MicroBio-Mid#2

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What are the 2 methods used to kill bacteria?: 1)Physical methods:
-heat
-radiation
-filtration

2)Chemical methods:
-antiseptics/disinfectants
-antibiotics
How does heat kill bacteria?: -denatures proteins, DNA
-melts lipids
-moist heat is better than dry heat b/c of faster penetration and better at protein denaturation

*used because easy to perform*
What is pasteurization?: -mild heat used to reduce the number of bacter
-doesn't kill all bacyteria present and doesn't kill lspores
-delays spoilage because fewer bacteria in product
What is the safety margin used by canning industries?: -canning companies use heating conditions designed to kill 10E12 spores
-therefore all pathogenic organisms are killed
What to forms of radiation are commonly used in sterilization?: 1)UV light: Cross links thymines in DNA, causes lethal mutations
-but has poor penetration props, good for surface only

2)Ionizing radiation(X and Gamma rays):
-dislodges electrons
-good penetration
-commercial uses only
What is Filter Sterilization?: -mechanical removal of bacteria (no kiling)
-used for heat sensitive liquids and gasses
-used in pharmaceuticals, growth media additives, surgical gasses, beverages
What is the purpose of chemical control methods?: -to reduce bacterial numbers to safe levels
-complete sterility may or may not be achieved
-use of germicides
-intended for use on surfaces
What are 2 types of germicides?: 1)Antiseptics: for use on biological surfaces (eg: skin)

2)Disinfectants: for use on inanimate surfaces
What are some general features of germicides?: -more than 14000 different formulations from 300 diff chemicals
-act by denaturing proteins, nucleic acids or degrading lipids
What does the efficiency of germicides depend on?: -conc used
-time of exposure
-presence of organic material
Give 8 examples of disinfectants and antiseptics: 1)Phenols: -denatures proteins, disrupts membranes
-disinfectant (eg: lysol)

2)Alcohols: -dissolve lipids
-anitseptic

3)Heavy Metals: -denatures proteins by binding -SH and -OH groups
-antiseptic (eg:dandruff shampoo)

4)Peroxides: -oxidizes lipids, proteins
-antiseptics (eg:anti acne cream)

5)Halogens: -denatures proteins & nucleic acids
-antiseptic (eg:iodine)
-disinfectant (eg:chlorine)

6)Quaternary ammonium compounds: -Cationic detergents based on NH4+, disrupts membranes
-antiseptic (eg:mouthwashes)

7)Gaseous agents: -denatures proteins and DNA by crosslinking organic groups
-disinfectant (eg:ethylene oxide, propylene oxide)

8)Chemicals as Preservatives: -used on food products to prevent microbial growth
-disinfectants (eg:organic acids, nitrates)
What are some sources to antibiotics? 3 of them.: -most are natural metabolic byproducts of soil microbes

-some are compounds synthesized in labs

-some are semisynthetic (chemically converted natural compounds to form a new antibiotic)
What are 3 general characteritics of antibiotics?: 1)Selective Toxicity: -must kill bacteria without harming host
-expressed as therapeutic index (TI)

2)Cidal vs Static Effect: -Bacteriocidal=kills bacteria
-Bacteriostatic=inhibits growth

3)Spectrum of Activity:
-narrow vs broad spectrum
What is the therapeutic index(TI)?: Dose which is toxic to host/Dose required to kill bacteria
What are the 5 ways that antibiotics work?: 1)Block DNA/RNA Synthesis
-eg rigamycin

2)Block protein synthesis
-eg erythromycin

3)Competitive metabolic inhibitors
-eg sulfonilamide

4)Disrupts cell membrane function
-eg polymyxin

5)Blocks peptidoglycan synthesis
-eg penicillin
What are the principles of combination antibiotics? 2 of them: -wants to avoid antagonism (one drug preventing the action of a second drug)

-wants to achieve synergism (enhanced effects vs either drug alone
What are some side effects to antibiotic use?: 1)Toxic to host

2)Allergic reactions (eg: penicillin)

3)Disruption of normal flora (eg: messes up gut boys and causes diarrhea)
What is bacterial antibiotic resistance?: -when a normally senstive bacteria is no longer affected by antibiotic
How do we know if bacteria are resistant?: 1)in vivo- patient is unresponsive to antibiotic therapy

2)in vitro- labs test for growth of bacteria in presence of the antibiotic
-determines: how resistant, correct antibiotic and proper dosage
What are 2 ways of measuring resistance?: 1)Disk Diffusion Method:
-put disc of antibiotic on infected plate, measure growth of inhibition

