Species identification of European forest pathogens of the genus Milesina (Pucciniales) using urediniospore morphology and molecular barcoding including M. woodwardiana sp. nov.

Abstract

Species of rust fungi of the genus Milesina (Pucciniastraceae, Pucciniales) are distributed mainly in northern temperate regions. They host-alternate between needles of fir (Abies spp.) and fronds of ferns (species of Polypodiales). Milesina species are distinguished based on host taxonomy and urediniospore morphology. In this study, 12 species of Milesina from Europe were revised. Specimens were examined by light and scanning electron microscopy for urediniospore morphology with a focus on visualising germ pores (number, size and position) and echinulation. In addition, barcode loci (ITS, nad6, 28S) were used for species delimitation and for molecular phylogenetic analyses. Barcodes of 72 Milesina specimens were provided, including 11 of the 12 species. Whereas urediniospore morphology features were sufficient to distinguish all 12 Milesina species except for 2 (M.blechni and M.kriegeriana), ITS sequences separated only 4 of 11 species. Sequencing with 28S and nad6 did not improve species resolution. Phylogenetic analysis, however, revealed four phylogenetic groups within Milesina that also correlate with specific urediniospore characters (germ pore number and position and echinulation). These groups are proposed as new sections within Milesina (sections Milesina, Vogesiacae M. Scholler & Bubner, sect. nov., Scolopendriorum M. Scholler & Bubner, sect. nov. and Carpaticae M. Scholler & Bubner, sect. nov.). In addition, Milesinawoodwardiana Buchheit & M. Scholler, sp. nov. on Woodwardiaradicans, a member of the type section Milesina, is newly described. An identification key for European Milesina species, based on urediniospore features, is provided.

Species diversity of Basidiomycota

“…New species of rust will certainly be found in isolated, under-explored, biodiverse areas, as well as through discovery of cryptic diversity in species complexes. For example, species diversity of rust fungi has increased through resolution of taxa in Chrysomyxa (Feau et al 2011), Coleosporium (McTaggart and Aime 2018), Dasyspora (Beenken et al 2012), Endoraecium (McTaggart et al 2015, Gymnosporangium (Zhao et al 2016a, b), Melampsora (Toome and Aime 2015;Zhao et al 2017a), Milesina (Bubner et al 2019), Neophysopella (Chatasiri and Ono 2008), Phakopsora (Beenken 2014;Maier et al 2016), Phragmidium (Liu et al 2018), Puccinia (Demers et al 2017;Hambleton 2010, 2013), Pucciniastrum (Liang et al 2006), Ravenelia Ebinghaus et al , 2020 and Uromycladium (Doungsa-ard et al 2018) (Fig. 12).…”

Studies on European rust fungi, Pucciniales: molecular phylogeny, taxonomy, and nomenclature of miscellaneous genera and species in Pucciniastraceae and Coleosporiaceae

“…Sequences were aligned with the CrustalW algorithm implemented in the programme Bio Edit, version 7.1.3.0 (Hall 1999), using the standard parameters offered by the programme. Two sequences of Milesina kriegeriana (Bubner et al 2019) were used as the outgroup. Alignments were used for phylogenetic reconstruction by three different methods:…”

“…Germ pores of urediniospores of Pucciniastrum epilobii s. lat. were visualised by the method described in Bubner et al (2019). For scanning electron microscopy (SEM), uredinia and urediniospores of dried specimens of P. epilobii s. lat.…”

“…and Rosaceae rusts within Coleosporiaceae using light and electron microscopy. These features have recently been documented for species and sectional delimitation in Milesina (Bubner et al 2019) but have not been considered in morphological studies of P. epilobii s. lat. (Klebahn 1900(Klebahn , 1914Bisby 1916;Pady 1933) and the rusts on Rosaceae (Ziller 1974;Helfer 2005;Qi et al 2019).…”

Delimiting species in Basidiomycota: a review

“…The LSU has been used as an alternative barcode for rust fungi instead, alone or with other markers (e.g., Bennett et al 2011;McTaggart et al 2015a;Maier et al 2016;Ebinghaus et al 2018). Depending on the genus, different barcodes may provide better species resolution (see discussion in Bubner et al 2019). McTaggart and Aime (2018) wrote for Coleosporium that the ITS2 spacer region may vary where LSU is unable to differentiate among species.…”