Pseudodidymellaceae fam. nov.: Phylogenetic affiliations of mycopappus-like genera in Dothideomycetes.

The familial placement of four genera, Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina, was taxonomically revised based on morphological observations and phylogenetic analyses of nuclear rDNA SSU, LSU, tef1, and rpb2 sequences. ITS sequences were also provided as barcode markers. A total of 130 sequences were newly obtained from 28 isolates which are phylogenetically related to Melanommataceae (Pleosporales, Dothideomycetes) and its relatives. Phylogenetic analyses and morphological observation of sexual and asexual morphs led to the conclusion that Melanommataceae should be restricted to its type genus Melanomma, which is characterised by ascomata composed of a well-developed, carbonaceous peridium, and an aposphaeria-like coelomycetous asexual morph. Although Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina are phylogenetically related to Melanommataceae, these genera are characterised by epiphyllous, lenticular ascomata with well-developed basal stroma in their sexual morphs, and mycopappus-like propagules in their asexual morphs, which are clearly different from those of Melanomma. Pseudodidymellaceae is proposed to accommodate these four genera. Although Mycodidymella and Xenostigmina have been considered synonyms of Petrakia based on sexual morphology, we show that they are distinct genera. Based on morphological observations, these genera in Pseudodidymellaceae are easily distinguished by their synasexual morphs: sigmoid, multi-septate, thin-walled, hyaline conidia (Mycodidymella); globose to ovoid, dictyosporus, thick-walled, brown conidia with cellular appendages (Petrakia); and clavate with a short rostrum, dictyosporus, thick-walled, brown conidia (Xenostigmina). A synasexual morph of Pseudodidymella was not observed. Although Alpinaria was treated as member of Melanommataceae in a previous study, it has hyaline cells at the base of ascomata and pseudopycnidial, confluent conidiomata which is atypical features in Melanommataceae, and is treated as incertae sedis.

Introduction

The family Melanommataceae (Pleosporales) was proposed for its type genus, Melanomma (Winter 1887). Currently, more than 20 genera with diverse ecological and morphological features are recognised in this family (Tian ). Petrakia and Xenostigmina have epiphyllous, lenticular ascomata with well-developed basal stroma, mycopappus-like propagules, and petrakia- or stigmina-like synasexual morphs, and were also accepted in Melanommataceae (Funk, 1986, Funk and Dorworth, 1988, Crous, 1998, Crous et al., 2009, Butin et al., 2013, Tian et al., 2015). Subsequently, two additional genera, Mycodidymella and Pseudodidymella, were reported to be phylogenetically related to this family (Gross ), although their morphological features were clearly different from those of Melanomma, which is characterised by carbonaceous ascomata, trabecular pseudoparaphyses, and aposphaeria-like coelomycetous asexual morphs (Barr, 1987, Barr, 1990, Lumbsch and Huhndorf, 2007, Kirk et al., 2008, Tian et al., 2015, Jaklitsch and Voglmayr, 2017).

The genus Petrakia was originally characterised by sporodochial conidiomata and muriform, brown conidia with cellular, hyaline appendages (Sydow and Sydow, 1913, Butin et al., 2013). Recently, the complete life cycle of Pe. echinata, which is the type species and a known causal agent of leaf blotch disease of Acer spp., was revealed (Butin ). Subsequently, phylogenetic analysis using large subunit nrDNA sequences indicated that this genus is related to Melanommataceae or Pleomassariaceae (Dothideomycetes; Butin ).

Xenostigmina zilleri, the type species of the genus, is a known pathogen that causes brown spot disease in Acer macrophyllum in Canada (Funk 1986). This species was originally described as Cercosporella aceris (Dearness 1917). Redhead & White (1985) introduced Mycopappus, and transferred two species to this genus, i.e. C. aceris and C. alni. The type species of Mycopappus, Mycop. alni, was suggested to be a member of Sclerotiniaceae (Helotiales, Leotiomycetes) based on its sclerotial morph and phylogenetic analyses using ITS sequences (Takahashi ). Mycopappus aceris was excluded from the genus, because the synasexual morph of this species is the dothideomycetous taxon X. zilleri (Funk and Dorworth, 1988, Crous, 1998, Wei et al., 1998, Crous et al., 2009). According to phylogenetic analysis, this genus was accepted as Melanommataceae (Phookamsak et al., 2014, Tian et al., 2015).

The genera Mycodidymella and Pseudodidymella are also members of Melanommataceae that produce mycopappus-like propagules in their asexual morphs (Wei et al., 1997, Wei et al., 1998, Gross et al., 2017). The genus Mycodidymella, which is based on the type species Mycod. aesculi, is known as a pathogen of concentric ring spot disease in Aesculus turbinata (Wei ). The life cycle of Mycod. aesculi is similar to those of Petrakia and Xenostigmina, except it has sigmoid and hyaline conidia in its synasexual morph. Although the synasexual morph of Petrakia seems to be clearly different from that of Mycodidymella and Xenostigmina, the latter two genera were synonymised with an older name, Petrakia (Jaklitsch & Voglmayr 2017).

The monotypic genus Pseudodidymella was established for Pseudod. fagi (Wei ). The species was found to be associated with brown leaf spots of Fagus crenata in Japan, and was originally characterised by lenticular ascomata with a well-developed basal stroma and a pycnopleiospora-like asexual morph, which is characterised by sporodochial conidiomata and conidia with appendages (Wei ). Mycodidymella is morphologically similar to this genus, but can be distinguished by its pycnopleiospora-like asexual morph (Wei ). Gross discovered Pseudod. fagi on F. sylvatica in Switzerland and suggested that the pycnopleiospora-like asexual morph has mycopappus-like propagules rather than individual conidia. Thus, morphological delimitation of these two genera is problematic and requires further research. According to a phylogenetic study using ITS sequences (Gross ), four genera with mycopappus-like propagules (Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina) formed a strongly supported clade within Melanommataceae sensu lato; however, familial placement and generic validity of each genus remain unresolved.

During our ongoing studies of ascomycetous fungi in Japan (Tanaka et al., 2010, Tanaka et al., 2011, Tanaka et al., 2015, Hashimoto et al., 2015a, Hashimoto et al., 2015b, Hashimoto et al., 2016, Hashimoto et al., 2017), we collected strains which are morphologically similar or phylogenetically related to Melanommataceae sensu lato. The main objectives of the present study were to clarify familial placement of genera in this family, and establish a taxonomic framework of Melanommataceae sensu lato based on morphological observations and molecular phylogenetic analyses of small subunit nrDNA (18S; SSU), large subunit nrDNA (28S; LSU), translation elongation factor 1-α (tef1), and DNA-directed RNA polymerase II second largest subunit (rpb2) sequences. ITS sequences were also obtained as DNA barcode markers.

Materials and methods

Isolates

All fungal structures were studied in preparations mounted in distilled water. Morphological characters were observed by differential interference and phase contrast microscopy (Olympus BX53, Japan), and images captured with an Olympus digital camera (DP21, Japan). A total of 28 single-spore isolates were used for morphological observation and phylogenetic analyses (Table 1).

Table 1

Specimens, isolates and new sequences used in this study.

Species	Original no.	Specimen no.1	Strain no.	Host/substrate	GenBank accession no.2
					SSU	LSU	tef1	rpb2	ITS
Alpinaria rhododendri	KT 2520	HHUF 30554	CBS 142901	Rhododendron brachycarpum	LC203314	LC203360	LC203388	LC203416	LC203335
Melanomma japonicum	KT 2076	HHUF 30539P	CBS 142902	dead wood	LC203290	LC203336	LC203364	LC203392	LC203318
	KT 3028	HHUF 30540P	CBS 142903	Fagus crenata	LC203291	LC203337	LC203365	LC203393	LC203319
	KT 3425	HHUF 30541P	CBS 142904	F. crenata	LC203292	LC203338	LC203366	LC203394	LC203320
	–	HHUF 26520H	CBS 142905	Dead wood	LC203293	LC203339	LC203367	LC203395	LC203321
			= JCM 13124
			= MAFF 239634
Me. pulvis-pyrius	KT 2110	HHUF 30542	CBS 142906	Acer sp.	LC203294	LC203340	LC203368	LC203396	LC203322
	KT 2113	HHUF 30543	CBS 142907	Dead wood	LC203295	LC203341	LC203369	LC203397	LC203323
	AH 375	HHUF 30544	CBS 142908	F. crenata	LC203296	LC203342	LC203370	LC203398	LC203324
	KH 27	HHUF 30545	CBS 142909	Dead wood	LC203297	LC203343	LC203371	LC203399	LC203325
	KH 77	HHUF 30546	CBS 142910	Dead wood	LC203298	LC203344	LC203372	LC203400	LC203326
	KH 86	HHUF 30547	CBS 142911	Dead wood	LC203299	LC203345	LC203373	LC203401	LC203327
	KH 197	HHUF 30548	CBS 142912	Dead wood	LC203300	LC203346	LC203374	LC203402	LC203328
Mycodidymella aesculi	KT 3060	HHUF 30549	CBS 142913	Aesculus turbinata	LC203301	LC203347	LC203375	LC203403	LC203329
	H 2610	HHUF 22892H	CBS 142914	A. turbinata	LC203302	LC203348	LC203376	LC203404	LC194192
	H 2620	–	CBS 142915	A. turbinata	LC203303	LC203349	LC203377	LC203405	LC203330
	AH 560	HHUF 30550	CBS 142916	A. turbinata	LC203304	LC203350	LC203378	LC203406	LC203331
Petrakia echinata	–	–	CBS 133072	Acer pseudoplatanus	LC203305	LC203351	LC203379	LC203407	–
	–	–	CBS 133070	A. pseudoplatanus	LC203306	LC203352	LC203380	LC203408	–
Pseudodidymella fagi	KT 3058	HHUF 30515	CBS 142917	F. crenata	LC203307	LC203353	LC203381	LC203409	LC150785
			= MAFF 245738
	KT 3074-3	HHUF 30516	CBS 142918	F. crenata	LC203308	LC203354	LC203382	LC203410	LC150786
			= MAFF 245739
	RF 5	HHUF 30517	CBS 142919	F. crenata	LC203309	LC203355	LC203383	LC203411	LC150788
			= MAFF 245741
	H 2579	HHUF 22903H	MAFF 245740	F. crenata	LC203310	LC203356	LC203384	LC203412	LC150787
	AH 561	HHUF 30553	CBS 142920	F. crenata	LC203311	LC203357	LC203385	LC203413	LC203332
Pseudod. minima	KT 2918	HHUF 30551H	CBS 142921	Fagus japonica	LC203312	LC203358	LC203386	LC203414	LC203333
			= MAFF 246249
	AH 556	HHUF 30552P	CBS 142922	F. japonica	LC203313	LC203359	LC203387	LC203415	LC203334
Xenostigmina aceris	–	–	CBS 124109	Acer macrophyllum	LC203315	LC203361	LC203389	LC203417	–
	–	–	CBS 115685	Acer sp.	LC203316	LC203362	LC203390	LC203418	–
	–	–	CBS 115686	Acer sp.	LC203317	LC203363	LC203391	LC203419	–

“H”: holotype, “P”: paratype.

