Delimitation, new species and teleomorph-anamorph relationships in Codinaea, Dendrophoma, Paragaeumannomyces and Striatosphaeria (Chaetosphaeriaceae)

Abstract The Chaetosphaeriaceae are a diverse group of pigmented, predominantly phialidic hyphomycetes comprised of several holomorphic genera including Chaetosphaeria, the most prominent genus of the family. Although the morphology of the teleomorphs of the majority of Chaetosphaeria is rather uniform, their associated anamorphs primarily exhibit the variability and evolutionary change observed in the genus. An exception from the morphological monotony among Chaetosphaeria species is a group characterised by scolecosporous, hyaline to light pink, multiseptate, asymmetrical ascospores and a unique three-layered ascomatal wall. Paragaeumannomyces sphaerocellularis, the type species of the genus, exhibits these morphological traits and is compared with similar Chaetosphaeria with craspedodidymum- and chloridium-like synanamorphs. Morphological comparison and phylogenetic analyses of the combined ITS-28S sequences of 35 isolates and vouchers with these characteristics revealed a strongly-supported, morphologically well-delimited clade in the Chaetosphaeriaceae containing 16 species. The generic name Paragaeumannomyces is applied to this monophyletic clade; eight new combinations and five new species, i.e. P. abietinussp. nov., P. eleganssp. nov., P. granulatussp. nov., P. sabinianussp. nov. and P. smokiensissp. nov., are proposed. A key to Paragaeumannomyces is provided. Using morphology, cultivation studies and phylogenetic analyses of ITS and 28S rDNA, two additional new species from freshwater and terrestrial habitats, Codinaea paniculatasp. nov. and Striatosphaeria castaneasp. nov., are described in the family. A codinaea-like anamorph of S. castanea forms conidia with setulae at each end in axenic culture; this feature expands the known morphology of Striatosphaeria. A chaetosphaeria-like teleomorph is experimentally linked to Dendrophoma cytisporoides, a sporodochial hyphomycete and type species of Dendrophoma, for the first time.


introduction
The family Chaetosphaeriaceae ) is a speciose, diverse group of pigmented, predominantly phialidic fungi some of which possess known teleomorphs (sexual and asexual morphs, hereafter teleomorph and anamorph respectively). Members of the family have a world-wide geographical distribution. They are essential components of biodiversity and play a role in decomposition of woody and herbaceous material and leaf litter, occur in soil, and some exhibit an endophytic lifestyle and have been isolated from living herbs and trees (e.g. Gams and Holubová-Jechová 1976;Hughes and Kendrick 1968;Réblová and Gams 1999;Réblová and Seifert 2003;Réblová 2004;Fernández and Huhndorf 2005;Huhndorf and Fernández 2005;Crous et al. 2012;Hashimoto et al. 2015;Yang et al. 2018;Lin et al. 2019;Luo et al. 2019).
Sexually reproducing fungi encompassed in the Chaetosphaeriaceae are perithecial ascomycetes that share several morphological traits such as similar anatomy of the brittle, melanised ascomatal wall, persistent paraphyses, unitunicate, thin-walled asci with a refractive, non-amyloid apical annulus, transversely septate ascospores that germinate by germ tubes and phialidic conidiogenesis. Several species produce both ascospores and conidia, ascomata are often associated with conspicuous conidiophores arranged in the juxtaposition. Most representatives of the family reproduce only asexually and are known as "anamorphic holomorphs" . They either permanently lost the ability to sexually reproduce and do not develop the teleomorph, or the latter remains to be discovered.
The majority of species accommodated in Chaetosphaeria possess ellipsoidal, fusiform to cylindrical-fusiform, 1-5-septate, hyaline, symmetrical ascospores with their length generally ranging from 6 to 40 μm. Ascomata are brown to black, papillate, often glossy with a two-layered ascomatal wall; the outer layer consisting of several rows of brick-like cells with dark brown, opaque walls. The transfer of a scolecosporous Lasiosphaeria raciborskii (Carroll and Munk 1964) with a three-layered ascomatal wall to Chaetosphaeria by Miller and Huhndorf (2004) expanded the concept of the genus. Huhndorf and Fernández (2005) introduced another four morphologically similar species based on ITS sequence data, i.e. Ch. ellisii (= Ch. longispora), Ch. lapaziana, Ch. panamensis and Ch. rubicunda, characterised by unique ascomatal wall anatomy, multiseptate scolecosporous ascospores and occurrence on decaying wood. Their ascomatal wall is composed of three layers. The typical chaetosphaeriaceous outer layer is present as the middle layer, while the outer layer consists of thin-walled, mostly globose cells. The ascospores are hyaline, cylindrical-filiform (up to 150 μm long), 7-16-septate, usually asymmetrical with a bluntly rounded apical end and tapering towards the basal end. These species were experimentally linked with a craspedodidymum-like anamorph, and some also form a chloridium-like synanamorph in axenic culture . Atkinson et al. (2007) and Perera et al. (2016) introduced another three Chaetosphaeria matching the diagnostic characters of this group. Among the known ascomycetes, the monotypic genus Paragaeumannomyces (Matsushima 2003), based on P. sphaerocellularis, is remarkably similar to these scolecosporous species of Chaetosphaeria in features of ascomata, asci and ascospores and ecology.
Our sampling of saprobic lignicolous fungi in terrestrial biotopes in various localities in Europe, New Zealand and North America revealed several species whose morphological characters best match those of the genus Paragaeumannomyces and other scolecosporous Chaetosphaeria, i.e. Ch. albida (Atkinson et al. 2007), Ch. longispora (Barr 1993;Huhndorf and Fernández 2005) and five unknown species. We also collected additional specimens that represent new species, an unknown Codinaea (Maire 1937;Hughes and Kendrick 1968) on submerged wood and leaves in France and United Kingdom and an undescribed Striatosphaeria (Samuels and Müller 1978) on decaying bark of a woody liana in French Guiana. Codinaea, typified by C. aristata, comprises fungi forming tufts of fertile or sterile setae accompanied by conidiophores terminating into a phialide and hyaline, aseptate, falcate conidia with setulae at both ends. Striatosphaeria is well distinguishable from other members of the family by brown, 1-septate ascospores with longitudinal ridges and furrows running the entire length of the ascospore and a codinaea-like anamorph with brown, 1-septate conidia.
