Biodiversity Data Journal : Taxonomic Paper
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Taxonomic Paper
Neomonodictys aquatica sp. nov. (Pleurotheciaceae) from a plateau lake in Yunnan Province, China
expand article infoSi-Ping Huang, Dan-Feng Bao§,‡,|, Hong-Wei Shen¶,‡,§, Hong-Yan Su, Zong-Long Luo
‡ College of Agriculture and Biological Sciences, Dali University, Dali, China
§ Center of Excellence in Fungal Research, Mae Fah Luang University, Chiang Rai, Thailand
| Department of Entomology and Plant Pathology, Faculty of Agriculture, Chiang Mai University, Chiang Mai, Thailand
¶ School of Science, Mae Fah Luang University, Chiang Rai, Thailand
Open Access

Abstract

Background

In this study, a new species Neomonodictys aquatica was collected from submerged decaying wood in Erhai Lake, Yunnan Province, China.

New information

Neomonodictys aquatica is characterised by acrogenous, solitary, oval, dictyospores (most are transverse septum, occasionally vertical septum, in immaturity the septum is clear, but when mature, the conidia becomes darker so the septum is not clear), smooth-walled conidia. The immature conidia are usually hyaline to olivaceous and mature conidia are usually darkened to black, sometimes with one pale basal cell. Phylogenetic analyses of combined ITS and LSU sequence data showed that the new collection is distinct from other Neomonodictys species. Description and illustration are provided as well.

Keywords

new species, asexual morph, freshwater fungi, phylogeny, taxonomy

Introduction

Pleurotheciales was introduced by Réblová et al. (2016), based on morphological characters and phylogenetic analyses. Members of Pleurotheciales are mostly saprobic on wood (Hyde et al. 2020) and some species have been identified as opportunistic human pathogens (Guarro et al. 2000, Chew et al. 2010, Réblová et al. 2020). Species of the order were collected on submerged decaying wood in lentic and lotic habitats in temperate, subtropical and tropical zones in Asia, Europe, Melanesia and North America (Matsushima 1971, Réblová et al. 2012, Réblová et al. 2016, Réblová et al. 2020, Hernández-Restrepo et al. 2017, Hyde et al. 2018, Hyde et al. 2020, Luo et al. 2018a).

Pleurotheciaceae is a single family of Pleurotheciales. It is typified by Pleurothecium with P. recurvatum as the type species (Morgan) Höhn (Réblová et al. 2016). Recently, Hyde et al. (2020) updated the phylogenetic tree for Pleurotheciales and introduced a new genus Neomonodictys Y.Z. Lu, C.G. Lin & K.D. Hyde. Currently, ten genera are accepted in this family (Réblová et al. 2016, Maharachchikumbura et al. 2016, Hernández-Restrepo et al. 2017, Hyde et al. 2020, Goh and Kuo 2021).

The monophyletic asexual genus Neomonodictys is established for a fungus (Neomonodictys muriformis) collected from a freshwater habitat in Thailand, which is morphologically similar to members of Monodictys S. Hughes (Hyde et al. 2020). Neomonodictys is characterised by holoblastic, monoblastic, integrated, terminal, determinate conidiogenous cells and muriform, subglobose to globose, smooth-walled, pale brown to darkened to black conidia (Hyde et al. 2020).

In this study, the fungus was isolated from submerged decaying wood in Erhai Lake, Yunnan Province in China. The morphology and phylogeny show that our collection is distinct from related species. We provide detailed descriptions, illustrations for Neomonodictys aquatica and a synopsis table for the morphology comparison.

Materials and methods

Isolation and morphological examination

Submerged decaying wood was collected from Erhai Lake, Dali City, Yunnan, China. The coordinates of sampling sites are 25°44′29.65″N, 100°09′49.33″E and at an altitude of 1966 m. Samples were returned to the laboratory in plastic bags. The samples were incubated in aseptic plastic boxes, lined with moistened tissue paper at room temperature for one week. Specimen observations and morphological studies were conducted following the protocols provided by Luo et al. (2018b).