Sequences generated in this study are shown in bold.

DNA isolation, amplification and phylogenetic analysis

DNA extraction was carried out with an ISOPLANT II kit (Nippon Gene, Japan) based on the manufacturer's protocol. Sequences of SSU, ITS, LSU, and tef1 and rpb2 were amplified by PCR with the primer pairs NS1/NS4, ITS1/ITS4 (White ), LR0R/LR7 (Rehner and Samuels, 1994, Vilgalys and Hester, 1990), EF1-983F/EF1-2218R (Rehner & Buckley 2005), and fRPB2-5F/fRPB2-7cR (Liu ), respectively. Amplifications were performed in 25 μL volumes that consisted of 2 μL DNA extract, 2.5 μL of 10 × TEMPase Buffer I, 10 mM dNTP mix, 1 μL of each 20-pM primer, 25 mL MgCl2, 14.5 μL MilliQ water, and 0.5 μL TEMPase Hot Start DNA polymerase (Ampliqon, Denmark). PCRs were carried out on a PC 320 thermo-cycler (ASTEC, Japan) as follows: 95 °C for 15 min; followed by 35 cycles of 1 min at 94 °C, 1 min at the designated annealing temperature (42.2 °C for SSU, 61.5 °C for ITS, 46 °C for LSU, 60 °C for tef1, and 58 °C for rpb2), and 1 min at 72 °C; and a final denaturation of 7 min at 72 °C. The PCR products were directly sequenced at SolGent (South Korea).

Newly generated sequences were deposited in GenBank (Table 1). Sequences of 73 taxa of Pleosporales and Hysteriales were also phylogenetically analysed (Table 1). Hysterium pulicare and Hysterobrevium mori (Hysteriaceae, Hysteriales) were used as outgroups. All sequences were aligned using the MUSCLE algorithm as implemented in the program MEGA v. 5 (Tamura ). Phylogenetic analyses were conducted using maximum likelihood (ML) and Bayesian methods. The optimal substitution models for each dataset were estimated by Kakusan4 (Tanabe 2011) based on the Akaike information Criterion (AIC; Akaike 1974) for ML analysis and Bayesian information Criterion (BIC; Schwarz 1978) for the Bayesian analysis.

The ML analysis was performed using TreeFinder Mar 2011 (Jobb 2011) based on the models selected with the AICc4 parameter (a proportional model among genes and codons): J2+G for SSU; GTR+G for LSU; F81+G for the tef1 first codon position, J1ef+G for the tef1 second codon position, and J2+G for the tef1 third codon position; and J2+G for the rpb2 first codon position, J1+G for the rpb2 second codon position, and J2+G for the rpb2 third codon position. Bootstrap percentages (BPs) were obtained by 1 000 bootstrap replications.

Bayesian analysis was performed with MrBayes v. 3.2.2 (Ronquist ) with substitution models for different regions selected with the BIC4 parameter (proportional model among loci and codons): K80+G for SSU; SYM+G for LSU; GTR+G for the tef1 first codon position, JC69+G for the tef1 second codon position, and GTR+G for the tef1 third codon position; and GTR+G for the rpb2 first codon position, GTR+G for the rpb2 second codon position, and GTR+G for the rpb2 third codon position. Two simultaneous, independent runs of Metropolis-coupled Markov chain Monte Carlo (MCMC) were performed for 2 M generations with trees sampled every 1 000 generations. Convergence of the MCMC runs assessed from the average standard deviation of split frequencies (<0.01) and effective sample size scores (all >100) using MrBayes v. 3.2.2 and Tracer v. 1.6 (Rambaut ), respectively. The first 25 % of trees were discarded as burn-in, and the remaining trees were used to calculate 50 % majority rule trees and determine posterior probabilities (PPs) for individual branches. The alignment was submitted to TreeBase under study number S20165.

Morphology

Colony characteristics of cultures grown on 2 % potato dextrose agar (PDA; Difco, France) were observed after 3 wk incubation at 20 °C in the dark. Colours were noted based on those described by Rayner (1970).

To induce sexual or asexual fructification in culture, 5 mm square mycelial agar discs were placed on water agar that included sterilised natural substrate, such as Aesculus turbinata and Fagus crenata leaves and rice straw, and the plates were incubated at 20 °C for 2 wk in the dark. When the substrate was colonised, the plates were incubated at 20 °C under blacklight blue illumination for 2 mo to observe sporulation. Cultures were deposited in the Westerdijk Fungal Biodiversity Institute (CBS), the Japan Collection of Microorganisms (JCM), and the Genebank Project of NARO, Japan (MAFF). Specimens were deposited in the Herbarium of Hirosaki University, Fungi (HHUF).

Results

Phylogeny

The ML and Bayesian phylogenetic analyses were conducted using an aligned sequence dataset composed of 941 nucleotides from SSU, 1 276 from LSU, 886 from tef1, and 1 021 from rpb2. The alignment contained a total of 73 taxa, which consisted of 59 taxa (80.8 %) in SSU, 73 (100 %) in LSU, 63 (86.3 %) in tef1, 51 (69.9 %) in rpb2 (Table 1, Table 2). No significant conflict was observed among individual gene phylogenies, but the familial and generic nodes mostly lacked significant support in SSU and LSU phylogenetic trees generated (data not shown). However, this combined dataset provided higher confidence values for the familial level than did those of the individual gene trees (data not shown). Of the 3 824 characters included in the alignment, 1 205 were variable and 2 844 were conserved. The ML tree with the highest log likelihood (−26580.8637) is shown in Fig. 1. The Bayesian likelihood score was −26638.727. The topology recovered by the Bayesian analysis was almost identical to that of the ML tree, except for the position of Aposphaeria corallinolutea, Bertiella macrospora, Herpotrichia macrotricha, Phragmocephala atra, Pseudostrickeria murigormis and Sarimanas shirakamiense.

Table 2

GenBank accession numbers of species used in the phylogenetic study.

Species name	Family	Strain no.1	GenBank accession no.
			SSU	LSU	tef1	rpb2
Alpinaria rhododendri	incertae sedis	ANM 73	–	GU385198	–	–
A. rhododendri	incertae sedis	CBS 141994E	KY190004	KY189973	KY190009	KY189989
Alternaria alternata	Pleosporaceae	CBS 916.96E	DQ678031	DQ678082	DQ677927	DQ677980
Aposphaeria corallinolutea	incertae sedis	CBS 131287H	–	JF740330	–	–
Bertiella macrospora	incertae sedis	IL 5005	–	GU385150	–	–
Beverwykella pulmonaria	incertae sedis	CBS 283.53H	KY190005	GU301804	–	GU371768
Byssosphaeria jamaicana	incertae sedis	SMH 1403	–	GU385152	GU327746	–
B. rhodomphala	incertae sedis	GKM L153N	–	GU385157	GU327747	–
B. salebrosa	incertae sedis	SMH 2387	–	GU385162	GU327748	–
B. schiedermayeriana	incertae sedis	SMH 3157	–	GU385163	GU327745	–
B. siamensis	incertae sedis	MFLUCC 10-0099H	KT289897	KT289895	KT962059	KT962061
B. villosa	incertae sedis	GKM 204N	–	GU385151	GU327751	–
Corynespora cassiicola	Corynesporascaceae	CBS 100822	GU296144	GU301808	GU349052	GU371742
Cyclothyriella rubronotata	Cyclothyriellaceae	CBS 141486E	KX650507	KX650544	KX650519	KX650574
Gemmamyces piceae	incertae sedis	CBS 141555	KY190006	KY189976	KY190011	KY189992
Herpotrichia diffusa	incertae sedis	CBS 250.62	DQ678019	DQ678071	DQ677915	DQ677968
H. juniperi	incertae sedis	CBS 200.31	DQ678029	DQ678080	DQ677925	DQ677978
H. macrotricha	incertae sedis	GKM 196N	–	GU385176	GU327755	–
H. vaginatispora	incertae sedis	MFLUCC 13-0865H	KT934256	KT934252	KT934260	–
Hysterium pulicare	Hysteriaceae	CBS 123377	FJ161161	FJ161201	FJ161109	FJ161127
Hysterobrevium mori	Hysteriaceae	CBS 123563	FJ161155	FJ161196	FJ161104	–
Leptosphaeria doliolum	Leptosphaeriaceae	CBS 505.75	GU296159	GU301827	GU349069	KT389640
Lophiostoma arundinis	Lophiostomataceae	CBS 621.86	DQ782383	DQ782384	DQ782387	DQ782386
Massaria inquinans	Massariaceae	CBS 125591E	HQ599442	HQ599400	HQ599340	–
Massarina eburnea	Massarinaceae	CBS 473.64	GU296170	GU301840	GU349040	GU371732
Melanomma populina	Melanommataceae	CBS 543.70E	EU754031	EU754130	–	–
M. populina	Melanommataceae	CBS 350.82	–	JF740265	–	–
M. pulvis-pyrius	Melanommataceae	CBS 124080E	GU456302	GU456323	GU456265	GU456350
M. pulvis-pyrius	Melanommataceae	CBS 109.77	FJ201987	FJ201986	GU456274	GU456359
M. pulvis-pyrius	Melanommataceae	CBS 371.75	FJ201989	FJ201988	GU349019	GU371798
Muriformistrickeria rubi	incertae sedis	MFLUCC 15-0681H	KT934257	KT934253	KT934261	–
Neoophiosphaerella sasicola	Lentitheciaceae	MAFF 239644E	AB524458	AB524599	AB539111	AB539098
Nigrograna obliqua	Nigrogranaceae	CBS 141475P	KX650512	KX650558	KX650530	KX650579
Phragmocephala atra	incertae sedis	MFLUCC 15-0021	KP698729	KP698725	–	–
Praetumpfia obducens	incertae sedis	CBS 141474E	KY190008	KY189984	KY190019	KY190000
Prosthemium betulinum	Pleomassariaceae	CBS 279.74	DQ678027	DQ678078	DQ677923	KT216532
Prosthemium canba	Pleomassariaceae	KT 2083-1	AB553646	AB553760	–	–
Pseudostrickeria	incertae sedis	MFLUCC 13-0764H	KT934258	KT934254	KT934262	–
muriformis
Pseudotrichia mutabilis	incertae sedis	PM 1	–	KY189988	KY190022	KY190003
Roussoella verrucispora	Thyridariaceae	CBS 125434H	AB524481	AB524622	AB539115	AB539102
Sarimanas shirakamiense	incertae sedis	KT 3000H	LC001712	LC001715	–	–
Seifertia azaleae	incertae sedis	DAOM 239136	–	EU030276	–	–
S. shangrilaensis	incertae sedis	MFLUCC 16-0238H	KU954102	KU954100	KU954101	–
Teichospora trabicola	Teichosporaceae	CBS 140730E	–	KU601591	KU601601	KU601600
Tumularia tuberculata	incertae sedis	CBS 256.84	–	GU301851	GU349006	–

