The a group of strains representing all phylogroups

The objective of the present study was to analyze the diversity structure of E. coli & K. pneumonia strains isolated from the River Yamuna & other water bodies traversing though the National Capital Territory of Delhi (India) and understand the molecular mechanisms underlying ?-lactamases mediated antibiotic resistance as well as co-resistance to one of the other important classes of antibiotics viz (fluro) quinolones, if any.

·         Water samples were collected from different Fifteen geographical sites of Delhi.

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·         The results of this study show that E. coli is more affected by the acquisition and

 stable integration of resistance genes in an aquatic environment than Klebsiella. 

·         Total Ninety-six E. coli & Eighty-four K. pneumoniae isolates were obtained from different-2 Geographical sites, Seven different -2 sites of Yamuna River, Three different-2 sites of Sewage water, Two different -2 sites of Ground water,  One sites of Canal water & Two different -2 sites of Other water bodies Faridabad waste water and Hindon River.

·          The isolated E. coli & K. pneumoniae strains when tested for their tolerance to AMR exhibited similar resistant patterns both in liquid and solid medium.

·         All the biochemical tests were same for different environmental strains of E. coli and K. pneumoniae. From this collection of E. coli & K. Pneumonia a group of strains representing all phylogroups was used further studies.

·         Total Ninety-six E. coli & Eighty-four K. pneumoniae isolates were obtained from different water samples, the isolated E. coli & K. pneumoniae strains when tested for their tolerance to AMR exhibited similar resistant patterns both in liquid and solid medium.

·         All the biochemical tests were same for different environmental strains of E. coli and K. pneumoniae.

·         From this collection of E. coli & K. Pneumonia a group of strains representing all phylogroups was used further studies. The Forty-eight E. coli & Forty-two K. pneumonia  strains were CONFIRMED by sequencing of their 16s ribosomal RNA (rRNA genes).

·         Correlation between antibiotic and AMR in collected AMR strains was examined, resistance pattern towards Ten different antibiotics i.e. Ceftazidime (30mcg), cefotaxime (30mcg), Cefpodoxim (10mcg), Azithromycin (15mcg), Metronidazole (4mcg), (Meropenem (10mcg), Amikacin (30mcg), Ampicilin (10mcg),  Ceftriaxone (30mcg), and  Ciprofloxacin (5mcg).

·         All the selected strains were resistant in E. coli strains almost Ampicillin (100%), Cefotaxime (72.91%), Ceftriaxone (70.83%), Ciprofloxacin (100%), Cefpodoxim(56.25%), Ceftazidime (83.33%), Metronidazole (70.83%), Meropenem (31.25%), Azithromycin (60.41%) and  Amikacin (12.5%) & K. pneumonia strains to Ampicillin (100 %), Cefotaxime (76.19 %), Ceftriaxone (80.95 %), Ciprofloxacin (100%), Cefpodoxim (88.095 %), Ceftazidime (85.71 %), Metronidazole (78.57%), Meropenem (50 %), Azithromycin (71.42 %) and Amikacin (23.80 %) & Ampicillin (100%).

·         Phenotypic Detection of ESBL to Cefotaxime, Ceftazidime, Cefepime Cefepirome alone and with Clavulanic acid. Almost all 76/90 (84%) of the E. coli (46) & K. pneumonia (30) were  showed ESBL & PMQR Positive.

·         We were detect the presence of protein in the strains of E. coli & K. pneumonia.

·         All the seventy six strains were screened for the presence of Protein. Following protein isolation, all the selected strains showed the presence of protein when subjected on SDS-PAGE.

·         All the seventy six strains were screened for the presence of plasmid. Following plasmid DNA isolation, all the selected strains showed the presence of plasmid when visualized on 1% agarose gel.

·         Our main effort was to characterize ESBL & PMQR genes from different isolates so that we could find out the divergence between them and then they can be further used for bioremediation of drug. Having established the plasmid borne nature of ESBL & PMQR gene in the selected strains, PCR amplification of

TEM, CTX-M, qnrS and aac (6′)-lb-cr)  genes were carried using specific primers, the expected length of PCR products for TEM gene corresponding to 780 bp was obtained from E. coli (46) 65.21 % & K. pneumonia (30) 66.66 % strains, CTX-M gene corresponding to 530 bp was obtained from E. coli (46) 34.7 % & K. pneumonia (30) 36.66 % strains, qnrS gene corresponding to 456bp was obtained from E. coli (46) 30.43 % and K. pneumonia (30) 19.56 % wild type isolated strains and aac(6′)-lb-cr) gene corresponding to 482bp FQRB respectively was obtained from only wild type isolated E. coli (46) 19.56 % & K. pneumonia (30) 23.33 % when checked on 1% agarose gel. Other PMQR gene such as qnrC were detected less frequently. Additionally no qnrD, gene was observed in this study.

·         DNA sequencing was performed in order to check the diversity and distribution of ESBL & PMQR genes. In the present study we found that all the nucleotide sequences for CTX-M gene fragments from selected strains were showing a great diversity (74-84%) but the sequences for all the selected TEM gene fragments showed a high level of similarity (upto 99%) and qnrS gene strain showed upto (94%) with other already reported sequences of that gene. 

·         A phylogenetic tree showed a different data with other already reported sequences of that gene.