Replication of the Influenza Virus

Viruses can only replicate in living cells. Influenza infection and replication is a multi-step process: firstly the virus has to bind to and enter the cell, then deliver its genome to a site where it can produce new copies of viral proteins and RNA, assemble these components into new viral particles and finally exit the host cell. Influenza viruses bind through hemagglutinin onto sialic acid sugars on the surfaces of epithelial cells; typically in the nose, throat and lungs of mammals and intestines of birds (Stage 1 in figure 1). After the hemagglutinin is cleaved by a protease, the cell imports the virus by endocytosis. Once inside the cell, the acidic conditions in the endosome cause two events to happen: first part of the hemagglutinin protein fuses the viral envelope with the vacuole's membrane, then the M2 ion channel allows protons to move through the viral envelope and acidify the core of the virus, which causes the core to dissemble and release the viral RNA and core proteins. The viral RNA (vRNA) molecules, accessory proteins and RNA-dependent RNA polymerase are then released into the cytoplasm (Stage 2). The M2 ion channel is blocked by amantadine drugs, preventing infection.

These core proteins and vRNA form a complex that is transported into the cell nucleus, where the RNA-dependent RNA polymerase begins transcribing complementary positive-sense vRNA (Steps 3a and b). The vRNA is either exported into the cytoplasm and translated (step 4), or remains in the nucleus. Newly synthesised viral proteins are either secreted through the Golgi apparatus onto the cell surface (in the case of neuraminidase and hemagglutinin, step 5b) or transported back into the nucleus to bind vRNA and form new viral genome particles (step 5a). Other viral proteins have multiple actions in the host cell, including degrading cellular mRNA and using the released nucleotides for vRNA synthesis and also inhibiting translation of host-cell mRNAs. Negative-sense vRNAs that form the genomes of future viruses, RNA-dependent RNA polymerase, and other viral proteins are assembled into a virion. Hemagglutinin and neuraminidase molecules cluster into a bulge in the cell membrane. The vRNA and viral core proteins leave the nucleus and enter this membrane protrusion (step 6). The mature virus buds off from the cell in a sphere of host phospholipid membrane, acquiring hemagglutinin and neuraminidase with this membrane coat (step 7). As before, the viruses adhere to the cell through hemagglutinin; the mature viruses detach once their neuraminidase has cleaved sialic acid residues from the host cell. Drugs that inhibit neuraminidase, such as oseltamivir, therefore prevent the release of new infectious viruses and halt viral replication. After the release of new influenza viruses, the host cell dies.

Figure 2. (A) If a cell is infected with two different influenza viruses, the RNAs of both viruses are copied in the nucleus. When new virus particles are assembled at the plasma membrane, each of the 8 RNA segments may originate from either infecting virus. The progeny that inherit RNAs from both parents are called reassortants. (B) The underlying concern is that swine are susceptible to a wide variety of influenza viruses, and are thought to make excellent `mixing vessels for influenza stains. Although the odds of it happening are probably very low, the worry is that a pig could be infected by two different influenza strains simultaneous, and a reassortment of the viruses could take place. The result could be a new, or mutated, flu virus. Because of the absence of RNA proofreading enzymes, the RNA-dependent RNA polymerase that copies the viral genome makes an error roughly every 10 thousand nucleotides, which is the approximate length of the influenza vRNA. Hence, the majority of newly manufactured influenza viruses are mutants; this causes "antigenic drift", which is a slow change in the antigens on the viral surface over time. The separation of the genome into eight separate segments of vRNA allows mixing or reassortment of vRNAs if more than one type of influenza virus infects a single cell (see figure 2). The resulting rapid change in viral genetics produces antigenic shifts, which are sudden changes from one antigen to another. These sudden large changes allow the virus to infect new host species and quickly overcome protective immunity. This is important in the emergence of pandemics.

Influenzavirus A
This genus has one species, influenza A virus. Wild aquatic birds are the natural hosts for a large variety of influenza A. Occasionally, viruses are transmitted to other species and may then cause devastating outbreaks in domestic poultry or give rise to human influenza pandemics. The type A viruses are the most virulent human pathogens among the three influenza types and cause the most severe disease. The influenza A virus can be subdivided into different serotypes based on the antibody response to these viruses. The serotypes that have been confirmed in humans, ordered by the number of known human pandemic deaths, are:

H1N1, which caused Spanish Flu in 1918, and Swine Flu in 2009 H1N2, endemic in humans, pigs and birds H2N2, which caused Asian Flu in 1957 H3N2, which caused Hong Kong Flu in 1968 H5N1, which caused Bird Flu in 2004 H7N2 H7N3 H7N7, which has unusual zoonotic potential H10N7
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Influenzavirus B
This genus has one species, influenza B virus. Influenza B almost exclusively infects humans and is less common than influenza A. The only other animals known to be susceptible to influenza B infection are the seal and the ferret. This type of influenza mutates at a rate 23 times slower than type A and consequently is less genetically diverse, with only one influenza B serotype. As a result of this lack of antigenic diversity, a degree of immunity to influenza B is usually acquired at an early age. However, influenza B mutates enough that lasting immunity is not possible. This reduced rate of antigenic change, combined with its limited host range (inhibiting cross speciesantigenic shift), ensures that pandemics of influenza B do not occur.

