title: [Establish and optimization of real-time fluorescent reverse transcription loop-mediated isothermal amplification for detection of avian influenza H5 hemagglutinin gene].
abstract: H5 subtype avian influenza (AIV-H5) is a major causative agent of animalloimia a rapid and sensitive molecular biological diagnosis is crucial to the control program of AIV-H5. AIV-H5 real-time fluorescent reverse transcription loop-mediated isothermal amplification (qRT-LAMP) was established by means of heat treatment of the samples. The sensitivity, specificity and repeatability of this method were assessed and the performance of Calcein,SYBR Green I,HNB,SYTO 81 in colorimetric detection was comparatively analyzed to screen the optimum dye. The results showed the sensitivity of this method was 100 times higher than that of standard real-time fluorescent RT-PCR, and the detection limit was one copy of the gene per reaction. This method had no cross-reactivity with other common avian respiratory tract infectious disease-related pathogens such as IBV and NDV. The present study suggested Calcein was the optimum dye. Small-scale tests suggested this method was reliable for survey monitoring of AIV-H5 on the spot, indicating its potential applications in field investigation.
authors.affiliation: Sichuan Agricultural University, Ya'an, 625014, China. melody_liuyun_19@126.com
authors.forename: Yun
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Jin-ming
authors.initials: JM
authors.lastname: Tang
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Hong
authors.initials: H
authors.lastname: Tao
authors.affiliation: Shenzhen Key Laboratory of Inspection Research & Development of Alien Pests, Shenzhen, 518045, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Sun
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Ti-kang
authors.initials: TK
authors.lastname: Lu
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Li-shan
authors.initials: LS
authors.lastname: Liao
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Jian-li
authors.initials: JL
authors.lastname: Liu
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Shao-ling
authors.initials: SL
authors.lastname: Zeng
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Chen-fu
authors.initials: CF
authors.lastname: Cao
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Cai-hong
authors.initials: CH
authors.lastname: Zhang
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Zhou-xi
authors.initials: ZX
authors.lastname: Ruan
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Jian-qiang
authors.initials: JQ
authors.lastname: Lv
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Jun-xing
authors.initials: JX
authors.lastname: Yang
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Qun-yi
authors.initials: QY
authors.lastname: Hua
authors.affiliation: Sichuan Agricultural University, Ya'an, 625014, China.
authors.forename: Zheng-li
authors.initials: ZL
authors.lastname: Chen
authors.affiliation: Shenzhen Entry-Exit Inspection & Quarantine Bureau, Shenzhen, 518045, China.
authors.forename: Zhi-feng
authors.initials: ZF
authors.lastname: Qin
mesh_terms: D000818:Animals; D002645:Chickens; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics*; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics / Q000302:isolation & purification*; D005585:Influenza in Birds / Q000175:diagnosis / Q000821:virology*; D011201:Poultry Diseases / Q000175:diagnosis / Q000821:virology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000295:instrumentation / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus; C109963:hemagglutinin, avian influenza A virus