On episode #28 of the podcast “This Week in Virology”, Vincent, Dick, Alan, and Eric F. Donaldson discuss a new test for influenza H5N1, poliovirus in Minnesota, Koala retrovirus, batteries made from viruses, and SARS.
Last week the US Food and Drug Administration approved a rapid test for determining whether a person is infected with an H5N1 subtype of influenza virus. Why is it better than previously available influenza diagnostic assays?
The format of the new assay, according to its developer, ArborVita, “is the familiar lateral-flow rapid antigen assay”. This type of diagnostic test is often designed to detect the presence of a protein in a clinical specimen, and is simple enough to be performed at home or in a physician’s office. The test sample is applied to a solid substrate, often a thin strip of nitrocellulose held in a plastic frame (illustrated). The nitrocellulose is impregnated with antibodies directed against the protein which is being assayed. These antibodies are labeled with colored latex or gold spheres to permit visual readout of the assay. The same antibody, minus the colored spheres, is bound along a line further down the nitrocellulose strip. When the clinical sample is applied to one end of the strip, it migrates by capillary action towards the opposite end. A protein that reacts with the labeled antibodies will also be bound by the immobilized antibodies as the complex moves down the strip. The result is a visible colored line on the filter indicating that protein is present in the sample. If no substance is bound to the antibody, it will move past the immobilized antibody and no color will develop.
A well-known example of a lateral-flow rapid antigen assay is the home pregnancy test, which is designed to detect human chorionic gonadotropin in urine.
The lateral-flow assay format has been used previously to detect influenza virus in clinical specimens, and depends on the presence of a viral protein called NP. Because the test involves the reaction of antibodies with proteins, it takes several hours to develop. Furthermore, the the kit must be refrigerated because antibodies lose activity at higher temperatures.
The new test for influenza H5N1 is novel because it does not depend upon an antibody-antigen reaction. Rather it uses the fact that another viral protein, called NS1, contains a binding sequence for a PDZ domain, a common structural motif found in many different proteins. PDZ domains bind with high specificity to short peptide sequences found in the carboxy-terminal regions of proteins. They are scaffolding proteins that play roles in signaling pathways involved in a variety of cell functions including membrane trafficking and cell polarity.
Examination of the protein sequences from many avian influenza virus isolates identified a PDZ ligand at the carboxy-terminus of NS1. The results of biochemical assays then showed that NS1 proteins from avian influenza viruses react with PDZ domains. Because the C-terminal region of the NS1 protein differs for each influenza subtype, a PDZ protein with high affinity for the NS1 protein of H5 viruses was selected for the assay. The NS1-specific PDZ domain is used in place of the antibody to capture the viral protein from clinical samples, such as a throat or nose swab. It is faster (40 minutes versus several hours) than antibody-based tests and does not require refrigeration.
The new assay is a simple, rapid, and stable test that can be used in many settings to monitor infection of humans with influenza H5N1 viruses. It is also a wonderful example of how unexpected research findings – in this case the discovery of a PDZ ligand in the viral NS1 protein – may lead to novel and effective clinical diagnostic reagents.
Obenauer, J. (2006). Large-Scale Sequence Analysis of Avian Influenza Isolates Science, 311 (5767), 1576-1580 DOI: 10.1126/science.1121586
The many human viral diseases that have crossed from other animal species Â – such as AIDS, Ebola, SARS, encephalitis and respiratory diesease caused by henipaviruses – demonstrate the pathogenic potential of the zoonotic pool. Are there also reverse zoonoses – diseases of humans that are transferred to other animal species?
An example of a reverse zoonosis occurred just last week at Lincoln Park Zoo in Chicago, where a 9 year old chimpanzee died of respiratory disease caused by human metapneumovirus. This member of the paramyxovirus family is responsible for approximately 10% of all respiratory tract infections. All seven members of a group of chimpanzees were infected with the virus, but only one became ill. The virus was likely transmitted to the chimps by humans, but precisely how and when is not known. Outbreaks of fatal respiratory disease in wild chimpanzees have been reported previously, and human metapneumovirus has been one of several human viruses isolated from these primates. Such infections are expected to occur more frequently in zoological parks, and ando also in game preserves as human encroachment of these facilities increases.
Influenza virus may also be frequently transmitted from humans to other animal species. An outbreak of influenza recently lead to the death of 6 bonobos (a species of chimpanzee) in a Congo wildlife sanctuary. Although the influenza subtype responsible for the outbreak has not been identified, it has been suggested that the source was one or more visitors during the February epidemic of influenza in Kinshasha.
Human influenza viruses are frequently isolated from pigs. Since its emergence in 1968, the H3N2 subtype has infected pigs many times throughout the world, and has often caused serious outbreaks. More recently the influenza H1N1 subtype has been shown to infect pigs. Influenza viruses remain in the pig population for long periods of time, and may serve as reservoirs for the recycling of older influenza virus strains back into the human population.
