Term
| What is the major change currently going on in microbiological biotech? What are the benefits of the change? |
|
Definition
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) |
|
|
Term
| Examples of biofuels used |
|
Definition
|
|
Term
| What is the most widely produced "bioproduct" in the world? |
|
Definition
|
|
Term
| Rationale for EtOH Fermentation |
|
Definition
Can oxygenate oil to improve combustion & reduce emission
Gasoline extender - import less oil (economic benefit)
Renewable transportation fuel |
|
|
Term
| What is the microbe used for ethanol fermentation? |
|
Definition
|
|
Term
| What is the benefit of a positive energy balance ratio? |
|
Definition
| Means that the product produces more energy than is used to make it |
|
|
Term
| What compound has the highest energy balance ratio and why? |
|
Definition
| Sugarcane has the highest ratio because the remnants from its fermentation are burned to act as feedstock for the distillation |
|
|
Term
| What is the problem with using biomass as a next-gen biofuel? |
|
Definition
| The presence of lignin bottlenecks the process; lignin coats the cellulose making it difficult to hydrolyze |
|
|
Term
| What are byproducts of EtOH fermentation and what are they used for? |
|
Definition
CO2 - sold to pop companies
DDG - protein from corn; feedstock for animals |
|
|
Term
| What is one of the limitations of S. cerevisiae regarding the sugars it can ferment? |
|
Definition
| Can only ferment 6C, up to the size of dimers |
|
|
Term
| What is end product inhibition in relation to ethanol fermentation? |
|
Definition
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 |
|
|
Term
| What is typically the highest % EtOH you can obtain via fermentation w/ S. cerevisiae? |
|
Definition
| Typically 10% EtOH/ 90% H2O because of the EPI that occurs |
|
|
Term
| Fermentation Conditions for S. cerevisiae |
|
Definition
Temperature = ambient (about 34 degrees C)
pH = 4-6 (slightly acidic)
Microaerobic conditions
Minimal additional nutrients need to be added
Highly exothermic reaction |
|
|
Term
| What is the typical EtOH yield in EtOH fermentation? |
|
Definition
|
|
Term
| EtOH Fermentation - Fermentation vs. Recovery |
|
Definition
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 |
|
|
Term
| Extractive Fermentation of EtOH (machine designed by Daugulis) |
|
Definition
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 |
|
|
Term
| Rationale for Biobutanol Use |
|
Definition
BuOH can be blended at higher concentrations w/ gasoline compared to EtOH
Higher energy content
Evaporates more slowly to give less emissions |
|
|
Term
| What is the organism used in Acetone-BuOH-EtOH (ABE) fermentation? |
|
Definition
| C. acetobutylicum (gram-positive obligate anaerobe) |
|
|
Term
| What is one of the problems with ABE fermentation? |
|
Definition
| Makes too many side-products |
|
|
Term
| Kinetics of ABE fermentation |
|
Definition
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) |
|
|
Term
| Yield of ABE fermentation? |
|
Definition
|
|
Term
| 3 main development phases in the formation of a biological process |
|
Definition
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 |
|
|
Term
| What types of antibiotics comprise the class of B-lactams? |
|
Definition
| Penicillins & Cephalosporins |
|
|
Term
| Why are almost all antibiotics made via biological processes? |
|
Definition
Typically there are no chemical processes that can be used instead
158/160 antibiotics being synthesized have only biological means of doing so |
|
|
Term
| General structure of B-lactams |
|
Definition
| All contain the same general structure - polyaromatics with the same central ring structure |
|
|
Term
| Difference in R group substitution between penicillins vs. cephalosporins |
|
Definition
Penicillins - 1 R group substitution possible
Cephalosporins - 2 R group substitutions |
|
|
Term
| What are semi-synthetic antibiotics? |
|
Definition
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 |
|
|
Term
| How can microbes be nudged into producing a certain type of antibiotic? |
|
Definition
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 |
|
|
Term
| What was the original species used to synthesize penicillin? What is the current species used? |
|
Definition
Old species = Penicillium notatum
Current = Penicillium chrysogenum |
|
|
Term
| Why was P. notatum not a good organism to be used for mass synthesis of penicillin? |
|
Definition
| Because it contained NO good process traits and could not be grown on liquid cultures (only grows on surfaces) |
|
|
Term
| Overview of Strain Development & Mutation |
|
Definition
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 |
|
|
Term
| Why are lactose and glucose both needed in the medium for P. chrysogenum? |
|
Definition
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 |
|
|
Term
| What is the typical length of P. chrysogenum fermentation? |
|
Definition
|
|
Term
| What is the purpose of starvation feeding? |
|
Definition
| 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) |
|
|
Term
| In the fermentation process of P. chrysogenum, what does most of the carbon supplied go towards? |
|
Definition
| Most carbon supplied (in the form of sugar) goes toward the antibiotic production (due to stress response induced by lactose) |
|
|
Term
| What property of penicillins allow them to be concentrated and purified? |
|
Definition
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 |
|
|
Term
| What type of metabolite is penicillin produced by P. chrysogenum considered to be? |
|
Definition
|
|
Term
| What is the concern with the overuse of antibiotics? |
|
Definition
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 |
|
|
Term
