Influenza Flashcards
Influenza Basics
What are they called?
What do they look like eg enveloped or not?
What time of genome?
Proteins in the particle?
Viruses are called orthomyxoviruses
They are enveloped (transmitted by bloody fluids and are easily inactivated by simple detergents)
Segmented, negative sense RNA genome (therefore they must have RNA dependent RNA polymerase packaged in the virus)
- Positive strand can then be used by the normal cellular machinery to make viral proteins
- make new minus strands from positive strands which form genetic material for new viruses
see agglutinate RBCs in the picture
Influenza Basics
What are the Surface (spike) proteins?
Hemagglutinin (HA)
- target for neutralizing antibodies, binds to sialic acid (the virus receptor) on the surface of cells, and mediates membrane fusion and virus entry
Neuraminidase (NA)
- cleaves sialic acid, enables new visions to be released from the host cell (otherwise they would just remain stuck on the surface by remaining bound to sialic acid)
Epidemics Timeline
Influenza—> seasonal epidemics: 35,000-40,000 people die of flu each year in the US
- usually people die from secondary bacterial pneumonia
- peak in winter
- influenza doesnt confer lifelong immunity becuase the virus constantly changes
- influenza is always ENDEMIC but there can be seasonal epidemics whose severity varies (track excess deaths to see if it is a bad influenza year
- rarely it is pandemic (worldwide epidemic)
Examples of occasional pandemics (worldwide epidemics)
1918 spanish flu, 500,000 deaths in the US> 40 million worldwide
1957 Asian flu 70,000 in US (could infect a lot of people but not kill them, less virulent than spanish)
1968 Hong Kong Flu 34,000 in the US
2009 Swine Flu 12,000 in the US
Key point: influenza virus is endemic throughout the world, some years incidence rates are higher causing epidemics and more rarely pandemics occur
Influenza Clinical findings
Respiratory transmission: replications in respiratory epithelium
Acute self, limited infection
Incubation period (1-4 days)
abrupt onset of fever, myalgia(headache), sore throat, nonproductive cough, generalized muscle aches and malaise
Viremia (virus detected in blood)rare; shed in respiratory secretions for 5 - 10 days
Otitis- ear infection frequent in children
Ab and cell mediated immunity clear viruses in 1 - 2 weeks though full recovery can take longer
Repair of respiratory damage > 1 month
Viral pneumonia - histology
- Relatively little intra-alveolar inflammation:
- viruses infect cells so you get interstitial inflammation (because they are intracellular this leads to mononuclear inflammatory infiltrate that is interstitial)
- bacteria grow in airspaces so you get alveolar inflammation
- bacterial pneumonia associated with productive cough
lymphocytes in alveolar walls
* - Variable interstitial inflammation, mostly chronic (lymphocytes)
- Often affects bronchioles/bronchi, sometimes without alveolar involvement
- Predisposes to 2ndary bacterial pneumonia (polys in airspaces)
- Sometimes cytopathic effects (CMV, RSV)
Viral Pneumonia Histology Vs Bacterial Pneumonia (slide 8)
Viral pneumonia’s: typically more diffuse, increased interstitial markings, airspaces not involved but alveolar walls are thickened
Bacterial pneumonia: more localized, more consolidation (whiting out) as airspaces are obliterated
Two major complications of Influenza
Secondary Bacterial pneumonia
- damage to ciliated epithelial cells—> decreased mucocilliary clearance—> increased risk for bacterial pneumonia (because you have lost your cilia)
- induces mucus section; S Pneumonia produces neuraminidase that cleave sialic acid from mucus, provides sugar source, rapid bacterial growth
- can rarely see brain infection (encephalopathy), heart infection (myocarditis, pericarditis)
- mortality: ~.1% of those infected; highest in < 1 year old, and elderly but higher in a pandemic
Reye’s syndrome
- combination of influenza + aspirin in an infant can result in Reye’s syndrome: fatty liver change, acute encephalitis, high mortality
- if you have a child that has the flu (any upper respiratory stuff) DO NOT give them aspirin
Diagnosis of Influenza
Many different types of viruses circulate during the winter months
- influenza A, B, respiratory Syncytial Virus (RSV), adenovirus, parainfluenza types 1, 2, 3
- accuracy of clinical diagnosis of influenza ~ 50%
- assisted by epidemiological information - is flue currently circulating in your community ?
