Clonality, outer-membrane proteins profile and efflux pump in KPC- producing Enterobacter sp. in Brazil.

BACKGROUND: Carbapenems resistance in Enterobacter spp. has increased in the last decade, few studies, however, described the mechanisms of resistance in this bacterium. This study evaluated clonality and mechanisms of carbapenems resistance in clinical isolates of Enterobacter spp. identified in three hospitals in Brazil (Hospital A, B and C) over 7-year. METHODS: Antibiotics sensitivity, pulsed-field gel electrophoresis (PFGE), PCR for carbapenemase and efflux pump genes were performed for all carbapenems-resistant isolates. Outer-membrane protein (OMP) was evaluated based on PFGE profile. RESULTS: A total of 130 isolates of Enterobacter spp were analyzed, 44/105 (41, 9%) E. aerogenes and 8/25 (32,0%) E. cloacae were resistant to carbapenems. All isolates were susceptible to fosfomycin, polymyxin B and tigecycline. KPC was present in 88.6% of E. aerogenes and in all E. cloacae resistant to carbapenems. The carbapenems-resistant E. aerogenes identified in hospital A belonged to six clones, however, a predominant clone was identified in this hospital over the study period. There is a predominant clone in Hospital B and Hospital C as well. The mechanisms of resistance to carbapenems differ among subtypes. Most of the isolates co-harbored blaKPC, blaTEM and /or blaCTX associated with decreased or lost of 35-36KDa and or 39 KDa OMP. The efflux pump AcrAB-TolC gene was only identified in carbapenems-resistant E. cloacae. CONCLUSIONS: There was a predominant clone in each hospital suggesting that cross-transmission of carbapenems-resistant Enterobacter spp. was frequent. The isolates presented multiple mechanisms of resistance to carbapenems including OMP alteration.

Multidrug-resistant Stenotrophomonas maltophilia: Description of new MLST profiles and resistance and virulence genes using whole-genome sequencing.

OBJECTIVES: Stenotrophomonas maltophilia is an opportunistic pathogen that has high intrinsic and acquired antimicrobial resistance, with great genetic diversity. The aim of this study was to characterise four S. maltophilia clinical isolates displaying different susceptibility profiles using whole-genome sequencing. METHODS: The whole genomes of four clinical isolates of S. maltophilia from three patients were sequenced using Ion Torrent™ PGM technology. The isolates presented different susceptibilities to trimethoprim/sulfamethoxazole (SXT) and levofloxacin. RESULTS: Three new multilocus sequence typing (MLST) profiles were identified (ST144, ST172 and ST173), differing in virulence and resistance genes. The ST172 isolate had more genes related to toxins than related to motility or adhesion and had different types of efflux pumps than the other isolates. The SXT-resistant strains belonged to ST172 or ST144 and did not harbour the sul1, sul2 or dfrA resistance genes. Strains I and II, from the same patient and belonging to the same ST but differing in resistance to SXT, had all of the resistance genes searched for in common, except for the SmeABC efflux pump complex genes that were only found in the SXT-resistant strain. All strains, including the strain susceptible to levofloxacin, harboured the qnrB gene, which may question the importance of this gene in determining levofloxacin resistance in S. maltophilia. CONCLUSION: Here we describe three new MLST profiles. Resistance to SXT in these strains appears to be associated with efflux pumps.

Molecular characterization of carbapenem-resistant Klebsiella pneumoniae isolates from a university hospital in Brazil.

INTRODUCTION: The emergence of Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae (KPC-Kpn) isolates is attracting significant attention in nosocomial infection settings. K. pneumoniae is the main pathogen that harbours blaKPC genes. METHODOLOGY: This study evaluated 54 K. pneumoniae carbapenem-resistant isolates from patients hospitalized at the University Hospital of Londrina, between July 2009 and July 2010. The isolates were phenotypically screened for carbapenemase production and submitted for genotypic confirmation by polymerase chain reaction (PCR) for KPC, metallo-ß-lactamases, OXA-48, and extended-spectrum beta-lactamase genes. The absence of outer membrane proteins (OMP) was investigated by SDS-PAGE. The susceptibility profile was determined by broth microdilution, according to Clinical and Laboratory Standards Institute protocol. RESULTS: All isolates were phenotypically positive for class A carbapenemase production, but negative for metallo-ß-lactamase activity. PCR analysis demonstrated that all isolates carried blaKPC genes and sequencing showed that all strains belonged to KPC-2 subtype. Four strains did not show porin expression, and all isolates were resistant to ertapenem, meropenem, and imipenem. Susceptibility rates reached 35.2% for gentamicin, 85.2% for polymixyn B, 87% for colistin, and 98.1% for both tigecycline and fosfomycin. Pulsed-field gel electrophoresis showed six clones, and three of them predominated among the isolates. CONCLUSIONS: KPC-2-producing K. pneumoniae is becoming predominant among carbapenem-resistant K. pneumoniae isolates at the hospital. The association of the enzyme KPC with other resistance determinants, such as loss of porins, may increase the severity of the situation of nosocomial infections. There is an urgent need to develop strategies for infection control and prevention.