Multiplex RT-PCR detection and distribution of four apple viruses in China

Apple trees are natural hosts of four economically important virus species in China. We used a simple, sensitive multiplex RT-PCR protocol with an internal control to simultaneously detect and differentiate four apple viruses: apple chlorotic leaf spot virus (ACLSV), apple mosaic virus (ApMV), apple stem pitting virus (ASPV), and apple stem grooving virus (ASGV). This multiplex RT-PCR could be used as an alternative to other routinely used detection methods. We used this protocol to evaluate the occurrence and distribution of the four apple viruses in China. The four viruses were widely distributed throughout the main apple production region of China, including Heilongjiang, Liaoning, Hebei, Beijing, Henan, Shanxi, Shaanxi, Shandong, Gansu, Ningxia, Xinjiang, Sichuan, and Yunnan. The four viruses, ApMV, ASPV, ASGV, and ACLSV, were present in 80.1%, 65.1%, 73.7%, and 69.7% of the samples, respectively. Two or more of the viruses were detected in most of the infections. The most frequent virus combinations were ApMV + ASGV + ASPV + ACLSV with an incidence of 27.22%, followed by ApMV + ASGV + ACLSV (14.37%), ApMV + ASGV + ASPV (12.54%), and ASGV + ApMV (11.01%). The incidence of the ASPV + ACLSV combination was the lowest (0.61%). This is the first extensive survey conducted in China for monitoring the four apple viruses, which provides important information for apple virus distribution and management in China.