![Coleodictyospora spp. (a,b from MFLU 18-1544, holotype. c-e redrawn from Matsushima [31] and Seifert et al. [42]). (a,b) C. muriformis (conidia with branched conidiophores); (c) C. micronesica (conidia growing on a short subulate conidiogenous cell which directly ascends from the basal mat of sporodochia); (d,e) C. cubensis (d sporodochia bearing a mass of conidia. e conidia with a long conidiogenous cell). Scale bars, (a-e) 30 μm. Index Fungorum number: IF558195; Facesoffungi number: FoF 09872 Etymology: in reference to the muriform conidia of the fungus Holotype: MFLU 18-1544](https://www.researchgate.net/profile/Wei-Dong-40/publication/354235901/figure/fig4/AS:1062713035878400@1630381940882/Coleodictyospora-spp-a-b-from-MFLU-18-1544-holotype-c-e-redrawn-from-Matsushima-31_Q320.jpg)
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Fungi
Journal of
Article
Five Novel Taxa from Freshwater Habitats and New Taxonomic
Insights of Pleurotheciales and Savoryellomycetidae
Wei Dong 1,2,3,4 , Rajesh Jeewon 5, Kevin D. Hyde 1,2, Er-Fu Yang 6, Huang Zhang 7, Xiandong Yu 8,
Gennuo Wang 9, Nakarin Suwannarach 3,4,* , Mingkwan Doilom 1,2 and Zhangyong Dong 1
Citation: Dong, W.; Jeewon, R.;
Hyde, K.D.; Yang, E.-F.; Zhang, H.;
Yu, X.; Wang, G.; Suwannarach, N.;
Doilom, M.; Dong, Z. Five Novel Taxa
from Freshwater Habitats and New
Taxonomic Insights of Pleurotheciales
and Savoryellomycetidae. J. Fungi
2021,7, 711. https://doi.org/
10.3390/jof7090711
Academic Editors: Samantha
C. Karunarathna and
Saowaluck Tibpromma
Received: 5 July 2021
Accepted: 4 August 2021
Published: 30 August 2021
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
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iations.
Copyright: © 2021 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
1Innovative Institute for Plant Health, Zhongkai University of Agriculture and Engineering,
Guangzhou 510225, China; dongwei0312@hotmail.com (W.D.); kdhyde3@gmail.com (K.D.H.);
j_hammochi@hotmail.com (M.D.); dongzhangyong@hotmail.com (Z.D.)
2Center of Excellence for Fungal Research, Mae Fah Luang University, Chiang Rai 57100, Thailand
3
Research Center of Microbial Diversity and Sustainable Utilization, Faculty of Science, Chiang Mai University,
Chiang Mai 50200, Thailand
4Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand
5Department of Health Sciences, Faculty of Medicine and Health Sciences, University of Mauritius,
Reduit 80837, Mauritius; r.jeewon@uom.ac.mu
6Research Center in Bioresources for Agriculture, Industry and Medicine, Chiang Mai University,
Chiang Mai 50200, Thailand; erfu20170431@gmail.com
7Faculty of Agriculture and Food, Kunming University of Science & Technology, Kunming 650500, China;
zhanghuang2002113@gmail.com
8School of Life Science and Technology, University of Electronic Science and Technology of China,
Chengdu 611731, China; yuxiandong@std.uestc.edu.cn
9Faculty of Environmental Science and Engineering, Kunming University of Science & Technology,
Kunming 650500, China; gennuo2021@gmail.com
*Correspondence: suwan.462@gmail.com
Abstract:
Pleurotheciales is the largest order in Savoryellomycetidae with a large proportion of
species known from freshwater habitats. In order to investigate the phylogenetic relationships of
taxa within Pleurotheciales and contribute to their diversity, submerged wood was collected from
freshwater habitats in China (Yunnan Province) and Thailand. Two dematiaceous, sporodochial
hyphomycetes and one annulatascales-like ascomycete with unusual morphology as compared
to extant ones were discovered. They were subjected to DNA-based phylogenetic analyses and
the results revealed three distinct lineages in Savoryellomycetidae. This morpho-phylo taxonomic
study supports the establishment of five novel taxa including two novel genera, Obliquifusoideum
and Saprodesmium, and three novel species, Coleodictyospora muriformis,Obliquifusoideum guttulatum
and Saprodesmium dematiosporum.Coleodictyospora muriformis and S. dematiosporum are placed in
Pleurotheciales, while O. guttulatum is referred to Savoryellomycetidae genera incertae sedis. The
phylogenetic relationships are also presented for Coleodictyospora and Pseudocoleodictyospora, which
raises an intriguing taxonomic issue. These two genera are positioned in two different classes, viz
Sordariomycetes and Dothideomycetes, although they are quite similar except for the presence of a
conidial sheath. This study expands our knowledge of the fungal diversity of freshwater fungi, and
also indicates that Pleurotheciales species are mostly found in freshwater habitats.
Keywords:
annulatascales-like; multi-locus phylogeny; Pseudocoleodictyospora; Sordariomycetes;
submerged wood; Thailand; Yunnan
1. Introduction
Savoryellomycetidae currently accommodates four orders, Conioscyphales, Fus-
cosporellales, Pleurotheciales and Savoryellales. This is based on evidence from phy-
logenetic analyses and divergence time studies with the order having a stem age estimated
as 268 MYA [
1
]. The four orders clustered as a robust clade in all studies [
1
–
3
]. Pleurothe-
ciales, with a single-family Pleurotheciaceae [
4
], is the largest order in Savoryellomycetidae.
J. Fungi 2021,7, 711. https://doi.org/10.3390/jof7090711 https://www.mdpi.com/journal/jof
J. Fungi 2021,7, 711 2 of 24
Pleurotheciaceae species have mostly been isolated from decaying wood or plant debris
as saprobes, while few species were also identified as opportunistic human pathogens
(Phaeoisaria clematidis) [
5
]. Multi-locus phylogenetic relationships of Pleurotheciaceae
species were investigated to better understand their taxonomy [
3
,
4
,
6
–
10
] and as a result,
eleven genera were accepted in the family [11].
Taxa of Pleurotheciaceae have perithecial ascomata with asexual dematiaceous hy-
phomycetous stages. Coelomyceteous asexual morphs have not been reported in the
family. Pleurotheciaceae is an assemblage of genera representing a highly diverse mor-
phology, especially in the asexual morphs. Pleurotheciella and Pleurothecium (type) gen-
erally have macronematous, unbranched conidiophores, holoblastic, sympodially prolif-
erating conidiogenous cells with a conspicuous rachis of denticles, and hyaline, septate
conidia [
4
,
9
,
10
,
12
,
13
]. Anapleurothecium has macronematous, unbranched conidiophores,
sympodial, denticulate conidiogenous cells which are the cases of Pleurotheciella and
Pleurothecium, but it has botuliform to cylindrical and brown conidia with a paler basal
cell [
7
]. Phaeoisaria has synnematous conidiophores with tiny aseptate conidia [
13
]. Phragmo-
cephala also has synnematous conidiophores, but the conidia are relatively large, ellipsoidal
to subglobose, with dark brown to black central cells and paler polar cells [
14
]. Sterigma-
tobotrys are distinct in the family by their well-defined stipe and a complex penicillate
conidiophore head consisting of series of penicillate branches [
6
]. While some other genera
lack conspicuous macronematous conidiophores, and the conidia directly arise from the
hyphae on the host substrate or from micronematous, subhyaline conidiophores, such as
Neomonodictys with subglobose to globose, muriform conidia [
8
] and Helicoascotaiwania
with helicoid conidia [2,15].
The diversity of morphology is also reflected in some sexual morphs of Pleurotheci-
aceae. The genera Adelosphaeria,Melanotrigonum,Pleurotheciella and Pleurothecium generally
have superficial ascomata with a short papilla, narrowly or broadly clavate asci with a
distinct, refractive apical ring and ellipsoidal-fusiform, septate ascospores [
3
,
4
,
9
], while
Phaeoisaria has immersed ascomata with a quite long neck, cylindrical asci and filiform,
multiseptate ascospores. Helicoascotaiwania is easily distinguished in the family in having
generally immersed ascomata lying horizontally or obliquely to the host substrate and
fusiform, versicolorous ascospores with darker central cells and paler polar cells [2,3].
The asexual–sexual morph connections were investigated based on cultural studies
with a combination of molecular data. Some hyphomycetes were linked as the life cycle of
known sexual morphs. The asexual morph of Pleurothecium recurvatum was first reported
from the artificial medium (WA) inoculated by an ascomycetous species Carpoligna pleu-
rothecii [
16
]. Réblováet al. [
9
] also found the asexual morphs from another medium (PCA)
inoculated by ascomycetous species Pleurotheciella rivularia and Pleurothecium semifecundum.
With DNA sequence data, Luo et al. [
10
] linked the asexual–sexual morph of Pleurotheciella
fusiformis based on two specimens collected from Erhai Lake, Yunnan, China.
Luo et al. [
6
] used multi-locus analysis to first report a sexual morph in Phaeoisaria,
namely Ph. filiformis, which was characterized by immersed ascomata with a long, cylin-
drical neck, and cylindrical asci containing hyaline and filiform ascospores. Although the
morphological traits associate Ph. filiformis as allied with Ceratosphaeria and Ophioceras in
Magnaporthales, phylogenetic analysis placed Ph. filiformis in Pleurotheciales. Neither
conidia nor conidiomatal structures were observed from the culture of Ph. filiformis [6].
The main objectives of this study were to revise the taxon diversity within Pleurothe-
ciales, report on novel taxa and provide new insights into the systematics of Savoryel-
lomycetidae. Two specimens of dematiaceous hyphomycetes were preliminarily identified
as Pleurotheciales species with micronematous conidiophores, holoblastic conidiogenous
cells and dark muriform conidia, but the morphologies were rather unusual as compared
to other members of the family. One specimen resembled annulatascaceae-like taxa in Dia-
porthomycetidae but is similar to the taxa of Pleurotheciaceae in Savoryellomycetidae. In
order to clarify the placement of these specimens, a multi-locus analysis of a concatenated
nuc 28S rDNA (LSU), nuc18S rDNA (SSU), internal transcribed spacer (ITS) and second-
J. Fungi 2021,7, 711 3 of 24
largest subunit of RNA polymerase II (rpb2) dataset were performed, and phylogenetic
relationships inferred.