2)Tube Dilution Method
-if growth occurs at high antibiotic concentration, then bacteria is resistant
-can find Minimal Inhibitory Conc (prevent growth)
-can find Minimal Bactericidal Conc (kills bacteria)
What are 3 mechanisms used to become resistant to antibiotic?: 1)Direct inactivation of antibiotic
-bacteria produces enzymes which break down antibiotic

2)Prevent uptake or promote excretion of anitbiotic

3)Alteration of antibiotic target:
a)overproduction of target overwhemls antibiotic
b)mutated target no longer recognized by antibiotic
How is resistance acquired by bacteria? 2 of them: 1)Spontaneous mutation in DNA (gives an altered target)
-low freq

2)Acquire new resistance gene (gene for inactivating enzyme)
-plasmids may carry multiple resistance genes, passed on from bac. to bac.
What factors lead to the spread of resistance? 3 of them: 1)Clinical overuse of antibiotics

2)Lack of patient compliance (ie:failure to take drug as prescribed). Therefore not all bac. killed and survivors develop reistance.

3)Widespread nontherapeutic use of antibiotics:
-gut basteria constantly exposed to antibiotics
-develops reservoir of resistant bacteria

**Key Points:
-if antibiotics are continously in enviro, then bacteria is constantly exposed, therefor continual selective pressure for resistant strains**
What would you do to limit the developement of resistance? 5 of them: -promote responsible clinical use of antibiotics
-restrict use of some antibiotics in human and animal health
-rotate use of different types of antibiotics
-use combo antibiotics to slow devel of resistance
-better public education in responsible antibiotic use
What are some new sources of antibiotics?: 1)New soil, water bacteria and fungi continuously being screend to find undiscovered antibiotic

2)Rejuvenate existing antibiotics: may be lethal to bac again

3)Novel nonmicrobial sources of antibiotics

4)Rational drug design
-pick specific bacteria and target it
-make a chem which kills it
-takes about 10 years to do
What is a virus?: -a non cellular infectious agent
-nucleic acid usually surrounded by a protein coat
-metabolically inert
-reproduce only within a living host cell
What are some key ways in which viruses differ from liveing cells? 5 of them: -much smaller in size
-simple (non-cellular) structure and organization
-contains either DNA or RNA, but never both
-not capable of independent reproduction
-2 distinct phases in life cycle (depends on whether inside or outside of the cell)
What is host range?: -range of organism used by virus as host
-viruses have host specificity
-host specificity determined by receptors on host cell and presence of suitable metabolic machinery in host cell
What is the structure of viruses?: a)has many types of nucleic acid
-ds DNA
-ss DNA
-ds RNA
-ss RNA ('plus' orientation = mRNA)
-ss RNA ('minus' orientation = anit mRNA)

***nucelic acid can be circular or linear
virus will ONLY HAVE ONE TYPE of nucleic acid***

b)Has 3 types of protein coats:
1-Helical
2-Icosahedral
3-Complex

c)Presence or absence of envelope
What is a capsomer? What is a capsid?: -indiviual protein subunits which comince to make a complete shell for viruses
What are the 3 basic shapes of protein coats for viruses?: 1)Helical: capsomeres coiled in rod shaped spiral form

2)Icosahedral: capsomers in a geometric conficuration of 12 corners and 20 faces ("geodesic dome")

3)Complex: helical 'tail' attached to icosahedral head (eg: bacteriophage)
What's the difference between a naked and and enveloped virus?: Naked virus:
-NA core enclosed only by capsid
-capsid may contain spikes (helps bind to receptors)

Enveloped virus:
-core + capsid + outer lipid envelope
-spikes associated with envelope
-parts of envelope are teaken from host cell plasma membrane as virus exits from host cell
What 3 factors are taken into account when classifying viruses?: 1)Type of nucleic acid

2)Type of protein coat

3)Presence or absence of envelope
Go over the life cycle of viruses.: 1)Adsorption: virion attaches to host cell

2)Penetration: intact virion (or it's NA) enters the cell

3)Uncoating: release of viral NA from capsid (skipped if NA alone penetrates)

4)Gene Expression: viral genes expressed to produce more viral NA and viral proteins

5)Assembly: new intact viruses assemble inside host cell

6)Release: virus particles exit host cell
What is a virion?: -a virus outside of the host cell
What is Lytic infection?: -lysis of bacterial host cell and release of new virion
What is lysogenic infection?: After attachment and penetration:
-vireus genome recombines into bacterial chromosome (prophage)
-phage genome replicates as part of chromosome
-all bacterial daughter cells carry virus in prophage state

**prophage may bust out of chromosome and enter lytic pathway**
What are the 2 possible outcomes of bacteriophage infection?: 1)Lytic infection

