Microbiology Module 1 & 2: Foundations, Cell Structure, Metabolism, Growth, and Control

Microbial World: Introduction and Diversity

Definition and Importance of Microorganisms

• Microorganisms are life forms too small to be seen by the naked eye.

• They are the oldest form of life and constitute a major fraction of Earth's biomass.

• Microbes inhabit plants, animals, and virtually every environment that supports life.

• They are diverse in form and function: single-celled, multicellular, and complex structures.

• Most microbes are not pathogenic; only a small fraction cause disease.

• Microbes are essential for nutrient cycling (carbon, nitrogen, phosphorus) and are used in biotechnology, genetic engineering, and bioremediation.

Studying Microbiology

• Microscopy is the primary tool for observing microbes.

• Culturing involves growing cells in/on nutrient media (liquid or solid) containing all required nutrients.

• Colony: a visible mass of cells derived from a single cell.

• Modern studies include molecular biology, biochemistry, genomics, and molecular genetics.

Evolutionary Timeline

• Earth: ~4.6 billion years old.

• First microorganisms: 3.8–4.3 billion years ago (anaerobic).

• Cyanobacteria (aerobic): ~2.6 billion years ago.

• Plants and animals: ~0.5 billion years ago.

Three Domains of Life

• Bacteria: Prokaryotes, usually undifferentiated single cells, 0.5–10 μm, 80+ phylogenetic lineages.

• Archaea: Prokaryotes, often in extreme environments, 12+ lineages, no known pathogens.

• Eukarya: Includes plants, animals, fungi; first unicellular, at least 6 kingdoms, diverse in size and physiology.

Viruses

• Obligate parasites; not cells; only replicate within host cells.

• No metabolism or independent replication.

• Classified by structure, genome, and host specificity.

Size and Surface Area-to-Volume Ratio

• As cell size increases, volume increases faster than surface area.

• Small cells have a higher surface area-to-volume (S/V) ratio, allowing faster nutrient/waste exchange.

• Order of size (smallest to largest): Viruses < Archaea < Bacteria < Eukaryotes.

Cell Shapes and Arrangements

• Cocci: Spherical

• Bacilli: Rod-shaped

• Coccobacillus: Short and plump

• Vibrio: Comma-shaped

• Spirillum: Rigid spiral

• Spirochete: Flexible spiral

• Arrangements: Staphylo- (clusters), Strepto- (chains), Diplo- (pairs)

Applications of Microbes

• Genetic engineering: e.g., biosynthetic insulin production.

• Bioremediation: Cleaning up oil spills and pollutants.

• Biotechnology: Food production (e.g., cheese, yogurt).

Historical Contributions

• Ignaz Semmelweis: Advocated handwashing to reduce puerperal fever.

• Joseph Lister: Introduced antiseptic surgery.

• Louis Pasteur: Disproved spontaneous generation, developed pasteurization, vaccines.

• Robert Koch: Established Koch's postulates, linked microbes to disease.

• Angelina Fanny Hesse: Introduced agar as a culture medium.

• Jane Hinton: Developed Mueller-Hinton agar for antibiotic testing.

• Albert Baez: Developed X-ray reflecting microscope.

• Sergei Winogradsky: Discovered chemolithotrophy, nitrogen fixation.

• Ester Lederberg: Discovered lambda phage, fertility factor F.

• Abigail Salyers: Pioneered microbiome research.

Microbial Cell Structure and Function

Prokaryotes vs. Eukaryotes vs. Archaea

• Bacteria: No nucleus, circular chromosome, no membrane-bound organelles, plasmids present.

• Archaea: No nucleus, circular chromosome, no membrane-bound organelles, plasmids, unique membrane lipids.

• Eukaryotes: Nucleus, linear chromosomes, membrane-bound organelles, sterols in membranes.

Cell Wall Composition

• Bacteria: Peptidoglycan (glycan chains cross-linked by peptides).

• Archaea: Pseudomurein or S-layer; isoprene monolayer.

• Fungi: Chitin.

• Cell walls maintain shape, rigidity, and prevent lysis.

Gram-Positive vs. Gram-Negative Bacteria

Feature	Gram-Positive	Gram-Negative
Peptidoglycan	Thick	Thin
Teichoic acids	Present	Absent
Outer membrane	Absent	Present (with LPS)
Lipid A/LPS	Absent	Present
Penicillin susceptibility	High	Lower

• Gram-positive bacteria are more susceptible to penicillin.

• Gram-negative bacteria produce endotoxin (Lipid A in LPS).

Glycocalyx

• Sticky polysaccharide coat outside the cell envelope.

• Functions: attachment, biofilm formation, protection from desiccation.

• Capsule: Tightly attached, organized.

• Slime layer: Loosely attached, easily deformed.

Pili, Fimbriae, and Hami

• Pili: Long protein appendages for attachment, genetic exchange (sex pili), or electron transfer.

• Fimbriae: Short, bristle-like; for adhesion to surfaces and biofilm formation.

• Hami: Archaeal grappling hook-like structures for attachment.

Flagella and Motility

• Flagella: Long, helical appendages for motility; powered by proton motive force (PMF).

