The completion of the genetic sequencing of the 1918 influenzaA virus by Taubenberger et al.1 and the subsequent recoveryof the virus by Tumpey et al.2 using reverse genetic techniquesare spectacular achievements of contemporary molecular biologyand provide important insights into the origin of pandemic influenza.The three pandemic viruses that emerged in the 20th century— the 1918 ("Spanish influenza") H1N1 virus, the 1957("Asian influenza") H2N2 virus, and the 1968 ("Hong Kong influenza")H3N2 virus — all spread rapidly around the world, butonly the 1918 virus was associated with mortality measured inthe thousands per 100,000 population.
In both 1957 and 1968, a new influenza virus emerged becauseof reassortment events involving two influenza viruses. Thesegmented genome allows each influenza A virus to exchange geneticmaterial with other influenza A viruses. In 1957, dual infectionof an individual animal — probably a human, but possiblyanother species, such as a pig — with an avian H2N2 influenzaand a human H1N1 influenza resulted in the emergence of a newinfluenza virus containing the hemagglutinin, the neuraminidase,and the gene for one of the polymerase proteins (PB1) from theavian virus, along with the remaining five genetic segmentsfrom the human H1N1 influenza virus.
The new reassortant virus circulated in humans until 1968, whenit was replaced by another reassortant virus, the H3N2 HongKong virus — created by the replacement of the hemagglutinin(H2) and polymerase (PB1) genes of the H2N2 virus with two newavian genes, H3 and a new PB1. Today, the descendants of thisvirus continue to cause the majority of influenza infectionsin humans (see diagram). Five of the genes of today's H3N2 influenzavirus have their origin in the 1918 pandemic.
The Two Mechanisms whereby Pandemic Influenza Originates.
In 1918, an H1N1 virus closely related to avian viruses adapted to replicate efficiently in humans. In 1957 and in 1968, reassortment events led to new viruses that resulted in pandemic influenza. The 1957 influenza virus (Asian influenza, an H2N2 virus) acquired three genetic segments from an avian species (a hemagglutinin, a neuraminidase, and a polymerase gene, PB1), and the 1968 influenza virus (Hong Kong influenza, an H3N2 virus) acquired two genetic segments from an avian species (hemagglutinin and PB1). Future pandemic strains could arise through either mechanism.
The startling observation of Taubenberger et al. was that the1918 virus did not originate through a reassortment event involvinga human influenza virus: all eight genes of the H1N1 virus aremore closely related to avian influenza viruses than to influenzafrom any other species, indicating that an avian virus musthave infected humans and adapted to them in order to spreadfrom person to person. Thus, pandemic influenza may originatethrough at least two mechanisms: reassortment between an animalinfluenza virus and a human influenza virus that yields a newvirus, and direct spread and adaptation of a virus from animalsto humans.
The concern at present relates to the widespread epidemic ofavian H5N1 influenza in domestic fowl, as well as wild birds,with sporadic transmission to humans. In the past decade, numerousinstances of bird-to-human transmission have been recognized(see table); although we have only recently become aware ofthem, these events are surely not new. What is new is the broadeningof the range of avian and nonavian species that have becomeinfected with the current H5N1 virus.3
Examples of Transmission of Avian Influenza Viruses to Humans.
The characterization of the recovered 1918 virus in tissue cultureand mice reveals at least two unique qualities. This virus isable to replicate and form plaques on tissue-culture monolayersin the absence of the protease trypsin. Normally, a proteasesuch as trypsin is required to activate the hemagglutinin inorder to initiate the infection of tissue culture, but the 1918virus can activate its own hemagglutinin through the actionof neuraminidase, either directly or indirectly (possibly byneuraminidase's binding of a host protease). The exact mechanismby which the neuraminidase takes on the protease activity hasnot been determined.
In addition, the 1918 virus is 100 times as lethal in mice asany other human influenza virus; the median lethal dose (LD50,or 103.5 to 3.75 median egg infectious doses [EID50]) is low,and the virus replicates rapidly so that high titers (>107EID50 per milliliter) are found in the lungs of infected mice.High virus inocula result in the death of mice as early as threedays after they have been infected. The 1918 virus was susceptibleto the adamantine compounds (amantadine and rimantadine) andneuraminidase inhibitors (oseltamivir and zanamivir), and theavailability of the recovered virus will facilitate studiesof other therapeutics. For example, the vigorous release ofcytokines in mice infected with the 1918 influenza virus isassociated with rapid onset of pulmonary disease and death.Compounds that block the action of specific cytokines can nowbe evaluated as therapeutics that might help to reduce the mortalityassociated with pandemic influenza.
