猪霍乱沙门氏菌的分离与鉴定以及PCR检测方法的建立

Isolation and Identification of Salmonella choleraesuis and the Establishment of Its PCR Detective Method

[目的]为了确定四川省某猪场初生仔猪大规模发病的原因。[方法]对发病仔猪进行细菌分离、培养特性及形态结构观察、生化鉴定、药敏试验、动物致病性试验和血清型鉴定,并建立PCR快速检测方法。[结果]从发病仔猪体内分离到1株优势菌,初步鉴定为沙门氏菌。根据沙门氏菌invB基因设计的1对引物进行PCR扩增,得到408bp的特异性条带,PCR产物经T-A克隆测序证实为invB基因,与参比序列具有非常高的同源性,说明该PCR检测方法具有较高的特异性和敏感性。经血清学试验确定该分离株为猪霍乱沙门氏菌,毒力试验和药敏试验结果表明分离菌致病性很强,对头孢三嗪、氯霉素、氧氟沙星等敏感。[结论]该研究为沙门氏菌病原鉴定与疾病诊断提供了依据。

[Objective] The aim of the research was to confirm the mass diseased cause of newborn piglets in a pig farm of Siehuan Province. [Method]The bacteria isolation, observation of cultured characters and morphologie structure, biochemical identification, drug sensitivity test were conducted on diseased piglets, and a rapid PCR detective method was established. [Result] One dominant strain was isolated from diseased piglets and was preliminarily identified as Salmonella. A specific band with 408 bp was obtained by PCR amplification with a pair of specific primers designed according to the Salmonella invB gene. PCR products were confirmed as invB genes by T-A clone and sequencing and had high homology compared with the reference sequences, which indicated that the PCR detective method had high specificity and sensitivity. The isolated strain was confirmed as Salmonella choleraesuis through serology test. And the results of toxicity test and drug sensitivity test showed that the isolated strain had a strong pathogenicity and was sensitive to eefiriaxone, ehloramphenieol and ofloxaein. [Conclusion] The study provided a basis for the pathogen identification of Salmonella and disease diagnosis.