Final Exam Concept Review Microbiology

1. What is a virus? Which structures are part of every virus? Which structures are found in some but not all viruses?

➢ Viruses

-are everywhere

-shape evolution

-can cause disease

-smaller than bacteria (require electron microscopy to be seen)

-require a host cell for their replication

--some have strange schemes for nucleic acid synthesis

➢Structures Part of Every Virus:

-genome: DNA or RNA; double- or single-stranded

-capsid: protein coat

--capsomers: individual subunits that comprise the capsid

--capsomers self-assemble into a capsid

➢ Structures Some Viruses Have:

-envelope: lipid bilayer derived from host cell membrane during viral budding

--virtually all bacteriophage are naked; animal viruses can be enveloped or naked

-spikes: viral proteins or glycoproteins in the envelope

-enzymes: for replication or pathogenesis

2. How are viruses different from bacteria? Contrast their size, structure, and how they reproduce.

Viruses vs. Bacteria

➢ Bacteria:

-living organism; unicellular; one cell

-larger (1000 nm)

-fission: a form of asexual reproduction

-cell wall: peptidoglycan or lipopolysaccharide

-has ribosomes

-DNA and RNA floating freely in cytoplasm

-"localized" infection

-usually treated w/ antibiotics

➢Viruses:

-not living; no cells

-smaller (20-400 nm)

-reproduction: invades a host cell, taking control and copies the viral DNA/RNA, then destroys the host cell and releases new viruses

-No cell wall; has protein coat (capsid)

-No ribosomes

-DNA or RNA enclosed inside a coat of protein

-"systemic" infection

-antibiotics will not effect the disease

3. How does a virulent phage replicate? Describe what occurs during each step in the phage lytic replication cycle.

1. Attachment (adsorption): the virus recognizes and binds to a host cell via a receptor molecule on the cell surface

-a specific ligand on the capsid (or spike) of the virus physically "sticks" on the membrane of the host cell

-this molecule, called a receptor, is usually a protein.

-a virus recognizes its host cells based on the receptors they carry, and a cell without receptors for a virus can't be infected by that virus

2. Penetration (entry): the virus or its genetic material enters the cell--release of the nucleic acid from the capsid

-one typical route for viral entry is fusion w/ the membrane, which is most common in Animal Viruses with envelopes

-Animal Viruses may also trick the cell into taking them in by bulk transport process called Endocytosis

-Phage usually inject their genome into the bacterial cell, leaving capsid outside

3. Synthesis: many variations of synthesis, depending on the viral genome structure (viral genomes can be DNA or RNA; single or double stranded; + or - strand ssRNA, so there's some variations in how their genomes have to be synthesized)

-dsDNA and ssDNA viruses: can use host enzymes to make new viral genomes

--the virus had a DNA genome, which can serve as a template for the enzyme DNA polymerase to use to make more DNA

-dsRNA and ssRNA viruses: need RNA replicase enzyme (RNA-dependent RNA polymerase) to make new viral genomes

--if the virus has an RNA genome, then it can use RNA replicase enzyme to synthesize more RNA

--RNA replicase is an RNA-dependent RNA polymerase, meaning it uses RNA as its template

-Retrovirus: + strand ssRNA genome but uses a dsDNA intermediate

--carries the enzymes reverse transcriptase and integrase

--DNA intermediate integrates into host genome

--Retroviruses have an RNA genome

--the whole aim of viruses is to make more virus particle

4. Compare and contrast the steps in the phage lytic replication cycle to the steps in an animal virus's replication cycle.

➢ Attachment Sites

-Bacteriophage: attachment of tail fibers to cell wall proteins

-Animal Virus: attachment of spikes, capsid, or envelope to plasma membrane protein

➢ Penetration

-Bacteriophage: injection of viral nucleic acid through bacterial cell wall

-Animal Virus: endocytosis or fusion

➢ Uncoating

-Bacteriophage: none needed

-Animal Virus: enzymatic digestion of viral proteins

➢ Synthesis

-Bacteriophage: In cytoplasm. Bacterial synthesis ceased. Viral DNA or RNA replicated, formation of viral mRNA. Viral components synthesized.

-Animal Virus: In cytoplasm (RNA) or nucleus (DNA). Host cell synthesis ceased. Viral DNA or RNA replicated, formation of viral RNA. Viral components synthesized.

➢ Maturation

-Bacteriophage: addition of collar, sheath, base plate, and tail fibers to viral nucleic acid-containing head

-Animal Virus: insertion of viral nucleic acid into capsid

➢ Release

-Bacteriophage: host cell lysis

-Animal Virus: Budding (enveloped viruses), cell rupture (non enveloped viruses)

➢ Chronic Infection

-Bacteriophage: lysogeny

-Animal Virus: Latency, chronic infection, cancer

5. Which viruses can rely on the host cell to supply the enzyme that replicates the viral genome? Which viruses cannot rely on the host cell to supply the enzyme that replicates the viral genome? Why?

-double stranded DNA and single stranded DNA viruses can use the host enzyme s to make new viral genomes b/c the DNA polymerase can use the DNA template to make more DNA

-double stranded RNA and single stranded RNA viruses need RNA replicase enzyme (RNA dependent RNA polymerase) to make new viral genomes. RNA replicase enzyme MUST be carried in the virion in a negative (-) strand RNA virus

6. How are "early viral proteins" different from "late viral proteins"? Give examples of each.

-Early Proteins: hijack the cell; force the synthesis of new viral genomes and viral coats (ex: sigma factor)

-Late Proteins: viral proteins that are part of the mature virion (ex: RNA replicase)

7. Define: "temperate phage" and "virulent phage". How do they differ?

Main difference: replication pathways

-Virulent phage is a lytic phage than can only use the lytic replication pathway (attachment-release process)

-Temperate phage can use both lytic and lysogenic replication pathways.

8. How is lysogenic replication different from lytic replication? Compare how the viral genome is replicated, what happens to viral gene expression, and what happens to the bacterial host.

Definition:

-Lysogenic replication: instead of phage continuing to the pathway of lysis (lytic), it enters the state of lysogeny, where it can remain for as long as it takes for it to continue. It does not necessarily lyse the cell. --> prophage

Gene expression: most phage genes are not expressed and phage genome is replicated in correspondence to bacterial genome

Bacterial host cell: not killed/lysed

9. Which protein is required to maintain lysogeny for lambda phage? How does phage lambda choose between lytic and lysogenic replication? Define "viral induction." What occurs during prophage induction?

-Protein needed to maintain lysogeny: lambda repressor protein (CI)

-Lambda decides btwn lytic and lysogenic pathways mostly based on environmental conditions

--ex: media's glucose concentration: Hfl protease can degrade the lambda repressor (lambda repressor protein is essential for lysogeny), so if there's a high glucose concnetrations, then cAMP concentration decreases, thus leading to more degradation. of lambda repressor

-Prophage induction: when bacteria is under stressful environmental conditions, such as UV radiation as it causes damage to DNA

10. True or false: A lysogenic phage has no effect on its host cell. Explain, describing the concept of "lysogenic conversion."

False.