| What was the first commercial recombinant DNA product? |
|
Definition
|
|
Term
| Pros vs. Cons of E. coli as a host to express recombined DNA |
|
Definition
Pros - fast and easy to grow, sequenced genome, many tools available
Cons - can't glcosylate proteins, high oxygen demand, cannot secrete proteins, produce endotoxins |
|
|
Term
| Pros vs. Cons of B. subtilis as a host in expressing recombined DNA |
|
Definition
Pros - fast growing & well understood, available promoter regions, secretes proteins
Cons - genetic instability, no good plasmids, secretes proteases which can degrade secreted proteins |
|
|
Term
| Pros vs. Cons of S. cerevisiae as a host to express recombinant DNA |
|
Definition
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 |
|
|
Term
| Pros vs. Cons of Mammalian Cell as a host in expressing recombinant DNA |
|
Definition
Pros - natural protein products, protein can be secreted
Cons - very slow growing, complex media formulation, can contain viruses, etc. |
|
|
Term
| What is a problem involved with using more complex organisms in a bioprocess? |
|
Definition
| Involves formulation of a more complex medium |
|
|
Term
| How can you reduce the occurrence of plasmid shedding from a cell to make recombination more likely? |
|
Definition
| 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 |
|
|
Term
|
Definition
| Know all substances and concentrations of substances present in the medium |
|
|
Term
|
Definition
| Don't know all exact concentrations of substances in medium |
|
|
Term
|
Definition
| 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 |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
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 |
|
|
Term
| Disposable Wave Bioreactor |
|
Definition
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) |
|
|
Term
| Is the preferred product intracellular or extracellular? |
|
Definition
| Want product to be extracellular because it is then easier to purify |
|
|
Term
| Formulation (Bioprocessing) |
|
Definition
| Additional preservatives added to the compound which are not part of the active ingredient |
|
|
Term
| Acute Care vs. Public Health Medicine |
|
Definition
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 |
|
|
Term
|
Definition
| Bacteria that are consistently associated with an animal/host; seen typically in people and are not pathogenic |
|
|
Term
| Which two normal flora are present nearly everywhere in the body? |
|
Definition
| Staphylococci and Coryenbacterium |
|
|
Term
| What is the leading cause of bacterial disease in humans? |
|
Definition
|
|
Term
|
Definition
Potential pathogen; normal flora colonize the nares
Colonizes compromised individuals to cause disease
Infect wounds, blood, and middle ear |
|
|
Term
|
Definition
| Methicillin resistant S. aureus - antibiotic resistant strain; NOT normal; artificially induced resistance |
|
|
Term
|
Definition
Found in the pharynx, mouth & intestines
Example - Strep. mutans
Forms OPPORTUNISTIC infections; forms plaques in the mouth and oral cavity |
|
|
Term
|
Definition
Consistent resident of the small inestine (normal flora there)
Causes - intestinal infection, UTIs, meningitis |
|
|
Term
|
Definition
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) |
|
|
Term
| Benefits to the host of having normal flora: |
|
Definition
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 |
|
|
Term
|
Definition
| The nature of the relationship between living things |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
| Relationship in which one of the two benefits, and the "host" is usually harmed (+/-) |
|
|
Term
|
Definition
| Microorganism capable of causing disease |
|
|
Term
|
Definition
| Ability of the microbe to cause disease |
|
|
Term
| What determines pathogenicity? |
|
Definition
| Both host and microbe factors |
|
|
Term
| What is the difference between OPPORTUNISTIC and OBLIGATE pathogens? |
|
Definition
Opportunistic - always present in the host
Obligate - only present in the host to cause disease (only present in disease state) |
|
|
Term
| 3 Factors that Pathogenicity is dependent upon |
|
Definition
The number of microorganisms infecting the host
The condition of the host
The virulence of the pathogen |
|
|
Term
|
Definition
| The DEGREE of the pathogenicity; dependent on the ability to invade and infect host |
|
|
Term
| Constituitive Host Defences |
|
Definition
Innate defenses; general protection that is inherent to the host
Non-specific responses
E.g. Skin, pH changes, lysozyme, mucus |
|
|
Term
|
Definition
Need to be INDUCED by exposure to the pathogen (not normally present in host state)
NOT immediate; usually quite specific (immune response) |
|
|
Term
|
Definition
| Ability of the organism to produce toxins |
|
|
Term
| Difference between Endotoxin & Exotoxin |
|
Definition
Endotoxin - LPS; associated w/ gram negative cell wall
Exotoxin - proteins released from bacterial cells that act at tissues in different locations |
|
|
Term
| DIFFERENCES between ENDO & EXO toxins |
|
Definition
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) |
|
|
Term
| When are exotoxins secreted by bacteria? |
|
Definition
| During the log phase of cell growth |
|
|
Term
| Modes of Action of Exotoxins: |
|
Definition
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 |
|
|
Term
| Effect of Diptheria exotoxin |
|
Definition
| Acts on EF-2 (elongation factor) to inhibit host cell protein synthesis |
|
|
Term
| Effect of BoNT (botulism neurotoxin) |
|
Definition
| Targets receptors on the presynaptic membranes of neurons in the PNS; causes no muscle contraction - flaccid paralysis |
|
|
Term
| Effect of TeNT (tetanus neurotoxin) |
|
Definition
| Affects CNS to cause SPASTIC paralysis |
|
|