Morphological observations were made by using a SMZ760 series stereomicroscope and photographed using a Nikon-80i microscope. The fungal structures were measured with Tarosoft (R) Image Frame Work programme and images were processed using Adobe Photoshop CS6 extended version 13.0 (Adobe Systems, USA). Single spore isolation was carried out following the method described in Chomnunti et al. (2014). Germinating conidia were transferred aseptically to PDA plates with 0.5 mg/l of Amoxicillin and incubated at room temperature under dark conditions. The colonies were checked every three days. A herbarium was deposited in the herbarium of Cryptogams Kunming Institute of Botany Academia Sinica (KUN-HKAS), Yunnan, China. Living cultures were deposited in Kunming Institute of Botany Culture Collection (KUNCC) and China General Microbiological Culture Collection Center (CGMCC). Facesoffungi numbers were registered as described in Jayasiri et al. (2015) and Index Fungorum numbers as in Index Fungorum(2021).

Molecular Phylogenetic Analyses

DNA Sequencing and Sequence Alignment

The appropriate fungal mycelium was scraped from the surface of colonies on Potato Dextrose Agar (PDA) plates with a scalpel into a 1.5 ml EP tube (Bao et al. 2018). Genomic DNA was extracted using the TreliefTM Plant Genomic DNA Kit (Beijing TsingKe Biological Technology and Services Co. Ltd, China) according to the manufacturer’s protocols.

The primers ITS4/ITS5 for Internal transcribed spacer (ITS) and LR0R/LR5 for Large subunit ribosomal ribonucleic acid (LSU rRNA) were selected for PCR amplification (Vilgalys and Hester 1990). Polymerase Chain Reaction (PCR) mixture was performed in a 25 μl system reaction containing 9.5 μl ddH2O, 12.5 μl of 2 × Power Taq PCR Master Mix, 1 μl of DNA template and 1 μl of each primer (10 μM) (Wang et al. 2019). The PCR thermal cycles for amplification of the ITS gene region were as per Su et al. (2015) and the LSU gene followed Sun et al. (2020). PCR amplifications were confirmed on 1% agarose electrophoresis gels stained with ethidium bromide.

Sequences were assembled with BioEdit. Sequences with high similarity indices were determined from a BLAST search to find the closest matches with taxa in Neomonodictys and from recently published data (Ariyawansa et al. 2015, Wanasinghe et al. 2015, Hyde et al. 2019, Hyde et al. 2020). All consensus sequences and the reference sequences were aligned in MAFFT v. 7 (http://mafft.cbrc.jp/alignment/server/index.html, Katoh and Standley 2013). Aligned sequences of each gene region (ITS and LSU) were combined and manually improved using BioEdit v. 7.0.5.2 (Hall 1999). Ambiguous regions were excluded from the analyses and gaps were treated as missing data.

Phylogenetic Analyses

Maximum Likelihood analysis was performed in the CIPRES Science Gateway v.3.3 (Miller et al. 2010) using RAxML v. 8.2.8 as part of the “RAxML-HPC2 on XSEDE” tool (Stamatakis 2006, Stamatakis et al. 2008). The final ML search was conducted using the GTRGAMMA + I model estimated using MrModeltest 2.2 (Nylander 2004), with ML bootstrap support being calculated from 1000 bootstrap replicates.

Bayesian analysis was performed using MrBayes v. 3.1.2. (Ronquist and Huelsenbeck 2003). The model of each gene was estimated using MrModeltest 2.2 (Nylander 2004), with GTR + I + G model being the best-fit model of ITS and LSU for Bayesian analysis. Posterior Probabilities (PP) (Rannala and Yang 1996) were performed by Markov Chain Monte Carlo sampling (MCMC) in MrBayes v.3.1.2 (Liu et al. 2012). Six simultaneous Markov chains were run for 50 million generations and trees were sampled every 5000th generation (resulting in 10,000 trees). The first 2000 trees, representing the burn-in phase of the analyses, were discarded and the remaining 8000 (post burning) trees were used for calculating posterior probabilities (PP) in the majority rule consensus tree (Cai et al. 2006, Liu et al. 2012).

Phylogenetic trees were visualised by FigTree v. 1.4.4 (Rambaut 2014) and edited in Microsoft Office PowerPoint 2016 (MicrosoftInc. United States). Newly-produced sequences in this study were submitted to GenBank (Table 1).

Table 1.

Isolates and sequences used in this study (newly-generated sequences are indicated in bold and with “*” after species name, the type strains are in bold).

Taxon

Strain

GenBank Accession No.

