- Email: [email protected]
Avian influenza: the next pandemic?
Clinical Microbiology Newsletter Vol. 28, No. 13
July 1, 2006
Avian Influenza: the Next Pandemic? Robert C. Moellering, Jr., M.D., Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachussetts
Abstract In 1997, a novel strain of influenza A (H5N1) was discovered in poultry in South Asia and China. Since that time, this avian influenza virus has undergone a number of mutational changes and has spread widely in poultry and birds. Its adaptation to migratory birds in 2005 provided the opportunity for even more widespread dissemination, which now includes 50 countries in Asia, Europe, and Africa. Although the virus has yet to obtain the necessary genes for efficient human-human transmission, it has already caused more than 100 human fatalities throughout the world. As the virus continues to propagate widely in birds and poultry, the likelihood of its acquiring the genetic mechanisms for human-human transmission increases. If this does occur, the ensuing worldwide pandemic could be catastrophic. Since 1997, an unprecedented epizootic avian influenza A (H5N1) virus that appears to be highly pathogenic has begun to cross the species barrier in Asia and elsewhere in the world and, as of April 2006, has caused over 100 human fatalities (1,2). In the ensuing 9 years, the virus has undergone a number of mutational changes that appear to have increased its pathogenicity and enlarged its host range, which now includes migratory birds that have begun to spread it widely along the major flyways of the world. The initial outbreaks were largely confined to China and South Asia, but as of April 2006, the virus had spread to birds (and some humans) in 50 countries in Asia, Europe, the Indian subcontinent, and Africa (2). Thus far, efficient humanto-human transmission of this virus has not occurred. However, as the virus continues to be spread widely among birds around the world, the likelihood
Mailing Address: Robert C. Moellering, Jr., M.D., Department of Medicine, Beth Israel Deaconess Medical Center, 110 Francis Street, Suite 6A, Boston, MA 02215. Tel.: 617-632-7437. Fax: 617-732-7439. Email: [email protected] Clinical Microbiology Newsletter 28:13,2006
increases that it will obtain the necessary mutations or acquire new genes via reassortment, which will enable efficient human-human spread and thus pose the threat of a worldwide pandemic.
Biology The virus of concern here, influenza virus, is an RNA virus of the family Orthomyxoviridae (3). There are three distinct types (A, B, and C) of influenza viruses, but only types A and B cause serious human disease. Influenza C generally causes mild, uncomplicated upper respiratory tract infections and is not responsible for pandemic spread. All of the influenza viruses have a host cell-derived envelope, envelope glycoproteins (hemagglutinins and neuraminidases) of critical importance for entry and egress from host cells, and a segmented RNA genome. Moreover, since they are RNA viruses, they have no proofreading mechanism and therefore are subject to a high mutation rate during replication. Influenza B viruses generally cause severe disease among older adults or persons at high risk due to immunocompromised status. Although they can cause epidemics, they do not cause worldwide pandemics. The © 2006 Elsevier
human is the only known host for influenza B viruses. Influenza A viruses, on the other hand, have a much broader host range. Their primary hosts are birds (especially ducks, chickens, and turkeys), but they can also easily infect swine and, with appropriate genetic changes, infect humans and other mammals. Because of their ability to undergo antigenic shift (due to the acquisition of completely new hemagglutinins and or neuraminidases), influenza A viruses have the ability to cause worldwide pandemics with significant mortality, even in young persons. Influenza viruses are designated by type (A, B, or C), site and date of first isolation, and the natures of the hemagglutinin (of which there are 15 distinct types in A viruses) and neuraminidase (of which there are 9 types in A viruses). Thus, influenza A/USSR/77 H1N1 is
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an influenza virus containing type 1 hemagglutinins and neuraminidases, which was first isolated in the former USSR in 1977.
Viral Entry and Replication On encountering an appropriate host, the hemagglutinin of the influenza virus binds to specific sialic acid-galactose receptors on the surfaces of respiratory epithelial cells. The virus then forms an indentation in the host cell membrane, which goes on to produce a vesicle, allowing the virus to enter the cell. As the pH in the vesicle decreases, there is a reconfiguration of the hemagglutinin, which dissolves the vesicle, allowing the genes of the virus to spill out and insert themselves into the epithelial cell nucleus. These genes basically take over the cell, producing new viral proteins, which are packaged with new copies of the viral genes. The neuraminidase destroys the sialic acid receptors on the epithelial cell surface, allowing the virus to escape and, at the same time, destroying the epithelial cell.