-prophage has the ability of conferring new properties to bacterial genome --> Lysogenic conversion

--ex: Cholera, which conferred toxin genes w/ pathogenicity to harmless bacterium, making them disease-oriented

---discovered by epidemiologist John Snow

Informative Slide: Lysogenic Cycle

Lysogenic cycle allows phage to reproduce w/out killing its host (lytic=lysis)

-phage lambda can switch btwn lytic and lysogenic cycle

Pathway

1. Attachment: same as Lytic cycle

2. Penetration (entry): same as Lytic cycle

3. Integration: once the phage DNA is inside the cell, it is not immediately copied/expressed to make useful proteins

-it recombines with a bacterial chromosome's region

-thus causing the phage DNA to be integrated into the chromosome

-circularization--so not degraded

-single crossover event btwn attP and attB catalyzed by pahge-encoded integrase

-Maintenance of Lysogeny:

--the lambda repressor protein (CI) is essential for lysogeny maintenance

--the prophage is maintained w/ most of its genes, such as genes for replication, excision, silenced by the lambda repressor

--the lambda repressor is a phage-encoded protein that represses the transcription of most phage genes except itself

4. Cell Division: continued division makes many cells w/ the prophage

-Prophage: the integrated phage DNA; it's not active, genes are not expressed, and doesn't drive production of new phages

-Phage can switch back to Lytic cycle --> Excision:

--Prophage Induction: occurs due to environmental conditions that are harmful to the bacteria

--for example, exposure of a lysogen to UV light causes DNA damage, which initiates the SOS response, which activates the protease activity of recA that degrades the lambda repressor.

Consequences of Lysogeny

-Specialized Transduction: occurs due to the inexact excision of the prophage that allows the phage to transfer bacterial genes from 1 bacterium to another

-Lysogenic conversion: the prophage alters the genome of the bacterium, conferring new properties

--ex: many prophages contain toxin genes which confer pathogenicity to an otherwise harmless bacterium

11. Describe the 4 different outcomes of infection of an animal cell by an animal virus. How do these differ from the outcomes of infection of a bacterial cell by phage?

-Latency is a characteristic of the herpesviruses, which include Herpes simple viruses, chickenpox virus (varicella), and the virus that causes mononucleosis (Epstein-Barr)

-Latent infection: virus present but not causing harm to cell; later emerges in lytic infection --> long term chronic infection

--the viral genome becomes an episome

--the expression of genes involved in lytic replication is suppressed

--reactivation can occur due to stress or immunosuppression

12. Describe the steps in the replication of the 2009 A(H1N1) influenza virus. What are the roles of the HA, NA, and PB proteins in the spread of the virus? Why is this virus considered a "quadruple reassortant"?

Roles of proteins in virus

-HA (hemagglutinin): binds to sialic acids on cells of respiratory tract; also plays an important role during virus uncoating in the endosome

-NA (neuraminidase): cleaves the sialic acid residues; role in release/shedding of virion during budding

-PB1 (viral RNA polymerase/RNA replicase): RNA replicase is only present in the virus's virion; it is an RNA dependent RNA Polymerase that uses the RNA as a template to make DNA.

-the 2009 A(H1N1) is considered a Quadruple Reassortment b/c it has 4 distinct sources of the gene segments in the virus

--Segments: Avianlike, Humanlike, Swinelike, & Eurasian like

13. Describe the steps in the replication of Coronaviruses. What are two pathways for Coronavirus fusion? What is the relationship between TMPRSS2 expression and hydroxychloroquine effectiveness as an antiviral against SARS-CoV2? Explain.

Replication of Coronaviruses

-virion binds cell surface receptor ACE2

-TMPRSS2 protease on host cell primes virus spike protein and permits entry via ACE2 binding

-depending on host cell type and maturation status of virion, viral envelope either fuses w/ plasma membrane or virion is endocytosed and envelope then fuses w/ endocytic vesicle

-positive ssRNA genome already capped and ready to go, so start translation immediately

-produce polyproteins 1a and 1b

-polyproteins processed by viral proteases to produce viral proteins, including subunits of RNA-dependent RNA polymerase (RNA replicase)

-virus replicates on double membrane vesicles in the cytoplasm

-positive ssRNA converted into negative ssRNA

-use negative ssRNA to make capped positive mRNAs and new positive ssRNA genomes

-glycoproteins S, E, M are translated into ER

-genome wrapped in nucleoprotein and then buds into golgi, then gets envelope and glycoproteins

-exits host via secretory pathway

The two pathways for Coronavirus fusion:

1. Fusion of their envelopes w/ the plasma membrane --> TMPRSS-mediated activation

-important for SARS-CoV-2

2. Fusion of their envelopes w/ the endosomal membrane --> cathepsin-mediated activation

Hydroxychloroquine Effectiveness:

-found to inhibit infection of cultured cells (VERO cells) by SARS-CoV-2 but has not been shown to be effective against SARS-CoV-2 in human cells

14. What was the question that Luria and Delbrück sought to answer with their experiment? Describe how their experimental results led to their conclusion. What results would have supported the alternate hypothesis? ***

-There was an ongoing debate abt whether Darwin's theory applied to microbes/bacteria as well

-Luria and Dulbrick showed that bacteria were subject to natural selection,

--based on an observation they made that whenever they plate E. coli on media w/ a lot of phage, most cells die but a few colonies grow. So, they questioned if the bacteriophage are causing these mutations to make them resistant or are the mutations occurring spontaneously and then selected for

--spontaneous mutation: a rare and random event

--resistant mutant: can arise by chance at nay time. if mutation happens early, then it has more time to pass down to offspring--many generations of offspring will contain that resistant mutation. if mutation occurs late, then has less time to pass down to offspring

Experiment: The Fluctuation Test

-took a wild type strain (in TSB), which is phage-sensitive

-they used this culture to inoculate (subculture) a 100 culture tubes and grew overnight cultures (16 hrs in TSB and no exposure to phage)

-after ON incubation, they took serial diluted each sample and plated it onto TSA plates w/ phage

--NOTE: bacteria were not exposed to phage until this step

-they found that growth of colonies fluctuated wildly (some even had none)

-plates w/ less colonies had mutation occur late

-plates w/ more colonies had mutation grow early as it had more generation time to pass down mutation to offspring

-plates w/ no colonies never had a mutation occur

-they fit these fluctuating numbers into mathematical models and showed that they were consistent w/ the mutations occurring at various times during growth of culture

-able to conclude that phage resistant mutations occur SPONTANEOUSLY and then are selected for (fitness advantage) by the phage, therefore, Darwin's laws/theory of evolution apply to bacteria too

-However, if

15. What is a prion and how is it believed to cause disease? How is a prion different from a virus?

Prions: not viruses! they're infectious proteinaceous particles

How they cause disease: when proteins fold incorrectly, it causes other prion proteins to be mis-folded too, causing aggregates that can kill cells

-the prion protein is normally in the alpha-helices form, where it is not harmful, but a mis-folding that leads to beta-sheets forming turn it harmful

-prion proteins are found in the brain and they kill neurons

Characteristics of prions (that make them different from viruses):

-prions are resistant to high temps, that would normally inactivate viruses

-not sensitive to radiation, which normally damages virus genomes

-not destroyed by enzymes that digest RNA or DNA

-sensitive to protein denaturing agents

1. Who are the eukaryotic microbes? Name three main groups of organisms that comprise the eukaryotic microbes. What compound is commonly found in the cell walls of fungi such as yeast?

Eukaryotic microbes have:

-a true nucleus and membrane-bounded organelles

-a characteristic cell envelope

The 3 main groups:

1. Fungi

-cell walls contain chitin

2. Protozoa

3. Algae

2. Besides E. coli, Saccharomyces cerevisiae (yeast) is probably the most widely used organism for laboratory study. What features does yeast have in common with E. coli that make them both great model organisms for biological study? What are advantages of using yeast rather than E. coli for research? What are advantages of using E. coli rather than yeast?

