A rapid multiplex nucleic acid detection system of airborne fungi by an integrated DNA release device and microfluidic chip.

ABSTRACT

Occupational health problems, such as asthma, in specific work environments arise from the presence of airborne fungi. Rapid detection of pathogenic airborne fungi is therefore important to reduce or avoid any adverse effects on staff health. Herein, we established a new integrated rapid Lyticase-Motor-Chemical reagent nucleic acid releasing (LMC) method for the release of fungal DNA. Aspergillus fumigatus, Aspergillus flavus, and Cryptococcus neoformans were chosen to evaluate the LMC method. The results of Loop-Mediated Isothermal Amplification (LAMP) analyses showed that this method could release the nucleic acid of 4 × 104 fungal spores, equaling to 400 copies per microliter. This rapid multiplex nucleic acid detection system of airborne fungi included an integrated DNA release device and a portable microfluidic chip. The integrated DNA release device combined mechanical lysing and biochemical reagent treatment to automate DNA release. The microfluidic chip was capable of multiplex nucleic acid detection. The detection limit of this system was 4 × 104 spores per test, meeting the requirement of early warnings. The whole analysis from the sample input to readout could be completed within 90 min, including 30 min for fungal DNA release and 45 min for LAMP analysis. The integrated DNA release device and microfluidic chip were portable, showing tremendous potential in point-of-care tests of airborne fungi.