2)Lysogenic infection
T/F Some viruses can act as antibiotics.: T
-viruses attack bacteria
-problem is controlling the virus afterwards
What steps are involved in adsorption?: -requires receptors on eucaryotic cell membrane
-receptors are recognized by viral spikes (proteins)
-spike+receptor interaction determines virus host rand and tissue specifictiy
-blocking receptor+spike interaction may prevent infection (eg: antibodies)
What 3 mechanisms are involved in Penetration?: 1)Direct penetration through host membrane by naked viruses

2)Fusion of virion envelope with host cell, parts of viral envelope left behind by enveloped viruses

3)Endocytosis: uses existing phagocytic mech of cell (enveloped viruses)
What steps are involved in uncoating?: -exposes virus nucleic acid to hosts biochemical machinery
-can occur during penetration or after penetration
-usually due to host hydrolytic enzymes which degrade capsid
What steps are involved in gene expression?: -viral nucleic acid is transcribed, translated, replicated
-usually completely dependent on host cell enzymes
-can be very complex, depending on type of NA in virion
What is the difference between 'early' and 'late' genes of the viral genome?: -Early genes-> transcribed immediately after entry into cell

-Late genes-> transcribed only after new viral RNA polymer is made
What steps are involved in assembly?: -building of new intact virus from component parts
-process varies depending on type of virus
>some spontaneous self assembly of capsid proteins
>sometimes requires enzymes from virus and/or host
>may occur in cytoplasm or nucleus
-poorly understood process for animal viruses
What steps are involved in release?: A)For Naked Viruses:
-virus particles assembled and accumulate in cytoplasm
-host cells biosynthesis stops
-host cells die and lyses
-virions are released

B)For Enveloped Viruses:
-virus glycoproteins are inserted into the inside of host cell membrane
-assembled virus capsid begins to bud out through the membrane
-envelope aquired while budding
*Budding may or may not kill the cell
*if host cell survives, may continue to shed virus
*some viral protinds may be left behind on cell surface, cell is killed by immune system
What are 4 possible outcomes of viral infections?: 1)Lytic Infection:
-host cell is killed as virions are released

2)Peresistent (Chronic) Infection:
-host cell survives
-virus shed for a long period of time
-infectious virions constantly present in patient

3)Latent Infection:
-host cell survives
-viral genome replicates as part of host chromosome
-very limited ecpression of viral genes
-no mature infectious virus released
-viral gene expression may be triggered at future time

4)Host Cell Transformation:
-formation of cancerous host cells by tumor viruses
What are 5 problems with antiviral agents?: 1) Side effect & toxicity to host cells: -viral life cycle is closely linked to that of the host cell
-it is hard to affect one without affecting the other
-many viruses still cannot be treated with anti-viral agents

2)Latent viruses may not be susceptible

3)Many viruses have long incubation periods

4)Resistance to some antivirals are developing

5)Developement of new agents is difficult
What are prions?: -infectious agents consisting only of protein
-an abnormally folded variant of a natural protein found in neurons
-causes slow diseases (ie: mad cow)
Whats the difference between PrP(C) and PrP(SC)?: -PrP(C) is a good prion (protease sensitive, does not aggregate)
-PrP(SC) is a bad prion (protease resistant, readily aggregates). Becomes spongy masses and clogs neurological system (causes mad cow)

-PrP(SC) can convert PrP(C) to PrP(SC) if they come in direct contact with eachother
What are some problems faced when diagnosing viral infections?: -symptoms of some viral disease mimic other microgbial diseases
-different types of viruses may produce similar symptoms
-diagnostic methods based on growth are more difficult for viruses than bac.
What is serology?: -looks for antibodies produced against viruses in patients blood
-used to diagnose viral infection

-can't tell diff b/w past and current infections
-might take long for antibodies to be made
What 3 methods are used to diagnose viral infections?: 1)Serology

2)Look for virus infected cells in host. Done adding antibodies tagged with fluorescent dye

3)Try to grow and identify virus directly from host.
-Requires use of Tissue Culture Cells (TCC)
-Add virus to TCC and allow infection
-look microscopically for ctyopathic effect on cells
-virus will cause change in morphology
>cell lysis
>rounding, fusing of cells
>inclusions inside cells

*slow process*
How do viruses cause symptoms of disease? 4 of them.: 1)Virus replication leads to host cell lysis

2)Virus infection alters host cell metabolism

3)Viral glycoproteins (spikes) insert into host cell membrane

4)Production of 'virokines' during virus infection
T/F Viruses often stay at the site of first contact with the host.: F
-Viruses spread via blood or lymphatic system
-may infect cells at other sites as long as suitable cell receptors are present
T/F Viruses can infect almost every body organ: T
-these bastards are versitile
What viruses are responsible for causing diarrhea?: -Enteric viruses (eg: Rotavirus, Norwalk virus, Adenovirus, Astrovirus)

