Microbiology Study Guide: Foundations, Cell Structure, Microscopy, Metabolism, and Growth

Chapter 1: A Brief History of Microbiology

Famous Scientists and Their Contributions

• Anton van Leeuwenhoek: Discovered microorganisms using handcrafted microscopes; considered the first to observe and describe microbes.

• Francesco Redi: Disproved spontaneous generation with his mason jar experiment, showing that maggots do not arise from meat without flies.

• Louis Pasteur: Father of microbiology; disproved spontaneous generation with swan-necked flask experiments, discovered fermentation, developed pasteurization, and attenuation (reducing virulence of pathogens).

• Robert Koch: Pioneered the study of disease causation (etiology); formulated Koch’s postulates for linking specific microbes to diseases.

• Edward Jenner: Developed the first successful smallpox vaccine using cowpox virus, founding immunology.

• Lady Montagu: Introduced variolation to Europe; Jenner’s method was safer.

• Joseph Lister: Developed aseptic techniques to reduce infection during surgery.

• Koch and Fanny Hesse: Introduced agar as a solid medium for bacterial culture.

• Florence Nightingale: Set hygiene standards in nursing; contributed to medical microbiology.

• Alexander Fleming: Discovered penicillin, the first antibiotic.

Koch’s Postulates

• 1. The suspected pathogen must be present in every case of the disease and absent from healthy hosts.

• 2. The pathogen must be isolated and grown in pure culture from a diseased host.

• 3. The cultured pathogen must cause the same disease when introduced into a healthy host.

• 4. The same pathogen must be re-isolated from the newly diseased host.

Purpose: To establish a causative relationship between a microbe and a disease.

Key Terms

• Pasteurization: Heating liquids to kill most contaminating bacteria without altering the liquid’s qualities.

• Attenuation: Reducing the virulence of a microorganism, often for vaccine development.

Types of Microorganisms

• Bacteria and Archaea: Prokaryotes lacking nuclei; bacteria have peptidoglycan cell walls, archaea have distinct cell wall components. Most reproduce asexually; some are beneficial, few are pathogenic.

• Viruses: Acellular, require host cells to replicate, consist of genetic material (DNA or RNA) within a protein coat (capsid).

• Fungi: Eukaryotic, with chitin cell walls. Yeasts are unicellular; molds are multicellular with hyphae forming mycelium. Non-photosynthetic.

• Protozoa: Unicellular eukaryotes, motile via cilia, flagella, or pseudopodia; diverse reproduction; some are pathogenic.

• Algae: Unicellular or multicellular, photosynthetic, classified by pigment and cell wall composition; generally nonpathogenic.

Chapter 2: The Chemistry of Microbiology

Basic Chemistry Concepts

• Atoms: Smallest units of matter, composed of protons, neutrons, and electrons.

• Isotopes: Atoms of the same element with different numbers of neutrons; radioactive isotopes can be used to kill microbes.

• Electron Configuration: Arrangement of electrons in shells; valence electrons determine chemical reactivity.

• Covalent Bonds: Electron sharing; can be nonpolar (equal sharing) or polar (unequal sharing).

• Ionic Bonds: Electron transfer creates charged ions (cations and anions) that attract each other.

• Hydrogen Bonds: Weak attractions between partial charges; important for stabilizing biological molecules.

• Synthesis (Anabolic) Reactions: Build larger molecules, require energy (often via dehydration synthesis).

• Decomposition (Catabolic) Reactions: Break down molecules, release energy (often via hydrolysis).

• Exchange Reactions: Involve both breaking and forming bonds; all reactions together constitute metabolism.

Organic Macromolecules

Macromolecule	Monomer/Type	Function/Notes
Lipids	Fatty acids, glycerol (not true polymers)	Energy storage, membrane structure, signaling (steroids)
Carbohydrates	Monosaccharides (e.g., glucose)	Energy storage, structural components, cell recognition
Proteins	Amino acids (20-21 types)	Structure, enzymes, regulation, transport, defense
Nucleic Acids	Nucleotides (phosphate, sugar, base)	Genetic information storage and expression

• Proteins: Built from amino acids via peptide bonds; function as enzymes, structural elements, transporters, and antibodies.

• Nucleic Acids: DNA and RNA; store and transmit genetic information; stabilized by hydrogen bonds between bases.

• Carbohydrates: Serve as energy sources (glucose), storage (starch, glycogen), and structural components (cellulose, peptidoglycan).

• Lipids: Hydrophobic molecules; include triglycerides, phospholipids (form membranes), and steroids (signaling).

Chapter 3: Cell Structure and Function

Prokaryotic Cell Structures

• Cell Wall: Provides shape and protection; made of peptidoglycan in bacteria. Gram-positive: thick peptidoglycan; Gram-negative: thin peptidoglycan plus outer membrane. Not all bacteria have cell walls.

