Epidemiology and pathogenesis of influenza

The orthomyxoviridae contains 3 types of influenza
There are however many hundreds of different influenza strains belonging in these 3 groups 

Naming 
1. Subtype
2. Location of the 1st isolation
3. Number of isolation
4. Year of 1st isolation 
5. Information of HA and NA proteins

e.g Current swine flu circulating 
-- A/California/04/2009 (H1N1)
e.g 1st case of H5N1 bird flu in humans 
- A/Hong Kong/156/97 (H5N1) 

There are currently 18 HA's, 9NA's

• Distinguished on basis of reactivity with a panel of antibodies
• Different HA's have over 30% AA differences
• All HA's proteins are structurally v.similar
• HA is cleaved by cellular proteases to give HA1 and HA2
  - Sequence changes between H1-H18 do not grossly affect overall structure. The function of all HA molecules is likely v.similar/identical.

Different HA groups can infect different hosts: not all infect humans

One determinant of host range is specificity of receptor binding (HA-SA) 
- HA must recognise sialic acid on the cell surface during virus entry
- SA can be linked to galactose via either alpha 2-3 or alpha 2-6 linkages

In general, bird viruses have HA that will preferentially attach to SA with alpha 2-3 linkages
Humans have mostly alpha 2-6 linked SA on their cells so are not infected 
- This sets up a species barrier that protects humans from infection by many avian influenza viruses

Many HA proteins are able to attach to both alpha 2-3 and alpha 2-6
- e.g H1, H2, H3
- Highest affinity for alph 2-3 SA (Avian cells) causing frequent infection in birds
-But some affinity for alpha 2-6 SA causing rare infections in humans

• Low pathogenicity 'seasonal' influenza virus strains tend to be H1, H2, H3
• Other low pathogenicity strains infecting animals ted to be H1-H4, H6, H8-18
• H5+H7 virus is different 
• Have repeatedly decimated chicken populations in China, hundreds of millions 
• In humans, H5+H7 strains are not restricted to respiratory tract- can infect other cell types (kidney, spleen, brain)
• Known as HPA1=High pathogenic avian influenza viruses.

• Different NA's are also at least 30% different 
• Also show differences in preferred host 
• Different activities relate to their ability to cleave hosts sialic acid 

All influenza virus types can infect birds, Only a few infect humans (N1+N2) (H1,2,3,5,7,9) 
Influenza virus has actually evolved as a bird virus 
And 1 that just happens to infect other animals 

1. Virus inhalation via respiratory tract
- A sneeze can contain 10,000 droplets
- 1 sneeze contains 1,000,000 virus particles at peak of virus shedding 
- Coughs tend to bring up deep lung material 
- Small droplets stay airborne for many minutes
- Exhaled breath contains individual virus particles
- Viruses directly enter respiratory tract and their intended cell targets
- It is very difficult to prevent transmission through virus inhalation 

2. Direct contact 
- Sneezing, coughing and breathing generate droplets that settle on surfaces (surfaces are called fomites)
- Influenza virus is relatively stable on hard surfaces 
- Stable on money for 17 days 
- Contact between these surfaces and mucosal cells causes direct inoculation
- Normally mediated by contaminated hands and fingers
-It is easy to prevent contact transmission by practicing good hygiene

Influenza virus targets respiratory tract

• Primary sites of infection are epithelial cells lining the entire respiratory tract
• Symptoms: 
  - Sore throat
  - Tiredness
  - Headache
  - Fever
  - Muscle aches
  - Runny nose 
  - Cough 
• Symptoms mostly due to host immune response 
• Rapid onset of symptoms, 24 hours after contact
• Symptoms peak at day 3, innate immunity peaks
• Decline in virus due to activity of innate immune system, finally removed by adaptive response

• Timing of flu season depends on hemisphere
• Incidence increases in both North and South Winters
  - Peak in N= Weel 7
  - Peak in S= Week 29 

Possible reasons for seasonal peaks 
- Sunlight, UV exposure
- Low temperature
- Low relative humidity
- Social behaviour: Indoor crowding 
- National holidays, return to school 

