Mungbean (Vigna radiata L.), an important leguminous food crop in China, is popularly grown in arid regions. The total area of mungbean production is 8.0 × 105 ha. In August and September 2010, wilted symptoms were observed in mungbean plants in Yulin, Shaanxi Province and Datong, Shanxi Province. Infected plants had silvery gray coloration of stems and lateral branching with senesced leaves still attached to the plant. Dark brown necrotic areas were observed on the exterior of the taproot underneath the epidermis and in the pith of the lower stems of wilted plants. Black spherical microsclerotia, 43.9 μm, were present in infected plant tissues. Six fungal isolates were cultured on potato dextrose agar at 25°C and the mycelium was initially hyaline but later became gray. Black microsclerotia, 60 to 80 × 75 to 123 μm, were observed in culture after 2 to 7 days of incubation. On the basis of field symptoms, colony color, and the size of microsclerotia, the fungus was identified as Macrophomina phaseolina (Tassi) Goid (3). To confirm the morphological identification, the rDNA internal transcribed spacer (ITS) regions of the six isolates were amplified with universal primers ITS1 and ITS4. The resulting ITS sequences of the six isolates (GenBank Accession Nos. HQ660589, HQ660590, HQ660591, HQ660592, HQ660593, and HQ660594) were aligned in GenBank, which showed 97 to 99% identity with 60 M. phaseolina isolates (e.g., Accession Nos. GU046867, FJ415067, and FJ960441). Using the PCR primers MpKF1 (5′-CCGCCAGAGGACTATCAAAC-3′) and MaKR1 (5′-CGTCCGAAGCGAGGTGTATT-3′) specific for M. phaseolina (1), a 350-bp PCR fragment was obtained, indicating that these isolates were M. phaseolina. Pathogenicity tests of six isolates were performed by inoculation of 3-week-old seedlings of cv. Zhonglv 8 using the hypocotyl inoculation technique, respectively (2). Each isolate was transferred to petri dishes containing PDA 2 days prior to inoculation. On the day of inoculation, an inoculum slurry was prepared by cutting agar with the pathogen into small strips and passing the strips through a 5-syringe until uniform. A small quantity of inoculum extruded into the vertical cut in each hypocotyl of at least eight seedlings in each pot, and the PDA was used as the control to extrude into the vertical cut in each hypocotyl of at least eight seedlings in another pot. The inoculated and control plants were incubated in the mist chamber at 25°C and 90 to 100% relative humidity for 48 h before growing in a greenhouse at 30°C. Six days after inoculation, all inoculated plants, wilted or dead, showed dark brown-to-black lesions. No symptoms were observed on the control plants. For each isolate tested, M. phaseolina was reisolated from inoculated plants, but was not isolated from the control plants. The fungus has been detected in 29 plant species of 23 genera in China, including the major crops Arachis hypogaea, Helianthus annuus, and Glycine max. Although M. phaseolina has caused great yield reduction of mungbean in many areas of Asia, to our knowledge, this fungus as a causal agent of mungbean charcoal rot has not previously been reported in China.
Reference: (1) B. K. Babu et al. Mycologia 99:797, 2007. (2) D. L. Pazdernik et al. Plant Dis. 81:1112, 1997. (3) G. S. Smith and T. D.Wyllie. Charcoal rot. Page 29 in: Compendium of Soybean Diseases. 4th ed. G. L. Hartmann et al., eds. The American Phytopathological Society, St. Paul, MN, 1999.