What makes Yeast a Model Organism:

-many genes are highly conserved from yeast to plants to humans

--divides by budding (leaves bud scar, so easy to track)

--stable haploid and diploid states, so useful for genetic experiments (effects if recessive mutations are evident in haploids)

-has a doubling time of about 2 hours in rich media

-easily transformed by foreign DNA. Yeast can be made competent artificially, and homologous recombination is very efficient in Yeast

What features Yeast has in common w/ E. coli:

-many vector plasmids that replicate in both Yeats and E. coli are available for recombinant DNA work

--grow rapidly

--have small, fully sequenced genomes

--both can be transformed to competent cells

Advantages of using Yeast rather than E. coli:

-Yeast has a small genome (14 million bases) about 3 times the size of E. coli genome

-recombinant eukaryotic proteins expressed in Yeast are more likely to be properly folded and post-translationally modified than the same proteins expressed in E. coli

--gives insight into human cell bio b/c yeast is eukaryotic like human cells

--more relevant to human health

--phenotypes can be observed

Advantages of using E. coli rather than Yeast:

-more streamlined (ex: gene regulation)

-insight into mechanisms for bacterial pathogenesis

3. Give at least 5 examples of research questions that could be studied using yeast but not E. coli (For example: How does the spliceosome function? How are chromosomes segregated during mitosis?). Give at least 5 examples of research questions that could be studied using E. coli but not yeast.

...

4. What are 4 major groups of protists and generally how are they distinguished? Which one includes Plasmodium, the parasite that causes malaria?

4 major groups of protists (classified based on motility) --means of locomotion:

1. Flagellates -- flagella

2. Amoeboids -- pseudopodia

3. Ciliates -- cilia

4. Apicomplexa (apicoplast) -- nonmotile

--includes plasmodium (causes malaria)

5. Describe the life cycle of Plasmodium. Inside which organs or tissues does it multiply? How does the sexual reproductive stage of the parasite occur? Which stage is targeted by the RTS,S Mosquirix malaria vaccine?

-The cycle begins with the bite of the Anopheles mosquito, which carries the Plasmodium parasites

-The Anopheles mosquito then releases sporozoites into the bloodstream. These sporozoites travel through the blood stream, to the liver, where they then invade the hepatocytes

-It is here that the parasites replicate, and develop over the course of a little more than a week

-After this period of replication, the so called merozoites burst forth from the hepatocyte, and begin a continual cycle of infecting red blood cells--this is the asexual stage of the parasite

-It is here that the parasites will invade red blood cells, and consume the hemoglobin inside as a food source

--This is where all the clinical manifestations of the disease occur and this is where most drugs attempt to eradicate the parasite

-Now, w/ some infrequent rate, by a mechanism that is frankly poorly understood, a small number of parasites will develop into a sexual form called gametocytes

-It is these gametocytes that re then picked up by a new Anopheles mosquito when taking a blood meal, and the entire cycle starts over again.

-If you can treat the disease at the asexual stage, or the block transmission, or block passing on the disease to new mosquitos, one can effectively control the disease

In Class Question: Which of the following would NOT undergo mitosis? Which of the following would contain murein (peptidoglycan)? Plasmodium falciparum, Saccharomyces cerevisiae (yeast), Entamoeba histolytica (amoeba), Staphylococcus epidermidis, Paramecium, Escherichia coli, Mycobacterium tuberculosis

NOT undergo Mitosis:

-Staphylococcus epidermidis

-Escherichia coli

-Mycobacterium tuberculosis

Contain Peptidoglycan:

-Staphylococcus epidermidis

-Escherichia coli

-Mycobacterium tuberculosis

Yeast Stories

Story 1: Yeast, cell division, and cancer

-Lee Hartwell performed a genetic screen and discovered the cell division cycle (CDC) genes, which control progress through the cell cycle

-the CDC genes are highly conserved in yeast, plants, and animals

-studying the cell cycle in yeast has improved our understanding of the causes of uncontrolled cell division and chromosome loss, which can lead to cancer in humans

Story 2: Yeast and Parkinson's disease

-Parkinson's disease- a neurodegenerative disease that affects movement

-αSynuclein protein (αSyn):

--accumulates in neurons of patients w/ Parkinson's disease

--missense mutations in αSyn gene or increased expression of the gene cause Parkinson's disease

-misfolding and accumulation of α-synuclein kills both neurons and yeast cells (b/c its cytotoxic)

-How αSyn accumulation kills cells: a genetic screen using led to the discovery that αSyn is toxic to cells b/c it blocked ER-to-Golgi trafficking

--Yeast cells do not express this protein naturally, but if added artificially, it leads to the yeast cells' death

Story 3: Yeast and genes that control aging

-counting bud scars (chitin rings) is a way to measure yeast cell age

--the oldest cells become sterile and divide more slowly

-a genetic screen identified mutants that had extended yeast cell lifespan

--when a cell buds off, it leaves a scar and the next one to bud off is adjacent to it (the cell w/. more bud scars is the oldest)

-a genetic screen found that the sir2 (sirtuins) gene influences aging in terms of longevity

--activation of the sir2 gene leads to hyper-activation of lifespan lengthening as sir2 encodes NAD-dependent deacetylase, which affects metabolism

--somewhat proved that calorie restriction increases lifespan

-Sirtuins are evolutionarily conserved in eukaryotes as well as yeast, worms, flies

1. What is "selective toxicity"? Does a good antimicrobial drug exhibit selective toxicity or not? Explain.

-a good therapeutic agent must have selective toxicity--the to ability to harm the microbial pathogen w/out causing significant harm to the host

-chemotherapeutic agents are used to directly or indirectly inhibit the uncontrolled growth and proliferation of cancer cells

-A chemotherapeutic agent needs selective toxicity so that it is only harmful to the pathogen and not the host (the person that is being treated)

2. What is "therapeutic index"? Order the following compounds in order of increasing therapeutic index: polymyxin, sulfa drugs, silver nitrate (note silver nitrate is a heavy metal compound, covered near the end of this packet). Explain your reasoning. Do the same for the following 2 compounds: chlorine bleach and penicillin. Explain.

-Therapeutic index: the ratio of the toxic dose to the therapeutic dose

--Toxic dose: the median dose at which the drug is lethal to the host

--Therapeutic dose: the median dose to effectively treat an infection

-The higher the Therapeutic Dose, the more specifically the drug harms the pathogen, but not the host

-Sulfa drugs vs polymyxins vs. silver nitrate:

(low therapeutic index --> high therapeutic index)

silver nitrate < polymyxins < sulfa drugs

-Reasoning: silver nitrate would have the lowest therapeutic index b/c it is a heavy metal compound, thus somewhat toxic to us. Then, based on their complex chemical structure, polymyxins were next. Sulfa drugs would have the highest therapeutic index as its structure if very similar to PABA structure.

-Chlorine bleach vs. penicillin:

(low therapeutic index --> high therapeutic index)

chlorine bleach < penicillin

-Reasoning: chlorine would have a lower therapeutic index b/c it is a halogen, therefore, it would be more reactive and toxic. However, penicillin would have a higher therapeutic index b/c it is particularly effective towards bacteria and used as an antibiotic.

3. What are the 5 common modes of action of antibacterial drugs? Give examples for each category. Which of these groups of antibacterial drugs are mainly effective against actively growing bacteria? Which of these groups exhibit high selective toxicity? Explain.

