MICR221 – Final – Flashcards
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| What is the major change currently going on in microbiological biotech? What are the benefits of the change? |
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| The displacement of chemical processes to biological ones Biological processes can be done at ambient temperature & pressure; have reduced emissions & energy usage; biological processes have renewable feedstock (biomass) |
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| Examples of biofuels used |
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| Ethanol & Butanol |
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| What is the most widely produced "bioproduct" in the world? |
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| Ethanol |
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| Rationale for EtOH Fermentation |
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| Can oxygenate oil to improve combustion & reduce emission Gasoline extender - import less oil (economic benefit) Renewable transportation fuel |
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| What is the microbe used for ethanol fermentation? |
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| Saccharomyces cerevisiae |
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| What is the benefit of a positive energy balance ratio? |
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| Means that the product produces more energy than is used to make it |
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| What compound has the highest energy balance ratio and why? |
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| Sugarcane has the highest ratio because the remnants from its fermentation are burned to act as feedstock for the distillation |
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| What is the problem with using biomass as a next-gen biofuel? |
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| The presence of lignin bottlenecks the process; lignin coats the cellulose making it difficult to hydrolyze |
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| What are byproducts of EtOH fermentation and what are they used for? |
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| CO2 - sold to pop companies DDG - protein from corn; feedstock for animals |
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| What is one of the limitations of S. cerevisiae regarding the sugars it can ferment? |
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| Can only ferment 6C, up to the size of dimers |
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| What is end product inhibition in relation to ethanol fermentation? |
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| When EtOH is produced by yeast, its build-up functions to inhibit the yeast Reaction kinetics - as the concentration of EtOH increases, the rate of the reaction decreases because of EPI |
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| What is typically the highest % EtOH you can obtain via fermentation w/ S. cerevisiae? |
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| Typically 10% EtOH/ 90% H2O because of the EPI that occurs |
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| Fermentation Conditions for S. cerevisiae |
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| Temperature = ambient (about 34 degrees C) pH = 4-6 (slightly acidic) Microaerobic conditions Minimal additional nutrients need to be added Highly exothermic reaction |
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| What is the typical EtOH yield in EtOH fermentation? |
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| 45-50% |
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| EtOH Fermentation - Fermentation vs. Recovery |
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| Fermentations that lead to the highest concentrations of EtOH have the lowest costs needed for distillation, however, more energy and time needs to be put in to get the maximal concentration due to EPI In the end RECOVERY > FERMENTATION; therefore, keep the reaction going as long as possible to get the highest concentration possible |
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| Extractive Fermentation of EtOH (machine designed by Daugulis) |
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| Functions to eliminate/reduce EPI Add H2O immiscible solvent with affinity for EtOH - this solvent extracts the ethanol produced Heat the solvent to boil off the ethanol, while the solvent gets recycled Decreases the effect of EPI by eliminating produced ethanol from fermentation vessel; increases reaction rates |
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| Rationale for Biobutanol Use |
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| BuOH can be blended at higher concentrations w/ gasoline compared to EtOH Higher energy content Evaporates more slowly to give less emissions |
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| What is the organism used in Acetone-BuOH-EtOH (ABE) fermentation? |
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| C. acetobutylicum (gram-positive obligate anaerobe) |
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| What is one of the problems with ABE fermentation? |
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| Makes too many side-products |
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| Kinetics of ABE fermentation |
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| Anaerobic (no O2 b/c acetobutylicum is an obligate anaerobe) Acidogenic phase (creation of acids), then solventogenic phase (formation of products) EPI causes low efficiency (same reason as EtOH) |
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| Yield of ABE fermentation? |
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| 2% |
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| 3 main development phases in the formation of a biological process |
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| Strain Development - find a microbe as a transforming agent Medium Development - need to know how to grow culture Process Development - need to maximize yield & profit |
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| What types of antibiotics comprise the class of B-lactams? |
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| Penicillins & Cephalosporins |
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| Why are almost all antibiotics made via biological processes? |
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| Typically there are no chemical processes that can be used instead 158/160 antibiotics being synthesized have only biological means of doing so |
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| General structure of B-lactams |
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| All contain the same general structure - polyaromatics with the same central ring structure |
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| Difference in R group substitution between penicillins vs. cephalosporins |
