A LAMP-based microfluidic module for rapid detection of pathogen in cryptococcal meningitis.

Cryptococcal meningitis (CM) is a global threat with significant attributable morbidity and mortality. Information on microfluidic detection for CM diagnosis is still limited. We developed a multifunctional microfluidic module that integrated the pathogen enrichment and on-chip nucleic acid extraction. The module adopted a simple filtration membrane to effectively capture Cryptococcus cells and simplify the process, and combined lyticase digestion, alkaline lysis and heating methods to optimize the strategy to achieve nucleic acid extraction. The entire process was operated in the module, which reduced the exposure risk of directly processing cryptococcal samples. A portable one-pot lyophilized LAMP reagent bead with no temperature limit was developed, which improved the flexibility of operation. This module did not require any additional instrument, and is promising to develop a simple, rapid, and efficient approach to realize the "sample in and answer out" detection of real CSF samples. This microfluidic module had practical prospects and is expected to replace LFA for efficacy evaluation and follow-up in the middle and late stages of CM treatment, and could be used as an auxiliary method to confirm cases with questionable LFA results in the early diagnosis of CM.