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Development and analysis of furazolidone‐resistant E scherichia coli mutants
Author(s) -
MartínezPuchol Sandra,
Gomes Cláudia,
Pons Maria J.,
RuizRoldán Lidia,
Torrents de la Peña Alba,
Ochoa Theresa J.,
Ruiz Joaquim
Publication year - 2015
Publication title -
apmis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.909
H-Index - 88
eISSN - 1600-0463
pISSN - 0903-4641
DOI - 10.1111/apm.12401
Subject(s) - furazolidone , nalidixic acid , mutant , efflux , nitroreductase , tetracycline , escherichia coli , chloramphenicol , mutation , bacterial outer membrane , biology , microbiology and biotechnology , chemistry , gene , biochemistry , antibiotics
Furazolidone‐resistant mutants were obtained from four clinical isolates of diarrhoeagenic Escherichia coli . The stability of the resistance and the frequency of mutation were established. The minimal inhibitory concentration of furazolidone, nitrofurantoin, nalidixic acid, ampicillin, chloramphenicol and tetracycline was established both in the presence and absence of the efflux pump inhibitor Phe‐Arg‐β‐Naphtylamyde. The presence of mutations in the nitroreductase genes nfsA and nfsB was analysed by PCR ; sequencing and their enzymatic activity was assessed by a spectrophotometric assay. Alterations in outer membrane proteins were studied by SDS ‐ PAGE . The frequency of mutation ranged from <9.6 × 10 −10 to 9.59 × 10 −7 . Neither an effect on efflux pumps inhibited by Phe‐Arg‐β‐Naphtylamyde nor cross‐resistance with the antibiotics studied was observed. Nineteen mutants (52.94%) presented mutations in the nitroreductase‐encoding genes: 17 in the nfsA gene (15 mutants with an internal stop codon, 2 with amino acid changes), 2 in the nfsB (all amino acid changes). Alterations in the outer membrane proteins OmpA and OmpW were also observed. Although more studies are necessary to find other resistance mechanisms, present data showed the low potential of selecting furazolidone‐resistant mutants, together with the lack of cross‐resistance with unrelated antimicrobial agents.