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Flashcards in 3: Viral properties and disease Deck (30):
1

Koch’s Postulate
-fx
1-4

-prove that a virus can cause disease (e.g. norovirus):
1. The micro-organism must be found in large no.s in all disease animals, but not in healthy ones
2. The organism must be isolated from a diseased animal and grown outside the body in a pure culture
3. When the isolated micro-organism is injected onto other healthy animals, it must produce the same disease
4. The suspected microorganism must be recovered from the experimental host, isolated, compared to the first microorganism and found to be identical

2

virus:
-genome
-replication
-size

virus: an infectious obligate intracellular parasite – obligate meaning that they cannot complete its life-cycle without it being in a host
-genome DNA or RNA
-viral genome replicated and directs the synthesis of more viral components and genomes
-small -100nm

3

Virus morphology
non
2
e.g. 3

en
3
e.g.

**

Non-enveloped:
-protein capsid
-more symmetrical
-e.g. adenovirus, picornvirus and calicvirus

Enveloped:
-protein around genome
-lipid envelop around derived from host membrane
-pleomorphic – lots of diff shapes
-e.g. Ebola virus

**herpes virus is a combo capsid and envelop

4

naming

disease, person, place, part of body, spreading

5

Baltimore Classification by

types 4

**

PRINT PIC

classified by type of genome by DNA or RNA (in various forms)

ds/ssDNA
+ve RNA strand – sense strand; directly translated
-ve RNA strand – antisense strand
dsRNA

**Some viruses have DNA or RNA but follow a different cycle for making it

6

The Central Dogma
norm

+ve
-ve 3

DNA --> RNA --> protein

+ve sense RNA strand – as soon as it enters the cell, ribosomes can translate that into a protein

-ve sense strand – cannot be translated straight away
-transcribed back into a +ve sense complimentary copy
-carry proteins/enzymes/machinery that will convert the genomes back into sense RNA
-Otherwise inert

7

Consequences of the viral genome type
RNA/retro
-own
-size
-specific size
-proteins

DNA
-size
-space for
-some
-but
**

RNA Viruses and Retroviruses:
o own polymerase to replicate: error prone & lack proofreading capacity --> high mutation rate; fast evolution
o genomes limited in size due to the instability of RNA
o Largest RNA viruses are around 30kb
o complicated coding strategies to encode more proteins than expected from a small RNA genome

• DNA Virus:
o Can be big because DNA is more stable
o There is space for accessory genes – things that viruses don’t inherently need to survive but may give some adv
o Some one long strand, some several little pieces
o segments: -ve: pieces of RNA must gather when virus tries to leave cell, +ve: opportunity to pick up new genes and evolve
**attenuated vaccines: accessory genes are lost in culture because there is no immune system

8

Generic Virus Replication Cycle 1-9

1. virus outside the cell, with it being inert
2. viral attachment protein binds to receptors on host cell surface
3. --> virus dock down onto the cell surface
4. Some viruses enter the cell by fusing with the host cell membrane and injecting their DNA or RNA into the cytoplasm
5. Other viruses enter via a series of vesicles such as endosomes
6. Once the viral genome is in the cell, it must be made into mRNA if it isn’t already
7. virus uses the host ribosomes to translate own mRNA & produce proteins
8. At the same time, the virus begins to replicate its own genome either using its own polymerase or host cell polymerase
9. The copies of the genome and the viral proteins assemble to form new virus particles which then leave the cell

9

Investigating Viruses in the Laboratory
-need

-effect

-can be due to
-i.e.
-after,
-no.
-1-4

-need host cells as viruses have to be grown inside host cells

-cytopathic effect (CPE): death of the cell as a result of being infected by the virus; change in cell shape --> cell death

-CPE could be due to the virus taking over genetic machinery --> cell unable to produce proteins to survive --> destined to die via apoptosis
-i.e. have to compete with host protein synthesis in aggressive way
-after death, viruses form plaques on cell monolayers from individual viruses infecting one cell, and then infecting other cells
-no. plaques measure how many viruses in the sample
-Plaque Assay:
1. Take a sample from the patient that contains the virus
2. Make a serial 10-fold dilution
3. Take a known volume of the dilutes and put it onto the susceptible cells
4. There will be plaues on the susceptible cells showing how many viruses there are in the dilution

10

Syncytia
-some
-syncytia define
-e.g.
-in lab,

-some don’t form plaques; form syncytia
-syncytia: big bundle of cells in the middle stuck together, effectively forming one massive cell
-e.g. HIV: fuse at cell membrane. HIV infected cell express HIV glycoproteins on their surface --> cell fusing as HIV proteins and the receptor on other cell interact
-in lab, can visualize whether presence of virus: drop sample of blood on some susceptible cells. if form syncytia, +ve

