Microorganisms
Eukaryotic Microorganisms & Opportunistic Pathogens
Eukaryotes: Organisms containing complex cellular structures with membrane-bound organelles (all organisms except bacteria).
Opportunistic Pathogens: Microorganisms that typically do not cause disease in healthy individuals but produce opportunistic infections in immunocompromised host systems with low resistance.
Treatment Challenges: Because eukaryotic micro-organisms share structural and functional similarities with human host cells, developing selective antimicrobials without severe host side effects is difficult.
Fungi Characteristics & Structure
General Properties: Eukaryotic, heterotrophic organisms with cell walls composed primarily of chitin rather than cellulose, making them more closely related to animals than plants.
Structural Components:
Hyphae: Threadlike filaments that secrete digestive enzymes to break down food externally, absorbing the released nutrients.
Mycelium: Interwoven network of repeated hyphal branches that forms the main vegetative feeding body of a fungus.
Mushrooms: Reproductive structures that produce and release airborne spores for sexual or asexual propagation.
Types of Fungi & Commercial Roles
Yeasts: Unicellular fungi that reproduce asexually through budding or spore formation. Examples include baking/brewing yeasts, fruit surface blooms, and Candida albicans.
Moulds: Multicellular filamentous fungi that reproduce via spores produced within sporangia or conidia.
Ecological & Commercial Value:
Primary decomposers (saprophytes) that recycle nutrients essential for plant growth.
Used commercially in food and beverage production (bread, beer, wine, cheese, edible mushrooms/truffles).
Source of pharmaceutical compounds, such as penicillin from Penicillium species.
Pathogenic Fungi & Health Issues
Dermatophytes (Microsporum, Trichophyton, Epidermophyton): Cause cutaneous infections including tinea pedis (athlete's foot) and tinea corporis (ringworm).
Candida albicans: Causes candidiasis (thrush) on mucous membranes and potential systemic disease in vulnerable hosts.
Pneumocystis jiroveci & Cryptococcus neoformans: Cause life-threatening pneumonia in immunocompromised individuals (e.g., AIDS patients).
Malassezia furfur: Causes superficial mycosis known as pityriasis versicolor (tinea versicolor).
Complications: Fungal spores cause airborne allergies, respiratory irritation, and systemic damage via mycotoxins such as aflatoxin.
Protozoa
Characteristics: Unicellular eukaryotic organisms enclosed by a cell membrane but lacking a rigid cell wall.
Classification by Motility Method:
Flagellates: Move using flagella (e.g., Giardia).
Ciliates: Move using cilia (e.g., Balantidium).
Sarcodina: Move via pseudopodia (e.g., Amoeba).
Sporozoa: Non-motile organisms (e.g., Plasmodium).
Clinical Pathogens:
Entamoeba histolytica: Ingested via contaminated food/water; causes amoebic dysentery.
Giardia lamblia: Causes gastrointestinal illness characterized by diarrhoea, pain, and malaise.
Plasmodium species: Transmitted by mosquito vectors; causes malaria.
Multicellular Parasites (Helminths)
Characteristics: Parasitic worms predominant in areas with poor sanitation and low socioeconomic status; exhibit complex life cycles often involving multiple host species.
Pathogenesis: Typically do not replicate inside the primary host, but shed produced eggs through faeces to maintain their transmission cycle.
Major Groups: Tapeworms, Roundworms, Flukes.
Ectoparasites & Vector Transmitted Diseases
Characteristics: External arthropod parasites residing on or within human skin surfaces.
Primary Examples:
Mites (Sarcoptes scabei): Burrow into skin causing scabies.
Lice (Pediculus humanus): Cause scalp irritation and redness.
Fleas & Ticks: Blood-feeding external vectors.
Vector Roles: Ectoparasites harbor and transfer secondary microorganisms (e.g., rickettsias) via bites or skin contamination, transmitting serious systemic diseases such as typhus, spotted fever, and bubonic plague.
Historical Milestones in Microbiology
Microscope Invention & Microscopy:
Robert Hooke (1665): Built the first compound microscope, initiating the structural observation of microscopic entities.
Anton van Leeuwenhoek (1673): Used simple hand lenses to observe live, motile microorganisms, which he termed "animalcules".
Electron Microscopy: Modern visualization of viruses requiring magnifications of approximately relies on electron microscopes.