Dendrophoma (Saccardo 1880;Crous et al. 2012) is characterised by superficial, stromatic, stipitate, cupulate conidiomata, phialidic conidiogenous cells arranged in terminal whorls and naviculate to botuliform, aseptate, hyaline conidia with polar appendages. Using DNA sequence data, Crous et al. (2012) confirmed its systematic placement in the Chaetosphaeriaceae. However, its teleomorph-anamorph relationship remains unknown. A collection of a chaetosphaeria-like species with glabrous, dark, erumpent, aggregated ascomata sometimes in caespitose clusters, stipitate asci with inconspicuous apical annulus and fusiform, hyaline, 1-septate ascospores was encountered in the cracks of the bark of twigs of Buxus sempervivens in Germany. The axenic culture derived from ascospores yielded an anamorph similar to Dendrophoma. A BLASTn search (Zhang et al. 2000) for possible relatives in GenBank (Sayers et al. 2019) suggested our isolate is similar to Dendrophoma cytisporoides, the type species of the genus.
The present study provides new data that improve our understanding of morphological and genetic diversity of the Chaetosphaeriaceae and its pleomorphism. Our longer term goals focus on identification of monophyletic, morphologically well-delimited natural lineages and the life history of species currently assigned to the family. To assess phylogenetic relationships of our isolates, we based the study on morphological and cultivation studies along with the analysis of DNA sequence data from the nuc rDNA internal transcribed spacer region (ITS1-5.8S-ITS2 = ITS) and nuclear large subunit 28S ribosomal DNA gene (28S).

Herbarium material and fungal strains
Material for this study was collected in north temperate regions of Europe (France, Germany and Ukraine) and North America (North Carolina, Tennessee), south subtropical and temperate climate zones of New Zealand, and in the neotropical regions of the Caribbean (Puerto Rico) and South America (French Guiana). An additional living culture was obtained from BCCM/MUCL Agro-food & Environmental Fungal Collection (MUCL), Université catholique de Louvain, Louvain, Belgium. Representative strains and ex-type strains were deposited at Westerdijk Fungal Biodiversity Institute (CBS), Utrecht, the Netherlands. Holotypes and other herbarium material (as dried voucher specimens) were deposited in the Fungarium of the Illinois Natural History Survey (ILLS), Champaign, Illinois, USA, the New Zealand Fungarium (PDD), Auckland, New Zealand and Herbarium of the Institute of Botany (PRA), Czech Academy of Sciences, Průhonice, Czech Republic. Isolates and specimens, their sources and GenBank accession numbers for ITS and 28S sequences generated in this study, are listed in Table 1.

Morphological characterisation
Morphological characteristics were obtained from fungi growing on natural substrate and growth media. Descriptions in the key are based on fungi growing on natural substrate. Herbarium material was rehydrated with tap water and examined with an Olympus SZX12 dissecting microscope (Olympus America, Inc., Melville, USA). Hand-sectioned ascomata and centrum material (asci, ascospores and paraphyses), conidiophores and conidia were mounted in 90 % lactic acid, Melzer's reagent, and lactophenol with cotton blue. All measurements were in Melzer's reagent. Means ± standard deviation (SD) based on a minimum of 20-25 measurements are given for dimensions of asci, ascospores and conidia. Micromorphological observations were made using an Olympus BX51 compound microscope with differential interference contrast (DIC) and phase contrast ( Leonian's agar (MLA) (Malloch 1981), oat-meal agar (OA) modified after Gooding and Lucas (1959) (30 g of oatmeal cooked in 1 L of distilled water for 15-30 min, filtered through cheesecloth, the filtrate was brought back to volume with distilled water, 15 g of agar, sterilized for 60 min at 121 C) and potato-carrot agar (PCA) . To induce sporulation, strains were also inoculated on CMA  with sterile stems of Urtica dioica. Descriptions of colonies are based on 4 wk old cultures grown in darkness at 22-23 C.

DNA extraction and amplification
Methods for the DNA extraction and amplification of samples with A.N.M., ILLS and S.M.H. prefixes followed Huhndorf et al. (2004) and Hustad and Miller (2015). Other samples were processed according to the following protocols. Total genomic DNA was extracted from mycelium removed from 3-wk-old cultures grown on MLA using the DNeasy® UltraClean® Microbial Kit (Qiagen GmbH, Germany) following the manufacturer's protocol for filamentous fungi. All PCR amplifications were carried out in 25 μL volume reactions using a Q5 High Fidelity DNA polymerase kit (New England Biolabs Inc., United Kingdom) according to the manufacturer's protocol. Primers used for the amplification included: V9G/LR8 (de Hoog and Gerrits van den Ende 1998; Vilgalys unpublished) for the internal transcribed spacers (ITS) of the nuclear rRNA cistron and D1, D2 and D3 domains (approx. 1900 bp of the 5' end) of the 28S rDNA gene. PCR was carried out in a BioRad C1000 thermal cycler (Bio-Rad Laboratories Inc., USA) as follows: 98 C for 30 s; 40 cycles of denaturation (98 C for 10 s), annealing (62 C for 30 s) and elongation (72 C for 90 s) and a final extension step at 72 C for 5 min. Amplicons were purified from agarose gels using a NucleoSpin® Gel and PCR Clean-up Kit (Macherey-Nagel GmbH & Co. KG, Germany) following the manufacturer's instructions, with an elution volume of 25 μL. The DNA concentration was assessed fluorimetrically using Quant-iT PicoGreen dsDNA Assay Kit and Qubit fluorometer (Invitrogen / Thermo Fisher Scientific, USA) to assure required sequencing concentrations adjusted for the length of amplicons/ number of reads required.

