RNA Viruses

Question 1

General Concepts in Medical Virology

Answer 1

• Viruses “target” specific cell types and organs
• We think (and learn) about them in a syndrome-specific manner
• Most are routinely diagnosed using molecular (PCR) or serology tests (rarely culture)
• High genomic diversity is a challenge for treatment and vaccines
• Treatments are often virus-specific
  
  • Not nearly as “broad-spectrum” as treatments for bacteria

All viruses have genetic information (a genome) and a capsid.
The others are variable (i.e., polymerase, envelope, and other proteins)

Question 2

Positive Sense vs. Negative Sense

Answer 2

Positive sense means the RNA genome can be read in a 5’ to 3’ direction by host translation machinery, i.e., is like messenger RNA. So it can be directly translated into protein. Whereas negative sense RNA you would need a complementary strand first.

Question 3

Some Medically Important RNA Viruses

Answer 3

• CNS: enteroviruses (picorna), West Nile (flavivirus),
  rabies virus (lyssavirus)
• Respiratory system: influenza, parainfluenza (orthomyxo/ paramyxovirus), hantavirus (bunya), coronaviruses (seasonal CoVs, SARS-CoV, SARS-CoV2, MERS-CoV), rhinoviruses (picorna), RSV
• Heart: coxsackievirus (picornavirus), others
• Gut: norovirus (calici), rotavirus (reo), others
• Liver: HAV (picornavirus), HCV (flavivirus), HDV, HEV.
  
  • HBV (actually a DNA virus), HCV, and HDV are transmitted sexually or via blood contact (parenteral transmission) HAV and HEV is transmitted via through stool (fecal oral transmission).
• Hemorrhagic fever: Ebola and Marburg viruses (filovirus), Lassa (arenavirus), YFV (flavivirus)

Very little correlation between virus family and what organ they affect/what symptoms they casue

Question 4

Rhinoviruses

Answer 4

• Genome type (e.g., DNA, RNA) and general structure (enveloped or non-enveloped)
  
  • Picornavirus (like enteroviruses and Hepatitis A)
  • Positive sense, single stranded RNA (looks like a human mRNA): + ssRNA
  • 7,200 bp
  • Not enveloped - Are MORE stable than enveloped viruses b/c the envelope is pretty susceptible to drying out and being destroyed by detergents.
  • HIGH genomic diversity
    
    • .Three groups (A,B,C). Many strains within a group
    • Makes it very difficult to develop long-term immunity as there will be another strain that is different enough to avoid the memory immune response.
  • IRES = Internal ribosome entry site: leads to translation of polyprotein
  • Four capsid proteins
  • Non-structural (including enzymes) proteins
    
    • Protease - cleaves the viral polyprotein
    • RNA-dependent RNA polymerase - copies the genome
• Important features of its life cycle and reproduction
  
  • 1) Binds to host cell and undergoes endocytosis and its uncoated so the genetic material is set free
  • 2) Directly translated by host ribosomes to make viral proteins, including
    
    • Capsid
    • RNA-dependent RNA polymerase (RdRP) - meaning a polymerase that makes RNA from RNA
  • 3) RNA is transcribed by RdRP to make more genome copies
  • 4) Have all the building blocks together and they assemble to make new viruses that then leave the cell.
• Target cells and organs
  
  • Infects epithelial cells
  • Upper respiratory tract including tonsils, some lower respiratory tract.
• Symptoms and clinical features
  
  • Causes the common cold (50%), pneumonia, exacerbations of chronic pulmonary disease, can lead to secondary bacterial infection•
• Transmission route
  
  • Transmitted by aerosol droplets, contact (main mode of transmission), and fomites
  • Highly transmissible; lasts on surfaces for days. This is partly b/c it doesn’t have an envelope, which makes it a lot more stable.
• Diagnosis
  
  • Diagnosed clinically; PCR can be used in research/surveillance
• Treatment (fecal-oral/fomites; parenteral)
  
  • No specific treatment (very difficult with the enormous amount of strains); maybe zinc or vitamin C. Jury still out.
• Prevention
  
  • No vaccine; prevent by hand-washing!

