Micro Final

Question 1

What are some characteristics of a virus?

Answer 1

They are very small and a virion, or complete virus, consists of a nucleic acid surrounded by a protein coat (capsid, the entire virus is a nucleocapsid)

They are metabolically inert and cannot reproduce on their own

They replicate within the host cell using the host cell machinery, where new copies of the genome are made and capsid is synthesized. The capsid is capable of self assembly

Infection is the process by which a virus is introduced to a host cell and reproduces

Question 2

What type of virus is sars-Cov-2?

Answer 2

It is a single stranded RNA (ss RNA) virus

Question 3

What are the steps of the bacteriophage “life” cycle?

Answer 3

1) Attachment to a cell (adsorption). Bacteriophages generally attach to structures such as pili, flagella, or receptors for transporters such as a maltose binding protein

2) Penetration where viral DNA enters and protein coat remains outside. Phages generally attach to a cell using a tail fiber that interacts with LPS. When tail pins make contact with the cell wall, a lysozyme-like protein makes a hole in the cell wall allowing for the injection of viral DNA

3) Replication where in the lytic cycle phages take over the host cell machinery. This involves the synthesis of viral enzymes, nucleic acid replication, synthesis of protein coats, assembly and packaging (not the most exact process), and release and lysis of host cell

Question 4

What is typically observed when mapping a viral growth curve within a cell?

Answer 4

1) First there is an adsorption period where a virus is added and attaches to host cells

2) Then there is a latent period where early enzymes, nucleic acids, and protein coats are formed, and this is the eclipse phase

3) Then rapid growth occurs during the maturation phase where assembly and release of viruses then occurs

Question 5

What are two enzymes viruses use in the “replication” process?

Answer 5

Lysozyme: an enzyme that makes holes in the bacterial cell wall allowing replicated virions to escape and genetic material to enter the cell

Reverse transcriptase: an enzyme in RNA viruses that replicate inside the host as DNA intermediates. Reverse transcriptase transcribes the viral RNA into a DNA intermediate

Question 6

What is the purpose behind a plaque assay and why is there so much variation in the results?

Answer 6

Researchers use a plaque assay to create a lawn of host cells and then look for zones of clearing (plaques) that indicate viral infection to generate a virulence factor

First phages and bacterial cells are poured onto an agar plate with nutrients and the mixture is sandwiched between a top agar and nutrient agar. Then phage plaques are usually observed within a lawn of host cells.

Each plaque theoretically represents one viral particle in the initial infection, and there is much variation since the efficiency of plating (no. of plaques per virion) and culture conditions vary along with virulence and characteristics of host cells

A viral titer is used to quantify the concentration of virulent viruses, and it refers to the concentration of virus suspension as measured by the number of plaque forming units (pfu)

Question 7

How can bacteriophages set up a state of coexistence with the host and become lysogenic (temperate phages can do this) rather than lytic (damaging to the host)? Why would viruses undergo the lysogenic pathway?

Answer 7

Bacteriophages have a lytic pathway where viral DNA can replicate, assemble, and then lyse the cell (rightward transcription, encodes for capsid/phage assembly) or they can undergo the lysogenic pathway where viral DNA can be integrated into host DNA (leftward transcription, encodes for excision/integration). A virus may undergo the lysogenic pathway when host cell replication is more likely to occur or when conditions aren’t favorable for the survival of viruses.

Question 8

How does the bacteriophage lambda integrate into the bacterial genome? How does it make the choice for lytic vs lysogenic pathway?

Answer 8

A site specific nuclease creates staggered ends of a phage and host allowing for the integration of lambda DNA, and gaps are then closed by DNA ligase.

Question 9

What genes are used in the process of choosing between lytic vs lysogenic pathway?

Answer 9

cI expression leads to lysogeny and cro expression leads to the lytic cycle. A lambda regulator region or “switch box” with three operators are used in this process.

The lambda repressor generally exists as a dimer where the N terminal domain binds to phage DNA and has a highest affinity for OR1 on the lambda genome. Cro additionally exists as a dimer and can bind to all three operators but has the highest affinity for the OR3 region.

Question 10

How does negative control of the lytic pathway and positive control of the lysogenic pathway using the lambda repressor occur?

Answer 10

Negative control occurs when the lambda repressor binds to OR2 since this prevents the transcription of PR and the cro transcript downstream. This is required for lytic growth.

Positive control occurs as the repressor at OR2 helps RNAP bind and increases the transcription of PRM and cI downstream approximately ten-fold, activating the lysogenic pathway.

Question 11

How does cooperative binding induce the expression of the lysogenic pathway?

Answer 11

The lambda repressor has the highest affinity for OR1, and this stimulates repressor binding to OR2 via protein-protein interactions between dimers. Repressor binding to OR2 induces lysogeny by blocking the expression of cro and lytic genes and stimulating RNAP binding and expression of the lysogenic pathway.

As the lambda repressor concentration increases, repressor binding to OR3 occurs via cooperative binding to a dimer far away. This is accomodated by DNA looping and it can cause lambda repressor to temporarily block its own expression.

Question 12

How does Cro induce the expression of the lytic pathway?

Answer 12

Cro has the highest affinity to the OR3 region and this blocks the expression of the lysogenic pathway via repression along with the expression of the lambda repressor, and this allows RNAP to bind to OR1. If the repressor binds to OR2 first, expression of Cro is blocked and RNAP can transcribe lysogenic genes.

If Cro wins the lambda enters the lytic cycle and if a repressor wins the host becomes a lysogen. If Cro concentration is too high it can shut off its own expression by binding to OR1 and preventing lytic transcription.

Question 13

How can the host defend against viral lyric infection?

Answer 13

The presence lambda repressors in the cytoplasm of a host can render it partially immune to infection by a lambda phage by blocking the lytic cycle.

Question 14

How is the phage able to induce the lytic cycle in times of stress?

Answer 14

In times of stress the phage needs a way to induce the lytic cycle and escape the host cell. DNA damage tends to occur from heat, UV radiation, etc. and this induces the SOS Repair system which activates a recA protein from host DNA. recA can cleave the lambda repressor preventing it from cooperatively binding as a dimer and this causes the expression of more lytic genes. This allows phages to quickly assemble and release from the cell escaping the stress inducing conditions.

Question 15

How can CRISPR serve as a “bacterial immune system” and retain a memory?

Answer 15

CRISPR/Cas systems can target either viral DNA or RNA and can interfere with viruses, plasmids, prophages, or other chromosomally encoded sequences within bacteria

When phage DNA or foreign DNA is detected, a Cas complex detects the foreign material and places a small segment of the foreign DNA into the leader end of a CRISPR locus which is then flanked by a CRISPR array to form a repeat spacer unit.

This repeat spacer is transcribed and processed into guide RNA, and this guide RNA binds to a nuclease known as CAS9. CAS9 uses this guide RNA to target foreign DNA and cleave it upon second exposure.

The PAM serves as a recognition site that stabilizes CAS9 temporarily during cleavage.

Question 16

How can CRISPR/Cas9 be used for gene editing?

Answer 16

1) An RNA guide molecule can be programmed to match any unique DNA sequence in the human genome

2) Cas9 can then be attached to the RNA guide and find the DNA target sequence

3) The RNA can then align with the target DNA sequence and Cas9 can then attach to and cut both strands of the DNA double helix

4) DNA cuts can be amended with an extra DNA insertion made up of added homologous DNA with a desired mutation. Mutations can also be induced since broken DNA can be repaired in an error prone process

Question 17

What are some differences between prokaryotes and eukaryotes, specifically when looking at mRNA and cell walls/membranes? Why is this important when looking at how eukaryotic viral infection differs from prokaryotic?

