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Diplodia scrobiculata and Diplodia pinea are endophytic fungi associated with dieback and cankers of mainly Pinus spp. in many parts of the world. These two fungi are closely related and have, in the past, been considered to represent two morphological forms (A and B morphotypes) of D. pinea. dsRNA elements are known to occur in both D. scrobiculata and D. pinea. Two dsRNA elements from D. pinea, SsRV1 and SsRV2, have been characterized previously. The aim of this study was to characterize a third dsRNA element that is most commonly associated with D. scrobiculata and to determine its phylogenetic relationship to other mycoviruses. The 5018-bp genome of this element was sequenced, and it is referred to as D. scrobiculata RNA virus 1, or DsRV1. It has two open reading frames (ORFs), one of which codes for a putative polypeptide with a high degree of similarity to proteins of the vacuolar protein-sorting (VPS) machinery, and the other for an RNA-dependent RNA polymerase (RdRp). Phylogenetic comparisons based on amino acid sequence alignments of the RdRp revealed that DsRV1 is closely related to a dsRNA element isolated from Phlebiopsis gigantea (PgV2), and they grouped separately from virus families in which mycoviruses have previously been described. Although D. pinea and D. scrobiculata are closely related, DsRV1 does not share high sequence identity with SsRV1 or SsRV2, and they probably have different recent evolutionary origins.

After recent changes to the taxonomy of the Botryosphaeriaceae species with diplodia-like (= dark, ovoid, often pigmented) conidia are considered to belong to at least three genera including Diplodia, Lasiodiplodia and Dothiorella. In a recent molecular phylogenetic study it became apparent that two groups of isolates with diplodia-like conidia required taxonomic revision. One group of isolates originated from Cupressus sempervirens in Greece and Cyprus and had been identified as D. pinea f. sp. cupressi based on morphological characteristics. The other isolates originated from a Casuarina sp. in Australia and were superficially similar to those in the first group based on their morphologically similar diplodia-like conidia. The aim of this study was to resolve the taxonomy of these two groups of isolates by combining the information from the multiple gene genealogies with morphological characters. The results showed that the isolates from C. sempervirens in Greece and Cyprus represent D. cupressi. The isolates from Casuarina in Australia belong to the more distantly related genus Dothiorella and represent a distinct species that is described here as Do. casuarini sp. nov.

Three anamorph genera of the Botryosphaeriaceae namely Diplodia, Lasiodiplodia and Dothiorella have typically dark, ovoid conidia with thick walls, and are consequently difficult to distinguish from each other. These genera are well-known pathogens of especially pine species. We generated a multiple gene genealogy to resolve the phylogenetic relationships of Botryosphaeriaceae with dark conidial anamorphs, and mapped host associations based on this phylogeny. The multiple gene genealogy separated Diplodia, Lasiodiplodia and Dothiorella and it revealed trends in the patterns of host association. The data set was expanded to include more lineages of the Botryosphaeriaceae, and included all isolates from different host species for which ITS sequence data are available. Results indicate that Diplodia species occur mainly on gymnosperms, with a few species on both gymnosperms and angiosperms. Lasiodiplodia species occur equally on both gymnosperms and angiosperms, Dothiorella species are restricted to angiosperms and Neofusicoccum species occur mainly on angiosperms with rare reports on Southern Hemisphere gymnosperms. Botryosphaeria species with Fusicoccum anamorphs occur mostly on angiosperms with rare reports on gymnosperms. Ancestral state reconstruction suggests that a putative ancestor of the Botryosphaeriaceae most likely evolved on the angiosperms. Another interesting observation was that both host generalist and specialist species were observed in all the lineages of the Botryosphaeriaceae, with little evidence of host associated co-evolution.

Sphaeropsis sapinea is an opportunistic pathogen causing serious damage to conifers, pre-disposed by adverse environmental conditions or mechanical damage. Three different morphological forms of the fungus have been described and are commonly referred to as the A, B and C morphotypes. Isolates of the different morphotypes have also been separated based on differences in pathogenicity and molecular characteristics. These differences, however, overlap and have not been considered sufficiently robust to justify the description of separate taxa. The aim of this study was to consider relationships between isolates representing different S. sapinea morphotypes, using multiple gene genealogies inferred from partial sequences of six protein-coding genes and six microsatellite loci. Genealogies generated for the protein-coding genes and microsatellite loci were not congruent but both consistently grouped isolates representing the A and C morphotypes in separate but closely related clades. In contrast, isolates of the B morphotype grouped together in a clade that was equally different to the A and C morphotypes as it was to the clade encompassing isolates of Botryosphaeria obtusa. These results provide strong evidence to show that the B morphotype isolates are distantly related to S. sapinea and represent a discrete taxon, which we describe here as Diplodia scrobiculata sp. nov.