- rapid diagnostic kits (use nasal swabs), low sensitivity and high specificity, can distinguish A from B but not type of influenza A
**real time PCR tests are used and can distinguish virus strains *** only use this though if child has difficutly breathing
Three types of Influenza
**antigenically they are different enough such that ab to influenza A do not cross react with influenza B or C **
C: infects only humans, no epidemics, relatively rare, only causes minor respiratory symptoms, not covered by vaccine
B: infects only humans, mostly children, milder disease, can cause minor epidemics, not many strains, included in vaccine
A: infects people and many different kinds of animals, especially birds (avian influenza), many different strains of influenza A that infect many kinds of animals (people, horses, birds etc)
- large animal reservoir, highly variable, many strains, included in vaccine
- cause sof most epidemics and all pandemics
A is worse than B which is worse than C
Influenza A virus
Influenza is an enveloped virus, what three virally encoded proteins are protruding from the viral membrane?
AKA what are the Three surface proteins?
Hemagglutinin = H(A)
- binds to sialic acid, the receptor for the virus
- at low pH, undergoes conformational changes that mediate membrane fusion and viral entry
- HA is the major target for neutralizing antibodies and is the most important component of the flu vaccine
Neuraminidase = N(A)
- enzyme that cleaves sialic acid (receptor HA binds to)
- when a new virus buds from the cell surface, it needs to be released
- since there is a lot of sialic acid (sugar found on lipids and many proteins) the new virus may just remind to the surface of the cell from which it has emerged
- with time, NA will cleave the sialic acid to which the virus is bound, releasing the virus
- target of antiviral drugs (Tamiflu, a drug)
M2 ion channel
- small integral membrane protein that forms a homotetramer
- is a drug target (amantadine)
- most influenza viruses are now resistant to it
key point: HA is the most important viral protein with regards to vaccines, while NA and M2 are drug targets (mostly NA)
Influenza Segmented Genome
Segmented genome = 8 pieces of RNA which complex with the nucleocapsid protein
- one copy of each RNA must be incorporated to make an infectious virus
**key point: influenza has a segmented genome which is important for the production of pandemic influenza strains that arise via ANTIGENIC SHIFT
Influenza variability
Influenza mutates a lot (every year for thousands of years)
Many different influenza A strains
- some infect people, others, pigs, birds etc
Strains called subtypes designated by numbers
If you get one subtype of A for example (like H1) you will get immunity to that but not to the other subtypes eg h2-4
Influenza Strains
Subtypes based on H & N proteins (HA AND NA)
Subtypes designated by numbers and appear in different combinations
Infection with one subtypes gives No immunity to other subtypes
16 different HA subtypes and 9 different NA subtypes
H1N1, H2N2, H3N2, H4N3, H5N1, H6N2, H7N9 etc, different HA and NA subtypes can appear in different combinations
Antibodies to H1 virus do not react to viruses with other subtypes and visa versa so infection with H1 virus does not give you immunity to H2, H3, H4, H5
Birds, especially water flows, are natural reservoirs for influenza A, all influenza subtypes can be found in bird populations
NB influenza B and C dont have subtypes, they do evolve each year so there are different B and C strains but not as variable as influenza A
- only infect people
Where can all influenza subtypes be found?
Bird populations
What is the seasonal Flu?
H1N1 and H3N2 influenza viruses have been circulating in humans (for 30 years) = seasonal flu
Viruses mutate every year
See card on ANtigenic drift
Describe Antigenic Drift as it relates to Flu Vaccines/ immunity
H1N1 and H3N2 = seasonal flu
ANTIGENIC DRIFT: If you get H3N2 seasonal flu this year you will develop immunity, next year the virus will be somewhat different but you will probably still be immune for that year plus 1 - 2 additional years but overtime seasonal flu accumulates enough mutations to be ‘new’
- plains why in the absence of vaccine, most people get infected with flu multiple times over the years
Summary:
- seasonal flu mutates
- changes a little every year
- seasonal flu vaccine changes each year to keep pace
- this is why you should get vaccinated each year
How does Antigenic Drift Work? (Influenza)
Amino acid changes (mutations) eliminate existing Ab binding to HA protein
There are 4 - 5 Ab binding sites on HA
With time all sites change and immunity is lost, lose antigenic structure
Leads to seasonal epidemics
Only antibodies to HA neutralize the virus
Eg if you get flu and become immune the next year 2/5 epitopes on Virus might have changed so Ab for those epitopes dont bind but the others will so you ares till immune but in another year or two those sites will have also changed so you are no longer immune
What causes Flu pandemics?