Answer 17

Transcription in prokaryotes is not compartmentalized and transcription is immediately followed by translation. DNA replication can also take place in the cytoplasm and no mRNA processing is required for translation.

In eukaryotes, DNA replication and transcription is limited to the nucleus and mRNA must be processed for transport out of the nucleus and translation is associated with a ribosome. There is a poly-A-tail on the 3’ end, a methylated guanosine triphosphate “cap” on the 5’ end, and splicing

Question 18

How does viral infection differ between bacterial cells and animal cells?

Answer 18

The bacterial cell wall tends to be rigid, and this often means that the viral genome is injected into cells during infection leaving a coat on the outside. But in animal cells no wall is present and the uptake of viruses generally takes place via endocytosis, meaning a whole viral particle usually enters the cell

During uptake, nucleocapsid of virus is uptaken into an animal cell via endocytosis and the viral envelope is lost. Eventually the capsid is uncoated and a viral nucleic acid allows for virus multiplication and other related processes

Question 19

What are some consequences of viral infection? How can they cause tumors or cancers after initial infection?

Answer 19

If progeny viruses are produced, cytopathic effects can occur where a cell dies after many virions are produced

Cells can also survive in the presence of progeny viruses and shed a low number of virions leading to chronic infection

Viral genome can also persist in the host cell leading to latent infection

Some viruses that have a genome that persists in the host cell and can “transform” animal cells post infection leading to uncontrolled proliferation. They are hypothesized to do this by blocking the activity of growth factors that can normally prevent growth and division in mature cells. This can contribute to cancer.

Question 20

What are challenges that eukaryotic viruses run into?

Answer 20

They need to get inside the cell

DNA replication and transcription is limited to the nucleus so it is hard for viral DNA to enter the genome or be expressed

mRNA must also be processed in order to associate with a ribosome and allow for protein production, and this involves a poly-A tail on a 3’ end, methylated guanosine triphosphate “cap” on the 5’ end, and splicing

Question 21

What are some characteristics of retroviruses and what is its replication cycle? What are three polyproteins that a viral mRNA can translate and what are each of their roles?

Answer 21

They contain an outer surface envelope protein and a lipid bilayer with transmembrane envelope proteins contained within them. They integrate into the host genome using RNA and replicate via a DNA intermediate using reverse transcriptase that is stored in the virion.

When the virus enters the host cell cytoplasm the genome is reverse transcribed into single stranded DNA which is then converted to double stranded DNA and integrated into the host genome.

The transcription of viral DNA then leads to the formation of viral mRNA and progeny genomes, and the viral mRNA is then translated into three polyproteins: Gag, Pol, and Env, and these are then processed into components of the viral capsid.

Gag is processed into the attachment proteins of the viral capsid, Pol is processed into a protease (cleaves amino acid chains), integrase (catalyzes integration of viral DNA with genome), and reverse transcriptase, and Env is processed into gp41 and gp120 which mediate the binding of viruses to host cells.

They are also potential vectors for gene therapy since they can integrate into the chromosome

Question 22

How does replication occur in retroviruses?

Answer 22

1) Retroviruses enter the host cytoplasm via endocytosis and uncoat in the cytoplasm allowing free viral RNA to be present in the cytoplasm

2) Reverse transcriptase forms ds DNA from the single strand of viral RNA which then travels to the nucleus and integrates with the host DNA

3) Host machinery can then produce viral mRNA from the integrated DNA which forms three polyproteins

4) Polyproteins are used to form a nucleocapsid around viral mRNA and progeny viral RNA

5) Encapsulated viral particle then buds and releases from the host cytoplasmic membrane via exocytosis

Question 23

What does reverse transcriptase do in retroviruses?

Answer 23

1) It can catalyze the synthesis of ssDNA with an RNA template

2) It can catalyze the synthesis of dsDNA from the synthesized ssDNA

3) It has ribonuclease H activity and can degrade the RNA strand of an RNA:DNA hybrid after DNA is synthesized from RNA

4) tRNA is used as a primer for reverse transcription

Question 24

What is the infection cycle of HIV? What proteins and receptors are involved in HIV infection?

Answer 24

HIV attacks Th cells and enters through tears/lesions in the mucosal layer. It replicates in monocytes and Th cells and numbers of these cells can decrease below 200 per mL (500-1000 is the normal level) after an asymptomatic period that can last up to 2 years. As Th cell numbers drop, infected individuals can exhibit an increased sensitivity to infection by other pathogens and eventually weight loss, diarrhea, sarcoma, cancer, dementia, etc.

1) First attachment involves the interaction between a gp120 protein and the CD4 and CCR5 on the surface of host cells (T helper cells and monocytes). HIV resistance can be linked to variations in CCR5

2) Then the fusion of the viral envelope with the host cell membrane is mediated by gp41

3) The nucleocapsid can then insert into the host cell cytoplasm via endocytosis and allow the genome to be reverse transcribed. Host cell machinery can be used to form viral mRNAs and synthesize the virus within the host cell

Question 25

How is HIV transmitted and what are key symptoms?

Answer 25

It enters through tears/lesions in the mucosal layer and can be transmitted through bodily fluids. Characterized by having a CD4 T cell number of less than 200/mL or one infection such as TB, wasting syndrome, pneumonia, etc.

Question 26

What are key characteristics of negative strand viruses and what does their genetic material look like? What are three examples?

Answer 26

They are single stranded RNA viruses and reproduce via exocytosis from a host cell where particles budding from a host cell membrane become enclosed in a membrane envelope. The RNA strand is negative and therefore complementary to the mRNA strand so host replication needs to occur in order for it to be transcribed. Rabies, Ebola, and Influenza are examples.

Question 27

How does replication occur in negative strand viruses?

Answer 27

When the virus is inside the host cell, the nucleocapsid migrates to the nucleus and the negative viral RNA molecules are transcribed to positive strand mRNAs using host primers. This is accomplished using viral endonucleases (catalyzes mRNA production and processing) and replicases (catalyzes the production of an mRNA strand from an RNA template from the virus). The replicase can also be used to catalyze the production of a second negative RNA strand using a positive strand mRNA as a template and more mRNA can be produced. Poly-A tails are then added to the viral mRNAs and are also transported out of the nucleus. The mRNA molecules produced from the viral negative template strand are translated and this leads to the production of structural proteins and the assembly of the nucleocapsid. The virus then assembles in the host cell cytoplasm and buds from the host cell.

Question 28

What are key characteristics of positive strand viruses and what does their genetic material look like? Give three/four examples?

Answer 28

They contain single strand RNA that can be coded into polyproteins and they tend to be small and consist of an icosahedral structure. 4 examples are poliovirus, rhinoviruses, hepatitis A, and corona viruses.

Question 29

How does replication in positive strand viruses occur?

Answer 29

The RNA contained within positive strand viruses can be directly translated into a polyprotein which forms viral proteins using host cell machinery. They can also synthesize minus strands which can be used to synthesize new plus strands. All of this takes place in the cytoplasm.

Question 30

What virus is responsible for the majority of colds? What does it mean if a virus is zoonotic and what are three zoonotic coronaviruses that have been deadly for humans?

Answer 30

Human coronaviruses cause approximately 30% of colds and three zoonotic viruses that spread between animals and humans in particular have been deadly: MERS-CoV (which caused Middle East Respiratory Syndrome), SARS-CoV (beta coronavirus that causes severe acute respiratory syndrome) and SARS-CoV-2 SARS-CoV in particular has a high fever, aches, respiratory symptoms, difficulty breathing, dry cough and transmission is believed to require direct contact with a 5.1% fatality rate.