5 Common Modes of Action of Antibacterial Drugs:

1. Inhibition of Cell Wall Synthesis

-penicillin = antibacterial & high selective toxicity

-bacitracin = antibacterial & high selective toxicity

-cephalosporin = antibacterial & high selective toxicity

-vancomycin = antibacterial & high selective toxicity

2. Disruption of Cell Membrane Function

-polymyxin = antibacterial & high selective toxicity

3. Inhibition of Protein Synthesis

-tetracycline = inhibits protein synthesis needed for bacterial growth & high selective toxicity

-erythromycin = inhibits protein synthesis needed for bacterial growth & high selective toxicity

-streptomycin = inhibits protein synthesis needed for bacterial growth & high selective toxicity

-chloramphenicol = inhibits protein synthesis needed for bacterial growth & high selective toxicity

4. Inhibition of Nucleic Acid Synthesis

-rifamycin (transcription) = antibacterial & low selective toxicity b/c it can lead to hepatotoxicity (liver toxicity)

-quinolones (DNA replication, metronidazole) = antibacterial & low selective toxicity b/c of side effects such as glucose metabolism dysfunction & increased risk for tendon rupture

5. Acting as an Antimetabolite

-sulfanilamide = antibacterial & high selective toxicity

-trimethoprim = antibacterial & high selective toxicity

4. Why is it important to finish a prescribed course of antibiotics?

-In every bacterial population, there is a natural cell-to-cell variation is susceptibility to an antibiotic

-If a course of antibiotics is not finished, then the bacteria that remain and multiply are the ones that are naturally more resistant. Any surviving bacteria have the opportunity to gain mutations or antibiotic resistance genes that make them even more resistant to antibiotics.

5. What are 5 common ways in which bacteria become resistant to antibiotics? Give an example of each. Which category is b-lactamase an example of?

There are 2 Broad Categories of Genetic Changes that confer resistance:

1. Spontaneous Chromosomal Mutation

-ex: altering the target enzyme's binding site for the drug

-usually only confers resistance to a single type of antibiotic

-antibiotics do not induce mutations, but they do select for antibiotics resistant strains

2. Extrachromosomal Resistance

-R (resistance) plasmids or R factors: some R plasmids carry 6 or 7 genes for resistance to different antibiotics

-R plasmids can be transferred rapidly via conjugation

Specific Effects of Mutations or Functions of R plasmid genes that make bacteria more resistant to antibiotics:

➢ 1. Mutation of the target enzyme or protein, so antibiotic can no longer bind

-ex: mutation of RNA Pol, so RMP (rifampicin) no longer binds

--conferred via a point mutation

➢ 2. Inactivation of antibiotic by chemical modification

-ex: Beta-lactamases, such as penicillinase (cleaves penicillin and inactivates it)

-ex: acetylation of amikacin and other aminoglycosides

--conferred via a point mutation

➢ 3. Preventing antibiotic entry into the cell by decreasing cell permeability

--conferred via a point mutation

➢ 4. By-passing the target enzyme

-ex: Sulfa-drug resistance if bacteria gained a new gene or enzyme that can synthesize folic acid. Synthesizes folic acid from PABA (inhibited by sulfa drugs), but if bacteria gains a gene that bypasses this inhibition and finds a new way to synthesize folic acid, so antibiotic doesn't have the same effect as before

--would need the acquisition of new genes

➢ 5. Pumping the drug out of the cell immediately after entry and before it has the chance to inhibit its target

-ex: bacterial charity

--conferred via a point mutation

6. Bacteria can gain antibiotic resistance via mutation or via gaining new genes. Which of the modes of antibiotic resistance that you named in #5 could be conferred via a point mutation? Which ones would require the acquisition of new genes?

^ in #5

7. Where did antibiotic resistance genes come from? Explain one theory.

-Bacteria eat antibiotic as a source of carbon, thus also contributing to antibiotic resistance b/c they use them as food

-Bacteria subsisting on antibiotics are surprisingly phylogenetically diverse & antibiotic-consuming isolates (of bacteria) are resistant to many antibiotics.

8. What is the newer way of looking for antibacterial drugs described in Moy et al., 2006? Contrast this method with the "old" way of looking for antibacterial drugs. What types of drugs could be discovered with the new method that would not be discovered with the old?

Older Way:

-Classic way of looking for new antibiotics (in vitro):

--have two tubes, one w/ bacteria + drug, the other w/ no drug.

--see if bacteria w/ drug dies, is fo see it has antimicrobial activity and then go on to test in animals

New Way:

-use a 96 well plate

-inoculate w/ C. elegans (worms) + E. faecalis (bacteria) -- normally the bacteria kills the worms

-no drug = worms die

-do worms live in presence of drug? If so, the drug could be a promising new antimicrobial

-this design takes into account Selective Toxicity--built in

-looking for effective curing of the infection

-this method allows the identification of compounds that target bacterial virulence factors

Drugs that can be discovered w/ new method & not old method:

-The findings from the paper indicate that the whole animal C. elegans screen identifies not only traditional antibiotics, but also compounds that target bacterial virulence or, stimulate host defense

9. What is a common mode of action of antifungal drugs?

-Mode of Action: many antifungal drugs exploit differences btwn. human and fungal cell membranes

--Human cell membranes contain cholesterol whereas fungal cell membranes contain ergosterol

--ex: Imidazoles (inhibit ergosterol synthesis), Nystatin (binds to ergosterol in fungal cell membranes, disrupting the cell membrane)

10. What is a common mode of action of antiviral drugs? Name three antiviral drugs against SARS-CoV2 and describe their mode of action. During which stage of COVID-19 should these be administered to be effective?

-Mode of Action: many antivirals are nucleotide analogs that inhibit viral nucleic acid synthesizing enzymes. They target enzymes of viruses instead of our enzymes, thus not inhibiting our activity.

3 Antiviral Drugs against SARS-CoV-2:

1. Molnupiravir: causes many mutations in SARS-CoV-2 genomes

-Mode of Action: gets incorporated into RNA, but it sort of shifts its structure to form base pairs w/ different nucleotides. Thus, the virus genome "mutates to death," so until virus is no longer functional. It is administered orally and accessible to many people.

--this drug is to reduce risk of hospitalization or death

2. REGEN-COV Monoclonal antibody therapy for post-exposure prophylaxis (prevention) for COVID-19

-Mode of Action: SARS-CoV-2 uses spike protein to attach to and enter human cells, which allows it to cause infection. Monoclonal antibodies bind to the spike protein, prevent the virus from attaching to human cells, and tag it for destruction. This may prevent development of severe COVID-19 ( for people who are already COVID Positive)

3. PFIZER's Novel COVID-19 Oral antiviral treatment, called PAXLOVID

-Mode of Action: a SARS-CoV-2 protease inhibitor, prevents protease from cleaving the polyprotein (a long that has been translated in one long line) that make the functional RNA replicase enzyme for SARS-CoV-2.

4. Fluvoxamine --> an Antidepressant has shown advances in treating early cases of COVID (study still in progress)

11. In general, is it more difficult to find good antibacterial drugs or good antiviral drugs? Why? In general, is it more difficult to find good antifungal drugs or good antibacterial drugs? Why?

-Antiviral Drugs: it is difficult to find good antiviral drugs b/c viruses have fewer potential targets and a smaller genome

-Antifungal Drugs: It is difficult to find good antifungal drugs b/c both fungi and human cells are eukaryotes, so it is difficult to target them w/out hurting our cells too. As opposed to bacteria, which are prokaryotes, so when targeted, they wont hurt our cells.

