A rapid low-cost real-time PCR for the detection of Klebsiella pneumonia carbapenemase genes.

Department of Laboratory Medicine, Beijing Tongren Hospital, Capital Medical University, No1, Dongjiaominxiang Road, Dongcheng District, Beijing, 100730, China.
Annals of Clinical Microbiology and Antimicrobials (Impact Factor: 2.19). 04/2012; 11(1):9. DOI: 10.1186/1476-0711-11-9
Source: PubMed


Klebsiella pneumonia carbapenemases (KPCs) are able to hydrolyze the carbapenems, which cause many bacteria resistance to multiple classes of antibiotics, so the rapid dissemination of KPCs is worrisome. Laboratory identification of KPCs-harboring clinical isolates would be a key to limit the spread of the bacteria. This study would evaluate a rapid low-cost real-time PCR assay to detect KPCs.
Real-time PCR assay based on SYBR GreenIwas designed to amplify a 106 bp product of the blaKPC gene from the 159 clinical Gram-negative isolates resistant to several classes of -lactam antibiotics through antimicrobial susceptibility testing. We confirmed the results of real-time PCR assay by the conventional PCR-sequencing. At the same time, KPCs of these clinical isolates were detected by the modified Hodge test (MHT). Then we compared the results of real-time PCR assay with those of MHT from the sensitivity and specificity. Moreover, we evaluated the sensitivity of the real-time PCR assay.
The sensitivity and specificity of the results of the real-time PCR assay compared with those of MHT was 29/29(100%) and 130/130(100%), respectively. The results of the real-time PCR and the MHT were strongly consistent (Exact Sig. (2-tailed) =1. 000; McNemar test). The real-time PCR detection limit was about 0.8 cfu using clinical isolates.
The real-time PCR assay could rapidly and accurately detect KPCs -harboring strains with high analytical sensitivity and specificity.

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    • "al - time PCR to detect KPC from 120 stool and 128 nasal swabs samples with an LOD of 80 CFU ml À1 , and attained a 100% and 99% sensitivity and specificity respectively ( Table 1 ) ( Wang et al . 2012a , b ) . This assay could not differentiate between various KPC variants , although it was fast ( <2 h ) . Subsequent studies by the same authors ( Wang et al . 2012b ) with 159 clinical isolates ( Kl . pneumoniae , 9 K . oxytoca , E . coli , Enterobacter cloacae , Serratia marces - cens , Proteus mirabilis , Ac . baumannii and Ps . aeruginosa ) were , however , 100% sensitive and specific , with an LOD of 0Á8 CFU ml À1 and a turnaround time of ≤4 h . The SYBR Green is a cheaper and more sensitive ( "
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