The "Flu"

Influenza is a respiratory infection characterized by fever, cough, and severe muscle aches. In the elderly and infirm, it is a major cause of disability and death (often as a result of secondary infection of the lungs by bacteria). Even in the young and healthy, influenza produces a prostrating disease of a few days duration and one not soon forgotten.

• a case of low fever and sniffles that keeps you home in bed for a day
• a gastrointestinal upset ("stomach flu")

In February 1997, Ann Reid, Jeffery Taubenberger and their colleagues reported (in the 21 March 1997 issue of Science) the partial sequences of 5 influenza genes recovered from the preserved lung tissue of a U.S. soldier who died from influenza in 1918.

• The soldier was one of an estimated 21 million people worldwide who died in the most devastating plague in human history. (A disease that attacks a large fraction of the population in every region of the world is called a pandemic.)
• No one at the time even knew what disease agent was causing the pandemic.
• Influenza is still with us. The knowledge of the genome of the 1918 virus may provide clues to help us avoid another pandemic.

This electron micrograph (courtesy of Dr. K. G. Murti) shows several influenza virus particles (at a magnification of about 284,000x). The surface projections are molecules of hemagglutinin and neuraminidase (see below).

• Influenza C
  Common but seldom causes disease symptoms
• Influenza B
  Often causes sporadic outbreaks of illness, especially in residential communities like nursing homes.
• Influenza A
  Responsible for regular outbreaks, including the one of 1918. Influenza A viruses also infect domestic animals (pigs, horses, chickens, ducks) and some wild birds.

The Influenza A Virus

• a globular particle (about 100 nm in diameter)
  
  
• sheathed in a lipid bilayer (derived from the plasma membrane of its host)
  
  
• Studded in the lipid bilayer are two integral membrane proteins
  
  
  • some 500 molecules of hemagglutinin ("H") and
  • some 100 molecules of neuraminidase ("N")
• Within the lipid bilayer are
  • some 3000 molecules of matrix protein
  • 8 pieces of RNA

• many copies of a nucleoprotein
• several molecules of three different RNA polymerases
• some "non-structural" protein molecules of uncertain function

The Genes of Influenza A

1. the hemagglutinin. 3 distinct hemagglutinins, H1, H2, and H3) are found in human infections. Nine others have been found in animal flu viruses.
2. the neuraminidase. 2 different neuraminidases (N1 and N2) have been found in human viruses; 7 others in other animals.
3. the nucleoprotein. Influenza A, B, and C viruses have different nucleoproteins.
4. two matrix proteins (encoded by different reading frames of the RNA)
5. two different non-structural proteins (also encoded by different reading frames)
6. one RNA molecule for each of the 3 RNA polymerases

The Disease

• Its hemagglutinin molecules bind to carbohydrate on the glycoproteins of the epithelial cells of the host.
• The virus is engulfed by receptor mediated endocytosis.
• The drop in pH in the endosome (endocytic vesicle) produces a change in the structure of the viral hemagglutinin enabling it to
• fuse the viral membrane with the vesicle membrane.
• This exposes the contents of the virus to the cytosol.
• The RNA enter the nucleus of the cell where fresh copies are made.
• These return to the cytosol where some serve as mRNA molecules to be translated into the proteins of fresh virus particles.
• Fresh virus buds off from the plasma membrane of the cell (aided by the neuraminidase) thus
• spreading the infection to new cells.

The result is a viral pneumonia. It usually does not kill the patient (the 1918 pandemic was an exception; some victims died within hours) but does expose the lungs to infection by various bacterial invaders that can be lethal. Before the discovery of the flu virus, the bacterium Hemophilus influenzae was so often associated with the disease that it gave it its name.

Pandemics and Antigenic Shift

• The "Asian" flu pandemic of 1957 and
• the "Hong Kong" flu pandemic of 1968.

(The pandemic of 1957 probably made more people sick that the one of 1918. But the availability of antibiotics to treat the secondary infections, that are the usual cause of death, resulted in a much lower death rate.)

The hemagglutinin of the 1918 flu virus was H1, its neuraminidase was N1, so it is designated as an H1N1 "subtype". Here are some others.

These data suggest that flu pandemics occur when the virus acquires a new hemagglutinin and/or neuraminidase. For this reason, when an H1N1 virus appeared in a few recruits at Fort Dix in New Jersey in 1976, it triggered a massive immunization program (which turned out not to be needed).

• Perhaps they have been retained by viruses hiding in isolated areas.
• Perhaps they have been acquired from some other animal.
  • H3 was found in birds and horses five years before it appeared on the Hong Kong virus.
• Perhaps the encoding of H and N by separate RNA molecules facilitates recombination in animals simultaneously infected by two different subtypes.
  • H3N2 virus has been recovered from pigs simultaneously infected with swine flu virus (H1N1) and the Hong King virus (H3N2).