2. Materials and Methods
2.1. Herbarium Material, Isolation and Morphology
Decayed woody twigs and branches submerged in freshwater streams in forests were
randomly collected in Yunnan Province, China, as well as Satun and Songkhla provinces in
Thailand where all places are in the Greater Mekong Subregion. Specimens were placed in
zip-lock plastic bags containing moistened cotton and taken to the laboratory. Sediment
on separated specimens was washed off with tap water and incubated in plastic boxes
lined with moistened tissue paper at room temperature (20–25
◦
C) for 1–2 weeks. The
ascomata and sporodochia developed on the specimens were examined with a Nikon
SMZ-171 dissecting microscope. Fungal structures were captured with a Nikon ECLIPSE
Ni compound microscope fitted with a Canon EOS 750D digital camera. Single spore
isolations were made from ascospore or conidium on potato dextrose agar (PDA, Shanghai
Bio-way technology Co., Ltd., Shanghai, China) at room temperature (20–25
◦
C). All
morphological approaches used herein were modified from Chomnunti et al. [
17
] and
Senanayake et al. [
18
]. Tarosoft (R) Image Frame Work program was used to measure the
fungal structures. Images were processed with Adobe Photoshop CS5 software (Adobe
Systems, San Jose, CA, USA). Herbarium specimens (dry wood with fungal materials) were
deposited in the herbarium of Mae Fah Luang University (MFLU), Chiang Rai, Thailand
and herbarium of Cryptogams, Kunming Institute of Botany Academia Sinica (HKAS),
Kunming, China. Living cultures were deposited in the Mae Fah Luang University Culture
Collection (MFLUCC) and Kunming Institute of Botany Culture Collection (KUMCC). The
novel taxa were registered in the databases Facesoffungi (http://www.facesoffungi.org,
accessed on 10 June 2021) [
19
] and Index Fungorum (http://www.indexfungorum.org/
names/names.asp, accessed on 21 June 2021).
2.2. DNA Extraction, PCR Amplification and Sequencing
Fungal mycelia were scraped from the colonies on PDA. The Biospin Fungus Genomic
DNA Extraction Kit (Bioer Technology Co., Hangzhou, China) was used to extract total
genomic DNA. The polymerase chain reaction (PCR) technique was utilized for the ampli-
fication of target DNA fragments. The primer pairs LR0R/LR5, NS1/NS4, ITS5/ITS4 and
fRPB2-5F/fRPB2-7cR were used to amplify LSU, SSU, ITS and rpb2 [
20
–
22
]. The amplifica-
tions were carried out as detailed in Dong et al. [
23
]. The PCR thermal cycle program for
the amplification of LSU, SSU and ITS was provided as initially 94 ◦C for 3 min, followed
by 35 cycles of denaturation at 94
◦
C for 30 s, annealing at 55
◦
C for 50 s, elongation at
72
◦
C for 90 s and a final extension at 72
◦
C for 10 min. The annealing was adjusted to
52
◦
C for rpb2. PCR products were checked on 1% agarose electrophoresis gels stained
with Gel Red. The sequencing reactions were carried out by Shanghai Sangon Biological
Engineering Technology and Services Co., Shanghai, China.
2.3. Molecular Phylogenetic Analyses
2.3.1. Sequence Selection and Phylogenetic Analyses Construction
The qualities of generated sequences were initially checked with Finch TV v. 1.4.0 and
each gene was subjected to a BlastN search in NCBI’s GenBank to assess the confidence level.
Phylogenetic placements of the unidentified fungi were resolved by analyzing four different
datasets: (1) two multi-locus datasets of a concatenated LSU, SSU, ITS and rpb2 sequences;
(2) two separate individual LSU and ITS datasets. The first multi-locus dataset was analysed
to infer the phylogenetic positions of unidentified fungi within the Savoryellomycetidae
along with sequences deposited from recent relevant publications [
3
,
6
,
24
]. In the second
multi-locus dataset, we included the other subclasses in Sordariomycetes to infer the
subclass status of an unidentified genus which could not be confirmed in any orders
in the subclass Savoryellomycetidae. Besides, Pseudocoleodictyospora and its related taxa
J. Fungi 2021,7, 711 4 of 24
were also included to show the phylogenetic relationships between Coleodictyospora and
Pseudocoleodictyospora. The individual LSU and ITS phylogenetic analyses were utilized
to auxiliarily assess the phylogenetic relationships of fungi in Savoryellomycetidae. All
sequences used in this study were listed in Table 1.
Table 1. Taxa used in the phylogenetic analyses and their corresponding GenBank accession numbers.
Taxon Voucher/Culture GenBank Accession Numbers
LSU SSU ITS rpb2
Achroceratosphaeria potamia CBS 125414 GQ996538 GQ996541 MH863679 KM588908
Adelosphaeria catenata CBS 138679TKT278707 KT278692 KT278721 KT278743
Anapleurothecium botulisporum FMR 11490TKY853483 - KY853423 -
Arecophila bambusae HKUCC 4794 AF452038 AY083802 - -
Ascotaiwania latericolla ICMP 22739TMN699407 - MN699390 MN704312
Ascotaiwania lignicola NIL 00005 HQ446364 HQ446284 - HQ446419
Bactrodesmiastrum obovatum FMR 6482 FR870266 - - -
Bactrodesmiastrum pyriforme FMR 11931 HE646637 - - -
Bactrodesmium diversum CBS 144080 MN699415 MN699371 MN699355 MN704294
Bactrodesmium obovatum CBS 144078 MN699425 MN699376 MN699396 -
Bactrodesmium pallidum CBS 145349 MN699429 MN699380 MN699364 MN704302
Brachysporiella setosa HKUCC 3713 AF132334 - - -
Canalisporium exiguum SS 00809 GQ390281 GQ390266 - HQ446436
Canalisporium grenadoideum BCC 20507 GQ390267 GQ390252 - HQ446420
Cercophora caudata CBS 606.72 AY999113 DQ368659 AY999135 DQ368646
Cercophora newfieldiana SMH 3303 AY780062 - - AY780167
Cercophora thailandica MFLUCC 12-0845 KU863127 KU872131 - KU940176
Cheilymenia stercorea AFTOL 148 AY544661 AY544705 - -
Coleodictyospora muriformis MFLUCC 18-1243TMW981648 MW981704 MW981642 -
Coleodictyospora muriformis MFLUCC 18-1279TMW981649 MW981705 MW981643 -
Conioscypha japonica CBS 387.84 AY484514 JQ437438 - -
Conioscypha lignicola CBS 335.93TAY484513 JQ437439 - JQ429260
Conioscypha peruviana ILL 41202TKF781539 - - -
Conioscypha varia CBS 604.70 MH871656 - MH859869 -
Cosmospora arxii CBS 748.69 MH871181 - NR-145062 HQ897725
Dematiosporium aquaticum MFLU 18-1641TMK835855 - - MN194029
Dematipyriforma aquilaria CGMCC 3.17268TKJ138623 KJ138622 KJ138621 -
Diaporthe cyatheae YMJ 1364 JX570891 JX570890 - JX570893
Diaporthe eres AR 3538 AF408350 - - -
Diaporthe xishuangbanica LC6744 KY011862 - - -
Doratomyces stemonitis AFTOL-ID 1380 DQ836907 DQ836901 - -
Entosordaria perfidiosa BW3 MF488992 - - MF489002
Fuscosporella pyriformis MFLUCC 16-0570TKX550896 KX550900 - KX576872
Fusicolla aquaeductuum KUMCC 18-0015 MH087221 - MH087219 -
Helicoascotaiwania farinosa DAOMC 241947 JQ429230 - JQ429145 -
J. Fungi 2021,7, 711 5 of 24
Table 1. Cont.
Taxon Voucher/Culture GenBank Accession Numbers
LSU SSU ITS rpb2
Helicoascotaiwania farinosa ILLS 53605TAY094189 - - -
Helicoascotaiwania farinosa P2-6 AY316357 - - -
Helicoascotaiwania lacustris CBS 145963TMN699430 MN699382 MN699399 MN704304
Helicoascotaiwania lacustris CBS 145964 MN699431 MN699383 MN699400 MN704305
Helicoascotaiwania lacustris CBS 146144 MN699432 MN699384 MN699401 MN704306
Melanotrigonum ovale CBS 138744 KT278710 KT278697 KT278725 KT278746
Melanotrigonum ovale CBS 138815 KT278711 KT278698 KT278722 KT278747
Melanotrigonum ovale CBS 138743TKT278709 KT278696 KT278724 KT278745
Microascus trigonosporus AFTOL-ID 914 DQ470958 DQ471006 DQ491513 -
Mucispora obscuriseptata MFLUCC 15-0618TKX550892 KX550897 - KX576870
Nectria nigrescens AR 4211 HM484720 JN939647 HM484707 JQ014123
Neoascotaiwania fusiformis MFLUCC 15-0621TKX550893 - MG388215 KX576871
Neoascotaiwania limnetica CBS 126576 KY853513 KT278689 KY853452 MN704308
Neoascotaiwania terrestris CBS 142291TKY853515 KY853547 KY853454 -
Neomonodictys muriformis MFLUCC 16-1136TMN644485 - MN644509 -
Neoroussoella alishanense MFLUCC 11-0190 MN028398 - MN028394 -
Neoroussoella bambusae MFLUCC 11-0124 KJ474839 - KJ474827 KJ474856
Neotorula submersa KUMCC 15-0280 KX789217 - KX789214 -
Neurospora crassa MUCL 19026 AF286411 X04971 - -
Obliquifusoideum guttulatum MFLUCC 18-1233TMW981650 MW981706 MW981645 -
Paracremonium binnewijzendii CBS 143277 MG250174 - NR-157491 -
Parathyridaria percutanea CBS 868.95 KF366449 KF366451 KF322118 KF366452
Phaeoisaria annesophieae MFLU 19-0531 MT559084 - MT559109 -
Phaeoisaria aquatica MFLUCC 16-1298TMF399254 - MF399237 MF401406
Phaeoisaria clematidis MFLUCC 18-1017 MW132065 MW132063 MW131990 -
Phaeoisaria fasciculata CBS 127885TKT278705 KT278693 KT278719 KT278741
Phaeoisaria fasciculata DAOM 230055 KT278706 KT278694 KT278720 KT278742
Phaeoisaria pseudoclematidis MFLUCC 11-0393TKP744501 KP753962 KP744457 -
Phaeoisaria sedimenticola CGMCC 3.14949TJQ031561 - JQ074237 -
Phragmocephala stemphylioides DAOM 673211 KT278717 - KT278730 -
Pisorisporium cymbiforme PRM 924378 KM588902 KM588899 - KM588905
Pleurotheciella aquatica MFLUCC 17-0464TMF399253 MF399220 MF399236 MF401405
Pleurotheciella centenaria DAOM 229631TJQ429234 JQ429246 JQ429151 JQ429265
Pleurotheciella erumpens CBS 142447TMN699435 MN699387 MN699406 MN704311
Pleurotheciella fusiformis MFLUCC 17-0113TMF399250 MF399218 MF399233 MF401403
Pleurotheciella guttulata KUMCC 15-0442 MF399256 MF399222 MF399239 MF401408
Pleurotheciella guttulata KUMCC 15-0296TMF399257 MF399223 MF399240 MF401409
Pleurotheciella krabiensis MFLUCC 16-0852TMG837013 MG837023 MG837018 -
Pleurotheciella lunata MFLUCC 17-0111TMF399255 MF399221 MF399238 MF401407
J. Fungi 2021,7, 711 6 of 24
Table 1. Cont.
