Highly sensitive and species-specific molecular detection of microbial pathogens

This study describes the development of a highly sensitive and specific endpoint PCR-based method for detecting microbial pathogens, including two Eukarya and one bacterial species. We developed an unbiased computer-assisted protocol to design oligonucleotides targeting repeated DNA sequences. This approach enables rapid endpoint PCR amplification using reduced sample input compared to current conventional methods. We successfully detected three human microbial pathogens: Candida glabrata, Candida albicans, and Pseudomonas aeruginosa with high specificity and sensitivity, enabling the detection of genomic DNA (gDNA) contained in as few as 1-10 cells of the two fungal pathogens tested. These findings suggest that this methodology is broadly applicable to any microorganism for which genome sequence data are available and that contains repeated DNA elements, highlighting its potential as a universal pathogen detection strategy.