Question 5

Rhinoviruses - Genome Type and General Structure

Answer 5

• Picornavirus (like enteroviruses and Hepatitis A)
• Positive sense, single stranded RNA (looks like a human mRNA): + ssRNA
• 7,200 bp
• Not enveloped - Are MORE stable than enveloped viruses b/c the envelope is pretty susceptible to drying out and being destroyed by detergents.
• IRES = Internal ribosome entry site: leads to translation of polyprotein
• Four capsid proteins
• Non-structural (including enzymes) proteins
  
  • Protease - cleaves the viral polyprotein
  • RNA-dependent RNA polymerase - copies the genome

Question 6

VIral Hepatitis

• HAV
• HBV
• HCV

Answer 6

• Hepatitis A
  
  • Picornavirus
  • Transmission is via fecal-oral / food
  • Very good vaccine available
• Hepatitis B
  
  • Hepadnavirus
  • Transmission is via blood / bodily fluids
  • Very good vaccine available
• Hepatitis C
  
  • Flavivirus
  • Transmission is via blood / bodily fluids
  • NO vaccine available

All cause hepatitis (liver inflammation)

These are the most common hepatitis viruses. Cannot have D without B and it’s HDV is not very common. HEV is extremely rare in the US, but could develop it after traveling.

Question 7

Hepatitis C

Answer 7

• Genome type (e.g., DNA, RNA) and general structure (enveloped or non-enveloped)
  
  • Flavivirus
  • +ssRNA genome
  • 9,500 bp
  • Enveloped
  • Many strains
• Important features of its life cycle and reproduction
  
  1. Virus envelope is used to bind to and fuse with the host cell in viral entry
  2. Uncoated
  3. Cell makes the viral proteins and replicate the RNA
  4. Virus envelope and budding utilize the cell’s lipoprotein synthesis pathways
• Target cells and organs
  
  • Infects liver cells (provide lots of nutrients)
• Symptoms and clinical features
  
  • Acute infection: often asymptomatic; can have fever, nausea, abdominal pain, jaundice
  • Chronic infection (85%): cirrhosis, liver cancer; leading reason for liver transplant in US. So 15% spontaneously clear the virus.
• Transmission route
  
  • Bloodborne (transfusions, IV drug use, tattoos, piercing, needlestick), sexual (rare), mother-to-child
    
    • 90% of people who use IV drugs have been exposed to hepatitis C
    • Change in 2020: screening now recommended for every adult (once in a lifetime, like HIV), (previously risk group based - baby boomers etc. - turns out need to screen more broadly)
• Diagnosis
  
  • Serology, followed by PCR if +
  • Q: Why would we use serology and then PCR?
    
    • A: Serology tells us if someone has ever been infected with hepatitis C; PCR tells us if they are still infected (85% of infected people), or whether they have cleared it (15%)
• Treatment (fecal-oral/fomites; parenteral)
  
  • Combination therapy: required due to high viral mutation rate
  • NS5B inhibitor
    
    • Nucleoside analogue: capitalizes on the error-prone RdRP
  • NS5A inhibitor
  • Virus-specific treatments developed ~2011. They are called direct acting antivirals.
  • Patients can now be cured in as little as 8 weeks and in >90%!!
• Prevention
  
  • No Vaccine
• History
  
  • Identified in 1989
  • Initially treated with interferon and ribavirin
    
    • Required long treatment course 6-12m
    • Significant side effects (“flu-like symptoms”), bone marrow suppression

Question 8

HCV - Genome type and general structure

Answer 8

• Flavivirus
• +ssRNA genome
• 9,500 bp
• Enveloped

Question 9

Recap of Rhinovirus and HCV

Answer 9

Question 10

HCV - Treatment

Answer 10

• Combination therapy: required due to high viral mutation rate
  
  • NS5B inhibitor
    
    • Nucleoside analogue: capitalizes on the error-prone RdRP
  • NS5A inhibitor

Question 11

Influenza

Answer 11

• Genome type (e.g., DNA, RNA) and general structure (enveloped or non-enveloped)
  
  • Orthomyxovirus
  • Negative -ssRNA, segmented genome
  • Has nucleoprotein “capsid” - the capsid does not form a shell like with the other viruses we discussed, it just binds along the length of the RNA.
  • Enveloped - so not very stable outside the body
  • Carries its RdRP in the virus particle, which Rhinovirus and HCV did not.
  • Eight segments (strands) of RNA, each of which encodes a different protein.
    
