Developing PCR-based novel molecular assays to quantitatively detect Fusarium solani in ginseng soil for assessing soil health in ginseng cultivation

The fungal species of Fusarium solani (F. solani) is closely associated with the soil borne root rot disease of Panax ginseng, leading to quality decline in the medicinal materials. In this study, we established experiment conditions of two PCR-based microbiological assays, a droplet digital PCR(ddPCR) method and a RT-qPCR method, and tested their efficacy of quick and quantitative detection of F. solani DNA in ginseng soil. These methods were further evaluated in quantifying F. solani for rhizosphere samples of ginseng plants with different status of pathogenic disease infection. The result showed that the two methods were both highly specific and reproducible in detecting and quantifying the gene of F. solani, without any observable cross-reaction with other species. Compared to the qPCR method in this study, ddPCR method exerted a higher sensitivity of detection (0.92 copies μL−1) than qPCR method (920 copies μL−1), and free of reliance on a standard calibration curve. With ddPCR method, the absolute quantification of nucleic acid samples is made possible by changing the detection standard from fluorescence intensity to the presence or absence of fluorescence, and the accuracy of detection has been significantly improved. The abundance of F. solani were 0−2,100 copies g−1 dws, 927.5−8,470 copies g−1 dws and 10,605−43,697 copies g−1 dws respectively in uncultured ginseng soil, soil from healthy ginseng and the soil from infected ginseng. The quantitative abundance of the pathogenic fungal species infected by the diseases but also track the dynamic change in the gene abundance of F. solani.in disease monitoring with ginseng growth, While the gene abundance of F. solani was highly accumulated in mid-August and decreased in September, the pathogenic fungal gene abundance tested was closely correlated to the incidence rate of ginseng root rot. Apparently, this pathogenic fungus is widely existing in ginseng soil even with the healthy plants in September, indicating a potentially great risk of high disease incidence for ginseng cultivation. Therefore, the developed dd PCR assay could be reliable and feasible, potentially applied in early diagnosis of the infection by the pathogenic fungal of F. solani in ginseng cultivation and in routine assessment of the disease occurrence with ginseng growth.