Clinical Disease Following a short incubation period of 1 to 2 days, patients infected with influenza develop sudden onset of fever, chills, headaches, myalgia, malaise, and anorexia (3). The clinical syndrome also includes dry cough, severe pharyngeal pain, substernal burning, and mild coryza without nasal obstruction. The fever generally lasts 4 to 8 days (mean, 3 days) and subsides by lysis rather than crisis. By the first day of infection, patients excrete large titers of viruses in their respiratory secretions and thus are highly infectious, contributing to the explosive spread of influenza among communities lacking primary immunity to the virus. Most patients infected with epidemic strains of influenza A or B
98
0196-4399/00 (see frontmatter)
virus do not develop primary viral pneumonia, but when this occurs, it is a serious and life-threatening complication. Secondary bacterial pneumonias may occur in the second week of the illness, and patients may develop myositis, myocarditis/pericarditis, peripheral neuropathies, and even encephalitis, but the last few complications are quite rare. Natural infection results in the production of antibodies to hemagglutinins and neuraminidases, as well as certain other structural proteins of the viruses (3). The antibodies are strain specific, but antibodies against heterologous strains may modify subsequent disease. Local mucosal IgA (and IgG) probably plays an important role in resistance to infection as do CD4+ cells and cytotoxic T cells, although the exact role of cellular immunity in influenza remains to be fully elucidated. Relatively minor antigenic changes occur frequently (every year or every few years) within the hemagglutinins and/or neuraminidases of the viruses circulating in the community. These changes provide relative resistance to antibodies directed against the earlier hemagglutinin or neuraminidase and are responsible for epidemic disease. The process by which this occurs is known as “antigenic drift.” “Antigenic shift,” in which new viruses arise via mutation or genetic reassortment and in which there is little or no relationship between the new hemagglutinin (or neuraminidase) antigens of the “old” and “new” viruses, is responsible for pandemic disease. Current vaccines are trivalent, containing one example each of influenza A (H1N1) virus, A (H3N2) virus, and influenza B virus thought to be most likely to cause disease in the upcoming season, based on antigenic analysis of circulating strains (usually in the south-
© 2006 Elsevier
ern hemisphere). Vaccines are subunit or split virus preparations made from inactivated virus grown in embryonated chicken eggs. A live-attenuated (coldadapted) influenza vaccine is also available and has been approved for individuals in the age group 5 to 49. This vaccine is administered intranasally.
Therapy Two types of antiviral agents are available. Amantadine and rimantadine inhibit viral uncoating and have potential activity against influenza A but not influenza B viruses. The influenza viruses currently circulating in the United States are resistant to amantadine and rimantadine. Zanamivir and oseltamivir are neuraminidase inhibitors, which prevent egress of the virus from infected cells. They are effective against both influenza A and B viruses (4). Both sets of antiviral agents are effective for prophylaxis of influenza. In order to be useful therapeutically, they must be given early in the course of infection. They are much less effective when given more than 24 to 48 hours after the onset of symptoms.
Animal Hosts As previously noted, the major hosts for influenza A viruses are birds. Avian strains are generally restricted in their ability to replicate in humans, primarily because their hemagglutinins bind preferentially to receptors in the epithelial cells of birds, which contain sialic acidgalactose of an α2→3 type, while human hemagglutinins “prefer” mammalian receptors with α2→6 linkages. Interestingly, swine contain receptors for both human and avian viruses, and therefore, the potential for co-infections with these viruses in swine is very real. There are two mechanisms by which viruses can
Clinical Microbiology Newsletter 28:13,2006
circumvent their barriers to interspecies transmission. The first is reassortment between avian viruses which provide novel surface glycoproteins, and human viruses, which provide genes allowing efficient replication in humans. This can potentially occur in swine, for instance. The H2N2 virus (Asian flu of 1957) and the H3N2 virus (Hong Kong flu of 1968) probably arose in this way. The second is adaptation of avian viruses to the human host by mutation and evolution in swine (the 1918 pandemic strain H1N1 most certainly was produced by this mechanism). In addition, avian viruses can sometimes be directly introduced into human populations (as is currently happening with the H5N1 avian flu strain circulating in South Asia). Because the present strains lack hemagglutinins with high affinity for receptors on human epithelial cells, efficient human-human spread has not yet occurred. In addition to the concern about the present influenza H5N1 virus, other recent avian influenza outbreaks in other parts of the world have also been associated with evidence of spread to humans. Thus, in 2003, an avian influenza virus (H7N7) outbreak in poultry was associated with 87 human cases and one fatality in The Netherlands. Between 1999 and 2003, avian influenza viruses H7N1 and H7N3 caused several outbreaks in poultry in northern Italy. Although there was no evidence of human seroconversion to H7N1, 3.8% of persons exposed to H7N3 (a “low-pathogenicity” strain) exhibited unequivocal evidence of seroconversion (without clinical disease), providing solid evidence of avian-human transmission (5,6). Studies of the 1997 influenza A H5N1 virus from patients in South Asia revealed that the virus contains a number of properties that may enhance its pathogenicity, including a highly cleavable hemagglutinin that is activated by multiple cellular proteases, specific substitutions in a polymerase basic protein 2 that enhance replication, and a specific substitution in protein 1 that confers increased resistance to inhibition by interferons and tumor necrosis factor alpha (TNF-α) and produces greater elaboration of cytokines (especially TNF-α) in human macrophages (1). Since 1997, studies reveal continued evolution of H5N1 that include changes Clinical Microbiology Newsletter 28:13,2006
Table 1. Countries with Bird/Poultry cases of H5/N1 avian influenza as of 7 April 2006a Afghanistan
Hong Kong
Pakistan
Albania
Hungary
Poland
Austria
Italy
Romania
Azerbaijan
India
Russia
Bosnia and Herzegovina
Indonesia
Serbia and Montenegro
Bulgaria
Iraq
Slovak Republic
Cambodia
Iran
Slovenia
Cameroon
Israel
South Korea
China
Japan
Sweden
Croatia
Jordan
Switzerland
Czech Republic
Kazakhstan
Thailand
Denmark
Laos
Turkey
Egypt
Malaysia
Ukraine
France
Mongolia
United Kingdom
Georgia
Myanmar (Burma)
Vietnam
Germany
Niger
Greece
Nigeria
a
Source: http://www.cdc.gov/flu/avian/outbreaks/current.htm.
