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A pseudovirus-based hemagglutination-inhibition assay as a rapid, highly sensitive, and specific assay for detecting avian influenza A (H7N9) antibodies

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      Human infection with a novel avian-origin influenza A (H7N9) virus.

      Infection of poultry with influenza A subtype H7 viruses occurs worldwide, but the introduction of this subtype to humans in Asia has not been observed previously. In March 2013, three urban residents of Shanghai or Anhui, China, presented with rapidly progressing lower respiratory tract infections and were found to be infected with a novel reassortant avian-origin influenza A (H7N9) virus. We obtained and analyzed clinical, epidemiologic, and virologic data from these patients. Respiratory specimens were tested for influenza and other respiratory viruses by means of real-time reverse-transcriptase-polymerase-chain-reaction assays, viral culturing, and sequence analyses. A novel reassortant avian-origin influenza A (H7N9) virus was isolated from respiratory specimens obtained from all three patients and was identified as H7N9. Sequencing analyses revealed that all the genes from these three viruses were of avian origin, with six internal genes from avian influenza A (H9N2) viruses. Substitution Q226L (H3 numbering) at the 210-loop in the hemagglutinin (HA) gene was found in the A/Anhui/1/2013 and A/Shanghai/2/2013 virus but not in the A/Shanghai/1/2013 virus. A T160A mutation was identified at the 150-loop in the HA gene of all three viruses. A deletion of five amino acids in the neuraminidase (NA) stalk region was found in all three viruses. All three patients presented with fever, cough, and dyspnea. Two of the patients had a history of recent exposure to poultry. Chest radiography revealed diffuse opacities and consolidation. Complications included acute respiratory distress syndrome and multiorgan failure. All three patients died. Novel reassortant H7N9 viruses were associated with severe and fatal respiratory disease in three patients. (Funded by the National Basic Research Program of China and others.).
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        Epidemiology of human infections with avian influenza A(H7N9) virus in China.

        The first identified cases of avian influenza A(H7N9) virus infection in humans occurred in China during February and March 2013. We analyzed data obtained from field investigations to describe the epidemiologic characteristics of H7N9 cases in China identified as of December 1, 2013. Field investigations were conducted for each confirmed case of H7N9 virus infection. A patient was considered to have a confirmed case if the presence of the H7N9 virus was verified by means of real-time reverse-transcriptase-polymerase-chain-reaction assay (RT-PCR), viral isolation, or serologic testing. Information on demographic characteristics, exposure history, and illness timelines was obtained from patients with confirmed cases. Close contacts were monitored for 7 days for symptoms of illness. Throat swabs were obtained from contacts in whom symptoms developed and were tested for the presence of the H7N9 virus by means of real-time RT-PCR. Among 139 persons with confirmed H7N9 virus infection, the median age was 61 years (range, 2 to 91), 71% were male, and 73% were urban residents. Confirmed cases occurred in 12 areas of China. Nine persons were poultry workers, and of 131 persons with available data, 82% had a history of exposure to live animals, including chickens (82%). A total of 137 persons (99%) were hospitalized, 125 (90%) had pneumonia or respiratory failure, and 65 of 103 with available data (63%) were admitted to an intensive care unit. A total of 47 persons (34%) died in the hospital after a median duration of illness of 21 days, 88 were discharged from the hospital, and 2 remain hospitalized in critical condition; 2 patients were not admitted to a hospital. In four family clusters, human-to-human transmission of H7N9 virus could not be ruled out. Excluding secondary cases in clusters, 2675 close contacts of case patients completed the monitoring period; respiratory symptoms developed in 28 of them (1%); all tested negative for H7N9 virus. Most persons with confirmed H7N9 virus infection had severe lower respiratory tract illness, were epidemiologically unrelated, and had a history of recent exposure to poultry. However, limited, nonsustained human-to-human H7N9 virus transmission could not be ruled out in four families.
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          Detection of antibody to avian influenza A (H5N1) virus in human serum by using a combination of serologic assays.

          From May to December 1997, 18 cases of mild to severe respiratory illness caused by avian influenza A (H5N1) viruses were identified in Hong Kong. The emergence of an avian virus in the human population prompted an epidemiological investigation to determine the extent of human-to-human transmission of the virus and risk factors associated with infection. The hemagglutination inhibition (HI) assay, the standard method for serologic detection of influenza virus infection in humans, has been shown to be less sensitive for the detection of antibodies induced by avian influenza viruses. Therefore, we developed a more sensitive microneutralization assay to detect antibodies to avian influenza in humans. Direct comparison of an HI assay and the microneutralization assay demonstrated that the latter was substantially more sensitive in detecting human antibodies to H5N1 virus in infected individuals. An H5-specific indirect enzyme-linked immunosorbent assay (ELISA) was also established to test children's sera. The sensitivity and specificity of the microneutralization assay were compared with those of an H5-specific indirect ELISA. When combined with a confirmatory H5-specific Western blot test, the specificities of both assays were improved. Maximum sensitivity (80%) and specificity (96%) for the detection of anti-H5 antibody in adults aged 18 to 59 years were achieved by using the microneutralization assay combined with Western blotting. Maximum sensitivity (100%) and specificity (100%) in detecting anti-H5 antibody in sera obtained from children less than 15 years of age were achieved by using ELISA combined with Western blotting. This new test algorithm is being used for the seroepidemiologic investigations of the avian H5N1 influenza outbreak.
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            Author and article information

            Affiliations
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            Author notes
            Correspondence to: Jianqing Xu, Email: xujianqing2014@ 123456126.com ; Tel: +86-21-37990333-7335; Fax: +86-21-57247094.

            : These authors contributed equally to this article.

            Contributors
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            From Shanghai Public Health Clinical Center & Institutes of Biomedical Sciences, Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, Shanghai Medical College, Fudan University, Shanghai, 201508, China
            Journal
            Infectious Diseases and Translational Medicine
            Infect. Dis. Transl. Med.
            Infect. Dis. Transl. Med.
            International Biological and Medical Journals Publishing House Co., Limited (Room E16, 3/f, Yongda Commercial Building, No.97, Bonham Stand (Sheung Wan), HongKong )
            2411-2917
            30 June 2015
            : 1
            : 1
            : 12-15
            10.11979/idtm.201501005

            This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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            Figures: 2, Tables: 2, References: 11, Pages: 4
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            Categories
            Letter: Detection Techniques

            Medicine, Infectious disease & Microbiology

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