Alignments and phylogenetic analyses
Two gene markers, ITS and 28S rDNA, were analysed to assess evolutionary relationships of the unknown fungi with members of the Chaetosphaeriaceae. Consensus se condary structure (2D) models for the ITS1 and ITS2 for members of the Chaetosphaeriaceae were built using the Ppfold program v.3.0 (Sukosd et al. 2012).
The obtained 2D consensus models were further improved using the program Mfold (Zuker 2003) and adjusted manually if necessary, based on comparison of homologous positions in the multiple sequence alignment. A predicted 2D model of the 28S of Saccharomyces cerevisiae (Gutell et al. 1993) was used to improve the alignment of this gene. The models were highly consistent in all taxa.
ITS and 28S sequences were aligned manually in Bioedit v.7.1.8 (Hall 1999). Gen-Bank accession numbers for ITS and 28S sequences of members of the Chaetosphaeriaceae retrieved from GenBank and published in other studies are listed in Table 2. Single-locus data sets of the Chaetosphaeriaceae (ITS: 89 sequences/602 characters including gaps, 28S: 86/1176) and Paragaeumannomyces (ITS: 35/489, 28S: 32/1104) were evaluated using PartitionFinder2 (Lanfear et al. 2016), implemented in the CIP-RES Science Gateway v.3.3 (http://www.phylo.org) (Miller et al. 2010), to find the best partitioning scheme for our datasets and to select best-fit models under corrected Akaike information criteria. Conflict-free data sets were concatenated into two alignments (deposited in TreeBASE 25964) that were subjected to subsequent phylogenetic analyses.
Three analyses were employed to estimate phylogenetic relationships. Bayesian Inference (BI) and Maximum Likelihood (ML) analyses were performed through the CIPRES Science Gateway v.3.3. ML analyses were conducted with RAXML-HPC v.8.2.12 (Stamatakis 2014) with a GTRCAT approximation. Nodal support was determined by non-parametric bootstrapping (BS) with 1 000 replicates. BI analyses were performed in a likelihood framework as implemented in MrBayes v.3.2.6 (Huelsenbeck and Ronquist 2001). Two Bayesian searches were performed using default parameters. The B-MCMCMC analyses lasted until the average standard deviation of split frequencies was below 0.01 with trees saved every 1000 generations. The first 25 % of saved trees, representing the burn-in phase of the analysis, were discarded. The remaining trees were used for calculating posterior probabilities (PP) of recovered branches. Maximum Parsimony (MP) analyses were conducted with PAUP 4.0a167 (Swofford 2003). A heuristic search was performed with the stepwise-addition option with 1000 random taxon addition replicates and TBR branch swapping. Because the secondary structures of the ITS and 28S were carefully studied when aligning the sequences, and regions with incomplete sequences were excluded from the analysis, we treated gaps as a fifth character state; they were given equal weight as the other characters. All characters were unordered. Branch support was estimated on the recovered topologies by performing a heuristic search of 1000 bootstrap replicates consisting of ten randomaddition replicates for each bootstrap replicate.

Phylogenetic analyses
Employing the predicted 2D structure of the variable ITS region and 28S enabled us to construct a reliable multiple sequence alignment of homologous positions at both Note: T, ET, IST and PT denote ex-type, ex-epitype, ex-isotype and ex-paratype strains.
helices and loops, thus eliminating potential ambiguous regions in the alignments. Initially, we compared trees from ML phylogenetic analyses of the two combined data sets (Chaetosphaeriaceae and scolecosporous species of Chaetosphaeria) after alignments were improved with the 2D structure, with and without applying Gblocks (Castresana 2000) using default options, to delimit and remove putative ambiguous regions. The phylogenetic trees based on datasets using Gblocks had lower support for nodes and relationships within and among several clades that could not be resolved (data not shown) compared to trees in which these regions remained. Therefore, the final phylogenies were based on datasets in which Gblocks was not employed. Evolutionary relationships of studied fungi were evaluated in the phylogenetic analysis based on the combined ITS and 28S sequences of 87 representative species of the Chaetosphaeriaceae. Leptosporella arengae, L. bambusae (Leptosporellaceae), and Tracylla eucalypti and T. aristata (Tracyllaceae) were used to root the tree. 76 nucleotides (nt) at the 5'-end and 606 nt at the 3'-end of 28S were excluded from the alignment because of missing data in the majority of sequences. The alignment had 1778 characters including gaps and 882 unique character sites (RAxML). In the MP analysis, 1021 characters were constant (proportion = 57.42 %), 134 variable characters were parsimony-uninformative, 623 characters were parsimony-informative (included); two most parsimonious trees were produced (length = 5066 steps, consistency index = 0.0.298, homoplasy index = 0.702, retention index = 0.631). For the BI analysis, GTR+I+G model was selected for ITS and 28S partitions. The ML tree (RAxML) is shown in Fig. 1. There were no conflicts among the trees generated by the three different phylogenetic analyses. The Chaetosphaeriaceae were resolved as a strongly supported clade; some of the nodes of the backbone tree, which obtained support in the ML and/or BI analyses, were not statistically supported in MP analysis. The 39 identified terminal clades corresponded to individual genera or natural groups of species. Codinaea was resolved as polyphyletic in three subclades. The unknown species was grouped in a clade (92 % ML BS/1.0 PP/92 % MP BS) containing seven Codinaea or Dictyochaeta species, three of which possess the typical Codinaea phenotype, while other morphologically similar species with setulate conidia clustered in the other two subclades, C. lambertiae, C. pini and C. simplex (100/1.0/100) and Dictyochaeta septata and D. cangshanensis (96/1.0/100). The new species Codinaea paniculata, based on four strains, was resolved as a monophyletic clade in all three analyses, although the statistical support varied. In ML and MP analyses the clade obtained 97 % and 100 % support, respectively, in the BI analysis it was weakly supported with 0.77 PP. The intraspecific variability of C. paniculata, based on ITS sequences, varied slightly. Three strains (CBS 145098 ex-type, CBS 126573, MUCL 34876) had identical ITS sequences, strain CBS 127692 differed from them by one base pair. The 28S sequence similarity of all strains of C. paniculata was 100 %. The undescribed Striatosphaeria was nested in the monophyletic Striatosphaeria (100/1.0/100) clade as sister to S. codinaeophora. It clustered in a subclade (93/0.98/99) with an endophytic isolate Striatosphaeria sp. monte6.2; their ITS exhibited 98.5 % sequence similarity. The five unknown chaetosphaeria-like species with scolecosporous ascospores were nested in a stronglysupported monophyletic clade (99/1.0/97). This clade contained eight additional morphologically similar species with scolecosporous ascospores and three-layered ascomatal wall. The clade is introduced as Paragaeumannomyces in this study. A chaetosphaerialike species grouped with the ex-type strain of Dendrophoma cytisporoides CBS 223.95 in a monophyletic clade (100/1.0/100). Dendrophoma was resolved as a member of a large, statistically weakly-supported grouping containing six other genera characterised by sporodochial conidiomata.
