Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, Jiangsu 225009, China.
Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou, Jiangsu 225009, China.
Poult Sci. 2018 Nov 1;97(11):4000-4007. doi: 10.3382/ps/pey254.
Salmonella enterica subsp. enterica serovar Gallinarum biovar Gallinarum (S. Gallinarum) and biovar Pullorum (S. Pullorum) are gram-negative bacteria, members of the most important infectious pathogens, and have caused common problems in the poultry industry, especially in the developing countries. O- and H-antigen specific anti-sera are commonly for slide and tube agglutination tests to identify Salmonella serovars. However, it is both labor intensive and time consuming, so there is an urgent need for a new technique for the rapid detection of the major Salmonella serovars. In this study, we developed a 1-step PCR assay to identify the serovar Gallinarum. This PCR-based assay was based on the SPUL_2693 gene, which was located in SPI-19 and found by comparing the genomes of the S. Pullorum and S. Gallinarum in the whole data of NCBI. The specificity of this gene was evaluated by bioinformatics analysis, and the results showed that the SPUL_2693 gene exists in all serovar Gallinarum. The specificity and sensitivity of this PCR assay were evaluated in our study. The developed PCR assay was able to distinguish the serovar Gallinarum from 27 different Salmonella serovars and 5 different non-Salmonella pathogens. The minimum limit of genomic DNA of S. Pullorum for PCR detection was 2.143 pg/μL, and the minimum limit number of cells was 6 CFU. This PCR assay was also applied to analyze Salmonella strains isolated from a chicken farm in this study. The PCR assay properly identified the serovar Gallinarum from other Salmonella serovars, and the results were in agreement with the results of a traditional serotyping assay. In general, the newly developed PCR-based assay can be used to accurately judge the presence of the serovar Gallinarum and can be combined with traditional serotyping assays, especially in the case of large quantities of samples.
鸡白痢沙门氏菌和鸡伤寒沙门氏菌是革兰氏阴性菌，是最重要的传染病病原体之一，在禽类养殖业中造成了常见的问题，特别是在发展中国家。O 抗原和 H 抗原特异性抗血清通常用于玻片和试管凝集试验来鉴定沙门氏菌血清型。然而，这种方法既费时又费力，因此迫切需要一种新的技术来快速检测主要的沙门氏菌血清型。在本研究中，我们开发了一种用于鉴定鸡白痢血清型的一步式 PCR 检测方法。这种基于 PCR 的检测方法是基于 SPUL_2693 基因，该基因位于 SPI-19 中，是通过比较整个 NCBI 中鸡伤寒沙门氏菌和鸡白痢沙门氏菌的基因组发现的。通过生物信息学分析评估了该基因的特异性，结果表明该 SPUL_2693 基因存在于所有鸡白痢血清型中。本研究还评估了该基因的特异性和敏感性。该开发的 PCR 检测方法能够将鸡白痢血清型与 27 种不同的沙门氏菌血清型和 5 种不同的非沙门氏菌病原体区分开来。PCR 检测鸡伤寒沙门氏菌的最低基因组 DNA 限量为 2.143 pg/μL，最低细胞数为 6 CFU。该 PCR 检测方法还应用于分析本研究中从鸡场分离的沙门氏菌菌株。PCR 检测方法能够正确地将鸡白痢血清型与其他沙门氏菌血清型区分开来，结果与传统血清分型检测方法的结果一致。总的来说，新开发的基于 PCR 的检测方法可以准确判断鸡白痢血清型的存在，并可与传统的血清分型检测方法相结合，特别是在大量样本的情况下。
