Topic 5 - Magor (Polio)

****How big is the Polio virus?

One of the smallest particle +strand RNA viruses that infects humans

***True or False: Polio = a Human only pathogen

TRUE

*****What type of Genetic material do Polio viruses have?

RNA

= RNA virus

• Picornavirus (small RNA virus)

****How infectious + Lethal is the polio virus?

Quite infectious = 3.5 (R0)

Quite lethal = 23%

******What type of RNA does Polio have? + or -?

(+)RNA

*******What is (+) RNA?

RNA that is already ready to be translated into protein

*******True or False: + strand RNA viruses do NOT need to package POLYMERASES + ENZYMES in the capsid?

• Why or why not?

TRUE

• Because they are + strand the polymerases + Enzymes can be directly translated from the (+) RNA = can synthesize their own

******Why Does an RNA+ Virus need to make a( -)RNA strand if protein can already be directly translated from the + strand?

For REPLICATION

• Need (-) to be a Template to synthesize more (+) strands

*******What are the things packages inside the capsid of the Poliovirus?

ONLY ONE RNA (+) strand

*******What is the Capsid of Polio virus made up of?

60 copies of 4 proteins (VP 1,2,3+4)

• 3 on outside

• 1 on inside

**************Give the steps of Polio Reproductive lifestyle

• from Binding to Exit

• How are things produced + replicated

Note: POLYPROTEIN synthesis

Note: RNA replication happens more on the surface/membrane of vesicle than within

*To what extent is the proliferation/replication of 1 RNA of Polio?

• # of copies in what amount of time

1 RNA = 50 000 copies in 24 hrs

• Very large amplification

******Difference between uninfected cells + cells infected with Polio

Vesicles (due to replication of RNA in the vesicles/vacuoles)

****What does the Polioviral +RNA strand encode for?

A single Polyprotein Precursor

********What are the 2 ENZYMES that cleave the polypeptide?

1. 2Apro

2. 3Cpro

2 virally encoded proteases

*****Which of the 2 ENZYMES cleaves the polypeptide first into Capsid vs. Proteases + RNA synthesis proteins

2Apro

*******How is the polyprotein initially cleaved if the protease enzymes are not yet translated + synthesized?

The pre-encoded proteins are able to cleave while still inside the polyprotein = AUTOCLEAVAGE/Self- cleavage

• Thus the initial first cleavage = slow

********What part of the polyprotein is cleaved into CAPSID PROTEINS (aka. what part of the protein = the capsid proteins)?

P1 = VP0 , 3 + 1

• VP0 = further cleaved into VP2 + 4

*****What part of the polyprotein Makes up the Polymerase

• AKA what is the Polio Polymerase called?

3Dpol

*******What type of POLYMERASE is the polio 3Dpol?

RNA dependent RNA-polymerase

• an enzyme that catalyzes the synthesis of RNA from an RNA template.

******What are the Characteristics of the polio 3Dpol polymerase?

• Does it exist in other organisms

• What is the Template

• proof-reading?

UNIQUE to VIRUS

Specific for RNA template (can differentiate between host + viral RNA + Know which to replicate)

• Copies Viral RNA + not host RNA

NO proofreading

********True or False: LIKE HIV polio = also a Quasispecies?

• WHY or WHY not? aka. what makes HIV a quasispecies?

TRUE: also = a quasispecies (NO WT)

• Because of Error prone characteristic = too many versions + No set most common

**********What is the Error rate of 3Dpol? (X/bp)

1/1000 Bp

******What is the Size of the polio RNA (in Bp)

• Why is it this size?

Small ~ 5kb

• WHY?: Error rate limits the size of the virus

  • Increase size = increase mistakes → high mistake = not viable

*******How is the 3Dpol Specific to viral RNA + does not copy host RNA? (how does it tell which is viral + host RNA)

• Red question in lecture

Viral RNA secondary shape = Unique to virus

*****What is the RNA secondary shape determined by?

The sequence of the RNA

• Base pairs with itself

********How does the polio virus address the End replication problem? (why we have telomerases)

• What strategy/mechanism does it use

Uses a PROTEIN as a Primer

*******What is the Protein Polio uses to address the end replication problem called?

3AB:Vpg

**********What is the Mechanism of using 3AB:Vpg as a Protein primer for the end replication problem

1. 3AB take RNA to Vesicle membrane + using a hydrophobic loop inserts into the membrane

2. RNA strand links to 3AB via a TYROSINE on the VPg portion of the protein (neutrophilic attack = covalent attachment of RNA to 3AB

3. Primer starts synthesis on the Tyrosine of 3AB

4. Elongation

5. Cleavage of VPg OFF 3AB

*******True or False: GENOMIC RNAs can serve as mRNAs for the synthesis of new Polio proteins?

TRUE

*******What is the First step of Translation of polio proteins?

REMOVAL of VPg protein

********How are Polio Virus Capsid assembled + packaged with RNA?

auto assembly

• When capsid proteins are in sufficient enough concentration (abundant) they automatically aggregate + package RNA genomes inside