Term
| Waterborne Disease Characteristics & Prevalence |
|
Definition
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) |
|
|
Term
| What was the initial precipitate which likely led to the Walkerton incident? |
|
Definition
| Started with heavy rainfall before any reported cases; could cause increased turbidity in drinking water (more contaminants) |
|
|
Term
| What was the E. coli strain that contaminated the water in Walkerton? |
|
Definition
|
|
Term
| What is believed to be the primary reservoir of E. coli O157:H7? |
|
Definition
|
|
Term
| Difference between coliforms and E. coli when testing water? |
|
Definition
Coliforms - used as a general indicator that the water may be contaminated
E. coli - direct fecal indicator |
|
|
Term
| Why does steak have a lower risk of E. coli contamination than ground beef? |
|
Definition
| Because steak originates only from one animal; ground beef is a mixture of hundreds of cattle, thereby increasing the likelihood of contamination |
|
|
Term
| Examples of waterborne microbial pathogens? |
|
Definition
| E. coli, Campylobacter, Salmonella, Shigella, viruses |
|
|
Term
| What is a ZOONOTIC organism? |
|
Definition
| An organism that is transmissible from animals to humans |
|
|
Term
| Risk factors of waterborne disease? |
|
Definition
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 |
|
|
Term
|
Definition
| Quantitative method used to test potable and non-potable water for the presence of indicator bacteria |
|
|
Term
| What are two indicators of contaminated water? |
|
Definition
|
|
Term
|
Definition
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 |
|
|
Term
| E. coli (in water testing) |
|
Definition
Easy to detect
Specific FECAL coliform - from human & animal waste
Grows as bright purple colonies based on production of B-glucouronidase |
|
|
Term
| What is the use of B. fragilis in water testing? |
|
Definition
| It is used as an indicator organism for contamination, and can be differentiated on a species level (bovine, human, neither) |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
Gram-negative bacteria prevalent in institutional settings
See increased rate of colonization in later winter/early spring
Causes purpuric rash |
|
|
Term
|
Definition
Virus - Paramyxoviridae; morbilivirus (- ssRNA virus)
Highly transmissible; have a known vaccine |
|
|
Term
| Incubational Period & Prodrome of Measles (Morbilivirus) |
|
Definition
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) |
|
|
Term
|
Definition
Can do throat/nasopharyngeal swab up to 4 days after rash onset
Can do urine test up to 7 days after rash onset |
|
|
Term
| Mumps (Viral Classification) |
|
Definition
| Paramyxoviridae, Rubulavirus (- ssRNA virus) |
|
|
Term
| Incubation Period and Prodrome for Mumps (Rubulavirus) |
|
Definition
Incubation period = 14-21 days
Prodrome = non-specific symptoms; can be asymptomatic; common to present as low respiratory tract infection |
|
|
Term
|
Definition
Coccus - round
Bacillus - elongated (rod)
Spirillum - spiraled
Streptocci - chains
Straphylococci - clumps |
|
|
Term
| Difference of Gram positives vs. negatives in the Gram stain (different colours)? |
|
Definition
Positive = stain a reddish colour (less dye retention)
Negative = stain a purple colour |
|
|
Term
|
Definition
| Media that has the addition of extracts to agar or broth to support growth of fastidious bacteria |
|
|
Term
|
Definition
| Used to test bacteria for particular metabolic activities, products, or requirements |
|
|
Term
| What is the purpose of the Kirby Bauer test? |
|
Definition
| To determine a given drug's efficacy in reducing microbial growth (susceptibility of bacteria to antibiotics) |
|
|
Term
| Influenza (Viral Classification) |
|
Definition
| Orthomyxoviridade, Influenzavirus |
|
|
Term
| Differences in the Types of Influenzavirus |
|
Definition
Types A, B, C
Type B - only infects humans
Type A - infects humans, birds, and mammals |
|
|
Term
| Where does influenza replicate in humans vs. water fowl? |
|
Definition
Humans - in RTE
Water fowl - in the GI tract epithelium |
|
|
Term
| Two types of viral proteins on the influenza envelope and their functions? |
|
Definition
Hemagglutinin (HA) - used for viral attachment & adsorption
Neuraminidase (NA) - used for viral release |
|
|
Term
|
Definition
Associated with antigenic SHIFT or emergence of recycled subtype
Little/no herd immunity
High attack rates & morbidity |
|
|
Term
|
Definition
Antigenic drift of existing strains
Often have a cross-reacting antibody for population
Variable mortality/morbidity |
|
|
Term
| What is believed to be the natural reservoir of avian influenza A? |
|
Definition
| Wild water fowl; virus believed to inhabit the GI tract of these animals |
|
|
Term
| Where is recombination believed to have occurred in the 2009 H1N1 outbreak? |
|
Definition
| 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 |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
| Got juice from infected leaves of plants and inoculated into healthy plants to cause disease transmission; unknown infecting agent |
|
|
Term
|
Definition
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 |
|
|
Term
|
Definition
| Used by Ivanofsky to discover "non-bacterial infectious agents"; made by Pasteur & Chamberland; made of unglazed porcelain and used as a bacterial strainer |
|
|
Term
|
Definition
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 |
|
|
Term
| What did Beijerinck name the organism involved in TMD? |
|
Definition
| Called it a "soluble living germ" (contagium vivum fluidum) |
|
|
Term
| What was the operational concept of a virus at the beginning of the 20th century? |
|
Definition
