Predictability of Phenotype in Relation to Common β-Lactam Resistance Mechanisms in Escherichia coli and Klebsiella pneumoniae

Abstract

cThe minimal concentration of antibiotic required to inhibit the growth of different isolates of a given species with no acquired resistance mechanisms has a normal distribution. We have previously shown that the presence or absence of transmissible antibiotic resistance genes has excellent predictive power for phenotype. In this study, we analyzed the distribution of six ␤-lactam antibiotic susceptibility phenotypes associated with commonly acquired resistance genes in Enterobacteriaceae in Sydney, Australia. Escherichia coli (n ‫؍‬ 200) and Klebsiella pneumoniae (n ‫؍‬ 178) clinical isolates, with relevant transmissible resistance genes (bla TEM , n ‫؍‬ 33; plasmid AmpC, n ‫؍‬ 69; extended-spectrum ␤-lactamase [ESBL], n ‫؍‬ 116; and carbapenemase, n ‫؍‬ 100), were characterized. A group of 60 isolates with no phenotypic resistance to any antibiotics tested and carrying none of the important ␤-lactamase genes served as comparators. The MICs for all drug-bacterium combinations had a normal distribution, varying only in the presence of additional genes relevant to the phenotype or, for ertapenem resistance in K. pneumoniae, with a loss or change in the outer membrane porin protein OmpK36. We demonstrated mutations in ompK36 or absence of OmpK36 in all isolates in which reduced susceptibility to ertapenem (MIC, >1 mg/liter) was evident. Ertapenem nonsusceptibility in K. pneumoniae was most common in the context of an OmpK36 variant with an ESBL or AmpC gene. Surveillance strategies to define appropriate antimicrobial therapies should include genotype-phenotype relationships for all major transmissible resistance genes and the characterization of mutations in relevant porins in organisms, like K. pneumoniae. E scherichia coli and Klebsiella pneumoniae are among the most important pathogens in community and hospital settings, and their increasing resistance to extended-spectrum (third-and fourth-generation) cephalosporin and carbapenem antibiotics is a major health threat. In Gram-negative bacteria, such as these, a variety of mechanisms may confer ␤-lactam resistance (1), but the spread of transmissible genes encoding extended-spectrum ␤-lactamases (ESBLs) (2), plasmid-mediated AmpC ␤-lactamases (pAmpCs) (3), and carbapenemases (4) is particularly significant. Carbapenems have been the antibiotics of choice for treating ESBLs and other multidrug-resistant strains, but carbapenem resistance is increasingly common (5). This is mostly attributed to the production of specific carbapenemases (6), but the expression of AmpC or ESBL enzymes in isolates with altered outer membrane porins is also associated with decreased susceptibility to carbapenems (7-9). Mutations in major outer membrane porins may be required for clinically relevant carbapenem resistance in K. pneumoniae isolates expressing carbapenemases, such as KPC and OXA-48-like enzymes, which rarely elicit clinically important antibiotic resistance in E. coli (6).The clinical definitions of antibiotic susceptibility are developed by the collection and analysis of...

The Inc FII Plasmid and its Contribution in the Transmission of blaNDM-1 and blaKPC-2 in Klebsiella pneumoniae in Egypt

“…Our results showed the predominance of both Inc FIIK and FII plasmids among the isolates and their transconjugants carrying bla KPC2 and/or bla NDM1 , suggesting that these types of plasmids mediate horizontal transmission and contribute to the dissemination of bla KPC2 and/or bla NDM1 in the environment of the Egyptian hospitals, which represents a great challenge and an important factor in the dissemination of resistance and treatment failure in cases of severe infections. Agyekum et al [39] reported that FIIK is the most common plasmid replicon found in K. pneumoniae. Another study done by Al-Marzooq et al [40] reported that the most common plasmid replicons were IncR and IncL/M.…”

Host adaptation and convergent evolution increases antibiotic resistance without loss of virulence in a major human pathogen

“…β-lactam antibiotics are among the most commonly prescribed for severe infections [80, 81] and the emergence of β-lactam resistance in K. pneumoniae has become a global health threat [82, 83]. In general, E. coli and K. pneumoniae carrying transmissible β-lactam resistance genes have predictable and normally distributed β-lactam MICs [21] but carbapenem MICs in K. pneumonia e are bimodally distributed with higher MICs correlating with OmpK36 defects [21]. OmpK36 loss or mutation is not uncommonly reported in highly resistant clinical isolates producing KPC, ESBL or AmpC β-lactamases [20, 84, 85].…”

“…Isolates were grown overnight in MHB. Bacteria were disrupted by sonication and outer membrane porins (OMPs) isolated with sarcosyl (Sigma-Aldrich, St. Louis, MO, USA), as previously described [21, 38]. Samples were boiled, analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (12% separating gels), and stained with Imperial protein stain (Thermo Scientific, Rockford, IL, USA), following the manufacturer’s instructions.…”

“…K. pneumoniae expresses two major nonspecific porins (OmpK35 and OmpK36) through which nutrients and other hydrophilic molecules such as β-lactams diffuse into the cell [8, 9]. The expression of these two major porins in K. pneumoniae are strongly linked with β-lactam susceptibility [6, 7] and strains lacking both porins exhibit high levels of resistance [10]. K. pneumoniae is commonly present in the human gut [1] but also grows in low-nutrient and low-osmolarity conditions, with decreased expression of the ‘osmoporin’, OmpK36, and increased expression of the ‘matrix porin’, OmpK35, which has the greater general permeability.…”

Antibiotic resistant Escherichia coli in southeastern Australian pig herds and implications for surveillance

“…Isolates resistant to 3GC or aminoglycosides were screened by multiplex PCR/reverse line blot (mPCR/RLB) (Agyekum et al., ; Ginn et al., ), to detect genes conferring resistance to β‐lactams ( bla TEM, bla SHV, bla CTX‐M, bla KPC, bla VEB, bla IMP, bla VIM, bla NDM, bla CMY‐2‐like, bla DHA, bla ACT, bla MIR, bla OXA‐10‐like, bla OXA‐30‐like, bla OXA‐48‐like genes) and aminoglycosides ( aac(3)‐II , aacA4 , aadB , armA , rmtB , rmtC , rmtF ). PCR using published primers (Ellem, Partridge, & Iredell, ; Perez‐Perez & Hanson, ; Woodford, Fagan, & Ellington, ) and sequencing was used to confirm resistance genes and determine the allele present.…”