title: Mutational and acquired carbapenem resistance mechanisms in multidrug resistant Pseudomonas aeruginosa clinical isolates from Recife, Brazil.
abstract: An investigation was carried out into the genetic mechanisms responsible for multidrug resistance in nine carbapenem-resistant Pseudomonas aeruginosa isolates from different hospitals in Recife, Brazil. Susceptibility to antimicrobial agents was determined by broth microdilution. Polymerase chain reaction (PCR) was employed to detect the presence of genes encoding β-lactamases, aminoglycoside-modifying enzymes (AMEs), 16S rRNA methylases, integron-related genes and OprD. Expression of genes coding for efflux pumps and AmpC cephalosporinase were assessed by quantitative PCR. The outer membrane proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The blaSPM-1, blaKPC-2 and blaGES-1 genes were detected in P. aeruginosa isolates in addition to different AME genes. The loss of OprD in nine isolates was mainly due to frameshift mutations, premature stop codons and point mutations. An association of loss of OprD with the overexpression of MexAB-OprM and MexXY-OprM was observed in most isolates. Hyper-production of AmpC was also observed in three isolates. Clonal relationship of the isolates was determined by repetitive element palindromic-PCR and multilocus sequence typing. Our results show that the loss of OprD along with overexpression of efflux pumps and β-lactamase production were responsible for the multidrug resistance in the isolates analysed.
authors.affiliation: Laboratório de Resistência Microbiana, Instituto de Ciências Biológicas, Universidade de Pernambuco, Recife, PE, Brasil.
authors.forename: Felipe Lira de Sá
authors.initials: FL
authors.lastname: Cavalcanti
authors.affiliation: Hospital Universitario Marqués de Valdecilla, Instituto de Investigación Marqués de Valdecilla, Santander, Spain.
authors.forename: Cristina Rodríguez
authors.initials: CR
authors.lastname: Mirones
authors.affiliation: Hospital Universitario Marqués de Valdecilla, Instituto de Investigación Marqués de Valdecilla, Santander, Spain.
authors.forename: Elena Román
authors.initials: ER
authors.lastname: Paucar
authors.affiliation: Hospital Universitario Marqués de Valdecilla, Instituto de Investigación Marqués de Valdecilla, Santander, Spain.
authors.forename: Laura Álvarez
authors.initials: LÁ
authors.lastname: Montes
authors.affiliation: Departamento de Genética, Centro de Ciências Biológicas, Universidade Federal de Pernambuco, Recife, PE, Brasil.
authors.forename: Tereza Cristina
authors.initials: TC
authors.lastname: Leal-Balbino
authors.affiliation: Laboratório de Resistência Microbiana, Instituto de Ciências Biológicas, Universidade de Pernambuco, Recife, PE, Brasil.
authors.forename: Marcia Maria Camargo de
authors.initials: MM
authors.lastname: Morais
authors.affiliation: Hospital Universitario Marqués de Valdecilla, Instituto de Investigación Marqués de Valdecilla, Santander, Spain.
authors.forename: Luis
authors.initials: L
authors.lastname: Martínez-Martínez
authors.affiliation: Hospital Universitario Marqués de Valdecilla, Instituto de Investigación Marqués de Valdecilla, Santander, Spain.
authors.forename: Alain Antonio
authors.initials: AA
authors.lastname: Ocampo-Sosa
mesh_terms: D000617:Aminoglycosides / Q000378:metabolism; D000666:Amphotericin B / Q000031:analogs & derivatives / Q000378:metabolism; D000935:Antifungal Agents / Q000378:metabolism; D001938:Brazil; D015780:Carbapenems / Q000378:metabolism*; D002510:Cephalosporinase / Q000145:classification / Q000378:metabolism; D018389:Codon, Nonsense / Q000378:metabolism; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004789:Enzyme Activation / Q000235:genetics; D016368:Frameshift Mutation / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D006801:Humans; D026901:Membrane Transport Proteins / Q000378:metabolism; D008780:Methyltransferases / Q000378:metabolism; D009154:Mutation; D009713:Nucleotidyltransferases / Q000378:metabolism; D017354:Point Mutation / Q000235:genetics; D018272:Porins / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification; D012091:Repetitive Sequences, Nucleic Acid; D018440:beta-Lactam Resistance / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics / Q000378:metabolism*
chemical_list: D000617:Aminoglycosides; D000935:Antifungal Agents; D015780:Carbapenems; D018389:Codon, Nonsense; D026901:Membrane Transport Proteins; D018272:Porins; C017501:methylamphotericin B; C077157:OprD protein, Pseudomonas aeruginosa; D000666:Amphotericin B; D008780:Methyltransferases; C047346:16S rRNA (adenine(1518)-N(6)-adenine(1519)-N(6))-dimethyltransferase; D009713:Nucleotidyltransferases; D002510:Cephalosporinase; D001618:beta-Lactamases