| An obligate intracellular parasite that is too small to observe in the light microscope, but can cause disease by multiplying in living cells |
|
|
Term
|
Definition
| Isolated 1st filterable agent from animals (foot & mouth disease; Picornaviridae) |
|
|
Term
|
Definition
| 1st filterable agent from humans (Yellow fever; Flaviviridae) |
|
|
Term
| How was it determined that viruses were like proteins/contained proteins? |
|
Definition
Methods to purify proteins could also purify TMV
TMV migrated in an electric field
TMV could be neutralized by rabbit antibodies |
|
|
Term
|
Definition
Concluded that bacteriophages were made of both protein AND DNA
1st suggestion that viruses were composed of nucleoprotein |
|
|
Term
| What is the largest virus known and how long is its genome? |
|
Definition
| Mimicking microbe virus (Mimivirus); genome = 1.2 megabases |
|
|
Term
|
Definition
Was working with shigella dysentery and noticed contamination of his plates by plaque formation
Credited with development of the principles of virology |
|
|
Term
| Virological Techniques Developed by d'Herelle |
|
Definition
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 |
|
|
Term
| Reservoir of Mumps virus? |
|
Definition
|
|
Term
| How is Mumps transmitted; what is its communicability (time period that it can be trasmitted? |
|
Definition
Transmitted through respiratory droplets
Can range from 3 days before, to 9 days after symptom onset |
|
|
Term
|
Definition
Buccal swab done within 4-5 days of symptom onset
Urine test done within 2 weeks of symptom onset |
|
|
Term
|
Definition
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 |
|
|
Term
| What is the viral plaque assay used to calculate? |
|
Definition
| Calculates the titre of the virus (concentration; PFU/mL) |
|
|
Term
| 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? |
|
Definition
Susceptible - virus found in pellet because it is adsorbed to bacterium
Non-susceptible - virus in supernatant because host-range specificity prevents binding to bacterium |
|
|
Term
|
Definition
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 |
|
|
Term
| What are relevant tissues in an embryonated egg in relation to virus replication/development? |
|
Definition
| Allantoic cavity & chorioallantoic membrane |
|
|
Term
|
Definition
| Cells have a finite life span in the culture; cells taken directly from the organism |
|
|
Term
|
Definition
| Cells are abnormal, often transformed; cells have been "immortalized" |
|
|
Term
| Cells produced from embryonic tissues are usually ________ cell lines. |
|
Definition
| Primary cell lines (cells have a finite life span) |
|
|
Term
| Cells retrieved from carcinoma tissue are usually considered to be __________ cell lines. |
|
Definition
| Continuous cell lines (because cells have been transformed into the malignant state; ability to continuously grow & divide) |
|
|
Term
| How are primary cell cultures generally prepared? |
|
Definition
| Start with tissue sample from organism; digest sample by trypsinization (serine protease); plate cells and feed with synthetic liquid media |
|
|
Term
| What was the first viral vaccine produced in cell culture? |
|
Definition
| Poliovirus vaccine (used monkey kidney cells) |
|
|
Term
| How are primary cell lines transformed into continuous cell lines? |
|
Definition
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) |
|
|
Term
| HeLa cells are an example of what type of cell line? Where were they originally cultured from? |
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Definition
| Example of a continuous cell line; cultured from malignant cells in a patient with cervical cancer |
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Term
| What does the particle-to-PFU ratio measure? |
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Definition
| 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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Term
| Difference between the count of viral PARTICLES vs. PFUs |
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Definition
Particle - the physical virion
PFU - the representation of infectious particles |
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Term
| Difference between viral infection and replication. |
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Definition
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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Term
| Ways viruses can be quantified: |
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Definition
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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Term
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Definition
| Single, folded polypeptide chain (gene product) |
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Term
| Structure Unit (Protomer) |
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Definition
| 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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Term
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Definition
| 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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Term
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Definition
| Morphological units seen on the surface of a particle via EM; seen as the morphological unit of an icosahedral capsid |
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Term
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Definition
| External protein shell around nucleic acid |
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Term
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Definition
| Internal portion; includes nucleic acid and closely associated proteins |
