An In-house PCR Workflow for Rapid Detection of <i>Candida </i>spp. from Blood Cultures of Septicemia Suspected Patients

Background: Candidemia is a leading cause of nosocomial bloodstream infections, with conventional culture methods often requiring 48 - 72 hours for species identification and showing suboptimal sensitivity. The rapid and accurate detection of Candida spp. is crucial for timely antifungal therapy and improved patient outcomes. Objectives: The present study aimed to design and validate a laboratory-developed polymerase chain reaction (PCR) assay for the rapid detection of Candida species in culture-positive blood specimens, specifically targeting growth-positive blood culture bottles. Methods: In this cross-sectional study, conducted between November 2024 and March 2025 at Imam Reza Hospital in Mashhad, 200 growth-positive blood culture bottles flagged by BacT/ALERT or conventional systems were analyzed using an in-house PCR assay targeting the hyphal wall protein 1 (HWP1) gene. Analytical sensitivity was determined through ten-fold serial dilutions of confirmed Candidaalbicans DNA, and the assay performance was compared to conventional culture results. Results: The HWP1-PCR assay successfully detected Candida spp. DNA in 10 of 200 samples (5.0%; 95% CI: 2.7 - 9.0%), all confirmed as C. albicans by sequencing. The assay demonstrated 100% sensitivity and specificity, with no false-positive or false-negative results. The limit of detection (LOD) was 0.0174 ng/μL for genomic DNA. Conclusions: The HWP1-PCR assay shows excellent efficacy for the rapid identification of C. albicans, reducing the identification time by over 48 hours compared to traditional methods. This assay enhances the timeliness of antifungal treatment and is suitable for resource-limited settings. Future research should focus on multicenter validation and antifungal susceptibility testing to assess the impact of these findings on treatment strategies.