It is not possible to know whether the current H5N1 is capableof adapting to humans so that it can spread with high efficiencythrough low-titer aerosol transmission to initiate an influenzapandemic. However, Taubenberger et al. provide some guidanceon the genetic changes that might be required for such an event.The role of PB1 must be critical, since in both 1957 and 1968,this polymerase gene was transferred along with the hemagglutininduring reassortment. By comparing the consensus sequence ofthe three avian influenza polymerase genes PA, PB1, and PB2with the 1918 sequence, as well as with more contemporary influenzaviruses, Taubenberger et al. have identified four amino acidsof PA, one of PB1, and five of PB2 that are found in human influenzaviruses (including the 1918 virus) but generally not in avianinfluenza viruses. In two instances, these amino acids are foundin nuclear localization signaling regions, suggesting that someor all of these amino acid differences are critical for thevirus to adapt to humans.
The genetic sequences of the 1997 Hong Kong H5N1 virus and the2004 Vietnam H5N1 virus reveal that several human isolates ofthese viruses contain one of the five amino acid changes inPB2 that have been identified as important to the ability ofthe 1918 virus to infect humans. This finding suggests thatseveral additional genetic changes must occur before these viruseswill begin to spread efficiently from person to person. Thegenetic sequences of avian viruses may provide a window throughwhich to monitor these sporadic transmissions for the potentialof the viruses to adapt to humans. The occurrence of additionalgenetic changes in the avian H5N1 virus circulating in birdsthat match the consensus sequence for PA, PB1, or PB2 in humaninfluenza would be cause for heightened concern.
On the basis of the rates of replacement of amino acids, Taubenbergeret al. estimated that avian influenza polymerase genes had beencirculating in humans as early as 1900. If this estimate iscorrect, then monitoring of the sequences of viruses isolatedin instances of bird-to-human transmission for genetic changesin key regions may enable us to track viruses years before theydevelop the capacity to replicate with high efficiency in humans.Knowledge of the genetic sequences of influenza viruses thatpredate the 1918 pandemic would be extremely helpful in determiningthe events that may lead to the adaptation of avian virusesto humans before the occurrence of pandemic influenza. We couldthen conduct worldwide surveillance for similar events involvingcontemporary avian viruses.
Source Information
Dr. Belshe is a professor of medicine, pediatrics, and molecular microbiology in the Division of Infectious Diseases and Immunology, Department of Internal Medicine, Saint Louis University, St. Louis.
An interview with Dr. Belshe can be heard at www.nejm.org.
References
Taubenberger JK, Reid AH, Lourens RM, Wang R, Jin G, Fanning TG. Characterization of the 1918 influenza virus polymerase genes. Nature 2005;437:889-893. [CrossRef][Medline]
Tumpey TM, Basler CF, Aguilar PV, et al. Characterization of the reconstructed 1918 Spanish influenza pandemic virus. Science 2005;310:77-80. [Free Full Text]
The Writing Committee of the World Health Organization (WHO) Consultation on Human Influenza A/H5. Avian influenza A (H5N1) infection in humans. N Engl J Med 2005;353:1374-1385. [Free Full Text]
Chen, Z., Wang, W., Zhou, H., Suguitan, A. L. Jr., Shambaugh, C., Kim, L., Zhao, J., Kemble, G., Jin, H.
(2010). Generation of Live Attenuated Novel Influenza Virus A/California/7/09 (H1N1) Vaccines with High Yield in Embryonated Chicken Eggs. J. Virol.
84: 44-51
[Abstract][Full Text]
Shih, S.-R., Horng, J.-T., Poon, L. L. M., Chen, T.-C., Yeh, J.-Y., Hsieh, H.-P., Tseng, S.-N., Chiang, C., Li, W.-L., Chao, Y.-S., Hsu, J. T.-A.
