Genotypic to Phenotypic Resistance Discrepancies Identified Involving β-Lactamase Genes, bla KPC, bla IMP, bla NDM-1, and bla VIM in Uropathogenic Klebsiella pneumoniae

Abstract

Introduction: Klebsiella pneumoniae carbapenemase (KPC) belongs to the Group-A β-lactamases that incorporate serine at their active site and hydrolyze various penicillins, cephalosporins, and carbapenems. Metallo-beta-lactamases (MBLs) are group-B enzymes that contain one or two essential zinc ions in the active sites and hydrolyze almost all clinically available β-lactam antibiotics. Klebsiella pneumoniae remains the pathogen with the most antimicrobial resistance to KPC and MBLs.

Methods: This research investigated the blaKPC, and MBL genes, namely, blaIMP, blaVIM, and blaNDM-1 and their phenotypic resistance to K. pneumoniae isolated from urinary tract infections (UTI) in Bangladesh. Isolated UTI K. pneumoniae were identified by API-20E and 16s rDNA gene analysis. Their phenotypic antimicrobial resistance was examined by the Kirby-Bauer disc diffusion method, followed by minimal inhibitory concentration (MIC) determination. blaKPC, blaIMP, blaNDM-1, and blaVIM genes were evaluated by polymerase chain reactions (PCR) and confirmed by sequencing.

Results: Fifty-eight K. pneumoniae were identified from 142 acute UTI cases. Their phenotypic resistance to amoxycillin-clavulanic acid, cephalexin, cefuroxime, ceftriaxone, and imipenem were 98.3%, 100%, 96.5%, 91.4%, 75.1%, respectively. Over half (31/58) of the isolates contained either blaKPC or one of the MBL genes. Individual prevalence of blaKPC, blaIMP, blaNDM-1, and blaVIM were 15.5% (9), 10.3% (6), 22.4% (13), and 19% (11), respectively. Of these, eight isolates (25.8%, 8/31) were found to have two genes in four different combinations. The co-existence of the ESBL genes generated more resistance than each one individually. Some isolates appeared phenotypically susceptible to imipenem in the presence of blaKPC, blaIMP, blaVIM, and blaNDM-1 genes, singly or in combination.

Conclusion: The discrepancy of genotype and phenotype resistance has significant consequences for clinical bacteriology, precision in diagnosis, the prudent selection of antimicrobials, and rational prescribing. Heterogeneous phenotypes of antimicrobial susceptibility testing should be taken seriously to avoid inappropriate diagnostic and therapeutic decisions.

Keywords: Bangladesh; Klebsiella pneumoniae; blaIMP; blaKPC; blaNDM-1; blaVIM; co-resistance; heteroresistance; urinary tract infections.

Figure 1

Gender and age distribution of K. pneumonia-positive and -negative UTI patients. Frequency distribution of urine culture with K. pneumonia positive (+ve) and negative (-ve) according to sex (male and female) and age group (1–20 YO, 21–40 YO and more than 40 YO) (n=142, Years Old = YO).

Figure 2

Impact of Extended Spectrum β-lactamase (ESBL) genes on the phenotypic susceptibilities of β-lactam antibiotics. The comparative susceptibilities of the ESBL-positive and -negative isolates were evaluated against selected b-lactam antibiotics, namely Amoxycillin+Clavulinic acid (AMC 30 µg), Cefepime (FEP 30 µg), Cefuroxime Sodium (CXM 30 µg), Cephalexin (CL 30 µg), Ceftriaxone (CRO 30 µg), Imipenem (IMP 10 µg). (A). The Y-axis values of black bars indicate the percentage of ESBL gene-carrying isolates showing as susceptible against respective antibiotics shown on the X-axis. Similarly, the white bars illustrate the percentages of susceptible isolates that do not carry the respective ESBL genes. Susceptibilities of (A) blaKPC-positive (n=9) and -negative (n=49), (B) blaIMP-positive (n=6) and -negative (n=52), (C) blaVIM-positive (n=11) and -negative (n=47), and (D) blaNDM1-positive (n=13) and -negative (n=45), isolates are shown separately.

Figure 3

Overlapping Extended Spectrum β-lactamase (ESBL) genes. The Venn-diagram shows the mutual prevalence of ESBL genes, such as blaKPC, blaIMP, blaVIM, and blaNDM-1 in the uropathogen Klebsiella pneumoniae. Each circle is labeled with the respective gene-name, and the number in the bracket indicates the total count of isolates positive for the specific gene. Numbers in the non-overlapping region show the count of isolates carrying respective single-type ESBL genes. Numerates in the overlapping region indicate the isolate number carrying the respective genes mutually.