Vaccination & Smallpox:
Edward Jenner (1796): Developed the smallpox vaccine by inoculating individuals with material taken from cowpox lesions on milkmaids' hands. Building on earlier observations by John Fewster and Benjamin Jesty, Jenner demonstrated that cowpox exposure conferred immunity against smallpox.
Infection Control & Hand Hygiene:
Surgical Infection Context: By the mid-19th century, post-operative sepsis accounted for the death of nearly half of all patients undergoing major surgery. Early surgeons operated in unwashed street clothes or blood-stained operating gowns, taking pride in accrued blood and pus as a mark of respect.
Ignaz Semmelweis (1840): Identified that medical student and physician hand contamination directly caused puerperal fever (a uterine Streptococcus infection striking post-partum women in hospital maternity wards). Demonstrating that antiseptic handwashing dramatically reduced maternal mortality, he earned the title "saviour of mothers".
Joseph Lister: Pioneered antiseptic surgery by advocating the application of carbolic acid (phenol) to sterilize instruments, surgical dressings, and wounds.
Biogenesis vs. Spontaneous Generation:
Spontaneous Generation (Generatio spontanea): Ancient belief (attributed to Greek mythology and Gaia) that living organisms spontaneously arose from non-living matter.
Francesco Redi: Proved that maggots developed from fly eggs laid on meat rather than spontaneously generating from decaying meat, earning recognition as the "father of parasitology".
Louis Pasteur (1861): Disproved spontaneous generation by demonstrating biogenesis (Omne vivum ex ovo - "all life comes from an egg"), proving that microorganisms originate only from pre-existing microbial life.
Germ Theory of Disease:
Robert Koch (1876): Established the Germ Theory of Disease by proving Bacillus anthracis caused anthrax. He formulated Koch's Postulates to definitively link a specific microbe to a disease:
The specific microorganism must be present in every case of the disease examined.
The microorganism must be isolated from the diseased host and maintained in a pure culture.
The cultured microorganism must reproduce the original infection when inoculated into a healthy, susceptible host.
The same microorganism must be re-isolated from the experimentally inoculated host and cultured again.
Antibiotic Discovery & Development:
Paul Ehrlich: Developed the first synthetic chemotherapeutic agent (salvarsan) capable of treating syphilis.
Alexander Fleming (1928): Discovered penicillin after noticing a contaminating mold colony (Penicillium notatum) on an unwashed agar plate had produced a clear zone of inhibition where surrounding bacterial growth was killed.
Beneficial Roles & Uses of Microorganisms
Ecological Roles:
Organic matter decomposition (saprophytic recycling).
Biogeochemical nutrient cycling, including atmospheric nitrogen fixation.
Primary oxygen production via algal photosynthesis.
Serving as the foundational base of aquatic and marine food chains.
Commercial & Food Production Uses:
Fermentation processes for bread, cheese, yoghurt, beer, wine, and vinegar.
Industrial synthesis of commercial chemicals, including citric acid, ascorbic acid (Vitamin C), and monosodium glutamate (MSG).
Production of therapeutic antibiotics and pharmaceuticals.
Therapeutic Applications:
Fecal Microbiota Transplant (FMT): Instillation of fecal material from a healthy donor into an unhealthy gastrointestinal tract to restore gut flora equilibrium and resolve recurrent Clostridium difficile infections.
Normal Flora, Transient Flora, & Sterile Body Sites
Normal Flora (Resident Microbiota):
Characteristics: Microorganisms permanently inhabiting specific body surfaces under varying moisture, temperature, pH, oxygen, and nutrient conditions.
Functions: Inhibit colonization by potential pathogens via nutrient competition and surface occupation (colonization resistance); synthesize useful vitamins and produce antibacterial chemicals.
Stability: Relatively stable, but subject to alteration by environment, diet, health status, or antibiotic therapy.
- Anatomical Distribution of Normal Flora:
- **Skin:** *Staphylococcus epidermidis*, *Staphylococcus aureus*, *Propionibacterium acnes*, *Corynebacterium* spp., *Micrococcus* spp., *Candida* spp.
- **Mouth:** *Streptococcus* spp., *Lactobacillus* spp., *Fusobacterium* spp., *Actinomyces* spp., *Bacteroides* spp., *Veillonella* spp.
- **Upper Respiratory Tract (Nasal Cavity / Pharynx):** *Staphylococcus* spp., *Streptococcus pneumoniae*, *Corynebacterium* spp., *Haemophilus influenzae*, *Branhamella catarrhalis*.