ANTIGENIC SHIFT (not Antigenic drift)
- major change in HA or NA subtype resulting from reassortment, occurs when a cell gets infected with two different viruses (each with different HA and or NA subtypes)
** this is why the segmentation of the genome is important becuase it makes reassortment easy**
- eg Avian influenza virus + Human influenza virus—> two viruses infect one cell and reassortment—> chemiric human influenza virus with avian surface HA proteins
- eg virus is infected with two different kinds of influenza (H3 and H2)
- since their genome is segmented one of the 8 segment codes for the HA
- a new virus can emerge that has all of the RNA segments for H2 virus, but instead of the piece of RNA that codes for the H2 HA protein, it picks up RNA that codes for the H3 Ha protein—> new virus that has an entirely new HA
- if H3 HA had never before been seen in people, then no one in the world would be immune so this can give rise to pandemic
Eg 1968 Hong Kong flu
Why is the Avian reservoir important? How can pigs provide a mixing vessel for human and avian flu
**there is a huge reservoir of influenza A viruses in birds and other animals - these serve as a source for new pandemic strains of influenza virus through antigenic shift **
- people and birds have different types of sialic cid so avian flu doesnt bind well to human sialic acid and vice versa, providing a barrier to cross-species transmission
- pigs have sialic acid receptors for both avian and human influenza in their tracheas
- domestic pig supports the growth of human and avian viruses, allowing it to provide a mixing vessel which can lead to reassortment and Ag shift, enabling the new virus to adapt (mutate) to use human sialic acid
- usually avian flu infects humans poorly but it can adapt in pigs to recognize human sialic acid
Eg avian H3 + human H2–> pig—> new human flu with avian H3 (derivation of the Hong Kong flu of 1968)
Distribution of sialic acid types in the human airway
Human sialic acid is found throughout the respiratory tract (human flue binds alpha 2, 6 sialic acid)
Avian sialic acid is found only deep in the respiratory tract (avian flu binds alpha2, 3 sialic acid), lower airway
- distribution explains why high inoculate of bird flu are needed to infect a person, explains why most who die from avian flue have direct contact with poultry/waterfowl because the only way to get infected is by breathing in a lot of the virus and then having some of that virus find its way deep in the lungs where the right sialic acid exists
Antigenic Shifts in History and Humans
Great spanish flu of 1918 - H1N1 virus
- highly virulent; killed millions, spread rapidly throughout the world
- for next 40 years it circulated in humans as seasonal flue, changing each year via antigenic drift
- Ag drif—> viral evolution with occasional epidemics
Asian Flu of 1957 was an H2N2 (replaced H1N1 and then became our seasonal flu with continual Ag drift)
Hong Kong Flu pandemic of 1968 (H3N2), HA changed, continue to circulate in humans as seasonal flu and continue to evolve by Ag drift
1977 Russian Flu: closely related to H1N1 (probs released from research lab in Russia or northern China) didn’t supplant H3N2 then circulating, mostly infected younger people since older people had immunity from earlier H1N1 infections
Swine flu of 2009 was an H1N1
- H1 subtype found in swine influenza strains introduced into human population in Mexico
- different enough from old H1N1 to cause a pandemic
- now the only H1N1 seasonal flu
- not super virulent
- virus skewed young - maybe due to cross protective immunity in older people from previous infections with H1N1
Avian Influenza - H5N1, since 1997 sporadic outbreaks in Southeast Asia,
And Since 2013 sporadic outbreaks of H7N9 bird flu
- pretty virulent, worry they will adapt to human sialic acid and cause new and deadly pandemic
What is the next pandemic?
