A simple double differential centrifugation-wash procedure to rapidly obtain bacterial identification and direct antimicrobial susceptibility testing from positive blood cultures / Método sencillo basado en una doble centrifugación diferencial para obtener una identificación bacteriana y sensibilidad a antibióticos rápida a partir de hemocultivos positivos

Introduction: This study proposes a simple and rapid method for both bacterial identification and direct antimicrobial susceptibility testing (AST) by using MALDI-TOF and a double differential centrifugation-wash procedure from positive blood cultures. Methods: Fifty-two positive blood cultures (37 gramnegative bacilli and 15 grampositive cocci) were studied by two methods for identification and AST: a reference method, and the rapid MALDI-TOF method obtaining a purified pellet by using a double differential centrifugation procedure. Results : A total of 1101 MIC values (mg/l) were interpreted according to EUCAST clinical breakpoints and compared using the two methods simultaneously. Discrepancies in 81 MIC values (7.35%) were detected. By analyzing standard parameters, we obtained 98.28% essential agreement and 92.65% categorical agreement considering all isolates tested. Conclusion: This method provides rapid bacterial identification and AST, offering definitive results 24–48h earlier than the conventional method (p<0.001) and improving the turnaround time in blood culture diagnostics, especially in laboratories without 24-h on-call.(AU)

Introducción: Este trabajo propone un método sencillo, rápido y barato de identificación bacteriana y sensibilidad antibiótica utilizando MALDI-TOF y una doble centrifugación diferencial a partir de hemocultivo positivo. Métodos: Se estudiaron 52 hemocultivos positivos (37 bacilos gramnegativos y 15 cocos grampositivos). Se compararon 2 métodos: un método convencional de identificación y determinación de sensibilidad a antibióticos automatizada partiendo de colonia crecida, y un método rápido utilizando MALDI-TOF, caracterizado por la obtención de un pellet purificado procedente de un hemocultivo positivo, mediante un procedimiento basado en una doble centrifugación diferencial. Resultados: Se analizaron y compararon 1.101 valores de CMI (mg/l) de acuerdo con los criterios establecidos por EUCAST y obtenidos por ambos métodos. Se detectaron discrepancias en 81 valores de CMI (7,35%). Considerando todos los aislados, la concordancia esencial fue del 98,28% y la concordancia categórica del 92,65%. Conclusión: Este método proporciona resultados de identificación y sensibilidad a antibióticos definitivos 24-48h antes que un método convencional (p<0,001), mejorando el tiempo de respuesta en el diagnóstico microbiológico de bacteriemias, especialmente en laboratorios sin servicio de guardias de 24h.(AU)

A simple double differential centrifugation-wash procedure to rapidly obtain bacterial identification and direct antimicrobial susceptibility testing from positive blood cultures.

INTRODUCTION: This study proposes a simple and rapid method for both bacterial identification and direct antimicrobial susceptibility testing (AST) by using MALDI-TOF and a double differential centrifugation-wash procedure from positive blood cultures. METHODS: Fifty-two positive blood cultures (37 gramnegative bacilli and 15 grampositive cocci) were studied by two methods for identification and AST: a reference method, and the rapid MALDI-TOF method obtaining a purified pellet by using a double differential centrifugation procedure. RESULTS: A total of 1101 MIC values (mg/l) were interpreted according to EUCAST clinical breakpoints and compared using the two methods simultaneously. Discrepancies in 81 MIC values (7.35%) were detected. By analyzing standard parameters, we obtained 98.28% essential agreement and 92.65% categorical agreement considering all isolates tested. CONCLUSION: This method provides rapid bacterial identification and AST, offering definitive results 24-48h earlier than the conventional method (p<0.001) and improving the turnaround time in blood culture diagnostics, especially in laboratories without 24-h on-call.

[Rapid identification of microorganisms by mass spectrometry in a blood culture system. Comparison of two procedures]. / Identificación rápida de microorganismos de frascos de hemocultivos por espectrometría de masas. Comparación de 2 procedimientos diagnósticos.