Taxon Voucher/Culture GenBank Accession Numbers
LSU SSU ITS rpb2
Pleurotheciella rivularia CBS 125238TJQ429232 JQ429244 JQ429160 JQ429263
Pleurotheciella rivularia CBS 125237 JQ429233 JQ429245 JQ429161 JQ429264
Pleurotheciella saprophytica MFLUCC 16-1251TMF399258 MF399224 MF399241 MF401410
Pleurotheciella submersa DLUCC 0739 MF399259 MF399225 MF399242 MF401411
Pleurotheciella submersa MFLUCC 17-1709TMF399260 MF399226 MF399243 MF401412
Pleurotheciella sympodia MFLUCC 18-1408 MW981652 - MW981644 -
Pleurotheciella sympodia MFLUCC 15-0996 MW981651 MW981703 MW981641 -
Pleurotheciella sympodia MFLUCC 18-0658 MT559086 MT559094 MT555418 -
Pleurotheciella sympodia MFLUCC 18-0983 MT555425 MT555734 MT555419 -
Pleurotheciella sympodia KUMCC 19-0213 MT555426 - MT555420 -
Pleurotheciella tropica MFLUCC 16-0867TMG837015 MG837025 MG837020 -
Pleurotheciella uniseptata DAOM 673210TKT278716 - KT278729 -
Pleurotheciella uniseptata KUMCC 15-0407 MF399248 - MF399231 MF401401
Pleurothecium aquaticum MFLUCC 17-1331TMF399263 - MF399245 -
Pleurothecium aquaticum B-27 MK835854 MK834786 - -
Pleurothecium floriforme MFLUCC 15-0628 KY697277 KY697279 KY697281 -
Pleurothecium obovoideum CBS 209.95 EU041841 - EU041784 -
Pleurothecium pulneyense MFLUCC 16-1293 MF399262 MF399228 - MF401414
Pleurothecium recurvatum DAOM 230069 JQ429238 JQ429252 JQ429157 JQ429269
Pleurothecium semifecundum CBS 131271TJQ429240 JQ429254 JQ429159 JQ429270
Podosordaria tulasnei CBS 128.80 KT281897 - - -
Pseudoascotaiwania persoonii A57-14C AY094190 - - -
Pseudocoleodictyospora sukhothaiensis MFLUCC 12-0554 KU764710 KU712471 KU712440 KU712493
Pseudocoleodictyospora tectonae MFLUCC 12-0385 KU764709 KU712461 KU712443 KU712491
Pseudocoleodictyospora tectonae MFLUCC 12-0387 KU764704 KU712462 KU712444 KU712492
Pseudocoleodictyospora thailandica MFLUCC 12-0565 KU764701 KU712472 KU712441 KU712494
Pseudoneurospora amorphoporcata CBS 626.80 FR774287 - - -
Rhexoacrodictys erecta HSAUPmyr4622 KX033556 KX033526 KU999964 -
Rhexoacrodictys erecta HSAUPmyr6489 KX033555 KX033525 KU999963 -
Rhexoacrodictys erecta KUMCC 20-0194 MT559123 - MT555421 -
Rhexoacrodictys fimicola HMAS47737 KX033553 KX033522 KU999960 -
Rhexoacrodictys fimicola HMAS43690 KX033550 KX033519 KU999957 -
Roussoella nitidula MFLUCC 11-0182 KJ474843 - KJ474835 KJ474859
Saprodesmium dematiosporum KUMCC 18-0059TMW981647 MW981707 MW981646 -
Savoryella aquatica SS 03801 HQ446372 HQ446290 - HQ446441
Savoryella lignicola NF00204 HQ446378 HQ446300 HQ446357 -
Savoryella longispora SAT 00322 HQ446380 HQ446302 HQ446359 HQ446450
Savoryella paucispora SAT 00866 HQ446381 HQ446303 - HQ446451
Savoryella verrucosa SS 00052 HQ446374 - HQ446353 HQ446445
J. Fungi 2021,7, 711 7 of 24
Table 1. Cont.
Taxon Voucher/Culture GenBank Accession Numbers
LSU SSU ITS rpb2
Savoryella yunnanensis MFLUCC 18-1395TMK411422 MK411423 - -
Sordaria fimicola CBS 508.50 AY681160 - - -
Sterigmatobotrys macrocarpa PRM 915682 GU017317 JQ429255 JQ429153 -
Sterigmatobotrys macrocarpa DAOM 230059 GU017316 - JQ429154 JQ429271
Sterigmatobotrys rudis DAOM 229838 JQ429241 JQ429256 JQ429152 JQ429272
Sterigmatobotrys uniseptata MFLUCC 15-0358TMK835850 MK834784 MK878379 -
Subglobosporium tectonae MFLUCC 12-0393 KU764703 KU712464 KU712445 KU712485
Subglobosporium tectonae MFLUCC 12-0390 KU764702 KU712463 KU712446 KU712495
Thyridaria broussonetiae CBS 141481 KX650568 - NR-147658 KX650586
Torula aquatica MFLUCC 16-1115 MG208146 - MG208167 MG207977
Torula herbarum CPC 24114 KR873288 - KR873260 -
Triadelphia uniseptata TA06NZ-142 KT278718 - - -
Tubakia seoraksanensis CBS 127490 KP260499 - - -
Xylaria hypoxylon CBS 122620 KY610495 - KY610407 KY624231
Zalerion maritima FCUL280207CP1 JN886806 KT347203 KT347216 -
Zalerion xylestrix 309156 EU848592 EU848591 - -
The ex-type cultures are indicated using “T” after strain numbers and newly generated sequences are indicated in bold.
2.3.2. Maximum Likelihood Analyses
Each dataset was aligned with MAFFT v. 7.409 online version [
25
] and manually
verified with BioEdit v. 7.2.5 Biological Sequence Alignment Editor (Ibis BioSciences, Carls-
bad, CA, USA), and then concatenated with Mesquite v. 3.11. The maximum likelihood
(ML) analyses were performed with RAxML-HPC v. 8 on XSEDE in CIPRES Science
Gateway [
26
,
27
], with the following changes from the default settings: maximum hours
to run: 5; model for bootstrapping phase: GTRGAMMA; analysis type: rapid bootstrap
analysis/search for best-scoring ML tree (-f a); bootstrapping type: rapid bootstrapping
(-x); bootstrap iterations: 1000 (the maximum value allowed).
2.3.3. Bayesian Inference Analyses
The Bayesian inference (BI) analyses were performed with MrBayes on XSEDE also in
CIPRES Science Gateway [
26
,
27
]. In the first analysis of Savoryellomycetidae, the best-fit
model was GTR+I+G for LSU, ITS and rpb2, and SYM+I+G for SSU. Six simultaneous
Markov chains were run for 965,100 generations and trees were sampled every 100th
generation. In total, 9651 trees were sampled and the first 25% of sampled trees representing
the burn-in phase of the analyses were discarded and the remaining 7239 trees were used
for calculating posterior probabilities (PP) in the majority rule consensus tree (critical value
for the topological convergence diagnostic is 0.01) [28].
In the second analysis, the best-fit model was GTR+I+G for all datasets. Six simul-
taneous Markov chains were run for 685,100 generations and trees were sampled every
100th generation. In total, 6851 trees were sampled and the first 25% of sampled trees
representing the burn-in phase of the analyses were discarded and the remaining 5139 trees
were used for calculating posterior probabilities (PP) in the majority rule consensus tree
(critical value for the topological convergence diagnostic is 0.01) [28].
Phylogenetic trees were viewed with FigTree v. 1.4.03 (http://tree.bio.ed.ac.uk/
accessed on 5 May 2021) and edited with Microsoft Office PowerPoint 2007 (Microsoft
Corporation, WA, USA).
J. Fungi 2021,7, 711 8 of 24
3. Results
3.1. Phylogenetic Analyses
In the first phylogenetic analysis (Figure 1), the representative homologous sequences
of Conioscyphales, Fuscosporellales and Savoryellales and sequences from all genera of
Pleurotheciales representing 90 isolates and two outgroup taxa (Doratomyces stemonitis
AFTOL-ID 1380 and Microascus trigonosporus AFTOL-ID 914) were included. The matrix had
2205 distinct alignment patterns, with 41.06% of completely undetermined characters and
gaps. In the RAxML tree, three distinct independent lineages were identified: (1) one new
genus Obliquifusoideum (no bootstrap support); (2) one new genus Saprodesmium (100% ML
BS/1.00 BI PP); (3) Coleodictyospora (62% ML BS/–) with one new species C. muriformis.