Culture characteristics. On CMD colonies 80-85 mm diam, circular, flat, margin fimbriate, aerial mycelium restricted mainly to the centre and margin of the colony, sparsely lanose, floccose centrally becoming mucoid towards the margin, cobwebby at the margin, colony centre whitish, pale brown to creamy towards the margin, pale brown pigment diffusing from the centre of the colony to the agar; reverse creamy. On MLA colonies 65-70 mm diam, circular, slightly raised, margin filiform, lanose, floccose, colony centre whitish becoming brown-grey towards the margin with a brown outer zone of submerged growth, pale brown pigment diffusing to the agar; reverse dark brown. On OA colonies 89-95 mm diam, circular, raised, margin filiform, aerial mycelium occasionally reduced or absent, colonies similar to those on MLA, lanose, floccose, locally mucoid and smooth or cobwebby, whitish becoming dark grey at the margin, a dark brown to burgundy brown pigment diffusing to the agar; reverse dark grey. On PCA colonies 78-89 mm diam, circular, flat to slightly raised, margin entire to weakly filiform, lanose, floccose, occasionally locally mucoid and smooth or with sparse decumbent aerial hyphae, cobwebby at the margin, whitish becoming brown towards the margin; reverse olivaceous brown. Sporulation on MLA, OA, CMD after 8 wk.
Other specimen examined. Habitat and distribution. All four isolates analysed in this study originated from the freshwater environment and occurred on decaying wood or leaves of Alnus glutinosa, Fraxinus excelsior and other unidentified hosts. Based on the BLASTn search of the ITS sequence of C. paniculata in GenBank, two isolates from roots of Elymus mollis (ITS: KU838460, KU839605, David et al. 2016), a native beach grass on the USA Pacific Northwest coast, and one environmental soil sample from ancient woodland enclosing a conifer plantation in the United Kingdom (ITS: KM374380, Johnson et al. 2014) showed 100 % sequence similarity. Based on these records, C. paniculata is known from the north temperate region in Europe in France and United Kingdom and North America in USA, Oregon.

Paragaeumannomyces
Notes. The holotype of P. sphaerocellularis (Japan, Schimizu-cho, Wakayama Pref., on decaying twig of unknown broadleaf tree, Apr. 2000, MFC-21077), the type species of Paragaeumannomyces (Matsushima 2003), was not available to us. A comparison of its protologue with our specimens and descriptions of other scolecosporous species of Chaetosphaeria (Carroll and Munk 1964;Huhndorf and Fernández 2005;Atkinson et al. 2007;Perera et al. 2016), combined with phylogenetic analysis of the ITS-28S sequences of 35 isolates, provided sufficient evidence to consider them congeneric. Paragaeumannomyces is proposed as the correct name for this morphologically and phylogenetically well-delimited group of chaetosphaeriaceous fungi. The width of the ascus is sometimes variable even within a single collection depending on the arrangement of ascospores in the sporiferous part, whether they are 2-3-seriate, 4-seriate endto-end or in a fascicle.

Key to
Culture characteristics. On CMD colonies 10-11 mm diam, circular, slightly convex, margin entire to weakly fimbriate, lanose, beige-brown with a dark brown outer zone of submerged growth, dark brown pigment diffusing from the colony margin to agar; reverse dark brown to black. On MLA colonies 12-15 mm diam, circular, slightly convex, margin entire, lanose, floccose, cobwebby at the margin, beige-brown with a dark brown outer zone of submerged growth, brown pigment diffusing from the colony margin to agar; reverse dark brown. On OA colonies 8-9 mm diam, circular, convex, margin entire, lanose, beige-brown, with a paler outer ring; reverse brown. On PCA colonies 14-15 mm diam, circular, convex, margin entire, lanose, floccose, cobwebby towards the margin, beige, pale brown towards the margin; reverse brown. Sporulation absent on all media, even after prolonged incubation (> 3 mo).
Other specimen examined. Ukraine Habitat and distribution. All specimens of P. abietinus occur on decaying wood of Abies alba. The species has been collected in mountain areas and is known in Europe in France and Ukraine.
Notes. Attempts to cultivate this species were unsuccessful for the Ukrainian specimens; the ascospores germinated over five days with long inflated germ tubes from both ends but did not grow after isolation on agar medium. The axenic culture derived from the ascospore isolate of the French material yielded sterile mycelium only.
Notes. For additional illustrations and description, see Atkinson et al. (2007).