11

Immunostaining the infected cells
3

virus produce diff products to cells
-If suspect RSV, induce sample onto a cell and wait for the cell to start producing a protein that is unique to the virus
-then use an antibody which can detect that protein

12

Viral Diagnosis

•Detecting viral genome: PCR
•Detecting viral antigens: Indirect Fluorescent Antibody, ELISA
•Detecting viral particles: Electron microscopy, Haemagglutination Assay
•Detecting virus cytopathic effect in cultured cells: Virus isolation
•Detecting antibodies to virus: Serology

13

Propagating Virus
-provide
-then
-->

some

-providing permissive cells
-Viruses may accumulate mutations that adopt them to the new host
--> attenuation (basis of generation of vaccines in the past)

some viruses have no permissive cell lines --> difficult to develop antiviral strategies against

14

Manipulating Viruses

Virus genome small; can be synthesized
--> reverse genetics – produce new viruses

15

tropism:
based on:
3

tropism: predilection of viruses to infect certain tissues and not others
based on:
o Susceptibility – receptor interactions
o Permissivity – ability to use the host cell to complete replication
o Accessibility – ability of the virus to reach the tissue

16

HIV

replication cycle 1-8

-receptor
-mutation
-tropism

HIV infects immune system and causes a decrease in immunity --> infection

replication cycle:
1. Virus is on the outside
2. GP120 glycoprotein on the virus capsule interacts with CD4 on the host cell membrane
3. interact with a co-receptor (CCR5 or CXCR4 depending on what cell the CD4 receptor is located) --> virus in close proximity with cell membrane
4. Virus membrane and host membrane fuse and the viral content will be inserted into the cell
5. Enzymatic copying of the viral genome by reverse transcriptase to produce viral DNA
6. The DNA moves into the nucleus and is integrated into the host genome
7. It is transcribed into mRNA and subsequently translated into a protein
8. All of these then come together at the cell membrane and produce a new virus particle which leaves the cell

o CD4 on HIV infects T cells and macrophages --> slight varying complimentary receptor
o Some have mutations in CCR5 --> resistant to HIV
o Some produce lots of chemokine - blocks the use of the co-receptors
o tropism switch during HIV replication – viruses evolve to bind to other receptor

17

Measles virus and its receptor

receptor i,ii
-
-

1-8

-H receptor (hemagglutinin) binds to:
i - SLAM (CD155)
ii - Nectin 4
-measles vaccine uses CD55
-SLAM and Nectin 4 are used at different stages

1) measles enter via the respiratory tract
2) binds to SLAM on immune cells (e.g. dendritic cells) which are surveying the area
3) hijack way to the lymph nodes
4) At the lymph nodes, virus is exposed to more cells expressing SLAM, which it then infects causing immunosuppression
5) When leaving host body, uses Nectin 4
6) carried from the lymph nodes to the lungs
7) crosses from blood to airspace by binding to Nectin 4 on bottom of airway epithelial cells
8) Viruses replicate in airway epithelial cells and burst out into the airspace to spread via the respiratory route

18

Influenza
life cycle
1.
- enveloped (**)
- theoretical
-tends to

2.
-pH
-only happens when
-only found
-viruses will

3-5

** -->
can be extended by

life cycle:
1. Spike proteins are used to attach to the surface of the cell
-Influenza is enveloped in hemagglutinin (HA) – latch onto surface of cells by binding to sialic acid (**tropism is not determined by receptor use)
-Sialic acid ubiquitous – it is everywhere; influenza can theoretically enter any cell
-tends to only affect the respiratory route, partly due to accessibility

2. virus engulfed and taken into an endosome
once bound onto sialic acid, enters the cell via endosome
-Low endosomal pH allows a massive conformational change for fuse with the endosome membrane and uncoat
-only happen if the protein of the virus has been sniped in two particular points
-only in the fluid that lines lungs that the right proteases are present
-Viruses mill around until they come into contact with the appropriate proteases

3. fuses and releases genome
4. Genome enters the nucleus where it replicates and directs synthesis of the proteins
5. form new virus particles

**--> influenza tropism determined by availability of host proteases
-tropism extended by mutation at the HA cleavage site: Some chicken viruses have mutated the region where the protease cleaves --> cleaved by different proteases found in other areas of the body; virus replication in every organ of the body

19

Transmission terminology 5

• Iatrogenic – due to medical care
• Nosocomial – acquired at hospital
• Vertical – from parent to offspring
• Horizontal – all other forms
• Germ line – part of the host genome

20

Viral Rashes

systemic infection – blood enter skin and deposit pathogen --> skin cells destroyed --> rash

21

Viral Dissemination from the site of entry

stages of a viral infection are:
1-5

chicken pox:
-symptoms occur at stage
-period
-1-4

stages of a viral infection are:
1. Local inflammation
2. Primary viraemia (virus in blood)
3. Amplification – increase the number of virus
4. Secondary Viraemia
5. Target Organ