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| Penicillins - 1 R group substitution possible Cephalosporins - 2 R group substitutions |
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| What are semi-synthetic antibiotics? |
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| They are anti-biotics that have the molecule produced by a microbe acquired, and then has its R group cleaved with the addition of a new R group to change the functionality Post-synthesis modulation of the R group |
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| How can microbes be nudged into producing a certain type of antibiotic? |
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| Can provide the precursor for the antibiotic in the medium to nudge microbe in favor of producing specific antibiotic Note that these are NOT semi-synthetic antibiotics |
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| What was the original species used to synthesize penicillin? What is the current species used? |
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| Old species = Penicillium notatum Current = Penicillium chrysogenum |
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| Why was P. notatum not a good organism to be used for mass synthesis of penicillin? |
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| Because it contained NO good process traits and could not be grown on liquid cultures (only grows on surfaces) |
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| Overview of Strain Development & Mutation |
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| Start with the chosen strain/organism and expose to general mutation; allow the survivors to grow on medium of choice (selective) Then take 200 colonies and suspend in sterile water (determines product traits); take best 50 and re-treat with mutations Continue mutations to obtain desired process traits and increase yield of desired compound |
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| Why are lactose and glucose both needed in the medium for P. chrysogenum? |
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| Glucose - creates little product, but a high amount of cell division (lots of biomass) Lactose - induces "stress state" in the organism; causes high production of antibiotic (due to stress response), but little biomass |
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| What is the typical length of P. chrysogenum fermentation? |
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| 6 days |
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| What is the purpose of starvation feeding? |
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| It is used to induce the stress response in bacteria in order to increase the amount of antibiotic produced (during stress situations, cell energy goes toward production of antibiotic) |
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| In the fermentation process of P. chrysogenum, what does most of the carbon supplied go towards? |
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| Most carbon supplied (in the form of sugar) goes toward the antibiotic production (due to stress response induced by lactose) |
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| What property of penicillins allow them to be concentrated and purified? |
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| Their carboxylic acid functionality; lower pH in solution below pKa of acid to get the undissociated species (neutral) This makes penicillin soluble in organic solvents, then once dissolved raise the pH to purify and repeat |
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| What type of metabolite is penicillin produced by P. chrysogenum considered to be? |
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| A secondary metabolite |
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| What is the concern with the overuse of antibiotics? |
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| Not all antibiotics are metabolized by the body, creating a selection pressure for antibiotic resistant organisms (selects for those with antibiotic resistance The more antibiotics used, the stronger the selection pressure - leads to development of more antibiotic resistant microbes |
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| What was the first commercial recombinant DNA product? |
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| Humulin - human insulin |
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| Pros vs. Cons of E. coli as a host to express recombined DNA |
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| Pros - fast and easy to grow, sequenced genome, many tools available Cons - can't glcosylate proteins, high oxygen demand, cannot secrete proteins, produce endotoxins |
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| Pros vs. Cons of B. subtilis as a host in expressing recombined DNA |
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| Pros - fast growing & well understood, available promoter regions, secretes proteins Cons - genetic instability, no good plasmids, secretes proteases which can degrade secreted proteins |
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| Pros vs. Cons of S. cerevisiae as a host to express recombinant DNA |
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| Pros - easy to grow & well understood, strong promoters available, eukaryotic - can be post-translationally modified (can activate proteins by glycosylation at a later time) Cons - grows slowly, poor rate of recombination |
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| Pros vs. Cons of Mammalian Cell as a host in expressing recombinant DNA |
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| Pros - natural protein products, protein can be secreted Cons - very slow growing, complex media formulation, can contain viruses, etc. |
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| What is a problem involved with using more complex organisms in a bioprocess? |
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| Involves formulation of a more complex medium |
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| How can you reduce the occurrence of plasmid shedding from a cell to make recombination more likely? |
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| Include an antibiotic resistance gene within the plasmid so that the cell gains a benefit by keeping the plasmid and not shedding it; otherwise, plasmid is seen as an energy burden to the cell and will likely be shedded |
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| Defined Medium |
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| Know all substances and concentrations of substances present in the medium |
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| Complex Medium |
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| Don't know all exact concentrations of substances in medium |
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| Casein Hydrolysate |
