Prokaryote Fundamentals & Virulence (Lecture 2)

Bacterial Morphology & General Structure

Prokaryotes (bacteria) possess a relatively simple, non-compartmentalized architecture. Their cytoplasm is a "bag" of ribosomes, enzymes and a single circular chromosome without a membrane-bound nucleus. A rigid peptidoglycan cell wall external to the plasma membrane maintains a defined cell shape, much like a balloon animal’s sculpted form.

• Common shapes
• Cocci (“berries”): round spheres – e.g.
• Staphylococcus aureus (clusters resemble grape bunches)
• Streptococcus pyogenes (chains, “string of pearls”)
• Bacilli: rod-shaped – e.g. Escherichia coli
• Spiral/filamentous: cork-screw or pasta-like – e.g. Treponema pallidum (syphilis)

Visualization & Culture Techniques

• Light microscopy (≤ 400\times) readily visualises bacteria, fungi, protozoa but not viruses (require electron microscopy).
• Solid media (agar) supply nutrients (blood, sugars, salts) ➜ colonies develop with distinct shapes, sizes, colours & textures.
• Blood agar: red; demonstrates haemolysis patterns.
• Chocolate agar: “cooked” blood; supports fastidious bugs (e.g. Streptococcus pneumoniae).
• Example cultures & clinical images
• Normal hand flora showing Bacillus spp., coliforms, staphylococci, Corynebacterium.
• S. pneumoniae colony growth correlating with right-lower-lobe consolidation on chest X-ray (pneumonia).
• Mixed S. aureus + S. pyogenes necrotising fasciitis in a diabetic patient ➜ extensive tissue debridement required.

Gram Stain: Principles & Procedure

1. Fixed smear on slide → sequential dyes & mordants (crystal violet, iodine, decolouriser, safranin).

2. Outcome
   • Gram-positive = purple (retain crystal-violet–iodine complex).
   • Gram-negative = pink/red (safranin counter-stain).

3. Provides two simultaneous taxonomic clues: shape (coccus vs rod) + cell-wall type.

4. Clinical utility: • visibility • empiric antibiotic direction • virulence insight.

Bacterial Cell Wall Architecture

• Key constituent = peptidoglycan.

Oxygen & Metabolic Preferences

• Obligate aerobes – require O2; grow at broth surface. • Obligate anaerobes – harmed by O2; sink to bottom.
• Facultative anaerobes – flexible; dispersed but denser at top.

Clinical pearls
• Deep, poorly perfused wounds (e.g. injection injury) support anaerobes such as Clostridium septicum producing black foul-smelling pus; specialised antibiotics needed.
• Human bite wounds = oral anaerobe mixture ➜ aggressive management.

Temperature Preferences (Extremophiles)

Microbes exhibit Goldilocks behaviour – each has an optimum range.

• Mesophiles ("middle lovers"): 20–45^\circ\text{C} – covers normal flora & human pathogens.
• Thermophiles: >45^\circ\text{C} (e.g. geothermal pools of Rotorua).
• Psychrophiles: <10^\circ\text{C} to -15^\circ\text{C}.

Outside preferred range they enter stasis or die.

Prokaryotic Protein Synthesis vs. Eukaryotic

• In bacteria transcription (RNA polymerase) & translation (70S ribosomes) happen side-by-side in cytoplasm – no nuclear envelope barrier.
• Eukaryotes: transcription in nucleus ➜ mRNA export ➜ translation on 80S ribosomes in cytoplasm/ER.
• Therapeutic relevance: many antibiotics selectively target bacterial ribosomes, polymerases, or cell-wall steps, sparing host cells.

Cell Division: Binary Fission & Growth Kinetics

1. Chromosome replication.

2. Cell elongation.

3. Septum (cross-wall) formation.

4. Separation into two identical daughters.

Under ideal conditions some species divide every 10\text{–}20\,\text{min}. Population after n divisions: 2^n – exponential surge explains rapid infection spread.

Bacterial Genetics & Horizontal Gene Transfer (HGT)

Genomic basics
• Single circular chromosome (haploid).
• Asexual, clonal fission ⇒ no meiotic recombination.

Three HGT "sex" mechanisms enable in-life evolution:

1. Conjugation – pilus-mediated "mating bridge" transfers plasmids between live cells.

2. Transformation – uptake of naked DNA/plasmids from lysed neighbours ("plug-and-play").

3. Transduction – bacteriophages act like mosquitoes, injecting DNA they picked up elsewhere.

Consequences: swift spread of antibiotic resistance, new toxins, immune-evasion tricks.

Virulence Factors: The Bacterial Superpowers

Adhesion & Evasion

• Pili/fimbriae and capsules (gelatinous polysaccharide halos) let bugs stick to mucosa & dodge phagocytes/antibiotics.
• Example: Klebsiella pneumoniae – prominent capsule.

Invasion Enzymes

• Collagenase – digests host collagen.
• Hyaluronidase – breaks down hyaluronic acid matrix.
⇒ Facilitates deeper tissue penetration & spread ➜ stronger inflammatory damage.

Haemolysins

• Secreted by S. pyogenes, S. aureus.
• Lyse erythrocytes; on blood agar produce α-, β- or γ-haemolysis (green clearing, complete clearing, none).
• Benefits: nutrient iron, camouflage in blood, vascular dissemination.

Spore Formation

• Internal "hibernation pod" forms when environment hostile (heat, salt, antibiotics).
• Highly resistant to boiling, freezing, radiation.
• Species: Clostridium tetani, C. botulinum, C. difficile, plus protozoans Cryptosporidium & Giardia.
• Spores germinate on sensing favourable conditions ⇒ sudden disease "surprise".

Biofilms

• Dense extracellular polysaccharide-protein matrix over tissue or foreign body (catheters, prosthetic joints/valves, IUDs).
• Blocks antibiotics & antibodies; impedes wound healing ➜ chronic infections, especially in diabetics.

Toxins

Systemic manifestations (fever, pain, malaise) largely toxin-mediated rather than due to sheer bacterial mass.

Case Highlights & Clinical Correlates

• Necrotising fasciitis (S. aureus + S. pyogenes) in diabetes – aggressive debridement + antibiotics.
• Clostridial anaerobic wound infection – black, foul "rotten fish" pus; O_2 avoidant.
• Tetanus in unvaccinated child – lockjaw, spasms due to tetanospasmin toxin.
• Meningococcal endotoxin – late bruise-like rash signals urgent treatment.

Key Take-Home Points

• Shapes: cocci, bacilli, spirals.
• Gram reaction: $\text{purple} = +$, $\text{pink} = -$; linked to peptidoglycan thickness & LPS content.
• Metabolism: obligate/facultative aerobes vs anaerobes.
• Temperature niches: psychrophile, mesophile, thermophile.
• Protein synthesis, cell division & walls differ from eukaryotes – antibiotic targets.
• Binary fission ⇒ 2^n growth; HGT (conjugation, transformation, transduction) drives evolution.
• Virulence arsenal: adhesion factors, enzymes, haemolysins, spores, biofilms, exotoxins & endotoxins – each with clinical implications.

End of Lecture 2 Notes