Quantitative assays of two soil-borne pathogens of Aconitum carmichaelii Debx., Sclerotium rolfsii and Mucor circinelloides, in the main cultivation areas of China

Aconitum carmichaelii Debx. is a typical traditional Chinese medicinal herb that has been cultivated for more than 1000 years in China. Sclerotium rolfsii and Mucor circinelloides have been reported to be pathogenic to A. carmichaelii, limiting the production and quality. Rapid diagnostic methods of these pathogens are required for disease prediction and management. Herein, we developed two real-time quantitative polymerase chain reaction (PCR) methods to detect and quantify S. rolfsii and M. circinelloides. The specificity of the primers was confirmed by using the NCBI online blast tool and by testing against phylogenetically closely related species and common soil-borne fungal pathogens. Sensitivity was determined, and the standard curves were generated for S. rolfsii and M. circinelloides, respectively, using the plasmid DNA containing the target amplicons. Using the developed methods, we performed quantitative detections on the soil and root samples of A. carmichaelii collected from 22 different fields in seven provinces of China. It was found that S. rolfsii and M. circinelloides were widely distributed in the main cultivation areas of A. carmichaelii, and M. circinelloides was a more abundant and more significant disease agent for A. carmichaelii. The developed techniques will be helpful in monitoring pathogens and facilitate the disease control and quality improvement of A. carmichaelii products.