Just as increasing human population, travel, and the global food business increase the likelhood that viruses will jump from animals into humans, the same factors ensure the reverse spread as well – often with dire consequences for zoological parks and wild and domestic animals. And the viruses we pass on may come back to haunt us another day.
Kaur, T., Singh, J., Tong, S., Humphrey, C., Clevenger, D., Tan, W., Szekely, B., Wang, Y., Li, Y., Alex Muse, E., Kiyono, M., Hanamura, S., Inoue, E., Nakamura, M., Huffman, M., Jiang, B., & Nishida, T. (2008). Descriptive epidemiology of fatal respiratory outbreaks and detection of a humanâ€related metapneumovirus in wild chimpanzees at Mahale Mountains National Park, Western Tanzania. American Journal of Primatology, 70 (8), 755-765 DOI: 10.1002/ajp.20565
Yu, H., Zhou, Y., Li, G., Zhang, G., Liu, H., Yan, L., Liao, M., & Tong, G. (2009). Further evidence for infection of pigs with human-like H1N1 influenza viruses in China Virus Research, 140 (1-2), 85-90 DOI: 10.1016/j.virusres.2008.11.008
de Jong, J., Smith, D., Lapedes, A., Donatelli, I., Campitelli, L., Barigazzi, G., Van Reeth, K., Jones, T., Rimmelzwaan, G., Osterhaus, A., & Fouchier, R. (2007). Antigenic and Genetic Evolution of Swine Influenza A (H3N2) Viruses in Europe Journal of Virology, 81 (8), 4315-4322 DOI: 10.1128/JVI.02458-06
I joined hostÂ Dr. Marc Pelletier on the TWiT podcast ‘Futures in Biotech’ to interview influenza virologist Dr. Peter Palese, professor and chair of microbiology at the Mount Sinai School of Medicine in New York. We talk about influenza and whyÂ Dr. Palese revived a virus that killed 50 million people.
On episode #26 of the podcast “This Week in Virology”, Vincent, Alan, and Rich Condit converse about induction of polyomavirus replication in multiple sclerosis patients treated with the MS drug Tysabri, the extent of human polyomavirus infection, selection of influenza vaccines for the 2009-10 season, cowpox virus transmission from animals to humans, vaccinia-like virus infecting humans and cattle in Brasil, and poxviruses.
The death of a dozen pigs from swine influenza last week in theÂ Philippines reminded me of an incident at Fort Dix, NJ in 1976. The infection of humans with a strain of swine influenza lead to a nationwide immunization campaign to curb a pandemic that never occurred.
An explosive outbreak of febrile respiratory disease raced through the 19,000 personnel at Fort Dix in January 1976. Virological laboratory studies revealed the presence of a new swine influenza strain which was named A/New Jersey/76 (Hsw1N1). The virus infected 230 soldiers and caused severe respiratory disease in 13, including one death.
At the time it was believed that a swine virus had caused the 1918-19 influenza pandemic. Therefore scientists were concerned that the virus had returned to Fort Dix and would soon cause another catastrophic outbreak. Dr. Edwin Kilbourne, a noted influenza researcher, and others convinced the US Public Health Service to contract for the production of 150 million doses of vaccine. In March of 1976 President Gerald Ford announced a program to inoculate every man, woman and child in the United States against swine flu. Immunizations began in October, but only 45 million doses had been distributed when the program was halted in December. By then it was clear that A/New Jersey/76 was going nowhere. An unfortunate consequence was that many individuals developed Guillain-BarrÃ© syndrome, a neurological disease involving muscle weakness, paralysis, and sometimes death.
Why didn’t A/New Jersey/76 spread to the general population? One factor was the limited contact between basic trainees and others who more frequently travel outside the facility. Older personnel may have been immune, because military influenza vaccine formulations from 1955 through 1969 contained a swine influenza component. Competition with concurrent circulating influenza virus strain, A/Victoria, might haveÂ limited the impact of A/New Jersey virus which is believed to transmit poorly among humans.
In retrospect, the swine flu program had many flaws. The vaccine should have been stockpiled until it was clear that an epidemic was taking place. Today we realize that the 1918 influenza virus is derived from an avian strain, not a swine strain – had this information been available in 1976, the immunization campaign would not have taken place. Presumably these and many other errors will not be repeated when the time comes to immunize against the next pandemic strain.
To this day the origin of A/New Jersey/76 virus is an enigma. One theory is that a swine virus was brought to Fort Dix early in 1976 as recruits returned after the holidays. However, none of the personnel who were interviewed admitted to having contact with pigs. The virus seems to have circulated at Fort Dix for about a month, then disappeared.