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Term
| What type of interaction occurs between capsid structural units? |
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Definition
NON-covalent bonds (protein protein interaction)
Contact between individual subunits determines overall 3D shape |
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Term
| When are arms and hinges important in the formation of a viral capsid? |
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Definition
| 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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Term
| Types of interactions that can exist between protein capsids: |
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Definition
| Protein-protein, protein-nucleic acid, protein-lipid |
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Term
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Definition
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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Term
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Definition
Spherical viruses; defined mathematically by their symmetry
ALL have 20 triangular faces and 12 vertices
Examples - Adenovirus, HSV-1, Poliovirus |
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Term
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Definition
Mixed elements
E.g. Bacteriophages (icosahedral head & complex tail), Poxviruses (brick shaped) |
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Term
| What codes for envelope proteins on virus envelopes? |
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Definition
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Term
| Functions of HA and NA spikes on influenza |
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Definition
HA = receptor binding & membrane fusion
NA = secondary uncoating & transcriptase; enzymatic activity |
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Term
| Mutations that cause year to year changes in Influenza typically occur in which proteins? |
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Definition
| HA and NA spikes on viral envelope |
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Term
| Why is (+) ssRNA immediately infectious (can be transfected into cells)? |
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Definition
| Because it can be immediately translated into proteins |
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Term
| Where do most DNA and RNA viruses replicate? Exceptions? |
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Definition
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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Term
| Taxonomic units of viral classification (broad to specific) |
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Definition
| Order -> Family -> Genus -> Species |
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Term
| Two main schemes of viral classification? |
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Definition
Classical System - based on shared physical properties
Baltimore System - based on genomics (replication strategies of virus) |
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Term
| What are the 4 characteristics used in the "Classical System of Classification" of Viruses? |
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Definition
Nature of genetic material in virion
Symmetry of capsid
Naked vs. Enveloped
Dimensions of Virus & Capsid |
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Term
| What is the Baltimore Classification System based on? |
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Definition
| Based on the Central Dogma of molecular biology (basically on viral replication strategies) |
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Term
| What is the one concept that is true for all viruses? |
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Definition
| ALL viruses must produce mRNA which can be translated by cellular ribosomes |
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Term
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Definition
linear ssDNA (Group II); non-enveloped, icosahedral particle
Replicates in the nucleus
NO enzymes in virion |
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Term
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Definition
circular dsDNA (Group I); non-enveloped icosahedral capsid
Replicates and assembles in nucleus
Examples - Simian virus type 40 (SV40) |
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Term
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Definition
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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Term
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Definition
Circular, partially gapped dsDNA (Group VII); enveloped icosahedral capsid
Contains viral DNA pol in the capsid; codes for RT |
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Term
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Definition
linear dsDNA (Group I); enveloped icosahedral capsid
Replicates in the nucleus |
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Term
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Definition
circular dsDNA (Group I); helical enveloped virus
Contains no polymerase in capsid
Replicates and assembles in cytoplasm |
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Term
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Definition
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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Term
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Definition
Segmented, dsRNA (Group III); non-enveloped icosahedral capsid