J. Fungi 2021, 7, x FOR PEER REVIEW 8 of 25
Rhexoacrodictyserecta KUMCC 20-0194
Melanotrigonumovale CBS 138744
Sterigmatobotrysrudis DAOM 229838
Pleurotheciella krabiensis MFLUCC 16-0852
T
Pleurotheciumpulneyense MFLUCC 16-1293
Pleurotheciellasympodia MFLUCC 18-1408
Pleurotheciella erumpens CBS 142447
T
Phragmocephalastemphylioides DAOM 673211
Neomonodictys muriformis MFLUCC 16-1136
T
Sterigmatobotrysmacrocarpa PRM 915682
Pleurotheciumsemifecundum CBS 131271
T
Pleurotheciellasaprophytica MFLUCC 16-1251
T
Rhexoacrodictys fimicola HMAS43690
Pleurotheciumaquaticum MFLUCC 17-1331
T
Pleurotheciellauniseptata DAOM 673210
T
Helicoascotaiwania farinosa DAOMC 241947
Pleurotheciellasympodia MFLUCC 18-0983
Pleurotheciella ri vularia CBS 125237
Pleurotheciella ri vularia CBS 125238
T
Pleurotheciella submersa MFLUCC 17-1709
T
Saprodesmium dematiosporium KUMCC 18-0059
T
Pleurotheciellasympodia KUMCC 19-0213
Pleurotheciella submersa DLUCC 0739
Coleodictyospora muriformis MFLUCC 18-1243
T
Pleurotheciellatropica MFLUCC 16-0867
T
Helicoascotaiwanialacustris CBS 145964
Pleurotheciumfloriforme MFLUCC 15-0628
Helicoascotaiwania lacustris CBS 145963
T
Pleurotheciellalunata MFLUCC 17-0111
T
Pleurotheciumobovoideum CBS 209.95
Anapleurotheciumbotulisporum FMR 11490
T
Melanotrigonumovale CBS 138815
Pleurotheciella gu ttulata KUMCC 15-0442
Pleurotheciella s ympodia MFLUCC 15-0996
Sterigmatobotrysuniseptata MFLUCC 15-0358
T
Pleurotheciella uniseptata KUMCC 15-0407
Rhexoacrodictyserecta HSAUPmyr6489
Melanotrigonumovale CBS 138743
T
Sterigmatobotrysmacrocarpa DAOM 230059
Pleurotheciellasympodia MFLUCC 18-0658
Pleurotheciumaquaticum B-27
Rhexoacrodictysfimicola HMAS47737
Pleurotheciellacentenaria DAOM 229631
T
Rhexoacrodictyserecta HSAUPmyr4622
Pleurotheciella fu siformis MFLUCC 17-0113
T
Coleodictyospora muriformis MFLUCC 18-1279
T
Pleurotheciella guttulata KUMCC 15-0296
T
“Brachysporiella setosa” HKUCC 3713
Pleurotheciumrecurvatum DAOM 230069
Adelosphaeriacatenata CBS 138679
T
Pleurotheciellaaquatica MFLUCC 17-0464
T
Helicoascotaiwanialacustris CBS 146144
Helicoascotaiwania farinosa ILLS 53605
T
Dematipyriforma aquilaria CGMCC 3.17268
T
Helicoascotaiwania farinosa P2-6
93/0.99
66/--
75/--
98/1.00
67/0.99
97/0.99
95/--
*
81/1.00
*
92/1.00
*
96/0.99
63/--
62/--
*
70/--
*
78/--
89/1.00
100/1.00
*
99/1.00
98/0.99
93/1.00
*
*
93/0.99
*
91/1.00
98/1.00
*
*
81/1.00
87/1.00
*
*
76/--
*
91/1.00
99/1.00
Pleurotheciales
*
--/0.98
*
Figure 1. Cont.
J. Fungi 2021,7, 711 9 of 24
J. Fungi 2021, 7, x FOR PEER REVIEW 9 of 25
Figure 1. RAxML tree of Savoryellomycetidae with four orders, Conioscyphales, Fuscosporellales,
Pleurotheciales and Savoryellales. The multi-locus tree is generated from combined LSU, SSU, ITS
and rpb2 sequence data. Bootstrap support values for maximum likelihood (the first value) equal to
or greater than 60% and Bayesian posterior probabilities (the second value) equal to or greater than
0.95 are placed near the branches as ML BS/BI PP. The asterisk “*” represents bootstrap support
values with 100% ML BS and 1.00 BI PP. The tree is rooted to Doratomyces stemonitis AFTOL-ID 1380
and Microascus trigonosporus AFTOL-ID 914. The ex-type cultures are indicated using “T” after strain
numbers and the new species introduced in this study are indicated in bold.
In the second multi-locus phylogenetic analysis (Figure 2), a total of seven subclasses
(Diaporthomycetidae, Hypocreomycetidae, Lulworthiomycetidae, Pisorisporiomyceti-
dae, Savoryellomycetidae, Sordariomycetidae and Xylariomycetidae) in Sordariomycetes,
as well as Pseudocoleodictyospora and its relatives in Dothideomycetes were included in the
dataset, representing 55 isolates and one outgroup taxon (Cheilymenia stercorea AFTOL
148). The matrix had 2068 distinct alignment patterns, with 45.12% of completely unde-
termined characters and gaps. In the RAxML tree, Coleodictyospora was phylogenetically
distant from Pseudocoleodictyospora, although their morphology was quite similar [29]. The
relationships of Obliquifusoideum were weak with four orders in Savoryellomycetidae (Fig-
ure 1), but it was shown to be a genus in Savoryellomycetidae with strong bootstrap sup-
port.
0.2
Neoascotaiwaniaterrestris CBS 142291
T
Neoascotaiwaniafusiformis MFLUCC 15-0621
T
Savoryella lignicola NF00204
Savoryellalongispora SAT 00322
Bactrodesmiastrum pyriforme FMR 11931
Savoryella paucisporaSAT 00866
Triadelphia uniseptataTA06NZ 142
Conioscypha peruviana ILL 41202
T
Bactrodesmiastrumobovatum FMR 6482
Conioscypha lignicola CBS 335.93
T
Fuscosporellapyriformis MFLUCC 16-0570
T
Phaeoisaria sedimenticolaCGMCC 3.14949
T
Phaeoisaria fasciculata CBS 127885
T
Bactrodesmiumdiversum CBS 144080
Ascotaiwania latericolla ICMP 22739
T
Neoascotaiwanialimnetica CB S 126576
Canalisporium grenadoideumBCC 20507
Phaeoisaria annesophieaeMFLU 19-0531
Doratomycesstemonitis AFTOL-ID 1380
Savoryella verrucosaSS 00052
Ascotaiwania lignicola NIL 00005
Pseudoascotaiwaniapersoonii A57-14C
Microascustrigonosporus AFTOL-ID 914
Savoryella yunnanensisMFLUCC 18-1395
T
Dematiosporiumaquaticum MFLU 18-1641
T
Conioscypha japonica CBS 387.84
Phaeoisaria pseudoclematidisMFLUCC 11-0393
T
Canalisporium exiguum SS 00809
Obliquifusoideum guttulatum MFLUCC 18-1233
T
Conioscypha varia CBS 604.70
Phaeoisaria fasciculataDAOM 230055
Bactrodesmiumpallidum CBS 145349
Mucispora obscuriseptataMFLUCC 15-0618
T
Phaeoisaria clematidis MFLUCC 18-1017
Savoryella aquatica SS 03801
Phaeoisaria aquatica MFLUCC 16-1298
T
Bactrodesmiumobovatum CBS 14 4078
*
96/1.00
84/--
*
96/1.00
70/--
83/0.97
*
*
68/0.95
95/1.00
100
82/0.99
*
90/--
98/0.99
74/0.98
*
Fuscosporellales
Conioscyphales
Outgroup
4X
4X
Pleurotheciales Savoryellales
*
*
*
*
--/0.99
*
*
*
*
*
*
Figure 1.
RAxML tree of Savoryellomycetidae with four orders, Conioscyphales, Fuscosporellales,
Pleurotheciales and Savoryellales. The multi-locus tree is generated from combined LSU, SSU, ITS
and rpb2 sequence data. Bootstrap support values for maximum likelihood (the first value) equal
to or greater than 60% and Bayesian posterior probabilities (the second value) equal to or greater
than 0.95 are placed near the branches as ML BS/BI PP. The asterisk “*” represents bootstrap support
values with 100% ML BS and 1.00 BI PP. The tree is rooted to Doratomyces stemonitis AFTOL-ID 1380
and Microascus trigonosporus AFTOL-ID 914. The ex-type cultures are indicated using “
T
” after strain
numbers and the new species introduced in this study are indicated in bold.
In the second multi-locus phylogenetic analysis (Figure 2), a total of seven subclasses
(Diaporthomycetidae, Hypocreomycetidae, Lulworthiomycetidae, Pisorisporiomycetidae,
Savoryellomycetidae, Sordariomycetidae and Xylariomycetidae) in Sordariomycetes, as
well as Pseudocoleodictyospora and its relatives in Dothideomycetes were included in the
dataset, representing 55 isolates and one outgroup taxon (Cheilymenia stercorea AFTOL 148).
The matrix had 2068 distinct alignment patterns, with 45.12% of completely undetermined
characters and gaps. In the RAxML tree, Coleodictyospora was phylogenetically distant from
Pseudocoleodictyospora, although their morphology was quite similar [
29
]. The relationships
of Obliquifusoideum were weak with four orders in Savoryellomycetidae (Figure 1), but it
was shown to be a genus in Savoryellomycetidae with strong bootstrap support.
J. Fungi 2021,7, 711 10 of 24
J. Fungi 2021, 7, x FOR PEER REVIEW 10 of 25
Figure 2. RAxML tree with taxa from two classes, Dothideomycetes and Sordariomycetes, to show
the phylogenetic relationships between Coleodictyospora and Pseudocoleodictyospora. The illustrations
of species in Coleodictyospora and Pseudocoleodictyospora are displayed near the generic names. The
multi-locus tree is generated from combined LSU, SSU, ITS and rpb2 sequence data. Bootstrap sup-
port values for maximum likelihood (the first value) equal to or greater than 60% and Bayesian
posterior probabilities (the second value) equal to or greater than 0.95 are placed near the branches
as ML BS/BI PP. The asterisk “*” represents bootstrap support values with 100% ML BS and 1.00 BI
PP. The tree is rooted to Cheilymenia stercorea AFTOL 148. The ex-type cultures are indicated using
“
T
” after strain numbers and the new species introduced in this study are indicated in bold.