Question 31

What is the infection cycle of coronavirus? What proteins/receptors are involved and how are they significant? What are two routes of entry and what are the specific roles of RNA-dependent RNA polymerase and three polyproteins that are initially translated?

Answer 31

1) Binding to a membrane fusion protein and viral entry via endocytosis occurs. For SARS-CoV-2 ACE2 is the primary membrane fusion receptor protein. There are abundant amounts of ACE2 on type 2 pneumocytes in the alveoli since ACE2 is important for regulating blood pressure, wound healing, and inflammation 2) The major route of entry is via the TMPRSS2 pathway where the ACE2 binds to the SARS-CoV-2 spike protein and TMPRSS2 (Transmembrane Serine Protease 2) cleaves this protein to allow for efficient fusion between the viral and host cell membrane and viral entry. There is also an endosomal or cathepsin pathway where Cathepsin L cleaves the spike protein 3) The positive strand can be translated by a host and also replicated via RdRp, which replicates positive mRNA into a negative strand template which can be used to form more positive strands, and polyproteins/viral components can be synthesized 4) A polyprotein can be encoded using the positive mRNA that become many smaller proteins including a protease, an RNA dependent RNAP, a Helicase, a Spike protein, and several accessory proteins such as an envelope, membrane, etc. The new viruses are then released via exocytosis

Question 32

What is the role of the Nsp15 endonuclease in SARS-CoV-2 infection, and what kinds of infections are interferons used in?

Answer 32

It inhibits the translation of immune signaling molecules by sitting at the entry tunnel of ribosomes and cleaving host mRNAs but ignoring viral mRNAs and allowing them to be translated due to the presence of a specific leader sequence. This has the effect of blocking the translation of immune signaling molecules such as Type I and Type III interferons.

Question 33

How does SARS-CoV-2 cause COVID symptoms?What conditions are risk factors for COVID and why? How do dexamethasone and Paxlovid work to mitigate COVID symptoms?

Answer 33

Shortness of breath is a key COVID symptom. This occurs since SARS-CoV-2 binds to an ACE2 receptor on lung epithelial cells infecting them and infected cells signal the immune system and activate CD8 T cells, macrophages, etc causing inflammation. This inhibits the transfer of oxygen in the alveoli and fluid can also build up in the lungs. These factors cause shortness of breath. Conditions that are associated with high levels of inflammation can lead to severe symptoms, such as old age, diabetes, autoimmune diseases, obesity since body is "primed" to respond more strongly to SARS-CoV-2. Dexamethasone - corticosteroid that reduces inflammatory overstimulation that is caused by SARS-CoV-2 and prevents collateral damage caused by an infected individual's immune system Paxlovid - contains two drugs 1) Ritonavir which can be used to slow the metabolism of antivirals in the liver since a CYP inhibitor prevents the antivirals from being metabolized too quickly by CYP enzymes in the liver 2) Nirmatrelvir which inhibits the viral protease that can separate the polyprotein precursor into individual proteins

Question 34

How does the mRNA vaccine work in detail?

Answer 34

A vaccine is produced using mRNA that encodes for the Spike protein that binds to an ACE2 receptor as an antigen. Upon infection, this protein can then be more easily targeted by immune cells.

Question 35

What is happening with the SARS-CoV-2 variants? What is the S-fuse assay and how is it involved in infection and research?

Answer 35

Under an S-Fuse Assay, a virus is plated with host cells and the GFP protein is split meaning fluorescence occurs when a virus, SARS-CoV-2, is fused with and ACE2 receptor on the host cell. The level of fluorescence can be used to estimate the virulence and level of binding. Variants have been shown to bind to ACE2 more strongly in an assay indicating it is more virulent.

Question 36

Define the terms endemic, epidemic, pandemic, prevalence, incidence, morbidity, and mortality

Answer 36

Endemic - disease that is always present at a low frequency in a population Epidemic - a larger than normal frequency of infection, likely to to rapid and direct human-human transmission Pandemic - epidemic that occurs over a wide geographical area Prevalence - total number of active cases Incidence - number of new cases Morbidity - having an illness Mortality - death from an illness

Question 37

What are some steps epidemiologists take to stop an epidemic?

Answer 37

1) Track diseases through systematic surveillance (reporting) or syndromic surveillance (tracking/observing emerging diseases) 2) Identify an outbreak 3) Track down a patient zero (index case) and identify close contacts 4) Determine disease etiology for emerging diseases (cause, transmission, infection) 5) Develop interventions/treatments

Question 38

What are three categories of drugs against HIV and what are their mechanisms?

Answer 38

1) Nucleoside analogs that inhibit reverse transcriptase-AZT (azidothymidine) are one example. Analogs can be added to a growing DNA strand synthesized by reverse transcriptase but prevent the addition of the next base and this prevents integration of viral DNA with genome 2) Nuceloside reverse transcriptase inhibitors interact directly with the reverse transcriptase enzyme that alter the catalytic site 3) Protease inhibitors bind to and block the activity of HIV protease preventing polyproteins from being processed The most effective therapy for HIV has been using several different drugs (usually a protease inhibitor and two nucleoside analogs)

Question 39

What issues are associated with HIV treatment and what are some barriers to treatment?

Answer 39

It is expensive (approximately $10,000 per year per patient) so a large number cannot afford treatment. There are also many side effects such as fatigue, anemia, body changes, neuropathy, etc. Patients who discontinue also have an immediate rebound in viral levels and drug resistant strains can also develop. Some barriers include politics (leaders can deny the presence of a health crisis) and false science/pseudoscience which can become widespread

Question 40

How can PrEP be used in the treatment/prevention of HIV?

Answer 40

It is called pre-exposure prophylaxis and it is a medication that reduces the risk of contracting HIV from sex (~99% decrease) and from IV drug use (~74% decrease). PrEP is administered by taking two oral medications and one approved weekly shot. Some side effects include a headache, nausea, apetite loss, possible kidney/liver damage. Different from PEP or post-exposure prophylaxis.

Question 41

What does 90/90/90 mean?

Answer 41

It is a UN target where 90% of infected of individuals should be aware of their HIV status, 90% of those individuals are on HIV treatment, and 90% of those individuals are virally suppressed.

Question 42

Why are vaccines difficult to develop for retroviruses, specifically in the case of HIV? What is the aim of Phase III trials in South Africa and what are the targets of these studies?

Answer 42

1) Live attenuated viruses can integrate into a host cell's DNA and cause disease 2) HIV has a high mutation rate and it is difficult to make a vaccine against all clades 3) mRNA vaccine technology is limited since HIV along with many other retroviruses are so diverse meaning multiple proteins will have to be targeted and a lot of mRNA will have to be used but the amount of mRNA that can be incorporated in a vaccine is limited

Question 43

What did the Frontier Phase III trials in South Africa suggest about the use of vaccines for HIV?

Answer 43

HVTN 702 trial enrolled 5400 HIV-uninfected people at risk for infection to test the safety/efficacy of the RV144 HIV vaccines. Two vaccines were tested, the ALVAC-HIV containing a viral vector with 3 HIV genes and a protein vaccine supplied by GSK which should create an immune response to a protein GP120 which facilitate entry into host cells. This was the most successful HIV trial to date and the efficacy was 31% against a single HIV clade.

Question 44

What are the symptoms of Ebola and how is it transmitted? What are isolation and safety protocols and what was significant about the 2015 Ebola outbreak?

