Efflux pumps expression and its association with porin down-regulation and β-lactamase production among Pseudomonas aeruginosa causing bloodstream infections in Brazil

Background: Multi-drug efflux pumps have been increasingly recognized as a major component of resistance in P. aeruginosa. We have investigated the expression level of efflux systems among clinical isolates of P. aeruginosa, regardless of their antimicrobial susceptibility profile.Results: Aztreonam exhibited the highest in vitro activity against the P. aeruginosa isolates studied (64.4% susceptibility), whereas susceptibility rates of imipenem and meropenem were both 47.5%. the MexXY-OprM and MexAB-OprM efflux systems were overexpressed in 50.8% and 27.1% of isolates studied, respectively. Overexpression of the MexEF-OprN and MexCD-OprJ systems was not observed. AmpC beta-lactamase was overexpressed in 11.9% of P. aeruginosa isolates. in addition, decreased oprD expression was also observed in 69.5% of the whole collection, and in 87.1% of the imipenem non-susceptible P. aeruginosa clinical isolates. the MBL-encoding genes bla(SPM-1) and bla(IMP-1) were detected in 23.7% and 1.7% P. aeruginosa isolates, respectively. the bla(GES-1) was detected in 5.1% of the isolates, while bla(GES-5) and bla(CTX-M-2) were observed in 1.7% of the isolates evaluated. in the present study, we have observed that efflux systems represent an adjuvant mechanism for antimicrobial resistance.Conclusions: Efflux systems in association of distinct mechanisms such as the porin down-regulation, AmpC overproduction and secondary beta-lactamases play also an important role in the multi-drug resistance phenotype among P. aeruginosa clinical isolates.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Universidade Federal de São Paulo, Div Infect Dis, BR-04039032 São Paulo, BrazilUniversidade Federal de São Paulo, Div Infect Dis, BR-04039032 São Paulo, BrazilFAPESP: FAPESP - 2006/01716-8CNPq: 307714/2006-3Web of Scienc

Diversity of β-Lactamases Produced by Ceftazidime-Resistant Pseudomonas aeruginosa Isolates Causing Bloodstream Infections in Brazil ▿

A retrospective survey was conducted to characterize β-lactamases in a collection of 43 ceftazidime-resistant Pseudomonas aeruginosa isolates recovered from patients with bloodstream infections hospitalized at a Brazilian teaching hospital between January and December 2005. Resistance rates for carbapenems, aminoglycosides, and quinolones were over 80%, with only colistin remaining active against all isolates. Pulsed-field gel electrophoresis analysis identified seven different genotypes. AmpC overproduction was found to be the sole β-lactamase-mediated mechanism responsible for ceftazidime resistance in four isolates (9.3%). Nine isolates (20.9%) produced an extended-spectrum β-lactamase (ESBL), either GES-1 (n = 7, 16.3%) or CTX-M-2 (n = 2, 4.6%). Carbapenemase activity was detected in 30 (70%) additional isolates. Among those isolates, two isolates (4.6%) produced the ESBL GES-5, possessing the ability to hydrolyze imipenem; a single isolate (2.3%) produced the metallo-β-lactamase (MBL) IMP-1; and 27 isolates produced the MBL SPM-1 (62.8%). None of the isolates coproduced both ESBL and MBL. Insertion sequence elements ISCR4 and ISCR1 were associated with blaSPM-1 and blaCTX-M-2 genes, respectively, whereas the blaGES-1 and blaGES-5 genes were part of class 1 integron structures. This study underlines the spread of MBL- and ESBL-producing P. aeruginosa isolates as an important source of ceftazidime resistance in Brazil

Irrigation Ponds as Sources of Antimicrobial-Resistant Bacteria in Agricultural Areas with Intensive Use of Poultry Litter

Poultry litter is widely used worldwide as an organic fertilizer in agriculture. However, poultry litter may contain high concentrations of antibiotics and/or antimicrobial-resistant bacteria (ARB), which can be mobilized through soil erosion to water bodies, contributing to the spread of antimicrobial resistance genes (ARGs) in the environment. To better comprehend this kind of mobilization, the bacterial communities of four ponds used for irrigation in agricultural and poultry production areas were determined in two periods of the year: at the beginning (low volume of rainfall) and at the end of the rainy season (high volume of rainfall). 16S rRNA gene sequencing revealed not only significantly different bacterial community structures and compositions among the four ponds but also between the samplings. When the DNA obtained from the water samples was PCR amplified using primers for ARGs, those encoding integrases (intI1) and resistance to sulfonamides (sul1 and sul2) and β-lactams (blaGES, blaTEM and blaSHV) were detected in three ponds. Moreover, bacterial strains were isolated from CHROMagar plates supplemented with sulfamethoxazole, ceftriaxone or ciprofloxacin and identified as belonging to clinically important Enterobacteriaceae. The results presented here indicate a potential risk of spreading ARB through water resources in agricultural areas with extensive fertilization with poultry litter

