Rapid detection of infectious laryngotracheitis virus isolates by loop-mediated isothermal amplification.
The objective of this study was to develop and evaluate a loop-mediated isothermal amplification (LAMP) method to detect infectious laryngotracheitis virus (ILTV) from commercial broiler and layer flocks in southern China. A set of six specific primers was designed to recognize six distinct genomic sequences of thymidine kinase (TK) from ILTV. The entire assay duration was recorded at 40 min under isothermal condition at 63.5 degrees C. The amplified products were analyzed by electrophoresis and visual judgment by the SYBR Green I dyeing. LAMP assay was 10-fold more sensitive than the routine PCR assay, with a detection limit of 46 copies per reaction. In detecting ILTV, the LAMP assay detected all 5 strains previously isolated, did not cross-react with other avian pathogens, and obtained a 100% sensitivity in 43 positive clinical samples with reference to virus isolation. Therefore, the LAMP assay may be a good alternative method for specific diagnosis of ILTV infection in primary care facilities, and in less well-equipped laboratories.
Xie
Qing-mei
QM
College of Animal Science, South China Agricultural University, Guangzhou 510642, China. qmx@scau.edu.cn
Ji
Jun
J
Pickens
Tristan Tyler
TT
Du
Li-qin
LQ
Cao
Yong-chang
YC
Li
Hong-mei
HM
Wang
Lin-guo
LG
Ma
Jing-yun
JY
Bi
Ying-zuo
YZ
D000818:Animals; D052160:Benzothiazoles; D002645:Chickens; D002681:China; D019411:Clinical Laboratory Techniques / Q000379:methods*; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D003959:Diamines; D004587:Electrophoresis, Agar Gel; D005456:Fluorescent Dyes; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D001352:Herpesvirus 1, Gallid / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009930:Organic Chemicals; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology; D011804:Quinolines; D012680:Sensitivity and Specificity; D013194:Staining and Labeling; D013997:Time Factors
D052160:Benzothiazoles; D017931:DNA Primers; D004279:DNA, Viral; D003959:Diamines; D005456:Fluorescent Dyes; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I