title: Functional Genome Screening to Elucidate the Colistin Resistance Mechanism.
abstract: Antibiogram profile of 1590 clinical bacterial isolates based on thirteen different antimicrobial compounds showed that 1.6% of the bacterial isolates are multidrug resistant. Distribution pattern based on 16S rRNA sequence analysis showed that Pseudomonas aeruginosa constituted the largest group (83.6%) followed by Burkholderia pseudomallei sp. A191 (5.17%), Staphylococcus sp. A261 (3.45%). Among the various antibiotics used, colistin appeared to be the most effective against the Gram negative bacteria. Burkholderia pseudomallei sp. A191 and Pseudomonas aeruginosa sp. A111 showed resistance to 1500 μg/ml and 750 μg/ml of colistin respectively which constitutes 7.7% of the bacterial population. A functional genomics strategy was employed to discover the molecular support for colistin resistance in Burkholderia pseudomallei sp. A191. A pUC plasmid-based genomic expression library was constructed with an estimated library size of 2.1 × 10(7)bp. Five colistin resistant clones were obtained after functional screening of the library. Analysis of DNA sequence of five colistin resistant clones showed homology to two component regularity systems (TCRS) encoding for a histidine kinase (mrgS) and its regulatory component (mrgR). Cross complementation assay showed that mutations in mrgS were sufficient enough to confer colistin resistant phenotype in a sensitive strain.
authors.affiliation: Biotechnology and Bioinformatics, NIIT University, Neemrana, Rajasthan 301705, India.
authors.forename: Mohit
authors.initials: M
authors.lastname: Kumar
authors.affiliation: Biotechnology and Bioinformatics, NIIT University, Neemrana, Rajasthan 301705, India.
authors.forename: Ashutosh
authors.initials: A
authors.lastname: Gupta
authors.affiliation: Centre of Biotechnology, Siksha 'O' Anusandhan University, Khandagiri, Bhubaneswar, Odisha 751003, India.
authors.forename: Rajesh Kumar
authors.initials: RK
authors.lastname: Sahoo
authors.affiliation: Department of Microbiology, IMS &SUM Hospital, Siksha 'O' Anusandhan University, Kalinga Nagar, Bhubaneswar, Odisha 751003, India.
authors.forename: Jayanti
authors.initials: J
authors.lastname: Jena
authors.affiliation: Department of Microbiology, IMS &SUM Hospital, Siksha 'O' Anusandhan University, Kalinga Nagar, Bhubaneswar, Odisha 751003, India.
authors.forename: Nagen Kumar
authors.initials: NK
authors.lastname: Debata
authors.affiliation: Centre of Biotechnology, Siksha 'O' Anusandhan University, Khandagiri, Bhubaneswar, Odisha 751003, India.
authors.forename: Enketeswara
authors.initials: E
authors.lastname: Subudhi
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001419:Bacteria / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000382:microbiology*; D001426:Bacterial Proteins / Q000235:genetics*; D002648:Child; D003001:Cloning, Molecular; D003091:Colistin; D024901:Drug Resistance, Multiple, Bacterial; D005260:Female; D016680:Genome, Bacterial; D000071677:Histidine Kinase / Q000235:genetics; D006801:Humans; D008297:Male; D008875:Middle Aged; D010802:Phylogeny; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D055815:Young Adult
chemical_list: D001426:Bacterial Proteins; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D000071677:Histidine Kinase; D003091:Colistin