12. What are examples of physical methods of microbial growth control? What are examples of chemical methods of microbial growth control? What are their modes of action?

➢ Bacteria have proteins, cell membranes and nucleic acids, which is why chemical methods are a great way for disinfecting surfaces. However, we are comprised of similar structures, which is why we cannot ingest them for treatment.

➢Chemical Antimicrobial Agents:

-halogens, such as iodine, chlorine, or halogenated compounds react w/ proteins, membranes, nucleic acids

-aldehydes react w/ proteins and nucleic acids

-heavy metals, such as silver, mercury, or copper react w/ proteins

-cationic detergents, such as quaternary ammonium compounds denature proteins, disrupt membranes. Detergents have a hydrophilic end and a hydrophobic end, which allows them to disrupt membranes and solubilize proteins

➢ Physical Antimicrobial Methods:

1. Heat: denatures proteins, degrades nucleic acids, disrupts cell membranes. Preferred method of sterilization/disinfection of items that can withstand high temps

↳Moist Heat (Steam--autoclaving): moist heat penetrates substances more quickly than dry heat.

↳Dry Heat (oven): killing is slower

↳Pasteurization: does NOT sterilize, but kills pathogens and decreases the number of microbes that cause spoilage.

2.Freezing or Refrigeration: these don't sterilize but do slow or inhibit cell growth. For long-term storage of microbes in research labs, bacteria or yeast can be frozen at -80 for yrs, w/ glycerol present to protect cell membranes.

3. Filtration: useful for sterilizing heat-sensitive solutions. Physically separates microbes from solutions based on size (filters w/ different pore sizes are available).

4. Radiation

↳UV radiation causes thymine dimers (DNA damage), but doesn't penetrate surfaces

↳ionizing radiation (gamma rays): extremely powerful mutagen; penetrates deep into objects and kills endospores. Used for irradiating meat, fruits, and vegetables.

13. In general, how would the therapeutic indices compare between the physical methods of growth control, the chemical methods of growth control, and the antimicrobial drugs? Explain.

-The physical and chemical methods of growth control are meant for surfaces outside the body, while antimicrobials are meant to inhibit microbial growth w/in the body.

-We cannot use physical or chemical methods of microbial growth to inhibit microbial growth w/in our body b/c they will damage our cells.

14. How is sterilization different from disinfection? How is bacteriostatic different from bactericidal?

Sterilization vs. Disinfection:

↳ Sterilization: the killing or removal of all living cells, spores, or viruses in a material or on an object

↳ Disinfection: killing, inhibition, or removal of pathogenic microbes

Bacteriostatic vs. Bactericidal:

↳ Bacteriostatic: inhibits bacterial growth; MIC (Minimum Inhibitory Concentration): lowest concentration of an antibacterial needed to inhibit bacterial growth

↳ Bactericidal: kills bacteria; MBC (Minimum Bactericidal Concentration): lowest concentration of an antibacterial needed to inhibit bacterial growth

In Class Question 1: Why is it worse than useless to take antibiotics for a cold or for the flu?

Taking antibiotics during viral infection can result in antibiotic resistant bacteria. They also have an affect on the microbiome (where the good bacteria is).

In Class Question 2: What effects would these physical or chemical agents have on human cells?

Damage human cells

In Class Question 3: How do you think the selective toxicity of a chemical agent (like phenols) compares to the selective toxicity of antimicrobial drugs?

-Antimicrobial drugs are higher in selective toxicity as they inhibit growth of bacteria, while causing as little harm to the host as they can.

-Chemical agents are much lower in selective toxicity as they can damage human cells.

1. Define and contrast: symbiosis, mutualism, parasitism, commensalism. Could a single organism be a commensal sometimes and a parasite at other times? Explain.

Symbioses = living together -- this is broken down into 3 categories:

↳ 1. Mutualistic: both partners benefit

-microbe-host mutualisms are common in nature

-together, symbiotic partners have new abilities that they do not have alone, allowing them to fill new environmental niches

↳ 2. Parasitic: one partner take nutrients/resources from the other

↳ 3. Commensal: one partner benefits, the other is unaffected

These definitions are not absolute, and can shift.

↳For example, a commensal relationship may become parasitic based on changed induced on the host, which may not be fully understood yet.

2. Why are mutualisms common in nature?

Mutualisms are crucial to the reproduction and survival of many plants and animals and to nutrient cycles in the ecosystem

3. What is an endosymbiosis? Between which types of organisms are endosymbioses commonly observed? Give two examples of an endosymbiont and its host, and describe what each partner gains from the endosymbiosis.

Endosymbiosis: a symbiosis in which the bacteria symbiont lives inside the host's cells

↳ Commonly observed in bacteria-invertebrate symbioses, and in some bacteria-plant symbioses

Examples of Endosymbiosis:

↳ 1. The origin of the Eukaryotic cells:

-Mitochondria evolved from Rickettsiae, a group of bacteria that are obligate intracellular parasites, even now

-Chloroplasts evolved from Cyanobacteria, a group of photosynthetic bacteria

-Genomes of Mitochondria (37 genes) and Chloroplasts (128 genes) are very much reduced in size compared to the genomes of modern-day Rickettsiae or Cyanobacteria

--Many genes were transferred to the nuclear genome os lost, as the endosymbiont came to rely on its host

Bacteria-like qualities of Mitochondria and Chloroplasts:

1. Divide by Binary Fission; some have FtsZ (conserved proteins) for septum formation

2. Protein Synthesis like bacteria: initiate w/ f-methionine; 70S ribosome (euk ribosomes are 80S)

3. Circular chromosome; no histones

↳ 2. Bacteria-Insect Symbioses

-Endosymbiont provides nutrients that host is unable to synthesize

-ex: Buchnera, the bacterial endosymbionts of aphids, can synthesize the 10 amino acids that the aphids cannot make

--Bacteriocytes are specialized insect cells in which the cytoplasm is packed w/ bacterial endosymbionts

4. How do Onchocerca (nematode) and Wolbachia (bacteria) influence river blindness?

↳Millions of people in tropical regions infected w/ Onchocerca, a parasitic worm transmitted by biting flies

↳ The worm lives in the human body for years and causes river blindness, a leading cause of blindness worldwide

↳ In 2002, it was discovered that work extracts lacking their usual endosymbiotic bacteria (Wolbachia) did not cause the disease in a mouse model, indicating that the worms' endosymbiotic bacteria are critical for causing the disease

5. What is the microbiota? Specifically, where is the microbiota commonly found? How can human beings be considered "supraorganisms"?

↳ The microbiota can be viewed as a metabolic "organ" exquisitely turned to our physiology that performs functions that we have not had to evolve on our own. These functions include the ability to process otherwise indigestible components of our diet, such as plant polysaccharides

↳ Most commonly found in the Gut

↳ Humans should be considered "supraorganisms." Each human has ~20,000 different genes, plus millions of genes from the microbiome. Together, these genes influence human health.

↳ Definition of Supraorganism: Individual organisms that together function like a single organism

6. Describe how the human microbiome can influence drug effectiveness. How did research by Wallace et al. seek to alleviate side effects of the colon cancer drug CPT-11 (irinotecan)?

How Human Microbiome can influence drug effectiveness:

↳CPT-11 is administered intravenously as a prodrug. Carboxylesterases (CE) in various tissues convert CPT-11 into SN-38, which kills cancer cells.

