The emergence and dissemination of an epidemic clone has contributed to the high carbapenem resistance rates among P. aeruginosa isolates in Brazil. In addition, the production of SPM MBL has an important role in carbapenem resistance in this region. This is the first report of dissemination of an SPM-1-like-MBL-producing strain of P. aeruginosa among unrelated Brazilian hospitals.

Pseudomonas aeruginosa strains that produce metallo-␤-lactamases (MBLs) are becoming increasingly prevalent. We evaluated the epidemiological and microbiological characteristics of monomicrobial bloodstream infections caused by MBL-producing P. aeruginosa isolates, as well as the clinical outcomes in patients with these infections.

BackgroundInfections are the major cause of morbidity and mortality in children with cancer. Gaining a favorable prognosis for these patients depends on selecting the appropriate therapy, which in turn depends on rapid and accurate microbiological diagnosis. This study employed real-time PCR (qPCR) to identify the main pathogens causing bloodstream infection (BSI) in patients treated at the Pediatric Oncology Institute IOP-GRAACC-UNIFESP-Brazil. Antimicrobial resistance genes were also investigated using this methodology.MethodsA total of 248 samples from BACTEC® blood culture bottles and 99 whole-blood samples collected in tubes containing EDTA K2 Gel were isolated from 137 patients. All samples were screened by specific Gram probes for multiplex qPCR. Seventeen sequences were evaluated using gender-specific TaqMan probes and the resistance genes blaSHV, blaTEM, blaCTX, blaKPC, blaIMP, blaSPM, blaVIM, vanA, vanB and mecA were detected using the SYBR Green method.ResultsPositive qPCR results were obtained in 112 of the blood culture bottles (112/124), and 90 % agreement was observed between phenotypic and molecular microbial detection methods. For bacterial and fungal identification, the performance test showed: sensitivity 87 %; specificity 91 %; NPV 90 %; PPV 89 % and accuracy of 89 % when compared with the phenotypic method. The mecA gene was detected in 37 samples, extended-spectrum β-lactamases were detected in six samples and metallo-β-lactamase coding genes in four samples, with 60 % concordance between the two methods. The qPCR on whole blood detected eight samples possessing the mecA gene and one sample harboring the vanB gene. The blaKPC, blaVIM, blaIMP and blaSHV genes were not detected in this study.ConclusionReal-time PCR is a useful tool in the early identification of pathogens and antimicrobial resistance genes from bloodstream infections of pediatric oncologic patients.

BackgroundEarly identification of pathogens and antimicrobial resistance in bloodstream infections (BSIs) decreases morbidity and mortality, particularly in immunocompromised patients. The aim of the present study was to compare real-time polymerase chain reaction (PCR) with commercial kits for detection of 17 pathogens from blood culture (BC) and 10 antimicrobial resistance genes.MethodsA total of 160 BCs were taken from bone marrow transplant patients and screened with Gram-specific probes by multiplex real-time PCR and 17 genus-specific sequences using TaqMan probes and blaSHV, blaTEM, blaCTX, blaKPC, blaIMP, blaSPM, blaVIM, vanA, vanB, and mecA genes by SYBR Green.ResultsTwenty-three of 33 samples identified by phenotypic testing were concordantly positive by BC and real-time PCR. Pathogen identification was discordant in 13 cases. In 12 of 15 coagulase-negative staphylococci, the mecA gene was detected and four Enterococcus spp. were positive for vanA. Two blaCTX and three blaSHV genes were found by quantitative PCR. The blaKPC and metallo-β-lactamase genes were not detected. Five fungal species were identified only by real-time PCR.ConclusionsReal-time PCR could be a valuable complementary tool in the management of BSI in bone marrow transplants patients, allowing identification of pathogens and antimicrobial resistance genes.

BackgroundBloodstream infections (BSIs) are the major cause of mortality in cancer patients. Molecular techniques are used for rapid diagnosis of BSI, allowing early therapy and improving survival. We aimed to establish whether real-time quantitative polymerase chain reaction (qPCR) could improve early diagnosis and therapy in paediatric cancer patients, and describe the predominant pathogens of BSI and their antimicrobial susceptibility.MethodsBlood samples were processed by the BACTEC system and microbial identification and susceptibility tests were performed by the Phoenix system. All samples were screened by multiplex 16 s rDNA qPCR. Seventeen species were evaluated using sex-specific TaqMan probes and resistance genes blaSHV, blaTEM, blaCTX, blaKPC, blaIMP, blaSPM, blaVIM, vanA, vanB and mecA were screened by SYBR Green reactions. Therapeutic efficacy was evaluated at the time of positive blood culture and at final phenotypic identification and antimicrobial susceptibility results.ResultsWe analyzed 69 episodes of BSI from 64 patients. Gram-positive bacteria were identified in 61 % of the samples, Gram-negative bacteria in 32 % and fungi in 7 %. There was 78.2 % of agreement between the phenotypic and molecular methods in final species identification. The mecA gene was detected in 81.4 % of Staphylococcus spp., and 91.6 % were concordant with the phenotypic method. Detection of vanA gene was 100 % concordant. The concordance for Gram-negative susceptibilities was 71.4 % for Enterobacteriaceae and 50 % for Pseudomonas aeruginosa. Therapy was more frequently inadequate in patients who died, and the molecular test was concordant with the phenotypic susceptibility test in 50 %.ConclusionsqPCR has potential indication for early identification of pathogens and antimicrobial resistance genes from BSI in paediatric cancer patients and may improve antimicrobial therapy.