Influenzavirus B is the only species of the genus influenzavirus B. The wide diversity in shapes and sizes of viruses are called morphology. Its protective protein shell, capsid, is enveloped; while the entire virion consists of an envelope, a matrix protein, a nucleoprotein complex, a nuecleocapsid, and a polymerase complex. Influenzavirus B comes in different shapes, usually spherical or filamentous. As far as humans know or have experienced is that Influenzavirus B can only be

found in seals and humans, or it is only known for causing diseases on these two species. Compared to influenzavirus A, it mutates three times slower, but humans still cannot have a long lasting immunity for it. The seasonal outbreak of flu is usually caused by both influenzavirus A and influenzavirus B. As the two are very similar, they cause the same spectrum of disease, only that influenzavirus B is not as powerful and does not cause a pandemic. So the international outbreak of flu must not be caused by influenzavirus B. The structures of influenzavirus B is very similar to the structure of that in influenzavirus A, it would be barely possible to distinguish a influenzavirus A and influenzavirus B under the electron microscope. But when looked carefully, it can be visualized that influenzavirus A has only three membrane proteins while the influenzavirus B has four.
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Influenzavirus C
This genus has one species, influenza C virus, which infects humans, dogs and pigs, sometimes causing both severe illness and local epidemics. However, influenza C is less common than the other types and usually only causes mild disease in children.

Influenzavirus C is very different from influenzavirus A and influenzavirus B. This virus possesses a receptor-destroying enzymatic activity. For the other two inlfuenzaviruses, they have proteins on their surface which match with the host cells surface, and thus are allowed to enter the host cell. But influenzavirus C has enzymes on its surface which catalyses the breakdown of the receptors on the surface of the host cells, making it more convenient to enter a host cell and replicate. Influenzavirus C can be found in humans and pigs, they rarely cause influenza in these two organisms, but can be deadly and may cause local epidemics.

Causes of the 1918 flu epidemic

The 1918 flu pandemic, also known as the Spanish Flu, remains the single most devastating epidemic in recorded human history, with estimates between 40 and 80 million deaths. Despite the severity of the disease, little is known about the causes, progression and eventual dissipation of the disease. However, some evidence has helped

researchers better understand how the disease originated and why it progressed so quickly. The 1918 Spanish Flu was an Influenza A strain of the H1N1 subtype. Influenza A is most well-known as the avian strain of influenza, which also includes the H5N1 subtype that has received recent publicity as the "avian bird-flu". The H1N1 strain is generally not among the most virulent or fatal of flu strains. In the last few years, variants of H1N1 have been responsible for over half of all flu infections worldwide, but a far smaller proportion of deaths. The most fascinating feature of the flu of 1918 may shed a great deal of insight into its origin and rapid spread. Most influenza strains target the immuno-compromised, the young, and the elderly. In these subpopulations, the immune system cannot effectively fight the influenza virus and, as a result, infection ensues. However, the Spanish flu featured a "cytokine burst" at the area of infection, which drastically changed how this flu interacted with the human host, targeting the healthiest members of the population. Cytokines are special cellular signalling molecules and many are involved with the human body's immune response. In victims of the Spanish flu, the virus actually causes levels of cytokines to spike dramatically, increasing the body's immune response. Leukocytes, or white blood cells, are recruited by the cytokine storm and cause damage to the tissue at the site of infection. For most humans, the infection site would be the lungs as the virus would be caught from air-borne particles. The lung tissues would be destroyed and fluids would fill the lung and impair breathing. The use of the body's own immune system as a weapon changes the infection pattern of the 1918 flu. Humans with the strongest immune systems, healthy 19-40 year olds, were most susceptible to infection and the resulting tissue damage. The young, elderly and immuno-compromised could not generate the powerful cytokine storms that weakened their healthier counterparts. With this in mind, it is easy to see how the disease could spread in the world of 1918. World War I was nearing its close, but soldiers, medics and