↳ SN-38 can be inactivated in the liver by UGT, generating SN-38G that is excreted via bile into the intestine.

↳ Although SN-38G is not toxic to the intestinal mucosa, Beta-glucuronidases (Beta-gluc) produced by gut flora metabolizes SN-38G to SN-38, which damages the gut mucosa.

How did Wallace et al. alleviate the side effects?

↳ Inhibitors, such as Inhibitor 1, of Beta-glucuronidases prevent the conversion of SN-38G to SN-38, and diminish Irinotecan (CPT-11) gut toxicity.

7. What is an opportunistic pathogen? Under what conditions would an opportunistic pathogen be able to cause disease? Define: microbial antagonism.

➣ Opportunistic pathogens (Opportunists): organisms that take advantage of particular opportunities to cause disease. These may be organisms that are part of the normal microbiota or not.

➣ Opportunities:

↳ 1. Failure of the host's normal defenses

--When the host is immunocompromised (ex: due to HIV, cancer treatment or immunosuppressant drugs)

↳ 2. Introduction of the organisms into the unusual body sites

--ex: E. coli in surgical wounds (rather than its usual site, the large intestine)

↳ 3. Disturbances in the normal microbiota

--Microbial Antagonism: normal microbiota ordinarily competes w/ pathogens for nutrients and space

--ex: relevance to Clostridium difficile

--microbes make it harder for invaders to come in. If these microbes were wiped out due to, for ex, antibiotic treatment, then you get rid of the microbes that help keep away invaders.

8. Define: virulence factor. Give examples of the various roles virulence factors may play.

➣ Virulence factors are structures or proteins produced by pathogens that help them to cause disease

↳Virulence factors are not meant to cause disease, but do so in the process

↳they are meant to protect, which is why they are not grouped w/ antibiotic resistance genes

➣ Examples of Bacterial Virulence Factors:

↳ 1. Structures important for adhesion to cells and tissues

--ex: adhesins, capsules, pili

↳ 2. Enzymes or structures that help in evading host defenses:

--Hyaluronidase: an enzyme produced by Streptococci that digests hyaluronic acid, a gluelike substance that helps hold the cells of certain tissues together

--antiphagocytic capsules: unable to be cleared very effectively by phagocytosis

↳ 3. Toxins that cause disease directly:

--Exotoxins: powerful secreted protein toxins

--Endotoxins: LPS in the gram-negative cell envelope, released only if the bacteria are dying, relatively weak toxin (LPS is part of the outer membrane, not secreted but only accessible)

--ex: Hemolysins are toxins that lyse red blood cells and other cells

↳ 4. Secretion systems for delivery of toxins or other effectors

--ex: Type 3 or Type 4 Secretion Systems

9. What is/are the virulence factor(s) of C. tetani, the bacteria that causes tetanus? How do those virulence factor(s) help C. tetani cause disease?

-Clostridium tetani is a Gram-positive bacteria that forms endospores; found in soil

-C. tetani is not invasive; enters the host by contaminating wounds

-C. tetani is a strict anaerobe and doesn't survive long in human tissues

➣ Tetanus:

↳ Produces a powerful exotoxin that interferes w/ neurotransmitters and causes muscle tightening (spastic paralysis) and asphyxiation; among the most potent toxins known; causes paralysis even at extremely low concentrations

--Virulence Factor = a powerful Exotoxin that causes (spastic) paralysis

10. Repeat #10 for E. coli O157:H7 and for B. anthracis.

➣ E. coli O157:H7:

↳ Virulence factors that allow E. coli O157:H7 to adhere and colonize the surface of the intestinal epithelium are adhesins and secretion systems

↳ E. coli makes the Tir protein, which binds to the Intimin (helps w/ adhesions), which is delivered via secretion system

↳ For E. coli O157:H7, infected cattle or people are the usual carriers

--a carrier is an infected individual who is a potential source of infection for others

➣ Anthracis:

↳ Antiphagocytic capsule (polyglutamyl) which inhibits phagocytosis of the actively growing bacteria

↳ Anthrax toxin--contains 3 different Exotoxins. Together, the exotoxin kill Macrophages, which releases toxic chemicals, leading to fever, internal bleeding, septic shock, and death.

--the genes for these toxins are found on a Virulence Plasmid

↳ In addition (although this would not be considered a "virulence factor"), Bacillus Anthracis can form Endospores. Anthrax endospores in carcasses of animals previosuly infected w/ anthrax can be found in the soil.

11. Unnecessary antibiotic use can lead to infection by which types of pathogens? Why?

➣ Opportunistic Pathogens

↳ When antibiotics are unnecessarily taken, it could lead to harmful effects such as hard to treat infectious due to the failure of the host's normal defenses

↳ The antibiotics ingested lead to the host having an immunocompromised system that is easily taken advantage of by the opportunistic pathogen

12. Would you consider a ribosomal protein (one of the proteins that makes a functional ribosome) to be a virulence factor? Why or why not?

➣ No

↳ If we were to knock ribosomal proteins, then mRNA will not be able to be translated, but this affect is not specifically for disease causing.

↳ Knocking out ribosomal proteins would not lead to decreased function b/c of mutation, but just death of cell.

1. What are the innate defenses? What triggers an innate response? How quickly do they respond?

➣ Innate Defenses:

↳ are present constitutively

↳ are virtually identical among all members of a species

➣ Triggers & Response Time:

↳ activated by antigens & their chemical properties

--respond QUICKLY

2. Which cells are responsible for the removal of most of the microbes your body encounters in your lifetime?

Phagocytes: white blood cells (leukocytes) that ingest and kill microbes

3. Define: complement. How is complement activated? What are the different functions of complement?

The Complement: system of ~30 soluble protein, usually inactive, but can be activated in a Proteolytic Cascade

3 Ways to Activate the Complement System:

↳ 1. Alternative pathway: surface of the pathogen

↳ 2. Binding of mannan on pathogen by serum lectins

↳ 3. Antigen-antibody complex

Functions of Complement:

↳1. Mediators of inflammation, phagocyte recruitment

↳ 2. Opsonization of bacteria

↳ 3. Membrane attack complex (MAC), lysis of cells

4. There are two main categories of phagocytes. What are they, and how do they differ from each other? How do these phagocytes recognize, then destroy microbes? Define: MAMPs, PRRs, Toll-like receptors, NETs.

Two main categories of phagocytes: Macrophages and Neutrophils

↳Macrophages: efficient phagocytic cells that can leave the circulatory system by moving across the walls of capillary vessels. The ability to roam outside of the circulatory system is important, b/c it allows macrophages to hunt pathogens w/ less limits. Macrophages can also release cytokines in order to signal and recruit other cells to an area w/ pathogens

↳ Neutrophils: phagocytic cells that are also classified as granulocytes b/c they contain granules in their cytoplasm. These granules are very toxic to bacteria and fungi, and cause them to stop proliferating or die on contact.

How do Phagocytes recognize & destroy Microbes:

➣ Key Concept: Phagocytes are cells that recognize pathogens and destroy them through phagocytosis. Recognition often takes place by the use of phagocyte receptors (Pattern Recognition Receptors--PRRs) that bind molecules commonly found on pathogens, known as MAMPs.

↳ Neutrophils and Macrophages have Toll Like Receptors (TLRs), a type of PRRs that bind to MAMPs.