Varicella Zoster Virus/ Chicken Pox:
-Symptoms occur after secondary Viraemia
-incubation period is around 14 days
-infection:
1. Virus enters the body
2. VZV can infect many cell types including peripheral blood mononuclear cells (PMBCs) & skin cells
3. From skin, infect sensory neurons where it remains latent
4. Virus can be reactivated when cellular immunity is impaired causing a painful rash at nerve endings - Shingles

22

Patterns of Viral Infection

a)
-afterwards,
-can be
-e.g.
-can be w/e.g. , (3)

b)
i-
ii- e.g.
1-3

c)
- may cause
- may encode
-by
e.g.
-H:
-E-B Virus:3
-P
-K and M
-H

a) Acute:
-afterwards, viral clearance, (e.g. colds / influenza) by adaptive immune system
-can be dangerous/ cause death – e.g. smallpox and dengue
-can be acute infection w/ accidental pathogenesis – e.g. polio (if infects the motor neuron), rubella (strong tropism for dividing neuronal cell tissue --> deafness, eye abnormalities [--> cataracts], congenital heart disease)

b) Persistent viral infections:
i - Chronic – low level replication in tissues which regen
ii - Latent – viral genomes maintained but no virus seen until episodes of reactivation when immunocompromised. e.g. Herpes Simplex Virus
1. enters your skin
2. into the nerve (don’t divide and live until death) in a latent state – moves into the soma cell by retrograde transport
3. When reactivation of virus, it begins to make new copies of itself and travels back to the end of the nerve and replicates through the skin (cold sore)

c)Oncogenesis:
-Viruses may cause cancer
-may encode an oncogene
-interferes with host cell cycle to enhance own replication: by making the host cell into a cancer the cell, virus can replicate more and move into new cells
e.g.:
-Hep B + C viruses: hepatocellular carcinomas
-Epstein-Barr Virus: Burkitt’s Lymphoma, Hodgkin’s Lymphoma, Nasopharyngeal Carcinoma
-Papilloviruses encode inhibitors of tumor suppressor genes and force the cell into S phase
-Kaposi Sarcoma-Associated Herpes virus and Merkel Cell Polyoma discovered by finding non-human genetic material in tumors
-HTLV-1: adult leukaemia

23

Outcome of Infection
8 (7)

• Viral sequence – may vary in virulence
• Viral Load – e.g. 2nd child higher dose
• Co-infections
• Co-morbidities:
o Asthmatics and respiratory viruses
o Obesity
o Immunosuppression
o Immunodeficiency
o Elderly
o Diabetes Mellitus
o Pregnancy
• Genetic resistance and susceptibility
• Other medications
• Host genetics
• Host age/gender

24

Evolution and Emergence of New Viruses

-viral quasispecies
-e.g.
-exists
1-4 -->

antigenic drift
e.g.
***

-viral quasispecies: grp of viruses related by similar mutation(s), competing within a highly mutagenic environment
-e.g.HIV
-quasispecies in a single infected host contains every mutation at every posn in genome
1 - virus may encounter bottleneck at transmission or during replication under limiting conditions
2 - high mutation rates & large progeny numbers & short replication time make viral evolution in response to selective pressures very fast
3 - Relative fitness of drug resistant virus vs wild type virus in vivo determines if drug resistant viruses proliferate
4 - Emergence of drug resistance as antiviral drug apply selection pressure; only resistant strains surviving
--> in some treatment, multiple drugs are used as most viruses not resistant to multiple drugs --> killed off by other drugs

Antigenic drift: spike proteins/hemagglutinin changes on a periodic basis --> current antibodies not being compatible; new antibodies needing to be generated to tackle the new strain of the virus
-e.g. influenza; changes roughly every year; best way to tackle this is through a yearly flu vaccination that best represents the current strain
***cold virus has all versions circulating at the same time, so vaccine cannot be made which contains all strains

25

new viruses develop
new virus =

5

New viruses are those which have only those recently discovered or detected

• Zoonosis - the crossing of pathogens, e.g. viruses from animals to humans.
• Increasing the genetic variation,
• By humans travelling around the world, it further increases the exposure.
• Further spread of the virus through vectors
• New discoveries

26

Global Influences on emerging infections
7

• Environmental modifications and demographics
• World Population
• Climate change
• Travel
• Farming practices and monocultures
• Immunosuppressed humans due to HIV
• Medical progress

27

Arboviruses: 4
i.e.