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| Complex source of N; hydrolyzed milk protein; can add as a general energy source if you don't know the exact metabolic needs of the cell |
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| Stirred-Tank Bioreactor |
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| Most common & typically used for bacteria (because they are robust and typically shear resistant) Need a cooling tank (because of exothermic reactions) Oxygen added at base to give means for energy for aerobic reaction |
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| Air-Lift Bioreactor |
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| NO mechanical agitation - used for shear-sensitive bacteria Air tube in middle section creates a convection current which gives mild to decent mixing of reaction components and no mechanical shear |
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| Disposable Wave Bioreactor |
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| Basically a giant plastic bag that rocks back and forth to form standing waves within it Waves provide nutrient mixing and oxygen transfer Typically used for mammalian cell cultures (low shear) |
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| Is the preferred product intracellular or extracellular? |
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| Want product to be extracellular because it is then easier to purify |
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| Formulation (Bioprocessing) |
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| Additional preservatives added to the compound which are not part of the active ingredient |
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| Acute Care vs. Public Health Medicine |
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| Acute Care - individually based; healthcare administered to the individual and emphasize on disease diagnosis; e.g. surgery Public Care - treat population as a whole; disease prevention for the who population |
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| Normal Flora |
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| Bacteria that are consistently associated with an animal/host; seen typically in people and are not pathogenic |
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| Which two normal flora are present nearly everywhere in the body? |
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| Staphylococci and Coryenbacterium |
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| What is the leading cause of bacterial disease in humans? |
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| Staphylococcus aureus |
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| S. aureus |
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| Potential pathogen; normal flora colonize the nares Colonizes compromised individuals to cause disease Infect wounds, blood, and middle ear |
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| MRSA |
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| Methicillin resistant S. aureus - antibiotic resistant strain; NOT normal; artificially induced resistance |
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| Streptococcus |
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| Found in the pharynx, mouth & intestines Example - Strep. mutans Forms OPPORTUNISTIC infections; forms plaques in the mouth and oral cavity |
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| E. coli |
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| Consistent resident of the small inestine (normal flora there) Causes - intestinal infection, UTIs, meningitis |
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| Lactobacilus |
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| Found in oral cavity & vagina Lowers the pH in the vagina to create an acidic environment to inhibit the growth of pathogens (provides defenses to the host) |
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| Benefits to the host of having normal flora: |
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| Flora synthesize and secrete vitamins - vitamin K and B12 from enteric bacteria Prevent colonization of pathogens - lactobacili in genitourinary tract Stimulate tissue development (lymphoid) - need pathogen exposure to build defense Stimulate production of cross-reactive antibodies |
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| Symbiosis |
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| The nature of the relationship between living things |
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| Mutualism |
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| A symbiotic relationship in which some reciprocal benefit accrues to both parties (+/+) E.g. ant, caterpillar, and plant - plant provides food for ant & caterpillar, ant protects the other two |
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| Commensalism |
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| A relationship in which one symbiont, the COMMENSAL, benefits, while the other, the HOST, is neither harmed or helped (+/0) E.g. Barnacles on whale; barnacles = commensal; whale = host |
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| Parasitism |
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| Relationship in which one of the two benefits, and the "host" is usually harmed (+/-) |
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| Pathogen |
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| Microorganism capable of causing disease |
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| Pathogenicity |
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| Ability of the microbe to cause disease |
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| What determines pathogenicity? |
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| Both host and microbe factors |
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| What is the difference between OPPORTUNISTIC and OBLIGATE pathogens? |
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| Opportunistic - always present in the host Obligate - only present in the host to cause disease (only present in disease state) |
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| 3 Factors that Pathogenicity is dependent upon |
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| The number of microorganisms infecting the host The condition of the host The virulence of the pathogen |
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| Virulence |
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| The DEGREE of the pathogenicity; dependent on the ability to invade and infect host |
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| Constituitive Host Defences |
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| Innate defenses; general protection that is inherent to the host Non-specific responses E.g. Skin, pH changes, lysozyme, mucus |
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| Inducible Host Defenses |