Formation of Neutrophil Extracellular Traps (NETs):

↳ When Neutrophil dies, it releases its chromatin, which creates this "net" that traps in microbes

↳ This is b/c when a cell is lysed, its DNA becomes viscous that is able to trap things in

5. What are some mechanisms that microbes use to undermine the innate defenses?

1. Anti-phagocytic Capsules: the capsule prevents phagocytosis

↳ ex: Haemophilus influenza and Bacillus anthracis

2. Inhibit fusion of lysosome w/ the phagosome, so the bacteria has a safe place to live inside the macrophage

↳ ex: Mycobacterium tuberculosis

3. Escape from the phagosome and divide in the cytoplasm of host cells. These bacteria then use the host cell's cytoskeletal proteins to propel them into neighboring cells

↳ ex: Listeria and Shigella

1. Contrast the innate and the adaptive defenses. Who are their main players? What triggers an innate or adaptive response? How quickly do they respond?

➣ Innate Defenses (Non-specific): a genetically programmed set of responses that can be mobilized immediately when an infection occurs

1. antigen independent

2. not antigen specific

3. exposure = quick response

4. no immunological memory

5. triggers: an innate response is activated by the presence of antigens and their chemical properties

➣ Adaptive Immunity (Specific):

1. antigen dependent

2. antigen specific

3. lag btwn. exposure and response time

4. immunological memory

5. triggers: adaptive immunity is initiated when an innate immune response fails to eliminate a new infection, and antigen & activated antigen-presenting cells are delivered to the draining lymphoid tissues

2. Define: antigen. Are all antigens proteins? How are antigens different from an epitope?

➣ Antigen: a substance the body identifies as foreign (non-self)

↳ many antigens are proteins or glycoproteins, but some are polysaccharides

➣ Epitope: a REGION on an antigen that is specifically recognized by antibodies or T-Cell Receptors

3. Define: antibody. Which cells produce antibodies? What are the functions performed by antibodies (be specific)? Are antibodies always soluble proteins, or are they ever found associated with cells? Explain. How do the IgG and IgM antibodies differ from each other?

➣ Antibody (aka Immunoglobulin): a protein that can bind specifically to epitopes on antigens

↳ Antibodies are produced by B lymphocytes (B Cells)

➣ Functions are performed by antibodies:

↳ 1. Opsonization: the process by which bacteria are altered by opsonins so as to become more readily and more efficiently engulfed by phagocytes (opsonins are a type of antibody)

↳ 2. Activation of complement (in innate defenses--the "classical pathway")

↳ 3. Agglutination: antibodies bind to microbe or viruses, forming clumps that are easily removed. For this to work, each antibody molecule has two binding sites so that they form clumps that can be easily removed

↳ 4. Neutralization: antibodies bind to toxins or antigens or viruses, so they can't attach to receptors on host cells

--Let's say you have a virus, and normally it attaches to a cell, but when surrounded by all these antibody molecules, they prevent them from attaching to the cell due to steric hindrance, so it cannot infect the cell.

--if antibodies bind to exotoxin, then exotoxin cannot bind to cell

--basically, antibodies bind to cell or exotoxin to prevent attachment step and infection, thus neutralizing it

➣ Are antibodies always soluble proteins?

↳ Antibodies can bind specifically to epitopes on antigens. They have two forms: 1). attaches to B Cell, 2). remains in soluble form, unattached in extracellular fluid

➣ IgG vs. IgM:

↳ IgG: the main class of antibodies found in the blood, accounts for as much as 20% if all plasma proteins

--produced in the largest quantities during a secondary response

--IgG can cross the placenta from mother to fetus and can be found in milk

--really effective in fighting off antigens

↳ IgM: found on the surface of B cells and secreted as a pentamer (connected by a J chain) by plasma cells

--the first antibody secret

4. Describe the general characteristics of the adaptive defenses. Define: diversity, immunological memory, clonal deletion, clonal selection, humoral immunity, cell- mediated immunity. What is a memory T or B cell, and how is it different from a "naïve" T or B cell?

➣ General Characteristics of Adaptive Immunity:

↳ 1. Specificity

--antibodies and T-Cell Receptors bind specifically to epitopes

↳ 2. Amazing diversity

--in total, B and T lymphocytes can recognize ~10^8 different epitopes

--each antibody or T-Cell Receptor recognizes just one epitope

↳ 3. Discrimination btwn. "Self" and "Non-self"

--clonal deletion-B and T cells that bind to "self" are destroyed during development and never make it out to circulation

--problems w/ distinguishing "self" from "non-self" result in autoimmune disease

↳ 4. Memory

--quick secondary response to previously encountered antigens

--involves Memory B and T cells

--Basis for vaccination

Definitions:

➣ Immunological Immunity: the secondary response is faster and more efficient due to the presence of memory B and T cells--long lived cells that may persist for decades, ready for activation upon encounter w/ the same antigen

➣ Clonal Selection: a single B or T cell that recognizes an antigen that enters the body is selected from the pre-existing cell pool of differing antigen specificities and then reproduced to generate a clonal cell population that eliminates the antigen.

➣ Humoral Immunity: carried out by antibodies produced by B lymphocytes

↳ most effective at defending against bacterial toxins, bacteria, and viruses circulating in the body, before they have entered cells

➣ Cell-Mediated Immunity: carried out by Cytotoxic T Lymphocytes (CTLs): most effective when antigens are already inside host cells and inaccessible to antibodies

↳ For example, cell-mediated immunity is involved in killing virus-infected cells

5. What is a T cell receptor? How is it different from an antibody? How is a T cell receptor similar to an antibody? Where are the CD4 and CD8 proteins found?

➣ Two classes of receptor proteins that mediate antigen recognition are Antibodies and T-Cell Receptors (TCRs).

↳ Antibodies are able to bind a diverse range of antigen shapes

↳ TCRs are specialized to recognize a cell surface protein, the Major Histocompatibility Complex (MHC)

➣ CD4 and CD8 Proteins:

↳ Helper T lymphocytes (CD4 T Cells) are activated when their T Cell Receptors bind to antigen + MHC Class II exhibited on the surface of antigen-presenting cells, such as Dendritic Cells, Macrophages, and B Cells

↳ Naive Cytotoxic T lymphocytes (CTLs or CD8 T Cells) are activated and induced to proliferate after their T Cell Receptors bind antigen + MHC Class I on Dendritic Cells and the CTL receives co-stimulatory signals from the Dendritic Cells

6. Contrast the helper T cells and the cytotoxic T lymphocytes (CTLs), in terms of how they function and how they are activated. What is the role of antigen- presenting cells? What happens after a helper T cell is activated? What happens after a cytotoxic T cell is activated?

➣ Helper T Cells (CD4 T Cells)

↳ Activation: when their T Cell Receptors bind to antigen + MHC class II on the surface of antigen presenting cells

--Antigen presenting cells are cells that can process a protein antigen, break it into peptides, and present it in conjunction w/ class II MHC molecules on the cell surface where it may interact w/ appropriate T Cell Receptors.

↳ Activated helper T cells have a critical role in producing Cytokines that activate and induce proliferation of B cells (so that the B cells can differentiate and proliferate to form plasma cells), Macrophages, and Killer T cells

↳ The critical role of Helper T Cells is seen AIDS, in which the Helper T Cells are killed by HIV and the patient loses immune system function

↳ Some CTLs also need help from Helper T Cells to be activated

➣ Cytotoxic T lymphocytes (CTLs or CD8 T cells)

↳ Activation: T cell receptors bind to antigen + MHC class I on Dendritic cells & the CTL receives co-stimulatory signals from the Dendritic cells

↳ An activated CTL recognizes an infected cell when the T Cell Receptor binds to antigen + MHC Class I on the infected cell

↳ The infected cell is killed directly by the CTL by releasing perforin and granzymes, which together induce apoptosis of the infected cell

↳ Activated CTLs kill virally-infected cells or other cells infected by intracellular pathogens

7. Describe the mechanism of B cell activation. How are helper T cells involved? Where does most B cell activation occur? What happens after a B cell is activated?