Answer 44

It is highly lethal where 50%-90% of infected individuals die usually due to organ failure/tissue death from inflammation. Symptoms are fever, bleeding, vomiting, muscle/joint pain, skin rash, diarrhea. Fruit bats are considered to be a vector and it is transmitted via bodily fluids, consumption of infected meats, contact. Incubation is a 2-21 day period In 2015, African health care workers were unfamiliar with the disease and mistook it for Lassa Fever/Malaria. There was a rapid spread to cities, stocked/staffed health care facilities were not present, WHO and other groups did not recognize the serious nature of the outbreak, and there was a failure of some to respond. The 2015 Ebola outbreak was widespread wheras in the past it was limited by scope and geography

Question 45

How can antibody therapy be used to treat Ebola? Is there an Ebola vaccine and does it work?

Answer 45

Ebola patients were recently treated with antibodies from survivors which was shown to be effective, and a Canadian company is developing a way to produce antibodies for Ebola in plants. rVSV-ZEBOV is also a recombinant vaccine that consists of a vesicular stomatitis virus which is engineered to express a glycoprotein from Zaire ebolavirus to provoke an immune response to Ebola and it was approved by health care workers in 2019.

Question 46

What are NTDs? How many people are affected by them and what are risk factors?

Answer 46

WHO defines them as disease that share a set of features that allow them to persist in conditions of poverty where they cluster and frequently overlap. Over 1 billion people suffer from one or more

Question 47

What is dengue/hemorrhagic fever? What type of mosquito usually transmits it, and how many viruses cause dengue? What are the moderate and severe symptoms?

Answer 47

It is a mosquito-borne disease and half the world's population lives in areas where dengue viruses can be transmitted. It tends to be transmitted by the Aedes agytpi mosquito. There are four distinct but related viruses and recovery from one provides lifelong immunity to that one but only partial protection from the other three. Vector control is the main area of prevention/control. Infants/young children may have a fever with a rash and older children/adults may have either a mild fever or a disease with a severe headache, pain, muscle/joint pains, rash. In severe cases it can lead to convulsions, liver enlargement, and haemorrhagic phenomena (bleeding through sweat glands), and circulatory failure

Question 48

What is the lifecycle of dengue (from mosquito to person and back to mosquito)?

Answer 48

Mosquitos usually acquire the virus after feeding on the blood of an infected person. After the virus is incubated for 8-10 days, an infected mosquito can transmit the virus to susceptible individuals for the rest of its life, and female mosquitos can also transmit the virus to offspring which can infect humans. Virus circulates the blood for 2-7 days (when they have fever) and this is when mosquitos acquire the virus. Dengue can also be transmitted from mother to child during pregnancy.

Question 49

How is dengue treated and what are some of the best prevention methods? How is this impacted by the increase in Aedes aegypti?

Answer 49

No treatment but symptoms can be managed and with intensive therapy, mortality can be reduced to less than 1%. Maintenance of fluid volume is most critical but pain/fever can also be treated. Vector control via environmental management and chemical methods have also been key. This includes solid waste disposal and improved water storage practices such as covering containers to prevent egg laying by female mosquitos. Led by community based programs Vaccine was also approved for children in 2022 for those with previous infection in areas where dengue is endemic

Question 50

What is Dracunculiasis and what is the life cycle of this parasite? How does infection occur and what symptoms does it cause?

Answer 50

It refers to guinea worm disease and it is transmitted using a flea as an intermediate host. Symptoms are a fever, rash, nausea/vomiting, diarrhea 1) It enters the body after a person drinks water containing water fleas that are infected with guinea worm larvae 2) The fleas are digested and the larvae are released into abdominal tissues where they can mate and develop 3) A female worm moves through the body, generally lower body, and grows forming a blister, and it can grow up to 3 feet long 4) The worm emerges from the blister it creates leaving the body and the victim rushes to cool the blister in water 5) Upon contact with the water, the worm releases clouds of larvae 6) The water fleas consume the worm larvae, but the larvae resist digestion and remain in the water fleas

Question 51

How is guinea worm disease treated and what are prevention measures?

Answer 51

There is no vaccine or medicine to treat or prevent this. Once the worm emerges from a blister, a health worker has to wrap the live worm around a piece of gauze little by little and this takes several visits Prevention programs have been shown to be effective and the most effective method is filtering out tiny water fleas from drinking water (pipe filters and small straw-like personal filters have been effective). Now cases are down 99% since 1986 and it is set to be the first parasitic disease to be eradicated since humans are a worm's only host.

Question 52

What are the six diseases targeted for eradication?

Answer 52

Dracunculiasis Poliomyelitis Mumps Rubella Lymphatic filariasis Cysticercosis

Question 53

What is parasitology and what is the global situation of parasitic diseases, why are they relevant? What are four main categories of parasites and what are three examples of soil-transmitted helminths?

Answer 53

Parasitology - the discipline concerned with the study of parasites and their relationships with their hosts Parasitic diseases are widespread, but there is relatively little research and it tends to not be well treated Four main types of parasites are Protozoa, Helminths, Ectoparasites, and Bacteria (not a traditional parasite) Soil-transmitted helminths are parasitic worms that infect their hosts through fecal contaminated soil. Three of them are Large roundworm, Whipworm, and Hookworm

Question 54

What was the main objectives hypotheses, and sample of the Schuar study?

Answer 54

Market Integration - the degree to which people consume from and produce for the global market economy Hypotheses - Two hypotheses were that infection risk will be highest in less market integrated Shuar and that parasite infection risk and intensity would be highest among participants that report factors that are linked with parasite transmission pathways (floor material, water source, bathroom type, H-SOL factors) Sample - There were 620 participants in 152 households across 10 communities Key measures - Kato-Katz thick smears were used to calculate species-specific infection status and eggs per gram which were key measurements of parasite load

Question 55

What was the infection prevalence for A. lumbricoides and T. trichiura and how did different factors affect infection risk and intensity? How did this compare to the hypothesis?

Answer 55

Infection prevalence - General infection (63%), A. lumbricoides (49%), T. trichiura (37%), Coinfection (27%) Infection risk/Intensity - More prevalent in children. More market integrated households had lower levels of infection and lower intensity as hypothesized. Water source and floor material were significant determinants of infection but bathroom type was not so support for hypothesis 2 was mixed Implications - This points to heterogenous effects of market integration and looks at other factors that are closely linked with disease risk Future directions - Researchers can collect environmental samples to better quantify eggs and parasite presence, collect data from Shuar living in more market integrated areas, test parasite patterns in relation to ongoing market integration over time

Question 56

What was the main objectives, methods/sample, results, and implications of the REACH study? What did the preliminary results from 2022 show and how does that compare to the "pilot results" from 2019? What are future directions?

Answer 56

Objectives - test whether intestinal infections are linked with elevated inflammation and household infrastructure, assess links between infection sources and parasitic diseases, test links between microbiota and intestinal parasite species, develop resources to remove inequities/barriers Sample - 91 adults, 49 in Mississippi and 42 in Illinois Methods - household interviews, using fecal calprotectin from stool samples to measure inflammation, using 18s rRNA sequencing to assess parasite infection rates (2019) and ELISA immunoassays to test H. pylori levels (2022) Results summary - living conditions associated with low-socioeconomic status are linked with high intestinal inflammation levels, higher levels of H. pylori. 2019 data suggests these were also linked with greater incidence of parasite infection. Implications - Altered immune profiles due to low socioeconomic status could result in increased pathogenesis and chronic health issues later in life, which could contribute to health inequities Future directions - Environmental hazard data, soil/water samples, gut microbes and parasite link, drivers of inflammation, comparative data in high income communities, develop community partnerships

Question 57

What does this study suggest about the web between environmental exposure, chronic psychosocial stressors, and the immune system?