In order to assess the phylogenetic position of Obliquifusoideum in Savoryellomyceti-
dae, we constructed individual LSU and ITS phylogenetic trees (shown as Supplementary
Figure 2.
RAxML tree with taxa from two classes, Dothideomycetes and Sordariomycetes, to show
the phylogenetic relationships between Coleodictyospora and Pseudocoleodictyospora. The illustrations
of species in Coleodictyospora and Pseudocoleodictyospora are displayed near the generic names. The
multi-locus tree is generated from combined LSU, SSU, ITS and rpb2 sequence data. Bootstrap
support values for maximum likelihood (the first value) equal to or greater than 60% and Bayesian
posterior probabilities (the second value) equal to or greater than 0.95 are placed near the branches as
ML BS/BI PP. The asterisk “*” represents bootstrap support values with 100% ML BS and 1.00 BI PP.
The tree is rooted to Cheilymenia stercorea AFTOL 148. The ex-type cultures are indicated using “
T
”
after strain numbers and the new species introduced in this study are indicated in bold.
J. Fungi 2021,7, 711 11 of 24
In order to assess the phylogenetic position of Obliquifusoideum in Savoryellomyceti-
dae, we constructed individual LSU and ITS phylogenetic trees (shown as Supplementary
Figures S1 and S2) to enable topological comparison with those derived from the multi-
locus datasets. The matrix of the LSU sequence comprised 92 isolates and had 537 distinct
alignment patterns, with 14% of undetermined characters or gaps. The matrix of ITS
sequence comprised 70 isolates and had 615 distinct alignment patterns, with 30.9% of un-
determined characters or gaps. The phylogenetic position of Obliquifusoideum was different
in all RAxML trees: it clustered with Savoryellales in individual LSU tree (Supplementary
Figure S1), clustered with Savoryellales and Pleurothecium species of Pleurotheciales in
individual ITS tree (Supplementary Figure S2), and clustered with Pleurotheciales in the
multi-locus phylogenetic tree (Figure 1); but without bootstrap support in all trees.
3.2. Taxonomy of Fungi Colonising Decaying Submerged Wood
3.2.1. Novel Taxa in Pleurotheciaceae
In this section, one new genus and two new species are introduced in Pleurotheciaceae.
These taxa are described alphabetically below.
Sordariomycetes O.E. Erikss. and Winka, Myconet 1(1): 10 (1997)
Savoryellomycetidae
Hongsanan, K.D. Hyde and Maharachch., Fungal Diversity 84:
35 (2017)
Pleurotheciales
Réblováand Seifert, in Réblová, Seifert, Fournier and Štˇepánek, Per-
soonia 37: 63 (2016)
Pleurotheciaceae
Réblováand Seifert, in Réblová, Seifert, Fournier and Štˇepánek,
Persoonia 37: 63 (2016)
Coleodictyospora
Charles ex Matsush., Matsushima Mycological Memoirs 5: 8 (1987)
Type species: Coleodictyospora cubensis Charles ex Matsush.
Notes:Coleodictyospora was introduced by Charles [
30
] with a single species C. cubensis,
but it lacked a Latin diagnosis. Matsushima [
31
] validated this genus and characterized it as
having cylindrical, simple, septate and hyaline conidiophores, monoblastic conidiogenous
cells, and transversely oblong or inverse reniform, muriform conidia surrounded by a
semi-gelatinous, hyaline sheath. Berkleasmium micronesicum was then transferred to Coleod-
ictyospora as C. micronesiaca based on its very similar morphological traits with C. cubensis,
but it differs in having smaller conidia (30–42
×
15–18
µ
m vs. 42–50
×
20–22
µ
m) and
reduced conidiophores [31].
Coleodictyospora cubensis was initially collected from North America [
30
] and subse-
quently recorded in Brunei [
32
] and Japan [
33
]. Nakagiri and Ito [
33
] named their specimen
IFO 32,660 as C. cubensis based on the dimensions of conidiophores, conidia and conidial
sheaths, as well as the number of conidial septa and the conidiophore attaching point,
although it had overlapping conidial size with C. micronesiaca. Nakagiri and Ito [
33
]
emphasized that IFO 32,660 might be a novel species considering the thinner conidia
(
28–48 ×13–19 µm
vs. 42–50
×
20–22
µ
m) and less number of septa (7–11 vs. 8–14) than
the type specimen of C. cubensis. However, this hypothesis could not be tested without the
re-examination of the specimen IFO 32,660 or the molecular data from similar specimens in
the same locality (Ishigaki Island, Japan).
Coleodictyospora micronesiaca is likely to be a cosmopolitan species as it was recorded in
several countries worldwide, including China (Hong Kong, Taiwan) [
31
,
34
], Cuba [
35
], USA
(Florida) [36], Mauritius [37], Mexico [38], Micronésia [39], Peru [40], Philippines [37] and
Thailand [
41
]. However, these records were diagnosed solely based on the morphology and
lack of support from molecular data, and the descriptions were omitted or briefly noted.
In this study, we isolated a Coleodictyospora species from decaying wood submerged
in freshwater and provide sequence data for it. Since C. cubensis and C. micronesiaca
lack sequence data in GenBank, we identified our new collection as a novel species in
Coleodictyospora based on the comparison of their morphology.
Coleodictyospora muriformis
W. Dong, Doilom and K.D. Hyde sp. nov.
(Figures 3and 4a,b)
J. Fungi 2021,7, 711 12 of 24
J. Fungi 2021, 7, x FOR PEER REVIEW 12 of 25
Figure 3. Coleodictyospora muriformis (MFLU 18-1544, holotype). (a,b) sporodochia with a mass of
conidia on natural substrate; (c) conidia, conidiogenous cells and conidiophores (arrow shows
branched conidiophore); (d,e) conidia with conidiogenous cells; (f,g) conidiophore bearing conidia;
(h,i) reniform conidia with semi-gelatinous sheaths (h clearly shows the dark brown bands at the
conidial transverse septa); (j) conidium in Indian Ink showing an irregular sheath; (k,l) colony on
PDA (left-front, right-reverse). Scale bars, (c–i) 20 μm; (j) 30 μm.
Figure 3.
Coleodictyospora muriformis (MFLU 18-1544, holotype). (
a
,
b
) sporodochia with a mass
of conidia on natural substrate; (
c
) conidia, conidiogenous cells and conidiophores (arrow shows
branched conidiophore); (
d
,
e
) conidia with conidiogenous cells; (
f
,
g
) conidiophore bearing conidia;
(
h
,
i
) reniform conidia with semi-gelatinous sheaths (h clearly shows the dark brown bands at the
conidial transverse septa); (
j
) conidium in Indian Ink showing an irregular sheath; (
k
,
l
) colony on
PDA (left-front, right-reverse). Scale bars, (c–i) 20 µm; (j) 30 µm.
J. Fungi 2021,7, 711 13 of 24
J. Fungi 2021, 7, x FOR PEER REVIEW 13 of 25
Figure 4. Coleodictyospora spp. (a,b from MFLU 18-1544, holotype. c–e redrawn from Matsushima
[31] and Seifert et al. [42]). (a,b) C. muriformis (conidia with branched conidiophores); (c) C. micro-
nesica (conidia growing on a short subulate conidiogenous cell which directly ascends from the basal
mat of sporodochia); (d,e) C. cubensis (d sporodochia bearing a mass of conidia. e conidia with a
long conidiogenous cell). Scale bars, (a–e) 30 μm.
Index Fungorum number: IF558195; Facesoffungi number: FoF 09872
Etymology: in reference to the muriform conidia of the fungus
Holotype: MFLU 18-1544
Figure 4.
Coleodictyospora spp. (
a
,
b
from MFLU 18-1544, holotype.
c
–
e
redrawn from Matsushima [
31
]
and Seifert et al. [
42
]). (
a
,
b
)C.muriformis (conidia with branched conidiophores); (
c
)C. micronesica
(conidia growing on a short subulate conidiogenous cell which directly ascends from the basal mat
of sporodochia); (
d
,
e
)C. cubensis (d sporodochia bearing a mass of conidia. e conidia with a long
conidiogenous cell). Scale bars, (a–e) 30 µm.
Index Fungorum number: IF558195; Facesoffungi number: FoF 09872
Etymology: in reference to the muriform conidia of the fungus
Holotype: MFLU 18-1544
Saprobic on decaying wood submerged in freshwater. Sexual morph: undetermined.
Asexual morph: hyphomycetous. Colonies on natural substrate, effuse, gregarious, puncti-
form, sporodochial, raised, black. Mycelium partly immersed in natural substrate, consisting
J. Fungi 2021,7, 711 14 of 24
of branched, septate, thin-walled, smooth, pale brown to brown hyphae. Conidiophores up to
55
µ
m long, 3
µ
m wide, micronematous, mononematous, ascending from the basal mat of
sporodochia, subcylindrical, branched, septate, hyaline to pale brown, smooth, thin-walled.
Conidiogenous cells 7.5–20
×
2.5–3.5
µ
m (
x
= 13.5
×
3
µ
m, n = 10), holoblastic, monoblas-
tic, integrated, determinate, terminal, cylindrical, hyaline, smooth, thin-walled. Conidia
32–44 ×15.5–19 µm
(
x
= 38.5
×
17
µ
m, n = 100), solitary, acrogenous, generally produced
in the middle position, occasionally laterally on conidiophores, and perpendicular to the
conidiophores, mostly cylindro-ellipsoidal, sometimes reniform, muriform, dictyoseptate,
with (7–)8–9 transverse and (2–)3 longitudinal septa, deeply constricted and with dark
brown bands at the transverse septa, slightly constricted and brown to dark brown at the
longitudinal septa, often distinctly constricted at the middle where conidiophore attaches to
form reniform, brown, smooth, thin-walled, with a hyaline, semi-gelatinous sheath. Sheaths
well-defined, ellipsoidal, thin at the beginning, 2
µ
m thick; becoming irregular-shaped,
uneven, larger after being mounted in the water, up to 55 µm thick in Indian Ink.
