CanID-PCR: a quick and low-cost PCR tool to identify Candida species on gDNA directly extracted from positive blood...

Candida bloodstream infection carries high mortality, and antifungal-resistant species, such as Candida auris and C. glabrata, are increasingly common. Delays in species identification can adversely affect patient outcomes. Conventional workflows by clinical microbiology laboratories require ~24 hours for Candida speciation to allow isolate growth on agar plates. We developed CanID-PCR, a simple, inexpensive PCR-based tool for rapid Candida identification directly from positive blood culture bottles. The assay targets the ACT1 gene, which contains a variable-length intron enabling discrimination among nine common Candida species. CanID-PCR was optimized for genomic DNA extracted directly from positive blood culture bottles and tested on 64 patient samples. Overall, species identification matched the clinical microbiology laboratory with 99% of concordance. One limitation was the inability to distinguish C. parapsilosis from C. metapsilosis, which are highly similar members of the C. parapsilosis complex. The unique discrepancy involved a failure to detect C. tropicalis in a mixed infection with C. glabrata. Among a total of seven samples with two species, one sample had a second species: C. fabianii (Cyberlindnera fabianii), detected only by CanID-PCR, but was missed by the clinical laboratory. These findings demonstrate that CanID-PCR can reveal polymicrobial Candida infections often overlooked when only a few selected colonies are analyzed. CanID-PCR provides rapid, accurate Candida speciation without the need for additional culture, requiring only standard laboratory equipment. This low-cost, straightforward approach is suitable for research laboratories, resource-limited clinical settings, or scenarios where timely species-level identification is critical.IMPORTANCEIdentifying species in Candida infections is of utmost importance to clinicians and researchers alike. Here, we describe CanID-PCR: a quick and low-cost PCR tool to identify Candida species on genomic DNA directly extracted from positive blood cultures. This technique uses basic laboratory equipment, and there is no need for high investment in special equipment. The tool allows identification of the nine most prevalent species in candidiasis. The technique is simple and can be used without the need for an additional culture to isolate the cells. It is noteworthy to mention that this tool can detect polymicrobial infections caused by more than one Candida species.