******Can Polio virus perform Recombination?

YES

********What is RNA recombination? in Polio virus?

Involves RNA polymerase TEMPLATE EXCHANGE

• Poly can switch template used as donor if current donor = not good

******It is estimated that ___-___% of polio viral templates recombine

10-20%

***In Polio Virus Does Viral template recombination happen more often when replicating the (-) or (+) strand?

When replicating the - strand

• IDK why

• I also don’t know if i’m interpreting/understanding this sentence correctly

**********What are the 2 types of Vaccines used in the Successful Vaccination campaign against polio?

1. Inactivated Vaccine

2. Oral Vaccine

************Compare + Contrast between the Inactivated vs. Oral Polio vaccine

• Method of delivery

• What type of Response does it induce

• What type of Vaccine is it

• Which works better

• IN WHAT SCENARIOS IS EACH ADMINISTERED

• Pros + Cons etc.

Inactivated: Injected (con) = administered in areas where there is no active polio

• CON = Doesn’t induce cellular immune response (T cells) + Often need booster to induce memory response + difficult to store (need to keep cold)

• Induces an ANTIBODY RESPONSE instead (B cell)

Oral: LIVE ATTENUATED VIRUS = Administer in areas where there is active Polio

• PRO: live virus = synthesizing new virions + viral proteins that target both humoral AND CELLULAR immune response(much better immunity) + easy to administer + store

• Used to eradicate polio in all countries (Except for 2)

****Which of the 2 types of polio vaccines is currently used in Canada?

Inactivated Polio Vaccine (IPV)

**********What is an attenuated vaccine?

living but weakened version of the virus or bacterium it protects against.

• Because the pathogen is alive, it can still replicate inside your body and trigger a strong immune response, but it has been modified so it can no longer cause serious disease.

• This approach produces some of the most powerful and longest-lasting immunity of any vaccine type

True or False: Vaccination eliminated poliovirus in Canada

TRUE

***When was the Last major Polio endemic in Canada?

• How many total cases

1959

• 190 000 polio infections

*****What can Polio cause?

Muscle weakness + Nerve inactivation = Paralysis

• if in the thoracic region = cannot breathe

eg. Paul Alexander age 72 Texas

Of the last major epidemic in 1959, How many of the 190 000 polio infections resulted in paralysis?

1 887

***When was the last active case of polio in Canada?

1996

• brought by visitor to Canada

********How was the Attenuated Vaccine made?

• In which organism was it proliferated/grown to make the vaccine?

• How does one ATTENUATE A VIRUS

Human viruses may be ATTENUATED by passage in NON-HUMAN cell lines

1. Pathogenic Virus = isolated from a patient + grown in human cultured cells

2. Cultured virus then used to infect a MONKEY

   1. 21 passage intracerebrally in monkey

   2. 8 passes in testicle

   3. 39 in kidney

   4. Plaque purification from monkey kidney cultures

3. Virus accumulated many mutations that allowed it to grow well in monkey cell

4. Virus no longer grows well in human cells = candidate for a vaccine

NOTE: High number of mutations = attenuates virus

• grown in monkey cells = adapted to monkeys = decrease effectiveness of infections in humans

• eg. can no longer assemble in humans

********WHAT is an ISSUE with ATTENUATED Vaccines?

REVERSION mutant of vaccine strain made an infectious virus that caused polio from an attenuated strain

Which VACCINE? (name) was the Reversion mutant strain derived from?

• What type of reversion mutation did it aquire?

P3 Sabin Vaccine

• Amino acid changes

• Although AA changes did not remake the original virus- the new proteins could assemble a functional capsid + be infectious

What does this map show?

AREAS of Polio activity

• 2 yellow countries = the 2 counties with active polio (Pakistan + Afghanistan)

• Dots = areas with Vaccine derived polio

********What are Vaccine-derived polio?

• How does it occur?

Reversion of Virulence = occurs due to People not Being vaccinated then getting infected by the vaccine version of the virus because they are not immune

Why was there a peak in vaccine derived polio in 2020?

Covid stopped vaccination regimes in some countries = less people vaccinated → more susceptible to vaccine strain

******************Does Vaccine derived polio affect the vaccinated? (in red in slides)

NO

*****What is the new Polio vaccine called

• What was the goal of developing this New vaccine?

OPV2

• Goal = Get rid of Vaccine-derived polio

****True or false: Vaccine-derived polio does exist in Canada

TRUE

• Detected in 2 Canadian waste water samples

**What are some Reasons not to vaccinate with the Inactivated vaccine? (?as an attempt to reduce vaccine-derived polio?)

MORE DIFFICULT

• 2 doses for memory

• Injection

• Need to be stored cold

Prob yes

Which 2 countries did Canada share important travel advisory updates + Why?

UK + Germany

• Live polio virus