Table 2. Human cases of H5/N1 avian influenza infection: 1 December 2003 – 7 April 2006a Country Azerbaijan
No. of cases
No. of deaths
7
5
Cambodia
5
5
China
16
11
Egypt
4
2
Indonesia
30
23
Iraq
2
2
Thailand
22
14
Turkey
12
4
Vietnam
93
42
Total
191
108
a
Source: Morbid. Mortal. Wkly. Rep. 55:370, 2006.
in antigenicity and the internal gene constellation, extended host range in avian species, an ability to infect felids, enhanced pathogenicity in mice and ferrets, and increased environmental stability (1,7,8). Indeed, it is the ability of the virus to extend its host range to migratory birds that is responsible for the current rapid spread of the virus to birds in Central Asia, Europe, the Indian subcontinent, and Africa. Table 1 lists those countries in which the avian influenza A (H5N1) virus has been definitively shown to have caused disease in the bird or poultry populations. Given the © 2006 Elsevier
remarkable range of migratory birds now carrying this virus along their flyways, the number of countries containing infected birds continues to expand. As of April 2006, there had been no reports of isolation of the virus from birds in North or South America or Australia. To date, the majority of human cases of H5N1 avian influenza infection have occurred in South Asia in patients with close contact with infected birds. Of the 191 human cases reported as of 7 April 2006, more than half (108) have resulted in death (Table 2) (2). In many of these cases, it appears that 0196-4399/00 (see frontmatter)
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primary viral pneumonia has been the cause of death. Indeed, more than 88% of the infected patients have had pulmonary infiltrates at the time of diagnosis (1). The current H5N1 virus is, for the most part, resistant to amantadine and rimantadine but is susceptible to the neuraminidase inhibitors oseltamivir and zanamivir (1). It appears that it may require higher doses of oseltamivir, however, for clinical effectiveness, and high-level resistance to oseltamivir resulting from a single-step substitution in N1 neuraminidase (His274Tyr) has emerged in several H5N1 patients treated with oseltamivir (9). These organisms, interestingly, are not cross-resistant to zanamivir (9).
Laboratory Diagnosis It is not known how sensitive the current influenza A enzyme immunoassay or direct fluorescent antibody tests will be to detect H5N1. Commercial rapid antigen tests appeared less effective in detecting H5N1 (only 36% of PCR-confirmed cases were positive by rapid antigen testing in Thailand in 1997) than for human influenza (1). A new, much more sensitive reverse transcription RT-PCR for H5N1 has recently been developed by the CDC and had been sent to 70 laboratories across the U.S. as of February 2006.
Vaccines As the H5N1 virus continues its relentless spread through the bird populations of the world, there are increasing concerns that the virus will acquire the necessary mutations (or new genes via reassortment) to enable its hemagglutinin to bind effectively to human epithelial receptors. Should this occur, it would set the stage for a massive pandemic, since there is currently little herd immunity to H5N1 viruses in the human population of the world. Vaccines have been made against the current H5N1 virus and have been employed with questionable success among poultry populations in South Asia (10,11). Inactivated subvirion influenza A (H5N1) vaccine has also recently been developed and administered to human volunteers. Investigators administered two intramuscular doses of H5N1 vaccine to 451 healthy adults and found the vaccine to be well tolerated. Potentially protective antibody titers (≥1:40) were seen in 54 to 58% of those who received 100
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the highest dose (90 µg) of the vaccine, and the authors concluded that “a conventional subvirion H5 influenza vaccine may be effective in preventing Influenza A (H5N1) disease in humans.” However, it required two doses of the vaccine to produce possibly protective titers, which were seen in only approximately half of the subjects vaccinated. Moreover, a vaccine such as this will not prevent human disease, since the currently circulating strains in birds are not those which would cause a human pandemic. It will be necessary to first identify the organism which has acquired receptors for hemagglutins that bind to human receptors before an effective vaccine can be made. Current estimates are that it will take at least 6 months to gear up to produce such a vaccine should the virus adapt itself to human receptors. Because a significant proportion of the population has antibodies to the type 1 neuraminidase found in the current avian flu strains (due to prior infection with circulating viruses or immunization with vaccines containing the N1 antigen), it is possible that there may be low levels of cross-protection against avian flu viruses conferred by these antibodies (12).