“H”: ex-holotype, “P”: ex-paratype, “E”: ex-epitype.

Fig. 1

Maximum-likelihood (ML) tree of Melanommataceae sensu stricto and Pseudodidymellaceae with its relatives. ML bootstrap percentages (BP) greater than 60 % and Bayesian posterior probabilities (PP) above 0.95 are presented at the nodes as ML BP/ Bayesian PP. A hyphen (“-”) indicates values lower than 60 % BP or 0.95 PP, and a node not present in the Bayesian analysis is shown with “x”. Ex-holotype, paratype, epitype, strains are indicated in with a superscript H, P and E, respectively. The newly obtained sequences are shown in bold. The scale bar represents nucleotide substitution per site.

Monophyly of the genera with mycopappus-like propagules (Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina) was well-supported (91 % ML BP/ 1.00 Bayesian PP). Although these four genera are phylogenetically related to Melanommataceae sensu lato, their morphological and ecological features are clearly distinct from those of the type genus Melanomma. Therefore, we establish a new family, Pseudodidymellaceae, to accommodate these genera with mycopappus-like propagules. Results from phylogenetic analyses of this study indicate that Alpinaria, formerly classified in Melanommataceae sensu lato (Jaklitsch & Voglmayr 2017), is phylogenetically distant from Melanommataceae sensu stricto (Fig. 1), but its familial placement is unresolved.

Taxonomy

Two families, including a new family (Pseudodidymellaceae), four genera, and seven species, including two new species and one new combination (Melanomma japonicum, Pseudodidymella minima, and Xenostimgmina aceris) are described below.

G. Winter [as ‘Melanommeae’], Rabenh. Krypt.-Fl., Edn 2 (Leipzig) 1.2: 220. 1887.

Saprobic on dead twigs of woody plants. Sexual morph: Ascomata globose to ovoid, immersed to superficial, gregarious, ostiolate. Peridium composed of thick-walled, pseudoparenchymatous, hyaline to brown cells. Pseudoparaphyses trabeculate, septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 8-spored. Ascospores olive brown, multi-septate, smooth. Asexual morph: Conidiomata pycnidial, globose to subglobose, superficial, black, ostiolate. Peridium composed of elongate, brown cells. Conidiophores absent. Conidiogenous cells holoblastic, ampliform to cylindrical, hyaline. Conidia ellipsoidal, hyaline, smooth, aseptate.

Type genus: Melanomma Nitschke ex Fuckel.

Notes: Melanommataceae was established by Winter (1887). Byssosphaeria, Keissleriella, Melanomma, Ostropella, and Strickeria have been referred to as members of Melanommataceae, and this family was characterised by gregarious ascomata composed of well-developed, carbonaceous or coriaceous peridium, trabecular pseudoparaphyses, and aposphaeria-like coelomycetous asexual morphs (Barr 1987). This familial concept was supported in “Outline of Ascomycota – 2007” for 18 genera (Lumbsch & Huhndorf 2007).

A study by Mugambi & Huhndorf (2009) on LSU and tef1 sequences showed that Melanommataceae is composed of Byssosphaeria, Herpotrichia, Melanomma, and Pseudotrichia, and previous familial concepts did not reflect natural relationships. Several genera, such as Keissleriella and Ostropella, were phylogenetically scattered in other Pleosporales (Mugambi and Huhndorf, 2009, Zhang et al., 2012, Tanaka et al., 2015), and Strickeria was placed in Sporocadaceae (Xylariales, Sordariomycetes) (Jaklitsch ). It was clear that the traditional concept of Melanommataceae is polyphyletic and needed revision (Kirk et al., 2008, Mugambi and Huhndorf, 2009, Hyde et al., 2013). Later, two genera, Tumularia (as Monotosporella) and Phragmocephala, which have mononematous or synnematous conidiophores in their asexual morphs, were reported in Melanommataceae (Schoch et al., 2009, Su et al., 2015). Wijayawardene et al., 2012, Wijayawardene et al., 2014 also listed additional dematiaceous genera, Exosporiella and Nigrolentilocus, as members of this family without molecular evidence. A broad concept of Melanommataceae was proposed by Tian and Jaklitsch & Voglmayr (2017), and Mycodidymella, Petrakia and Xenostigmina were treated as members of this family. However, the results of our phylogenetic analyses and morphological observations indicate that Melanommataceae should be restricted to its type genus, Melanomma.

Nitschke ex Fuckel, Jb. nassau. Ver. Naturk. 23–24: 159. 1870 (1869–1870).

Synonym: Moriolopis Norman ex Keissl., Nytt Mag. Natur. 66: 88. 1927.

Saprobic on dead twigs of woody plants. Sexual morph: Ascomata globose to ovoid, immersed or erumpent to superficial, gregarious, with a short ostiolar neck. Peridium composed of thick-walled, pseudoparenchymatous, hyaline to brown cells. Pseudoparaphyses trabecular, septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 8-spored. Ascospores olive brown, sometimes with paler ends, strongly or slightly curved, multi-septate, smooth. Asexual morph: Conidiomata pycnidial, globose to subglobose, superficial, black, with a papillate ostiole. Peridium composed of elongate, brown cells. Conidiophores absent. Conidiogenous cells holoblastic, ampliform to cylindrical, hyaline, smooth. Conidia ellipsoidal, hyaline, smooth, aseptate.

Type species: Melanomma pulvis-pyrius (Pers.) Fuckel.

Notes: The genus Melanomma was established by Fuckel (1870). Species in this genus are known to be saprobes on decaying plant material or weak plant pathogens (Chesters, 1938, Holm, 1957, Zhang et al., 2008). Melanomma pulvis-pyrius is a well-studied, widespread species in this genus. However, other species have rarely been reported or have not been recorded since their initial description. Only a few species have received modern taxonomic treatment (Holm, 1957, Mathiassen, 1989, Mathiassen, 1993, Barr, 1990), although approximately 300 epithets are listed in Index Fungorum (http://indexfungorum.org). Asexual morphs of this genus were reported to be aposphaeria-like coelomycetes or Nigrolentilocus (Ichinoe, 1970, Sivanesan, 1984, Castañeda-Ruiz et al., 2001, Sánchez and Bianchinotti, 2015, Tian et al., 2015).

A. Hashim. & Kaz. Tanaka, sp. nov. MycoBank MB819613; Fig. 2.

Fig. 2

Melanomma japonicum.A, B. Ascomata on substrate. C. Ascoma in longitudinal section. D. Lateral peridium of ascoma. E. Basal peridium of ascoma. F. Ascus. G. Apex of ascus. H. Stipe of ascus. I. Pseudoparaphyses. J–L. Ascospores. M–O. Conidiomata in culture. P. Conidioma in longitudinal section. Q. Peridium of conidioma. R, S. Conidiogenous cells. T. Conidia. U. Germinating conidia. A, C–J from HHUF 26520; B, K, L from HHUF 30540; M–O from culture CBS 142903; P–U from culture CBS 142905 = JCM 13124 = MAFF 239634. Scale bars: A, M = 500 μm; B = 200 μm; C, N–P = 100 μm; D, E, G–L, R–U = 5 μm; F, Q = 10 μm.

Etymology: Referring to its country of origin, Japan.