Efflux pumps expression and its association with porin down-regulation and β-lactamase production among <it>Pseudomonas aeruginosa </it>causing bloodstream infections in Brazil

Abstract Background Multi-drug efflux pumps have been increasingly recognized as a major component of resistance in P. aeruginosa. We have investigated the expression level of efflux systems among clinical isolates of P. aeruginosa, regardless of their antimicrobial susceptibility profile. Results Aztreonam exhibited the highest in vitro activity against the P. aeruginosa isolates studied (64.4% susceptibility), whereas susceptibility rates of imipenem and meropenem were both 47.5%. The MexXY-OprM and MexAB-OprM efflux systems were overexpressed in 50.8% and 27.1% of isolates studied, respectively. Overexpression of the MexEF-OprN and MexCD-OprJ systems was not observed. AmpC β-lactamase was overexpressed in 11.9% of P. aeruginosa isolates. In addition, decreased oprD expression was also observed in 69.5% of the whole collection, and in 87.1% of the imipenem non-susceptible P. aeruginosa clinical isolates. The MBL-encoding genes blaSPM-1 and blaIMP-1 were detected in 23.7% and 1.7% P. aeruginosa isolates, respectively. The blaGES-1 was detected in 5.1% of the isolates, while blaGES-5 and blaCTX-M-2 were observed in 1.7% of the isolates evaluated. In the present study, we have observed that efflux systems represent an adjuvant mechanism for antimicrobial resistance. Conclusions Efflux systems in association of distinct mechanisms such as the porin down-regulation, AmpC overproduction and secondary β-lactamases play also an important role in the multi-drug resistance phenotype among P. aeruginosa clinical isolates.</p

Cloverleaf test (modified Hodge test) for detecting carbapenemase production in Klebsiella pneumoniae: be aware of false positive results

The aim of this study was to evaluate the presence of carbapenemases in a Klebsiella pneumoniae collection and the performance of the modified Hodge test (MHT) to correctly identify this phenotype.Twenty-eight K. pneumoniae clinical isolates with reduced susceptibility to carbapenems were evaluated. Antimicrobial susceptibility and molecular typing were performed by agar dilution and PFGE, respectively. the MHT was performed using both standard and high inoculum of test organisms. Imipenem hydrolysis was investigated by spectrophotometric assays and carbapenemase-encoding genes were identified by PCR and amplicon sequencing. Porin loss was investigated by both PCR and SDS-PAGE.Susceptibility rates for imipenem, meropenem and ertapenem were 93%, 57% and 11%, respectively. the PFGE analysis showed seven unrelated genotypes. By testing standard inoculum and ertapenem or meropenem discs, 25% (n = 7) and 21% (n = 6) of the isolates were classified as carbapenemase producers, respectively. When a higher inoculum was employed, these rates increased to 54% (n = 15) and 43% (n = 12), respectively. No imipenem hydrolysis was detected. PCRs identified bla(CTX-M) in 27 (96%) isolates, of which 2 isolates also carried bla(GES-1.) SDS-PAGE and PCR assays revealed that all isolates had lost at least one outer membrane protein, except for a single isolate that was found to express both OmpK35 and OmpK36.False detection of carbapenemase production was observed by the MHT possibly as a result of extended-spectrum beta-lactamase (ESBL) production coupled with porin loss as reported before. Clinical laboratories must be aware of this fact, especially in geographical areas where ESBL-producing isolates are highly prevalent.Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Universidade Federal de São Paulo, Lab ALERTA, São Paulo, BrazilUniversidade Federal de São Paulo, Lab ALERTA, São Paulo, BrazilCNPq: 307714/2006-3Web of Scienc

Outbreak of OXY-2-Producing Klebsiella oxytoca in a Renal Transplant Unit▿

We describe a Klebsiella oxytoca infection outbreak in a renal transplant unit that involved seven patients. All strains belonged to a single pulsed-field gel electrophoresis pattern and were resistant to amoxicillin-clavulanate, cefuroxime, piperacillin-tazobactam, and aztreonam but susceptible to ceftriaxone, ceftazidime, cefepime, and imipenem. Chromosomal β-lactamase hyperproduction was caused by a point mutation in the blaOXY-2 gene promoter region

Detection of GES-5-producing Klebsiella pneumoniae in Brazil

Universidade Federal de São Paulo, Lab ALERTA, São Paulo, BrazilUniversidade Federal de São Paulo, Lab ALERTA, São Paulo, BrazilWeb of Scienc