Answer 57

Chronic psychosocial stress, limited access to resources, and increased pathogen exposure are associated with low socioeconomic status and are linked with immune system dysregulation This is linked with an increased risk of pathogenic infection which can cause a loss of "good" bacteria and more "bad" bacteria along a greater inflammatory immune response (also partially due to more toxin exposure) The increased proliferation of "bad" bacteria and increased inflammatory response leads to gastrointestinal inflammatory conditions which contribute to damaged gut structure/function and malnutrition, and these perpetuate health inequities

Question 58

What are some biosynthetic processes that antibiotics target?

Answer 58

- Cell wall synthesis - DNA gyrase - DNA directed RNA polymerase - Protein synthesis (30S and 50S inhibitors) - Cell wall - Cytoplasmic membrane structure - Folic acid metabolism - Cytoplasmic membrane

Question 59

What role do antibiotics play in treating infection?

Answer 59

They target and kill growing cells and give your immune system a fighting chance against a pathogen. When an antibiotic is present a very small number of cells survive and then when antibiotics are cleared, microbes repopulate

Question 60

What are some generalized resistant mechanisms that bacteria have developed against antibiotics?

Answer 60

- Efflux pumps that pump out antibiotics - Decreased uptake of antibiotics - Inactivating enzymes - Generating alternative enzymes that antibiotics don't target - Target alterations

Question 61

How do B-lactam antibiotics function? What are some examples and what kinds of bacteria do they target?

Answer 61

They serve as substrate mimics and bind to active transpeptidases that are essential for crosslinking glycan linked peptide chains in the bacterial cell wall permanently inactivating them (Penicillin and Ampicillin)

Question 62

What are B-lactamases and how does augmentin and cephalexin function to combat B-lactamases?

Answer 62

B-lactamases are bacterial enzymes that break the B-lactam ring of an antibiotic and inactivate B-lactam antibiotics Augmentin is a combination of amoxicillin, a B-lactam antibiotic, and Clavulanic acid, a B-lactamase inhibitor. Clavulanic acid binds to B-lactamase irreversibly via its B-lactam ring which protects amoxicillin from denaturation. Typically prescribed for infections that do not respond to other antibiotics Cephalexin is similar to a B-lactam antibiotic but resistant to typical B-lactamases

Question 63

How do fluoroquinolones function as antibiotics? What kinds of bacteria do they target and what are some examples?

Answer 63

They are synthetic compounds that interfere with gyrase and prevent the packaging of bacterial DNA. They also inhibit topoisomerase IV which prevent DNA replication and cause rapid bacterial death. Quinolones generally function against a wide range of bacteria and they are frequently used to treat non-complicated infections (UTI, traveler's diarrhea, etc) Resistance can occur when mutations in bacterial topoisomerase IV and DNA gyrase prevent fluroquinolone from binding and allowing genome replication to continue. It can also occur when bacterial modifications to an efflux pump allow bacteria to pump fluoroquinolone out of the cell (via target modifications and efflux pumps)

Question 64

How do macrolides function as antibiotics? How can resistance develop? What kinds of bacteria do they target?

Answer 64

They are protein synthesis inhibitors that act on a 50S ribosomal subunit and inhibit the formation of a peptide bond. They contain lactone rings that are connected to sugar moieties. Resistance occurs via efflux pumps and ribosomal target modification

Question 65

Why is antibiotic resistance a problem?

Answer 65

1) Bacteria are everywhere (hospitals, farms, organisms, etc.) 2) Antibiotics can kill bacteria that cause infection but this can have the effect of selecting for resistant bacteria 3) These resistant bacteria can multiply and some can transfer resistant genes to other bacteria 4) Antibiotics can also kill helpful bacteria that protect us causing more resistant microbes to circulate 5) Antibiotic resistant microbes can spread to new settings and transfer genetic material to even "harmless" microbes

Question 66

How can resistant genes spread between bacteria (specifically)?

Answer 66

Resistance genes can be transferred using plasmids via conjugation (DNA circles that can move between cells), transposons via transformation (from nearby dead organisms) and conjugation (pieces of DNA that go into a cell and change the overall DNA), and phages via transduction (viruses that attack germs and carry DNA from germ to germ

Question 67

What are the mechanisms of phage therapy and what is its intended function? Is this a viable solution to antibiotic resistance?

Answer 67

The aim is to isolate and/or engineer bacteriophages to attack bacterial pathogens, specifically resistant organisms. There has been some success but it is still in its early stages

Question 68

What is the mechanism and function of vaccines?

Answer 68

They are modulated by Th2 stimulation of B cell proliferation. When a vaccine is administered, a primary response occurs when Th2 cell is activated when an APC presenting MHC II activates a B cell with the same MHC II antigen complex on its surface Direct B cell stimulation then occurs where upon the second encounter with an antigen after a pathogen invades, an antigen binding to antibodies on a B cell surface activate a B cell directly and this initiates a rapid response Antibodies function via opsinization, where a toxin or virus binds to a receptor on a host cell, when a complement cross-links antibodies that are attached to a microbe, the complement becomes activated, and microbes can also be trapped in mucin

Question 69

Primary vs Secondary Response

Answer 69

Primary - following antigen introduction there is a latent period before an antibody appears in the blood. Then a gradual increase in antibody titer and a slow fall occurs Secondary - upon second exposure to an antigen, the antibody titer rises to 10-100 times of the primary response. There is also a class switch from IgM to IgG antibodies

Question 70

Herd immunity

Answer 70

Vaccinating a population can protect unvaccinated individuals via herd immunity since vaccines reduce the incidence and prevalence of a disease reducing the transmission

Question 71

What are four different types of immunity and how are they acquired?

Answer 71

Natural Active - infection renders an individual immune Artificial Active (Immunization) - injection of antigen to generate antibodies Artificial Passive - injection of antiserum from individual who has developed active immunity Natural Passive - when newborns retain IgG antibodies in their bloodstream

Question 72

What are 4 traditional types of vaccines?

Answer 72

Killed or inactivated pathogens - made from whole organisms (silk polio) Attenuated - organism has been cultured to reduce pathogenicity but it retains its ability to infect and can induce the immune system (sabin polio) Toxoid - denatured toxin is used to make the vaccine and antibodies are raised against the toxin and inactivate it upon infection (tetanus) Surface molecules - based on the idea that antibodies are most effective when they are directed at surface molecules (flu, COVID)

Question 73

How does the Sinovac, mRNA, Novovax, and Jannsen vaccines function against COVID?

Answer 73

Sinovac - consists of the inactivated virus. 66% effective at preventing symptomatic infections and 100% against severe COVID but not sterilizing (breakthrough infections can occur) mRNA - contains spike protein mRNA so that immune cells can target the spike protein Novovax - contains the spike protein along with an adjuvant to boost immune response Jannsen - contains an adenovirus vector where the code for the SARS-CoV-2 spike protein is carried inside the adenovirus. This adenovirus can enter the host cell and induce it to display spike. Hypothesized to induce a more complete response since it displays an antigen that is similar to that of an infected cell

Question 74

What are mumps? What virus causes them and what is the infection cycle?

Answer 74

It is a negative strand RNA virus spread by aerosols that replicates in salivary glands. Symptoms are a fever/headache, muscle aches, swollen/tender salivary gland under the ears, may also cause inflammation of pancreas Vaccine contains a live attenuated virus and it is typically given as part of the MMR vaccine. Community protection relies on the principle of herd immunity. There was an outbreak in 2006 due to vaccine failure and it is linked with a breakdown in herd immunity

Question 75

What is polio and what is its infection cycle? What is a barrier to achieving the complete eradication of polio?

Answer 75

It is a positive strand RNA virus that is transmitted between humans via the fecal-oral route, so it tends to disproportionately occur in areas with poor sanitation systems. It multiples in the intestinal tract and makes its way through blood and lymph. It destroys motor neurons resulting in flaccid paralysis and axon regeneration can restore muscle function. Antivax movements and poor sanitation systems serve as barriers to eradication.