Replicates in the cytoplasm |
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Term
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Definition
(+) ssRNA (Group IV); non-enveloped, icosahedral capsid
Replicates in the cytoplasm
E.g. - Rhinovirus, Poliovirus |
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Term
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Definition
non-segmented (+) ssRNA virus (Group IV); enveloped, icosahedral capsid
Replicates in the cytoplasm; receives envelope by budding from ER |
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Term
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Definition
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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Term
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Definition
Non-segmented (+) ssRNA virus (Group IV); enveloped, helical nucleocapsid
Cytoplasmic replication; buds from ER & Golgi |
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Term
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Definition
non-segmented (-) ssRNA (Group V); enveloped, helical capsid (Bullet shaped)
Cytoplasmic replication, buds from CM |
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Term
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Definition
segmented (-) ssRNA (Group V); enveloped, helical nucleocapsid (HA and NA spikes)
Replicates in the NUCLEUS; buds from CM |
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Term
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Definition
non-segmented (-) ssRNA (Group V); enveloped, helical nucleocapsid
Replicates in the cytoplasm, buds from CM |
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Term
| What is one structure/step in viral replication that no virus can code for? |
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Definition
| Ribosomal RNA; viruses must use host ribosomes as translational machinery |
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Term
| Eclipse Period vs. Latent Period |
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Definition
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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Term
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Definition
Sputnik Virophage; need host cell to be co-infected w/ Mimivirus
- Has a circular dsDNA genome & icosahedral capsid |
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Term
| Picornaviridae Adsorption (Poliovirus/Rhinovirus) |
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Definition
| Use canyon at the bottom of a surface depression that is accessible by receptor on host cell surface |
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Term
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Definition
| Fibers extending from capsid (non-enveloped) act as anti-receptors |
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Term
| Influenzavirus Adsorption |
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Definition
| HA + NA spikes on envelope; HA interacts with sialic acid residues to promote host cell binding |
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Term
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Definition
| HSV glycoproteins (gpB and C) interact with carbohydrate residues; get secondary reaction with antireceptor, membrane fusion and viral penetration |
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Term
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Definition
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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Term
| 2 general pathways for penetration |
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Definition
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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Term
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Definition
| DIRECT translocation through CM (rare); or receptor-mediated endocytosis |
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Term
| Different types of viral fusion proteins and viruses |
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Definition
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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Term
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Definition
| Membrane fusion w/ envelope -> mediated by gp41 & gp120 fusion proteins w/ CD4 and CCR-5 receptors |
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Term
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Definition
| 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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Term
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Definition
| Receptor med. endocytosis; low pH in endosome causes degradation and nucleic acid is released directly into nucleus |
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Term
| General strategies for uncoating: |
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Definition
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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Term
| Sites where capsid uncoating can occur |
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Definition
| At the plasma membrane (HIV), in the cytoplasm (Influenza), at the nuclear membrane (Adenovirus) |
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Term
| Early Gene Products vs. Late Gene Products |
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Definition
Early Gene Products = regulatory proteins (transcription factors)
Late Gene Products = structural proteins
Need expression of early genes to get late gene expression |
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