Detection of blaKPC and blaNDM genes by duplex PCR with lateral flow dipsticks from sterile body fluid samples.

ABSTRACT

Duplex polymerase chain reaction with lateral flow dipsticks (duplex PCR-LFD) was developed for the simultaneous detection of beta-lactamase Klebsiella pneumoniae carbapenemase (blaKPC ) and beta-lactamase New Dehli metallo-beta-lactamase (blaNDM ) genes in body fluid samples. This method was validated using well-characterized isolates. The assessment of the specificity of duplex PCR-LFD showed that there was no cross-reactivity with other targets. The detection limit of the duplex PCR-LFD assay was 20 CFU per ml for blaKPC and blaNDM . Among 177 sterile body fluid samples tested by the duplex PCR-LFD assay, 40 were blaKPC -positive and five were blaNDM -positive. The results obtained from 122 corresponding Gram-negative bacteria which were isolated from these clinical samples and tested by duplex PCR-LFD assay showed that there were 37 strains carrying blaKPC genes in 40 blaKPC -positive samples and three strains carrying blaNDM genes in five blaNDM -positive samples. Statistical analysis indicated that there was no significant difference between the direct detection of blaKPC and blaNDM genes in clinical sterile body fluid samples and their corresponding clinical isolates. Therefore, duplex PCR-LFD can be effective for the simultaneous detection of blaKPC and blaNDM in clinical isolates and directly from clinical samples, which may be helpful for the administration of appropriate antimicrobial treatment.