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| Need to be INDUCED by exposure to the pathogen (not normally present in host state) NOT immediate; usually quite specific (immune response) |
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| Toxigenesis |
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| Ability of the organism to produce toxins |
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| Difference between Endotoxin & Exotoxin |
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| Endotoxin - LPS; associated w/ gram negative cell wall Exotoxin - proteins released from bacterial cells that act at tissues in different locations |
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| DIFFERENCES between ENDO & EXO toxins |
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| ENDO - LPS, found on OM, NOT denatured by boiling, antigenic, low potency & specificity, no enzymatic activity EXO - protein secreted, denatured by boilding, antigenic, high potency & specificity, enzymatic activityl toxin is usually specific to the site of damage (e.g. neurotoxin) |
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| When are exotoxins secreted by bacteria? |
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| During the log phase of cell growth |
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| Modes of Action of Exotoxins: |
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| Damage CM - causes cells lysis and bacterial spread Inhibit protein synthesis Actiavte 2nd messengers - alter function without killing cell Inhibit neurotransmitter release Activate host immune response |
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| Effect of Diptheria exotoxin |
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| Acts on EF-2 (elongation factor) to inhibit host cell protein synthesis |
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| Effect of BoNT (botulism neurotoxin) |
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| Targets receptors on the presynaptic membranes of neurons in the PNS; causes no muscle contraction - flaccid paralysis |
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| Effect of TeNT (tetanus neurotoxin) |
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| Affects CNS to cause SPASTIC paralysis |
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| Waterborne Disease Characteristics & Prevalence |
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| Disease outbreaks rare in well engineered systems (more common in rural compared to urban Outbreaks associated with increase in rainfall/runoff (due to increased turbidity) Peak occurrence during Spring/Summer months (higher temperature in wells to allow for more bacteria growth) |
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| What was the initial precipitate which likely led to the Walkerton incident? |
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| Started with heavy rainfall before any reported cases; could cause increased turbidity in drinking water (more contaminants) |
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| What was the E. coli strain that contaminated the water in Walkerton? |
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| E. coli O157:h7 |
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| What is believed to be the primary reservoir of E. coli O157:H7? |
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| Cattle |
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| Difference between coliforms and E. coli when testing water? |
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| Coliforms - used as a general indicator that the water may be contaminated E. coli - direct fecal indicator |
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| Why does steak have a lower risk of E. coli contamination than ground beef? |
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| Because steak originates only from one animal; ground beef is a mixture of hundreds of cattle, thereby increasing the likelihood of contamination |
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| Examples of waterborne microbial pathogens? |
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| E. coli, Campylobacter, Salmonella, Shigella, viruses |
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| What is a ZOONOTIC organism? |
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| An organism that is transmissible from animals to humans |
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| Risk factors of waterborne disease? |
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| High animal density - E. coli infection is higher in rural areas of Canada with high cattle density Human exposure to animals Poor source drinking water |
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| Membrane Filtration |
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| Quantitative method used to test potable and non-potable water for the presence of indicator bacteria |
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| What are two indicators of contaminated water? |
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| E. coli and coliforms |
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| Coliforms |
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| Grow as pink colonies based on ability to ferment lactose NON-specific indicators of contamination (suggest general contamination) Can be from environmental sources or from feces |
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| E. coli (in water testing) |
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| Easy to detect Specific FECAL coliform - from human & animal waste Grows as bright purple colonies based on production of B-glucouronidase |
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| What is the use of B. fragilis in water testing? |
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| It is used as an indicator organism for contamination, and can be differentiated on a species level (bovine, human, neither) |
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| Giardiasis |
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| Most commonly identified waterborne pathogen in N. America Protozoa w/ cyst & trophozite forms; causes backpacker's diarrhea & beaver fever Best removed from water by slow sand filtration |
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| Meningococcus |
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| Gram-negative bacteria prevalent in institutional settings See increased rate of colonization in later winter/early spring Causes purpuric rash |
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| Measles |
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| Virus - Paramyxoviridae; morbilivirus (- ssRNA virus) Highly transmissible; have a known vaccine |
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| Incubational Period & Prodrome of Measles (Morbilivirus) |
