DNA sequence; mutant is an organism with altered genotype.

• Reversion: Mutation that restores original phenotype; revertant is the organism.

• Selectable vs Non-Selectable Mutations: Selectable mutations confer growth advantage; non-selectable do not.

• Wild-Type Strain: Reference strain with typical genotype.

• Auxotroph: Mutant unable to synthesize a required compound.

• Replica Plating: Technique to screen for mutants.

• Types of Mutations:

  • Silent: No effect on protein

  • Nonsense: Creates stop codon

  • Missense: Changes amino acid

  • Frameshift: Alters reading frame

• Mutation Rates: Not constant; influenced by environmental factors.

• Mutagens: Agents that increase mutation rate; include chemicals, radiation.

• DNA Repair: Mechanisms to correct DNA damage.

• Recombination: Exchange of genetic material; observed experimentally by gene transfer.

• Horizontal Gene Transfer:

  • Transformation: Uptake of free DNA

  • Transduction: Transfer by bacteriophage

  • Conjugation: Direct cell-to-cell transfer

• Transposable Elements: DNA sequences that move within genome.

• CRISPR: Adaptive immune system in bacteria; stores viral DNA sequences for defense.

Example: Replica plating is used to identify auxotrophic mutants.