- **Gastrointestinal Tract:**
- **Stomach:** Highly acidic; harbors very few microbes (*Lactobacillus* spp.).
- **Intestines / Colon:** Contains over 300 distinct species, including *Bacteroides* spp., *Enterococcus* spp., *Enterobacter* spp., *Escherichia coli*, *Proteus* spp., *Klebsiella* spp., *Bifidobacterium* spp., *Eubacterium* spp., *Clostridium perfringens*, *Fusobacterium* spp., and *Candida* spp.
- **Genitourinary Tract & Vagina:** *Lactobacillus* spp., *Corynebacterium* spp., *Enterococcus* spp., *Candida albicans*, and various strict anaerobes.
Transient Flora:
Microorganisms that temporarily colonize anatomical sites for brief periods.
Cleared rapidly by mechanical washing from body secretions, immune defense clearance, or competitive exclusion by established normal flora.
Sterile Body Areas:
Anatomical zones naturally completely devoid of microorganisms:
Foetus in utero.
Whole blood and circulatory system.
Serous membrane cavities (pericardial, pleural, and peritoneal cavities).
Internal solid organs (e.g., kidneys, liver, pancreas, deep lung parenchyma, lower trachea, and bronchi).
Terminology & Classifications of Infection & Disease
Core Definitions:
Infection: Multiplication and colonization of microorganisms within or on a body site.
Disease: Any alteration to the normal physiological or metabolic state of the host body.
Infectious Disease: A disease caused directly by an infectious microorganism or its toxic products.
Pathogen: Any organism capable of producing disease.
Infection Source Classifications:
Endogenous Infection: Originates from inside the host body (e.g., opportunistic overgrowth or translocation of resident normal flora).
Exogenous Infection: Originates from pathogens acquired from the external environment.
Healthcare-Associated Infection (HCAI / Nosocomial): Exogenous infection acquired during a patient's stay in a hospital or healthcare facility.
Extent & Anatomical Distribution Classifications:
Localised Infection: Microorganisms remain restricted to the initial site of entry.
Disseminated Infection: Pathogens spread beyond the primary entry site to involve adjacent tissues or secondary organs.
Systemic Disease: Pathogens spread through the circulatory system, affecting multiple organ systems simultaneously.
Mixed Infection: Infection involving two or more distinct species acting synergistically to create favorable conditions for one another.
Primary vs. Secondary Infection: Primary infection is an initial acute illness caused by a primary pathogen; secondary infection is a subsequent opportunistic infection arising when host defenses are weakened by the primary disease.
Clinical Classifications:
Superinfection: Secondary infection occurring after normal protective microbial flora is destroyed by broad-spectrum antibiotic therapy.
Subclinical (Inapparent) Infection: Infection where the host displays no overt clinical symptoms, despite inducing a detectable antibody immune response.
Persistent Infections: Long-term infections persisting after initial disease resolution (categorized as slow, latent, chronic, or oncogenic infections).
Opportunistic Infections:
Caused by non-pathogenic microbes or normal flora under specific permissive conditions:
Host immune defenses are compromised (e.g., chemotherapy, HIV, or immunosuppressive therapy).
Microorganisms gain access to abnormal anatomical locations (e.g., surgical wound contamination or intravenous catheter sites).
Alteration of physiological homeostatic conditions (e.g., broad-spectrum antibiotic treatment altering local pH and microbiota balance).
Pathogenicity, Virulence, & Stages of Disease Progression
Determinants of Microbial Pathogenesis:
Pathogenicity: The qualitative capacity of an organism to cause disease by gaining host entry, attaching to host tissues, multiplying, evading host defenses, and causing structural or functional tissue damage.
Virulence: The quantitative degree or intensity of disease caused by a pathogen. Dependent on specific virulence factors including capsules, enzymes (e.g., hyaluronidase, hemolysins), and exotoxins/endotoxins.
Infective Dose: The threshold quantity of pathogen cells or particles required to successfully initiate disease in a host.
Host Susceptibility Factors:
Specific and non-specific physical defenses (intact skin barriers, mucous membranes, innate and adaptive immunity).
Predisposing environmental factors (climate, seasonal variations, air pollution, housing, personal hygiene).
Individual host status (age, nutritional status, genetic constitution, underlying health conditions, lifestyle, mental stress).