Used hamsters as an infection model system 
Incubated infected hamsters alongside infected
Varied incubation conditions 
- Showed transmission best at low temperature, below 5 degree celsius 
- Transmission best at low humidity, 20-35%

Low temperature 
- More virus released
- Virus particles most stable at low temperatures
- More virus contact

Low humidity 
- Exhaled droplets smaller -stay airborne longer
- Increased chance of droplet inhalation
- More virus contact

Conditions found in these weeks. These neatly explains the seasonal transmission in temperate regions. Winters are cold with low humidity- thus virus can exist in droplets and on surfaces for longer

• Caused by an exceptional virus 
• It infected 30% of worlds population 
• Mortality was so high it caused a massive dropl in global life expectancy 

- Fatality rate was around 10%

This was the most deadly event in recorded human history
No live 1918 virus stored as medical samples
There were many attempts 1951-2005 to find stored clinical samples- All failed due to sample decomposition 

Jonah Hutlin identified mass grave in Alaska that remained frozen for almost 100 years
They were monks from a missionary, 72/80 ied in 5 days in 1918
2005, bodies exhumed and preserved lungs were removed and RNA was extracted

After much effort the complete genome of over 13,000 nucleotides sequenced. It was a HIN1 virus 
- Recreated virus exhibited profound pathogenicity in mice, ferrets and macaques
- Swapped segments with seasonal virus to identity pathogenesis factors. 

• Seasonal flu strain gives only mild disease in mice
• Chimera with HA segment from 1918 virus 
• 100% lethal infection in mice

Seasonal flu strains gives only mild disease in ferrets
- NP, PB1, PB2, PA segments from 1918 virus added to seasonal flu virus
- Chimeria behaved same as 1918 strain 
- Severe pathogenesis of 1918 virus is polygenic- HA, NP, polymerase

Reasons for severity of 1918 pandemic are still not well understood
- One factor appears to be cleavability of HA protein
- HA0 must be cleaved into HA1 and HA2.
- Protease required for most HA cleavage is only in respiratory tract cells- Human airway trypsin (HAT)
- This confines most flu viruses to respiratory tract cells
- 1918 influenza virus has HA protein that could be cleaved by other proteases found within many cell types. 
- Virus could infect cells of other organs and tissues, HA in H5 and H7 virus can also be cleaved in this way. 

• For every virus that enters a cell, 10,000 come out 
• Influenza virus RdRp makes mistakes when copying 
  - Error rate is around 1x10-4 
  - 1 mistake in 10,000 nucleotides
  - Influenza genome is around 13,000 nucleotides
  - Each virus is different 
• Most drift changes are on HA1
• HA1 has low functional restrictions- mutations do not always affect function 
• Some mutations are a benefit- The host immune response cannot recognised new viruses 
• Most drift changes are outside receptor binding site 
• There are few changes here as this would affect HA's ability to bind SA

• Occurs when segments mix
• Depends on 2 different influenza viruses infected the same cell
• Reassortment between human-human or avian-avian is more likely 
• Species barrier largely prevents human and avian viruses infecting same cell

The pig often allows humans and avian strains to mix
- Pigs have both alpha 2-3 and alpha 2-6 linked sialic acids on cells
- Human and avian viruses can infect pig cells at the same time 
- Resulting viruses are very different to previous species 
- Little immunity 
- Causes serious disease, pandemic

• Shift is a relatively rare event 
• Shift has been responsible for all 4 pandemic events 
• 1918 H1N1- Spanish flu ( 50 million deaths)
• 1957 H2N2- Asian flu (2 million deaths) 
• 1968 H3N2- Hong Kong (2 million deaths)
• 2009 H1N1 swine flu ( 0.5 million deaths) 

1918 and 2009 are closely related 

Example of shift: 
Current H1N1 swine flu virus has segmented from 4 different viruses
1. Initial triple event occured between avian, swine and human viruses
2. A 2nd reassortment event occured later 

Virus emerged in 2009, still predominant circulating strain