• Types: Peritrichous (all over), Lophotrichous (tuft at one end), Monotrichous (single), Amphitrichous (one at each end).

• Run: Counterclockwise rotation (flagella bundle together).

• Tumble: Clockwise rotation (flagella separate).

• Pseudopods: Eukaryotic, arm-like projections for movement and feeding (e.g., amoebas).

Types of Taxis

• Chemotaxis: Response to chemicals.

• Phototaxis: Response to light.

• Osmotaxis: Response to ion concentration.

• Hydrotaxis: Response to water.

• Aerotaxis: Response to oxygen.

Endospores

• Dormant, highly resistant structures formed by some Gram-positive bacteria (e.g., Bacillus, Clostridium).

• Resistant to heat, radiation, chemicals, desiccation, and starvation.

• Formed under unfavorable conditions; germinate when conditions improve.

Viral Structure

• Genome (DNA or RNA), capsid (protein shell), sometimes envelope (phospholipid bilayer from host).

• Surface proteins for host attachment.

• Enzymes: lysozyme, neuraminidase, polymerases, reverse transcriptase (in retroviruses).

• Shapes: Helical, icosahedral, complex (e.g., bacteriophage).

Cell Membrane and S-Layer

• All cells have a phospholipid bilayer; eukaryotes have sterols, archaea have isoprene monolayers.

• S-layer: protein/glycoprotein layer, outermost in some bacteria/archaea, provides strength and protection.

Cell Inclusions

• Carbon storage: Poly-β-hydroxybutyrate (PHB), glycogen.

• Mineral storage: Polyphosphate, elemental sulfur, carbonate minerals.

• Gas vesicles: Buoyancy.

• Magnetosomes: Orientation in magnetic fields (magnetotaxis).

Microbial Metabolism

Types of Phosphorylation

• Substrate-level phosphorylation: Direct transfer of phosphate to ADP from a high-energy substrate.

• Oxidative phosphorylation: Electron transport chain (ETC) creates a proton motive force (PMF) to drive ATP synthesis.

• Photophosphorylation: Light energy generates PMF for ATP synthesis.

• Oxidative phosphorylation is the most efficient (30–32 ATP per glucose).

Metabolic Classes

Class	Energy Source	Carbon Source
Chemoorganotroph	Organic chemicals	Organic compounds (heterotroph)
Chemolithotroph	Inorganic chemicals	CO2 (autotroph)
Phototroph	Light	CO2 (photoautotroph) or organics (photoheterotroph)

Glycolysis

• Breakdown of glucose (6C) to two pyruvate (3C) molecules.

• Occurs in two stages: energy investment (uses 2 ATP) and energy payoff (produces 4 ATP).

• Net gain: 2 ATP (substrate-level phosphorylation), 2 NADH.

• Does not require oxygen.

Tricarboxylic Acid (TCA) Cycle

• Oxidizes pyruvate to CO2; generates NADH and FADH2 for ETC.

• Each turn produces 1 ATP (2 per glucose), 3 NADH, 1 FADH2.

• Most energy stored in electron carriers.

Electron Transport Chain (ETC) and ATP Synthesis

• Electrons from NADH/FADH2 passed through ETC, creating a proton gradient across the membrane.

• Protons flow back via ATP synthase, generating ATP (chemiosmosis).

• Oxygen is the final electron acceptor in aerobic respiration; alternatives (e.g., nitrate, sulfate) in anaerobic respiration.

• Key equations:

  • Generation time: $g = \frac{t}{n}$

  • Cell number: $N_t = N_0 2^n$

  • Growth rate: $k = \frac{0.693}{g}$

Fermentation

• ATP generated by substrate-level phosphorylation only.

• Regenerates NAD+ for glycolysis.

• End products: lactic acid, ethanol, acetic acid, CO2.

• Used in food/beverage production (e.g., cheese, yogurt, beer).

Calvin Cycle (Autotrophy)

• Light-independent reactions of photosynthesis.

• Uses ATP, NADPH, and CO2 to synthesize glucose.

• One glucose requires 6 CO2, 18 ATP, 12 NADPH.

Nitrogen Fixation

• Conversion of N2 gas to ammonia (NH3).

• Requires nitrogenase enzyme; inhibited by oxygen.

• Performed by some bacteria (e.g., Rhizobium).

Biosynthetic Pathways

• Gluconeogenesis: Synthesis of glucose from non-carbohydrate precursors.

• Pentose phosphate pathway: Generates pentose sugars and NADPH.

• Amino acid, nucleotide, fatty acid, and lipid biosynthesis: Use intermediates from glycolysis and TCA cycle.

Microbial Growth and Its Control

Binary Fission and Growth Curves

• Binary fission: Cell enlarges, replicates DNA, forms septum, and splits into two daughter cells.

• Growth curve phases: Lag, exponential (log), stationary, death/decline.

• Exponential phase is ideal for antibiotic targeting.

Culture Media

• Defined media: Exact chemical composition known.

• Complex media: Contains digests of animal/plant products.

• Selective media: Inhibits some microbes, allows others.