Clinical Manifestations:
Signs: Measurable, objective physical changes observed by a clinician (e.g., elevated body temperature, rash, laboratory blood test results).
Symptoms: Subjective manifestations experienced and reported by the patient (e.g., pain, malaise, nausea, headache).
Fever: Induced by endogenous pyrogens (released by host white blood cells) or exogenous pyrogens (e.g., bacterial endotoxins).
Inflammation: Characterized by the classical cardinal signs: heat (calor), pain (dolor), redness (rubor), and swelling (tumor).
- Stages in the Course of Infectious Disease:
1. **Incubation Period:** Time interval between initial pathogen entry and the first appearance of signs or symptoms. Pathogen overcomes initial host barriers and multiplies.
2. **Prodromal Period:** Short phase characterized by early, mild, non-specific symptoms (malaise, headache, fatigue).
3. **Acute (Invasive) Phase:** Disease is at its peak severity; patient exhibits characteristic signs and symptoms. Failure of host defenses or medical intervention during this phase can lead to a fatal outcome.
4. **Decline Phase:** Pathogen numbers decrease and clinical symptoms subside due to successful host immune response or antimicrobial treatment.
5. **Convalescence:** Recovery phase during which tissue repair occurs and bodily functions return to normal.
Epidemiology & Chain of Infection
Epidemiological Categories of Disease:
Communicable Disease: Transmissible directly or indirectly from one host to another. (All communicable diseases are infectious).
Contagious Disease: A communicable disease that is exceptionally easily and rapidly transmitted from person to person (e.g., Hepatitis B, measles).
Non-Communicable Disease: Disease not transmitted host-to-host. Acquired from environmental sources, normal flora translocation, or insect vectors (e.g., tetanus, type 2 diabetes, cardiovascular disease).
Epidemiological Terminology:
Morbidity: Incidence of illness or sickness in a population.
Mortality: Death rate resulting from a disease in a population.
Incidence: Number of new disease cases emerging in a defined population over a specified time period.
Prevalence: Total number of active (old and new) disease cases present in a population at a specific point in time.
Sporadic: Disease occurring occasionally and irregularly in isolated cases.
Endemic: Disease constantly present at a baseline level in a specific geographic area.
Epidemic: Sudden, significant increase in disease cases above expected endemic levels in a population.
Pandemic: Worldwide epidemic affecting multiple countries or continents.
Infectious Diseases in New Zealand:
High incidence of post-streptococcal rheumatic fever (disproportionately affecting Pacific Island populations).
Elevated incidence of Tuberculosis (TB), particularly among Māori, Pacific Island, and non-European ethnic groups.
Recurrent epidemics of meningococcal meningitis (predominantly in Māori and Pacific Island communities).
High rates of viral hepatitis (Hepatitis A in Pacific Islanders; Hepatitis B in Māori).
High rates of foodborne campylobacteriosis among European New Zealanders.
High prevalence of otitis media leading to permanent hearing loss in of school children.
Approximately 50 legally notifiable diseases required by law to be reported upon diagnosis (e.g., campylobacteriosis, gastroenteritis from a common source, giardiasis, salmonellosis, rheumatic fever, TB, AIDS, hepatitis, meningococcal disease, pertussis, mumps, measles, typhoid, malaria).
- The Chain of Infection:
- **Causative Agent:** Bacteria, virus, fungi, parasite.
- **Reservoir:**
- *Human:* Symptomatic patients, asymptomatic carriers, latent viral carriers, resident flora.
- *Animal:* Hosts harboring zoonoses (often requiring biological vectors for human transfer).
- *Non-living:* Soil, water, fomites (contaminated equipment, surgical instruments, linen, dressings).
- **Portal of Exit:** Respiratory tract secretions, gastrointestinal faeces, skin lesions/dust, urogenital secretions, blood.
- **Mode of Transmission:** Direct contact, indirect contact, common vehicle, or vector.
- **Portal of Entry:** Cuts/breaks in skin, respiratory inhalation, gastrointestinal ingestion, urogenital mucous membranes.
- **Susceptible Host:** Elderly, infants, immunocompromised, or any unvaccinated/vulnerable individual.
Portals of Entry, Exit, & Modes of Transmission
Portals of Entry Details:
Skin: Cutaneous damage (cracks, abrasions, animal/insect bites, surgical incisions, intravenous line insertions).
