7.7 Microbiology: Bacterial Genetics & Viral Life Cycles

Prokaryotic Morphology, Physiology & Cell Wall Biochemistry

Bacteria are single-celled prokaryotic organisms lacking a membrane-bound nucleus or membrane-bound organelles. Their cellular structure and metabolic adaptations distinguish them from eukaryotes.

Gram-Positive vs. Gram-Negative Cell Envelope

The bacterial cell envelope is classified using the Gram stain technique (Crystal Violet, Iodine, Alcohol Wash, Safranin Counterstain).

  Gram-Positive Envelope                    Gram-Negative Envelope
  +---------------------------+             +---------------------------+  Outer Membrane
  | Thick Peptidoglycan       |             | LPS (Lipopolysaccharide)  |  (LPS Endotoxin)
  | Layer (Teichoic Acid)     |             +---------------------------+
  +---------------------------+             | Periplasmic Space / Thin  |  Thin Peptidoglycan
  | Plasma Membrane           |             | Peptidoglycan Layer       |  & Periplasm
  +---------------------------+             +---------------------------+
                                            | Inner Plasma Membrane     |
                                            +---------------------------+

Bacterial Growth Dynamics

Bacterial populations reproduce by binary fission, following a predictable four-phase logarithmic growth curve when cultured in a closed system:

1. Lag Phase: Bacteria adapt to the new environment; active protein synthesis occurs without cell division.
2. Log (Exponential) Phase: Exponential growth and rapid binary fission ($2^n$). Bacteria are most susceptible to cell-wall targeting antibiotics during this phase.
3. Stationary Phase: Growth rate equals death rate as essential nutrients are depleted and toxic metabolic wastes accumulate.
4. Death (Decline) Phase: Toxic waste buildup causes exponential cell mortality.

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Mechanisms of Bacterial Horizontal Gene Transfer

Bacteria generate genetic diversity through horizontal gene transfer (HGT) via three distinct mechanisms.

  Transformation  --->  Direct uptake of naked environmental DNA
  Conjugation     --->  Direct cell-to-cell transfer via Sex Pilus (F factor)
  Transduction    --->  Bacteriophage virus serves as vector

1. Transformation

Direct uptake and integration of exogenous, naked DNA fragments from the surrounding environment by a competent bacterial cell. Competence can be natural or induced artificially in laboratory settings using chemical shock ($CaCl_2$) or electroporation. Classic historical evidence was demonstrated by Frederick Griffith's 1928 experiment using Streptococcus pneumoniae.

2. Conjugation

Bacterial "sexual" transfer involving direct cell-to-cell contact via a specialized sex pilus bridge. It is mediated by plasmid genes, most notably the Fertility factor (F factor).

• $F^+ \rightarrow F^-$ Cross: An $F^+$ cell (donor containing the extrachromosomal F plasmid) extends a sex pilus to attach to an $F^-$ recipient cell. The F plasmid undergoes rolling-circle replication, transferring a single strand of plasmid DNA into the recipient. The recipient synthesizes a complementary strand, converting it into an $F^+$ cell.
• High-Frequency Recombination ($Hfr$) Strains: The F factor plasmid integrates into the main bacterial host chromosome via homologous recombination. When an $Hfr$ cell undergoes conjugation, it attempts to transfer its entire integrated chromosome into the recipient. Because the conjugation bridge usually breaks before the entire chromosome is transferred, the recipient receives chromosomal genes adjacent to the insertion site, remaining $F^-$.

3. Transduction

The transfer of bacterial host DNA from one cell to another mediated by a bacteriophage virus vector.

• Generalized Transduction: Occurs during the lytic cycle. Bacteriophage enzymes degrade host DNA into fragments. During viral packaging, a fragment of bacterial host DNA is accidentally packaged into a newly formed viral capsid instead of viral DNA. When this transducing phage infects a new host, it injects the bacterial DNA.
• Specialized Transduction: Occurs during the transition from the lysogenic cycle to the lytic cycle. A prophage integrated into the host genome excises imprecisely, carrying adjacent bacterial host genes with it into all newly synthesized viral progeny.

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Viral Structure & Genome Classification

Viruses are non-cellular obligate intracellular parasites that cannot synthesize ATP or proteins independently, requiring host cell machinery for replication.

Virion Structure

• Nucleic Acid Core: DNA or RNA genome (never both); single-stranded (ss) or double-stranded (ds).
• Capsid: Protein shell composed of repeating subunit monomers called capsomeres.
• Envelope: Optional outer lipid bilayer derived from host cell membranes during viral budding, containing viral glycoprotein spikes (e.g., Influenza hemagglutinin). Enveloped viruses are sensitive to heat and detergents; non-enveloped (naked) viruses are resilient.