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| Incubation = 6-18 days Prodromal Phase = 3-4 days; fever, catarrhal inflammation, Koplik spots in oral cavity (THIS is pathopneumotic, can use to directly diagnose) |
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| Culturing of Measles |
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| Can do throat/nasopharyngeal swab up to 4 days after rash onset Can do urine test up to 7 days after rash onset |
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| Mumps (Viral Classification) |
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| Paramyxoviridae, Rubulavirus (- ssRNA virus) |
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| Incubation Period and Prodrome for Mumps (Rubulavirus) |
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| Incubation period = 14-21 days Prodrome = non-specific symptoms; can be asymptomatic; common to present as low respiratory tract infection |
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| Shapes of Bacteria |
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| Coccus - round Bacillus - elongated (rod) Spirillum - spiraled Streptocci - chains Straphylococci - clumps |
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| Difference of Gram positives vs. negatives in the Gram stain (different colours)? |
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| Positive = stain a reddish colour (less dye retention) Negative = stain a purple colour |
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| Enrichment Media |
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| Media that has the addition of extracts to agar or broth to support growth of fastidious bacteria |
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| Characteristic Media |
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| Used to test bacteria for particular metabolic activities, products, or requirements |
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| What is the purpose of the Kirby Bauer test? |
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| To determine a given drug's efficacy in reducing microbial growth (susceptibility of bacteria to antibiotics) |
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| Influenza (Viral Classification) |
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| Orthomyxoviridade, Influenzavirus |
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| Differences in the Types of Influenzavirus |
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| Types A, B, C Type B - only infects humans Type A - infects humans, birds, and mammals |
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| Where does influenza replicate in humans vs. water fowl? |
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| Humans - in RTE Water fowl - in the GI tract epithelium |
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| Two types of viral proteins on the influenza envelope and their functions? |
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| Hemagglutinin (HA) - used for viral attachment & adsorption Neuraminidase (NA) - used for viral release |
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| Pandemic Flu |
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| Associated with antigenic SHIFT or emergence of recycled subtype Little/no herd immunity High attack rates & morbidity |
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| Seasonal Flu |
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| Antigenic drift of existing strains Often have a cross-reacting antibody for population Variable mortality/morbidity |
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| What is believed to be the natural reservoir of avian influenza A? |
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| Wild water fowl; virus believed to inhabit the GI tract of these animals |
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| Where is recombination believed to have occurred in the 2009 H1N1 outbreak? |
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| Believed to have occurred in pigs because they are susceptible to both avian and human strains of influenza; act as a mediator for recombination between human and avian strains |
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| Koch's Postulates |
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| Same pathogen present in every case Pathogen must be isolated from diseased host and grown in pure culture From pure culture, pathogen must be able to infect healthy organism Same organism needs to be present in newly infected organism |
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| Mayer's Work on TMV |
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| Got juice from infected leaves of plants and inoculated into healthy plants to cause disease transmission; unknown infecting agent |
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| Ivanofsky's Work on TMV |
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| Repeated Mayer's experiments, but added another step - took juice from plants and strained through bacterial filter Took filtrate and found that it could still infect healthy plants Provided an operational definition for viruses - non-bacterial infectious agents |
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| Chamberland Filter |
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| Used by Ivanofsky to discover "non-bacterial infectious agents"; made by Pasteur & Chamberland; made of unglazed porcelain and used as a bacterial strainer |
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| Beijerinck |
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| Collaborated w/ Mayer and found that the filtered sap from plants could be diluted and then regain its titer after transfer through living plants Knew the agent had to be able to grow in the plant |
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| What did Beijerinck name the organism involved in TMD? |
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| Called it a "soluble living germ" (contagium vivum fluidum) |
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| What was the operational concept of a virus at the beginning of the 20th century? |
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| An obligate intracellular parasite that is too small to observe in the light microscope, but can cause disease by multiplying in living cells |
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| Loeffler & Frosch |
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| Isolated 1st filterable agent from animals (foot & mouth disease; Picornaviridae) |
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| Reed et al |
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| 1st filterable agent from humans (Yellow fever; Flaviviridae) |
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| How was it determined that viruses were like proteins/contained proteins? |
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| Methods to purify proteins could also purify TMV TMV migrated in an electric field TMV could be neutralized by rabbit antibodies |
