title: Development and evaluation of a reverse transcription loop-mediated isothermal amplification assay for H3N8 equine influenza virus.
abstract: Reverse transcription loop-mediated isothermal amplification (RT-LAMP) was applied to the detection of equine influenza virus (EIV). Because equine influenza is caused currently by EIV of the H3H8 subtype, the RT-LAMP primer set was designed to target the hemagglutinin gene of this subtype. The detection limit of the RT-LAMP assay was a virus dilution of 10(-5); which was 10(3) times more sensitive than the Espline Influenza A&B-N test and 10 times more sensitive than a reverse transcription polymerase chain reaction (RT-PCR) assay. The specificity of the RT-LAMP assay was examined by using several equine pathogens and nasal swabs collected from horses with fever in 2010 after EIV was eradicated in Japan. No cross-reactions were observed. Using 100 nasal swabs collected from horses with fever during an EIV outbreak in 2007, the RT-LAMP assay detected EIV in 52 samples, whereas the Espline test and the RT-PCR assay detected EIV in only 17 and 41 samples, respectively. These results indicate that the RT-LAMP assay is specific for EIV and more sensitive than the Espline test and the RT-PCR assay. Because it provides high sensitivity and ease of manipulation without the need for a thermal cycler or gel electrophoresis, the RT-LAMP assay should be applicable for laboratory diagnosis of EIV.
authors: Epizootic Research Center, Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. nemoto_manabu@epizoo.equinst.go.jp
authors: Manabu
authors: M
authors: Nemoto
authors: Takashi
authors: T
authors: Yamanaka
authors: Hiroshi
authors: H
authors: Bannai
authors: Koji
authors: K
authors: Tsujimura
authors: Takashi
authors: T
authors: Kondo
authors: Tomio
authors: T
authors: Matsumura
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics; D006734:Horse Diseases / Q000175:diagnosis* / Q000821:virology; D006736:Horses; D053123:Influenza A Virus, H3N8 Subtype / Q000235:genetics / Q000302:isolation & purification*; D007564:Japan; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009297:Nasal Mucosa / Q000821:virology; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009976:Orthomyxoviridae Infections / Q000175:diagnosis* / Q000821:virology; D012680:Sensitivity and Specificity; D014730:Veterinary Medicine / Q000379:methods*; D014773:Virology / Q000379:methods*
chemical_list: D017931:DNA Primers