H5N1 Avian flu
- starting in 1997, yearly outbreaks in humans
- high mortality rates, kills young adults without bacterial superinfection so it is super virulent
- mostly individuals who work directly with poultry and close contacts in Southeast Asia
- spreads human to human poorly, fear they will adapt to humans and cause a new pandemic
Or H7N9 avian flu
Huge reservoir for avian influenza viruses + cultural practices (live bird markets, pop density, small farms with poultry and pigs) make Southeast Asia an ideal location for generation of new pandemic influenza
Will avian flu cause a serious new pandemic?
What is needed for a serious pandemic? (Does H5N1 have it)
Novel virus - little or no immunity (yep- H5 is not part of our vaccine regiment)
Capable of causing disease in humans (yep)
Highly pathogenic/virulent (needed for bad pandemic) (yep)- doesnt have to be virulent because novel H1N1 was not terribly virulent
Capable of sustained person to person transmission (not yet because sialic acid isnt adapted for people
Spanish flu pandemic 1918
- young and old died but also a spike in deaths of young adults is what made the 1918 flu unusual
- one of the greatest pandemics, infected > 500 million people, about 50 million died
- people betweeen 15 and 34 years old most differentially affected - death rate 20x higher than previous years for people WITHOUT a bacterial superinfection (which is what the avian flus are now doing which is concerning)
- approximately half of soldiers who died in WWI died of flu
- 2.5% mortality rate compared to normal .1% rate
**highly virulent, often killed people directly without bacterial superinfection
Death carts in 1918 in philadelphia
Resurrecting the 1918 influenza virus
Molecular archeology!
Pathological specimen (circa 1918) because the army doesnt throw anything out so you could find people who died back then and get their preserved lung tissues —> gene sequencing and PCR to amplify flu genes —> gene reconstructions —> reverse genetics—> phenotypic characteriziation in tissue cultures and animal models
- Jeff Taubenberger
After seeing this molecular archeologists Johan Hilton excavated a mass grave in Alaskas Seward Peninsula where in 1918 72/80 ppl died from flu in 6 day period—> mass grave preserved by permafrost
- found an obese woman in her mid20s when she died of influenza, whose tissues were rather well preserved
1918 Virus - what do we know now from the reconstructed viruus
Far more virulent in animals
Triggers aberrant innate immune responses:
- less IFN production, so likely more virus replication
- much higher levels of cytokines - leads to vascular permeability, pulmonary edema (induces cytokine storm leading to pulmonary edema and respiratory fluid )
Flu Vaccine
Expensive guess as to what the seasonal flu is going to look like a year later
Decisions about what strains to include are made in Jan/feb
Virus selection—> production begins—> FDA testing licensure—> filling and packaging—> product release—> vaccine begins oct-nov (immunity develops 2 weeks after vaccination
All flu vaccines contain the HA proteins from 1 or 2 influenza B, one H1N1 and one H3N2 virus, these change on a yearly basis
WHO has a worldwide network of inelunza surveillance labs, in Jan/Feb of each year, decision is made about the three viruses that will compose the influenza vaccine in the coming winter, takes 6 mo to make the vaccine
Vaccine contents and different types (influenza vaccine)
All contain the HA proteins from 3 or 4 influenza strains, these can change each year
Three types of vaccines that can deliver these HA proteins
Inactivated vaccine: produced in eggs (flu is grown in embryonated chicken eggs, partially purified and inactivated)
- partially purified, inactivated, most common
- delivered IM and is the oldest and most common
Life ATtenuated (FluMist) - produced in eggs, virus is adapted to only grow at lower temperatures, and then is engineered to express whatever HA proteins are recommended for the yearly vaccine
- virus replicates in the nose and induces good immunity
- produced in eggs
- squirt this mix of viruses up your nose and because it is adapted to grow at lower temperatures it wont really spread
- dont give to young, old, pregnant, immunosuppressed
Subunit Recombinant (FluBlok) - licensed in 2013, new, relevant HA proteins are produced in insect cells, so you use this vaccine for people with egg allergies