-The immune system can produce antibodies against millions of different antigens. B cells, which are constantly produced from lymphoid stem cells are found in the blood and lymphoid tissues

-Each B cell can synthesize only pone of the millions of possible antibodies and displays this antibody on its surface

-When an antigen meets a B cell having a surface antibody of the proper specificity, it complexes w/ the antibody. B cells that re not activated do not develop further

-Stimulation of only the B cells that carry antibodies that react w/ antigen it referred to as clonal selection

-Following antigen-antibody interaction, antigen-antibody complexes aggregate on the surface of the cell. This is called capping. The antigen-antibody complexes are then internalized.

-The activated B cell swells and begins to divide rapidly, producing a B cell clone

-The B cells then differentiate into plasma cells and memory cells. Plasma cells produce the specific antibody that provoked its formation, whereas memory cells remain in circulation, but do not produce antibodies. Memory cells can become activated by a later challenge by the same antigen

-Many antibody responses require signals from T helper cells. The T-helper cell is stimulated by an interaction w/ an antigen presenting cell such as a Macrophage

-The Macrophage ingests the antigen, digests it and presents it on the cell surface via a Class II MHC

-The Macrophage then activates a T-helper cell that carries a T-Cell Receptor capable of recognizing the antigen on the Class II MHC of the Macrophage

-The B cell presents the antigen on its surface using Class II MHC

-The activated T-Helper Cell interacts w/ the B cell and secretes chemical signals called Interleukins to stimulate the B cell to differentiate into plasma cell and produce antibodies

8. HIV specifically targets the helper T cells. Explain why the destruction of helper T cells is so devastating, referring to the roles of the helper T cells in the adaptive defenses.

➣ The critical role of Helper T cells is seen in AIDS, in which the Helper T cells are killed by HIV and the patient loses immune system function

➣ HIV: shut down the immune system by attacking the Helper T cells, central players in adaptive immunity

➣ Once Helper T cells are destroyed, the body can no longer launch a specific immune response and becomes susceptible to many opportunistic infections.

9. Explain how vaccines can protect against diseases. What are some materials that serve as effective vaccines?

➣ What makes a good vaccine?

↳ something that teaches your immune system to recognize the "intruder" w/out making you sick, usually with the help of memory cells.

➣ Materials that serve as effective vaccines:

↳ Live-attenuated virus

↳ Inactivated virus

↳ Protein subunit

↳ Virus-like particles

↳ DNA and RNA

↳ Viral vector

↳ Antigen-presenting cells

10. Which cells are currently used to replicate the virus to make the flu vaccine? How is the injected flu shot different from the nose-sprayed flu vaccine? What is the difference between a trivalent and quadrivalent flu vaccine?

➣ New approaches that grow the viruses in mammalian cells have received regulatory approval and will probably soon replace the egg-grown vaccine.

➣ Flu shot vs. Nose-spray:

↳ Flu shot: the form of the vaccine that is injected, does not contain viable viruses. It contains viruses that have been inactivated so that they cannot invade cells and reproduce

↳ Nose-spray: contains viable virus, but has been chemically weakened ("attenuated"), so that it reproduces poorly

➣ Trivalent vs. Quadrivalent:

↳ Trivalent: virus contains three different inactivated influenza strains matching those that circulated the previous year: influenza A H1N1 and H3N2 and the most commonly detected influenza B strain

↳ Quadrivalent: vaccine containing inactivated viruses of both influenza B strains in addition to the H3N2 and H1N1 influenza A strains

11. Contrast different types of COVID-19 vaccine candidates and how they work. Regarding mRNA vaccines: how are they manufactured? How do they result in immunological memory? What are hypothesized benefits of mRNA vaccines?

➣ Types of COVID19 Vaccine Candidates:

↳ 1. Weakened virus: a virus is conventionally weakened for a vaccine by being passed through animal or human cells until it picks up mutations that make it less able to cause disease. Codagenix in Farmingdale, NY, is working w/ the Serum Institute of India, a vaccine manufacturer in Pune, to weaken SARS-CoV-2 by altering its genetic code so that viral proteins are produced less efficiently.

↳ 2. Inactivated Virus: in these vaccines, the virus is rendered uninfectious using chemicals, such as formaldehyde, or heat. Making them, however, requires starting w/ large quantities of infectious virus.

↳ 3. Replicating Viral Vector (such as weakened measles): the newly approved Ebola vaccine is an example of a viral-vector vaccine that replicates w/in cells. Such as vaccines tend to be safe and provoke a strong immune response. Existing immunity to the vector could blunt the vaccine's effectiveness, however.

↳ 4. Non-replicating Viral Vector (such as adenovirus): No licensed vaccines use this method, but they have a long history in gene therapy. Booster shots can be needed to induce long-lasting immunity. US-based drug giant Johnson & Johnson is working on this approach.

↳ 5. Protein Subunits: Twenty-eight teams are working on vaccines w/ viral protein subunits--most of them are focusing on the virus's spike protein or a key part of it called the receptor binding domain. Similar vaccines against the SARS virus protected monkeys against infection but haven't been tested in people. To work, these vaccines might require adjuvants--immune-stimulating molecules delivered alongside the vaccine--as well as multiple doses.

↳ 6. Virus-like Particles: empty virus shells mimic the coronavirus structure, but aren't infectious b/c they lack genetic material. Five teams are working on "virus

12. What are ways in which bacteria can be cleared from the body by the innate and adaptive defenses? What are ways in which viruses can be cleared from the body by the innate and adaptive defenses?

➣ Bacteria

↳ Innate: Complement Activation results in the rapid clearance of bacteria by immune cells

↳ Adaptive: humoral immunity carried out by antibodies produced by B lymphocytes

↳ Adaptive: Cell-mediated immunity carried out by cytotoxic T lymphocytes (CTLs)

➣ Viruses

↳ Innate: viruses initially activate the innate immune response which recognizes the viral immune response

13. What are some mechanisms microbes use to undermine the innate or the adaptive defenses?

➣ Direct attacks against the Helper T cells, thus shutting down the main components of the adaptive immune response

↳ ex: HIV

➣ Malnutrition that depletes the health of active Helper T cells, leading to immunosuppression

↳ ex: measles

➣ Mutation that leads to consistent changes in surface antigens, therefore antibodies cannot attack (Antigenic Variation--variation in antigens!)

↳ ex: HIV, influenza

14. What is a superantigen? Describe the "superantigen hypothesis" for SARS- CoV2 and MIS-C. Explain how the superantigen hypothesis could lead to novel treatments for MIS-C and severe COVID-19.

*Cheng et al.

➣ Superantigen: the hyperactivation of T cells, where antigens and antibodies are no longer speciec to each other but a wide variety, leading to an abnormal activation levels of T cells (mechanism works like self-intoxication/attack, where body experiences shock)

➣ Treatments:

↳ The "superantigen" trait is observed by SARS-CoV-2, thus activating the SAg segment of the virus could lead to the self-inflammation of itself (specifically the S protein, which is cleaved by Monoclonal antibodies), thus inhibiting the virus's viral activities.