Respiratory Tract: Inhalation of airborne particles, infectious droplets, aerosols, fungal spores, or via contaminated medical equipment (ventilators, nebulisers).
Gastrointestinal Tract: Ingestion of fecally contaminated food or fluid.
Urogenital Tract: Skin/GI flora translocation, urinary catheters, sexual contact.
Portals of Exit Details:
Upper Respiratory Tract: Expelled during talking, coughing, or sneezing. A single sneeze expels air from the nose at , releasing approximately bacteria into air currents.
Gastrointestinal Tract: Excreted in faeces, contaminating soil and municipal or marine water supplies.
Skin: Exudate from skin lesions or shedding of contaminated epidermal dust flakes.
Blood & Bodily Fluids: Bloodborne pathogen transmission (e.g., HIV, Hepatitis B, Hepatitis C).
- Modes of Transmission:
- **Contact Transmission:**
- *Direct Contact:* Physical body-to-body touch (skin, mucous membranes, perspiration, direct fluid contact).
- *Indirect Contact:* Transmission via inanimate intermediate contaminated objects called **fomites** (door handles, utensils, equipment).
- *Faecal-Oral Transmission:* Ingestion of pathogens due to inadequate handwashing after bowel movements.
- *Droplet Transmission:* Short-range respiratory droplets projected via coughing, sneezing, or talking.
- **Common Vehicle Transmission:**
- *Airborne:* Distant transport via air currents, dust particles, or dried droplet nuclei (aerosols). Mitigated by wet-mopping and damp dusting.
- *Water-borne:* Ingestion of water contaminated with sewage or faeces.
- *Food-borne:* Consumption of food contaminated via poor hygiene, improper thermal storage, or undercooking.
- **Vector Transmission:**
- *Mechanical Vector:* Passive transport of pathogens on the feet or body parts of insects (e.g., flies landing on faeces then food).
- *Biological Vector:* Active transmission where the pathogen multiplies or completes part of its lifecycle within the vector before injection via bites or stings (e.g., mosquitoes transmitting malaria, ticks transmitting Lyme disease).
Physical & Chemical Methods of Microbial Control
Antiseptics:
Chemical substances applied directly to living tissue or skin to reduce microbial numbers, sepsis, or putrefaction (e.g., Dettol, Savlon).
All antiseptics possess some level of host tissue toxicity or irritation; modern clinical practice prioritizes preventative aseptic technique.
Disinfectants & Sanitisers:
Disinfectants: Antimicrobial chemicals applied strictly to inanimate, non-living surfaces/objects to destroy vegetative microorganisms (e.g., Pine O Cleen).
Sanitisers: Chemical agents that reduce microbial contamination on surfaces to safe public health levels (e.g., Janola bleach, Napisan).
Note: Neither disinfectants nor sanitisers achieve complete sterilisation.
Sterilisation Methods:
Definition: The complete elimination or destruction of all living microorganisms, including vegetative bacteria, fungi, viruses, and bacterial endospores.
Autoclaving (Moist Heat Under Pressure): Uses saturated steam under pressure at for a minimum of . Kills all microbial life, including bacterial endospores.
Boiling Water: Exposure to boiling water () for kills vegetative bacteria and viruses, but is ineffective against bacterial endospores.
Dry Heat Sterilisation: Requires longer duration due to slower heat penetration (e.g., exposure to for ). Suitable for anhydrous powders, heat-stable glassware, and narrow glass pipettes.
Flaming: Direct exposure of metal inoculating loops and wires to an open Bunsen burner flame until glowing red hot.
Ultraviolet (UV) Radiation: Non-ionizing radiation used to sterilize exposed air, smooth surfaces, and transparent fluids. Overexposure causes human skin damage and carcinogenesis.
Food Preservation Techniques:
Hermetic sealing (canning/bottling) to exclude air.
Thermal processing (pasteurisation/heat treatment).
Refrigeration and freezing (slows metabolic/enzymatic activity; does not kill bacteria).
Desiccation (dehydration/drying).
Preservative chemical additives and acidification (e.g., pickling in vinegar to reduce pH below growth thresholds).
Immunity & Immunisation
Immunity: State of protection from infectious disease, categorized as natural or acquired.
Natural Immunity:
Inherent Resistance: Physiological or genetic unsuitability of host tissues for specific pathogen growth.