-infected due to ingestion of infected food and water
What is Rotavirus?: -naked, dsRNA, double layered capsid (acid resistant)
-infects villi
-leads to lysis which causes fever, vominting, diarrhea
-up to 10E9 virion/ml of feces released
-symptoms last approx 1wk
-affects young kids, 90% of adults have antibodies against rotavirus
What is Hepatits C Virus (HCV)?: -a blood borne virus
-inflames the liver
-causes: weight loss, fever, jaundice, fatigue, muscle and joint pain, loss of lever function, death

-Enveloped, ssRNA
-fast replication
-infected by blood or body fluids only!!!
-20% of acute patients eliminate virus w/o further probs with treatment
-80% acute of patients enter chronic state (infection persists for >6mths)
What is Herpes Simplex Virus (HSV)?: -large, enveloped, dsDNA virus
-a nervous system virus
-has a huge host range
-infection begins at breaks in mucosal surfaces or skin (ie: needs contact with infected animal)
-HSV enters epithelial cells (causing fluid filled blisters)
-HSV may spread to neurons causing Latent infections:
>analogous to 'prophage' state
>little viral gene expression
>no damage to host cell
What are the 5 types of Herpes infections?: 1)Oral herpes (cold sores) - HSV-1

2)Herpes keratitis - infects eyes

3)Herpes whitlow - fingers

4)Genital Herpes - (HSV-2)
-point of entry is dink or vagi
>vesicular lesions occur (itch, pain, discharge)
>first infection, get flu like symptoms
>virus becomes latent, can be reactivated whenever

5)Herpes encephalitis - migration of HSV to brain. F's up behavior, seizure, fever, nausea
How do you control Herpes Simplex Virus (HSV)?: -acyclovir: not a cure, shortens duration of active leions

-avoid contact with active lesions

-can kill with a good soap wash. Kills lipid envelope
What is West Nile Virus(WNV)?: -a virus of the nervous system
-enveloped, ssRNA
-transmitted by mosquitoes, breast milk, organ transplants, blood transfusion
-may cause fever, headache, rash, swollen glands
-lasts 3 to 4 days, no long term effects

-patients >50yrs are at risk of developing encephalitis
>inflames brain
>headache, high fever, disorientation, stiff neck, muscle aches, paralysis, convulsions
>symptoms last for weeks then subside

*no real vaccine
What is the influenze virus?: -a respiratory tract virus
-enveloped, ssRNA, segmented RNA(8 pieces)

2 types of spike proteins on envelope
1)Neuraminidase(N):
-degrades mucus that lines epithelial cells or resp. tract
-required for release of mature virion from host cell
2)Hemagglutinin(H):
-for attachment
Go over the life cycle of the influenza virus.: -attaches to epithelial cell receptos
-virion envelope fuses with host membrane
-releases ssRNA into cytoplasm and migrates to the nucleus
-assembly of mature virion in cytoplasm
-released by budding

**transmitted by aerosols or hand to nose contact**
**damages host cells and kills them**
What are the 3 types of influenza virus?: Type A:
-most common
-broad host range (humans, birds, pigs)
-can cause pandemics(world wide spread)

Type B,C:
-infect only humans
-don't cause pandemics(small outbreaks)
-usually milder symptoms
How do different flu strains arise? 2 mechanisms.: 1)Antigenic Drift:
-spontaneous mutations in H or N genes which give rise to minor changes in H or N proteins

2)Antigenic Shift:
-completely new H and/or N protein acquired from another flu virus
-possible becusae of segmented genome
What is the ebola virus?: -only found naturally on African continent
-Enveloped; 1 ssRNA with 12 genes
-broad host range
-causes viral hemorrhagic fever
-virion enters directly into blood or inhaled
-tend to hang out in blood, lungs, and nasal secretion

*2 days of infection: high fever, headache, fatigue, bloody diarrhea
*7-10 days of infection: mucosal bleeding, circ system collapse, shock, blindness, death
What do we know about Ebola's transmission? 3 of them: 1)Zoonotic disease:
-animal to human transmission

2)Human to human transmission:
-via physical contact with secretions, blood of patients
-airborne spread possible but rare

3)International outbreaks are possible (ie: beyond Africa)
What hypothesis were made regarding the genesis of viruses? 3 of them.: 1)Retrograde evolution:
-cells which have continually lost funstion so as to become completely dependent on another host

2)Cellular origin
-combos of cellula macromolecules (protein, NA) that gained ability to self assemble and replicate

3)Parallel evolution of cells and viruses when life first began

**No strong evidence for any of the above**
Why are viruses important from a scientific viewpoint? 3 reasons: -models for gene organization and expression

-used as vectors for introducing new genes into animal cells

-used as possible agents of human gene evolution

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