Elsevier

Mycological Research

Volume 111, Issue 11, November 2007, Pages 1268-1276
Mycological Research

The polyphyletic nature of Pleosporales: an example from Massariosphaeria based on rDNA and RBP2 gene phylogenies

https://doi.org/10.1016/j.mycres.2007.08.014Get rights and content

Abstract

Massariosphaeria is a loculoascomycetous fungus currently accommodated within the Pleosporales. However, based on morphology alone, it has been difficult to assess its familial position and its affinities to other fungi with bitunicate asci. In order to establish its evolutionary relationships, two regions of the rDNA (18S and 28S) and two regions of the RPB2 protein-coding gene were sequenced and analysed phylogenetically. Multigene phylogenies revealed that Massariosphaeria is not monophyletic and results are in disagreement with existing morphological-based classification schemes. Characters, such as ascomatal shape and ascospore morphology, have evolved more than once within the Pleosporales. The familial placement of several species is still obscure, except M. grandispora, which could be confidently assigned to the Lophiostomaceae. M. typhicola is closely related to Trematosphaeria hydrela (Melanommataceae), whereas M. triseptata is related to Melanomma radicans but shares close affinities to the Sporormiaceae. The placement of M. roumeguerei is still unresolved, and it does not appear to have any close evolutionary relationship to any known melanommataceous or pleosporaceous genera. Our molecular data also refute the monophyly of Kirschsteiniothelia, Massarina, Melanomma, and Pleospora, and support previous phylogenetic hypotheses that Melanommataceae is polyphyletic. There is a need for more phylogenetic (and taxonomic) studies within the Pleosporales, especially incorporation of more anamorphic taxa and type species.

Introduction

The genus Massariosphaeria (Pleosporales, Dothideomycetes) is characterised by a typical pleosporaceous morphology (Leuchtmann 1987), which include: black, bean-shaped or elongated, immersed (sometimes semi-immersed) ascomata; black and papillate ostioles; a thin-walled, smooth peridium (textura prismatica) with black to grayish cells; abundant, hyaline, septate hamathecium filaments; and bitunicate, cylindric-clavate asci. It is morphologically a relatively well-characterised genus, easily recognisable by the fusiform, multiseptate ascospores with relatively large thick walls, yellow to brown colour, some with transverse septum only, with the cell above the septum the largest, and surrounded by a prominent mucilaginous sheath (Tanaka and Harada, 2004, Van, 2005). All species have a tendency to form red pigments, on the host and especially in culture (Van 2005). The genus was first established as a section of Leptosphaeria (Müller 1950), but was later given generic status. This taxonomic arrangement has largely been accepted (Leuchtmann, 1984, Huhndorf et al., 1990), although Barr (1989) included Massariosphaeria in Chaetomastia. Currently, there are about 28 accepted species Index Fungorum, June 2006 and both phragmosporous and dictyosporous taxa are included (Leuchtmann, 1987, Shoemaker and Babcock, 1989, Tanaka and Harada, 2004).

Lophiostoma, Leptosphaeria, Melanomma, and Trematosphaeria are pleosporaceous genera with species that are morphologically similar to various Massariosphaeria species. This has resulted in taxonomic uncertainty with species being transferred from one genus to another (e.g. Shoemaker and Babcock, 1989, Huhndorf et al., 1990, Tanaka and Harada, 2003, Tanaka and Harada, 2004). The familial position of Massariosphaeria is still not well defined. This is partly because the circumscription of other families within the order Pleosporales is still unclear. Massariosphaeria is generally accepted to belong to the family Lophiostomaceae (Eriksson and Hawksworth, 1991, Kirk et al., 2001). However, it has also been referred to the family Dacampiaceae by Barr (1992), no doubt because she included some species of Massariosphaeria in Chaetomastia (Barr 1989). Most of the known anamorphic Massariosphaeria species are aposphaeria-like, which is also the anamorph produced by Melanomma (Melanommataceae) (Kirk et al., 2001, Tanaka and Harada, 2004).

On morphological grounds, it has been very difficult to predict the familial placement, and to date, only one species of Massariosphaeria (M. phaeosphaeria, the type species), has had its 18S rDNA partially analysed (Liew et al. 2000). It is thus unknown whether the genus is mono- or polyphyletic. This issue needs to be addressed as there have been a few reports on polyphyly of some pleosporaceous genera. (e.g. Kodsueb et al., 2006a, Liew et al., 2002). This paper is a continuity of several taxonomic studies on the Pleosporales where we have targeted important dothideomycetous genera, such as Leptosphaerulina, Letendraea, Pleospora, Tubeufia, Pyrenophora, and Wettsteinina (Kodsueb et al., 2006a, Pinnoi et al., 2007). The present work had three objectives: (1) to determine whether Massariosphaeria represents a natural group; (2) to verify the familial placement of Massariosphaeria; and (3) to discuss phylogenetic findings with respect to morphological-based classification schemes.

Section snippets

DNA extraction, amplification and sequencing

Cultures of fungi used in this study were obtained from the Centraalbureau voor Schimmelcultures (Massariosphaeria grandispora = CBS613.86; M. roumeguerei = CBS612.86; M. triseptata = CBS614.86; M. typhicola = CBS609.86). Isolates were grown on potato-dextrose agar (PDA) and malt-extract agar (MEA) for two to four weeks and total genomic DNA was extracted from mycelia following the protocols as outlined by (Cai et al. Cai et al., 2005, Cai et al., 2006a, Cai et al., 2006b). GenBank accession numbers

Ribosomal DNA phylogenies

The 18S rDNA dataset comprises 60 taxa with 1022 characters (16 % parsimony informative). WP analyses yielded two trees that were identical in topology and not statistically different, based on KH tests. One of them is shown in Fig 1 (TL = 957.95, CI = 0.568, RI = 0.756, RC = 0.430, HI = 0.432). The trees generated in UP analyses were identical to those from WP but had slightly less nodal support. Phylogenies obtained show that Massariosphaeria spp. do not constitute a monophyletic lineage. M. roumeguerei

Discussion

The objective of this study was to assess the phylogenetic relationships of Massariosphaeria and establish its familial placement within the Pleosporales. Based on 18S rDNA phylogenies, Liew et al. (2000) found that M. phaeospora (the type species) was basal to other members of the Massarinaceae (as circumscribed by Eriksson & Hawksworth 2003). Inderbitzin et al. (2002) could not confidently assess the proper affinities of M. phaeospora as its phylogenetic placement was largely dependent upon

Acknowledgements

The University of Hong Kong (RGC 7322/04M-awarded to R.J. and K.D.H.) is thanked for funding this research and providing H.K.W. with a training stipend to visit Hong Kong. This study was also partially supported by the National Science foundation of China (No. 30300229).

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