ITS

LSU

Adelosphaeria catenata

CBS 138679

NR_145396

MH877664

Anapleurothecium botulisporum

FMR 11490

NR_153582

KY853483

Ascotaiwania mitriformis

HKUCC3706

AF132324

Ascotaiwania sawadae

SS00051

HQ446340

HQ446363

Bactrodesmiastrum obovatum

FMR 6482

NR_152537

FR870266

Bactrodesmiastrum pyriforme

FMR 11931

HE646636

HE646637

Brachysporiella setasa

HKUCC 3713

AF132334

Canalisporium caribense

SS03683

GQ390284

GQ390269

Canalisporium elegans

SS00895

GQ390286

GQ390271

Canalisporium exiguum

SS00809

GQ390296

GQ390281

Canalisporium pulchrum

SS03982

GQ390292

GQ390277

Conioscypha japonica

CBS 387.84

AY484514

Conioscypha lignicola

CBS 335.93

AY484513

Conioscypha minutispora

CBS 137253

NR_137847

NG_066275

Conioscypha peruviana

ILL 41202

NG_058867

Conioscypha varia

CBS 113653

AY484512

Fuscosporella pyriformis

MFLUCC 16-0570

NR_152555

NG_059711

Helicoascotaiwania farinosa

DAOM 241947

JQ429145

JQ429230

Melanotrigonum ovale

MR 3685

KT278726

KT278712

Melanotrigonum ovale

CBS 138744

KT278725

KT278710

Melanotrigonum ovale

CBS 138815

KT278722

KT278711

Melanotrigonum ovale

CBS 138743

NR_145397

NG_058197

Melanotrigonum ovale

CBS 138742

KT278723

KT278708

Mucispora obscuriseptata

MFLUCC 15-0618

NR_152556

NG_059709

Neomondictys aquatica *

KUNCC21-10708

MZ686200

OK245417

Neomonodictys muriformis

MFLUCC 16-1136

NR_168231

NG_068916

Parafuscosporella moniliformis

MFLUCC 15-0626

NR_152557

NG_059710

Phaeoisaria aquatica

MFLUCC 16-1298

NR_160592

NG_066194

Phaeoisaria clematidis

MFLUCC 16-1273

MF399229

MF399246

Phaeoisaria clematidis

DAOM 226789

JQ429155

JQ429231

Phaeoisaria clematidis

MFLUCC 17-1968

MG837022

MG837017

Phaeoisaria clematidis

MFLUCC 17-1341

MF399230

MF399247

Phaeoisaria fasciculata

DAOM 230055

KT278720

KT278706

Phaeoisaria fasciculata

CBS 127885

NR_145395

NG_064241

Phaeoisaria guttulata

MFLUCC 17-1965

MG837021

MG837016

Phaeoisaria loranthacearum

CBS 140009

NR_56593

NG_064294

Phaeoisaria pseudoclematidis

MFLUCC 11-0393

NR_155648

NG_059559

Phaeoisaria sedimenticol

CGMCC 3.14949

MK878380

MK835851

Phaeoisaria sparsa

FMR11939

HF677179

HF677185

Phaeoisaria microspora

MFLUCC 16-0033

MF671987

Pleurotheciella aquatica

MFLUCC 17-0464

NR_160591

NG_066193

Pleurotheciella centenaria

DAOM 229631

NR_111709

NG_060098

Pleurotheciella lunata

MFLUCC 17-0111

NR_160593

NG_066195

Pleurotheciella rivularia

CBS 125238

NR_111711

NG_057950

Pleurotheciella rivularia

CBS 125237

JQ429161

JQ429233

Pleurotheciella fusiformis

KUMCC 15-0192

MF399234

MF399251

Pleurotheciella fusiformis

MFLUCC 17-0113

MF399233

MF399250

Pleurotheciella fusiformis

MFLUCC 17-0115

MF399232

MF399249

Pleurotheciella fusiformis

MFLUCC 16-1356

MF399235

MF399252

Pleurotheciella guttulata

KUMCC 15-0442

MF399239

MF399256

Pleurotheciella guttulata

KUMCC 15-0296

NR_160594

NG_066399

Pleurotheciella krabiensis

MFLUCC 16-0852

MG837018

MG837013

Pleurotheciella krabiensis

MFLUCC 16-0858

MG837019

MG837014

Pleurotheciella saprophytica

MFLUCC 16-1251

NR_160595

NG_066196

Pleurotheciella submersa

MFLUCC 17-1709

NR_160596

MF399260

Pleurotheciella submersa

DLUCC 0739

MF399242

MF399259

Pleurotheciella submersa

MFLUCC 17-0456

MF399244

MF399261

Pleurotheciella tropica

