Overview
This lecture covers bacteriophage (phage) biology, focusing on phage life cycles, their role in horizontal gene transfer, bacterial resistance mechanisms, and modern applications in genetic engineering and medicine.
Introduction to Phages
- Phages are viruses that infect bacteria or archaea.
- They have two main life cycles: lytic (destroy host) and lysogenic (integrate into host genome).
- Phages can transfer genes, including toxins, between bacteria.
Phage Life Cycles
- Lytic cycle involves phage replication, bacterial lysis, and release of new phages (e.g., T4 phage).
- Lysogenic cycle involves phage DNA integrating into the bacterial genome and replicating with the host (e.g., lambda phage).
- Decision between cycles depends on host cell conditions: abundant resources favor lytic; poor conditions favor lysogenic.
Phage Structure and Infection
- Phages have a head (contains nucleic acid) and tail (recognizes host cell).
- Infection starts with specific attachment, followed by penetration (DNA injection) aided by lysozyme.
- Early phage genes hijack host machinery and destroy bacterial DNA, sparing modified phage DNA.
Phage Detection: Plaque Assay
- Plaque assays mix bacteria and phages on agar to visualize zones of lysis (plaques).
- Plaques indicate the presence of lytic phages.
Phages and Horizontal Gene Transfer
- Lysogenic phages can mistakenly carry host genes during excision, spreading traits like toxins.
- Example: E. coli O157:H7 (EHEC) harbors a shigga toxin gene from a lysogenic phage, causing disease.
E. coli O157:H7 Outbreaks
- EHEC infects through contaminated food; only ~10 bacteria needed for infection.
- Shigga toxin binds receptors on intestinal and kidney cells, inhibiting protein synthesis and causing disease.
- Cattle are common reservoirs, shedding EHEC in feces, often contaminating food.
Bacterial Defense Mechanisms
- Restriction enzymes cut foreign DNA at specific sequences; host DNA is protected by methylation.
- CRISPR systems archive foreign DNA segments as spacers, guiding Cas enzymes to cut matching invaders in subsequent infections.
Applications: Genetic Engineering and Phage Therapy
- CRISPR-Cas systems allow precise genome editing in various organisms.
- Phages are being developed as antibacterial therapies, potentially complementing antibiotics, though resistance can arise.
Key Terms & Definitions
- Phage (Bacteriophage) — virus that infects bacteria.
- Lytic Cycle — phage replication causing host cell lysis.
- Lysogenic Cycle — phage DNA integrates into host genome.
- Plaque Assay — method to detect and quantify lytic phages.
- Restriction Enzyme — protein that cuts DNA at specific sequences.
- CRISPR — bacterial DNA sequences used to recognize and destroy foreign DNA.
- Cas9 — CRISPR-associated nuclease for cutting DNA.
- Shigga Toxin — toxin encoded by a phage, responsible for EHEC symptoms.
Action Items / Next Steps
- Review diagrams of phage life cycles and defense systems.
- Read about CRISPR applications in genome editing.
- Examine case studies on EHEC outbreaks and phage therapy development.