In this study, 40 strains of gram negative rods were used to identify and detect CPE, CRPA, and CRAB, which are threats to public health, and to identify and detect quickly and accurately.
Phenotypic tests, molecular diagnostics of the blaKPC blaNDM blaIMP blaVIM blaOXA-48-like genes in the carbapenemase genotypes were performed to determine the distribution patterns of related genes.
As a result of the phenotypic test, MHT was positive in 18 of 40 strains, CIT was positive in 7 strains of meropenem+PBA and 15 strains in meropenem+EDTA were positive.
PCR results showed that the amplified products were identified in 18 strains of blaKPC, 14 strains of blaNDM, 2 strains of blaIMP, and 1 strain of blaVIM, and no amplified products were found in blaVIM blaOXA-48-like. Also, we confirmed 8 strains of blaKPC+blaNDM, 1 strain of blaKPC+blaIMP, and 1 strain of blaNDM+blaIMP+blaVIM. The amplified gene was subjected to sequence analysis, and homology of 99% or more was confirmed for all genes used for the analysis. It was confirmed that the melting curve analysis result using real-time PCR was 100% consistent with the PCR result.
As a result of comparative analysis of phenotypic analysis results and PCR results, 61.1% false negatives for blaKPC, 6.6% false positives for blaNDM, and 100% false positives for blaOXA-48-like. were confirmed. In phenotypic test, IMP and VIM genes could not be identified, and non-carbapenemase was found to have a 37.5% discrepancy.
In conclusion, rapid and specific melting curve analysis using real-time PCR is the key to gene identification through early diagnosis of CPE, CRPA, and CRAB can improve monitoring, diagnosis, and treatment of CRE, which is a public health threat with limited treatment options, high mortality and morbidity, and effective antibacterial treatment and control timely infections to prevent transmission.