MFLUCC 16-0867

MG837020

MG837015

Pleurotheciella uniseptata

KUMCC 15-0407

MF399231

MF399248

Pleurothecium aquaticum

MFLUCC 17-1331

NR_160597

NG_066197

Pleurothecium floriforme

MFLUCC 15-0628

NR_156614

NG_059791

Pleurothecium obovoideum

CBS 209.95

EU041784

EU041841

Pleurothecium pulneyense

MFLUCC 16-1293

MF399262

Pleurothecium recurvatum

CBS 138686

KT278727

KT278715

Pleurothecium recurvatum

CBS 138747

KT278728

KT278714

Pleurothecium recurvatum

CBS 131646

JQ429150

JQ429236

Pleurothecium recurvatum

CBS 131272

JQ429149

JQ429237

Pleurothecium recurvatum

CBS 101581

JQ429148

Pleurothecium semifecundum

CBS 131482

JQ429158

JQ429239

Pleurothecium semifecundum

CBS 131271

NR_111710

NG_057951

Savoryella aquatica

SS 03801

HQ446349

HQ446372

Savoryella lignicola

NF00204

HQ446357

HQ446378

Savoryella longispora

SAT00322

HQ446359

HQ446380

Savoryella paucispora

SAT00866

Savoryella verrucosa

SS 00052

HQ446353

HQ446374

Sterigmatobotrys macrocarpa

PRM 915682

JQ429153

_

Sterigmatobotrys macrocarpa

DAOM 230059

GU017316

Sterigmatobotrys rudis

DAOM 229838

JQ429152

JQ429241

Triadelphia uniseptata

DAOMC 250376

KT278718

Taxon treatment

Neomonodictys aquatica D.F. Bao, S.P. Huang & Z.L. Luo, sp. nov.

Material    Download as CSV 
Holotype:
  1. scientificName:
    Neomonodictys aquatica
    ; kingdom:
    Fungi
    ; phylum:
    Ascomycota
    ; class:
    Sordariomycetes
    ; order:
    Pleurotheciales
    ; family:
    Pleurotheciaceae
    ; genus:
    Neomonodictys
    ; waterBody:
    Erhai Lake
    ; locality:
    Baitaiyi
    ; verbatimElevation:
    1966 m
    ; locationRemarks:
    China, Yunnan Province, Dali, saprobic on submerged decaying wood in Erhai Lake
    ; verbatimLatitude:
    25 44 29.65N
    ; verbatimLongitude:
    100d 09' 49.33'' E
    ; year:
    2020
    ; habitat:
    freshwater, submerged decaying wood
    ; recordedBy:
    Longli Li; Siping Huang
    ; collectionID:
    2EH 3-17-1 H
    ; collectionCode:
    L127.

Description

Sexual morph Undetermined. Asexual morph Hyphomycetous (Fig. 1) sporodochia. Colonies on natural substratum superficial, scattered, black, glistening. Mycelium immersed in the substrate, composed of septate, smooth, thin-walled, light to dark brown, 2–3 μm wide hyphae. Conidiophores lacking. Conidiogenous cells short or occasionally missing, suborbicular, holoblastic, monoblastic, integrated, terminal, determinate, hyaline to pale brown. 3.7–6.4 × 2.9–4.7 μm (x̄ = 5.1 × 3.8 μm, n = 10). Conidia 23.1–29.5 × 8.5–11.5 μm (x̄ = 26 × 10 μm, n = 30), acrogenous, acrospore, oval, ellipsoidal to obovoid, muriform, smooth-walled, hyaline when young, becoming dark brown at maturity sometimes with one pale basal cell.

Figure 1.  

Neomonodictys aquatica (KUN-HKAS 115806, holotype). a Colonies on submerged wood; b-e Conidiophores with conidia; f-j Conidiogenous cells with conidia; k-p Conidia; q Germinating conidium; r, s Colony on PDA. Scale bars: b-c, e, k = 25 μm; d, f-i, l-p = 20 μm; j, q = 30 μm.

Culture characteristics: Conidia germinate on PDA in 36 h. Colonies growing on PDA, subglobose, with flat surface, edge jagged, reaching 3 cm long and 2.5 cm wide in 12 weeks at 28°C, dark grey in PDA medium. Mycelium superficial and partially immersed, branched, septate, hyaline to pale brown, smooth.

