title: Detection of Haemophilus parasuis isolates from South China by loop-mediated isothermal amplification and isolate characterisation.
abstract: Haemophilus parasuis is the etiological agent of Glässer's disease, which is characterised by ﬁbrinous polyserositis, meningitis and polyarthritis, causing severe economic losses to the  swine industry. In this study, a loop-mediated isothermal ampliﬁcation (LAMP) test was  developed to improve the speciﬁcity, facility and speed of diagnosis of H. parasuis isolates. The LAMP assay rapidly ampliﬁed the target gene within 50 min incubation at 63 °C in a  laboratory water bath. The LAMP amplicon could be visualised directly in the reaction tubes  following the addition of SYBR Green I dye. The detection limit of this LAMP method was  10 CFU/mL, which was 10 times more sensitive than the earlier 16S rRNA polymerase chain  reaction (PCR) test conducted by Oliveira, Galina and Pijoan (2001), and no cross-reactivity  was observed from other non-H. parasuis strains. This LAMP test was evaluated further on  187 clinical specimens from pigs suspected of being infected with H. parasuis. Forty-three were found positive by bacterial isolation of H. parasuis, as well as by the 16S rRNA PCR and LAMP tests. The 43 H. parasuis isolates were classiﬁed into 9 serovars and had 37 genetic patterns when analysed by pulsed-ﬁeld gel electrophoresis (PFGE). This displayed that various H. parasuis serovars and genotypes were widely distributed in South China. Therefore, the speed, speciﬁcity and sensitivity of the LAMP test, the lack of a need for expensive equipment, and the visual readout showed great potential for a correct clinical diagnosis of H. parasuis in favour of controlling Glässer's disease.
authors: The Key Laboratory of Animal Disease Control and Prevention of the Ministry   of Agriculture, South China Agricultural University. chgangxu@126.com.
authors: Jian-Min
authors: JM
authors: Zhang
authors: Hai-Yan
authors: HY
authors: Shen
authors: Ming
authors: M
authors: Liao
authors: Tao
authors: T
authors: Ren
authors: Li-Li
authors: LL
authors: Guo
authors: Cheng-Gang
authors: CG
authors: Xu
authors: Sai-Xiang
authors: SX
authors: Feng
authors: Hui-Ying
authors: HY
authors: Fan
authors: Jing-Yi
authors: JY
authors: Li
authors: Ji-Dang
authors: JD
authors: Chen
authors: Bin
authors: B
authors: Zhang
mesh_terms: D000818:Animals; D002681:China; D016521:Electrophoresis, Gel, Pulsed-Field / Q000662:veterinary; D006192:Haemophilus Infections / Q000175:diagnosis / Q000662:veterinary*; D044137:Haemophilus parasuis / Q000145:classification / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods / Q000662:veterinary*; D016133:Polymerase Chain Reaction / Q000662:veterinary; D012336:RNA, Ribosomal, 16S / Q000737:chemistry; D012680:Sensitivity and Specificity; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis*
chemical_list: D012336:RNA, Ribosomal, 16S