Pandemic Risks Concerns over the possibility of a worldwide pandemic are related primarily to knowledge of the 1918 influenza pandemic. This was caused by an H1N1 virus that very likely arose via mutation (or genetic reassortment) in swine and then spread from the U.S. (where it apparently originated) around the world in three distinct waves in 1918 and 1919. It is estimated that up to 50 to 100 million deaths occurred worldwide secondary to this particular virus. It was initially spread to Europe by American troops mobilized for World War I and from there spread rapidly to other parts of the world. The mortality rate from infections with this virus approached 70 to 90% in certain populations, such as those in Oceania and Alaska. Many of the patients appeared to die from overwhelming viral pneumonia with other serious complications. Recently, the 1918 H1N1 virus has been recreated in the laboratory. Specifically genomic RNA of the 1918 virus recovered from archived formalin-fixed lung autopsy © 2006 Elsevier
material and from unfixed lung tissues of a 1918 influenza victim buried in the Alaska permafrost have been used via reverse genetics to generate an influenza virus bearing all eight gene segments of the original pandemic virus (13). In stark contrast to contemporary H1N1 viruses, the 1918 pandemic virus has the ability to replicate in the absence of trypsin, causes death in mice and embryonated chicken eggs, and displays a high-growth phenotype in human bronchial epithelial cells. Data accumulated thus far suggest that coordinated expression of the 1918 virus genes confers a unique high-virulence phenotype that was clearly responsible for the remarkable mortality rate in the 1918 pandemic (13). Given the remarkable virulence of this organism, concerns over the potential human adaptation of the present H5N1 avian influenza virus appear more than justified. It is clear that a major pandemic of influenza, which could infect as many as 300,000,000 people worldwide, would overwhelm the health care facilities of every country in the world. At present in the U.S., there are few excess hospital beds. At any given time, it is estimated that 75% of all of the respiratory assist devices in the U.S. are being utilized. Thus, an influenza pandemic would create an influx of patients that would overwhelm current health care facilities. Moreover, most hospitals in the U.S. exercise “just in time” inventories which means that they do not have long term supplies on hand and are dependent upon rapid delivery of needed supplies. In the event of an influenza pandemic, there is no assurance that the personnel necessary for such a system to work would even be available. Finally, there is currently not nearly enough supply of antiviral agents, such as oseltamivir or zanamivir, to meet the needs should a pandemic occur. Thus, it will take a major effort on the part of all health care workers and ancillary personnel as well as government relief agencies, to deal with the overwhelming problems such a pandemic would present (14). References 1. WHO. 2005. The Writing Committee of the World Health Organization Consultation on Human Influenza A/H5. Avian
Clinical Microbiology Newsletter 28:13,2006
influenza A (H5N1) infection in humans. N. Engl. J. Med. 353:1374-1385. 2. WHO. 2006. Update: Influenza activity — United States, March 19-25, 2006. Morb. Mortal. Wkly. Rep. 55:368-370. 3. Treanor, J.J. 2005. Influenza virus, p. 2060-2085. In G.L. Mandell, J.E. Bennett, and R. Dolin (ed.), Principles and practice of infectious diseases, 6th ed. Elsevier, Philadelphia. 4. Moscona, A. 2005. Neuraminidase inhibitors for influenza. N. Engl. J. Med. 353:1363-1373. 5. Puzelli S. et al. 2005. Serological analysis of serum samples from humans exposed to avian H7 influenza viruses in Italy between 1999 and 2003. J. Infect. Dis. 192:1318-1322.
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6. Hayden, F., and A. Croisier. 2005. Transmission of avian influenza viruses to and between humans. J. Infect. Dis. 192:1311-1314. 7. Tiensin, T. et al. 2005. Highly pathogenic avian influenza H5N1, Thailand, 2004. Emerg. Infect. Dis. 11:1664-1672. 8. World Health Organization Global Influenza Program Surveillance Network. 2005. Evolution of H5N1 avian influenza viruses in Asia. Emerg. Infect. Dis. 11:1515-1521. 9. de Jong, M.D. et al. 2005. Oseltamivir resistance during treatment of influenza A (H5N1) infection. N. Engl. J. Med. 353:2667-2672. 10. Schwartz, B. and B. Gellin. 2005. Vaccination strategies for an influenza pandemic. J. Infect. Dis. 191:1207-1209.
© 2006 Elsevier
11. Lipatov, A.S. et al. 2005. Efficacy of H5 influenza vaccines produced by reverse genetics in a lethal mouse model. J. Infect. Dis. 191:1216-1220. 12. Epstein, S.L. 2006. Prior H1N1 influenza infection and susceptibility of Cleveland family study participants during the H2N2 pandemic of 1957: an experiment of nature. J. Infect. Dis. 193:49-53. 13. Tumpey, T.M. et al. 2005. Characterization of the reconstructed 1918 Spanish influenza pandemic virus. Science 310:77-80. 14. Lipsitch, M. and B.R. Bloom. 2006. Avian flu preparing for a pandemic. Harvard Pub. Health Rev. Winter 2006:4-7.