Culture characteristics: on PDA, colony irregular, reaching 15 mm diam. in 25 days at
room temperature (25–30
◦
C), surface rough, with dense mycelia, velvety, dry, umbonate
in the middle from the side view, edge undulate; from above, dark gray at the margin, pale
gray at the middle; from below, dark brown to black at the margin, pale gray at the middle;
not producing pigmentation in culture.
Material examined: THAILAND, Satun Province, Khuan Kalong District, Thung Nui
Sub-District (6
◦
55
0
19” N 100
◦
08
0
17” E), on decaying wood submerged in Chang stream
originated from Panan Waterfall, 10 May 2018, W. Dong, hat284 (MFLU 18-1544, holotype),
ex-type living culture MFLUCC 18-1243 = MFLUCC 18-1279; ibid., HKAS 105018, isotype,
ex-isotype living culture KUMCC 19-0034 = KUMCC 19-0052.
Habitat and distribution: stream is located in tropical rainforest in Southern Thailand
with hot and humid climate conditions, shallow and clear, flowing slowly from the Panan
Waterfall, surrounded by angiosperms.
Notes:Coleodictyospora muriformis belongs in Coleodictyospora based on the punctiform,
sporodochial colonies on the natural substrate, monoblastic conidiogenous cells, and
cylindro-ellipsoidal, muriform conidia produced perpendicularly to the conidiophores
and with a hyaline, semi-gelatinous sheath [
30
,
31
]. Coleodictyospora muriformis is easily
distinguished from the type species C. cubensis by its shorter conidiophores (up to 55
µ
m
long vs. 70–85
µ
m long), smaller conidia (32–44
×
15.5–19
µ
m vs. 42–50
×
20–22
µ
m) and
fewer conidial transverse septa ((7–)8–9 vs. 8–14). The transverse septa of the conidia of
C. muriformis are filled with dark brown bands, while they were neither described nor
illustrated in C. cubensis [
30
,
31
,
33
,
42
]. Coleodictyospora muriformis has overlapping conidial
dimensions with C. micronesiaca, but it differs in having long, branched conidiophores (up
to 55
µ
m long) and longer conidiogenous cells (7.5–20
×
2.5–3.5
µ
m), and the conidiophores
attach to the middle of the conidia. In contrast, C. micronesiaca lacks conidiophores and the
conidiogenous cells are shorter (2–8 ×3–4 µm), which directly ascend from the basal mat
of sporodochia; the conidiogenous cells often attach to the end of the conidia. In addition,
the conidiogenous cells in C. micronesiaca are mostly short subulate, while they are long
cylindrical in C. muriformis. We therefore introduce C. muriformis as new to the genus.
A morphological comparison of Coleodictyospora species is summarized in Table 2and a
combined figure plate of three species is illustrated in Figure 4.
Saprodesmium W. Dong and Doilom gen. nov.
Index Fungorum number: IF558196; Facesoffungi number: FoF 09873
Etymology: “saprus” = saprobic, referring to the saprobic lifestyle of the fungus;
“desmós” = bond, link, referring to the aggregated conidia in sporodochia
Saprobic on decaying wood submerged in freshwater. Sexual morph: undetermined.
Asexual morph: hyphomycetous. Colonies on natural substrate, effuse, gregarious, punc-
tiform, sporodochial, raised, black. Mycelium partly immersed in natural substrate, con-
sisting of branched, septate, thin-walled, smooth, pale brown to brown hyphae. Conidio-
phores micronematous, mononematous, unbranched, vesiculate, septate. Conidiogenous cells
J. Fungi 2021,7, 711 15 of 24
holoblastic, monoblastic, integrated, determinate. Conidia solitary, obovoid to ellipsoidal,
clearly muriform, olivaceous when young, becoming quite blackish with age and obscur-
ing the septa, with several subhyaline basal cells, smooth, thin-walled. Conidial secession
schizolytic.
Table 2. Morphological comparison of Coleodictyospora species (on natural substrate) (update from Nakagiri and Ito [33]).
C. cubensis C. micronesica C. muriformis
Conidiophores 70–85 ×3.5–5 µm
Reduced, 2–8
×
3–4
µ
m (on CMA)
Up to 55 µm long, 3 µm wide
Conidiophore attaching point Middle End Middle
Conidia 42–50 ×20–22 µm 30–40 ×13–16 µm 32–44 ×15.5–19 µm
Conidial sheaths 55–60 ×40–45 µm Present, but not measured Up to 55 µm thick in Indian
Ink
Conidial transverse septa 8–14 6–9 (7–)8–9
Conidial septa with or
without dark bands Not mentioned Not mentioned With dark brown bands
Type species: Saprodesmium dematiosporum W. Dong, Doilom and K.D. Hyde
Notes: The BlastN search of NCBI’s GenBank using the LSU sequence shows
Saprodesmium dematiosporum has the closest hits with several genera in Pleurotheciaceae,
i.e., Rhexoacrodictys erecta (KUMCC 20-0194, similarity = 96.68%), Neomonodictys muri-
formis (MFLUCC 16-1136, similarity = 94.11%) and Pleurothecium obovoideum (CBS 209.95,
similarity = 93.73%
). The closest hits using SSU sequence are Rhexoacrodictys erecta (KUMCC
20-0194,
similarity = 99.54%
), Dematipyriforma aquilaria (3-11-1, similarity = 99.49%) and
Pleurothecium aquaticum (B-27, similarity = 99.06%). Based on ITS BlastN search, the closest
relatives are however Phaeoisaria sp. (BAB-4787, similarity = 97.11%) and Pleurothecium re-
curvatum (CBS 138686, similarity = 96.11%). Saprodesmium dematiosporum clusters as an inde-
pendent branch between Dematipyriforma and Rhexoacrodictys with high bootstrap support
in concatenated LSU-SSU-ITS-rpb2 phylogeny (100% ML BS/1.00 BI PP,
Figure 1
) and indi-
vidual ITS phylogeny (100% ML BS/1.00 BI PP, Supplementary Figure S2). Saprodesmium
dematiosporum clusters with Rhexoacrodictys species in individual LSU phylogeny, but no
bootstrap support (Supplementary Figure S1).
Dematipyriforma is an endophytic genus comprising a single species D. aquilaria [
43
].
Dematipyriforma shares similar morphological characteristics with Saprodesmium in hav-
ing micronematous conidiophores, holoblastic conidiogenous cells and septate conidia.
However, they are entirely different genera in the following aspects. The conidiophores of
Dematipyriforma are hypha-like [
43
], while they are vesiculate in Saprodesmium which are
also unique in the family Pleurotheciaceae. The conidia of Dematipyriforma are elongate
pyriform, 4–5 transverse septate, sometimes 1–2 longitudinal septate, pale grey oliva-
ceous to pale brown, and has rhexolytic conidial secession [43]. In contrast, the conidia of
Saprodesmium are obovoid to ellipsoidal, irregularly muriform and olivaceous when young,
becoming quite blackish with age and obscuring the septa, with several subhyaline basal
cells, and has schizolytic conidial secession. In addition, Saprodesmium species is saprobe,
while Dematipyriforma species is endophyte [43].
Rhexoacrodictys, typified by R. erecta, was introduced for several hyphomycetes charac-
terized by macronematous, long cylindrical conidiophores with percurrent proliferating,
monoblastic, integrated, terminal conidiogenous cells, and obovoid, oval or subspherical,
muriform, brown to dark brown conidia often with a paler basal cell bearing a small
marginal frill derived from the upper portion of the conidiophores and with rhexolytic
conidial secession [
44
]. Saprodesmium shares some morphological traits with Rhexoacrodictys
especially with regards to its muriform and obovoid conidia. Saprodesmium, however, has
olivaceous conidia and when mature it has quite a blackish pigmentation obscuring the
conidial septa. Rhexoacrodictys is featured by rhexolytic conidial secession with conidia
that have a conspicuous paler basal cell bearing a small marginal frill, while the conidia
of Saprodesmium secede schizolytically and it instead has several subhyaline, depressed
J. Fungi 2021,7, 711 16 of 24
subglobose cells at the base. Rhexoacrodictys has macronematous, long cylindrical conidio-
phores with percurrent proliferating [
44
,
45
], whereas Saprodesmium has micronematous,
short, vesiculate, determinate conidiophores.
Based on the multi-locus and individual phylogenetic analyses, as well as the mor-
phological comparison with the similar taxa in the family, we introduce Saprodesmium as a
novel genus in Pleurotheciaceae.
Saprodesmium dematiosporum W. Dong, Doilom and K.D. Hyde sp. nov. (Figure 5)
J. Fungi 2021, 7, x FOR PEER REVIEW 17 of 25
Figure 5. Saprodesmium dematiosporum (HKAS 101710, holotype). (a) sporodochia with a mass of co-
nidia and scattered conidia on natural substrate; (b) conidia and conidiophores; (c) conidia attach
on pseudoparenchyma of sporodochia; (d,e) conidia with conidiogenous cells (arrows); (i) conidial
surface showing muriform patttern; (j,k) colony on PDA (left-front, right-reverse). Scale bars, (b–
e,h) 15 μm; (f,g,i) 10 μm.
Culture characteristics: on PDA, colony circular, reaching 50 mm diam. in 30 days at
room temperature (25–30 °C), surface rough, with dense mycelia, dry, raised from the side
view, edge entire; from above, dark gray at the margin, pale gray to white at the middle;
from below, black at the margin, dark olivaceous at the middle; not producing pigmenta-
tion in culture.
Material examined: CHINA, Yunnan Province, Pingbian District (22°59′13″ N
103°40′30″ E), on decaying wood submerged in an unnamed stream originated from
Figure 5.
Saprodesmium dematiosporum (HKAS 101710, holotype). (
a
) sporodochia with a mass of
conidia and scattered conidia on natural substrate; (
b
) conidia and conidiophores; (
c
) conidia attach
on pseudoparenchyma of sporodochia; (
d
,
e
) conidia with conidiogenous cells (arrows); (
i
) conidial
surface showing muriform patttern; (
j
,
k
) colony on PDA (left-front, right-reverse). Scale bars, (
b
–
e
,
h
)
15 µm; (f,g,i) 10 µm.