Material examined: China, Yunnan Province, Dali, sprobic on submerged decaying wood in Erhai Lake, September 2020, S. P. Huang, L-127 (KUN-HKAS 115806, holotype), ex-type living culture, KUNCC 21-10708 = CGMCC3.20681.

Etymology

Name reflects the aquatic habitat of this fungus

Notes

Morphologically, Neomonodictys aquatica is easily distinguished from N. muriformis. Neomonodictys muriformis has wider conidia than N. aquatica (15–25 vs. 8–12.2 μm). In addition, conidia of N. aquatica are oval or ellipsoidal to obovoid, while N. muriformis has subglobose to globose conidia. In the phylogenetic analysis, N. aquatica clustered with N. muriformis with strong support (99% ML and 1.00 PP) (Fig. 2). ITS comparison between our strain and MFLUCC 16-1136 revealed 57 bp difference in a total of 539 bp. LSU comparison between our strain and MFLUCC 16-1136 revealed 13 bp difference in total of 829 bp (Jeewon and Hyde 2016). Therefore, we introduce our new isolate as a new species.

Figure 2.  

Phylogenetic tree based on RAxML, generated from a combined ITS and LSU dataset. Bootstrap support values for Maximum Likelihood (ML, black) higher than 75% and Bayesian posterior probabilities (BYPP, red) greater than 0.95 are indicated above the nodes as ML/PP. The tree is rooted to Leotia lubrica. The type-derived sequences are indicated in bold and new isolates are in red. Bootstrap values for Maximum Likelihood (ML) equal to or greater than 75% and clade credibility values greater than 0.90 from Bayesian-inference analysis labelled on the nodes. Ex-type strains are in bold and black, the new isolate is indicated in bold and red. (Fig. 2).

Analysis

Phylogenetic analyses

The phylogram generated from Maximum Likelihood analysis, based on combined ITS and LSU sequence data, represents Pleurotheciales and the closely related orders. Seventy-nine strains are included in the combined analyses, which comprise 2039 characters (ITS: 849 bp, LSU: 1190 bp) after aligning. Leotia tubrica (AFTOL-1) is the outgroup taxon in this phylogentic tree. The best RAxML tree with a final likelihood value of -12803.740107 is presented. The matrix had 698 distinct alignment patterns with 34.21% undetermined characters or gaps. Estimated base frequencies were as follows: A = 0.222096, C =0.295691, G = 0.272214, T = 0.209999; substitution rates AC = 1.588217, AG = 2.820721, AT = 2.535737, CG = 1.003016, CT = 5.905028, GT = 1.000000; gamma distribution shape parameter α = 0.570011.

In the phylogenetic analysis, our new isolate Neomonodictys aquatica clustered as a sister taxon with N. muriformis with strong bootstrap support (99 ML/1.00 PP, Fig. 2).

Discussion

Up to now, two species are accepted in Neomonodictys, including the newly-introduced species. Both of them are collected from submerged wood in freshwater habitats (Hyde et al. 2020) and only asexual morphs are reported. Morphologically, Neomonodictys is similar to Monodictys in having solitary, dictyospores conidia and monoblastic, hyaline to brown conidiogenous cells (Ellis 1971, Seifert et al. 2011). Compared with the diaphragms of them, Neomonodictys aquatica have a mostly transverse septum, less of the vertical septum, but the transerve and vertical septa of N. muriformis are evenly distributed. The significant difference between Neomonodictys and Monodictys is conidiophores, which are shorter than in the former (Kukwa and Diederich 2005). Phylogenetically, they are distinct (Hyde et al. 2020). In the phylogenetic analysis, Monodictys was placed in Dothideomycetes (Day et al. 2006, Seifert et al. 2011, Wijayawardene et al. 2020), while Neomonodictys was placed in Sordariomycetes (Hyde et al. 2020).

Acknowledgements

This study was financed and supported by National Natural Science Foundation of China (Project ID: 32060005) and Yunnan Fundamental Research Project (grant NO. 202101AU070137). Si-Ping Huang thanks Zheng-Quan Zhang, Jie Gao, Long-Li Li and Rui Gu for the assistance in sample collection and thanks to Long-Li Li and Xi Fu on DNA extraction and PCR amplification.

References