Paragaeumannomyces albidus is the only species of the genus characterised by a wide range of ascoma colours that change when ascomata are young and fresh or mature and dried. Different colours were used by Atkinson et al. (2007) to distinguish P. albidus from closely related P. bombycinus. Paragaeumannomyces albidus differs from the latter species in having distinctly papillate ascomata, which are almost white, yellowish-white, areolate and translucent when young except for the black papilla (Fig. 5A, B). In older specimens and after drying, ascomata often become laterally pinched, dark yellow, buff, tawny to dark ginger-brown ( Fig. 5C-E). The ascomatal wall of P. albidus is thicker than that of P. bombycinus, with an outer layer containing an external melanised section. In our material, asci were longer than those in the original description, 270-295 × 18.5-20.5 μm long and 155-225 μm long in the sporiferous part vs 220-260 × 16-20 μm fide Atkinson et al. (2007). The size and septation of ascospores matched those given in the protologue, though being slightly longer in the upper range: (3-)5-11-septate, (55-)69-86 × 5-6.5(-7) μm vs (5-)7(-12)-septate, (47-)60-80 × 5-7 μm fide Atkinson et al. (2007). The ascospores exhibited a dextrinoid reaction in Melzer's reagent turning reddish-brown except for the tips of the end cells that remain hyaline.
Attempts to isolate our specimens of P. albidus in living culture were not successful. Therefore, the DNA was extracted from herbarium material of all three collections, but only ITS1 of PDD 118738 could be amplified and sequenced. Comparison of the ITS1 sequences of our specimen and the holotype of P. albidus revealed 100 % similarity (Fig. 2). Habitat and distribution. The species has been collected on decaying wood of Nothofagus sp. and is known from New Zealand (Atkinson et al. 2007).
Notes. For description, illustration and holotype information, see Atkinson et al. (2007). Although P. albidus and P. bombycinus share identical ITS sequences and the size of their ascomata, asci and ascospores considerably overlap, the latter species was distinguished by characters in the ascomatal wall, ascoma appearance and ascospore septation. The ascomata of P. bombycinus are light fawn-grey and non-areolate when fresh, the ascomatal wall lacks the external melanised layer or the melanisation is only weakly present, and the black papilla is lacking or indistinct being covered by the outer layer. The ascospores of P. bombycinus are (7-)11(-13)-septate compared to (5-)7(-12)-septate ascospores of P. albidus. Atkinson et al. (2007) considered the ITS sequence identity uninformative in the light of distinct morphologies between the two species. Another explanation to this peculiar case could be that P. bombycinus is a described anomaly within P. albidus based on a single collection. More specimens of the "bombycinus" phenotype need to be examined, and their ITS and possibly other loci analysed. Etymology. Elegans (L) elegant, referring to elegant and lovely ascomata adorned with setae.
Other Habitat and distribution. The present species is a saprobe on decaying wood of Nothofagus sp. and other unidentified hosts, known from New Zealand (Atkinson et al. 2007; this study).
Notes. Paragaeumannomyces elegans is distinguishable from other members of the genus by densely setose, dull glistening brown ascomata with a light grey tinge, which gives them an almost grey appearance when dried. The new species resembles P. garethjonesii (Perera et al. 2016) and P. panamensis  in 7-septate ascospores and setose ascomata with acute, stiff, opaque setae scattered over the entire surface, but differs from them in larger ascomata, asci and wider ascospores (for a detailed comparison see the key).
Comparison of the ITS sequence of the holotype of P. elegans with available Paragaeumannomyces sequences revealed 100 % sequence similarity with a specimen PDD 92561 (New Zealand, Taupo, Ohakune, ITS: EUO37895) tentatively identified as P. raciborskii (Atkinson et al. 2007) (Fig. 2). Habitat and distribution. Paragaeumannomyces garethjonesii was collected on a Fabaceae seed pod and is known from Asia in Thailand (Perera et al. 2016).
Notes. For description, illustration and holotype information see Perera et al. (2016). Paragaeumannomyces garethjonesii resembles P. panamensis  in size of ascomata, which are the smallest (up to 250 μm diam and 270 μm high) within the genus, setae scattered over the entire ascoma, overlapping length of their asci and 7-septate ascospores, but it differs by shorter (38-47 μm) setae, slightly wider (10.7-13.3 μm) asci and the absence of aleuriospore-like cells in culture (for a detailed comparison see the key). Etymology. Granulum (L), granule, small grain, diminutive of granum, referring to the roughened surface of the ascomatal wall composed of globose cells, which appears granulose in the surface view.
Habitat and distribution. A saprobe on decaying wood, known from New Zealand. Notes. Paragaeumannomyces granulatus most closely resembles P. abietinus in the ascoma appearance, pink content of the ascoma centrum, ascospores with usually more than seven septa and positive dextrinoid reaction in Melzer's reagent but both species are separated by size of asci and ascospores. The ascospores of P. abietinus are (5-)7-9(-11)-septate and shorter [(62-)65-87 μm] and asci are shorter and narrower [(185-)195-240 × 12-14.5(-15.5 Habitat and distribution. Paragaeumannomyces lapazianus is common on decaying wood in the neotropics and is known from the Caribbean in Puerto Rico and Jamaica, from Central America in Costa Rica, and from South America in French Guiana (Carroll and Munk 1964;Huhndorf and Fernández 2005).
Notes. For description, illustration and holotype information see Carroll and Munk (1964) and Huhndorf and Fernández (2005). Paragaeumannomyces lapazianus has 7-septate ascospores and the largest ascomata in the genus, (400-)500-950 μm diam and 525-825(-1025) μm high fide Huhndorf and Fernández (2005), and forms a craspedodidymum-like anamorph in vitro. The anamorph is characterised by inflated, pigmented conidiogenous cells with a flared collarette and oblate to horizontally oblong conidia with a short abscission scar or frill and without setulae.   Barr, Mycotaxon 46: 48. 1993 Habitat and distribution. The species occurs on decaying wood and is known from the north temperate region in the USA (Indiana, New Jersey, North Carolina, South Carolina, Tennessee) (Barr 1993;Huhndorf and Fernández 2005; this study).