Rapid identification of microorganisms is critical in hospitalized infected patients. Blood culture is currently the gold standard for detecting and identifying microorganisms causing bacteremia or sepsis. The introduction of mass spectrometry by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) in microbiology laboratories, especially in microorganisms growing in blood culture bottles, provides rapid identification. This study evaluates the performance of the Maldi Sepsityper Biotyper procedure (hereinafter, MS) compared to that of an in-home method (hereinafter, HF). Eight hundred and forty (840) positive blood culture bottles were processed using the HF procedure, 542 of which were also processed using MS. The organisms were identified in 670 (79.76%) and 391 (72.14%) bottles respectively (p = 0,0013). This study demonstrates the effectiveness of both procedures for identifying microorganisms directly from positive blood culture bottles. However, the HF procedure proved to be more effective than MS, especially in the presence of Gram positive organisms.

Identificación rápida de microorganismos de frascos de hemocultivos por espectrometría de masas: Comparación de 2 procedimientos diagnósticos / Rapid identification of microorganisms by mass spectrometry in a blood culture system: Comparison of two procedures

La identificación rápida de microorganismos es crítica, en especial en pacientes sépticos hospitalizados. La espectrometría de masas conocida como matrix-assisted laser desorption/ionization time-of- flight mass spectrometry (MALDI-TOF MS) permite la identificación directa desde botellas de hemocultivos positivos en forma rápida y sencilla. Este estudio evaluó el desempeño del procedimiento basado en el sistema MALDI Biotyper que utiliza el kit comercial MALDI Sepsityper de Bruker Daltonics (en adelante, MS) frente a uno artesanal (en adelante, HF). Se procesaron 840 botellas de hemocultivos positivos con HF y 542 de estas fueron evaluadas también con MS. Se logró la identificación de los microorganismos en 670 (79,76 %) y 391 (72,14 %) botellas, respectivamente (p = 0,0013). Se demostró la efectividad de ambos procedimientos para la identificación de microorganismos desde frascos de hemocultivos positivos. Sin embargo, el procedimiento HF fue superior al MS, en especial frente a bacterias gram positivas.

Rapid identification of microorganisms is critical in hospitalized infected patients. Blood culture is currently the gold standard for detecting and identifying microorganisms causing bacteremia or sepsis. The introduction of mass spectrometry by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) in microbiology laboratories, especially in microorganisms growing in blood culture bottles, provides rapid identification. This study evaluates the performance of the Maldi Sepsityper Biotyper procedure (hereinafter, MS) compared to that of an in-home method (hereinafter, HF). Eight hundred and forty (840) positive blood culture bottles were processed using the HF procedure, 542 of which were also processed using MS. The organisms were identified in 670 (79. 76 %) and 391 (72. 14 %) bottles respectively (p = 0,0013). This study demonstrates the effectiveness of both procedures for identifying microorganisms directly from positive blood culture bottles. However, the HF procedure proved to be more effective than MS, especially in the presence of Gram positive organisms.

O método rápido de avaliação de programas de saúde / Rapid evaluation method of health programs

O Método Rápido de Avaliação é uma ferramenta de apoio à gestão e às equipes locais no planejamento e na tomada de decisão. Alguns dos seus princípios são: a agilidade, o uso de múltiplas fontes de evidência e o baixo custo. Pode ser adaptado às necessidades e condições de cada local. A utilização de uma matriz lógica possibilita um olhar objetivo sobre o desenho do projeto/programa e seus componentes, mostrando a interdependência deste com o contexto em que se insere. A construção do Método Rápido de Avaliação é compartilhada com os profissionais e usuários envolvidos no programa. A coleta de dados de diversas fontes e a utilização de diferentes métodos permite a triangulação de dados, o que possibilita a verificação contínua da confiabilidade, da validade e da interpretação da informação coletada.

The Rapid Evaluation Method (REM) is a support tool for management and local teams in the planning and decision making process. Some of its principles are: agility, the use of multiple sources of evidence, and low cost. It can be adapted to the needs and conditions at the local level. The use of a logic matrix allows for an objective view of the project/program design and its components. It also shows the interdependence between the program and the environment where it is located. REM construction is shared between the professionals and users involved in the program. Data collection from various sources and the use of diverse methods allows data triangulation and continuous verification of the reliability, validity, and interpretation of the information collected

