Lecture 4

Question 1

Some infection basics…

Answer 1

Transmission is important for the viral genome to establish itself in a host population and endure
- We live and prosper in a cloud of viruses: most infections have no consequence and many of them are inapparent

Question 2

What is the West Nile virus?

Answer 2

West Nile virus in 1999, had spread throughout the USA in less than 4 years, around 1 million infections by October of 2004, it’s a febrile illness but only about 20% develop symptoms, hard to stop the epidemic because most people were asymptomatic, about 1% had neuroinvasive symptoms

Question 3

What is pathogenesis?

Answer 3

• The process of producing a disease: how does the virus enter? what is the host response? where does replication occur? how does the infection spread? what tissues are infected? Acute or chronic infection? transmission? etc.
• The two main components are the effects of the viral replication in the host and the effects of the host response on the virus

Question 4

What are the requirements of a successful infection?

Answer 4

• Sufficient quantity of virus (each virus has a timer at which it is efficient in infecting or not
• Accessible, susceptible and permissive cells
• Local antiviral response must be absent or overcome
• our tears, saliva, motion, mucus and dead skin cells all are defences agains virus or pathogen entry

Question 5

What is a susceptible cell?

Answer 5

• has a functional receptor for the given virus (virus may or may not be able to replicate in cell)

Question 6

What is a resistant cell?

Answer 6

No receptor (may or may not be able to support viral replication if virus finds another way to penetrate)

Question 7

What is a permissive cell?

Answer 7

Cell has the capacity to replicate virus (may or may not be susceptible (so no receptor but the virus enters another way and replicates anyway)

Question 8

What is a susceptible and permissive cell?

Answer 8

This is the only cell type that can take up the virus and replicate it.

Question 9

How does the virus gain access for transmission?

Answer 9

• Penetrate the skin barrier, the mucosal surfaces like the respiratory tract (ciliated cells), alimentary tract, urogenital tract, the eyes, the fetus can get sick transplacentally (mom to fetus in utero) or perinatally (during birth like HIV)

Question 10

What is transmission?

Answer 10

• Spread from one susceptible host to another (required to maintain the chain of infection)
• Two general patterns: direct transmission (i.e. human to human) and vector-borne (i.e. mosquito to human)

Question 11

Some transmission terminology: what is isolation vs quarantine?

Answer 11

• Isolation: separates sick people with a contagious disease from people who are not sick
• Quarantine: separates and restricts the movement of people who were exposed or potentially exposed to a contagious disease in case they become sick (these people may have been exposed to the disease or may have the disease but do not have symptoms)

Question 12

What is a nosocomial vs iatrogenic infection?

Answer 12

• When an individual is infected in a hospital or healthcare facility (common due to concentration of people in hospital)
• Iatrogenic: when the activity of a healthcare worker leads to infection (ex: reusing needles, not washing hands, BAD practices, etc.)

Question 13

What is horizontal vs vertical vs germ line transmission?

Answer 13

• Horizontal: between members of the same species (zoonotic if different species)
• Vertical: transfer of an infection from parent to offspring (perinatal or transplacental)
• Germ line: transmitted as part of the genome (ex: proviral DNA)

Question 14

Viral spread within the organism

Answer 14

• After replication at the site of entry, viruses may remain localized, others may spread beyond the primary site (disseminated).
• If many organs are infected => systemic infection
• Physical and immune barriers must be breached

Question 15

What is viremia?

Answer 15

The presence of virus particles in the blood, kind of like the viral titer. (example: Hep C transmission through blood supply in early 80s before we screened for viremia because we had not yet discovered Hep C)

Question 16

Explain the course of infection in an animal, refer to slide 13 of lecture 4.

Answer 16

• Lots of virus enter (up), not all replicate (down), those who replicate at primary site of infection (up), leave and go in bloodstream (down), replication at distal/secondary sites (up) so on and so forth (down again when leave through bloodstream or I guess healing whichever comes first)

Question 17

Viral spread of mouse pox.

Answer 17

Gets in through injuries in skin of mouse foot, direct access to lymph node because it replicates there, then it pours into the bloodstream to reach other organs. In the case of mouse pox, the virus spreads to the spleen and liver and then goes in blood again and can disseminate to other organs or back to skin, makes a rash and virus is shed through skin for transmission.

