Detection and identification of Candida spp. in bloodstream infections of pediatric patients admitted in intensive care units: validation of a multiplex PCR

Abstract

Disseminated candidiasis is an important cause of morbidity and mortality in immunocompromised and critically ill pediatric patients admitted at intensive care units. Laboratory diagnosis of candidemia is troublesome due to the lack of blood cultures sensitivity. Detection of Candida DNA by polymerase chain reaction (PCR) in tiny blood volumes may provide a more rapid and reliable candidemia detection in critically ill children. The aim of this study is to validate a multiplex PCR technique for detection and identification of seven Candida species in pediatric patients bloodstream at risk to develop fungemia, and comparing results with those obtained by blood cultures. Blood samples collected from 30 newborns admitted at Neonatal Intensive Care Unit and 30 patients admitted at Pediatric Intensive Care Unit of Instituto da Criança-HCFMUSP, presenting with predisposing conditions to develop candidemia will be analyzed. A control group composed of 30 non infected children, admitted at the pediatric surgery unit of the same hospital to undergo small cirurgical interventions will be included. The limit of detection and the specificity of the multiplex PCR will be determined employing species-specific primers designed in the present study. DNA extracted from blood samples will be submitted to amplifications and PCR products analyzed by agarose gel electrophoresis. Agreement between both methods will be assessed by the McNemar test. If PCR positive results are not confirmed by blood cultures, DNA sequencing will be performed. (AU)