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We evaluated the performance of five phenotypic tests [Modified Hodge Test (MHT); combined-disk test (CDT) using phenylboronic acid, EDTA, and cloxacillin; CarbaNP and CarbAcinetoNP; Blue-Carba, Carbapenembac™ and Carbapenembac Metallo™] for carbapenemase detection in Gram-negative bacilli (GNB). A total of 73 carbapenemase producers and 27 non-carbapenemase producers were tested. All GNB were subcultured onto Müeller-Hinton agar (MHA), MacConkey agar (MAC), and sheep blood agar (SBA). High sensitivity (100%) and specificity (100%) was observed for MHA using CarbaNP, Blue-Carba, and Carbapenembac™. The sensitivity and specificity of CarbaNP (98.6%/100%), Blue-Carba (97.1%/91.0%), and Carbapenembac™ (100%/96.5%) were slightly lower for SBA. In contrast, unacceptable sensitivity rates of CarbaNP (71.1%) and Blue-Carba (66.6%), but not Carbapenembac™ (97.3%), were observed for MAC. The colorimetric methods showed high sensitivity and specificity to detect carbapenemase production from isolates grown on MHA or SBA. However, colonies obtained from MAC must not be tested for carbapenemase detection by colorimetric methods.

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Carbapenem-resistant Enterobacteriaceae (CRE) is a major international health problem, and its identification in developing countries is based exclusively on phenotypic methods. The aim of this study was to assess the sensitivity and related parameters of the modified Hodge test (MHT). The assessment was performed in a large number of isolates obtained from different hospitals in several cities of a south Brazilian state. Bacterial species were identified using an automated method. The MHT was performed according to the guidelines set by the CLSI. The gene blaKPC was amplified in order to confirmation CRE expression. The sensitivity, specificity, positive, and negative predictive values were calculated. A total of 942 isolates were submitted to the reference laboratory for confirmation; 143 showed a negative MHT (15.18%) result, while 784 were positive (83.23%), and 15 samples displayed an indeterminate MHT (1.59%) result. All samples expressed the KPC-2 enzyme. Sensitivity, specificity, positive, and negative predictive percentiles were 99%, 89%, 98%, and 99% respectively. We conclude that the modified Hodge test is a reliable test for the prediction of KPC-producing bacteria.

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Introducción: la producción de enzimas con actividad hidrolítica frente a carbapenémicos, denominadascarbapenemasas, es uno de los principales mecanismos de resistencia a carbapenémicos en Pseudomonas aeruginosa. El test de Hodge modificado es la prueba fenotípica más empleada para la detección de carbapenemasas; sin embargo, de acuerdo con el CLSI, este método sólo puede emplearse en Enterobacteriáceas ya que presenta limitaciones en Bacilos Gram negativos no fermentadorescomo Pseudomonas aeruginosa. Objetivo: evaluar la eficacia de variaciones del test de Hodge modificado para la detección de carbapenemasas en aislados de Pseudomonas aeruginosa. Materiales y métodos: se evaluó el desempeño del test 3D y tres variaciones del test de Hodge modificado, tomando como prueba de referencia la detección de los genes de las carbapenemasas KPC, VIM, IMP, NDM y OXA-48 mediante reacción en cadena de la polimerasa. Las variaciones consistieron en emplear: a) agar MacConkey en lugar de Mueller Hinton, b) Klebsiella pneumoniae ATCC 700603 como cepa indicadora sensible a carbapenémicos en lugar de Escherichia coli ATCC 29212 y c) las dos condiciones anteriores simultáneamente. Resultados: de los ensayos evaluados, la tercera variación, que empleó tanto agar MacConkey como la cepa indicadora de Klebsiella pneumoniaeATCC 700603, mostró los mejores valores de sensibilidad y especificidad para la detección de carbapenemasas en Pseudomonas aeruginosa (90,0% y 85,7%, respectivamente). Conclusiones: la implementación de las variaciones del test de Hodge modificado podría ser una alternativa económica y de fácil realización para la detección fenotípica de carbapenemasas en Pseudomonas aeruginosa en los laboratorios de Microbiología Clínica.

Introduction: one of the most important mechanisms of carbapenem resistance in Pseudomonas aeruginosa is the production of carbapenem-hydrolyzing enzymes known as carbapenemases. The modified Hodge test is the most frequently used phenotypic screening test for carbapenemases. Nevertheless,CLSI recommends using modified Hodge test only in members of Enterobacteriaceae, sincethe test has demonstrated limitations in other Gram-negative bacilli and non-glucose-fermenters as Pseudomonas aeruginosa. Objective: To evaluate the performance of 3D test and three variations of modified Hodge test to detect carbapenemases in Pseudomonas aeruginosa isolates. Materials and methods: The efficacy of 3D test and three variations in modified Hodge test were evaluated taking polymerase chain reaction for carbapenemases KPC, VIM, IMP, NDM and OXA-48 as the gold standard. Variations consisted in using a) MacConckey agar instead of Mueller Hinton, b) Klebsiellapneumoniae ATCC 700603 as indicator carbapenem-sensitive strain instead of Escherichia coli ATCC 29212 and c) last two conditions simultaneously. Results: Of the variations tested, the third assay using both MacConckey agar and Klebsiella pneumoniae ATCC 700603 showed the best sensitivity and specificity (90.0% and 85.7%, respectively). Conclusions: The implementation of variations in modified Hodge test could be a cheap and easy to perform alternative for phenotypic carbapenemase detection in Pseudomonas aeruginosa in Clinical Microbiology laboratories.

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OBJECTIVE: Enterobacteriaceae bacteria harboring Klebsiella pneumoniae carbapenemase are a serious worldwide threat. The molecular identification of these pathogens is not routine in Brazilian hospitals, and a rapid phenotypic screening test is desirable. This study aims to evaluate the modified Hodge test as a phenotypic screening test for Klebsiella pneumoniae carbapenemase. METHOD: From April 2009 to July 2011, all Enterobacteriaceae bacteria that were not susceptible to ertapenem according to Vitek2 analysis were analyzed with the modified Hodge test. All positive isolates and a random subset of negative isolates were also assayed for the presence of blaKPC. Isolates that were positive in modified Hodge tests were sub-classified as true-positives (E. coli touched the ertapenem disk) or inconclusive (distortion of the inhibition zone of E. coli, but growth did not reach the ertapenem disk). Negative results were defined as samples with no distortion of the inhibition zone around the ertapenem disk. RESULTS: Among the 1521 isolates of Enterobacteriaceae bacteria that were not susceptible to ertapenem, 30% were positive for blaKPC, and 35% were positive according to the modified Hodge test (81% specificity). Under the proposed sub-classification, true positives showed a 98% agreement with the blaKPC results. The negative predictive value of the modified Hodge test for detection was 100%. KPC producers showed high antimicrobial resistance rates, but 90% and 77% of these isolates were susceptible to aminoglycoside and tigecycline, respectively. CONCLUSION: Standardizing the modified Hodge test interpretation may improve the specificity of KPC detection. In this study, negative test results ruled out 100% of the isolates harboring Klebsiella pneumoniae carbapenemase 2. The test may therefore be regarded as a good epidemiological tool.

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