Saprobic on dead twigs of woody plants. Sexual morph: Ascomata globose to ovoid, superficial, gregarious, 190–320 μm diam, 200–340 μm high. Ostiolar neck short papillate, composed of carbonaceous, thick-walled, black cells. Peridium 40–60 μm thick of two layers at side; outer layer 25–40 μm thick of elongate, thin-walled, 12–20 × 3–4 μm, brown cells; inner layer 12.5–30 μm thick of globose to rectangular, 10–17.5 × 5–7 μm, hyaline cells; base of ascomata 40–53 μm thick, of two layers; outer layer 15–30 μm thick of elongate, thin-walled, 3.5–7.5 × 3.5–5 μm, brown cells; inner layer 10–30 μm thick of globose to rectangular, 7–10.5 × 6–9 μm, brown cells. Pseudoparaphyses trabeculate, 0.5 μm wide, septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 73–105 × 5.5–9 μm ( = 89.9 × 7 μm, n = 26), with a short stipe (7–16 μm long,  = 10.3 μm, n = 20), apically rounded with an ocular chamber, 8-spored. Ascospores fusiform, with broad rounded ends, straight to slightly curved, 12–19 × 3–7 μm ( = 15.1 × 4.6 μm, n = 151), l/w 2.5–4.9 ( = 3.4, n = 151), 3-septate, with a primary septum nearly median (0.44–0.57,  = 0.51, n = 75), olive brown, sometimes with paler ends, constricted at the septa, smooth. Asexual morph: Conidiomata pycnidial, globose to subglobose, up to 230 μm high in section, 150–250 μm diam, semi-immersed, solitary. Ostiolar neck short papillate, composed of thick-walled, black cells. Peridium 12–33.5 μm wide, composed of 8.5–16.5 × 3.5–7.5 μm, rectangular, brown cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic, 8–13.5 × 2–3 μm, cylindrical, hyaline, smooth. Conidia cylindrical with rounded ends, 3–4 × 2–2.5 μm ( = 3.3 × 2.2 μm, n = 50), l/w 1.1–2.1 ( = 1.5, n = 50), hyaline, smooth, aseptate, guttulate when young.

Culture characteristics: Colonies on PDA attaining 25–27 mm diam within 21 d in the dark, floccose, centrally raised, smoke grey (Rayner 1970), grey olivaceous at centre; reverse smoke grey, grey olivaceous at margin (Fig. 8A); asexual morph formed.

Fig. 8

Colony characters of Melanomma spp. and Pseudodidymellaceae spp. used in this study on PDA within 3 wk at 20 °C in the dark (left: upper, right: reverse). A.Melanomma japonicum (CBS 142905 = JCM 13124 = MAFF 239634, ex-holotype culture). B.Me. pulvis-pyrius (CBS 142908). C.Mycodidymella aesculi (CBS 142914, ex-holotype culture). D.Pseudodidymella fagi (MAFF 245740, ex-holotype culture of Pycnopleiospora fagi). E.Pseudod. minima (CBS 142921 = MAFF 246249, ex-holotype culture). F.Alpinaria rhododendri (CBS 142901). Scale bar: A–F = 1 cm

Specimens examined: Japan, Aomori, Hakkoda, Okiagetai, on dead twigs of woody plant, 15 Apr. 2006, K. Tanaka, KT 2076 (HHUF 30539 paratype, ex-paratype living culture CBS 142902); Akita, Kazuno, Hachimantai, Yakeyama, Mousen pass, on dead twigs of Fagus crenata, 24 Jun. 2012, K. Tanaka, KT 3028 (HHUF 30540 paratype, ex-paratype living culture CBS 142903); Kagoshima, Tarumizu, Mt. Oonogara, on dead twigs of Fagus crenata, 25 Oct. 2013, K. Tanaka, KT 3425 (HHUF 30541 paratype, ex-paratype living culture CBS 142904); Aomori, Hakkoda, near Yunotai, on dead twigs of woody plant, 21 Jul. 2001, Y. Harada (HHUF 26520 holotype designated here, ex-holotype living culture CBS 142905 = JCM 13124 = MAFF 239634).

Notes: This species is morphologically closest to Me. pulvis-pyrius in ascospore size, but the size of conidia of this species is slightly longer and slenderer (3–4 μm vs. (2–)2.5–3.5 μm long; 1.1–2.1 vs. 1.0–1.7 length/width). ITS sequences of these two species differed by 13 positions with one gap.

(Pers.) Fuckel, Jb. nassau. Ver. Naturk. 23–24: 160. 1870 (1869–1870). Fig. 3.

Fig. 3

Melanomma pulvis-pyrius.A, B. Ascomata on substrate. C. Ascoma in longitudinal section. D. Lateral peridium of ascoma. E. Basal peridium of ascoma. F. Pseudoparaphyses. G. Ascus. H. Apex of ascus. I. Stipe of ascus. J, K. Ascospores. L. Germinating ascospore. M, N. Conidiomata in culture. O. Conidioma in longitudinal section. P. Peridium of conidioma. Q, R. Conidiogenous cells. S. Conidia. T. Germinating conidia. A–F, J–L from HHUF 30544; G–I from HHUF 30543; M–Q, T from culture CBS 142912; R, S from culture CBS 142908. Scale bars: A, M = 500 μm; B, N = 200 μm; C, O = 100 μm; D, E, P = 10 μm; F–L, Q–T = 5 μm.

Saprobic on dead twigs of woody plants. Sexual morph: Ascomata globose to ovoid, 210–310(–410) μm diam. Ostiolar neck short papillate, composed of carbonaceous cells. Peridium 75–88 μm thick of two layers at side; outer layer 35–45 μm thick; inner layer 30–40 μm thick, 65–75 μm thick at base. Pseudoparaphyses trabeculate, 1–1.5 μm wide. Asci 71–92 × 5–8.5 μm ( = 82.1 × 6.3 μm, n = 14), with a short stipe (5–8 μm long,  = 5.7 μm, n = 12). Ascospores 11.5–15.5 × 4–5 μm ( = 13 × 4.2 μm, n = 75), l/w 2.5–3.6 ( = 3.1, n = 75), 3-septate, with a primary septum nearly median (0.45–0.58,  = 0.50, n = 75). Asexual morph: Conidiomata pycnidial, globose to subglobose, 160–300 μm diam, with a papillate ostiolar neck. Peridium 18.5–22 μm wide, composed of 4–16.5 × 2.5–5 μm, rectangular, brown cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic, 8–17.5 × 1.5–4 μm, cylindrical, hyaline, smooth. Conidia cylindrical with rounded ends, (2–)2.5–3.5 × 2–2.5(–3) μm ( = 2.9 × 2.3 μm, n = 50), l/w 1.0–1.7 ( = 1.3, n = 50), hyaline, smooth, aseptate, guttulate when young.

Culture characteristics: Colonies on PDA attaining 22–24 mm diam within 21 d, floccose, fasciculate, centrally raised, pale olivaceous grey; reverse greyish sepia, olivaceous buff at margin (Fig. 8B); asexual morph formed.

Specimens examined: Japan, Aomori, Minamitsugaru, Owani, on dead twigs of Acer mono var. mayrii, 1 Jul. 2006, K. Tanaka, KT 2110 (HHUF 30542, culture CBS 142906); Hirosaki, Zatoishi, on dead twigs of woody plant, 8 Jul. 2006, H. Yonezawa, KT 2113 (HHUF 30543, culture CBS 142907); Noheji, near Mt. Eboshi, on dead twigs of Fagus crenata, 2 Sep. 2015, A. Hashimoto et al., AH 375 (HHUF 30544, culture CBS 142908); Nishimeya, Ooshirosawa stream, on dead twigs of woody plant, 25 Jun. 2007, K. Hirayama et al., KH 27 (HHUF 30545, culture CBS 142909); on dead twigs of woody plant, 21 Jul. 2007, K. Hirayama et al., KH 77 (HHUF 30546, culture CBS 142910); Kawaratai, Ookawazoe, on dead twigs of woody plant, 28 Aug. 2007, K. Hirayama et al., KH 86 (HHUF 30547, culture CBS 142911); on dead twigs of woody plant, 30 Aug. 2008, K. Hirayama et al., KH 197 (HHUF 30548, culture CBS 142912).

Notes: The above specimens were identified as Me. pulvis-pyrius, the type species of Melanomma. The size of ascospores in our materials was almost identical to that of Me. pulvis-pyrius reported by Holm (1957), who observed the neotype of this species. The rpb2 sequences of our isolates were identical or had one or two differences compared with those of Me. pulvis-pyrius (GU456350) obtained from the ex-epitype culture (CBS 124080).

Melanomma pulvis-pyrius is a well-studied species in Melanomma; its taxonomy and ontogeny of sexual morphs have been described (Chesters 1938), and it has been reported worldwide (Holm, 1957, Sivanesan, 1984, Vassilieva, 1987, Vasyagina et al., 1987, Romero, 1998, Mathiassen, 1989, Mathiassen, 1993, Zhang et al., 2008, Mugambi and Huhndorf, 2009, Jaklitsch and Voglmayr, 2017). However, this is the first report of Me. pulvis-pyrius from Japan. This species was epitypified by Zhang based on a specimen collected from Salix caprea in France.

In the phylogenetic tree, Me. pulvis-pyrius clustered with Me. populina (CBS 543.70 and CBS 350.82) with moderate to strong support (93 % ML BP/ 1.00 Bayesian PP). Because we could not compare the characters of these two species, further study is needed in the future to confirm whether these two species are conspecific.

A. Hashim. & Kaz. Tanaka, fam. nov. MycoBank MB819614.

Parasitic on living leaves of woody plants. Sexual morph: Ascomata subglobose to lenticular, immersed, ostiolate. Peridium pale brown to brown, distinctly thickened at base. Pseudoparaphyses septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 8-spored. Ascospores fusiform with rounded ends, straight, 1-septate, hyaline, smooth. Spermatia cylindrical, hyaline. Asexual morph: Propagules epiphyllous, white to yellowish, globose to subglobose, multicellular, with numerous, flexuous, cylindrical, multi-septate hyphal appendages, detached at stroma-like base composed of subglobose to oblong, hyaline to yellow cells. Synasexual morph: Conidiomata sporodochial, superficial. Stromata composed of globose to subglobose cells. Conidiophores reduced. Conidiogenous cells annellidic or holoblastic. Conidia clavate, sigmoid or rounded to oval or broadly ellipsoidal, phragmosporous to muriform, hyaline to brown, falcate to sigmoid.

