title: Rapid detection of duck hepatitis A virus genotype C using reverse transcription loop-mediated isothermal amplification.
abstract: A one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was used and optimized to develop a rapid and sensitive detection system for duck hepatitis A virus genotype C (DHAV-C) RNA. A set of four specific primers was designed against highly conserved sequences located within the 3D gene from DHAV (strain GX1201). Under optimal reaction conditions, the sensitivity of DHAV-C-specific RT-LAMP was 100-fold higher than that of reverse transcriptase-polymerase chain reaction (RT-PCR), with a detection limit of 0.3pg (6.59×10(4) copies) per reaction. No cross-reactivity was observed from the samples of other duck viruses, which is in good accordance with RT-PCR. Furthermore, a positive reaction can be visually inspected by observing turbidity or color change after the addition of SYBR green I dye. The DHAV-C-specific RT-LAMP assay was applied to the samples and compared with RT-PCR. The positive-sample ratios were 26.7% (12 of 45) by RT-LAMP and 20% (9 of 45) by RT-PCR. Therefore, the newly developed RT-LAMP assay is a rapid, specific, sensitive, and cost-effective method of DHAV-C detection. This assay has potential applications in both clinical diagnosis and field surveillance of DHAV-C infection.
authors.affiliation: Division of Avian Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China.
authors.forename: Chuanfeng
authors.initials: C
authors.lastname: Li
authors.affiliation: Division of Avian Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China.
authors.forename: Zongyan
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Division of Avian Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China.
authors.forename: Chunchun
authors.initials: C
authors.lastname: Meng
authors.affiliation: Division of Avian Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, China. Electronic address: liugq@shvri.ac.cn.
authors.forename: Guangqing
authors.initials: G
authors.lastname: Liu
mesh_terms: D000818:Animals; D003362:Cost-Benefit Analysis; D017931:DNA Primers / Q000235:genetics; D005838:Genotype; D004371:Hepatitis Virus, Duck / Q000235:genetics / Q000302:isolation & purification*; D025202:Molecular Diagnostic Techniques / Q000191:economics / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000191:economics / Q000379:methods*; D010850:Picornaviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology; D012367:RNA, Viral / Q000235:genetics; D048348:Reverse Transcription; D012680:Sensitivity and Specificity; D013997:Time Factors; D014730:Veterinary Medicine / Q000191:economics / Q000379:methods*; D014773:Virology / Q000191:economics / Q000379:methods*
chemical_list: D017931:DNA Primers; D012367:RNA, Viral
keywords: Duck hepatitis A virus; Loop-mediated isothermal amplification; RNA-dependent RNA polymerase