Question 76

Who was Albert Sabin and Jonas Salk and what was their role in the development of polio vaccines? Compare and contrast their work

Answer 76

Albert Sabin - made a vaccine consisting of an attenuated virus that could provide lifetime immunity Jonas Salk - made a vaccine that could be given by mouth. This was easier to administer in countries with underdeveloped sanitation systems

Question 77

Bioremediation

Answer 77

The cleanup of oil or other pollutants by microorganisms (oil spills, pesticides, dichlorination, sewage treatment). Anaerobic bioremediation involves the use of chlorinated solvents that are hazardous and not easily degraded to remove chloride ions.

Question 78

Hydrocarbon decomposition

Answer 78

The process by which naturally occurring bacteria, fungi, and green algae are able to oxidize hydrocarbons. These can associate with oil, concentrating at the oil water interface and oxidize the oil to CO2. These bacteria develop on oil films and slicks.

Question 79

How can microbes be selected for hydrocarbon decomposition?

Answer 79

1) Specific mobility/adhesion mechanisms that attach to petroleum droplets 2) Extra-cellular emulsifying agents are able to survive low O2 environments 3) Cell-bound inducible enzymes are produced that depolymerize nutrient sources 4) Desorption mechanisms release from petroleum droplets 5) Biochemical pathway with an enzyme production that degrades target wastes 6) Bacteria can oxidize ~80% of nonvolatile components within a year and increase in number 10^3-10^6 shortly after a spill. Antimicrobial agents are actually needed in gas storage tanks to prevent degradation of fuel 7) Addition of nutrients such as phosphates and nitrogen to a spill area increases the rates of bioremediation

Question 80

Radioactive bioremediation

Answer 80

Bioremediation that occurs under radioactive conditions due to the use of radioactive microbes

Question 81

What is an example of a radioactive microbe and how does function?

Answer 81

D. radiodurans can withstand 1.5 million rads (700 rads is lethal for humans). In most organisms, exposure to high levels of ionizing radiation or UV lead causes extensive DNA damage, but D. radiodurans can repair its own DNA via a specialized DNA repair protein that binds to the ends of DNA generated from radiation and prevents damage from endonucleases.

Question 82

How were DNA repair genes identified in D. radiodurans?

Answer 82

Microarrays were used to identify a single ORF whose expression was induced 2-fold following the treatment of D. radiodurans with ionizing radiation. RecA, another DNA repair protein, was increased 1.5 fold under these conditions and the deletion of this ORF resulted in an increased sensitivity to DNA damage via radiation and a lower surviving fraction over time.

Question 83

How has D. radiodurans been engineered to detoxify waste?

Answer 83

A mercuric reductase gene from E. coli was cloned into D. radiodurans in order to detoxify the ionic mercury found in radioactive waste that is generated from nuclear weapon manufacture. This strain reduces Hg(II) to elemental mercury which is less toxic (it has also been used to detoxify toluene)

Question 84

What are the stages of wastewater treatment?

Answer 84

Primary treatment - consists of the removal of solids by passing through a series of screens and also letting an effluent stand to let solids settle to the bottom Secondary treatment - employs microbes to reduce the organic load of the wastewater to environmentally acceptable levels Tertiary treatment - uses chemical methods (precipitation, filtration, chlorination) to remove inorganics such as phosphate from wastewater

Question 85

How are bacteria used in wastewater treatment via anoxic secondary treatment?

Answer 85

A set of digestive and fermentative reactions that are carried out by different bacterial and archaeal species. Its efficiency is measured in terms of the percentage decrease in biological oxygen demand, and this refers to the amount of oxygen that would be consumed if all the organics in one liter were oxidized by bacteria, archaea, and protozoa (good BOD is 95%). This process takes place in "sludge digestors" or "bioreactors" During this treatment, macromolecules are broken down into soluble components (monosaccharides, amino acids, fatty acids) and these are then further broken down into acetate, H2, and CO2 which are substrates for methanogenic bacteria. The major products are CO2 and methane. Methane is either burned off or collected for use in electric generators to heat/cool the treatment plant.

Question 86

How do sludge digestors operate?

Answer 86

Digestor contains a sludge inlet where a supernatant layer, scum layer over it, gas later on top with a gas outlet, digesting sludge below it, and stabilized sludge at the bottom which moves through a sludge outlet. The scum and supernatant layers are also removed Complex polymers are hydrolyzed into monomers by microbial enzymes and acetate and H2 + CO2 are produced by fermentation. CH4 and CO2 are then produced via methanogenesis

Question 87

What are two methods of aerobic sewage treatment?

Answer 87

Tracking filter method - wastewater is sprayed onto a 2m deep bed of crushed rocks. As the liquid slowly trickles through this bed, organic matter then adsorbs into the rocks and allows for microbial growth on the surface. The microbes then mineralize the organic matter into CO2, NH4, nitrate, sulfate, and phosphate Activated sludge method - wastewater is mixed in a large tank and slime forming bacteria grow and form flocs which then attract protozoa, other small animals, and some bacteria/fungi. Effluent containing flocs is then pumped into a clarifier where flocs then settle and some of it is then returned to an incubator where it then functions as an innoculum. The rest of it is sent to a sludge digestor.

Question 88

What are some problems that come up with activated sludge treatment?

Answer 88

Generally not enough time is used for the complete oxidation of organic materials. Since the adsorption to the floc is the primary means of reducing BOD in the liquid, the main reduction of BOD occurs after the transfer of the floc to a sludge digestor In addition it is energy intensive, it removes nitrogen rom the water cycle and releases it in the form of N2O contributing to global warming, and it rarely produces portable water

Question 89

Thermophilic sludge digestion

Answer 89

In some sludge digestors head is added to the anaerobic digestion step, and this doubles/triples the energy recovery by producing more biogas but more nutrients are released so tertiary treatment is complicated

Question 90

How is water currently purified and how can these methods work with bioremediation?

Answer 90

1) Water is pumped from its source to sedimentation basins where gravel and sand can settle out 2) Water is transferred from the sedimentation basin to a coagulation basin and chemicals can be added to trap microorganisms and absorb organic matter, which can then be removed from the water 3) The water is clarified and filtered through thick layers of sand and ~99% of the bacteria from the original water had been removed at that point 4) Chlorination then eliminates the remaining organisms and oxidizes organic compounds which can neutralize them

Question 91

How do biofuels operate and what role do microbes have in this? What are some pros and cons of biofuels? What are two common biofuels?

Answer 91

Biofuel - conversion of biomass to liquid fuels, ethanol (sourced from corn and sugarcane) and biodiesel (sourced from vegetable oil, animal fat, or recycled cooking grease) are common biofuels that can be used in a similar way to gasoline Biomass - plant material, vegetation, or agricultural waste Microbes can be used to quickly generate biomass while producing desired products Ethanol cons - it can be toxic at high concentrations, an energy intensive purification process is required, and it does not have the full fuel value of gasoline. It is also not transported using traditional means

Question 92

What is the process of biomass to biofuel? What is lignocellulose and how is it relevant in the context of biofuels?

Answer 92

Lignocellulose - plant dry matter which is the most abundant raw material available on Earth, made up of carbohydrate polymers that are linked by an aromatic polymer lignin and it is a biomass that can be converted to fuels To convert biomass to biofuels, plants are used to make carbohydrate polymers, such as lignocellulose, and enzymes then convert these to monomers. Microbes can then be used to digest these monomers and form biofuels Challenges - Lignin is present in lignocellulose and it can be resistant to depolymerization. In addition, the composition of cellulose and hemicellulose can impact fuel production since cellulases are used to depolymerize but the process can be slow and there is a limited set that can be used. It also requires energy catalysts and big plants.

