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Development of a reverse transcription recombinase polymerase amplification combined with lateral-flow dipstick assay for avian influenza H9N2 HA gene detection.
Wang, Zeng; Yang, Pan-Pan; Zhang, Yu-Han; Tian, Kai-Yue; Bian, Chuan-Zhou; Zhao, Jun.
Afiliação
  • Wang Z; College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, China.
  • Yang PP; College of Veterinary Medicine, Henan University of Animal Husbandry and Economy, Zhengzhou, China.
  • Zhang YH; College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, China.
  • Tian KY; College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, China.
  • Bian CZ; College of Veterinary Medicine, Henan University of Animal Husbandry and Economy, Zhengzhou, China.
  • Zhao J; College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, China.
Transbound Emerg Dis ; 66(1): 546-551, 2019 Jan.
Article em En | MEDLINE | ID: mdl-30403438
ABSTRACT
H9N2 avian influenza viruses (AIVs) have been detected from wild birds and domestic poultry worldwide. Serious diseases combined with secondary infection have caused high mortality and great economic losses to poultry industry. Therefore, simple, rapid, sensitive and accurate methods suitable for field detection of H9N2 AIVs are crucial to efficiently control virus infection and spread in time. In this study, an isothermal reverse transcription recombinase polymerase amplification with lateral-flow dipstick (RT-RPA-LFD) assay for detection of hemagglutinin (HA) gene of H9 subtype influenza viruses was developed. The optimal forward and reverse primers targeting HA gene of H9 subtype influenza viruses were labeled with fluorescein isothiocyanate (FITC) and biotin at the 5'-end, respectively. The amplification reaction could be finished in 20 min at a wide temperature range of 30-42°C, and then the products could be visualized with naked eyes. The developed H9 RT-RPA-LFD was able to detect 0.15 pg of H9N2 AIV RNA, which was 10 times more sensitive than that of conventional RT-PCR. The H9 RT-RPA-LFD assay did not detect nucleic acids extracted from H9 negative samples or from other poultry respiratory pathogens. The clinical performance of H9 RT-RPA-LFD was determined by testing 120 cloacal samples collected from chickens with respiratory syndromes. The coincidence rate of the detection results between RT-RPA-LFD and conventional RT-PCR was 95.8%. Therefore, the developed RT-RPA-LFD assay provides a rapid, reliable and sensitive method for field diagnosis of H9 subtype AIVs.
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Texto completo: 1 Base de dados: MEDLINE Assunto principal: Doenças das Aves Domésticas / Glicoproteínas de Hemaglutininação de Vírus da Influenza / Técnicas de Amplificação de Ácido Nucleico / Transcrição Reversa / Vírus da Influenza A Subtipo H9N2 / Reação em Cadeia da Polimerase em Tempo Real / Influenza Aviária Tipo de estudo: Diagnostic_studies Limite: Animals Idioma: En Ano de publicação: 2019 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Doenças das Aves Domésticas / Glicoproteínas de Hemaglutininação de Vírus da Influenza / Técnicas de Amplificação de Ácido Nucleico / Transcrição Reversa / Vírus da Influenza A Subtipo H9N2 / Reação em Cadeia da Polimerase em Tempo Real / Influenza Aviária Tipo de estudo: Diagnostic_studies Limite: Animals Idioma: En Ano de publicação: 2019 Tipo de documento: Article