Sphaeria longispora (Ellis 1877) is a later homonym of S. longispora (Currey 1859) and S. longispora (Karsten 1873). Barr (1993) revised the holotype of S. longispora Ellis (USA, New Jersey, Newfield, on fallen branch of Kalmia latifolia, 20 Jul 1874, J.B. Ellis, NY) and concluded that the fungus is better placed in Lasiosphaeria due to filiform, septate ascospores and setose ascomata and proposed a replacement name, Lasiosphaeria ellisii as a nomen novum. This species was later transferred to Chaetosphaeria by Huhndorf and  as Ch. ellisii. Kirk (2014) considered the first combination of S. longispora in Ophioceras by Saccardo (1883) as the earliest legitimate name of the taxon in the same rank (Art. 41.3) to replace Sphaeria longispora Ellis. Ophioceras longisporum Sacc. is, therefore, a basionym for all future combinations. Kirk (2014) proposed a new combination Chaetosphaeria longispora but erroneously cited S. longispora as the basionym, which does not affect the valid publication of the new combination (Art. 41.8c).
Notes. For description, illustrations and holotype information see Huhndorf and Fernández (2005) and Perera et al. (2016). Paragaeumannomyces panamensis is similar to P. sphaerocellularis in ascoma and is more or less comparable in size of asci, but differs by shorter, always 7-septate ascospores and occurrence in the tropics. For detailed comparison, see notes to P. sphaerocellularis. Habitat and distribution. Paragaeumannomyces raciborskii has been collected on culms of Chusquea bamboo and other unidentified bamboo species, on palm wood and fruit, and decaying wood of unknown trees. The species has a pantropical geographical distribution and is probably the most commonly encountered species of the genus; it is known from Indonesia in Java and Central America in Costa Rica (Penzig and Saccardo 1897;Carroll and Munk 1964). Other collections published under this name, which may represent different species, originate from Asia in Thailand, the Caribbean in Cuba, Jamaica and Puerto Rico, Central America in Costa Rica and Panama, and South America in Ecuador, French Guiana and Venezuela .
Habitat and distribution. Paragaeumannomyces rubicundus occurs on decaying wood and is known from the Caribbean in Puerto Rico and from Central America in Costa Rica .
Notes. For description, illustration and holotype information see Huhndorf and Fernández (2005). Paragaeumannomyces rubicundus is distinguished from other species of the genus by ascomata with red surface crystals not dissolving in water, 3 % KOH or lactophenol. Similar to Paragaeumannomyces sp. 4 (S.M.H. 3119), the craspedodidymum-like anamorph forms conidia with three setulae.  Etymology. The species epithet is proposed in honour of Sabine M. Huhndorf for her contribution to mycology and studies in Chaetosphaeria.
Other specimen examined. Habitat and distribution. A saprobe on decaying wood, so far known from North America in the USA (North Carolina, Tennessee) (Ellis 1887;Huhndorf and Fernández 2005).
Notes. Huhndorf and Fernández (2005) reported P. longisporus (as Ch. ellisii) from numerous collections from North America; the phylogenetic analysis of ITS sequences of six specimens resolved this species as a statistically unsupported clade with two strongly supported subclades. Although Huhndorf and Fernández (2005) described P. longisporus with setae scattered over the entire ascoma, in discussion, they admitted the presence of setae also around the ostiole: "In C. ellisii, C. raciborskii and C. panamensis the setae tend to be scattered over the entire surface of the ascomata, however some specimens of C. ellisii may have setae concentrated only at the apex." Barr (1993) described the ostiole of the holotype of S. longispora surrounded by a crown of dark brown, stiff setae. We examined three collections tentatively identified as P. longisporus from North America (ILLS00121384, ILLS00121385, ILLS00121386) and in each the ostiole was delimited by densely aggregated acute setae. Apart from the ostiolar setae, additional setae were scattered over the entire ascoma, but they differed by their density among collections. The ascomata and asci of these three collections are comparable in size; the main difference lies in the ascospore length. The specimen ILLS00121384 has longer [(64.5-)68.5-86.5(-88.5) μm] ascospores compared to ILLS00121385 and ILLS00121386, which have shorter [(50.5-)52.5-68 μm] ascospores corresponding to the size given by Barr (1993) for the S. longispora holotype.

Paragaeumannomyces smokiensis
Habitat and distribution. A saprobe on decaying wood, known only from the USA. Notes. The present species is most similar to P. abietinus, the only member of the genus known from Europe. They share dark reddish-brown, glabrous ascomata with short setae surrounding the ostiole. Although the size of ascospores of both species overlap, P. abietinus differs from P. smokiensis by longer [(185-)195-240 × 12-14.5(-15.5) μm] asci and slightly longer and wider ascospores [(62-)65-87 × (3.5-)4-5.5 μm] with usually less septa [(5-)7-9(-11)]. Habitat and distribution. The species was described from dead twigs of an unknown broadleaf tree and is so far known only from the subtropical climate zone of the northern hemisphere in Japan, Wakayama Prefecture (Matsushima 2003).
Culture characteristics. On CMD colonies 23-25 mm diam, circular, flat, margin entire, lanose, floccose, funiculose at the centre, cobwebby towards the periphery, whitish with irregular pale brown spots due to pigmented funiculose mycelium, with an isabelline outer zone of submerged growth; reverse beige. On MLA colonies 22-25 mm diam, circular, raised, margin entire, lanose, floccose, zonate, with grey, brown and white zones, with an isabelline outer zone of submerged growth; reverse dark grey. On OA colonies 31-33 mm diam, circular, flat, margin entire, sparsely lanose, floccose, cobwebby at the margin, zonate, whitish, colony centre with irregular dark brown spots due to pigmented submerged mycelium, pale brown towards the margin, with an olivaceous outer zone of submerged growth, dark brown pigment diffusing to agar at the colony centre; reverse olivaceous grey. On PCA colonies 17-19 mm diam, circular, slightly convex centrally, margin entire, lanose, floccose becoming cobwebby towards the periphery, isabelline to light beige with irregular brown spots due to pigmented mycelium; reverse light beige. Sporulation abundant on CMD, CMA with Urtica stems and PCA, sparse on MLA and OA.
