Bacteriophage.docx

Different shapes Genetic composition can be single, double stranded. Can be RNA or DNA. Their genome can code as few as 4 genes or as any as 100s. Analysis and isolation of phage’s Plaque assay is used. Sample from the environment is taken. The sample is diluted. Mix with appropriate host cells. Pou...

Different shapes Genetic composition can be single, double stranded. Can be RNA or DNA. Their genome can code as few as 4 genes or as any as 100s. Analysis and isolation of phage’s Plaque assay is used. Sample from the environment is taken. The sample is diluted. Mix with appropriate host cells. Pour on agar plate. Incubate to let them grow. Plaque forms a clear zone in the bacterial lawn. Structure of bacteriophages Can be icosahedral. Filamentous Complex 2 types of bacteriophages Lytic phage’s. can only undergo lytic cycles. Temperate phage’s. can undergo lytic and lysogenic cycle Lytic cycle Adsorption/attachment- Virus attaches to the cell surface receptors. Penetration- The viral genome is injected into the bacterial cell. Replication and synthesis- the viruses nucleic acid gets replicated as it hijacks the machinery. Assembly- the bacteriophage components assemble. Release- enzymes break the cell peptidoglycan causing osmotic lysis. Lysogenic cycle Attachment/Adsorption Penetration Prophage formation- integrates into the cells genome and forms a prophage. Maintenance- every time the bacterium divides, it replicates the prophage genome. Spontaneous induction- phage genome is excised from host cell. Replication and synthesis Assembly Release The only difference between the 2 cycles is the prophage part. How does phage decide between lytic and lysogenic cycle? The genome of the phage contains different transcription factors called C1 and cro. These facilities synthesis of genes. C1 facilitates the synthesis of genes needed for lysogenic cycle and cro facilitates the synthesis of genes needed for lytic cycle. The region between the C1 and cro is operator sequences that the C1 and cro interacts with and physically compete. More C1 binding to operator region means it will outcompete cro and will undergo lysogenic cycle and vice versa. How does the phage switch between these 2 cycles? Another protein called CII. Involved in monitoring health of the cell and decided which cycle to choose accordingly. CII is sensitive to degradation by host proteases- HfLA When cells are actively growing and have enough nutrients, they produce high levels of the protease HfLA, CII is degraded by HfLA so the cells undergo lytic growth. If there are not enough nutrients, the cells won’t produce as much protease, so CII is activated and activates transcription of CI and promotes lysogenic growth. Non-virulent bacteria can transform into highly virulent pathogens through lysogenic conversion, Lysogenic bacteriophages may contribute to development of antibiotic resistance. To treat pathogenic infections. Phage therapy -use of lytic bacteriophages 4 principles Phage must be lytic. A single dose of phage should treat infection. Phage is non-toxic and highly specific for targeted bacterial populations. Antibiotic-resistant bacteria remain sensitive to phage-mediated lysis. Bacteriophages Antibiotics Usually targets a very narrow host range, the disease-causing bacterium must be identified. Target both pathogenic microorganisms and normal microflora. Ten-fold lower rate Resistance to antibiotics is not limited to targeted bacteria. New phage’s can be accomplished in days or weeks Make take several years to develop Few minor side effects Multiple side effects including secondary infections Despite the lack of immune system, bacteria possess different ways to fight against the viral attack. Restriction endonucleases Antiviral system CRISPR. Restriction endonucleases Recognises short and specific sequences and cut them. Protects hosts own DNA by modifications such as methylation. CRISPR = clustered regularly interspaced short palindromic repeats Short DNA sequences in the genome of bacteria that are left behind by the phage that has previously infected the bacteria. Applications in gene editing.