Question 93

What do first generation biofuels do and what are strengths and problems?

Answer 93

These are conventional biofuels that are constructed from food products such as sugar, starch, and vegetable oil. The biofuels produced are not structurally different from second or third generation biofuels the source just changes. Some problems are that they threaten the food chain, they can increase carbon emissions, especially when they are outside traditional agricultural settings, and they have intense growth requirements.

Question 94

What do second generation biofuels do and what are strengths and problems? What makes them unique?

Answer 94

They are "advance biofuels" produced from sustainable feedstock that is available, low impact on greenhouse gas emissions, low impact on land use, and minimally threatens the food supply To qualify for second generation, a feedstock should grow on marginal or non-agricultural land and it should not require a great amount of water or fertilizer. In addition food products can become second generation when they are no longer useful for consumption such as waste vegetable oil

Question 95

What do third generation biofuels do and what are strengths and problems? How are they generated?

Answer 95

This is an unofficial category and these biofuels are derived from algae. It was moved into its own category since algae are capable of higher yields with lower resource inputs and require a unique production mechanism. Third generation biofuels could also potentially offer solutions that can mitigate most of the drawbacks of 1st and 2nd gen biofuels.

Question 96

What is the microbial circular economy? What are two examples?

Answer 96

The aim of the circular economy is resource efficiency, optimizing the value of biomass over time, and sustainability. In a circular bioeconomy, biological waste products are recovered, recycled, and reused to produce goods and biofuels sustainably One example of this is wastewater treatment where wastewater treated microbial economies can produce carbon and nitrogen wastes that can be used for power generation and agricultural fertilizer. This creates a circular bioeconomy Another example is fish feed where microbes can be used to produce bioflocs from fish waste and carbohydrate supplements which can then be used to produce fish feed

Question 97

How does Klebsiella cause UTI infection? How is this process similar/different to E. Coli?

Answer 97

It is an important cause of nosocomial/opportunistic infections such as pneumonia, UTI, bloodstream infection (second leading cause behind E. Coli) It progresses through an intracellular bacterial communities (IBC) and filaments can be found in K. pneumoniae infected mice. It doesn't always express pili and attach to cells like E. Coli

Question 98

How do researchers (Dr. Rosen) combat Klebsiella antibiotic resistance?

Answer 98

Rosen lab first understands the mechanism behind infection along with the host's adaptive immune response to Klebsiella in order to develop treatments aimed at inhibiting virulence and eliciting adaptive immunity of human hosts The Rosen Lab has projects aimed at producing bioconjugate vaccines, studying innate and adaptive immunity such as T cell responses and effectors, and virulence factor regulation (FimK and RfaH)

Question 99

What does the BARNARDS study suggest about the link between antibiotic misuse and childhood mortality? What disparities does it suggest are present with antibiotics?

Answer 99

Neonatal sepsis was studied in 7 low/middle income countries: Bangladesh, India, Pakistan, Ethiopia, Rwanda, Nigeria, and South Africa. Overall incidence of sepsis was 47/1000 live-births and in comparison the US incidence was 1-2/1000 live births. Resistant Klebsiella pneumoniae accounted for >10% of culture cases. Points to a systemic failure to deliver appropriate antibiotic therapy since ampicillin-gentamicin was used as therapy for neonatal sepsis but only 3% of the isolate microbes linked to infection were susceptible to ampicillin and only 30% were susceptible to gentamicin. Effectiveness was higher when neonates were treated with ceftazidime-amikacin (93%)

Question 100

What does the BARNARDs study suggest about what we should do prevent antimicrobial resistance and childhood deaths?

Answer 100

1) Use antibiotics globally more effectively (antimicrobial stewardship programs, limiting agricultural use of antibiotics, using location specific guidelines for antibiotics, increasing access to diagnostic testing) 2) Increasing access to safe water and sanitation 3) Identifying which pathogens are responsible for childhood deaths 4) Vaccines

Question 101

What is a verbal autopsy and how can it be used to determine a cause and source of an illness? Is it complete?

Answer 101

A verbal autopsy has mostly been used to get data on the causes of childhood death where an oral interview is done on a mother/family member, but there are pitfalls 1) Data tends to be incomplete since data tends to be gathered less in the highest mortality countries 2) It tends to also be poor quality since it processes used to collect data are non-standardized and it is difficult to integrate analyses 3) In addition it is delayed where data is released every 2-3 years

Question 102

What are alternative methods of generating childhood mortality data?

Answer 102

CHAMP surveillance strategy - an alternative way of gathering childhood mortality data. Assessed using verbal autopsies, minimally invasive tissue sampling, cultures, clinical records, diagnostic testing, and DeCoDe panels (determination of cause of death) to assess for causation DeCoDe panels were based on demographic data, verbal autopsy, clinical abstractions from available medical records, maternal abstraction, MITS collection data, Molecular and other diagnostics, and pathology results

Question 103

How can vaccines be used to try and halt global antimicrobial resistance? How is a K. pneumoniae vaccine hypothesized to help this?

Answer 103

After PCV7 was introduced to children in the US, the incidence of invasive AMR-pneumococcal infection dropped 80% in young children and over 50% across all ages. This meant a decrease of 35 prescriptions for 100 unvaccinated children was present which is equivalent to 1.4 million less antibiotic prescriptions annually in children.

Question 104

What is hypervirulent K. pneumoniae (hvKp)? What are defining characteristics and how can we use testing/vaccinations to prevent its spread?

Answer 104

Hypervirulent Klebsiella strains can cause more community acquired infections with more severe symptoms There is no universally agreed-upon marker for hypervirulence but it is associated with 1) Hyper-mucoviscosity (capsule of bacteria contains more mucus, a string test can be used where a 5-mm string of mucus can be produced from a colony on a plate 2) More severe clinical symptoms (liver abscess, meningitis/epidural abscess, pneumonia, necrotizing skin, bacteremia) 3) Distinct sequences and capsules (K. pneumoniae encodes for 134 K types, mostly K1 and K2 and ST23 is enriched in hvKp strains) 4) More virulence associated genes

Question 105

What are virulence genes associated with hvKp and what roles do plasmids and ICEs play in the spread of these genes?

Answer 105

Some genes are a virulence plasmid with pK2044, pLVPK, 14 ICEs (integrative and conjugative elements), specific genes such as rmpA, magA, KvrA, KvrB, luc (aerobactin), Iro (salmochelin), colibactin toxin

Question 106

What is a conjugate vaccine and what is the goal of conjugate vaccines for K. pneumoniae? How can hvKp be targeted specifically as opposed to E. Coli? What roles do antibodies play in this?

Answer 106

Hypervirulent Klebsiella has K type capsular polysaccharides (>100) where 24 cover ~60% and 11 serotypes of O-antigen lipopolysaccharide on the cell wall where 4 cover ~80%. There is currently a phase I trial of a vaccine for hvKp in Germany containing 4 O antigens. Antibodies against K and O proteins assist in the killing of Klebsiella and this varies by strain.

Question 107

What are the advantages of bioconjugation over chemical conjugation in producing vaccines?