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| Schlesinger |
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| Concluded that bacteriophages were made of both protein AND DNA 1st suggestion that viruses were composed of nucleoprotein |
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| What is the largest virus known and how long is its genome? |
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| Mimicking microbe virus (Mimivirus); genome = 1.2 megabases |
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| d'Herelle |
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| Was working with shigella dysentery and noticed contamination of his plates by plaque formation Credited with development of the principles of virology |
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| Virological Techniques Developed by d'Herelle |
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| Used serial dilutions & plaque assay to determine the TITRE of the virus (quantify infectious units, PFUs); developed the viral plaque assay Showed the 1st step of viral infection had to be adsorption (virus present in only the pellet, not supernatant) See host range specificity with viruses (can be seen w/ centrifuge) Described cell lysis and virus release |
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| Reservoir of Mumps virus? |
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| Humans |
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| How is Mumps transmitted; what is its communicability (time period that it can be trasmitted? |
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| Transmitted through respiratory droplets Can range from 3 days before, to 9 days after symptom onset |
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| Diagnosis of Mumps |
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| Buccal swab done within 4-5 days of symptom onset Urine test done within 2 weeks of symptom onset |
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| Viral Plaque Assay |
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| Take OG virus samples and do ten fold dilutions; add 0.5 mL of each dilution to different plates w/ host cells Incubate plants then count plaques; as dilution increases, less plaques get formed |
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| What is the viral plaque assay used to calculate? |
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| Calculates the titre of the virus (concentration; PFU/mL) |
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| If susceptible bacterium is mixed w/ a solution of virus and then centrifuged, where will the virus be found? What is a non-susceptible bacterium is used instead? |
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| Susceptible - virus found in pellet because it is adsorbed to bacterium Non-susceptible - virus in supernatant because host-range specificity prevents binding to bacterium |
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| The Phage Group |
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| Founded by Delbruck & Luria Viewed bacteriophage/bacteria model as a way to understand cancer viruses & developmental biology Used phages as probes to understand how genetic information could determine biology |
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| What are relevant tissues in an embryonated egg in relation to virus replication/development? |
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| Allantoic cavity & chorioallantoic membrane |
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| Primary Cell Culture |
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| Cells have a finite life span in the culture; cells taken directly from the organism |
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| Continuous Cell Culture |
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| Cells are abnormal, often transformed; cells have been "immortalized" |
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| Cells produced from embryonic tissues are usually ________ cell lines. |
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| Primary cell lines (cells have a finite life span) |
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| Cells retrieved from carcinoma tissue are usually considered to be __________ cell lines. |
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| Continuous cell lines (because cells have been transformed into the malignant state; ability to continuously grow & divide) |
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| How are primary cell cultures generally prepared? |
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| Start with tissue sample from organism; digest sample by trypsinization (serine protease); plate cells and feed with synthetic liquid media |
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| What was the first viral vaccine produced in cell culture? |
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| Poliovirus vaccine (used monkey kidney cells) |
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| How are primary cell lines transformed into continuous cell lines? |
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| Start with primary cell culture and grow on medium; some cells die, others continue to grow; subculture the cells that have continued growth (treat w/ trypsin to avoid contact inhibition) During growth, some cells in primary cell line become mutagenized to cause transformation Because there is no selective pressure, they continue to grow and retain their mutations (cells become immortalized to form a continuous cell line) |
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| HeLa cells are an example of what type of cell line? Where were they originally cultured from? |
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| Example of a continuous cell line; cultured from malignant cells in a patient with cervical cancer |
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| What does the particle-to-PFU ratio measure? |
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| It is a measure of infectivity; a high ratio corresponds to a low infectivity (shows not all particles can successfully replicate) |
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| Difference between the count of viral PARTICLES vs. PFUs |
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| Particle - the physical virion PFU - the representation of infectious particles |
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| Difference between viral infection and replication. |
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| All viruses have the ability to infect a cell, but not all can successfully replicate within the cell Infection - association with host Replication - complete production of new virions |
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| Ways viruses can be quantified: |
