A HYPOVIRULENT ISOLATE OF CRYPHONECTRIA-PARASITICA WITH MULTIPLE, GENETICALLY UNIQUE DSRNA SEGMENTS

Many different dsRNA-containing isolates of Cryphonectria (Endothia) parasitica, the chestnut blight fungus, have been identified in surveys conducted in the central Appalachians. Isolate C-18, collected in 1989 from a virulent canker in southern West Virginia, was found to contain 11 segments of dsRNA ranging from 1 to 5 kb in size. Virulence experiments, using Golden Delicious apples, excised dormant American chestnut stems, and American chestnut sprouts, determined that the dsRNA associated with isolate C-18 significantly reduced virulence and altered cultural morphology when isogenic dsRNA-containing and dsRNA-free asexual progeny were compared. The 11 dsRNA segments were transmitted into other isolates of C. parasitica via hyphal anastomosis and into conidia in an all-or-none fashion. Complementary DNA clones from a library representing C-18 dsRNA did not hybridize to other dsRNAs associated with hypovirulent isolates D2 (Pennsylvania), Ep-713 (Europe), NB58-88 (New Jersey), to another multisegmented dsRNA from West Virginia (9B-2-1), or to wound tumor virus. None of the cDNA clones tested hybridized to more than one dsRNA segment by Northern blot analysis. Eight of the 11 dsRNA segments were demonstrated to be genetically unique. Using a phosphate buffer extraction protocol, icosahedral particles approximately 60 nm in diameter were purified from the mycelial tissue of isolate C-18. The dsRNA virus associated with hypovirulence of isolate C-18 is therefore fundamentally different from other hypovirulence-associated viruses characterized to date.