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July 1, 2006
Avian Influenza: the Next Pandemic? Robert C. Moellering, Jr., M.D., Department of Medicine, Beth Israel Deaconess Medical Center, Boston, Massachussetts
Abstract In 1997, a novel strain of influenza A (H5N1) was discovered in poultry in South Asia and China. Since that time, this avian influenza virus has undergone a number of mutational changes and has spread widely in poultry and birds. Its adaptation to migratory birds in 2005 provided the opportunity for even more widespread dissemination, which now includes 50 countries in Asia, Europe, and Africa. Although the virus has yet to obtain the necessary genes for efficient human-human transmission, it has already caused more than 100 human fatalities throughout the world. As the virus continues to propagate widely in birds and poultry, the likelihood of its acquiring the genetic mechanisms for human-human transmission increases. If this does occur, the ensuing worldwide pandemic could be catastrophic. Since 1997, an unprecedented epizootic avian influenza A (H5N1) virus that appears to be highly pathogenic has begun to cross the species barrier in Asia and elsewhere in the world and, as of April 2006, has caused over 100 human fatalities (1,2). In the ensuing 9 years, the virus has undergone a number of mutational changes that appear to have increased its pathogenicity and enlarged its host range, which now includes migratory birds that have begun to spread it widely along the major flyways of the world. The initial outbreaks were largely confined to China and South Asia, but as of April 2006, the virus had spread to birds (and some humans) in 50 countries in Asia, Europe, the Indian subcontinent, and Africa (2). Thus far, efficient humanto-human transmission of this virus has not occurred. However, as the virus continues to be spread widely among birds around the world, the likelihood
Mailing Address: Robert C. Moellering, Jr., M.D., Department of Medicine, Beth Israel Deaconess Medical Center, 110 Francis Street, Suite 6A, Boston, MA 02215. Tel.: 617-632-7437. Fax: 617-732-7439. Email: [email protected] Clinical Microbiology Newsletter 28:13,2006
increases that it will obtain the necessary mutations or acquire new genes via reassortment, which will enable efficient human-human spread and thus pose the threat of a worldwide pandemic.
Biology The virus of concern here, influenza virus, is an RNA virus of the family Orthomyxoviridae (3). There are three distinct types (A, B, and C) of influenza viruses, but only types A and B cause serious human disease. Influenza C generally causes mild, uncomplicated upper respiratory tract infections and is not responsible for pandemic spread. All of the influenza viruses have a host cell-derived envelope, envelope glycoproteins (hemagglutinins and neuraminidases) of critical importance for entry and egress from host cells, and a segmented RNA genome. Moreover, since they are RNA viruses, they have no proofreading mechanism and therefore are subject to a high mutation rate during replication. Influenza B viruses generally cause severe disease among older adults or persons at high risk due to immunocompromised status. Although they can cause epidemics, they do not cause worldwide pandemics. The © 2006 Elsevier
human is the only known host for influenza B viruses. Influenza A viruses, on the other hand, have a much broader host range. Their primary hosts are birds (especially ducks, chickens, and turkeys), but they can also easily infect swine and, with appropriate genetic changes, infect humans and other mammals. Because of their ability to undergo antigenic shift (due to the acquisition of completely new hemagglutinins and or neuraminidases), influenza A viruses have the ability to cause worldwide pandemics with significant mortality, even in young persons. Influenza viruses are designated by type (A, B, or C), site and date of first isolation, and the natures of the hemagglutinin (of which there are 15 distinct types in A viruses) and neuraminidase (of which there are 9 types in A viruses). Thus, influenza A/USSR/77 H1N1 is
0196-4399/00 (see frontmatter)
97
an influenza virus containing type 1 hemagglutinins and neuraminidases, which was first isolated in the former USSR in 1977.
Viral Entry and Replication On encountering an appropriate host, the hemagglutinin of the influenza virus binds to specific sialic acid-galactose receptors on the surfaces of respiratory epithelial cells. The virus then forms an indentation in the host cell membrane, which goes on to produce a vesicle, allowing the virus to enter the cell. As the pH in the vesicle decreases, there is a reconfiguration of the hemagglutinin, which dissolves the vesicle, allowing the genes of the virus to spill out and insert themselves into the epithelial cell nucleus. These genes basically take over the cell, producing new viral proteins, which are packaged with new copies of the viral genes. The neuraminidase destroys the sialic acid receptors on the epithelial cell surface, allowing the virus to escape and, at the same time, destroying the epithelial cell.
