title: Development of Multiplex-Mismatch Amplification Mutation-PCR Assay for Simultaneous Detection of Campylobacter jejuni and Mutation in gyrA Gene Related to Fluoroquinolone Resistance.
abstract: Campylobacter jejuni, a foodborne pathogen, is the major cause of enteritis in humans worldwide, however, its increasing resistance to fluoroquinolones reported recently is of a major concern. In the present study, multiplex-mismatch amplification mutation assay-polymerase chain reaction (MMAMA-PCR) was developed for the first time with the aim to quickly identify C. jejuni and to detect the single nucleotide mutation (C-257 to T) frequently observed in gyrA gene, associated with the acquisition of resistance to fluoroquinolones. In this assay, mismatch amplification mutation primers for the detection of gyrA mutation in C. jejuni were coupled with primers for the hip gene encoding for hippuricase and 16S rRNA gene of C. jejuni, respectively, in the multiplex PCR assay. The specificity and accuracy of this method were analyzed by the use of 78 C. jejuni strains with previously confirmed resistance phenotypes and the mutation (C-257 to T) in gyrA gene, as well as 107 clinical isolates of various bacterial species, including 29 C. jejuni isolates. This study indicates that MMAMA-PCR is a promising assay for the rapid identification of C. jejuni with a specific mutation in gyrA gene, responsible for the resistance to fluoroquinolones.
authors: 1 Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Veterinary Medicine, China Agricultural University , Beijing, China .|2 Key Laboratory of Detection for Veterinary Drug Residue and Illegal Additive, MOA, College of Veterinary Medicine, China Agricultural University , Beijing, China .
authors: Mingquan
authors: M
authors: Cui
authors: 1 Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Veterinary Medicine, China Agricultural University , Beijing, China .|3 China Institute of Veterinary Drug Control , Beijing, China .
authors: Chenbin
authors: C
authors: Wu
authors: 1 Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Veterinary Medicine, China Agricultural University , Beijing, China .|2 Key Laboratory of Detection for Veterinary Drug Residue and Illegal Additive, MOA, College of Veterinary Medicine, China Agricultural University , Beijing, China .
authors: Peng
authors: P
authors: Zhang
authors: 1 Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Veterinary Medicine, China Agricultural University , Beijing, China .|2 Key Laboratory of Detection for Veterinary Drug Residue and Illegal Additive, MOA, College of Veterinary Medicine, China Agricultural University , Beijing, China .
authors: Congming
authors: C
authors: Wu
mesh_terms: D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002169:Campylobacter Infections / Q000382:microbiology; D016123:Campylobacter jejuni / Q000145:classification / Q000187:drug effects* / Q000302:isolation & purification* / Q000378:metabolism; D002681:China; D027081:DNA Gyrase / Q000235:genetics* / Q000378:metabolism; D024901:Drug Resistance, Multiple, Bacterial; D004751:Enteritis / Q000382:microbiology; D024841:Fluoroquinolones / Q000494:pharmacology*; D005517:Foodborne Diseases / Q000382:microbiology; D005798:Genes, Bacterial; D006801:Humans; D057230:Limit of Detection; D008826:Microbial Sensitivity Tests; D058889:Molecular Typing / Q000379:methods; D060885:Multiplex Polymerase Chain Reaction; D009692:Nucleic Acid Heteroduplexes; D017354:Point Mutation; D012329:RNA, Bacterial / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D015203:Reproducibility of Results
chemical_list: D001426:Bacterial Proteins; D024841:Fluoroquinolones; D009692:Nucleic Acid Heteroduplexes; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D000581:Amidohydrolases; C019672:aminoacylase I; D027081:DNA Gyrase
keywords: Campylobacter jejuni; fluoroquinolones resistance; gyrA mutation; rapid identification