Title : Development of a syndromic molecular diagnostic assay, using barcoded magnetic bead technology for tick-borne pathogens
Abstract:
The infectious disease diagnostics often depends on costly serological testing with poor sensitivity, low specificity, and long turnaround time. Here In this manuscript, we demonstrate the proof of the principle for simultaneous detection of two tickborne pathogens from a single patient sample using barcoded magnetic bead technology on the BioCode® 2500 system. Specific primer sets complementary to the conserved genes of Anaplasma phagocytophilum and Borrelia Burgdorferi were used in PCR amplification of the target, followed by the hybridization of the resulting biotinylated PCR products with specific probes tethered to the barcoded magnetic beads for simultaneous detection, using a fluorophore with high quantum yield. The assay has extremely high signal to background ratio and detects up to two copies of the target in a test reaction with high sensitivity and specificity. The assay demonstrated 100 percent positive and negative agreements on performance evaluation using patient specimens and blood samples spiked with tittered pathogens. No cross reactivity was observed with other related tickborne pathogens and genomic DNA of human, cattle and canine origin. The assay can be upgraded to a sensitive and cost-effective multiplex diagnostic approach that can simultaneously detect all clinically important tickborne pathogens in a single sample with a short turnaround time.