-impacted by 4
-normal hosts
-vector
-dead end hosts

WNF
-belongs to
-discovered
-outbreaks
-animals ill
-analysis
-further analysis
-cause

DHF
-areas
-serotypes
-risk factors 5
-antibodies

arboviruses: yellow fever, dengue, West Nile and chikingunya
-i.e. Flaviviruses (grp of RNA viruses that cause a number of serious human diseases)

-impacted by global warming, decrease in mosquito control, imports, stagnant water in large cities and dams
-Wild birds act as the normal hosts,
-mosquitoes act as the vectors, transferring it to other birds, humans & horses
-Humans and horses are dead-end hosts as the bacteria cannot multiply as effectively in these organisms; mosquitoes cannot get the infection from humans or horses

West Nile fever in New York:
o belongs to Japanese encephalitis group of flaviviruses
o discovered in 1937, but only caused mild diseases
o several reported outbreaks of the condition, e.g. Eastern Europe, New York in 1990 (7 unexplained deaths)
o Concurrently, birds in Bronx Zoo & wild crows became ill
o A subtractive differential analysis used to diagnose a human brain sample revealed WNV DNA
o RT-PCR showed sequence identity with a virus circulating in Israel
o exact cause was unknown as to why it spread to New York from Japan

DHF (Dengue Hemorrhagic Fever):
o Areas with the mosquito species Aedes aegypti linked to dengue fever
o DHF as 4 serotypes of dengue fever: x-reactive, but no cross protective; antibodies will not protect the person against variant fever strain
o Risk factors:
Virus strain
Pre-existing anti-dengue antibody from either previous infections or maternal antibodies in infants
Age
Higher risk in secondary infections
Higher risk in locations with 2 or more serotypes circulating simultaneously at high levels
o Dengue antibodies can either neutralize infection or enhance it: may not block but instead bind to virus --> have Fc receptors --> enter other cells which it would not normally enter --> infection to spread--> DHF

28

Human Viruses from animal sources

-e.g. S
-e.g. H
-e.g. other 3

**

-e.g. SARS-CoV: virus from bats / civets
-e.g. HIV: from chimpanzees and sooty mangabeys
-e.g. ebola, Hendra, Nipah (can spill over, but don’t transfer as efficiently)

**Bats are particular a problem, as harbor many viruses – and increased exposure of human populations and domestic animals has increased the number of viruses which humans can get infected by

29

Coronaviruses as a threat to human health

main grps of coronaviruses: group 1 + 2 contain mammalian viruses and can cause disease in humans, and group 3 which only contains avian viruses
-Human coronaviruses NL63 and HKU1 --- more severe hospitalized cases

SARS:
o virus form of coronavirus: enveloped, interact with the receptor ACE-2 protein in humans
o almost identical virus isolated from masked plam civets & raccoon doa in wet markets
o Chinese horseshoe bats harbor SARS-2 like coronaviruses that can use bat and human ACE-2 as receptors
o S protein is highly plastic and can adapt to different receptors overcoming host range barriers
o variety of cases beginning in China, 2002
o transmitted by infected respiratory droplets **conc virus sources in lift buttons / bathroom drains. amplified by nebulizers in hospitals
o Children only experienced a mild disease, patients over 60 had a high mortalityo most infectious around 10 days after catching it; same time symptomatic; increased spread rapidly
o Destruction of lung tissue from over exuberant immune response
o emergency response coordinated, which was international and rapid

MERS:
o tourism & business travel has spread the virus beyond the middle east
o closely related to HKU4 and HKU5 – both bat coronaviruses
o high seropsotivity in camels – i.e. most prevalent in camels
o ARDS in older infected people but can be asymptomatic in infected younger people
o The DPP4 receptor in the lungs is targeted – involved by diabetes; diabetics greater risk
o not readily transmitting between people but people come together for pilgrimages; could easily spread due to the large number of people who attend
o limited transmission, but diverse clinical signs i.e. hard to diagnose, no vaccine and no antiviral treament

30

Influenza

-influenza virus potential to become problem because of the variety of viruses

-Due to the similarities, reassortment could lead to antigenic shift: 2 viruses come together to form a new virus
--> variety of pandemics over the years

e.g. Swine Flu Pandemic in 2009:
o subtypes caused swine flu pandemic in the 1920’s was the same in pigs and humans
o as humans live longer, surviving viruses slowly mutated --> unrecognizable; new virus i.e. pandemic in 2009
o as pigs are killed rather quickly, virus remained the same in them

e.g. H7N9 Influenza:
o growing concern: increasing no. of reported cases in China
o effected high proportion of elderly men inked with infected poultry in wet markets
o poultry showed no symptoms
o virus already shown human adaptive changes e.g. mutations in HA receptor binding sites and polymerase adapted for human cells
o limited transmission, no vaccines (but some underway), antivirals available but well tolerated

-Man’s intervention as the cause of emerging viruses:
o Myxoma virus released to control rabbit populations in Australia – but mutated: rabbits becoming resistant to it --> problem
o Flu conrovery Spurs research moratorium genetically manipulating H5N1 influenza viruses to see the effect of adding in mutations – this may create a deadly or lethal strain