Rapid bacteria identification and susceptibility profile in blood culture from patients attending at a tertiary care hospital in South of Brazil

Background. Bloodstream infection is a critical disease with high mortality rate. Adequate treatment required rapid microbiological identification and timely administration of appropriate antibiotics. Rapid diagnostic tests that accurately identify infection-causing pathogens and the effective antimicrobials against these pathogens can increase the likelihood that patients are treated appropriately. Rapid diagnostic tests can also be used to help clinicians discontinue unnecessary antibiotics or de-escalate broad-spectrum antimicrobial therapy to a narrower-spectrum option. Current technologies employed in blood culture routine diagnostics are precise and sensitive but rather slow, most depending on bacterial growth. Objective. In this study, we evaluated a rapid bacterial identification (rID) and a rapid antimicrobial susceptibility testing by disk diffusion (rAST) from positive blood culture to overcome the limitations of the conventional methods. We investigated a rapid workflow to reduce the turnaround time in bloodstream infections diagnostics in a routine microbiology laboratory. Methods. The study included hemocultures flagged as positive by BACT/ALERT® (Biomérieux, Marcy l’Etoile, France) between 7-12:00 a.m. and inoculated on Chocolate agar at our clinical microbiology laboratory. At 16.00 p.m., after 4-6h incubation at 35-38°C (5% CO2), identification by MALDI-TOF MS (VITEK MS® system,Biomérieux, Marcy-l’Étoile, France) and AST (disk diffusion method) were performed. Results were compared to identification (sID) and AST (sAST) results after 24h incubation. An identification score value of > 96% was considered as a correct species identification. For AST categorical agreement (CA), very major errors (VME, false-susceptible result of rapid AST), major errors (ME, false-resistant result of rapid AST), minor errors (mE, false categorization involving intermediate result) were investigated. Results. We identified a total of 526 bacterial isolated from blood cultures obtained from patients attended at a tertiary hospital in the south of Brazil, 246 Gram-negative (GN) and 279 Gram-positive (GP) aerobes. The overall concordance between rID and sID was 88.6% and was highest for GN (95.5%). K. pneumoniae and E. coli presented 96.0% and 98.5% rate of concordance, respectively. Total of 2196 and 1476 antimicrobial agents’ comparisons were obtained for GN and GP, respectively. Evaluating rAST, the CA, VME, ME and mE were 97.7, 0.7, 0.5 and 1.1% for GN and 98.0, 0.5, 0.7 and 0.8% for GP, respectively. Meropenem CA, VME and ME were 98.3, 0.5 and 0.5%, respectively; no mE was observed. Oxacillin CA, ME and mE were 97.4, 1.6 and 0.6%, respectively; no VME was observed. Sensitivity and specificity of rAST method were calculated for each antimicrobial agents. Meropenem presented 99.2 and 98.1% and Oxacillin presented 96.9 and 97.9% of sensibility and specificity, respectively. Overall Kappa scores of the comparisons results demonstrated the high agreement between rAST and sAST. Conclusions. The rapid methods for bacterial identification and for rapid AST results proposed in this study were distinguished from standards ones by feasible modifications and the accuracy of rAST were comparable with the standard method. Identification and AST of aerobic bacteria from positive blood cultures after a shortened incubation on solid blood agar is a fast and reliable method that may improve management of bloodstream infections, allowing us to save up to 24 h to identifying bacteria and supply useful information to adapt antibiotic therapy when necessary.Introdução. A infecção da corrente sanguínea é uma doença crítica com alta taxa de mortalidade. O tratamento adequado exige identificação microbiológica rápida e administração oportuna de antibióticos apropriados. Testes diagnósticos rápidos que identifiquem com precisão os patógenos causadores de infecções e os antimicrobianos eficazes contra esses patógenos podem aumentar a probabilidade dos pacientes serem tratados adequadamente. As tecnologias atuais empregadas nos diagnósticos de rotina da hemocultura são precisas e sensíveis, mas bastante lentas, e dependente do crescimento bacteriano em cultura em meio sólido. Objetivo. Neste estudo, nós avaliamos a identificação rápida bacteriana (rID) e um teste rápido de susceptibilidade antimicrobiana por disco-difusão (rAST) de hemocultura positiva para otimizar a rotina de hemoculturas positivas, superando as limitações dos métodos convencionais. Nós investigamos um fluxograma rápido para reduzir o tempo de liberação dos resultados no diagnósticos de infecções da corrente sanguínea em um laboratório de microbiologia clínica. Métodos Hemoculturas positivas pelo BACT / ALERT® (Biomérieux, Marcy l'Etoile, França) entre as 7 - 12:00h eram processadas e inoculadas em ágar Chocolate. Após 4-6h de incubação a 35-38 ° C (5% CO2), identificação por MALDI-TOF MS (sistema VITEK MS®, Biomérieux, Marcy-l'Étoile, França) e AST (método de difusão em disco) foram realizados. Os resultados foram comparados com os resultados de identificação (sID) e AST (sAST) a partir de culturas de 24h. Para a concordância categórica AST (CA), erros muito importantes (VME, resultado falso-suscetível da AST rápida), erros maiores (ME, resultado falso-resistente da AST rápida), erros menores (mE, falsa categorização envolvendo resultado intermediário) foram investigados. Resultados. Identificamos um total de 526 bactérias isoladas de hemoculturas obtidas de pacientes atendidos em um hospital terciário no sul do Brasil, 246 aeróbios Gram-negativos (GN) e 279 Gram-positivos (GP). A concordância global entre rID e sID foi de 88,6% e foi mais alta para GN (95,5%). K. pneumoniae e E. coli apresentaram taxa de concordância de 96,0% e 98,5%, respectivamente. Total de 2196 e 1476 comparações de agentes antimicrobianos foram obtidas para GN e GP, respectivamente. Avaliando rAST, o CA, VME, ME e mE foram 97,7, 0,7, 0,5 e 1,1% para GN e 98,0, 0,5, 0,7 e 0,8% para GP, respectivamente. Para meropenem CA, VME e ME foram 98,3, 0,5 e 0,5%, respectivamente; nenhum mE foi observado. Para oxaciina CA, ME e mE foram 97,4, 1,6 e 0,6%, respectivamente; nenhum VME foi observado. A sensibilidade e especificidade do método rAST foram calculadas para cada agente antimicrobiano. Meropenem apresentou 99,2 e 98,1% e a Oxacilina apresentou 96,9 e 97,9% de sensibilidade e especificidade, respectivamente. Os escores Kappa gerais dos resultados das comparações demonstraram a alta concordância entre rAST e sAST. Conclusão. Os métodos rápidos para identificação bacteriana e para teste de sensibilidade aos antimicrobianos propostos neste estudo diferenciam-se dos padrões por modificações no tempo de cultura e a precisão do rAST foi comparável com o método padrão. Identificação e AST de bactérias aeróbias de hemoculturas positivas após uma incubação rápida em meio sólido é um método rápido e confiável que pode otimizar o diagnóstico de infecções de corrente sanguínea, permitindo-nos reduzir em até 24 h a liberação da identificação bacteriana e o perfil de sensibilidade, assim fornecer informações de forma mais rápida e precisas a equipe assistencial

