Multiplex PCR scheme for variant plasmid mediated class C β-lactamase typing

J Clin Lab Anal. 2018 Mar;32(3):e22298. doi: 10.1002/jcla.22298. Epub 2017 Jul 18.

Abstract

Background: An increasing of prevalence and diversification of plasmid-mediated AmpC (pAmpC) has been emerged worldwide. The incidence of pAmpC resulted in increasing β-lactamase production and conferred resistance to almost all β-lactam antibiotics excluding carbapenems. The lack of standard method for pAmpC identification and classification exert a challenge in epidemiological surveillance and infection control practices.

Methods: A robust, single tube multiplex PCR has been developed to classify six different pAmpC groups including CIT (CMY-2 like, LAT and CFE), ECB (ACT, MIR), MOX & CMY-1 like, DHA, ACC, and FOX. The developed method was optimized and validated by testing of sensitivity and specificity.

Results: Developed method can detect crude extracted DNA template at nano-scale (2.5 ηg) and has high discriminatory power as compared to phenotypic and commercial genotypic method.

Conclusion: The developed method can be utilized for tracking the changes of clinically important resistance patterns and further investigation of occurrence and distribution of plasmid-mediated AmpC types.

Keywords: Enterobacteriaceae; Plasmid-mediated AmpC β-lactamases; cefoxitin resistance; genotypic method; multiplex PCR.

MeSH terms

  • Enterobacteriaceae / enzymology
  • Enterobacteriaceae / genetics*
  • Molecular Typing / methods*
  • Multiplex Polymerase Chain Reaction / methods*
  • Phenotype
  • Plasmids / genetics*
  • Reproducibility of Results
  • Sensitivity and Specificity
  • beta-Lactamases / genetics*

Substances

  • beta-Lactamases