Type genus: Pseudodidymella C.Z. Wei et al.

Notes: Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina have mycopappus-like propagules in their life cycles. Although sexual morphs of these genera were reported, and several molecular studies were performed, the phylogenetic placement of these genera remains unresolved (Crous et al., 2009, Butin et al., 2013, Li et al., 2016, Gross et al., 2017). According to the multi-locus phylogenies, these genera are closely related to each other (Li et al., 2016, Gross et al., 2017, Jaklitsch and Voglmayr, 2017). Based on phylogenetic study, Phookamsak proposed to include Petrakia and Xenostigmina in Melanommataceae. Tian accepted these two genera in Melanommataceae in a subsequent study. In our study, the monophyly of these four genera with mycopappus-like propagules was strongly supported (91 % ML BP/ 1.00 Bayesian PP; Fig. 1). Therefore, we introduce a new family, Pseudodidymellaceae, to accommodate the above four genera. Species in this family bear several common features, including sexual morphs with lenticular and subcuticular ascomata erumpent from host tissue, asexual morphs with mycopappus-like propagules, and with or without a synasexual morph that has sporodochial conidiomata. Pseudodidymellaceae can be distinguished from Melanommataceae sensu stricto based on the presence of mycopappus-like propagules.

C.Z. Wei et al., Mycologia 90: 336. 1998.

Synonym: Blastostroma C.Z. Wei et al., Mycologia 90: 337. 1998.

Parasitic on living leaves of woody plant. Sexual morph: Ascomata subglobose to lenticular, immersed, ostiolate. Peridium with rim-like side wall, composed of rectangular, thin-walled, pale brown cells, well-developed at base. Pseudoparaphyses septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 8-spored. Ascospores fusiform, 1-septate, hyaline, smooth. Spermatia cylindrical, hyaline. Asexual morph: Propagules epiphyllous, white to yellowish, globose to subglobose, multicellular; main bodies subglobose to oblong, bearing numerous, unbranched, flexuous, cylindrical, multi-septate hyphal appendages. Synasexual morph: Conidiomata sporodochial, white to yellowish. Stromata composed of globose to subglobose cells. Conidiophores absent. Conidiogenous cells holoblastic, hyaline. Conidia falcate to sigmoid, hyaline, multi-septate, obtuse at the apex, truncate at the base.

Type species: Mycodidymella aesculi C.Z. Wei et al.

C.Z. Wei et al., Mycologia 90: 336. 1998. Fig. 4.

Fig. 4

Mycodidymella aesculi.A, B. Ascomata on substrate. C. Ascoma in longitudinal section. D. Peridium of ascoma. E. Ascus. F. Apex of ascus. G. Stipe of ascus. H, I. Ascospores. J–L. Spermatogonia in culture. M. Spermatogonium in longitudinal section. N. Perdium of spermatogonium. O, P. Spermatogenous cells. Q, R. Spermatia. S, T. Leaves of Aesculus turninata with necrotic brown spots. U, V. Propagules on the leaf surface. W, X. Propagules. Y. Appendage of propagule. Z, AA. Sporodochia on the leaf surface. AB. Sporodochium in longitudinal section. AC. Stroma of sporodochium. AD. Conidiogenous cells. AE, AF. Conidia. A–I from HHUF 22892. J–R from culture CBS 142913. S, T from HHUF 30550. U–Y from HHUF 30549. Z–AF from HHUF 23078. Scale bars: A, J, T, Z = 500 μm; B, K, L, U, V = 250 μm; C, M, W, X, AA, AB = 50 μm; D, E, N, Y, AC, AE, AF = 10 μm; F–I, O–R, AD = 5 μm.

Synonyms: Blastostroma aesculi C.Z. Wei et al., Mycologia 90: 338. 1998.

Mycopappus aesculi C.Z. Wei et al., Mycologia 90: 336. 1998.

Petrakia aesculi (C.Z. Wei et al.) Jaklitsch & Voglmayr, Sydowia 69: 91. 2017.

Parasitic on living leaves of Aesculus turbinata. Sexual morph: Ascomata subglobose to lenticular, solitary to 3–5 grouped, immersed, up to 210 μm high, 260–380 μm diam. Ostiolar neck short papillate, composed of thick-walled, black cells. Peridium 17.5–27.5 μm thick at side, with rim-like side wall, composed of rectangular, thin-walled, 10–13.5 × 6–9 μm, pale brown cells, at base 105–140 μm thick, composed of 8.5–11.5 × 6.5–8.5 μm, hyaline to pale brown cells. Pseudoparaphyses numerous, trabeculate, 0.8–1.3 μm wide, septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 45.5–60 × 7–12.5 μm ( = 53.3 × 10 μm, n = 20), with or without a short stipe, apically rounded with an ocular chamber, 8-spored. Ascospores fusiform with rounded ends, straight, 16–21.5 × 3–4.5 μm ( = 18.6 × 3.9 μm, n = 21), l/w 4.3–5.3 ( = 4.7, n = 21), with a septum nearly median (0.44–0.55,  = 0.51, n = 21), constricted at the septum, hyaline, smooth, guttulate when young. Spermatia 3–5 × 1–2 μm ( = 3.6 × 1.5 μm, n = 50), l/w 1.7–3.7 ( = 2.5, n = 50), cylindrical, hyaline. Asexual morph: Propagules epiphyllous, white to yellowish, globose to subglobose, 200–565 μm diam ( = 331.9 μm, n = 30); main bodies subglobose to oblong, 85–193 × 116–228 μm ( = 127.6 × 152.4, n = 30), composed of 7.5–10 μm diam cells; hyphal appendages 19 to 37, unbranched flexuous, cylindrical, 3–7-septate, 72–150 × 3.5–5.5 μm ( = 111.5 × 4.6, n = 30). Synasexual morph: Conidiomata sporodochial, white to yellowish. Stromata 15–20 μm thick, composed of hyaline, globose to subglobose cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic, hyaline, smooth, 9–12 × 4–5.5 μm. Conidia falcate to sigmoid, 57–94 × 5.5–8.5 μm ( = 75.8 × 6.8, n = 50), hyaline, 8–13-septate, obtuse at the apex, truncate at the base.

Culture characteristics: Colonies on PDA attaining 31–40 mm diam within 21 d, velvety, floccose, centrally raised, buff, grey olivaceous at centre; reverse buff; grey olivaceous at centre (Fig. 8C); spermatial, asexual and synasexual morphs formed.

Specimens examined: Japan, Aomori, Minamitsugaru, Owani, on living leaves of Aesculus turbinata, 12 Aug. 2012, K. Tanaka et al., KT 3060 (HHUF 30549, culture CBS 142913); Nishimeya, Kawaratai, Ookawazoe, near Annmon waterfall trail, on living leaves of Aesculus turbinata, 4 Oct. 1995, C. Z. Wei & Y. Harada (HHUF 23078 holotype of Blastostroma aesculi); 10 Sep. 2016, A. Hashimoto, AH 560 (HHUF 30550, culture CBS 142916); Hirakawa, Ikarigaseki, on living leaves of Aesculus turbinata, 18 Apr. 1995, C. Z. Wei & Y. Harada, H 2610 (HHUF 22892 holotype of Mycodidymella aesculi, ex-holotype living culture CBS 142914); on living leaves of Aesculus turbinata, 18 Apr. 1995, C. Z. Wei & Y. Harada, H 2620 (culture CBS 142915).

Notes: The genus Mycodidymella was established to accommodate a single species, Mycod. aesculi, and this species causes large concentric leaf spots on Aesculus turbinata in Japan (Wei ). This species is morphologically characterised by lenticular ascomata and 1-septate, hyaline ascospores in the sexual morph, mycopappus-like propagules in the asexual morph, and blastostroma-like sigmoid conidia in the synasexual morph. The sexual morph of this species morphologically resembles those of Didymella or Pseudodidymella. Wei assigned this genus to Phaeosphaeriaceae based on morphology. Later, familial placement of this genus was treated as incertae sedis in Dothideomycetes (Lumbsch & Huhndorf 2007). Recently, Butin described the sexual morph of Pe. echinata, which is the type species of Petrakia; they found that the sexual morphology of Petrakia matches that of Mycodidymella and thus synonymised Mycodidymella with Petrakia (Butin ). This proposal was accepted by subsequent studies (Tian et al., 2015, Li et al., 2016, Jaklitsch and Voglmayr, 2017). However, Mycod. aesculi was not included in their analyses. Our phylogenetic study revealed that their monophyletic status was not supported in any analyses (below 60 % ML BP/ 0.95 Bayesian PP, Fig. 1). We retained Mycodidymella as a natural genus in Pseudodidymellaceae (discussed below).

C.Z. Wei et al., Mycologia 89: 496. 1997.

Synonym: Pycnopleiospora C.Z. Wei et al., Mycologia 89: 496. 1997.

Parasitic on living leaves of Fagus spp. Sexual morph: Ascomata subglobose to lenticular, solitary to grouped, immersed, ostiolate. Peridium composed of rectangular, thin-walled, pale brown cells, well-developed at base. Pseudoparaphyses septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 8-spored. Ascospores fusiform with rounded ends, 1-septate, hyaline, smooth. Spermatia cylindrical, hyaline. Asexual morph: Propagules epiphyllous, white to yellowish, globose, multicellular; main bodies globose, subglobose, hyaline to yellow, bearing numerous, unbranched, flexuous, multi-septate hyphal appendages.