Index Fungorum number: IF558197; Facesoffungi number: FoF 09874
J. Fungi 2021,7, 711 17 of 24
Etymology: in reference to the dematiaceous conidia
Holotype: HKAS 101710
Saprobic on decaying wood submerged in freshwater. Sexual morph: undetermined.
Asexual morph: hyphomycetous. Colonies on natural substrate, effuse, gregarious, puncti-
form, sporodochial, raised, black. Mycelium partly immersed in natural substrate, consisting
of branched, septate, thin-walled, smooth, pale brown to brown hyphae. Conidiophores
micronematous, mononematous, vesiculate, consisted of 1–4 subglobose, smooth, hyaline
cells (each cell 8.5–12
µ
m diam.), unbranched, septate, constricted at the septa, smooth,
thin-walled. Conidiogenous cells 8–11
µ
m diam. (
x
= 9.7
µ
m, n = 10), holoblastic, monoblas-
tic, integrated, determinate, terminal, subglobose, hyaline, smooth, thin-walled. Conidia
21–36 ×14.5–27 µm
(
x
= 27.5
×
21.5
µ
m, n = 70), solitary, acrogenous, obovoid to ellip-
soidal, subglobose, clearly muriform, olivaceous when young, becoming quite blackish
with age and obscuring the septa, with several subhyaline to pale brown basal cells, smooth,
thin-walled. Conidial secession schizolytic.
Culture characteristics: on PDA, colony circular, reaching 50 mm diam. in 30 days at
room temperature (25–30
◦
C), surface rough, with dense mycelia, dry, raised from the
side view, edge entire; from above, dark gray at the margin, pale gray to white at the
middle; from below, black at the margin, dark olivaceous at the middle; not producing
pigmentation in culture.
Material examined: CHINA, Yunnan Province, Pingbian District (22
◦
59
0
13” N 103
◦
40
0
30” E),
on decaying wood submerged in an unnamed stream originated from Dawei Mountain Nature
Reserve, 20 September 2017, W. Dong, WF23A (HKAS 101710, holotype), ex-type living culture
KUMCC 18-0059; ibid., MFLU 18-1165, isotype.
Habitat and distribution: stream is nearby Nature Reserve in Southern Yunnan of
Yunnan-Kweichow Plateau, shallow and clear, flowing rapidly from the Dawei Mountain,
surrounded by angiosperms.
3.2.2. Novel Taxa in Savoryellomycetidae
In this section, one new genus with one new species are introduced and phylogeneti-
cally referred to Savoryellomycetidae genera incertae sedis.
Obliquifusoideum W. Dong, Doilom and K.D. Hyde gen. nov.
Index Fungorum number: IF558198; Facesoffungi number: FoF 09875
Etymology: in reference to its neck growing oblique to the host substrate and fusoid
ascospores
Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata super-
ficial, ellipsoidal, black, coriaceous, ostiolate, with a lateral neck. Necks hyaline to dark,
subcylindrical, oblique or horizontal to the host substrate. Peridium thin, soft, comprising
several layers of brown, thin-walled cells of textura angularis.Paraphyses tapering towards
the apex, dense, hypha-like, septate, unbranched, hyaline. Asci 8-spored, unitunicate,
cylindrical, short pedicellate, with a small, refractive, barrel- or jar-shaped, apical ring,
persistent. Ascospores uniseriate, fusoid, septate, hyaline, thin-walled. Asexual morph:
undetermined.
Type species: Obliquifusoideum guttulatum W. Dong, Doilom and K.D. Hyde
Notes: The BlastN search of NCBIs GenBank using LSU sequence shows Obliquifu-
soideum guttulatum has the closest hits with several genera in Pleurotheciaceae, but with low
percentage similarity, i.e., Melanotrigonum ovale (CBS 138743, similarity = 92.06%), Pleurothe-
ciella saprophytica (MFLUCC 16-1251, similarity = 92.03%), Phaeoisaria annesophieae (MFLU
19-0531, similarity = 91.86%) and Sterigmatobotrys rudis (DAOM 229838,
similarity = 91.83%
).
The closest hits using SSU sequence are several genera in Pleurotheciaceae, i.e., Dematipyri-
forma aquilaria (3-11-1, similarity = 98.13%), Phaeoisaria clematidis (MFLUCC 18-1017, similar-
ity = 97.99%) and P. fasciculata (DAOM 230055,
similarity = 97.92%
). The closest hits using
ITS sequence are however several genera in Conioscyphales and Pleurotheciales, i.e., Pleu-
rothecium recurvatum (DAOM 230069, similarity = 98.75%), Conioscypha varia (CBS 604.70,
similarity = 94.89%) and Neomonodictys muriformis (MFLUCC 16-1136,
similarity = 93.19%
).
J. Fungi 2021,7, 711 18 of 24
The placement of Obliquifusoideum guttulatum is different in multi-locus and individual
LSU and ITS phylogenetic trees and lacks significant support in all trees. Obliquifusoideum
guttulatum is revealed as a sister taxon of Pleurotheciales in the multi-locus analysis of
concatenated LSU-SSU-ITS-rpb2 matrix (Figure 1), while it clusters with Savoryellales in
the individual LSU phylogeny (Supplementary Figure S1); Savoryellales and Pleurothecium
species in the individual ITS phylogeny (Supplementary Figure S2). Obliquifusoideum
is similar to Helicoascotaiwania in Pleurotheciales, and Ascotaiwania,Neoascotaiwania and
Savoryella in Savoryellales. They generally have dark ascomata with a lateral neck, which is
oblique or horizontal to the host substrate, and septate ascospores. However, the ascospores
of the four genera are mostly ellipsoidal and versicolorous with dark middle cells and
hyaline polar cells [
2
–
4
,
46
,
47
]. In contrast, Obliquifusoideum has fusoid and evenly hyaline
ascospores. The morphological differences and the independent lineage in the multi-locus
and individual phylogenetic trees therefore support Obliquifusoideum as a new genus.
Although the relationships of Obliquifusoideum were weak with four orders in Sa-
voryellomycetidae (Figure 1), it was shown to be a genus in Savoryellomycetidae with
strong bootstrap support (Figure 2). We consider it is wise to refer Obliquifusoideum to
Savoryellomycetidae genera incertae sedis for now, until its phylogeny is better resolved
with additional taxon sampling followed by divergence time estimates studies.
Obliquifusoideum guttulatum W. Dong, Doilom and K.D. Hyde sp. nov. (Figure 6)
Index Fungorum number: IF558199; Facesoffungi number: FoF 09876
Etymology: in reference to the guttulate ascospores of the fungus
Holotype: MFLU 18-1575
Saprobic on decaying wood submerged in freshwater. Sexual morph: Ascomata
100–120
µ
m high, 155–170
µ
m diam., scattered, superficial, ellipsoidal, black, coriaceous,
ostiolate, with a lateral neck, ejecting asci and ascospores soon during incubation and
becoming empty. Necks 160–180
µ
m long, 17–30
µ
m wide, hyaline to black, subcylindrical,
oblique or horizontal to the host substrate. Peridium thin, 8–20
µ
m thick, soft, comprising
several layers of pale brown, thin-walled cells of textura angularis, dark brown outwards. Pa-
raphyses 3.5–5
µ
m wide, tapering towards the apex, dense, hypha-like, septate, unbranched,
hyaline, embedded in a gelatinous matrix. Asci 97–110
×
7.3–7.7
µ
m (
x
= 105
×
7.5
µ
m,
n = 10
), 8-spored, unitunicate, cylindrical, slightly narrower and truncate at the apex, short
pedicellate, with a small, distinct, refractive, barrel- or jar-shaped, apical ring, 2 ×2.7 µm,
persistent. Ascospores 14–17.5
×
4.3–5
µ
m (
x
= 15.5
×
4.6
µ
m, n = 10), overlapping uniseriate,
fusoid, straight or slightly curved, one median septate, with two additional obscure septate
at two sides, guttulate, hyaline, thin and smooth-walled, without a gelatinous sheath.
Asexual morph: undetermined.
Culture characteristics: on PDA, colony circular, reaching 8 mm diam. in 48 days at
room temperature (25–30
◦
C), surface rough, with dense mycelia, dry, rigid, umbonate
from the side view, edge entire; from above, creamy at the margin, dark grey to dark brown
at the middle, brown at the center; dark brown from below; not producing pigmentation
in culture.
Material examined: THAILAND, Songkhla Province, Rattaphum District, Khao Phra
Sub-District (7
◦
00
0
03” N 100
◦
08
0
33” E), on decaying wood submerged in a stream originated
from Borriphat Waterfall, 10 May 2018, W. Dong, hat138 (MFLU 18-1575, holotype), ex-type
living culture MFLUCC 18-1233; ibid., HKAS 105007, isotype, ex-isotype living culture
KUMCC 19-0023.
Habitat and distribution: stream is located in tropical rainforest in Southern Thailand
with hot and humid climate conditions, shallow and clear, flowing slowly from the Bor-
riphat Waterfall, surrounded by angiosperms.
J. Fungi 2021,7, 711 19 of 24
J. Fungi 2021, 7, x FOR PEER REVIEW 20 of 25
Figure 6. Obliquifusoideum guttulatum (MFLU 18-1575, holotype). (a,b) ascomata lying on submerged
wood; (c) section of ascoma; (d) peridium; (e) section of neck; (f) paraphyses; (g,h) unitunicate asci;
(i,j) apical rings; (k–m) ascospoers (m ascospore in Indian Ink); (n,o) colony on PDA (up-front,
down-reverse). Scale bars, (c) 50 μm; (d,f) 20 μm; (e) 30 μm; (g,h) 15 μm; (i,j) 5 μm; (k–m) 10 μm.
4. Discussion
Doilom et al. [29] established a novel genus Pseudocoleodictyospora to accommodate
three hyphomycetous species collected from the bark of living Tectona grandis (teak) and
distinguished them from Coleodictyospora by the absence of a hyaline sheath. This estab-
lishment, however, lacks the support from the DNA sequence data of Coleodictyospora. The
Figure 6.