• Cytoplasmic Membrane: Phospholipid bilayer with proteins; controls substance movement; present in all bacteria.

• Cytoplasm: Contains cytosol, DNA (nucleoid), ribosomes (70S), inclusions, and cytoskeleton.

• Ribosomes: 70S, site of protein synthesis; target for antibiotics.

• Glycocalyx: External layer; capsule (organized, resists phagocytosis) or slime layer (loose, aids attachment). Not all bacteria have these.

• Flagella: Motility structures; not present in all bacteria. Movement via rotation (runs and tumbles).

• Fimbriae: Short, numerous projections for adhesion and biofilm formation.

• Pili: Long tubes for DNA transfer (conjugation); usually one or two per cell.

• Endospores: Dormant, highly resistant structures (e.g., Bacillus, Clostridium); formed under harsh conditions.

Eukaryotic Cell Structures

• Nucleus: Membrane-bound, contains DNA; site of gene expression control.

• Cytoplasmic Membrane: Phospholipid bilayer with sterols; controls transport, allows endocytosis/exocytosis.

• Cell Wall: Present in fungi, plants, algae, some protozoa; made of cellulose, chitin, or other polysaccharides.

• Ribosomes: 80S, larger than prokaryotic; often attached to rough ER.

• Flagella and Cilia: Motility structures; flagella undulate, cilia are short and numerous.

• Cytoskeleton: Microtubules, microfilaments, intermediate filaments; shape, movement, organelle transport.

• Membranous Organelles: Mitochondria (ATP production), chloroplasts (photosynthesis), ER (synthesis), Golgi (processing), lysosomes, peroxisomes, vacuoles, vesicles.

Osmotic Effects on Cells

• Hypertonic Solution: Higher solute concentration outside; water leaves cell, causing shrinkage (crenation).

• Hypotonic Solution: Lower solute concentration outside; water enters cell, causing swelling and possible lysis.

Endospore Formation

• Produced by Bacillus and Clostridium under unfavorable conditions.

• Process: DNA is copied, surrounded by membrane, thick peptidoglycan, and spore coat.

• Function: Survival under extreme conditions; not for reproduction.

• Germination: Return to vegetative state when conditions improve.

Chapter 4: Microscopy, Staining, and Classification

Types of Light Microscopy

• Bright-field: Standard; for stained or pigmented specimens; shows cell shape, size, arrangement.

• Dark-field: Increases contrast for pale/colorless cells; specimen appears bright on dark background; good for motility observation.

• Phase-Contrast: Visualizes living, unstained cells; highlights internal structures by refractive index differences.

• Differential Interference Contrast (DIC): Uses polarized light and prisms for pseudo-3D images; enhances detail in transparent specimens.

Oil Immersion and Light Refraction

• Oil immersion increases resolution by reducing light refraction, allowing more light to enter the objective lens and improving image clarity.

Staining Techniques

Stain Type	Purpose	Examples
Simple	Single dye; reveals size, shape, arrangement	Methylene blue, crystal violet
Differential	Multiple dyes; distinguishes cell types/structures	Gram, Acid-fast, Endospore stains
Special	Highlights specific structures	Negative (capsule), Flagellar, Fluorescent stains
Electron Microscopy	Contrast for electron microscopy	Osmium tetroxide, heavy metals

• Gram Stain: Differentiates Gram-positive (purple) and Gram-negative (pink/red) bacteria based on cell wall structure.

• Acid-Fast Stain: Identifies bacteria with waxy cell walls (e.g., Mycobacterium).

• Endospore Stain: Uses heat and strong dyes to stain resistant endospores.

• Negative (Capsule) Stain: Stains background, leaving capsules as clear halos.

• Flagellar Stain: Visualizes flagella arrangement.

Gram Staining Steps and Cell Differences

1. Crystal Violet (Primary Stain): Stains all cells purple.

2. Gram’s Iodine (Mordant): Forms complex with crystal violet, fixing dye inside cells.

3. Alcohol/Acetone (Decolorizer): Dehydrates thick peptidoglycan in Gram-positive cells (retains dye); dissolves outer membrane in Gram-negative cells (dye washes out).

4. Safranin (Counterstain): Stains Gram-negative cells pink/red; Gram-positive remain purple.

• Gram-positive: Thick peptidoglycan, no outer membrane, purple after staining.

• Gram-negative: Thin peptidoglycan, outer membrane with LPS, pink/red after staining.

Binomial Nomenclature and Bacterial Shapes

• Binomial Nomenclature: Genus (capitalized) + species (lowercase), both italicized (e.g., Escherichia coli).

• Abbreviations: After first use, genus can be abbreviated (e.g., E. coli).

• Taxonomic Keys: Dichotomous keys use paired statements to identify organisms.

• Bacterial Shapes:

  • Cocci: Spherical; arrangements include diplococci, streptococci, staphylococci.