Type species: Pseudodidymella fagi C.Z. Wei et al.

Notes: The genus Pseudodidymella, based on the type species Pseudod. fagi, has lenticular ascomata and a pycnopleiospora-like asexual morph which is characterised by sporodochial conidiomata and appendage-bearing conidia (Wei ). Because its sexual morph morphologically resembles that of Didymella, this genus was considered a member of Phaeosphaeriaceae (Wei ). The sexual morph of this genus superficially resembles that of Mycodidymella, but it can be distinguished based on its pycnopleiospora-like asexual morph (Wei ). Since then, this genus has been treated as incertae sedis in Dothideomycetes (Lumbsch & Huhndorf 2007). Gross discovered Pseudod. fagi on Fagus sylvatica in Switzerland; they noted that the asexual morph of this species was previously recorded as Pycnopleiospora, but actually has mycopappus-like propagules rather than individual conidia, and the original description (Wei ) seemed to misinterpret over-mature propagules. They also confirmed that Pseudodidymella is phylogenetically related to other mycopappus-forming genera, such as Mycodidymella, Petrakia, and Xenostigmina, based on the ITS phylogeny. Thus, morphological delimitation of Pseudodidymella and Mycodidymella is problematic and requires further research. In the present study, we recollected Pseudod. fagi from its type locality, and compared the fresh materials to the holotype of Py. fagi. Based on morphological and phylogenetic comparisons of these specimens, we also conclude that Wei misinterpreted the pieces of broken overmatured mycopappus-like propagules (Fig. 5AA and AB) as conidia of Pseudodidymella, but Pseudodidymella actually has mycopappus-like propagules in its asexual morph.

Fig. 5

Pseudodidymella fagi.A, B. Ascomata on substrate. C. Ascoma in longitudinal section. D. Peridium of ascoma. E. Ascus. F. Apex of ascus. G. Stipe of ascus. H, I. Ascospores. J–L. Spermatogonia in culture. M. Spermatogonium in longitudinal section. N. Perdium of spermatogonium. O, P. Spermatogenous cells. Q, R. Spermatia. S, T. Leaves of Fagus crenata with necrotic brown spots. U, V. Propagules on the leaf surface. W, X. Propagules. Y–AB. Appendages of propagule. A–I from HHUF 22903. J–R from culture CBS 142917 = MAFF 245738. S, T from HHUF 30553. U, X, AA, AB from HHUF 30516. V, W, Z from HHUF 23672. Y from HHUF 30517. Scale bars: A, J, T = 500 μm; B, K, L, U, V = 250 μm; C, M, W, X = 50 μm; D, E, N, Y–AB = 10 μm; F–I, O–R = 5 μm.

Species in this genus bear common features, with more than 60 hyphal appendages in mycopappus-like propagules. Although other related genera have sporodochial synasexual morphs, no synasexual morph is known from Pseudodidymella (Wei et al., 1997, Gross et al., 2017, present study). Morphologically, Pseudodidymella resembles Mycodidymella, but can be distinguished based on the rim-like walls of the ascomata, and numerous hyphal appendages in the asexual morph.

C.Z. Wei et al., Mycologia 89: 496. 1997. Fig. 5.

Synonym: Pycnopleiospora fagi C.Z. Wei et al., Mycologia 89: 496. 1997.

Parasitic on living leaves of Fagus crenata. Sexual morph: Ascomata subglobose to lenticular, solitary to 3–5 grouped, immersed, up to 175 μm high, 200–300 μm diam. Ostiolar neck short papillate, composed of thick-walled, black cells. Peridium 20–22 μm thick at side, composed of rectangular, thin-walled, 7.5–10.5 × 6.5–8.5 μm, pale brown cells, at base 58–67 μm thick, composed of 10–13.5 × 5–11.5 μm, hyaline to pale brown cells. Pseudoparaphyses numerous, 1–2 μm wide, septate, branched and anastomosed. Asci bitunicate, fissitunicate, cylindrical, 49–76.5 × 10–14 μm ( = 60.3 × 11.5 μm, n = 20), with a short stipe (3.5–8 μm long,  = 6.1 μm, n = 20), apically rounded with an ocular chamber, 8-spored. Ascospores fusiform with rounded ends, straight, 18.5–24 × 4–5 μm ( = 20.5 × 4.3 μm, n = 20), l/w 4.3–5.6 ( = 4.8, n = 20), with a septum nearly median (0.47–0.58,  = 0.52, n = 20), constricted at the septum, hyaline, smooth, guttulate when young. Spermatia 3–5 × 1–1.5 μm ( = 3.9 × 1.2 μm, n = 50), l/w 2.1–4.8 ( = 3.3, n = 50), cylindrical, hyaline. Asexual morph: Propagules epiphyllous, white to yellowish, globose, 290–500 μm diam ( = 387.2 μm, n = 30); main bodies globose, 160–315 μm diam ( = 227.4 μm, n = 30), composed of subglobose, hyaline to yellow, 11.5–15 × 7.5–11.5 μm cells; hyphal appendages 63 to 138, unbranched, flexuous, cylindrical, 1–4-septate, 67–133 × 3–5 μm ( = 97.1 × 3.7 μm, n = 52).

Culture characteristics: Colonies on PDA attaining 27–37 mm diam within 21 d, velvety, plane, buff to olivaceous black at centre; reverse buff to olivaceous black at centre (Fig. 8D); spermatial and asexual morphs formed.

Specimens examined: Japan, Aomori, Nakatsugaru, Onikawabe, on living leaves of Fagus crenata, 12 Aug. 2012, K. Tanaka et al., KT 3058 (HHUF 30515, culture CBS 142917 = MAFF 245738); Nishimeya, Ookawazoe, near Annmon waterfall trail, on living leaves of Fagus crenata, 2 Sep. 2012, K. Tanaka et al., KT 3074-3 (HHUF 30516, culture CBS 142918 = MAFF 245739); 2 Sep. 2012, R. Fujimoto et al., RF 5 (HHUF 30517, culture CBS 142919 = MAFF 245741); 10 Sep. 2016, A. Hashimoto, AH 561 (HHUF 30553, culture CBS 142920); Hirakawa, Ikarigaseki, on living leaves of Fagus crenata, 28 Apr. 1995, C. Z. Wei & Y. Harada, H 2579 (HHUF 22903, holotype of Pseudodidymella fagi, ex-holotype living culture MAFF 245740); artificial inoculation on leaves of Fagus crenata, 30 Sep. 1996, C. Z. Wei (HHUF 23672, holotype of Pycnopleiospora fagi).

Notes: This species was originally reported to cause brown leaf spots on Fagus crenata in Japan. More recently, it was reported from a new host, F. sylvatica (Gross ). To elucidate its host spectrum, further surveys for this fungus and other species on Fagus is needed.

A. Hashim. & Kaz. Tanaka, sp. nov. MycoBank MB819615. Fig. 6.

Fig. 6

Pseudodidymella minima.A–C. Leaves of Fagus japonica with necrotic brown spots. D–F. Propagules on the leaf surface. G, H. Propagules. I. Appendages of propagule. A–C, H from HHUF 30552. D, E, G, I from HHUF 30551. F from culture CBS 142921 = MAFF 246249. Scale bars: D–F = 250 μm; G, H = 50 μm; I = 5 μm.

Etymology: Referring to the smaller-sized propagules observed in this species.

Parasitic on living leaves of Fagus japonica. Sexual morph: Unknown. Asexual morph: Propagules epiphyllous, white to yellowish, globose, 110–220(–240) μm diam ( = 164.4 μm, n = 60); main bodies globose, multicellular, 78–168 μm diam ( = 115 μm, n = 60), composed of subglobose, 7.5–10 μm diam, hyaline to yellow cells; hyphal appendages 65 to 135, unbranched, flexuous, cylindrical, 1–2-septate or rarely aseptate, 27–44 × 3–6 μm ( = 35.5 × 4.4 μm, n = 59).

Culture characteristics: Colonies on PDA attaining 32–38 mm diam within 21 d, floccose, plane, smoke grey; reverse honey to isabelline (Fig. 8E); asexual morph formed.

Specimens examined: Japan, Iwate, Hanamaki, near Dai spa, on living leaves of Fagus japonica, 9 Oct. 2011, K. Tanaka, KT 2918 (HHUF 30551 holotype designated here; ex-holotype living culture CBS 142921 = MAFF 246249); 3 Sept. 2016, A. Hashimoto, AH 556 (HHUF 30552 paratype, ex-paratype living culture CBS 142922).

Notes: This species on Fagus japonica is easily distinguished from Pseudod. fagi on F. crenata by its much smaller propagules. Sequence differences between these two species were found at six nucleotide positions with one gap in the ITS sequences.

We did not observe the sexual or synasexual morph of Pseudod. minima. Further surveys are therefore needed to reveal the ecological features of this species.

(Dearn. & Barthol.) A. Hashim. & Kaz. Tanaka, comb. nov. MycoBank MB821403.

Basionym: Cercosporella aceris Dearn. & Barthol., Mycologia 9: 362. 1917.

Synonyms: Mycopappus aceris (Dearn. & Barthol.) Redhead & G.P. White, Canad. J. Bot. 63: 1430. 1985.

Petrakia aceris (Dearn. & Barthol.) Jaklitsch & Voglmayr, Sydowia 69: 90. 2017.

Stigmina zilleri A. Funk, Canad. J. Bot. 65: 482. 1987.

Xenostigmina zilleri (A. Funk) Crous, Mycol. Mem. 21: 155. 1998.

Mycosphaerella mycopappi A. Funk & Dorworth, Canad. J. Bot. 66: 295. 1988.

Didymella mycopappi (A. Funk & Dorworth) Crous, Mycol. Mem. 21: 152. 1998.