    • Each coated with nucleoprotein
    • Packaged with RdRP
    • No protease, since no polyprotein (though protease activity is required from host enzymes at some points in the virus life cycle, beyond the scope of this course)
• Important features of its life cycle and reproduction
  
  1. Binding to receptor and cell entry
  2. Transcription of -RNA genome to +RNA using the RdRP it carries. The positive sense RNA can be copied back into negative sense RNA by RdRP to make more genome copies and also serves as messenger RNA to make proteins.
  3. After translation of the viral proteins, they can be assembled in the cell.
  4. It buds from the cell, takes a part of the cell membrane with it to form the envelope.
• Target cells and organs
  
  • Infects epithelial cells in the upper and lower respiratory tract
• Symptoms and clinical features
  
  • Symptoms: fever, chills, body aches, difficulty breathing, and cough (In contrast, with Rhinovirus often times you won’t even develop a cough because the virus may just be limited to the nasal mucous membranes) but it’s very typical with the flu.
  • Most of the symptoms are due to the immune response, specifically interferon and cytokines
  • Duration: <1 week, but full recovery can take ~1 month
  • Complications: bacterial superinfection. Can kill a lot of people.
• Transmission route
  
  • Respiratory droplets; less by fomites (touch)
• Diagnosis
  
  • Performed on nasopharyngeal swab
  • Rapid test – result in minutes
  • PCR – result in hours
• Treatment (fecal-oral/fomites; parenteral)
  
  • Oseltamivir, zanamivir - just need to remember these neuraminidase inhibitors
    
    • Block neuraminidase (one of the envelope proteins of the flu virus)
    • Prevent the virus from budding off the host cell. Cannot leave the cell.
    • Need to be tacken very early in infection.
  • Amantadine, rimantadine
    
    • Target the M2 protein on the viral envelope
    • Prevent entry of the virus into the cell
    • Current influenza strains are resistant
• Prevention
  
  • Injected (protein) or intranasal (LAIV – live attenuated influenza virus)
    
    • Spray is generally less effective in adults.
    • Virus is made weaker in the spray (attenuated) by a mutation that adapts it so it doesn’t work in warmer temperatures (i.e., temperature of human body).
    • Injection is better for those that are immunocompromised b/c it’s not live attenuated.
  • Mixture of 4 strains (quadrivalent) concocted every year based on surveillance data; LAIV or injected - inactivated vaccine (egg- or cell culture based) or recombinant; in recent years:
    
    • 1 influenza A (H1N1) virus (variant of 2009 strain)
    • 1 influenza A (H3N2) virus (adapting more often)
    • 2 influenza B viruses
  • Recommended YEARLY for everyone >6 months old

Question 12

Influenza - Genome and Structure

Answer 12

• Orthomyxovirus
• ssRNA, segmented genome
• Has nucleoprotein “capsid” - the capsid does not form a shell like with the other viruses we discussed, it just binds along the length of the RNA.
• Enveloped
• Carries its RdRP in the virus particle, which Rhinovirus and HCV did not.
• Eight segments (strands) of RNA, each of which encodes a different protein.
  
