Rapid detection of Mycobacterium tuberculosis based on antigen 85B via real-time recombinase polymerase amplification

Abstract Tuberculosis (TB), as a common infectious disease, still remains severe challenge to public health. Due the unsatisfied clinical needs of currently available diagnostic vehicles, it is desired establish new approach for universally detecting Mycobacterium tuberculosis. Herein, we designed real-time recombinase polymerase amplification (RPA) technology identifying M. tuberculosis within 20 min at 39°C via custom-designed oligonucleotide primers and probe, which could specifically target antigen 85B (Ag85B). Particularly, F4-R4 produced fastest fluorescence signal with probe among four pairs in RPA assays. The optimal primers/probe combination effectively identify detection limit 4·0 copies per ?l, not show positive genomic DNA from other mycobacteria or pathogens. Ag85B-based determine extracted high reliability (100%, 22/22). More importantly, when testing sputum samples, displayed an admirable sensitivity (90%, 95% CI: 80·0-96·0%) specificity (98%, 89·0-100·0%) compared traditional smear microscopy, was similar assay Xpert MTB/RIF. This based Ag85B provides promising strategy rapid universal diagnosis TB.