Maternal Passive Immunity: Transplacental antibody transfer in utero or immunoglobulin ingestion via breast milk protecting neonates.
Acquired Immunity:
Active Immunity: Long-term immunity stimulated by exposure to foreign antigens. The host produces specific memory B and T lymphocytes and antibodies.
Natural Active: Induced by recovering from an actual clinical infection.
Artificial Active (Vaccination): Induced by administering vaccines containing weakened (attenuated), inactivated (killed) pathogens, toxoids, or subunit antigens.
Passive Immunity: Short-term protection conferred by administering pre-formed, ready-made antibodies (e.g., hyperimmune serum or antitoxins). Does not generate host immunological memory.
Bacteria: Morphology, Arrangements, & Cellular Structure
General Properties: Single-celled prokaryotic organisms lacking a true membrane-bound nucleus and complex membrane-bound organelles. Highly adaptable; reproduce rapidly via binary fission.
- Bacterial Shapes (Morphology):
- **Coccus** (plural: *Cocci*): Spherical bacterium.
- **Bacillus** (plural: *Bacilli*): Rod-shaped bacterium.
- **Coccobacillus:** Intermediate oval shape between a coccus and bacillus.
- **Spirillum:** Rigid wavy or spiral shape.
- **Spirochaete:** Flexible, tightly coiled helical spiral.
- **Vibrio:** Curved, comma-shaped rod.
Bacterial Cellular Arrangements:
Diplococci / Diplobacilli: Pairs of cells.
Staphylococci: Irregular, grape-like clusters (formed by division in multiple random planes).
Streptococci / Streptobacilli: End-to-end linear chains (formed by division in a single plane).
Bacterial Ultra-Structure & Organelles:
Glycocalyx (Capsule / Slime Layer): Outer gelatinous sticky polysaccharide/polypeptide layer. Enhances surface attachment (e.g., Streptococcus mutans adhering to tooth enamel to produce plaque) and protects pathogens against host phagocytosis.
Cell Wall: Rigid complex peptidoglycan network protecting against osmotic lysis and maintaining cell shape.
Plasma Membrane: Phospholipid bilayer containing transport proteins beneath the cell wall, regulating nutrient and waste transport.
Nucleoid & Chromosome: Single, circular double-stranded DNA chromosome localized in an unenclosed nucleoid region.
Plasmids: Small, extra-chromosomal circular DNA molecules replicating independently of the main chromosome; carry non-essential genes conferring advantages such as antibiotic resistance or toxin production.
Ribosomes: prokaryotic ribosomes responsible for protein translation.
Flagella: Rigid rotating protein filaments powered by ATP driving bacterial motility. (e.g., Proteus vulgaris exhibits peritrichous flagella enabling swarming motility that forms concentric rings on agar).
Pili & Fimbriae: Hair-like protein surface projections. Fimbriae facilitate surface attachment; conjugation pili enable horizontal gene/plasmid transfer between bacterial cells.
Gram-Positive vs. Gram-Negative Cell Walls & Gram Staining
Gram-Positive Cell Wall:
Consists of a thick, multi-layered, dense peptidoglycan layer.
Interspersed with teichoic acids and lipoteichoic acids anchored to the underlying plasma membrane.
Contains outer S-layer proteins in specific species (e.g., Staphylococcus aureus).
Retains primary crystal violet dye, staining purple under the microscope.
Gram-Negative Cell Wall:
Consists of a very thin peptidoglycan layer located within a gel-like periplasmic space.
Surrounded by an Outer Membrane containing lipopolysaccharide (LPS) molecules composed of Lipid A (endotoxin), core polysaccharide, and O-specific side chains.
Decolorized by alcohol wash and takes up safranin counterstain, staining pink/red under the microscope (e.g., Escherichia coli).
Gram Stain Procedure (Differential Staining):
Primary Stain: Cover heat-fixed bacterial smear with Crystal Violet (1 minute); all bacterial cells stain purple.
Mordant: Apply Gram's Iodine (1 minute); forms a large, insoluble Crystal Violet-Iodine (CV-I) complex inside the cell wall.
Decolorization: Wash with Alcohol or Acetone/Alcohol mixture (briefly). Dehydrates thick Gram-positive peptidoglycan, trapping CV-I; dissolves lipid-rich Gram-negative outer membrane, leaching out CV-I complex.