Obliquifusoideum guttulatum (MFLU 18-1575, holotype). (
a
,
b
) ascomata lying on submerged
wood; (
c
) section of ascoma; (
d
) peridium; (
e
) section of neck; (
f
) paraphyses; (
g
,
h
) unitunicate
asci; (
i
,
j
) apical rings; (
k
–
m
) ascospoers (m ascospore in Indian Ink); (
n
,
o
) colony on PDA (up-front,
down-reverse). Scale bars, (c) 50 µm; (d,f) 20 µm; (e) 30 µm; (g,h) 15 µm; (i,j) 5 µm; (k–m) 10 µm.
4. Discussion
Doilom et al. [
29
] established a novel genus Pseudocoleodictyospora to accommodate
three hyphomycetous species collected from the bark of living Tectona grandis (teak) and
distinguished them from Coleodictyospora by the absence of a hyaline sheath. This establish-
ment, however, lacks the support from the DNA sequence data of Coleodictyospora. The
presence of conidial sheath as a criterion for delimiting two genera is interesting as this
J. Fungi 2021,7, 711 20 of 24
is often used for species delimitation in classification, such as species in Astrosphaeriella,
Dictyosporium,Kirschsteiniothelia and Natipusilla [
48
]. In this study, we collected a freshwater
hyphomycetous species which has very similar morphs to Pseudocoleodictyospora, but is
characterized by a hyaline sheath. This peculiar phenotype further confirms it as a novel
species in Coleodictyospora, namely C. muriformis (see notes of C. muriformis). On the basis
of DNA-based phylogeny, Coleodictyospora muriformis is phylogenetically distant from Pseu-
docoleodictyospora (Pseudocoleodictyosporaceae, Pleosporales) and clusters as a member of
the Pleurotheciales (Figure 2). This study further confirms the Doilom et al. [
29
] taxonomic
assumption of establishing a novel genus based on the presence of conidial sheath with
the support from the DNA sequence data. Amazingly, Coleodictyospora and Pseudocoleod-
ictyospora are positioned in two different classes Sordariomycetes and Dothideomycetes,
respectively (Figure 2), though they are quite similar except in terms of the conidial sheath.
Nevertheless, it is not advisable to use conidial sheath as a criterion segregating species at a
higher taxonomic level as it is often an unstable characteristic, especially among freshwater
species such as Caryospora submersa and Pseudoastrosphaeriella bambusae [48].
In our multi-locus phylogenetic tree (Figure 1), Coleodictyospora is affiliated to Neomono-
dictys muriformis and Pleurothecium obovoideum.Coleodictyospora is similar to Neomonodictys
in having muriform conidia, but they are entirely different genera. The conidia of Neomono-
dictys are subglobose to globose, comprising several subglobose cells, which are irregularly
arranged in the conidia, pale brown when immature, producing black pigmentation and
obscuring the conidial septa, with a protruding basal cell which attaches to the conid-
iophore [
8
]. In contrast, Coleodictyospora has cylindro-ellipsoidal conidia, with (7–)8–9
transverse and (2–)3 longitudinal septa, deeply constricted and with dark brown bands
at the transverse septa, generally produced in the middle position and are perpendicular
on the conidiophore. Pleurothecium obovoideum was proposed based on a known species,
Ramichloridium obovoideum [
49
]. In the phylogenetic tree of Arzanlou et al. [
49
], they showed
that the strain CBS 209.95 of R. obovoideum clustered with the sexual morph of Pleurothecium
recurvatum (type species) and its morphological characteristics fit well with Pleurothecium
and R. obovoideum was therefore transferred to Pleurothecium, namely P. obovoideum. How-
ever, with more species in Pleurothecium,P. obovoideum was reported to be distant from
P. recurvatum and clustered with Neomonodictys in a well-supported clade [
6
,
8
]. The re-
assessment of P. obovoideum is pending, however, its pleurothecium-like morphological
characteristics [49] warrant it cannot be congeneric with Coleodictyospora.
Interestingly, we found that the conidia of Coleodictyospora are quite similar to the
ascospores of a sexual species Boerlagiomyces websteri.Boerlagiomyces were recognized in
Tubeufiaceae [
11
,
50
,
51
] and confirmed with DNA sequence data derived from a reference
specimen of Boerlagiomyces macrospora [
29
]. However, B. websteri represented by a puta-
tive strain BCC 3834 clustered with several apothecial taxa in Pezizomycotina [
52
], and
Boonmee et al. [
50
] had some doubts on this species because of its perithecial characteristic.
Therefore, the accurate phylogenetic position of B. websteri is still questionable. Although
the soft, membranous, setose ascomata and large dictyosporous ascospores of B. websteri fit
with the features of Boerlagiomyces [
50
], the two-spored asci are unusual compared with
the eight-spored asci of the type species B. velutinus [
50
]. Whether B. websteri has close
phylogenetic relationships with Coleodictyospora in Savoryellomycetidae is pending and
has to be resolved.
On the basis of morphology, we initially considered Obliquifusoideum as a member
of Annulatascales due to its black ascomata with a lateral neck which is oblique or hor-
izontal to the host substrate, hypha-like paraphyses with tapering apex, cylindrical asci
with a distinct, refractive apical ring, and fusoid, hyaline ascospores. It is of interest,
however, that Obliquifusoideum clusters in Savoryellomycetidae with relationships to Pleu-
rotheciales and Savoryellales, which is distant from Annulatascales and annulatascales-like
taxa in Diaporthomycetidae (Figure 2). It is not unexpected that Obliquifusoideum with
annulatascales-like morphology can be discovered in another subclass Savoryellomyceti-
dae as Annulatascales is commonly recognized to be polyphyletic and the species are
J. Fungi 2021,7, 711 21 of 24
often encountered from freshwater habitats [
53
,
54
]. It is reasonable that Obliquifusoideum
is placed in Savoryellomycetidae due to its dark ascomata with an oblique or horizontal
neck, and phragmoseptate conidia which are the sexual features of the other two members
Pleurotheciales and Savoryellales.
DNA-based phylogeny has helped to provide better insights into the taxonomy of
Pleurotheciales and a recommendation of species boundaries was established [
55
], lead-
ing to recent classification updates. Rhexoacrodictys, which was treated as a genus in
Savoryellales by Xia et al. [
56
], was accepted in Pleurotheciales in a later phylogenetic
study [
6
]. Our phylogenetic results corroborate those of Luo et al. [
6
], and our new genus
Saprodesmium forms a well-supported lineage basal to Rhexoacrodictys in Pleurotheciales
(Figure 1). The BlastN search of Saprodesmium using SSU sequence in NCBIs GenBank
reveals a high similarity (99.49%) with a hyphomycetous species Dematipyriforma aquilaria.
Dematipyriforma, typified by D. aquilaria, was isolated as an endophyte from the trunk of
Aquilaria crassna, producing dark muriform conidia [
43
]. Dematipyriforma was placed in
Savoryellales in the phylogenetic tree of Sun et al. [
43
], however, they did not include other
related orders (Conioscyphales, Fuscosporellales and Pleurotheciales) in Savoryellomyceti-
dae. Our multi-locus phylogeny places Dematipyriforma as a sister genus to Rhexoacrodictys
and Saprodesmium with good bootstrap support in Pleurotheciales (
99% ML BS/1.00 BI PP
,
Figure 1). On the other hand, the muriform conidia of Dematipyriforma are similar to
Neomonodictys in Pleurotheciales. According to this morphological trait and phylogenetic
result, we accept Dematipyriforma in Pleurotheciales. Besides saprobes and few opportunis-
tic human pathogens, Dematipyriforma is the only presently known endophytic genus in
the order, which increases our understanding of different life modes of Pleurotheciales.
Freshwater fungi are a unique group of organisms with a special ability to survive
and grow on submerged wood in water by producing soft rot cavities [
57
,
58
]. There is
very little overlap between the fungi growing on wood submerged in freshwater and those
on adjacent stream sides [
59
,
60
]. Thus, we are continually finding novel taxa from this
unique habitat and since streams are often disparate elements, we are likely to discover
many more which will improve our understanding of fungal classification [
61
]. Freshwater
appears to be an ecologically important niche for species in Pleurotheciales [
62
]. Almost all
Pleurotheciella species were collected from freshwater [
3
,
4
,
6
,
9
,
10
,
13
], as well as some species
from Helicoascotaiwania,Phaeoisaria,Pleurothecium and Sterigmatobotrys [6,10,12,15,63].
Supplementary Materials:
The following are available online at https://www.mdpi.com/article/
10.3390/jof7090711/s1, Figure S1: Phylogenetic tree generated from maximum likelihood analysis
(RAxML) based on LSU sequence data, Figure S2: Phylogenetic tree generated from maximum
likelihood analysis (RAxML) based on ITS sequence data.
Author Contributions:
Conceptualization, W.D. and M.D.; resources, N.S., M.D. and Z.D.; writing—
original draftpreparation, W.D.; writing—review and editing, W.D., R.J., K.D.H., E.-F.Y., H.Z., X.Y., G.W.,
N.S., M.D. and Z.D.; supervision, N.S., M.D. and Z.D.; funding acquisition, Z.D. and N.S. All authors
have read and agreed to the published version of the manuscript.
Funding:
This research was funded by the Key Realm R&D Program of Guangdong Province, grant
no: 2018B020205003. This work was partly supported by Chiang Mai University.
Data Availability Statement:
The data generated from this study can be found in the Index Fungo-
rum (http://www.indexfungorum.org/names/names.asp, accessed on 1 August 2021) and GenBank
(https://www.ncbi.nlm.nih.gov/nuccore, accessed on 1 August 2021).
Acknowledgments:
R.J. thanks the University of Mauritius for support. K.D.H thanks the Thailand
Research grant entitled Impact of climate change on fungal diversity and biogeography in the Greater
Mekong Subregion (grant no: RDG6130001). H.Z. thanks the National Natural Science Foundation of
China (Project ID: NSF 31500017). W.D. thanks Shaun Pennycook (Landcare Research, New Zealand)
for assistance in new epithets. Mahboob Mahboobullah is thanked for his preliminary analyses and
suggestions on the manuscript.
Conflicts of Interest: The authors declare no conflict of interest.
J. Fungi 2021,7, 711 22 of 24
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