Clinical Disease Following a short incubation period of 1 to 2 days, patients infected with influenza develop sudden onset of fever, chills, headaches, myalgia, malaise, and anorexia (3). The clinical syndrome also includes dry cough, severe pharyngeal pain, substernal burning, and mild coryza without nasal obstruction. The fever generally lasts 4 to 8 days (mean, 3 days) and subsides by lysis rather than crisis. By the first day of infection, patients excrete large titers of viruses in their respiratory secretions and thus are highly infectious, contributing to the explosive spread of influenza among communities lacking primary immunity to the virus. Most patients infected with epidemic strains of influenza A or B
98
0196-4399/00 (see frontmatter)
virus do not develop primary viral pneumonia, but when this occurs, it is a serious and life-threatening complication. Secondary bacterial pneumonias may occur in the second week of the illness, and patients may develop myositis, myocarditis/pericarditis, peripheral neuropathies, and even encephalitis, but the last few complications are quite rare. Natural infection results in the production of antibodies to hemagglutinins and neuraminidases, as well as certain other structural proteins of the viruses (3). The antibodies are strain specific, but antibodies against heterologous strains may modify subsequent disease. Local mucosal IgA (and IgG) probably plays an important role in resistance to infection as do CD4+ cells and cytotoxic T cells, although the exact role of cellular immunity in influenza remains to be fully elucidated. Relatively minor antigenic changes occur frequently (every year or every few years) within the hemagglutinins and/or neuraminidases of the viruses circulating in the community. These changes provide relative resistance to antibodies directed against the earlier hemagglutinin or neuraminidase and are responsible for epidemic disease. The process by which this occurs is known as “antigenic drift.” “Antigenic shift,” in which new viruses arise via mutation or genetic reassortment and in which there is little or no relationship between the new hemagglutinin (or neuraminidase) antigens of the “old” and “new” viruses, is responsible for pandemic disease. Current vaccines are trivalent, containing one example each of influenza A (H1N1) virus, A (H3N2) virus, and influenza B virus thought to be most likely to cause disease in the upcoming season, based on antigenic analysis of circulating strains (usually in the south-
© 2006 Elsevier
ern hemisphere). Vaccines are subunit or split virus preparations made from inactivated virus grown in embryonated chicken eggs. A live-attenuated (coldadapted) influenza vaccine is also available and has been approved for individuals in the age group 5 to 49. This vaccine is administered intranasally.
Therapy Two types of antiviral agents are available. Amantadine and rimantadine inhibit viral uncoating and have potential activity against influenza A but not influenza B viruses. The influenza viruses currently circulating in the United States are resistant to amantadine and rimantadine. Zanamivir and oseltamivir are neuraminidase inhibitors, which prevent egress of the virus from infected cells. They are effective against both influenza A and B viruses (4). Both sets of antiviral agents are effective for prophylaxis of influenza. In order to be useful therapeutically, they must be given early in the course of infection. They are much less effective when given more than 24 to 48 hours after the onset of symptoms.
Animal Hosts As previously noted, the major hosts for influenza A viruses are birds. Avian strains are generally restricted in their ability to replicate in humans, primarily because their hemagglutinins bind preferentially to receptors in the epithelial cells of birds, which contain sialic acidgalactose of an α2→3 type, while human hemagglutinins “prefer” mammalian receptors with α2→6 linkages. Interestingly, swine contain receptors for both human and avian viruses, and therefore, the potential for co-infections with these viruses in swine is very real. There are two mechanisms by which viruses can
Clinical Microbiology Newsletter 28:13,2006
circumvent their barriers to interspecies transmission. The first is reassortment between avian viruses which provide novel surface glycoproteins, and human viruses, which provide genes allowing efficient replication in humans. This can potentially occur in swine, for instance. The H2N2 virus (Asian flu of 1957) and the H3N2 virus (Hong Kong flu of 1968) probably arose in this way. The second is adaptation of avian viruses to the human host by mutation and evolution in swine (the 1918 pandemic strain H1N1 most certainly was produced by this mechanism). In addition, avian viruses can sometimes be directly introduced into human populations (as is currently happening with the H5N1 avian flu strain circulating in South Asia). Because the present strains lack hemagglutinins with high affinity for receptors on human epithelial cells, efficient human-human spread has not yet occurred. In addition to the concern about the present influenza H5N1 virus, other recent avian influenza outbreaks in other parts of the world have also been associated with evidence of spread to humans. Thus, in 2003, an avian influenza virus (H7N7) outbreak in poultry was associated with 87 human cases and one fatality in The Netherlands. Between 1999 and 2003, avian influenza viruses H7N1 and H7N3 caused several outbreaks in poultry in northern Italy. Although there was no evidence of human seroconversion to H7N1, 3.8% of persons exposed to H7N3 (a “low-pathogenicity” strain) exhibited unequivocal evidence of seroconversion (without clinical disease), providing solid evidence of avian-human transmission (5,6). Studies of the 1997 influenza A H5N1 virus from patients in South Asia revealed that the virus contains a number of properties that may enhance its pathogenicity, including a highly cleavable hemagglutinin that is activated by multiple cellular proteases, specific substitutions in a polymerase basic protein 2 that enhance replication, and a specific substitution in protein 1 that confers increased resistance to inhibition by interferons and tumor necrosis factor alpha (TNF-α) and produces greater elaboration of cytokines (especially TNF-α) in human macrophages (1). Since 1997, studies reveal continued evolution of H5N1 that include changes Clinical Microbiology Newsletter 28:13,2006
Table 1. Countries with Bird/Poultry cases of H5/N1 avian influenza as of 7 April 2006a Afghanistan
Hong Kong
Pakistan
Albania
Hungary
Poland
Austria
Italy
Romania
Azerbaijan
India
Russia
Bosnia and Herzegovina
Indonesia
Serbia and Montenegro
Bulgaria
Iraq
Slovak Republic
Cambodia
Iran
Slovenia
Cameroon
Israel
South Korea
China
Japan
Sweden
Croatia
Jordan
Switzerland
Czech Republic
Kazakhstan
Thailand
Denmark
Laos
Turkey
Egypt
Malaysia
Ukraine
France
Mongolia
United Kingdom
Georgia
Myanmar (Burma)
Vietnam
Germany
Niger
Greece
Nigeria
a
Source: http://www.cdc.gov/flu/avian/outbreaks/current.htm.
