Detection of clinically relevant carbapenemase encoding genes in carbapenem-resistant Enterobacter cloacae complex and Klebsiella pneumoniae isolated from farmed freshwater fish.

AIMS: The present study was aimed to detect clinically relevant carbapenemase encoding genes in carbapenem-resistant Enterobacter cloacae complex (CR-ECC), Klebsiella pneumoniae (CR-KP), and Serratia plymuthica (CR-SP) isolated from farmed freshwater fish. METHODS AND RESULTS: Out of 243 spatially diverse freshwater fish samples analysed, 5.3% were contaminated with CR-ECC, 1.6% with CR-KP, and 0.4% with CR-SP. The CR-ECC was further identified as E. asburiae (38.5%), E. mori (23.1%), E. cloacae (15.4%), E. hormaechei (15.4%), and E. kobei (7.7%) by 16S rRNA gene sequencing. The CR-ECC were resistant to carbapenems and cefoxitin, whereas CR-KP and CR-SP were multi-drug resistant (MDR). The CR-ECC harboured the carbapenemase gene blaIMI alone or in combination with blaTEM, blaEBC, blaCIT, blaACC, and tet(E). Whereas, CR-KP harboured carbapenemase gene, blaNDM-5 along with blaOXA-48, blaSHV, blaOXA-1, blaCTX-M-15, tet(A), sul1, and qnrB. No carbapenemase-encoding genes were detected in CR-SP. The MLST analysis showed that CR-KP belonged to ST231 and ST1561 lineages, while CR-ECC did not show exact match with any reported STs. The plasmid replicons predominantly detected were IncF and IncI1. Broth mating assays of CR-KP and CR-ECC with recipient Escherichia coli J53 indicated that blaNDM-5 was transferable but not blaIMI. CONCLUSION: This study highlights the low-level contamination of carbapenem-resistant Enterobacterales (CRE) harbouring clinically relevant carbapenemase-encoding genes in farmed freshwater fish from India. The CR-ECC of fish origin did not show the potential to spread carbapenem resistance.

Molecular characterization of carbapenem resistant E. coli of fish origin reveals the dissemination of NDM-5 in freshwater aquaculture environment by the high risk clone ST167 and ST361.

Aquatic environment can act as reservoir and disseminator of antimicrobial resistance and resistant pathogens. Novel high-risk carbapenem resistant E. coli (CREC) are continuously emerging worldwide; however, the occurrence of CREC in freshwater aquaculture environment is largely unexplored. To fill this gap, large scale sampling of freshwater pond sites and retail fish markets was done between Oct 2020 and Oct 2021 to investigate the CREC contamination in fish. The frequency of CREC contamination in the freshwater fish was 6.99% (95% CI: 3.78-10.20%). All the isolates were MDR and harbored carbapenemase encoding gene, blaNDM-5 along with other antimicrobial resistance genes (ARGs), blaTEM (64.7%), blaCTX-M-15 (35.3%), blaOXA-1 (5.9%), tet(A) (100%), sul1 (94.1%), qnrS (82.3%), cat1 (35.3%), and cat2 (23.5%). The isolates belonged to phylogroup C and showed low virulence gene profile. ERIC-PCR grouped the isolates into five clusters (I-V). The isolates of clusters I, II, and III were identified as ST167 (76.4%) and of cluster IV as ST361 (17.6%). This is the first report documenting the contamination of NDM-5 producing E. coli ST167 and ST361 of clinical/livestock lineage in freshwater fish from India. The blaNDM-5 was significantly associated with ARGs, tet(A), and sul1; and plasmid replicons, IncF, IncI1, and IncP, signifying the presence of blaNDM-5 and associated ARGs on these transferable plasmids. These findings were validated by the successful conjugal transfer of blaNDM-5 and associated ARGs into non-CREC strain (J53). Our study highlights the ability of CREC to disseminate antimicrobial resistance which has health implications and environmental concerns.

Sperm motility in the fishes of pesticide exposed and from polluted rivers of Gomti and Ganga of north India.

Investigation of lethal dose of gamma-HCH (gamma isomer of hexachlorocyclohexane), DDT (dichlorodiphenyltrichloroethane) and chlorpyrifos on spermatozoa motility after 40 days exposure in catfish, Heteropneustes fossilis was done under laboratory conditions. The sperm motility was done in the fishes captured from unpolluted ponds of Gujartal considering as reference site and polluted rivers Gomti and Ganga of north India at pre-spermiating stage. Results indicate that 1ppm of gamma-HCH, DDT and chlorpyrifos was lethal dose on sperm motility. The motility of spermatozoa decreased in insecticide exposed fish as well as in the fishes of polluted rivers when compared with their respective controls. The sperm motility was highest at 1:2000 (testicular milt: extender) dilution and duration of sperm motility was 90s after post-activation. The duration of motility also declined in the fishes captured from polluted rivers when compared with the same species captured from the reference site. It is concluded that the insecticides decrease the sperm motility and its duration in exposed fish as well as in the captured fishes from polluted rivers causing the decline in fish population of riverine systems due to influence of xenobiotics on the endocrine system.