In 1999, a severe outbreak (i.e., 100% infection) of a virus disease was observed in a single field of common bean in Colusa County, CA. The symptoms included a yellow mosaic, leaf epinasty and, in some plants, a systemic necrosis. This field was adjacent to a clover field that had been harvested early in the development of the bean plants. A preliminary serological test (enzyme-linked immunosorbent assay, ELISA) suggested that the virus infecting these bean plants was Peanut mottle virus (PeMoV). This would represent the first report of this virus in California. A range of common bean cultivars (Black Turtle Soup, Topcrop, California Early Light Red Kidney, and Sutter Pink) were inoculated with sap prepared from symptomatic leaves collected from this field. Symptoms developing on these plants ranged from systemic necrosis (cvs. Sutter Pink and Black Turtle Soup) to strong yellow green mosaic and leaf distortion (cvs. Topcrop and California Early Light Red Kidney). Furthermore, inoculated primary leaves of cv. Topcrop failed to develop local lesions, which is characteristic of PeMoV. ELISAs on all symptomatic plants with antisera against PeMoV, BYMV, BCMV, and BCMNV as well as reverse transcription polymerase chain reaction (RT-PCR) analysis with primer pairs specific for PeMoV, BYMV, BCMV, and BCMNV were negative. To further investigate the nature of this virus, a minipurification method was used to purify virions from symptomatic leaves of all four cultivars. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis of purified virions from these cultivars revealed a 32-kDa band consistent with infection by a potyvirus. Transmission electron microscopy analysis of these preparations revealed the presence of potyvirus-like flexous rods (approximately 750 nm long and 10 nm wide). We next designed a primer pair specific for the coat protein gene of Clover yellow vein virus (ClYVV) and RT-PCR with these primers resulted in the amplification of a 630-bp DNA fragment from four isolates of the unknown potyvirus. No fragments were amplified from an uninfected control. The PCR-amplified fragments were direct-sequenced, and sequence comparisons revealed that the sequences of all four isolates were 95% identical to that of ClYVV (Genbank accession number D89541). Subsequently, a ClYVV antiserum was obtained from Simon Scott (Department of Plant Pathology, Clemson University), and ELISAs performed on leaves infected with all four isolates were positive. Finally, to assess whether the virus was seed-transmitted, seed harvested from this field was planted in a greenhouse (two lots of 400 seed each). None of the plants from these seeds developed virus symptoms, suggesting that the virus was not seed-transmitted. Together, these results indicate that the virus disease outbreak in this bean field was caused by ClYVV rather than PeMoV. The inoculum source for the virus was probably the adjacent clover field. This is the first report of ClYVV infecting common bean in California.
Outbreak of Clover yellow vein virus in a Bean Field in Colusa County, California
