1 Poultry Institute , Chinese Academy of Agricultural Sciences, Yangzhou, China .
2 Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonose, Yangzhou University , Yangzhou, Jiangsu, China .
Foodborne Pathog Dis. 2018 Jul;15(7):413-419. doi: 10.1089/fpd.2017.2410. Epub 2018 Apr 30.
Antimicrobial resistance genes play an important role in mediating resistance to sulfonamide in Gram-negative bacteria. While PCR is the current method to detect sulfonamide resistance genes (sul1, sul2, sul3), it is time-consuming and costly and there is an urgent need to develop a more convenient, simpler and rapid test for the sul. In this study, we describe a multiplex loop-mediated isothermal amplification (m-LAMP) assay we developed for the rapid and simultaneous detection of three sul. This m-LAMP assay successfully detected seven reference strains with different sul genotypes, but was negative for nine sul-negative reference strains. The m-LAMP products were verified by HinfI restriction enzyme digestion and the detection limit of the test was 0.5 pg genomic DNA per reaction. Testing 307 sulfonamide-resistant Enterobacteriaceae clinical isolates with the m-LAMP revealed all were positive for the sul with sul2 (79.5%) and sul1 (64.5%) being most prevalent, and sul3 the least (12.1%). Of the Enterobacteriaceae isolates tested, the Salmonella Indiana, a newly emerging serovar resistant to numerous antimicrobials, were most commonly positive with 33% having sul3.
抗菌药物耐药基因在介导革兰氏阴性菌对磺胺类药物的耐药性方面发挥着重要作用。虽然聚合酶链反应（PCR）是目前检测磺胺类药物耐药基因（sul1、sul2、sul3）的方法，但该方法既耗时又昂贵，因此迫切需要开发一种更方便、更简单、更快速的 sul 检测方法。在本研究中，我们描述了一种用于快速同时检测三种 sul 的多重环介导等温扩增（m-LAMP）检测方法。该 m-LAMP 检测方法成功地检测到了七种具有不同 sul 基因型的参考菌株，但对九种 sul 阴性参考菌株呈阴性。m-LAMP 产物通过 HindIII 限制性内切酶消化进行验证，该检测方法的检测限为每个反应 0.5 pg 基因组 DNA。用 m-LAMP 对 307 株磺胺类药物耐药性肠杆菌科临床分离株进行检测，结果均为 sul 阳性，其中 sul2（79.5%）和 sul1（64.5%）最为常见，sul3 则最少（12.1%）。在所检测的肠杆菌科分离株中，新型出现的对多种抗菌药物耐药的沙门氏菌印第安纳血清型最常阳性，其中 33%携带 sul3。