Epidemics and Antigenic Drift

Missense mutations in the hemagglutinin (H) gene.

• wait until a new crop of susceptible young people comes along
• change the epitopes on the hemagglutinin molecule (and, to a lesser degree, the neuraminidase) so that they are no longer recognized by the antibodies circulating in the bodies of previous victims.
  • By 1972, the H3 molecules of the circulating strains differed in 18 amino acids from the original "Hong Kong" strain
  • By 1975, the difference had increased to 29 amino acids.

The gradual accumulation of new epitopes on the H (and N) molecules of flu viruses is called antigenic drift. Spontaneous mutations in the H (or N) gene give their owners a selective advantage as the host population becomes increasingly immune to the earlier strains.

Flu Vaccines

Although a case of the flu elicits a strong immune response against the strain that caused it, the speed with which new strains arise by antigenic drift soon leaves one susceptible to a new infection. Immunization with flu vaccines has proved moderately helpful in reducing the size and severity of new epidemics.

• an A strain of the H1N1 subtype
• an A strain of the H3N2 subtype and
• a B strain.

Because of antigenic drift, the strains used must be changed periodically as new strains emerge that are no longer controlled by people's residual immunity.

A new kind of vaccine?

• induces antibodies directed against one of the matrix proteins, a molecule that has not been found to undergo either antigenic drift or antigenic shift.
• is given as a nasal spray rather than by injection. Presumably this tilts the antibody response to IgA production, a better defense against infection by inhaled viruses than blood-borne IgG antibodies.

Other weapons against flu

Amantadine and Rimantadine

Zanamivir and Oseltamivir

Antibiotics

Antibiotics are of absolutely no value against the flu virus. However, they are often given to patients to combat the secondary bacterial infections that occur and that are usually the main cause of serious illness and death.

Why so few drugs?

There are far fewer anti-viral drugs than anti-bacterial drugs because so much of the virus life cycle is dependent on the machinery of its host. There are many agents that could kill off the virus, but they would kill off host cell as well. So the goal is to find drugs that target molecular machinery unique to the virus. The more we learn about these molecular details, the better the chance for developing a successful new drug.

The Hong Kong "bird flu"

In May of 1997, a 3-year-old boy in Hong Kong died of influenza. It turned out that the virus that killed him was the same one that had been killing chickens in the region but had not been implicated in any human infections. The virus is H5N1. The H5 molecule is common among bird influenza viruses but has never before been seen on flu viruses that cause human disease.

As a glance at the tables above will show, humans have had long experience with infections and vaccines by both H1 and H3 flu viruses. But the human population has absolutely no immunity against any H5 viruses. Was the boy's disease an isolated incident or had the ground been prepared for another worldwide pandemic?

• Only 17 additional cases (producing five more deaths) were eventually found.
• All eight genes, not just the H5 gene, are characteristic of bird influenza viruses and neither human nor pig flu viruses. So the virus does not seem to be well adapted to infect human cells.
• Detailed analysis of the viruses recovered from seven of the patients revealed two different strains of the H5N1 virus. This suggests that most human cases are acquired directly from sick chickens, not from each other. (Just as several strains of H1N1 and H3N2 are currently causing illness in humans, so different strains of bird flu can be responsible for different outbreaks among chickens.)
• Over half the people showing signs of infection with H5N1 were poultry workers who were not in contact with any human cases of the disease.
• Many people in close contact to patients sick with bird flu (e.g., family members and neighbors) were tested and showed no signs of being infected by the virus. So person-to-person spread seems not to occur or to occur only rarely.

Despite this good news, work is going forward on an H5N1 vaccine just in case!

Searching for the "Spanish flu" virus

An international team, led by Canadian scientist Dr. Kirsty Duncan, travelled to Longyearbyen, Norway in 1998 to collect tissue samples from the bodies of six miners who died there in 1918, presumably of the "Spanish" flu. It was hoped that the bodies had been buried six feet deep in the permafrost and so would have remained frozen since 1918. However, it turned out that the bodies were in a shallow grave and would have been repeatedly thawed and frozen. Despite that, tissue samples have been removed for analysis in laboratories in Canada, the United States, Great Britain, and Norway. The goal is to attempt to recover fragments of the viral genome in the hope of understanding why that flu virus was so lethal. The samples will be handled under strict conditions of biological containment.

To find out more about this project, the case in Hong Kong, and much more about the flu, read the superlative account by Malcolm Gladwell ("The Dead Zone") in the September 29, 1997 issue of The New Yorker.

As for the sequence analysis by Reid and Taubenberger, their results suggest that the hemagglutinin (H) gene of the 1918 virus came from birds and moved into both humans and pigs.