Integrating bacterial identification and susceptibility testing : a simple and rapid approach to reduce the turnaround time in the management of blood cultures

We evaluated a rapid bacterial identification (rID) and a rapid antimicrobial susceptibility testing by disk diffusion (rAST) from positive blood culture to overcome the limitations of the conventional methods and reduce the turnaround time in bloodstream infection diagnostics.-e study included hemocultures flagged as positive by bacT/ALERT®, identification by MALDI-TOF MS, and rAST.-eresults were compared to identification and antimicrobial susceptibility testing (AST) results by current standard methods, after 24 h incubation. For rAST categorical agreement (CA), very major errors (VME), major errors (ME), and minor errors (mE) were calculated. A total of 524 bacterial samples isolated from blood cultures were obtained, including 246 Gram-negative (GN) and 278 Gram-positive (GP) aerobes. -e overall concordance of rID was 88.6%, and it was highest among GN (96%). A total of 2196 and 1476 antimicrobial agent comparisons were obtained for GN and GP, respectively. Evaluation of rAST, CA, VME, ME, and mE disclosed 97.7, 0.7, 0.5, and 1.1% for GN and 98.0, 0.5, 0.7, and 0.8% for GP, respectively. Meropenem CA, VME, and ME were 98.3, 0.5, and 0.5%, respectively; mE was not observed. Oxacillin CA, ME, and mE were 97.4, 1.6, and 0.6%, respectively; VME was not observed. Overall, kappa scores of the results of the comparisons demonstrated the high agreement between rAST and the standard method. Identification and ASTof aerobic bacteria from positive blood cultures after a short period of incubation on solid blood agar is a fast and reliable method that may improve the management of bloodstream infections

Occurrence and genetic characterization of Listeria spp. in minimally processed vegetables commercialized in Porto Alegre, Brazil

Minimally processed vegetables go through many steps before they are refrigerated, selection, washing, peeling, cutting, disinfection and finally packaging. However, if no care is taken at the origin of the raw materials and in the processing stages, there is a chance of finding pathogenic bacteria, such as Listeria monocytogenes, which are able to grow at low temperatures. The aim of this research was to verify the occurrence of Listeria sp. in minimally processed vegetables sold in Porto Alegre, Brazil, and the genetic relationship among the isolates. Minimally processed salads were sampled monthly from local supermarkets and analyzed by inoculation on Listeria Enrichment Broth and subsequent seeding on two selective media, Palcam and Modified Oxford Agar. The typical colonies were identified to species level and their intergenic region 16S-23S rDNA were amplified in order to verify the genetic variability. Species of Listeria were found in 23 of the 52 processed salad samples analyzed and L. monocytogenes was found in seven. The presence of L. monocytogenes in the samples is a health concern, as these salads are eaten without further treatment by the consumer. The amplification of the intergenic region 16S-23S rDNA, showed a great genetic diversity among the isolates, with 43 different patterns, proving the usefulness of this technique in epidemiologic studies

Evaluation of identification and susceptibility for Candida spp. isolated directly from positive blood culture bottles

Determination of the susceptibility profile of isolates of Candida from blood culture bottles is extremely important for correctly guiding patient pharmacotherapy. ,e aim of this study was to compare the results of analysis of Candida isolated directly from blood culture bottles by the VITEK MS MALDI-TOF identification system and the fluconazole disk diffusion assay with those of standard identification methods. Testing directly from the bottle allowed results 24 to 48 hours quicker than the standard method. ,ere was a categorical agreement of 51.64% (47 of 91 samples) between the results of analysis directly from the bottle and analysis by the standard method. Regarding species identification, there was 96.15% agreement for Candida parapsilosis (25 of 26 samples). Categorical agreement between the rapid and standard disk diffusion methods was 95%, and the agreement between the rapid disk diffusion method and the broth microdilution method was 97%. Only minor errors in the rapid method were observed: 3 (5%) in the standard disk diffusion method and 2 (3%) in the broth microdilution method. Our study concluded that the rapid disk diffusion method for fluconazole is a fast, easy, reproducible, and consistent method. Its timely implementation for testing antifungal agents in the clinical microbiology laboratory can help reduce profile release times, thus helping to determine the most appropriate antifungal treatment