Answer 107

Conjugation is used to isolate a protein/compound for use in vaccines. Chemical conjugation is 1) Labor intensive - multiple rounds of purification and detoxification have to be used 2) Heterogenous product - a homogenous product is not formed so there is variation within each batch 3) Harsh chemicals are used which can alter/destroy polysaccharides 4) Biohazard - it is required for large batches of a pathogenic organism to be grown As opposed to chemical conjugation, a more purified bioconjugate can be obtained via glycoengineering in bacteria. This is cost effect and flexible, streamlined with it can be performed with lab safe E. Coli, and the supply is scalable. Klebsiella bioconjugates of K1/K2 capsules can be produced in E. Coli using a plasmid rmpA and possibly made into a vaccine. K vaccines can be better than O vaccines in some strains but a K and O combo is likely needed. In this process, a gene cluster encoding for polysaccharide synthesis proteins, a carrier protein, and a conjugating enzyme (oligosaccharyltransferase) can be transformed into E. Coli and a purified bioconjugate can be formed which can be made into a vaccine.

Question 108

What are some of the current predictions for a K. pneumoniae vaccine?

Answer 108

1) To prevent neonatal sepsis: broad vaccine given during pregnancy 2) To prevent hypervirulent infection: K1/K2 vaccine given to children in southeast Asia 3) To prevent antimicrobial resistant nosocomial infection: KL106/KL107/KL102 vaccine given to immunocompromised patients

Question 109

How are 16S rRNA sequencing, metagenomic sequencing, and transcriptomics/RNAseq/metabolomics used to study microbiomes?

Answer 109

16S rRNA amplicon sequencing - identifies which microbes are present Metagenomic sequencing - identifies which microbes are present and their functions Transcriptomics/RNA seq and metabolomics - identify the functions that microbes are actively doing

Question 110

What are the goals of the Human Microbiome Project?

Answer 110

1) Determine whether individuals share a core human microbiome 2) Understand whether or not changes in the human microbiome are correlated with changes in human health 3) Develop new technological and bioinformatic tools to support these goals 4) Address ethical, legal, and social implications raised by human microbiome research

Question 111

How does it go about achieving these goals?

Answer 111

1) Microbiome samples were collected from 300 healthy individuals 2) These samples were subjected to 16S RNA sequencing in order to understand the complexity of these communities 3) Metagenomics will be used to provide insights into functions that are preformed by the human microbiome (Sequencing center is at WashU genome center)

Question 112

What does a healthy microbiome look like? What does the lean vs obese phenotype study suggest about how our microbiome impacts our health?

Answer 112

A mouse study was conducted with two twins, one lean and one obese. When a microbial transplant from the obese twin was given to a recipient mouse, increased adiposity was present, but no change was observed when a microbial transplant was given from the lean twin. This suggests that our microbial communities impact our health and phenotype. Short chain fatty acids (SCFA) can also be by-products of microbial fermentation that can help the body. Some SCFAs can trigger anti-inflammatory pathways and promote health (butyrate and propionate)

Question 113

What are factors that impact microbial diversity at the population-level and individual-level?

Answer 113

1) Vertical transmission of microbes from mother to child serves as a "seed" for the microbiome 2) Priority effects can occur where the first colonized microbes shape the succeeding microbial community 3) Horizontal transmission via the environment/geography/parasite infection status shapes microbial diversity (environmental differences are linked with differences in microbes) 4) Selection via diet, disease, environmental exposure, lifestyle, etc. determines which microbes persist

Question 114

What are important milestones for the development of the human gut microbiome?

Answer 114

In utero effects - it is a sterile environment but metabolites and immune factors can be present due to the mother's microbiome and influence subsequent colonization Birth - gestational age and birth mode can influence microbe colonization Milk consumption - breastmilk vs formula vs both Weaning age - consumption of solid food then allows microbes to massively grow. Fiber content is an extremely important factor

Question 115

What are four phenotypes that point to ways in which we have evolved with our microbes?

Answer 115

Lactase non-persistent genotype has a stronger selection for lactose digesting microbiome if lactose is present. Low LCT expression Lactase persistent genotype has a weaker selection for lactose digesting microbiome and high LCT expression Low salivary amylase genotype has a weaker selection for resistant starch-digesting microbiome. Low AMY1 gene copy numbers High salivary amylase genotype has stronger selection for resistant starch-digesting microbiome. High AMY1 gene copy numbers.

Question 116

What are taxonomic diversity and functional redundancy in the context of the microbiome?

Answer 116

Sometimes even if taxa in a microbiome vary in abundance between individuals and there is a wide variety (taxonomic diversity), the actual overall function of the microbiome may not (functional redundancy)

Question 117

Dysbiosis

Answer 117

An imbalance in bacterial composition, change in bacterial metabolic activities, or change in bacterial distribution in a microbiome community that is linked with a "diseased" state

Question 118

What is the relationship between the gut microbiome and hypertension (give detail)? What is the role of inflammation and the nervous system and how is this hypothesized to begin?

Answer 118

Bacteria produce short-chain fatty acids such as acetate, propionate, and butyrate which feeds colonocytes in the liver. As they get distributed to the body, blood pressure decreases since they can decrease inflammation. There are two links between gut microbiome and blood pressure 1) G-protein interactions in the intestines can modulate neural pathways that decrease inflammation and blood pressure. Gut-cardiorenal axis can modulate blood pressure when active by increasing GPCR ligand binding, circadian rhythm, and the intestinal immune network for IgA production. It can also decrease pro-inflammatory cytokine IL-1 signaling along with cardiovascular disease Gut-autonomic nervous system-cardiorenal axis can increase sympathetic neuronal communication between the gut and hypothalamus and this can increase permeability decreasing blood pressure 2) Depending on if you have a 'leaky gut' --- based on the microbes obtained a birth Leakier the gut, the more likely pathogens can leave the intestine --> triggering an immune response --> triggering an increase in blood pressure Early limited breastfeeding (compared to never being breastfed), has decreased blood pressure  Summary: the microbes obtained early in life have an impact on later phenotypes

Question 119

What is the relationship between the gut microbiome and asthma (give detail)?

Answer 119

When comparing people who do not have asthma and severe asthma, they found a different gut microbiome. People who have severe asthma have a loss in some microbe (clostridiaceae).

Question 120

How can fecal transplants be used in a clinical setting? What infection is it most commonly associated with?

Answer 120

Involves the transfer of microbiota from healthy individual to diseased individual by transferring fecal matter. Highly effective for C. diff but not reliable for other diseases.

Question 121

What are two examples of animals that rely on their microbiomes to digest toxic substances?

Answer 121

Giant pandas have gut microbiomes that degrade cyanide, a toxin found in bamboo Pack rats have gut microbes that degrade creosote toxins. Both Neotoma bryanti and N. lepida were shown to have more different types of microbes when consuming creosote

Question 122

How do soil microbes help plants grow? What does this suggest about why crops are changed at different times?

Answer 122

A soil microbiome provides plants with nutrients and helps exclude pathogens, Rhizobium was just one.

Question 123

How has human activity in changed how we interact with our microbiomes?

Answer 123

Agricultural management practices, plant domestication, environmental factors, and the application of microbes are all human factors that change the soil characteristics and compounds expelled by roots along with the abundance, diversity, functionality, and colonization of microorganisms in the soil. This goes on to affect plant genotype Farming practices such as changing the timing of cover crop planting changes the soil microbiome, and cultivating the soil microbiome has increasingly become key in sustainable farming

Question 124

What is the “microbiome of the built environment”? Give several examples. What are two ways this can impact antibiotic resistance?

Answer 124

Chemicals we use in building materials and day to day life affect the microorganisms around us, and microbes can even use chemical products to produce microbial volatile organic compounds Stagnant water in plumbing systems can be conducive to microorganism biofilm formation and change microbial communities. Biofilms allow for the transfer of more genes which can contribute to antibiotic resistance Microbes in build environments experience more stress due to UV light exposure, low humidity, temperature variation, chemical, and antibiotic exposure. This stress may induce transfer of mobile genetic elements (key factor in antibiotic resistance) and it can select for bacteria with a resistance to stress