Counterstain: Apply Safranin (1 minute); pink dye stains decolorized Gram-negative cells, while Gram-positive cells remain deep purple.
Endospore Formation (Sporulation)
Endospores:
Highly resistant, dehydrated, non-reproductive dormant structures produced inside specific Gram-positive bacteria (Clostridium spp. and Bacillus spp.).
Formed to survive extreme environmental stress (heat, fire, desiccation, chemical disinfectants, radiation, freezing) for centuries.
Germinate into active vegetative bacterial cells upon rehydration when favorable environmental conditions return.
- Steps of Sporulation (Figure 4.8):
1. **Incipient Stage:** Bacterial DNA replicates; a invaginated **spore septum** isolates the newly replicated DNA and a small portion of cytoplasm.
2. **Engulfment:** Plasma membrane grows inward around the isolated DNA, cytoplasm, and membrane, starting to enclose it.
3. **Forespore Formation:** Spore septum completely encloses the isolated portion, forming a **forespore** bounded by two distinct membranes.
4. **Peptidoglycan Cortex Synthesis:** A thick peptidoglycan cortex layer synthesizes between the two forespore membranes.
5. **Spore Coat Formation:** A protective protein **spore coat** forms around the peptidoglycan cortex, and the spore dehydrates.
6. **Release:** The mother cell lyses, releasing the mature **free endospore**.
Bacterial Growth Requirements & Reproduction
Physical Growth Requirements:
Temperature: Most human-associated bacterial species are mesophiles preferring temperatures between and . Temperatures above kill most vegetative bacteria; freezing halts growth without killing.
pH: Optimum bacterial growth occurs near neutral pH (). Highly acidic environments (e.g., pickling at low pH) inhibit bacterial growth.
Moisture & Osmotic Pressure: Aqueous environments are necessary to transport dissolved nutrients across the cell membrane.
Oxygen Classifications:
Obligate Aerobes: Require molecular oxygen for cellular respiration ("obliged" to have oxygen).
Obligate Anaerobes: Destroyed or inhibited by molecular oxygen ("obliged" to live without oxygen).
Facultative Anaerobes: Capable of growing in either the presence or absence of molecular oxygen.
Reproduction via Binary Fission:
Asexual reproduction generating genetically identical progeny cells.
Cell elongates, duplicates its circular DNA, and segregates one copy to each end of the cell.
The cell wall and plasma membrane grow inward (septation) to divide the parent cell into two daughter cells.
Doubling time (generation time) can be as short as under optimal conditions.
Pathogenic Bacteria & Associated Diseases
Staphylococcus species (Staphylococcus aureus):
Normal flora of skin and nasal passages.
Causes localized boils, impetigo, folliculitis, surgical wound infections, scalded skin syndrome, toxic shock syndrome, and food poisoning via enterotoxins.
Readily develops multidrug resistance (e.g., Methicillin-Resistant Staphylococcus aureus / MRSA).
Streptococcus species:
Normal flora of the throat and upper respiratory tract.
Causes pharyngitis (strep throat), tonsillitis, otitis media, cellulitis, pneumonia, and wound infections.
Untreated throat infections can trigger post-streptococcal autoimmune sequelae, including acute glomerulonephritis and rheumatic fever.
Toxigenic strains cause scarlet fever, necrotising fasciitis ("flesh-eating disease"), and quinsy (peritonsillar abscess).
Clostridium species:
Strict obligate anaerobic, endospore-forming Gram-positive bacilli.
Produce potent exotoxins causing tetanus (Clostridium tetani), botulism (Clostridium botulinum), gas gangrene (Clostridium perfringens), and pseudomembranous colitis/diarrhoea (Clostridium difficile).
Haemophilus influenzae (Hib):
Resident flora of the upper respiratory tract; causes otitis media, acute epiglottitis, and bacterial meningitis in young children.
Escherichia coli:
Dominant facultative anaerobe of intestinal normal flora; primary indicator organism for faecal water contamination.
Translocation causes urinary tract infections (UTIs); enterotoxigenic strains cause severe gastroenteritis and diarrhoea.
Proteus species (Proteus vulgaris):
Intestinal bacterium that translocates to cause urinary tract infections. Highly motile via flagella, exhibiting swarming behavior that produces concentric growth rings on agar media.
Intracellular Bacteria
Obligate Intracellular Bacteria: Extremely small bacteria incapable of reproducing outside a host eukaryotic cell.