Phage amplification assay as rapid method for Salmonella detection

The application of rapid methods is crucial for the HACCP program implantation in food industry. In this context, Phage Amplification Assay is a good candidate because is based on the interactions of phage and their host bacteria. This method using phage P22 was applied with to detect Salmonella cells in chicken breast. Samples of 25 g of chicken breast were diluted and the appropriate dilutions were used in phage amplification assay for Salmonella detection. After 3-4 h of incubation, it was observed a phage titre of approximately 10(4) pfu mL-1, indicating that there were Salmonella cells which were naturally present in the meat. The presence of Salmonella cells were verified by using direct plating on XLD agar and by conventional enrichment procedure. The colonies suspected to be Salmonella were serologically tested and were identified as belonging to the serogroups B (S. typhimurium group) and D (S. enteritidis group). It can be concluded that this method provides a rapid and alternative application for Salmonella detection in food samples reducing both time and laboratory work to 3-4 hours.

A aplicação de métodos rápidos é crucial para a implantação de programas de HACCP em indústrias de alimentos. Neste contexto, o método de amplificação de bacteriófagos é um instrumento de diagnóstico importante porque está baseado na interação dos bacteriófagos com suas células hospedeiras. Este método, usando o bacteriófago P22, foi aplicado para detectar Salmonella em peito de frango. Amostras de 25 g de peito de frango foram diluídas e as diluições apropriadas foram usadas no método de amplificação de bacteriófagos na detecção de Salmonella. Após 3-4 horas de incubação, foi observado uma titulação de partículas virais de, aproximadamente, 10(4) ufp mL-1 (unidades formadoras de placas virais), indicando a presença de células de Salmonella na carne de frango. A comprovação da presença de Salmonella neste produto foi verificada usando-se plaqueamento direto em ágar XLD e procedimento de enriquecimento convencional. As colônias suspeitas de Salmonella foram sorologicamente testadas e identificadas como pertencendo aos sorogrupos B (grupo de S. typhimurium) e D (grupo de S. enteritidis). Portanto, concluiu-se que este método pode ser aplicado, na detecção de Salmonella em alimentos, porque fornece rápido e conclusivo resultado, reduzindo o tempo de análise e o trabalho laboratorial para 3-4 horas.

Phage amplification assay as rapid method for Salmonella detection

The application of rapid methods is crucial for the HACCP program implantation in food industry. In this context, Phage Amplification Assay is a good candidate because is based on the interactions of phage and their host bacteria. This method using phage P22 was applied with to detect Salmonella cells in chicken breast. Samples of 25 g of chicken breast were diluted and the appropriate dilutions were used in phage amplification assay for Salmonella detection. After 3-4 h of incubation, it was observed a phage titre of approximately 10(4) pfu mL-1, indicating that there were Salmonella cells which were naturally present in the meat. The presence of Salmonella cells were verified by using direct plating on XLD agar and by conventional enrichment procedure. The colonies suspected to be Salmonella were serologically tested and were identified as belonging to the serogroups B (S. typhimurium group) and D (S. enteritidis group). It can be concluded that this method provides a rapid and alternative application for Salmonella detection in food samples reducing both time and laboratory work to 3-4 hours.

A aplicação de métodos rápidos é crucial para a implantação de programas de HACCP em indústrias de alimentos. Neste contexto, o método de amplificação de bacteriófagos é um instrumento de diagnóstico importante porque está baseado na interação dos bacteriófagos com suas células hospedeiras. Este método, usando o bacteriófago P22, foi aplicado para detectar Salmonella em peito de frango. Amostras de 25 g de peito de frango foram diluídas e as diluições apropriadas foram usadas no método de amplificação de bacteriófagos na detecção de Salmonella. Após 3-4 horas de incubação, foi observado uma titulação de partículas virais de, aproximadamente, 10(4) ufp mL-1 (unidades formadoras de placas virais), indicando a presença de células de Salmonella na carne de frango. A comprovação da presença de Salmonella neste produto foi verificada usando-se plaqueamento direto em ágar XLD e procedimento de enriquecimento convencional. As colônias suspeitas de Salmonella foram sorologicamente testadas e identificadas como pertencendo aos sorogrupos B (grupo de S. typhimurium) e D (grupo de S. enteritidis). Portanto, concluiu-se que este método pode ser aplicado, na detecção de Salmonella em alimentos, porque fornece rápido e conclusivo resultado, reduzindo o tempo de análise e o trabalho laboratorial para 3-4 horas.