(2010). BPR2-D2 targeting viral ribonucleoprotein complex-associated function inhibits oseltamivir-resistant influenza viruses. J Antimicrob Chemother
65: 63-71
[Abstract][Full Text]
Sym, D., Patel, P. N, El-Chaar, G. M
(2009). Seasonal, Avian, and Novel H1N1 Influenza: Prevention and Treatment Modalities. The Annals of Pharmacotherapy
43: 2001-2011
[Abstract][Full Text]
Munster, V. J., de Wit, E., van den Brand, J. M. A., Herfst, S., Schrauwen, E. J. A., Bestebroer, T. M., van de Vijver, D., Boucher, C. A., Koopmans, M., Rimmelzwaan, G. F., Kuiken, T., Osterhaus, A. D. M. E., Fouchier, R. A. M.
(2009). Pathogenesis and Transmission of Swine-Origin 2009 A(H1N1) Influenza Virus in Ferrets. Science
325: 481-483
[Abstract][Full Text]
Stein, R. A.
(2009). Lessons From Outbreaks of H1N1 Influenza. ANN INTERN MED
151: 59-62
[Abstract][Full Text]
MOSSAD, S. B.
(2009). The resurgence of swine-origin influenza A (H1N1). Cleveland Clinic Journal of Medicine
76: 337-343
[Abstract][Full Text]
McSharry, J. J., Weng, Q., Brown, A., Kulawy, R., Drusano, G. L.
(2009). Prediction of the Pharmacodynamically Linked Variable of Oseltamivir Carboxylate for Influenza A Virus Using an In Vitro Hollow-Fiber Infection Model System. Antimicrob. Agents Chemother.
53: 2375-2381
[Abstract][Full Text]
Chaipan, C., Kobasa, D., Bertram, S., Glowacka, I., Steffen, I., Solomon Tsegaye, T., Takeda, M., Bugge, T. H., Kim, S., Park, Y., Marzi, A., Pohlmann, S.
(2009). Proteolytic Activation of the 1918 Influenza Virus Hemagglutinin. J. Virol.
83: 3200-3211
[Abstract][Full Text]
Gray, G. C., Kayali, G.
(2009). Facing pandemic influenza threats: The importance of including poultry and swine workers in preparedness plans. Poult. Sci.
88: 880-884
[Abstract][Full Text]
Li, F C K, Choi, B C K, Sly, T, Pak, A W P
(2008). Finding the real case-fatality rate of H5N1 avian influenza. J. Epidemiol. Community Health
62: 555-559
[Abstract][Full Text]
Huang, I-C., Li, W., Sui, J., Marasco, W., Choe, H., Farzan, M.
(2008). Influenza A Virus Neuraminidase Limits Viral Superinfection. J. Virol.
82: 4834-4843
[Abstract][Full Text]
Wan, X.-F., Chen, G., Luo, F., Emch, M., Donis, R.
(2007). A quantitative genotype algorithm reflecting H5N1 Avian influenza niches. Bioinformatics
23: 2368-2375
[Abstract][Full Text]
Yen, H.-L., Lipatov, A. S., Ilyushina, N. A., Govorkova, E. A., Franks, J., Yilmaz, N., Douglas, A., Hay, A., Krauss, S., Rehg, J. E., Hoffmann, E., Webster, R. G.
(2007). Inefficient Transmission of H5N1 Influenza Viruses in a Ferret Contact Model. J. Virol.
81: 6890-6898
[Abstract][Full Text]
Thomas, J. K., Noppenberger, J.
(2007). Avian influenza: A review. Am J Health Syst Pharm
64: 149-165
[Abstract][Full Text]
Skowronski, D. M., Li, Y., Tweed, S. A., Tam, T. W.S., Petric, M., David, S. T., Marra, F., Bastien, N., Lee, S. W., Krajden, M., Brunham, R. C.
(2007). Protective measures and human antibody response during an avian influenza H7N3 outbreak in poultry in British Columbia, Canada. CMAJ
176: 47-53
[Abstract][Full Text]
Kramer, J. S., Durham, C., Schroeder, T., Garrelts, J. C.
(2006). Effectiveness of half-dose versus full-dose influenza vaccine in health care workers.. Am J Health Syst Pharm
63: 2111-2115
[Abstract][Full Text]
Bartlett, J. G.
(2006). Planning for Avian Influenza. ANN INTERN MED
145: 141-144
[Abstract][Full Text]
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