A multiplex assay for characterization of antimicrobial resistance in Neisseria gonorrhoeae using multi-PCR coupled with mass spectrometry.

The Journal of Antimicrobial Chemotherapy
Yamei LiJunping Peng

Abstract

Complicated mechanisms and variable determinants related to drug resistance pose a major challenge to obtain comprehensive antimicrobial resistance (AMR) profiles of Neisseria gonorrhoeae. Meanwhile, cephalosporin-resistant mosaic penA alleles have been reported worldwide. Therefore, it is urgent to monitor the expansion of cephalosporin-resistant mosaic penA alleles. To develop a comprehensive high-throughput method to efficiently screen AMR determinants. We developed a method based on multiplex PCR with MALDI-TOF MS, which can simultaneously screen for 24 mutations associated with multiple antimicrobial agents in 19 gonococcal AMR loci (NG-AMR-MS). The performance of the NG-AMR-MS method was assessed by testing 454 N. gonorrhoeae isolates with known MICs of six antibiotics, eight non-gonococcal Neisseria strains, 214 clinical samples and three plasmids with a confirmed mosaic penA allele. The results show that NG-AMR-MS had a specificity of 100% with a sensitivity as low as 10 copies per reaction (except for PorB A121D/N/G, 100 copies per reaction). For clinical samples with gonococcal load >5 copies/μL, the method can accurately identify 20 AMR mutations. In addition, the method successfully detected specific cephalosporin-r...Continue Reading

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