Habitat and distribution. Striatosphaeria castanea occurs on the bark of woody liana and as an endophyte of Encyclia ghillanyi. It is known from South America in Brazil and French Guiana.
Habitat and distribution. Saprobic on decaying wood and bark of Buxus sempervivens, Deutzia scabra, Rhododendron sp., and Ulmus sp. The species is known from Europe in France, Germany and the Netherlands (Saccardo 1879;Sutton 1965;Crous et al. 2012; this study).
Notes. Our strain sporulated in vitro only after prolonged incubation. On OA it formed globose to pulvinate fertile conidiomata, while on PCA the conidiomata often became confluent. The comparison of ITS and 28S sequences of our strain with those of the epitype strain of D. cytisporoides CBS 223.95 (Crous et al. 2012) confirmed they are conspecific; the two strains share 100 % sequence similarity.

Discussion
Based on morphology, cultivation studies and phylogenetic analysis of the combined ITS-28S loci, seven new species and eight new combinations are introduced in the Chaetosphaeriaceae. Paragaeumannomyces (Matsushima 2003) is proposed for the monophyletic, strongly supported clade of former Chaetosphaeria species (Figs 1, 2) characterised by scolecosporous, multiseptate, asymmetrical, hyaline to light pink ascospores and unique three-layered ascomatal wall. The unusual wall was first brought to attention by Carroll and Munk (1964). The colour of the outer wall ranges from white, whitish-yellow, ginger-brown to reddish-brown, russet to dark brown and is composed of thin-walled cells of textura angularis, which may contain pale purple pigment when fresh, occasionally with red surface crystals. Setae, if present, are scattered over the entire ascoma or only surround the ostiole. They are stiff, acute, dark brown with opaque walls, arise from the middle, carbonaceous layer and penetrate the outer layer of globose cells. Members of the genus occur on decayed plant material, especially on strongly decayed decorticated wood.
Examination of our material of Paragaeumannomyces revealed that ascospores of four species exhibit a strong dextrinoid reaction in Melzer's reagent, namely P. abietinus, P. albidus, P. granulatus and P. smokiensis. The ascospores turned reddishbrown except for the end cells, which remained partially hyaline, especially at the tips. Interestingly, these species share glabrous, non-setose ascomata or only minute setae are arranged around the ostiole. The chemical reaction is visible in ascospores without guttules, which otherwise fill the cells and obscure the colour. The ascospores of P. elegans, P. longisporus and P. sabinianus exhibit a negative or weak dextrinoid reaction; some mature ascospores turned light pink-brown. These species share setose ascomata, sometimes with ostioles surrounded by minute setae. Although more species need to be examined to evaluate this character, we hope it is not premature to argue that the dextrinoid reaction of ascospores is species-specific and has the potential to become another diagnostic feature facilitating species identification. Because we did not examine all known species of Paragaeumannomyces, this character has not been used in the key, but it is mentioned in the species descriptions, if known.
Paragaeumannomyces, typified by P. sphaerocellularis, encompasses 18 species. The present phylogenetic tree (Fig. 2) contains 12 of them and four subclades labelled Paragaeumannomyces sp. 1-4, which represent separate, yet undescribed taxa at the species rank. The molecular data of P. raciborskii and P. sphaerocellularis are unavailable. The closest relatives to Paragaeumannomyces are species of Exserticlava and Chaetosphaeria lignomollis with a kylindria-like anamorph, characterised by septate, versicolorous or hyaline ascospores, respectively (Fig. 1).
Members of Paragaeumannomyces are not easily cultivated and only seldom sporulate in vitro. Huhndorf and Fernández (2005) noted that even isolates from the same specimen varied in their ability to produce the anamorph in culture. The craspedodidymum-like anamorph with usually semi-macronematous to micronematous conidiophores, inflated phialides, deeply flared, cup-shaped collarettes and hyaline nonseptate conidia, with or without setulae, was reported for P. lapazianus, P. longisporus, P. panamensis, P. rubicundus, and Paragaeumannomyces sp. 1-4, while the chloridium-like synanamorph is known only in P. longisporus and Paragaeumannomyces sp. 2 Perera et al. 2016). The systematic placement of Craspedodidymum (Holubová-Jechová 1972), typified by C. elatum, is unknown. The genus was erected for a hyphomycete forming effuse colonies on an old petiole of Phoenix canariensis in a green house in the Czech Republic. To date, 15 binomials were introduced in the genus (Index Fungorum). Craspedodidymum elatum differs from Paragaeumannomyces anamorphs by macronematous, dichotomously branched conidiophores and non-septate, dark brown conidia with a basal hilum. In hyaline, unicellular, globose or triangular conidia with setulae, the Paragaeumannomyces anamorphs also resemble Bahusutrabeeja (Subramanian and Bhat 1977) and Nawawia (Marvanová1980), respectively. Bahusutrabeeja and Nawawia are similar to each other but differ in shape of conidia. Bahusutrabeeja, typified with B. dwaya, forms globose conidia on solitary conidiophores, while Nawawia, based on N. filiformis, have conidia triangular, round-tetrahedral or obpyramidal-shaped on conidiophores arising from small stromata. Their molecular data Vu et al. 2019) suggest a distant relationship to Paragaeumannomyces.
