Fungal DNA-barcoding on a chip: Magnetoresistive biosensors for yeast infection diagnosis

The worldwide burden of fungal infection has been increasing due to the expansion of the endemic borders, the host range of certain mycosis, and the emergence of novel opportunistic, drug and multi-drug-resistant fungal pathogens. Many deaths can be attributed to slow, inaccurate, or absent diagnostic testing and a lack of timely administration of effective antifungal treatment. Panfungal PCR followed by sequence identification has the practical advantage of rapidly and accurately detecting slow-growing, unculturable and even rare, unexpected or novel pathogens. We have developed and tested a robust and flexible biosensing diagnostic strategy, consisting of panfungal rDNA amplification, DNA hybridization, and magnetic labeling on a portable microfluidic setup. We designed DNA-capture probes specific for Candida albicans, Candida auris, Nakaseomyces glabratus, and Cryptococcus neoformans, and evaluated them for diagnostic sensitivity and specificity assessment, first in silico against a dataset of 58 pathogenic species (1239 DNA sequences), and then in vitro against the four species and 30 unsequenced clinical isolates of N. glabratus. On-chip, our assay performed with 95% accuracy, 91% sensitivity, and 98% specificity under a limit of detection of 600 single-stranded amplified ITS DNA sequences. Our results demonstrate the diagnostic value of DNA-barcoding and magnetoresistive biosensing, emphasizing the revolutionizing potential of DNA-sequence-based technologies in clinical practice.