  • Each coated with nucleoprotein
  • Packaged with RdRP
  • No protease, since no polyprotein (though protease activity is required from host enzymes at some points in the virus life cycle, beyond the scope of this course)
• Not important to know all 8 segments, but do know Hemagglutinin and Neuraminidase

Question 13

Influenza - Genomic Diversity

• Hemagglutinin
• Neuraminidase

Answer 13

• Three types: Influenza A; Influenza B; Influenza C (rare)
• Multiple subtypes: current Influenza A subtypes are H1N1 and H3N2
  
  • Within each subtype, there are different strains. Every year, these change a little
    
    • Named for the type/place of original isolation/date of original isolation/antigens. Some examples include:
      
      • A/Michigan/…/2015/H1N1
      • A/Singapore/…/2016/H3N2
      • B/Colorado/…/2017
      • B/Phuket/…/2013
• Hemagglutinin (18 subtypes)
  
  • The viral attachment protein
  • Binds to sialic acid on host epithelial cells
  • Promotes fusion of the virus with the host cell
  • Elicits the host antibody response
• Neuraminidase (11 subtypes)
  
  • Cleaves sialic acid
  • Allows the virus to bud off the cell
  • Also elicits antibody response
• All HA and NA subtypes infect wild birds, only some infect pigs and/or humans

Question 14

Influenza rapid test (also called lateral flow assay)

Answer 14

1. Add sample and have virus-specific antibodies, which bind to virus antigen if positive, forming antigen-antibody complexes.
2. Diffuse through the paper through the paper by capillary action to the “test area” which contains immobilized antibodies which catch the antigen-antibody complexes. A positive result is indicated by a color change from the binding of antigen-antibody complex to the result line.
3. Test validity ensured by color change from the binding of antibody to the control line.

Question 15

Antigenic Drift vs. Antigenic Shift

Answer 15

Antigenic Drift:

• Changes in hemagglutinin and neuraminidase proteins due to mutations in the viral genome
• Not recognized as well by existing antibodies
• Happens continuously during replication.

Antigenic Shift:

• When two strains of influenza infect the same cell, the RNA segments can re-assort randomly, creating new viral strains
• Not recognized at all by existing antibodies
• Can lead to a pandemic
• Pigs are an excellent “mixing vessel” for human influenza. Pigs can be infected with pig, bird, and human flu viruses. If this happen simultaneously, viruses can rearrange and swap genes.
• What characteristics of influenza make it possible for antigenic shift to occur:
  
  1. A single cell can be infected with multiple strains at once.
  2. Its genome is in multiple strands of nucleic acid (a segmented genome)

Question 16

Epidemic vs. Pandemic

Answer 16

• Epidemic: outbreak of an infectious disease that spreads quickly and affects many individuals
• Pandemic: an epidemic that spreads through diverse populations across a large geographic area
  
  • H1N1 1918
    
    • Estimated 50 million deaths, 2.5% mortality
  • H2N2 1957
    
    • Estimated 1 million deaths, 0.1% mortality
  • H3N2 1968
    
    • Estimated 2 million deaths, 0.1% mortality
  • H1N1 2009 (swine flu)
    
    • Estimated 200,000 deaths, 10% mortality
    • Comparable to seasonal influenza
• Endemic: Means disease/virus is always present in a population or region
  
  • R0, basic reproduction number, and endemic means the R0 is pretty similar to 1.
  • R0 of 1 means that one person passes the disease to one other person on average. R0 of 2 means that one person passes the disease to two people on average. R0 of .5 out of 2 people who have the disease only one person is infected.

Question 17

2009 H1N1 Pandemic “Swine Flu”

Answer 17

This type is still circulating as seasonal flu and is included in yearly vaccines

Question 18

Avian Influenza “Bird Flu”

Answer 18

• Some influenza viruses can be transmitted directly from birds to humans and this is what is known as “bird flu.”
• So far, this has just occurred via sporadic transmissions, without sustained human to human transmission. The fear is that one of these strains could adapt to humans (via antigenic drift) to become more easily spread from person to person.
• High mortality
• Little person-to-person spread

Question 19

Recap of Influenza

Answer 19

• •Enveloped -ssRNA virus with 8 genome segments
• HA and NA are the key envelope proteins that elicit protective antibodies
• Antigenic drift (minor genetic changes; seasonal variation; need for yearly flu vaccine) vs. shift (reassortment; pandemics)
• NA is the target of oseltamivir