Notes: Xenostigmina zilleri is the name that has been commonly used for this pathogen, although the epithet of Cercosporella aceris is older than that of Stigmina zilleri (Crous, 1998, Crous et al., 2009, Phookamsak et al., 2014, Tian et al., 2015, Gross et al., 2017). Therefore, we proposed a new combination, Xenostigmina aceris.

Incertae sedis

Jaklitsch & Voglmayr, Sydowia 69: 84. 2017.

Saprobic on dead twigs of woody plants. Sexual morph: Ascomata globose to ovoid, immersed to superficial, gregarious, sometimes confluent, ostiolate. Peridium composed of elongate, thin-walled, brown cells, at base composed of elongate, hyaline cells. Pseudoparaphyses septate, branched and anastomosed. Asci bitunicate, cylindrical, 8-spored. Ascospores fusiform, multi-septate, smooth. Asexual morph: Conidiomata pseudopycnidial, globose to cylindrical, sometimes deformed, septate, confluent, multiloculate, scattered, semi-immersed, black, with one to two non-papillate ostiole. Peridium rectangular, brown cells. Conidiophores absent. Conidiogenous cells holoblastic, cylindrical, hyaline, smooth. Conidia cylindrical with rounded ends, hyaline, smooth, aseptate.

Type species: Alpinaria rhododendri (Niessl) Jaklitsch & Voglmayr.

(Niessl) Jaklitsch & Voglmayr, Sydowia 69: 84. 2017. Fig. 7.

Fig. 7

Alpinaria rhododendri.A, B. Ascomata on substrate. C. Ascoma in longitudinal section. D. Lateral peridium of ascoma. E. Ascus. F. Apex of ascus. G. Stipe of ascus. H. Pseudoparaphyses. I–K. Ascospores. L, M. Conidiomata in culture. N. Conidiomata in longitudinal section. O. Peridium of conidioma. P, Q. Conidiogenous cells. R. Conidia. S. Germinating conidia. A–K from HHUF 30554. L–S from culture CBS 142901. Scale bars: A, L = 500 μm; B, C, M, N = 100 μm; D, E, O = 10 μm; F–K, P–S = 5 μm.

Basionym: Cucurbitaria rhododendri Niessl, Verh. Nat. Ver. Brünn 10: 200. 1872.

Synonyms: Gibberidea rhododendri (Niessl) Petr., Ann. Mycol. 32: 330. 1934; nom. illegit.

Melanomma rhododendri Rehm, Ber. Naturhist. Ver. Augsburg 26: 48. 1881.

Gibberidea rhododendri (Rehm) Petr., Krypt. Forsch. (München) 2: 160. 1931.

Gibberidea rhododendri (Rehm) Kirschst., Hedwigia 81: 206, 1944; nom. illegit.

Saprobic on dead twigs of ericaceous plants. Sexual morph: Ascomata globose to ovoid, immersed, becoming largely erumpent to superficial, gregarious, sometimes confluent, 140–190 μm high, 110–250 μm diam. Ostiolar neck short papillate, composed of carbonaceous, thick-walled, black cells. Peridium 55–75 μm thick at side composed of elongate, thin-walled, 12–13 × 5–6.5 μm, brown cells, 87–102 μm thick at base composed of elongate, thin-walled, 4–6 μm diam, hyaline cells. Pseudoparaphyses trabeculate, 1–1.5 μm wide, septate, branched and anastomosed. Asci bitunicate, cylindrical, 100–118 × 7–9 μm ( = 109.5 × 7.8 μm, n = 11), with a short stipe (3.5–10 μm long,  = 7 μm, n = 11). Ascospores fusiform, 13–21 × 5–6 μm ( = 16.5 × 5.6 μm, n = 50), l/w 2.2–4.2 ( = 3.0, n = 50), 3-septate, with a primary septum nearly median (0.42–0.57,  = 0.50, n = 50) and constricted, smooth, without sheath. Asexual morph: Conidiomata pseudopycnidial, globose to cylindrical, sometimes deformed, septate, confluent, multiloculate, scattered, semi-immersed, black, up to 190 μm high, 110–250 μm diam. Ostiolar neck mainly single, occasionally two, non-papillate. Peridium 20–25 μm wide, composed of 7.5–11.5 × 5–7 μm, rectangular, brown cells. Conidiophores reduced to conidiogenous cells. Conidiogenous cells holoblastic, 6–10.5 × 3–4.5 μm, cylindrical, hyaline, smooth. Conidia cylindrical with rounded ends, 2–4 × 1–2 μm ( = 3 × 1.6 μm, n = 50), l/w 1.1–2.6 ( = 1.9, n = 50), hyaline, smooth, aseptate, guttulate when young.

Culture characteristics: Colonies on PDA attaining 26–31 mm diam within 21 d, velvety, wet, olivaceous black, smoke grey at margin; reverse olivaceous black at centre (Fig. 8F); asexual morph formed.

Specimen examined: Japan, Iwate, Hachimantai, Yakeyama, near Goshogake spa, on leaf bud of Rhododendron brachycarpum, 9 Jul. 2008, Y. Harada, KT 2520 (HHUF 30554; culture CBS 142901).

Notes: The ascospore size in the material mentioned above is identical to that of A. rhododendri reported by Jaklitsch & Voglmayr (2017), who designated the epitype of this species. The ITS, tef1 and rpb2 sequences from our material are completely identical to those from the ex-epitype strain of this species (CBS 141994). This species has been reported from twigs or buds of Rhododendron spp. in the Asia (R. chrysanthum; Müller 1959), Europe (R. ferrugineum and R. hirsutum; Jaklitsch & Voglmayr 2017), and North America (Rhododendron sp.; Mugambi & Huhndorf 2009). In addition, we collected this species on R. brachycarpum from the subalpine zone in Japan. Alpinaria rhododendri appears to be a relatively common species in the subalpine to alpine zone worldwide.

Alpinaria was recently established to accommodate a single species A. rhododendri, which was transferred from Melanomma because this species is phylogenetically distinct from the type species of Melanomma, and possesses ascomata with a roughened surface view of textura prismatica and textura angularis (Jaklitsch & Voglmayr 2017). Furthermore, they treated the genus as a member of Melanommataceae (Jaklitsch & Voglmayr 2017). Although no asexual morph was reported for this species (Müller, 1959, Mugambi and Huhndorf, 2009, Jaklitsch and Voglmayr, 2017), we newly observed its asexual morph in culture (Fig. 7L–S). As a result of our observation of the asexual morph, as well as the sexual morph, we clarified that this species has atypical features for Melanommataceae; its ascomata are composed of hyaline cells at the base, and are pseudopycnidial. Confluent conidiomata are not found in sexual/asexual morphs of Melanommataceae. In our phylogenetic tree, the genus placement is confirmed outside Melanommataceae sensu stricto (Fig. 1). Therefore, we treat Alpinaria as incertae sedis in Pleosporales in this study; additional taxa related to this genus will be needed to resolve its familial placement.

Discussion

Re-circumscription of Melanommataceae sensu stricto

Melanommataceae has been extensively studied in recent years based on phylogenetic evidence (Mugambi and Huhndorf, 2009, Schoch et al., 2009, Wijayawardene et al., 2012, Wijayawardene et al., 2014, Butin et al., 2013, De Gruyter et al., 2013, Su et al., 2015, Tian et al., 2015, Li et al., 2016, Gross et al., 2017, Jaklitsch and Voglmayr, 2017). The characters emphasised for members of this family include a carbonaceous peridium of ascomata and trabecular pseudoparaphyses. These species are known saprobes on decaying plant material, or, rarely, as plant pathogens. The familial concept of Melanommataceae was revised and expanded after in a study by Mugambi & Huhndorf (2009), who applied a molecular approach. A recent monograph of Melanommataceae was based on morphological and multi-gene phylogenetic data (Tian ). Although monophyly of Melanommataceae was confirmed in previous studies, statistical support for Melanommataceae sensu lato was lacking (Mugambi and Huhndorf, 2009, Schoch et al., 2009, Tian et al., 2015). Additionally, previous authors did not examine the asexual morphs, although various asexual morphs, such as those with mononematous, synnematous, and pycnidial conidiomata, are known to occur in this family. Two of the most striking genera are Petrakia and Xenostigmina, which have mycopappus-like propagules as asexual morphs, and were reported to be foliicolous necrotrophs (Funk, 1986, Funk and Dorworth, 1988, Crous, 1998, Crous et al., 2009, Butin et al., 2013), whereas species of Melanomma, the type genus of this family, have aposphaeria-like pycnidial asexual morphs and are known to be saprobes on twigs of various plant hosts (Chesters, 1938, Romero, 1998, Zhang et al., 2008). Our multi-gene phylogenetic analyses of this family clearly showed the poly- and paraphyletic nature of Melanommataceae sensu lato (Fig. 1), and morphological observations of sexual and asexual morphs led to the conclusion that Melanommataceae should be restricted to the type genus Melanomma. In addition, four genera with mycopappus-like propagules in their asexual morphs (Mycodidymella, Petrakia, Pseudodidymella, and Xenostigmina) are separated from Melanommataceae sensu stricto, and we thus establish a new family, Pseudodidymellaceae, to accommodate these genera.