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| Counting physical particles by EM (counts all particles) Genome copy # / PCR (quantitative; count number of virus particles) Hemagglutinin assay (counts number of virions, not infectivity) Plaque assay (measures infectivity of the virus) |
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| Subunit |
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| Single, folded polypeptide chain (gene product) |
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| Structure Unit (Protomer) |
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| Collection of one or more non-identical subunits which form building blocks of the larger assembly (e.g. VP1 of poliovirus) |
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| Assembly Unit |
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| Set of subunits OR structure units that is an intermediate in formation of a larger structure; usually symmetrical (e.g. hexamers & pentamers) |
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| Capsomere |
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| Morphological units seen on the surface of a particle via EM; seen as the morphological unit of an icosahedral capsid |
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| Capsid |
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| External protein shell around nucleic acid |
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| Core |
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| Internal portion; includes nucleic acid and closely associated proteins |
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| What type of interaction occurs between capsid structural units? |
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| NON-covalent bonds (protein protein interaction) Contact between individual subunits determines overall 3D shape |
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| When are arms and hinges important in the formation of a viral capsid? |
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| Arms and hinges only are important AFTER the individual subunits have come into contact; symmetry is caused by the repeated contact between identical subunits |
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| Types of interactions that can exist between protein capsids: |
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| Protein-protein, protein-nucleic acid, protein-lipid |
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| Helical Symmetry |
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| Rod-like structures; hollow cylinder As subunits assemble, the nucleic acid is packaged simultaneously (e.g. TMV) Examples - Influenza, TMV, Baculovirus |
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| Icosahedral Symmetry |
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| Spherical viruses; defined mathematically by their symmetry ALL have 20 triangular faces and 12 vertices Examples - Adenovirus, HSV-1, Poliovirus |
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| Complex/Mixed Symmetry |
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| Mixed elements E.g. Bacteriophages (icosahedral head & complex tail), Poxviruses (brick shaped) |
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| What codes for envelope proteins on virus envelopes? |
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| Viral genes |
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| Functions of HA and NA spikes on influenza |
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| HA = receptor binding & membrane fusion NA = secondary uncoating & transcriptase; enzymatic activity |
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| Mutations that cause year to year changes in Influenza typically occur in which proteins? |
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| HA and NA spikes on viral envelope |
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| Why is (+) ssRNA immediately infectious (can be transfected into cells)? |
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| Because it can be immediately translated into proteins |
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| Where do most DNA and RNA viruses replicate? Exceptions? |
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| DNA - most replicate in the nucleus; EXCEPTION - Poxviruses replicate in the cytoplasm RNA - most viruses replicate in the cytoplasm; EXCEPTION - Influenza replicates in the nucleus |
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| Taxonomic units of viral classification (broad to specific) |
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| Order -> Family -> Genus -> Species |
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| Two main schemes of viral classification? |
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| Classical System - based on shared physical properties Baltimore System - based on genomics (replication strategies of virus) |
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| What are the 4 characteristics used in the "Classical System of Classification" of Viruses? |
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| Nature of genetic material in virion Symmetry of capsid Naked vs. Enveloped Dimensions of Virus & Capsid |
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| What is the Baltimore Classification System based on? |
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| Based on the Central Dogma of molecular biology (basically on viral replication strategies) |
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| What is the one concept that is true for all viruses? |
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| ALL viruses must produce mRNA which can be translated by cellular ribosomes |
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| Parvoviridae |
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| linear ssDNA (Group II); non-enveloped, icosahedral particle Replicates in the nucleus NO enzymes in virion |
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| Polyomaviridae |
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| circular dsDNA (Group I); non-enveloped icosahedral capsid Replicates and assembles in nucleus Examples - Simian virus type 40 (SV40) |
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| Adenoviridae |
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| linear dsDNA (Group I); non-enveloped, icosahedral symmetry Have a fiber protein projecting from each vertex Replicates and assembles in nucleus |
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| Hepadnaviridae |
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| Circular, partially gapped dsDNA (Group VII); enveloped icosahedral capsid Contains viral DNA pol in the capsid; codes for RT |
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| Herpesviridae |
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| linear dsDNA (Group I); enveloped icosahedral capsid Replicates in the nucleus |
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| Baculoviridae |