• Differential media: Distinguishes microbes by metabolic reactions.

Continuous Culture and Chemostats

• Chemostat: Device for continuous culture; maintains steady-state growth by adding fresh medium and removing spent culture.

• Growth rate controlled by limiting nutrient; population density by dilution rate ($D = \frac{F}{V}$).

Environmental Adaptations

Type	Optimal Condition	Example Environment
Psychrophile	≤15°C	Polar ice, deep ocean
Mesophile	~39°C	Soil, human body
Thermophile	45–80°C	Hot springs
Hyperthermophile	>80°C	Hydrothermal vents
Acidophile	pH ≤5.5	Acidic hot springs
Neutrophile	pH 5.5–8	Human body
Alkaliphile	pH ≥8	Soda lakes
Halophile	3–6% NaCl	Salt lakes
Halotolerant	Low–moderate salt	Human skin
Osmophile	High sugar	Honey, syrups
Xerophile	Dry	Deserts

Oxygen Requirements

• Aerobes: Require oxygen.

• Anaerobes: Oxygen is toxic.

• Facultative anaerobes: Can grow with or without oxygen.

• Microaerophiles: Require low oxygen.

• Aerotolerant: Indifferent to oxygen.

• Capnophiles: Require CO2.

Biofilms

• Communities of microbes attached to surfaces, embedded in a polysaccharide matrix.

• Stages: Attachment, colonization, development, dispersal.

• Biofilms are highly resistant to antibiotics and disinfectants.

• Involved in medical device infections, dental plaque, and industrial fouling.

Antibiotic Resistance Mechanisms

• Altered targets (e.g., MecA in MRSA).

• Efflux pumps (e.g., AcrAB-TolC in E. coli).

• Enzymatic degradation (e.g., beta-lactamases).

Peptidoglycan Synthesis and Antibiotic Targets

• Peptidoglycan synthesis involves transpeptidase and transglycosylase enzymes.

• Antibiotics target cell wall synthesis (e.g., beta-lactams, vancomycin, bacitracin).

Control of Microbial Growth

• Decontamination: Making objects safe to handle.

• Sanitizer: Reduces microbial numbers.

• Disinfection: Kills/inhibits pathogens on inanimate objects.

• Sterilization: Kills all microbes, including endospores.

• Antiseptic: Reduces microbes on living tissue.

• Heat (autoclave) is the most widely used sterilization method.

• Radiation (UV, gamma, X-rays) and filtration are used for heat-sensitive materials.

• Decimal reduction time (D): Time to reduce viability by 90% at a given temperature.

• Minimum inhibitory concentration (MIC): Lowest concentration of an agent that inhibits growth.

Types of Antimicrobial Agents

• -cidal: Kills microbes.

• -static: Inhibits growth.

• -lytic: Kills by lysis.

Viruses and Their Multiplication

Lytic Cycle (Virulent Infection)

• Virus replicates and destroys host cell.

• Steps: Attachment, penetration, synthesis, assembly, release.

• Host cell lysis releases new virions (burst size).

• One-step growth curve: latent period (eclipse + maturation), then release.

• Early proteins: DNA replication and gene expression.

• Middle/late proteins: Structural components and enzymes for assembly/release.

Lysogenic Cycle (Temperate Infection)

• Viral genome integrates into host chromosome (prophage).

• Host cell survives and divides, passing on viral DNA.

• Induction (triggered by stress) excises viral DNA, entering lytic cycle.

• Lysogeny maintained by repressor proteins.

Viral Replication in Eukaryotes

• Entire virion enters animal cell (via fusion or endocytosis).

• Replication often occurs in the nucleus; formation of viroplasms.

• Plant viruses: Non-enveloped, transmitted via wounds or vectors.

• Modes of infection: Virulent, latent, persistent, transformation (cancer).

Host Range and Cytopathic Effects

• Host range: Spectrum of cells a virus can infect (e.g., Hepatitis B infects human liver cells).

• Cytopathic effects: Visible changes in host cells (size, shape, lysis, inclusion bodies, multinucleation, transformation).

Applied Microbiology: Sustainable Agriculture

Microbial Applications in Agriculture

• Use of nitrogen-fixing bacteria (e.g., Rhizobium, Azotobacter, Clostridium, Frankia) to convert atmospheric nitrogen into plant-usable forms.

• Cyanobacteria (e.g., Nostoc, Anabaena) colonize root cortex, aiding nutrient uptake.

• Defense bacteria (e.g., Clavibacter, Agrobacterium, Bacillus, Pseudomonas) synthesize antibiotics or trigger plant defenses.

• Fungi (e.g., Glomeromycota, Ascomycota, Basidiomycota) assist in nutrient transfer and defense.

Manipulation and Research

• Genetic and molecular techniques (e.g., 16S rDNA analysis) to study and manipulate plant-microbe interactions.

• Applications include biofertilizers, crop improvement, and sustainable agriculture.

• Challenges: Loss of symbiotic activity in domesticated crops, potential for opportunistic pathogens.

Additional info: This section integrates applied microbiology with ecological and evolutionary concepts relevant to microbial symbioses and biotechnology.