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Establishment of a Multiplex Loop-Mediated Isothermal Amplification Method for Rapid Detection of Sulfonamide Resistance Genes (sul1, sul2, sul3) in Clinical Enterobacteriaceae Isolates from Poultry

Author:
Gong, Jiansen, Zhuang, Linlin, Zhang, Di, Zhang, Ping, Dou, Xinhong, Wang, Chengming
Source:
Foodborne pathogens & disease 2018 v.15 no.7 pp. 413-419
ISSN:
1556-7125
Subject:
DNA, Gram-negative bacteria, Salmonella Indiana, anti-infective agents, antibiotic resistance genes, detection limit, genotype, loop-mediated isothermal amplification, polymerase chain reaction, poultry, rapid methods, restriction endonucleases, serotypes
Abstract:
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.
Agid:
6458606