Table 2. Human cases of H5/N1 avian influenza infection: 1 December 2003 – 7 April 2006a Country Azerbaijan
No. of cases
No. of deaths
7
5
Cambodia
5
5
China
16
11
Egypt
4
2
Indonesia
30
23
Iraq
2
2
Thailand
22
14
Turkey
12
4
Vietnam
93
42
Total
191
108
a
Source: Morbid. Mortal. Wkly. Rep. 55:370, 2006.
in antigenicity and the internal gene constellation, extended host range in avian species, an ability to infect felids, enhanced pathogenicity in mice and ferrets, and increased environmental stability (1,7,8). Indeed, it is the ability of the virus to extend its host range to migratory birds that is responsible for the current rapid spread of the virus to birds in Central Asia, Europe, the Indian subcontinent, and Africa. Table 1 lists those countries in which the avian influenza A (H5N1) virus has been definitively shown to have caused disease in the bird or poultry populations. Given the © 2006 Elsevier
remarkable range of migratory birds now carrying this virus along their flyways, the number of countries containing infected birds continues to expand. As of April 2006, there had been no reports of isolation of the virus from birds in North or South America or Australia. To date, the majority of human cases of H5N1 avian influenza infection have occurred in South Asia in patients with close contact with infected birds. Of the 191 human cases reported as of 7 April 2006, more than half (108) have resulted in death (Table 2) (2). In many of these cases, it appears that 0196-4399/00 (see frontmatter)
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primary viral pneumonia has been the cause of death. Indeed, more than 88% of the infected patients have had pulmonary infiltrates at the time of diagnosis (1). The current H5N1 virus is, for the most part, resistant to amantadine and rimantadine but is susceptible to the neuraminidase inhibitors oseltamivir and zanamivir (1). It appears that it may require higher doses of oseltamivir, however, for clinical effectiveness, and high-level resistance to oseltamivir resulting from a single-step substitution in N1 neuraminidase (His274Tyr) has emerged in several H5N1 patients treated with oseltamivir (9). These organisms, interestingly, are not cross-resistant to zanamivir (9).
Laboratory Diagnosis It is not known how sensitive the current influenza A enzyme immunoassay or direct fluorescent antibody tests will be to detect H5N1. Commercial rapid antigen tests appeared less effective in detecting H5N1 (only 36% of PCR-confirmed cases were positive by rapid antigen testing in Thailand in 1997) than for human influenza (1). A new, much more sensitive reverse transcription RT-PCR for H5N1 has recently been developed by the CDC and had been sent to 70 laboratories across the U.S. as of February 2006.
Vaccines As the H5N1 virus continues its relentless spread through the bird populations of the world, there are increasing concerns that the virus will acquire the necessary mutations (or new genes via reassortment) to enable its hemagglutinin to bind effectively to human epithelial receptors. Should this occur, it would set the stage for a massive pandemic, since there is currently little herd immunity to H5N1 viruses in the human population of the world. Vaccines have been made against the current H5N1 virus and have been employed with questionable success among poultry populations in South Asia (10,11). Inactivated subvirion influenza A (H5N1) vaccine has also recently been developed and administered to human volunteers. Investigators administered two intramuscular doses of H5N1 vaccine to 451 healthy adults and found the vaccine to be well tolerated. Potentially protective antibody titers (≥1:40) were seen in 54 to 58% of those who received 100
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the highest dose (90 µg) of the vaccine, and the authors concluded that “a conventional subvirion H5 influenza vaccine may be effective in preventing Influenza A (H5N1) disease in humans.” However, it required two doses of the vaccine to produce possibly protective titers, which were seen in only approximately half of the subjects vaccinated. Moreover, a vaccine such as this will not prevent human disease, since the currently circulating strains in birds are not those which would cause a human pandemic. It will be necessary to first identify the organism which has acquired receptors for hemagglutins that bind to human receptors before an effective vaccine can be made. Current estimates are that it will take at least 6 months to gear up to produce such a vaccine should the virus adapt itself to human receptors. Because a significant proportion of the population has antibodies to the type 1 neuraminidase found in the current avian flu strains (due to prior infection with circulating viruses or immunization with vaccines containing the N1 antigen), it is possible that there may be low levels of cross-protection against avian flu viruses conferred by these antibodies (12).
