Quinolone co-resistance in ESBL- or AmpC-producing Escherichia coli from an Indian urban aquatic environment and their public health implications.

Quinolone and ß-lactam antibiotics constitute major mainstay of treatment against infections caused by pathogenic Escherichia coli. Presence of E. coli strains expressing co-resistance to both these antibiotic classes in urban aquatic environments which are consistently being used for various anthropogenic activities represents a serious public health concern. From a heterogeneous collection of 61 E. coli strains isolated from the river Yamuna traversing through the National Capital Territory of Delhi (India), those harboring blaCTX-M-15 (n = 10) or blaCMY-42 (n = 2) were investigated for co-resistance to quinolones and the molecular mechanisms thereof. Resistance was primarily attributed to amino acid substitutions in the quinolone resistance-determining regions (QRDRs) of GyrA (S83L ± D87N) and ParC (S80I ± E84K). One of the E. coli strains, viz., IPE, also carried substitutions in GyrB and ParE at positions Ser492→Asn and Ser458→Ala, respectively. The phenotypically susceptible strains nevertheless carried plasmid-mediated quinolone resistance (PMQR) gene, viz., qnrS, which showed co-transfer to the recipient quinolone-sensitive E. coli J53 along with the genes encoding ß-lactamases and led to increase in minimal inhibitory concentrations of quinolone antibiotics. To the best of our knowledge, this represents first report of molecular characterization of quinolone co-resistance in E. coli harboring genes for ESBLs or AmpC ß-lactamases from a natural aquatic environment of India. The study warrants true appreciation of the potential of urban aquatic environments in the emergence and spread of multi-drug resistance and underscores the need to characterize resistance genetic elements vis-à-vis their public health implications, irrespective of apparent phenotypic resistance.

Analysis of iron acquisition and storage-related genes in clinical and non-clinical strains of Yersinia enterocolitica biovar 1A.

Possession of mechanisms for iron acquisition and its storage enhances the ability of the bacteria to survive in the iron-limiting environment of the host. In this study, 81 strains of Yersinia enterocolitica biovar 1A isolated from various clinical (n = 51) and non-clinical (n = 30) sources were investigated for the presence of the genes related to iron acquisition and storage. Important genes which were present in more than 85% of the strains included hasA, foxA, bfr, bfd, ftnA, and hmsT as well as the fhuCDB, fepBDGCfesfepA, feoAB, yfuABCD, hemPRSTUV, and hmsHFRS gene clusters. Majority of these genes is being reported for the first time in biovar 1A strains and showed significant homology with genes present in the known pathogenic biovars of Y. enterocolitica. However, no significant difference was observed in the distribution of iron acquisition and storage-related genes among clinical and non-clinical biovar 1A strains. Thus, it may be suggested that the presence of iron acquisition and storage-related genes per se might not be responsible for the supposedly better ability of clinical biovar 1A strains to cause infections in humans. However, in the backdrop of this data, the need to undertake functional studies are highly recommended.

Proteomic analysis of Yersinia enterocolitica biovar 1A under iron-rich and iron-poor conditions indicate existence of efficiently regulated mechanisms of iron homeostasis.

The pathogenicity of Yersinia enterocolitica biovar 1A strains is controversial as these lack most of the known virulence factors. Acquisition of iron and presence of well-regulated iron homeostasis in bacteria represents an important virulence trait. Differential abundance of proteins was examined under iron-rich and iron-poor conditions in a clinical Y. enterocolitica biovar 1A strain IP27407. Whole cell protein profiles were analysed by 2D gel electrophoresis (2D-GE). Following statistical and MALDI-TOF MS analyses, 28 differentially abundant proteins were identified. Significant iron-responsive changes were observed in the proteins involved in iron acquisition or storage namely, hemin receptor (HemR), periplasmic Fe(2+) transport protein (Tpd), periplasmic chelated iron-binding protein (YfeA) and bacterioferritin (Bfr). Quantitative real-time PCR (qRT-PCR) of eight mRNA transcripts revalidated the differential protein abundance. In silico analysis of iron homeostasis mediated by the bacterioferritin and bacterioferritin-associated ferredoxin (Bfr-Bfd) complex suggested two pathways for the release of reserve iron which might be operating under conditions of different iron availability. The study, for the first time, showed the existence of highly competent iron homeostasis mechanisms in Y. enterocolitica biovar 1A and identified the key proteins involved thereof. Such mechanisms might have implications for the pathogenicity of Y. enterocolitica biovar 1A strains. BIOLOGICAL SIGNIFICANCE: Although, a few studies have identified the differentially abundant bacterial proteins in response to iron starvation, little information is available in this regard for Y. enterocolitica (especially, the biovar 1A strains). In the present study, differential abundance of several proteins was identified under iron-rich and iron-poor conditions by 2D-GE and MALDI-TOF/MS analysis. These included proteins which may not only be directly implicated in iron acquisition or storage but also play crucial role in cellular metabolism. Given the absence of most known virulence factors in Y. enterocolitica biovar 1A strains, demonstration of well-regulated mechanisms for efficient iron homeostasis constitutes an important observation. The proteins, as identified in the present study, provide useful insights to further unravel the potential pathogenicity of the biovar 1A strains.

Distribution and molecular characterization of genes encoding CTX-M and AmpC ß-lactamases in Escherichia coli isolated from an Indian urban aquatic environment.

Aquatic environments harboring antibiotic resistant Escherichia coli constitute an important public health concern. Thus, it is important to characterize the resistance genetic elements of waterborne E. coli. It is also important to identify the predominant clonal groups/phylogroups represented by resistant strains to understand the epidemiology of antibiotic resistant E. coli in natural environments, and to identify the role of well-established genotypes in the spread of resistance in a particular geographical area through natural environments. In the present investigation, E. coli strains (n=126) isolated from various points along the river Yamuna traversing through the National Capital Territory of Delhi (India) were grouped phylogenetically. A collection of 61 strains representing all phylogroups was investigated for extended-spectrum ß-lactamase (ESBL) and AmpC production. blaTEM, blaSHV and blaCTX-M genes were detected and analyzed, promoter/attenuator mutations associated with chromosomally-mediated AmpC overexpression were identified, and plasmid-mediated ampC was determined. blaTEM was the most widespread (100%) gene followed by bla(CTX-M) (16%), and plasmid-mediated ampC (3%). bla(CTX-M-15) and bla(CMY-42) were identified as the genes encoding CTX-M type ESBL and CIT type AmpC ß-lactamases, respectively. CTX-M-15 ESBL phenotype was most common in phylogroup D (50%), followed by phylogroups B1 (30%), and A (20%). E. coli that produce plasmid-mediated AmpC were rare and present only in phylogroup D. Presence of multi ß-lactam resistance, bla(CTX-M-15) and bla(CMY-42) in waterborne E. coli belonging to virulence-associated phylogroup D highlights the need for routine surveillance of resistance determinants in aquatic environments. This is also the first report for the presence of bla(CMY-42) in waterborne E. coli.