Microbiology 6 - Propagation of Viruses.pdf

Microbiology Propagation of Viruses Microbiology| Propagation of Viruses Contents : Anti-Viral Therapy 3 Interferons 11 Detection of virus-Infected cells 16 Interactions 20 Microbiology| Propagation of Viruses Anti-Viral Therapy : When to use antiviral therapy: 1. when vaccines are not available or...

Microbiology Propagation of Viruses Microbiology| Propagation of Viruses Contents : Anti-Viral Therapy 3 Interferons 11 Detection of virus-Infected cells 16 Interactions 20 Microbiology| Propagation of Viruses Anti-Viral Therapy : When to use antiviral therapy: 1. when vaccines are not available or not highly effective 2. multiplicity of serotypes (e.g., rhinoviruses) 3. constantly changing virus (e.g., influenza, HIV). Who need to treat with antiviral agents: 1. when vaccines would not be effective 2. immunosuppressed patients. Microbiology| Propagation of Viruses A. Nucleoside and Nucleotide Analogs : Synthetic Inhibit cellular enzymes. Virus encoded enzymes. Competes with normal nucleotides for incorporation into viral DNA or RNA. Microbiology| Propagation of Viruses A. Nucleoside and Nucleotide Analogs : Synthetic Associates with viral polymerase, irreversible, Causes chain termination. Acylovir HSV VZV Gancyclovir Cytomegalovirus Microbiology| Propagation of Viruses Nucleotide analogs differ from nucleoside analogs in having an attached phosphate group. persist in cells for long periods of time. Resistance arise. The use of combinations of antiviral drugs can delay the emergence of resistant (e.g. HAART) Microbiology| Propagation of Viruses B. Reverse Transcriptase Inhibitors : Characteristics It does not require phosphorylation for activity. does not compete with nucleoside triphosphates. Action Binds to reverse transcriptase. disrupting the enzyme’s catalytic site. Ex: Nevirapine Microbiology| Propagation of Viruses C. Protease Inhibitors : Synthetic. designed by computer. Action inhibit the viral proteases. Prevent mature virion core formation. Prevent activation of RT. Microbiology| Propagation of Viruses Yields noninfectious virus particles. Ex: Saquinavir for HIV indinavir and ritonavir Saquinavir Microbiology| Propagation of Viruses D. Other Types of Antiviral Agents : Fuzeon: blocks the virus and cellular membrane fusion (e.g. HIV1) amantadine and rimantadine: blocking viral un-coating (Inf. A ) Foscarnet: inhibits viral DNA polymerases and reverse transcriptases Methisazone: blocked a late stage in viral replication result non-infectious virus (e.g. poxvirus) Microbiology| Propagation of Viruses Interferons : Alpha, Beta & Gamma. Induced during viral infection. Used for HBV & HCV. Microbiology| Propagation of Viruses RNA viruses are stronger inducers than DNA. Degrade viral mRNA. Inhibit protein synthesis of the virus. prompting the synthesis of other proteins in the host cell which inhibit viral replication. Enhance MHC I, II expression to present viral antigens. Induce nitric oxide synthetase. Microbiology| Propagation of Viruses Isolation and cultivation of Viruses : Growth of virus in animals is still used for the primary isolation of certain viruses and for 1. study pathogenesis. 2. study viral oncogenesis. 3. Diagnostic to recover virus from clinical samples. 4. for research. Microbiology| Propagation of Viruses In vivo Animals: cows; chickens; mice; rats. Embryonated eggs. In vitro Organ and tissue culture cells derived from an animal. a) Primary cultures (trypsin), grow to one or few passages in culture. Microbiology| Propagation of Viruses b) Diploid cultures have a finite lifespan. They usually can undergo a maximum of 50 passages before they senesce. Normal human cells, such as human skin fibroblasts, are one example of diploid cells. c) Continuous cell lines are immortalized cell lines with an infinite lifespan. These usually either come from tumor tissue or have been deliberately immortalized or transformed. However, many rodent cell lines spontaneously transform. The type of cell culture used for viral cultivation depends on the sensitivity of the cells to a particular virus. Microbiology| Propagation of Viruses Detection of Virus-Infected Cells : 1. CPE: Viral Cytopathological Effects cell lysis or necrosis, inclusion formation, giant cell formation, and cytoplasmic vacuolization 2. Appearance of a virus-encoded protein, such as the hemagglutinin of influenza virus. 3. Detection of virus-specific nucleic acid (PCR). 4. Adsorption of erythrocytes to infected cells, called hemadsorption. Microbiology| Propagation of Viruses 5. Viral growth in an embryonated chick egg. EX: death of embryo (e.g., encephalitis viruses) production of pocks or plaques (e.g., herpes, smallpox, vaccinia) development of hemagglutinins in embryo (e.g. Influenza) Microbiology| Propagation of Viruses Cell death (Cell rounding/Degeneration/Aggregation) Inclusion bodies in the nucleus or cytoplasm (react to acid dyes) situated in the nucleus (herpes) in the cytoplasm (poxvirus) In both (measles virus) Cell surface changes Viral antigen expression Hemadsorption (hemagglutinin expression) Microbiology| Propagation of Viruses Normal Cell and CPE : Microbiology| Propagation of Viruses Interactions : when two genetically distinct viruses infect a cell different phenomena can result antigenic drift: (minor changes) individual bases in the DNA or RNA mutate to other bases. Most of these point mutations are "silent" – they do not change the protein that the gene encodes – but others can confer evolutionary advantages such as resistance to antiviral drugs. Antigenic shift occurs when there is a major change in the genome of the virus. Microbiology| Propagation of Viruses A. Recombination : The classic mechanism is that the nucleic acid strands break, and part of the genome of one parent is joined to part of the genome of the second parent. Recombination involves the breakage and rejoining of two chromosomes (M and F) to produce two re-arranged chromosomes (C1 and C2). Yield progeny similar to itself upon replication. Microbiology| Propagation of Viruses Reassortment ( segmented genomes) RNA viruses : with segmented genomes (e.g., influenza virus) the formation of recombinants is caused by reassortment of individual genome fragments rather than by an actual crossover event. Microbiology| Propagation of Viruses B. Complementation : Occurs among defective viruses. defective interfering virus particles: have lost essential segments of genome but contain normal capsid proteins. they require infectious homologous virus as helper for replication. they interfere with the multiplication of that homologous virus. Microbiology| Propagation of Viruses C. Phenotypic Mixing (special type of complementation) : Phenotypic mixing usually occurs between different members of the same virus family genotype with a heterologous phenotype. Occur when genome of one virus becomes randomly incorporated within capsid proteins of another virus. phenotypic mixing also can occur between enveloped viruses (Not necessary closely related viruses) “phenotypic masking” or “transcapsidation”: If the genome is encased in a completely heterologous protein coat