Virulence and double-stranded RNA in Sphaeropsis sapinea

Forty wildtype isolates of Sphaeropsis sapinea were grouped into the morphotypes A and B based on previously defined differences in cultural and morphological criteria as well as restriction sites for Dde I and Bst UI endonucleases in nuclear ribosomal DNA amplicons. Thirteen of 20 type A isolates and nine of 20 type B isolates contained detectable dsRNA (55%) of different molecular weight and size. dsRNA was transmitted into conidia at a frequency of 71-100%. By selecting single conidia, dsRNA-free subcultures were obtained from six of 22 isolates containing dsRNA. Pathogenicity tests on expanding buds of landscape trees of three species of Pinus showed highly significant statistical interactions between isolate virulence, Pinus species, and year. Pine species-year had a profound impact on virulence. The pattern in the interactions was revealed by principal component analysis of the interaction sums of squares of the anova (Additive Main Effects and Multiplicative Interaction; AMMI). Pinus sylvestris was highly interactive in its susceptibility to S. sapinea with seasonal effects. P. nigra and P. resinosa were more stable. The interactivity analysis was used to apportion interaction to specific isolates to improve the accuracy of the estimates of virulence. Estimates of the relative virulence of isolates were predicted over five different Pinus species-years. Isolates were ranked in virulence and interactivity using the AMMI model. This model permitted mean separation tests of the relative virulence among isolates over the combined Pinus species-years. One isolate was identified as potentially having dsRNA-mediated hypovirulence based on the significantly greater virulence of its isogenic, dsRNA-free subculture, as expressed over the three Pinus species and 2 years. Type A isolates containing dsRNA ranged from stable to highly interactive and from low to high in virulence. Type B isolates containing dsRNA were similar in interactivity but virulence ranged from avirulent to moderate, seldom exceeding the mean for S. sapinea . dsRNA-free isogenic subcultures tended not to express higher virulence than their dsRNA-containing parent strains but often changed in interactivity. Therefore, in one year a dsRNA-free subculture might be more virulent than its dsRNA-containing parent. In another year the dsRNA-free subculture might be less virulent.