A new combination for Microcyclospora rumicis in Sphaerulina based on molecular and morphological data

The cercosporoid species, Microcyclospora rumicis Arzanlou & Bakhshi was recently introduced on Rumex crispus L., from Iran (Guilan province, Talesh) (Arzanlou & Bakhshi 2011). Based on the morphological features, this fungus was described as a species of the genus Microcyclospora J. Frank et al. (Arzanlou & Bakhshi 2011). During a phylogenetic study of cercosporoid fungi from north of Iran,the genomic DNA of thetype strain (CBS 131546) of the species, was extracted using the protocol developed by Möller et al. (1992). Amplification and sequencing of the internal transcribed spacer regions and intervening 5.8S nrRNA gene of the nrDNA operon (ITS) and partial nuclear ribosomal large subunit (LSU) were done with primers V9G + ITS4 and LR0R + LR5 respectively (Quaedvlieg et al. 2013). The resulting sequences were subjected to a BLAST search to find most similar sequences in GenBank. The obtained sequences from GenBank together with the novel generated sequences of this study, were aligned and subjected to multi-gene DNA sequence analyses using MrBayes v. 3.2.2. Based on the phylogenetic analyses, this fungus clusters together with Sphaerulina Sacc. species in a clade that is distinct (Fig. 1). Therefore, the fungus was transferred to the genus Sphaerulina and, a new combination, Sphaerulina rumicis is introduced here with following characteristics:

Sphaerulina rumicis (Arzanlou & Bakhshi) M. Bakhshi & Arzanlou, comb. nov. — MycoBank MB 823634.

Basionym: Microcyclospora rumicis Arzanlou & Bakhshi, Mycotaxon 118: 182. 2011.

Specimens examined. Iran, Guilan, Talesh, on leaves of Rumex crispus L., Sept. 2010, M. Bakhshi, CCTU 1 = CBS 131546.

The LSU and ITS sequences of the type strain (CBS 131546) were deposited under GenBank accessions No. MG561649 and No. MG561648, respectively.

Notes: Based on its morphological features including microcyclic conidiation, conidiophores reduced to conidiogenous cells, scolecosporous, guttulate conidia and, unthickened nor darkened hila, Arzanlou & Bakhshi (2011) placed this fungus in Microcyclospora. To resolve the phylogenetic placement of this fungus, we generated phylogenetic tree based on LSU and ITS DNA sequence data (Fig 1). These data clearly revealed that the fungus was unrelated to Microcyclospora and is shown to belong to Sphaerulina. The phylogeny of the genus Sphaerulina (with type speciesSphaerulina myriadea (DC.) Sacc.) has been clarified recently by Verkley et al. (2013) and Quaedvlieg et al. (2013). Other than the lack of pycnidial conidiomata, the morphological characters of the Sphaerulina rumicis are similar to other species of Sphaerulina (Verkley et al. 2013, Quaedvlieg et al. 2013). The placement of Microcyclospora rumicis in Sphaerulina according to the absence of pycnidial conidiomata, is somewhat notorious; however this phenomenon has also been observed in other species of fungi. For example, Crous et al. (2013) have shown that Septoria pistacinaAllesch., a major pathogen of pistachio, despite of producing pycnidial conidiomata, belongs to Pseudocercospora (P. pistacina (Allesch.) Crous). Therefore, this is another example that questions the value of conidiomatal structure at generic level, and emphasizes the need for integration of morphological features with molecular data for accurate identification of fungal species.

Fig. 1. Phylogenetic tree inferred by Bayesian analysis of the combined two-loci (LSU and ITS) sequence alignment using MrBayes v.3.2.2. The scale bar indicates 0.1 expected changes per site. Cladosporium allicinum (CPC 5101) was used as an outgroup.