The original P. longisporus clade with two strongly supported subclades was recognized as two species, the short-spored P. longisporus and the long-spored P. sabinianus. In general, a high degree of ITS sequence variability and more or less uniform teleomorphic phenotype pose special problems in species identification, especially in P. raciborskii, which was resolved as polyphyletic ; this study). Morphology of this species was studied by Huhndorf and Fernández (2005) based on more than 100 species, mostly from the neotropics. The broad species concept of P. raciborskii fide Huhndorf and Fernández (2005) includes specimens with wider ascomata, longer ascospores with less septa and longer asci than reported in the protologue (Penzig and Saccardo 1897) and re-description of this species prepared by Carroll and Munk (1964); for details see above. Although no significant variability among ascomata, asci and ascospores was encountered, Huhndorf and Fernández (2005) reported intraspecific variability regarding setae, which were lacking in some specimens. On the other hand, certain variability at the anamorphic level, typical of many natural groups of the Chaetosphaeriaceae, also occurs in P. raciborskii fide Huhndorf and Fernández (2005) and to some extent delimits the four subclades.  . A close comparison of the descriptions of the holotype of P. raciborskii (Penzig and Saccardo 1897;Carroll and Munk 1964) with collections gathered by Huhndorf and Fernández (2005) and identified as P. raciborskii confirms that none of the four subclades inferred in the ITS-28S ML tree (Fig. 2) could be delimited as P. raciborskii s. str. From a biogeographical perspective it is more likely that they belong to different species entirely. Therefore, the name P. raciborskii for these strains was rejected in our phylogeny; instead, the four subclades were designated Paragaeumannomyces sp. 1-4.
Codinaea (Maire 1937) is one of the largest genera of the Chaetosphaeriaceae with a turbulent taxonomic history. Based on a cluster analysis of phialidic dematiaceous hyphomycetes, Arambarri and Cabello (1989) considered Codinaea, with usually falcate, septate or non-septate conidia bearing setulae at both end, and Dictyochaeta (Spegazzini 1923) with non-setulate, non-septate, cylindrical and asymmetrical conidia, congeneric. Since then, Dictyochaeta (syn. Codinaea) became a broadly circumscribed genus with more than 100 species and the new morphological concept was followed by many mycologists. Réblová and Seifert (2007) confirmed with DNA sequence data that Dictyochaeta fuegiana, the type species of the genus, is a member of the Chaetosphaeriaceae. Based on the ITS-28S phylogeny (Fig. 1), D. fuegiana is unrelated to morphologically similar species with setulate conidia, classified in Codinaea or Dictyochaeta, and resolved as polyphyletic, which is in agreement with Lin et al. (2019) and Réblová et al. (2020). However, in the absence of molecular DNA data of Codinaea aristata, the generic type, it is difficult to delimit Codinaea phylogenetically. The morphological traits delimiting the new species C. paniculata, i.e. presence of setae, unbranched and shorter conidiophores growing at the base of the setae with monophialidic conidiogenous cells in vivo and falcate, non-septate conidia with setulae at both ends, best match those of Codinaea. In the subclade where C. paniculata was clustered, several species sharing the same Codinaea morphotype were present, namely C. assamica (Hughes and Kendrick 1968), D. siamensis ) and D. terminalis .
Based on published records, Striatosphaeria is an uncommon lignicolous genus with a known distribution in the neotropics. It was introduced by Samuels and Müller (1978) based on two Brazilian collections of S. codinaeophora. Additional specimens of S. codinaeophora known to us were collected on decaying wood of Dacryodes excelsa and Nectandra turbacensis and unidentified hosts in Costa Rica, French Guiana and Puerto Rico (Réblová and Winka 2000;Fernández et al. 2006;S.M. Huhndorf pers. data). The codinaea-like anamorph develops only in axenic culture. The conidia are asymmetrical, brown, 1-septate with minute, hyaline setulae at each end. Although S. codinaeophora was described with non-setulate conidia (Samuels and Müller 1978), a photograph of conidia with setulae accompanied the S. codinaeophora lineage on a phylogenetic tree (Fernández et al. 2006: fig. 1, 6g). The setulate conidia were also present in the new species, S. castanea. It is probable that setulae are formed later, after conidia detach from the conidiogenous cells. The conidia with setulae at both ends are formally introduced in Striatosphaeria for the first time in this study.
The genus Dendrophoma with a single species, D. cytisporoides, was proposed by Saccardo (1880) for fungi with phoma-like fruit bodies, botuliform hyaline conidia and conidiogenous cells arranged in a verticillate fashion. Sutton (1965) lectotypified Phoma cytisporoides (Saccardo 1879) and reported additional characters not mentioned by Saccardo in the protologue, i.e. dark brown, acute setae accompanying conidiomata and minute, unbranched setulae at both ends of conidia. Sutton (1965) compared D. cytisporoides with Dinemasporium graminum, the type species of the genus, and reduced Dendrophoma to synonymy with Dinemasporium (Léveillé 1846). Using nuclear ribosomal loci, Crous et al. (2012) re-established Dendrophoma and placed the genus in the Chaetosphaeriaceae, where it emerged as a separate lineage from Dinemasporium. The anamorph-teleomorph relationship of Dendrophoma has been established for the first time in this study. The teleomorph is morphologically similar to Chaetosphaeria (Tulasne and Tulasne 1863), but differs in having immersed to erumpent ascomata, densely branched ascogenous hyphae (Fig. 12H), the ascal apex lacking a visible discharge mechanism, which can only be seen as a minute apical ring with PC illumination (Fig. 12I) and a morphologically distinct anamorph forming stromatic, stipitate, cupulate sporodochial conidiomata.
Saül in 2018 and 2019 in connection with the inventorial ABC project, during which a new species of Striatosphaeria castanea was collected. We thank René Schumacher for a collection of Dendrophoma cytisporoides. We thank both reviewers for their valuable comments and suggestions.
This study was supported by the project of the Czech Science Foundation (GAČR 20-14840S), and as long-term research development projects of the Czech Academy of Sciences, Institute of Botany (RVO 67985939) (M.R.) and the University Hospital Hradec Králové MH CZ -DRO (UHHK, 00179906) (J.N.). This study was also supported by a National Science Foundation award (DEB-0515558) to A.N.M. The WM Keck Center at the University of Illinois Urbana-Champaign is thanked for sequencing services. The field work of M.R. in New Zealand was partly supported by Studienstiftung für mykologische Systematik und Ökologie (2003)