Question 18

What is tropism and what determines it?

Answer 18

• The spectrum of tissues infected by a virus (enterotropic, neurotropic, hepatotropic, etc.) ranches from limited like hep C almost exclusively hepatocytes to pantropic like polio which infects pretty much all cell types
• Determinants: susceptibility, permissivity, accessibility, host defenses, etc.

Question 19

How does the virus shed?

Answer 19

Respiratory secretions like aerosols, nasal secretions, mucosal shedding, skin lesions, blood/blood supply, urine, semen, feces, insect vectors, germiline/vertical. Oftentimes, the number and viral load of aerosols produced through speaking and other expiratory activities can be higher than those of droplets.

Question 20

Aerosols vs droplets?

Answer 20

• Aerosols: within and beyond 1 meter, can float in air for hours, can be inhaled <5 microns 5-100 microns. If they can be inhaled they are aerosols (1 min speaking is about 1000 aerosols)
• Droplets: can travel less than 1 meter, fall to the ground in under 5 seconds, cannot be inhaled >100 microns

Question 21

How can we reduce airborne transmission?

Answer 21

• Wearing masks, keeping distance, reducing indoor occupancy
• Good ventilation especially in crowded areas like schools since the tiniest suspended particles can remain airborne for hours

Question 22

True or false, aerosol transmission can also occur at short distances.

Answer 22

True, but people have for some reason equated short range to droplet transmission. People worry more about controlling droplet/fomite transmission. Some strategies work also for aerosol Pedro like we need to focus more on that. If Covid was just droplet, distancing alone would’ve ended the pandemic but here we are.

Question 23

How long can Covid stay suspended as aerosols?

Answer 23

T1/2 = 1-3h

Question 24

Measles viral transmission

Answer 24

• Most transmissible virus
• 1 infected person will infect on average 9 other people
• creates infectious centres on human airway epithelial cells (mucosal surfaces) that peel off go through mucus are expelled through like sneezes so not only visions involved in transmission but whole ass chunks of cells (virus factories) getting out there

Question 25

How does geography affect viral transmission? Example?

Answer 25

- Geography can restrict presence of virus for example through specific vectors or animal reservoirs only present in certain parts of the world/different temperature - Chikungunya virus only present in Europe since carried by aedes aegypti but 1 aa acid change in envelope so now aedes albopictus can also carry this virus and these are found in North America

Question 26

How does seasonality affect viral transmission?

Answer 26

- Difference in temperature, humidity and our behaviour changes with the changing seasons (ex: concentrating indoors in winter)

Question 27

How do viruses find the right cell to infect?

Answer 27

- too big to diffuse so they need specific pathways - 1) adhere to a cell surface (electrostatics) so no specificity just for proximity 2) attach to specific receptors on cell surface 3) penetration 4) transport and uncoating (transfer genome inside the cell)

Question 28

The importance of cellular receptors...

Answer 28

- Essential for all viruses except those of fungi, because there is no extracellular phase and plants because entry is through mechanical damage - 1985: silica acid was the first viral receptor discovered (for influenza virus) - It was the development of technologies that facilitated the ID of viral receptors (i.e. monoclonal antibodies, recombinant DNA technology (engineering new receptors that allow virus entry), etc.

Question 29

True or false, viral receptors are made specifically for viral entry.

Answer 29

False, all receptors have their own cellular functions, viruses just take advantage of them.

Question 30

Spit the facts about viral receptors... (Hint: 4 of them)

Answer 30

- Different viruses can have the same receptor (ex: adenovirus DNA and coxsackie B3 RNA both bind CAR) - Viruses in the same family may bind different receptors (ex: Rhinoviruses at least 3 Dif, retroviruses at least 16 Dif) - One virus may bind multiple receptors (example: HSV has 3 co receptors HVEM, nectin-1 and nectin-2) - A virus may use different receptors on different cell types (ex: zika virus uses Gas6/AXL to enter astrocytes and glial cells but uses a different entry mechanism in neurons) so it binds Gas6 a soluble protein and piggy backs on it to bind to natural receptor AXL

Question 31

How do non-enveloped viruses bind to the cell membrane?