Seasonal Flu Vaccine Examples/ Trivalent/Tetravalent influenza virus vaccine
Seasonal flue vaccine, influenza B, A H1 and A H3 viruses
Increasing number of vaccines include four viruses (two influenza Bs)
Virus Recommendations
Recommended for all persons > 6 months
Children 6 mos - 8 yrs: two doses 4 weeks apart
High dose shot used for persons > 65 yo
Life attenuated preferred for children 2 -8 yrs
Life attenuated shouldn’t be used for
- < 2yo, > 49 yo
- pregnant women
- immunocompromised
- persons with history of egg allergy
- children 2 - 4 with asthma
Persons with egg allergies:
- can give standard vaccine to people who have mild reactions to egg products
- can use recombinant flu vaccine
WHEN IN DOUBT—> inactivated vaccine can be given to anyone save for those with significant egg allergies
Differences in use
Live attenuated: small pox, yellow fever, mumps, measles, rubella, flu mist, rotavirus, chickenpox, Sabin polio
Killed/inactivated: influenza, hep A, Salk polio, rabies
Subunit: HepB, HPV
Two Antiviral flu drugs
amantadine
M2 ion channel is the target for amantadine
- “uncoating” - M2 ion channel inhibitors - amantadine
- when virus enters the cytoplasm, capsid falls apart buut then a few hours later same RNA segments and NP proteins assemble to form new capsids
- M2 protein is a proton channel and tells the NP protein to dissasemble and assemble
- when virus enters an endoscope, not only is HA triggered to cause fusion,, but protons flow through the M2 Channel into the viral core—> acid bath to the viral capsid and triggers it to dissociate so that nuclear transport can then occur
- newly synthesized NP protein has never seen acid pH so it is ‘different’ than NP that enters to the cell
- acid pH is the switch that regulates viral assembly and disassembly, causes a conformational change in HA
NB influenza is unique in that it replicates in the nucleus but normally RNA viruses replicate in the cytoplasm and DNA in the nucleus
Amantadine (symmetries) and Rimantidine (flumadine) = small molecule inhibitors of M2 protein, bind to channel and prevent acidification of viral core—> prevent virus disassembly and prevent infection, but most drugs resistant to this now
Two Antiviral drugs:
Neuraminidase inhibitors
Oseltamivir and related molecules are both NA inhibitors
Can ameliorate symptoms but dont stop infection
By inactivating NA they limit virus spread - virus particles emerging from an infected cell bind to the host cell sialic acid, and cannot ‘cleave’ themselves off of the receptor so spread to adjoining cells is inhibited
Influenza Antivirals
M2 ion channel inhibitors
amantadine (symmetrel), flue type A, used for treatment and prevention, treatment age > 1 yr
Rimantidine (flumadine), flu type A, used for treatment and prevention, treatment in adults
Neuraminidase inhibitors - block virus release from cell surface
- zanamivir (relenza) treat A/B, used for treatment, > 7 yo, inhalation
- oseltamivir (Tamiflu), treat A/B > 1 yr for brevention and treatment, oral
Most viruses circulating are resistant to amantadine, drugs that target NA are usually effective in limiting virus spread
Practice Q:
Antigenic Shift is influenced by all except
A. Animal reservoirs for influenza virus
B. Accumulation of mutations in Ab epitopes
C. Presence of a segmented genome
D. Confection of cells
E. Sialic Acid linkages
B becuase this is antigenic drift and leads to epidemics
A. .presence of such reservoirs is why flu A but not flu B or c can cause pandemics
C. Makes reassortment possible
D. Without this you cant have reassortment
E. For viruses to coninfect a cell, they have to recognize sialic acid linkages on that cell
Mr. Johnson, 60 yo retired pilot from Thailand gets fever, myalgia, non productive cough after a severe and sudden attack of tiredness
Influenza! Prescribe rest and analgesics and he feels better
BUT then he develops sudden shaking chill and cough producing rusty sputum
One is feeling better and then worse…. what’s going on
A. Influenza pneumonia
B. Other viral pneumonia
C. Rhinotracheitis from influenza
D. Bacterial pneumonia
D!!
For influenza, al low the following are targets for antivirals or neutralizing Ab except:
A. Hemagglutinin
B. Polymerase
C. Neuraminidase
D. M2 protein
B!!
A. Target of neutralizing AB
C. Tamiflu- prevent virus release
D. M2 amantadine prevent virus uncoating by preventing protons from flowing through M2 ion channel in the viral membrane following endocytosis