Relationships among genera in Pseudodidymellaceae

Mycodidymella and Xenostigmina are retained as natural genera in the present study. Butin found that the sexual morph of Mycodidymella is similar to that of Petrakia, and thus recognised Petrakia in a broad sense and included Mycodidymella as a synonym. This treatment was supported by a later study (Li ). Gross showed these three genera are closely related based on an ITS phylogeny, but no taxonomic conclusions about their generic validities were made. Recently, Jaklitsch & Voglmayr (2017) proposed that Mycodidymella and Xenostigmina are synonyms of Petrakia. They considered that phylogenetic relatedness of Xenostigmina and Petrakia, and morphological similarity of the sexual morph and mycopappus-like propagules among these genera are strong arguments for synonymising them (Jaklitsch & Voglmayr 2017). Our phylogenetic analysis including Mycodidymella as well as Xenostigmina and Petrakia clarified that their monophyletic status was not well supported in any analyses (below 60 % ML BP/ 0.95 Bayesian PP, Fig. 1). Their sexual morphs are superficially similar as indicated by Jaklitsch & Voglmayr (2017), but Mycodidymella has deeper and more well-developed ascomata (up to 210 μm high) than those of Petrakia (up to 150 μm high) and Xenostigmina (up to 100 μm high). Additionally, their morphological characters of their synasexual morphs are also different; hyaline, up to 20 μm thick sporodochia, holoblastic conidiogenous cells, and sigmoid, multi-septate, thin-walled, hyaline conidia (Mycodidymella; this study); brown, up to 30 μm thick sporodochia, annellidic conidiogenous cells, and globose to ovoid, dictyosporus, thick-walled, brown conidia with cellular appendages (Petrakia; Butin et al., 2013, Li et al., 2016); and brown to black, up to 45 μm high sporodochia, holoblastic conidiogenous cells, and clavate with a short rostrum, dictyosporus, thick-walled, brown conidia (Xenostigmina; Funk, 1986, Crous, 1998). Therefore, we treat these genera as distinct based on morphological differences of sexual and synasexual morphs.

Synasexual morphs of these three genera are produced after leaves fall in late autumn (Funk and Dorworth, 1988, Wei et al., 1997, Butin et al., 2013, Gross et al., 2017). Conidia of synasexual morphs were not observed on overwintered leaves for Petrakia and Mycodidymella, and their function in the disease cycle during the winter season has not been clarified (Wei et al., 1997, Butin et al., 2013). No synasexual morph is known from Pseudodidymella, despite their close relationship to the other three genera. Further studies on the Pseudodidymella synasexual morph are needed to elucidate the whole life cycle of this genus and produce robust taxonomic classifications for Pseudodidymellaceae.

Form and function of mycopappus-like propagules

The genus Mycopappus was established based on its type species Mycop. alni (on Alnus, Betula, Crataegus, and Pyrus; Redhead and White, 1985, Braun et al., 2000, Takahashi et al., 2006), which produces epiphyllous, multicellular propagules in its asexual morph (Redhead & White 1985). Later, three species were assigned to in this genus: Mycop. aceris (on Acer macrophyllum; Redhead & White 1985), Mycop. aesculi (on Aesculus turbinata; Wei ), and Mycop. quercus (on Quercus acutissima; Suto & Kawai 2000). Two species, Mycop. alni and Mycop. quercus, produce microconidia and sclerotia in culture (Redhead and White, 1985, Suto and Kawai, 2000), and the sexual morph of the latter species is characterised by stipitate apothecia and inoperculate asci (Suto & Suyama 2005). Mycopappus alni was suggested to be a member of Sclerotiniaceae (Helotiales, Leotiomycetes) based on its sclerotial morph and phylogenetic analyses using ITS sequences (Takahashi ). The two other species, Mycop. aceris and Mycop. aesculi, were excluded from Mycopappus sensu stricto, because their sexual morphs belong to the dothideomycetous taxa, namely Xenostigmina aceris (Funk and Dorworth, 1988, Crous, 1998, Crous et al., 2009) and Mycodidymella aesculi (Wei ), respectively. Morphological differences in mycopappus-like propagules among these lineages were indicated in a previous study (Suto & Kawai 2000). The main bodies of sclerotiniaceous species (Mycop. alni and Mycop. quercus) are composed of multi-septate claviform cells (Suto and Kawai, 2000, Suto and Suyama, 2005, Takahashi et al., 2006), whereas those of dothideomycetous species (Mycod. aesculi and X. aceris) are composed of aseptate globose cells (Redhead and White, 1985, Wei et al., 1998). The morphological resemblance of mycopappus-like propagules between leotiomycetous and dothideomycetous lineages appears to be the result of convergent evolution due to similar ecological function, such as rain-splash dispersal across the leaf surface. A similar situation was reported in two phylogenetically distinct genera, Spiroplana (Dothideomycetes) and Spirosphaera (Leotiomycetes), which have spirally coiled, buoyant conidia that resulted in adaptation to water dispersal in terrestrial or aero-aquatic environments (Voglmayr ).

The mycopappus-like propagules of Pseudodidymellaceae may contribute to secondary infection of host leaves with high inoculum potential. Wei suggested that this morph plays an important role in disease development. Morphological variation of the propagules at the generic level was observed, but the taxonomic significance was not been examined in several studies (Redhead and White, 1985, Wei et al., 1998, Butin et al., 2013, Gross et al., 2017, Jaklitsch and Voglmayr, 2017). Our observations revealed that morphological features of propagules differed between Mycodidymella, Petrakia, and Xenostigmina (with few appendages), and Pseudodidymella (with numerous appendages). The hyphal appendages of Pseudodidymella could enhance fungal encounters with Fagus leaves that have conspicuous wax ornamentation (Denk 2003), as is the case of asexual fungi with conidial appendages (Nag Raj, 1993, Hashimoto et al., 2015a). The morphological variation of propagules is also observed at the species level: Pseudod. fagi on F. crenata has a larger main body with longer appendages (Fig. 5W and X), and Pseudod. minima on F. japonica has a smaller main body with shorter appendages (Fig. 6G and H). These morphological variations of their propagules may be correlated with presence (in F. japonica) or absence (in F. crenata) of leaf papillae (Denk 2003) as a result of adaptation to the host surface.

A phoma-like morph is known in the life cycle in Petrakia (Butin ). This morph is also observed in Mycodidymella and Pseudodidymella after fructification of mycopappus-like propagules (Fig. 4, Fig. 5J–R). The conidial-like structures of this morph appear to be spermatia, because they do not germinate in water agar or glucose agar.

Speciation through host switching and host jumping

Plant pathogens frequently infect phylogenetically related hosts (Jackson, 2004, Giraud et al., 2008, Walker et al., 2010, Walker et al., 2012, Mejía et al., 2011). The genus Pseudodidymella was originally established as a monotypic genus composed of the type species Pseudod. fagi, which was reported to be a pathogen of F. crenata (Fagaceae, Fagales) in Japan (Wei ). Most recently, this species was re-discovered and reported to be a disease agent of F. sylvatica in Germany and Switzerland (Gross ). A new species of this genus, Pseudod. minima, occurs on F. japonica. Members of Pseudodidymella appear to be host-specific on Fagus. Close host/fungus associations and coevolution were reported in members of Gnomoniaceae, Phaeosphaeriaceae, and Sclerotiniaceae (Jackson, 2004, Walker et al., 2012, Ertz et al., 2015). Although ITS sequences of Pseudod. fagi were 100 % identical among isolates from F. crenata and F. sylvatica (Gross ), those of Pseudod. minima differed from Pseudod. fagi based on six nucleotide positions and one gap in ITS sequences (this study). This result was compatible with host phylogeny: F. crenata and F. sylvatica are closely related to each other, but F. japonica is phylogenetically distantly related to the other species (Denk ).

Alternatively, three genera, Mycodidymella, Petrakia, and Xenostigmina, are host-specific for Acer spp. or Aesculus (Sapindaceae, Sapindales), which are distantly related to Fagales (APG IV 2016). It has been recognised that several plant pathogens switch to unrelated host plants (Reddy et al., 1998, Takamatsu et al., 2000, Jackson, 2004). Gross also found that host switching occurred in members of Pseudodidymellaceae, and members of this family evolutionally diversified by host switching. Similar evolutionary processes that led to speciation through host jumping are known from Clavicipitaceae, which includes plant pathogens, insect pathogens, and mycoparasites (Kepler ).

Future studies

The asexual genus Seifertia on Rhododendron spp. is characterised by synnematous conidiomata with cladosporium-like conidia (Li ). Phylogenetic relatedness of this genus to members of Pseudodidymellaceae was suggested (Li et al., 2016, Gross et al., 2017). However, we prefer to not include this species in Pseudodidymellaceae and place it incertae sedis, because of the lack of mycopappus-like propagules in the life cycle. This genus might represent a new family; however, analysis of its sexual morph and further taxa related to this genus are needed to determine its familial placement. Another genus, Alpinaria, was originally established to accommodate the type species, A. rhododendri, which was segregated from Melanomma (Jaklitsch & Voglmayr 2017). They regarded the genus as a member of Melanommataceae, based on phylogenetic analyses (Jaklitsch & Voglmayr 2017). In the present study, we newly observed the asexual morph of Alpinaria, which had not been reported in previous studies (Müller, 1959, Holm, 1968, Mugambi and Huhndorf, 2009, Jaklitsch and Voglmayr, 2017). According to our phylogenetic analyses and morphological observations, this species is distantly related to Melanommataceae sensu stricto (Fig. 1) and has atypical features for Melanommataceae, such as hyaline cells at the base of ascomata and pseudopycnidial conidiomata. Several melanomma-like fungi that possess well-developed carbonaceous ascomata may have evolved several times within Pleosporales, such as in Cyclothyriellaceae, Ohleriaceae, Nigrogranaceae, Teichosporaceae, Thyridariaceae (Jaklitsch and Voglmayr, 2016, Jaklitsch et al., 2016b). It seems that familial circumscriptions based merely on sexual morph characters is insufficient to distinguish the members of Melanommataceae sensu lato.

The present study revealed unexpected diversity of Melanommataceae sensu Tian . Our approaches, which combined morphological features of both sexual and asexual morphs with molecular phylogenetic analyses, enabled a re-circumscription of Melanommataceae sensu stricto and the establishment of Pseudodidymellaceae. To build a comprehensive taxonomic framework, further discovery of more specimens along with additional morphological and molecular data would help elucidate other unresolved lineages of Melanommataceae sensu lato.