Answer 31

- Bind via capsid surface (ex: poliovirus binds through concave part to receptor CD155) or protrusions like fibres at vertices of adenovirus that bind to CAR receptor - Then either injection of genome or endocytosis and uncoating happens after escape from endosome

Question 32

What is the role of co-receptors in viral infection?

Answer 32

- Sometimes viral receptors are not accessible at the surface of the cell - Example: coxsackie group B viruses require two receptors: DAF (decay-accelerating factor) and CAR (coxsackie-adenovirus receptor) DAF drags to CAR which is in a tight junction for entry of virus

Question 33

Give two examples of enveloped viruses that fuses with the plasma membrane.

Answer 33

- Paramyxoviruses (HN protein binds to receptor which triggers a conformational change in the F or fusion protein so it sticks into the host cell surface and hairpin motion brings membranes close until they can fuse) - HIV (one viral protein interacts with CD4 so conformational change to interact with CCR so other conformational change so fusion peptide (GP41) can touch surface and fusion of membranes can happen when exclusion of water)

Question 34

Fusion with plasma membrane must happen at ... pH.

Answer 34

Neutral

Question 35

Two types of endocytosis are...

Answer 35

- Macropinocytosis: ruffling of membrane and uptake of a bunch of extracellular stuff in large vacuoles - Receptor-mediated endocytosis: clathrin mediated or other independent pathways

Question 36

Fusion in the endoscope can be triggered at ... pH.

Answer 36

Low, between 6.5 and 5.3, conformational changes are triggered for fusion in the endosome.

Question 37

Example of virus that fuses with the endosome membrane...

Answer 37

Influenza virus has HA molecule which is a trimer and low pH triggers conformational change so insertion of fusion membrane into endosome membrane and then hairpin motion until membranes are brought together and fusion happens and 8 genome segments are released into the host cell At cell membrane HA binds to silica acid and host cell proteases activate the receptor => receptor mediated endocytosis

Question 38

What are class I fusion proteins?

Answer 38

- Perpendicular to membrane (spikes) - Mostly alpha-helical - Form trimers

Question 39

What are class II fusion proteins?

Answer 39

- Parallel to the membrane (lie flat on the surface and THEN extend conformational change) - Mostly Beta-sheets - Form dimers (usually trimers of dimers) - Example: Dengue virus in endosome pH drops and receptors go from parallel to stick up and into membrane, hairpin and fusion of membranes, release of genome into cell

Question 40

What is an unusual entry mechanism? Give an example.

Answer 40

- The initial entry receptor for cell is unknown, might be macropinocytosis. Once in the endosome, an endosomal protein NPC1 (Newman Pick C1) is the one that interacts with the virus and leads to membrane fusion vs usually being mediated by a virus fusion protein. - If people have Newman pick disease, lack of receptor and no longer susceptible to ebola

Question 41

What can be said about the regulation of fusion?

Answer 41

- Must not occur in the wrong location - Plasma membrane: neutral pH, usually second protein receptor interaction - Endosome: low pH, proteolytic cleavage activates the fusion receptor for fusion (Class I), cleavage of a second protein (class II) activates the fusion protein, endosome fusion receptor like ebola

Question 42

How do non-enveloped viruses enter the cell?

Answer 42

- Typically rely on rupture of cell membranes. - Adenovirus (ruptures endosome, pokes holes until endosome bursts and then BAM in cell) - Poliovirus creates a membrane channel where it can inject its genome - Retroviruses, HSV: direct cell-to-cell transmission, virological synapse, cellular conduits and extracellular viral assemblies

Question 43

The cytoplasm is quite active, how do viral particles move around in it?

Answer 43

- Movement of particles above 500 kDa does not occur through passive diffusion => viral particles are transported actively within endosomes, lysosomes or simply the free virion via the cytoskeleton (actin, tubulin)

Question 44

How does the virus access the nucleus?

Answer 44

During cell division since there is the breakdown of the nuclear membrane or zip through the nuclear pore complex thanks to NLS (influenza) OR docking at the nuclear pore and injection of genome (HSV-1/adenovirus (second one gets transported to the nuclear pore with dynein along MT and then really tight binding to nuclear pore so when kinesis walks in the opposite direction the capsid gets ripped apart and the viral genome can be released into nucleus across pore) OR nuclear membrane disruption (Parvovirus pokes holes).