Pandemic Risks Concerns over the possibility of a worldwide pandemic are related primarily to knowledge of the 1918 influenza pandemic. This was caused by an H1N1 virus that very likely arose via mutation (or genetic reassortment) in swine and then spread from the U.S. (where it apparently originated) around the world in three distinct waves in 1918 and 1919. It is estimated that up to 50 to 100 million deaths occurred worldwide secondary to this particular virus. It was initially spread to Europe by American troops mobilized for World War I and from there spread rapidly to other parts of the world. The mortality rate from infections with this virus approached 70 to 90% in certain populations, such as those in Oceania and Alaska. Many of the patients appeared to die from overwhelming viral pneumonia with other serious complications. Recently, the 1918 H1N1 virus has been recreated in the laboratory. Specifically genomic RNA of the 1918 virus recovered from archived formalin-fixed lung autopsy © 2006 Elsevier
material and from unfixed lung tissues of a 1918 influenza victim buried in the Alaska permafrost have been used via reverse genetics to generate an influenza virus bearing all eight gene segments of the original pandemic virus (13). In stark contrast to contemporary H1N1 viruses, the 1918 pandemic virus has the ability to replicate in the absence of trypsin, causes death in mice and embryonated chicken eggs, and displays a high-growth phenotype in human bronchial epithelial cells. Data accumulated thus far suggest that coordinated expression of the 1918 virus genes confers a unique high-virulence phenotype that was clearly responsible for the remarkable mortality rate in the 1918 pandemic (13). Given the remarkable virulence of this organism, concerns over the potential human adaptation of the present H5N1 avian influenza virus appear more than justified. It is clear that a major pandemic of influenza, which could infect as many as 300,000,000 people worldwide, would overwhelm the health care facilities of every country in the world. At present in the U.S., there are few excess hospital beds. At any given time, it is estimated that 75% of all of the respiratory assist devices in the U.S. are being utilized. Thus, an influenza pandemic would create an influx of patients that would overwhelm current health care facilities. Moreover, most hospitals in the U.S. exercise “just in time” inventories which means that they do not have long term supplies on hand and are dependent upon rapid delivery of needed supplies. In the event of an influenza pandemic, there is no assurance that the personnel necessary for such a system to work would even be available. Finally, there is currently not nearly enough supply of antiviral agents, such as oseltamivir or zanamivir, to meet the needs should a pandemic occur. Thus, it will take a major effort on the part of all health care workers and ancillary personnel as well as government relief agencies, to deal with the overwhelming problems such a pandemic would present (14). References 1. WHO. 2005. The Writing Committee of the World Health Organization Consultation on Human Influenza A/H5. Avian
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influenza A (H5N1) infection in humans. N. Engl. J. Med. 353:1374-1385. 2. WHO. 2006. Update: Influenza activity — United States, March 19-25, 2006. Morb. Mortal. Wkly. Rep. 55:368-370. 3. Treanor, J.J. 2005. Influenza virus, p. 2060-2085. In G.L. Mandell, J.E. Bennett, and R. Dolin (ed.), Principles and practice of infectious diseases, 6th ed. Elsevier, Philadelphia. 4. Moscona, A. 2005. Neuraminidase inhibitors for influenza. N. Engl. J. Med. 353:1363-1373. 5. Puzelli S. et al. 2005. Serological analysis of serum samples from humans exposed to avian H7 influenza viruses in Italy between 1999 and 2003. J. Infect. Dis. 192:1318-1322.
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6. Hayden, F., and A. Croisier. 2005. Transmission of avian influenza viruses to and between humans. J. Infect. Dis. 192:1311-1314. 7. Tiensin, T. et al. 2005. Highly pathogenic avian influenza H5N1, Thailand, 2004. Emerg. Infect. Dis. 11:1664-1672. 8. World Health Organization Global Influenza Program Surveillance Network. 2005. Evolution of H5N1 avian influenza viruses in Asia. Emerg. Infect. Dis. 11:1515-1521. 9. de Jong, M.D. et al. 2005. Oseltamivir resistance during treatment of influenza A (H5N1) infection. N. Engl. J. Med. 353:2667-2672. 10. Schwartz, B. and B. Gellin. 2005. Vaccination strategies for an influenza pandemic. J. Infect. Dis. 191:1207-1209.
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11. Lipatov, A.S. et al. 2005. Efficacy of H5 influenza vaccines produced by reverse genetics in a lethal mouse model. J. Infect. Dis. 191:1216-1220. 12. Epstein, S.L. 2006. Prior H1N1 influenza infection and susceptibility of Cleveland family study participants during the H2N2 pandemic of 1957: an experiment of nature. J. Infect. Dis. 193:49-53. 13. Tumpey, T.M. et al. 2005. Characterization of the reconstructed 1918 Spanish influenza pandemic virus. Science 310:77-80. 14. Lipsitch, M. and B.R. Bloom. 2006. Avian flu preparing for a pandemic. Harvard Pub. Health Rev. Winter 2006:4-7.
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