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| circular dsDNA (Group I); helical enveloped virus Contains no polymerase in capsid Replicates and assembles in cytoplasm |
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| Poxviridae |
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| linear dsDNA, X linked (Group I); complex (brickshaped), enveloped capsid Contains DNA dependent RNA polymerase in capsid Replicates and assembles in CYTOPLASM |
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| Reoviridae |
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| Segmented, dsRNA (Group III); non-enveloped icosahedral capsid Replicates in the cytoplasm |
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| Picornaviridae |
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| (+) ssRNA (Group IV); non-enveloped, icosahedral capsid Replicates in the cytoplasm E.g. - Rhinovirus, Poliovirus |
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| Flaviviridae |
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| non-segmented (+) ssRNA virus (Group IV); enveloped, icosahedral capsid Replicates in the cytoplasm; receives envelope by budding from ER |
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| Retroviridae |
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| Diploid (+) ssRNA genome (Group VI); enveloped, icosahedral capsid Packaged RT and Integrase within the capsid Integration of viral dsDNA intermediate into host chromosome |
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| Coronaviridae |
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| Non-segmented (+) ssRNA virus (Group IV); enveloped, helical nucleocapsid Cytoplasmic replication; buds from ER & Golgi |
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| Rhabdoviridae |
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| non-segmented (-) ssRNA (Group V); enveloped, helical capsid (Bullet shaped) Cytoplasmic replication, buds from CM |
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| Orthomyxoviridae |
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| segmented (-) ssRNA (Group V); enveloped, helical nucleocapsid (HA and NA spikes) Replicates in the NUCLEUS; buds from CM |
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| Paramyxoviridae |
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| non-segmented (-) ssRNA (Group V); enveloped, helical nucleocapsid Replicates in the cytoplasm, buds from CM |
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| What is one structure/step in viral replication that no virus can code for? |
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| Ribosomal RNA; viruses must use host ribosomes as translational machinery |
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| Eclipse Period vs. Latent Period |
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| Eclipse - period between adsorption and appearance of first virions within the cell Latent - time between adsorption to cell and mature virion release from cell Eclipse = latent if the maturation of the virus coincides with its release |
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| Example of a virophage |
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| Sputnik Virophage; need host cell to be co-infected w/ Mimivirus - Has a circular dsDNA genome & icosahedral capsid |
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| Picornaviridae Adsorption (Poliovirus/Rhinovirus) |
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| Use canyon at the bottom of a surface depression that is accessible by receptor on host cell surface |
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| Adenovirus Adsorption |
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| Fibers extending from capsid (non-enveloped) act as anti-receptors |
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| Influenzavirus Adsorption |
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| HA + NA spikes on envelope; HA interacts with sialic acid residues to promote host cell binding |
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| Herpesvirus Adsorption |
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| HSV glycoproteins (gpB and C) interact with carbohydrate residues; get secondary reaction with antireceptor, membrane fusion and viral penetration |
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| HIV Adsorption |
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| gp120 & 41 are envelope glycoproteins which interact with CD-4 receptors on cell surface to cause conformational changes Fusion peptide on gp41 extends into CM |
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| 2 general pathways for penetration |
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| Receptor-Mediated Endocytosis - e.g. Influenzavirus Surface fusion with viral envelope - need fusion proteins in envelope for this to occur (e.g. Mumps, HSV, HIV) |
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| Poliovirus Penetration |
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| DIRECT translocation through CM (rare); or receptor-mediated endocytosis |
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| Different types of viral fusion proteins and viruses |
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| Influenza - conformational change in HA due to low pH in endosome triggers fusion HIV - binding of gp41/120 to CD-4 causes conformational change |
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| HIV Penetration |
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| Membrane fusion w/ envelope -> mediated by gp41 & gp120 fusion proteins w/ CD4 and CCR-5 receptors |
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| Influenza Penetration |
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| Receptor mediated endocytosis; once in endosome, low pH causes conformational change in HA spike to trigger membrane fusion and nucleic acid release |
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| Adenovirus Penetration |
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| Receptor med. endocytosis; low pH in endosome causes degradation and nucleic acid is released directly into nucleus |
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| General strategies for uncoating: |
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| HIV - uncoats at plasma membrane; membrane fusion releases nucleic acid into cell Influenza - uncoats in the endosome Adenovirus - uncoats at nuclear membrane, nucleic acid released into nucleus via nuclear pore |
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| Sites where capsid uncoating can occur |
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| At the plasma membrane (HIV), in the cytoplasm (Influenza), at the nuclear membrane (Adenovirus) |
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| Early Gene Products vs. Late Gene Products |
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| Early Gene Products = regulatory proteins (transcription factors) Late Gene Products = structural proteins Need expression of early genes to get late gene expression |