  • Bacilli: Rod-shaped; can form chains (streptobacilli).

  • Spiral: Includes spirilla (rigid), spirochetes (flexible), vibrio (comma-shaped).

Chapter 5: Microbial Metabolism

Metabolism, Catabolism, and Anabolism

• Metabolism: All chemical reactions in a cell.

• Catabolism: Breakdown of complex molecules; releases energy (exergonic).

• Anabolism: Synthesis of complex molecules; requires energy (endergonic).

Enzymatic Reactions

• Enzymes: Biological catalysts; speed up reactions without being consumed.

• Components: Apoenzyme (protein), cofactors (metal ions or coenzymes), active site (binds substrate).

Core Metabolic Pathways

• Glycolysis: Glucose → 2 pyruvate + ATP + NADH.

• Krebs Cycle: Acetyl-CoA → CO2 + NADH + FADH2 + ATP.

• Electron Transport Chain (ETC): NADH/FADH2 donate electrons; ATP generated via oxidative phosphorylation.

ATP Formation Mechanisms

• Substrate-level phosphorylation: Direct transfer of phosphate to ADP.

• Oxidative phosphorylation: Electron transport chain and chemiosmosis.

• Photophosphorylation: Light-driven ATP synthesis (photosynthetic organisms).

Respiration and Fermentation

• Aerobic Respiration: Oxygen is the final electron acceptor.

• Anaerobic Respiration: Other molecules (e.g., nitrate, sulfate) are final electron acceptors.

• Fermentation: Organic molecule from within the cell is the final electron acceptor; partial oxidation of sugar; regenerates NAD+.

Non-Carbohydrate Energy Sources

• Lipids: Broken down by lipases; glycerol enters glycolysis, fatty acids enter Krebs cycle as acetyl-CoA.

• Proteins: Proteases release amino acids, which are converted to metabolic intermediates.

Chapter 6: Microbial Nutrition and Growth

Microbial Growth Requirements

• Essential Nutrients: Water, carbon (sugars), nitrogen (proteins/amino acids), sulfur, phosphorus, oxygen.

Oxygen Requirements

Type	Oxygen Requirement	Examples
Obligate Aerobes	Require oxygen	Most human pathogens
Obligate Anaerobes	Cannot tolerate oxygen	Clostridium
Facultative Anaerobes	Grow with or without oxygen	E. coli
Microaerophiles	Require low oxygen	Microaerophilic bacteria

Temperature Requirements

• Mesophiles: 30–37°C; most human pathogens.

• Psychrophiles: <20°C; cold environments.

• Thermophiles: >45°C; hot environments.

• Hyperthermophiles: >80°C; extreme heat.

• Psychrotolerant: Grow at 0°C, optimal 20–40°C.

pH and Water Effects

• Neutrophiles: pH 6.6–7.5; most bacteria.

• Acidophiles: Grow best in acidic habitats.

• Alkalinophiles: Grow in alkaline environments (up to pH 11.5).

• Osmotic Pressure: High salt/sugar restricts growth; halophiles tolerate high salt.

• Hydrostatic Pressure: Barophiles require high pressure (deep-sea environments).

Culture Media Types

Type	Description	Examples
Defined	Exact chemical composition known	Minimal media
Complex	Contains extracts, not chemically defined	Blood agar, chocolate agar
Selective	Inhibits some microbes, allows others	MacConkey (Gram-negative), phenylethanol (Gram-positive)
Differential	Distinguishes microbes by property	Blood agar (hemolysins)
Transport	Maintains viability during transport	Transport media

Obtaining and Storing Pure Cultures

• Streak Plate: Isolates colonies for pure cultures.

• Pour Plate: Used for quantification.

• Membrane Filtration/Centrifugation: Concentrate bacteria from specimens.

• Storage: Refrigeration (short-term), freezing, lyophilization (long-term).

Counting Bacteria

• Counting Chamber: Direct microscopic count; includes dead cells.

• Plate Counts: Counts live colonies; uses serial dilution.

• Turbidity: Measures light scattering; estimates cell density.

Microbial Growth and Binary Fission

• Binary Fission: Asexual reproduction; cell divides into two identical daughter cells.

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

• Exponential Growth: Population doubles each generation; described by $2^n$ (n = number of generations).

Bacterial Growth Curve

1. Lag Phase: Adjustment, metabolic activity increases, no division.

2. Log (Exponential) Phase: Rapid cell division, population increases geometrically.

3. Stationary Phase: Nutrient depletion/waste accumulation slows growth; cell death equals cell division.

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

Types of Cultures

• Broth Culture: Closed system; fixed volume; goes through all growth phases.

• Chemostat: Open system; continuous culture; maintains population in a specific growth phase; used in industrial microbiology.

Hemolysins

• Enzymes produced by some bacteria that lyse red blood cells; detected on blood agar as clear zones (hemolysis).