statesmen were still traveling abroad with greater frequency than any other time. Most of the armed forces of the countries involved in World War I were young men, fitting the description of that group most susceptible to the influenza. The wartime stress and horrid living conditions for soldiers across Europe further exacerbated the spread of the flu and the resulting casualties. New transportation technology made travel easier for everyone, setting the stage for a global outbreak. Japan and American Somoa were among the few nations that took

extreme measures to quarantine infected individuals and close ports to travelers from other countries. As a result, Somoa didn't report a single death and Japan had a much lower mortality rate than any other Eastern Asian nation. As our world continues to grow into a global economy, it is important that we are aware of the dangers of global outbreaks of disease. There is a great deal to learn from the 1918 flu epidemic that could prove useful in protecting humanity from future devastation. From understanding the mechanism of pandemic flu infections, including the cytokine storms and infection of the healthy, to a better model of disease travel in a connected world, the origins of the flu of 1918 provide greater insight into epidemic creation and spread.

Involve changes in the influenza polymerase A polymerase of an influenza virus: composed of viral proteins PB1, PB2, and PA, assembles with viral RNA and nucleoprotein (NP) to mediate transcription and replication of the viral genome. General acknowledgements on adaptive mutation Viruses isolated from birds generally contain polymerases with the avian-signature glutamic acid at amino acid 627 in the PB2 subunit. These polymerases have restricted activity in human cells. An adaptive change in this residue from glutamic acid to the human-signature lysine confers high levels of polymerase activity in human cells.

glutamic acid-to-lysine mutation facilitates escape from an inhibitory factor that restricts the function of avian-derived polymerases in human cells .The identity of the putative inhibitor and the molecular basis for the activity associated with changes at amino acid 627 have not yet been established genetic reassortment allows new viruses to evolve under both natural conditions and in artificial cultures is the mixing of the genetic material of one species into new combinations in different individuals: e.g. When two different Influenza A viruses co-infect the same host cell, new virions are released that contain segments from both parental strains. can only occur between influenza viruses of the same type. We dont know why influenza B and C cannot exchange RNA segments---- the reason is probably linked to the packaging mechanisim that ensures that each influenza

virion contains at least one copy of each RNA segment Process of genetic reassorment When an influenza virus infects a cell, the individual RNA segments enter the nucleus. There they are copied many times to form RNA genomes for new infectious virions. The new RNA segments are exported to the cytoplasm, and then are incorporated into new virus particles which bud from the cell. If a cell is infected with two different influenza viruses, the RNAs of both viruses are copied in the nucleus. When new virus particles are assembled at the plasma membrane, each of the 8 RNA segments may originate from either infecting virus. (The progeny that inherit RNAs from both parents are called reassortants. ) a cell that is co-infected with two influenza viruses L and M. The infected cell produces both parental viruses as well as a reassortant R3 which inherits one RNA segment from strain L and the remainder from strain M.

Antigenic shift occurs only in influenza virus A infects more than just humans the process by which at least two different strains of a influenza virus (or different viruses), combine to form a new subtype having a mixture of the surface antigens of the two original strains. Antigenic shift is a specific case of reassortment or viral shift that confers a phenotypic change.

Antigenic drift How influenza viruses evade infection-fighting antibodies by constantly changing the shape of their major surface protein, antigen Dr. Yewdell(Scott Hensley, Ph.D., Jonathan W. Yewdell, M.D., Ph.D., ). ---------According to the prevailing theory, drift occurs as the virus is passed from person to person and is exposed to differing antibody attacks at each stop. With varying success, antibodies recognize one or more of the four antigenic regions in hemagglutinin, the major outer coat protein of the flu virus. Antibodies in person A,

for example, may mount an attack in which antibodies focus on a single antigenic region. Mutant viruses that arise in person A can escape antibodies by replacing one critical amino acid in this antigen region. These mutant viruses survive, multiply and are passed to person B, where the process is repeated. natural mutation over time of known strains of influenza (or other things, in a more general sense) which may lead to a loss of immunity, or in vaccine mismatch

(Genetic drift)

change in the frequency of a gene variant (allele) in a population due to random sampling(related to statistics). The alleles in the offspring are a sample of those in the parents, and chance has a role in determining whether a given individual survives and reproduces. A population's allele frequency is the fraction of the copies of one gene that share a particular form.

main pt of antigenic drift and antigenic shift new forms antigen is different from the old antigen, antibodies can no longer bind to the receptors and viruses with these new antigens can evade immunity to the original strain of the virus. When such a changes occurs, people who have had the illness in the past will lose their immunity to the new strain and vaccines against the original virus will also become less effective new forms of antigen binding site on antibodies cannot bind with the new forms of antigen of the same virus anymore, since the body may not encounter this new form of antigen before and then blahblahblah this is the mechanisim for antigenic dirft and shift