Genomic Replication Strategies

According to the Baltimore classification system, viral genome structure dictates replication pathways:

  +ssRNA Virus    --->  Functions directly as mRNA (Host ribosome translates)
  -ssRNA Virus    --->  Must carry pre-packaged RNA-dependent RNA Polymerase (RdRp)
  Retrovirus      --->  Carries Reverse Transcriptase (Converts ssRNA into dsDNA provirus)

• Positive-Sense Single-Stranded RNA ($+\text{ssRNA}$): The viral genome functions directly as mature mRNA. Upon entering the host cytosol, host ribosomes immediately translate viral proteins, including RNA-dependent RNA polymerase (RdRp).
• Negative-Sense Single-Stranded RNA ($-\text{ssRNA}$): The viral genome is complementary to mRNA. Human host cells do NOT possess enzymes capable of transcribing RNA from an RNA template. Therefore, $-\text{ssRNA}$ virions MUST package pre-formed RdRp enzyme inside their capsid to transcribe the negative strand into a complementary $+\text{ssRNA}$ mRNA before translation can occur.
• Retroviruses ($+\text{ssRNA}$ with DNA intermediate, e.g., HIV): Enveloped viruses carrying two single-stranded $+\text{ssRNA}$ molecules and three essential enzymes:
  1. Reverse Transcriptase (RNA-dependent DNA polymerase): Synthesizes double-stranded cDNA from the viral RNA genome.
  2. Integrase: Integrates viral cDNA into the host nuclear chromosome, forming a permanent provirus.
  3. Protease: Cleaves polyprotein precursors during virion maturation.

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Bacteriophage Life Cycles: Lytic vs. Lysogenic

Bacteriophages are viruses that infect bacteria, displaying two distinct developmental pathways.

  Lytic Cycle      --->  Rapid viral multiplication -> Host cell lysis (Virulent)
  Lysogenic Cycle  --->  Viral genome integrates as Prophage -> Passive replication (Temperate)

1. The Lytic Cycle (Virulent Phages)

1. Attachment & Injection: Phage tail fibers bind specific bacterial cell wall receptors and inject viral DNA into the cytoplasm.
2. Synthesis: Viral genes take over host machinery, hydrolyzing host chromosomal DNA and synthesizing viral mRNA, capsids, and genomic copies.
3. Assembly: Viral components self-assemble into mature progeny virions.
4. Lysis: Viral lysozyme degrades the peptidoglycan cell wall. Water enters osmotic swelling, causing the host cell to burst (lyse) and release hundreds of infectious phages.

2. The Lysogenic Cycle (Temperate Phages)

1. Integration: Phage DNA integrates into the host bacterial chromosome via site-specific recombination, becoming a dormant prophage.
2. Passive Replication: The bacterium reproduces normally via binary fission, copying the prophage along with its own chromosome and transmitting it to all daughter cells.
3. Induction: Environmental stressors (e.g., UV radiation, chemical damage) trigger excision of the prophage from the host chromosome, initiating the lytic cycle.

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Subviral Particles: Prions & Viroids

Subviral particles are infectious entities simpler than classic viruses.

1. Prions (Infectious Misfolded Proteins)

Prions are infectious agents consisting entirely of protein molecules containing NO nucleic acids (no DNA or RNA).

• Pathogenic Mechanism: Normal cellular prion protein ($\text{PrP}^c$) rich in $\alpha$-helices is expressed in neuronal membranes. The pathogenic isoform ($\text{PrP}^{Sc}$) adopts a disease-causing conformation rich in $\beta$-pleated sheets.
• Cascade Effect: $\text{PrP}^{Sc}$ binds $\text{PrP}^c$ and acts as a template, inducing normal $\alpha$-helical proteins to refold into pathological $\beta$-sheet conformations. Misfolded $\text{PrP}^{Sc}$ aggregates into insoluble, protease-resistant amyloid plaques.
• Diseases: Transmissible spongiform encephalopathies (TSEs) such as Creutzfeldt-Jakob disease (CJD), Kuru, and Bovine Spongiform Encephalopathy (Mad Cow Disease).

2. Viroids

Viroids are small, circular single-stranded RNA molecules lacking a protein capsid. They primarily infect plant hosts, base-pairing with host mRNA transcripts to induce gene silencing via RNA interference.