Rickettsias:
Transmitted to humans via arthropod vectors (lice, fleas, ticks).
Cause systemic infections, including epidemic typhus and spotted fevers.
Chlamydias:
Transmitted via direct human-to-human contact or sexual contact.
Cause trachoma, non-gonococcal urethritis, pelvic inflammatory disease (PID), and atypical pneumonia.
Mycoplasmas:
Extremely small bacteria completely lacking a peptidoglycan cell wall (pleomorphic shape).
Naturally resistant to cell-wall targeting beta-lactam antibiotics (e.g., penicillin).
Cause atypical walking pneumonia and intrauterine infections.
Viruses: Structure, Classification, & Host Specificity
General Properties: Non-cellular sub-microscopic infectious agents measuring in size. Lack metabolic machinery and organelles; obligate intracellular parasites dependent on host cells for replication.
- Viral Structure:
- **Genome Core:** Nucleic acid consisting of either single- or double-stranded DNA or RNA (e.g., Influenza contains 8 segmented RNA molecules).
- **Capsid:** Protein coat surrounding the genome core, composed of repeating protein subunits called capsomeres.
- **Envelope:** Outer lipid bilayer membrane acquired from host cell membranes during viral budding; contains embedded viral glycoprotein spikes:
- **Hemagglutinin (H Spike):** Facilitates viral attachment to host cell surface receptors.
- **Neuraminidase (N Spike):** Facilitates viral release from infected host cells.
Viral Classification Criteria: Classified by nucleic acid type (DNA vs. RNA), capsid symmetry/geometry, presence or absence of an envelope, and replication strategy.
DNA Viruses: Parvoviridae, Papovaviridae (Papillomavirus causing warts), Adenoviridae, Herpesviridae, Poxviridae.
RNA Viruses: Picornaviridae, Togaviridae, Reoviridae, Coronaviridae (Coronavirus), Myxoviridae / Paramyxoviridae, Rhabdoviridae, Retroviridae.
Host Specificity: Highly specific host cell tropism mediated by precise binding between viral attachment spikes and complementary host cell membrane receptors (glycoproteins or glycolipids). Exceptions capable of cross-species transmission (zoonoses) include Rabies, Zika, Avian Influenza, and Swine Influenza.
Viral Replication & Pathogenic Viruses
Six-Step Viral Replication Cycle (in Human Cells):
Attachment: Viral surface spikes bind specifically to complementary glycoprotein receptor sites on the target host cell membrane.
Penetration: The virus enters the host cell via receptor-mediated endocytosis or direct membrane fusion.
Uncoating: Host or viral enzymes breakdown the protein capsid, releasing viral nucleic acid into the cytoplasm.
Biosynthesis: Viral nucleic acid enters the host cell nucleus/cytoplasm, hijacking host ribosomes, ATP, and enzymes to transcribe viral mRNA, replicate the viral genome, and synthesize viral structural proteins.
Assembly: Viral nucleic acid genomes and synthesized capsomeres self-assemble into mature daughter virions.
Release: Mature virions exit the cell via cell lysis (destroying the cell) or budding (taking part of the host plasma membrane as an envelope).
Selected Pathogenic Viruses & Clinical Conditions:
Herpesviridae (DNA):
Herpes simplex virus (HSV): Type 1 causes oral cold sores; Type 2 causes genital lesions.
Varicella-zoster virus (VZV): Primary infection causes chickenpox; reactivation causes shingles (herpes zoster).
Epstein-Barr virus (EBV): Causes infectious mononucleosis (glandular fever).
Myxoviridae & Paramyxoviridae (RNA):
Influenza virus: Acute febrile respiratory illness.
Rubeola virus: Causes measles.
Mumps virus: Causes painful inflammation of the parotid salivary glands.
Respiratory Syncytial Virus (RSV): Causes severe bronchiolitis and pneumonia in infants.
Retroviridae (RNA):
Human Immunodeficiency Virus (HIV): Infects helper T-lymphocytes, leading to Acquired Immunodeficiency Syndrome (AIDS).
Papillomaviridae (DNA):
Human Papillomavirus (HPV): Causes skin warts and cervical/mucous membrane neoplasia.
Poliovirus: Target of the Global Polio Eradication Initiative, which reduced global cases by over since 1988 (leaving only Afghanistan and Pakistan polio-endemic as of 2023; New Zealand's last case was reported in 1998).