Use of the Genealogical Sorting Index (GSI) to delineate species boundaries in the Neofusicoccum parvum–Neofusicoccum ribis species complex

[Display omitted] ► First application of the Genealogical Sorting Index to identify fungal species. ► Description of new species; Neofusicoccum occulatum in the N. parvum-ribis complex. ► Species from disturbed environments requires analysis from a global perspective. Neofusicoccum is a recently described genus of common endophytes and pathogens of woody hosts, previously placed in the genus Botryosphaeria. Many morphological characteristics routinely used to describe species overlap in Neofusicoccum, and prior to the use of molecular phylogenetics, isolates from different hosts and locations were often misidentified. Two cryptic species Neofusicoccum ribis and Neofusicoccum parvum were initially described from different continents and recently another four species within this complex were described using fixed nucleotide polymorphisms for differentiation. In a survey of eucalypt cankers in eastern Australia, a collection of morphologically similar Neofusicoccum isolates were obtained. This collection was analysed within the framework of the morphological (MSRC), ecological (ESRC) and phylogenetic (PSRC) species recognition concepts. Morphological data based on spore measurements (MSRC), together with pathogenicity trials (ESRC) were considered alongside molecular analysis (PSRC), which included multiple gene phylogenies constructed from four nuclear gene regions. We also used the Genealogical Sorting Index method to provide objective evidence for the status of terminal taxa in the phylogenetic analysis. The isolates examined exhibited overlapping morphological and culture characteristics, similar pathogenicity to excised stems and shared hosts within the same locations. Phylogenetic analysis separated isolates into 8 clades corresponding to six described species: N. ribis, N. parvum, Neofusicoccumkwambonambiense, Neofusicoccum cordaticola, Neofusicoccum umdonicola, Neofusicoccum batangarum, and two new species. GSI support indicated combined phylogenetic data were monophyletic for all clades and all p-values were significant allowing us to reject the null hypothesis that all groups were from a single mixed group. Consequently the description of Neofusicoccum occulatum is presented.