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Submitted 5 July 2022, Accepted 31 January 2023, Published 4 April 2023
Corresponding Author: Kevin D. Hyde – e-mail – kdhyde3@gmail.com 195
Freshwater fungal biology
Calabon MS1,2,3, Hyde KD1,2,4,5,6*, Jones EBG7, Bao DF1,8,9, Bhunjun CS1,2,
Phukhamsakda C1,10,11, Shen HW1,2,8, Gentekaki E1,2, Al Sharie AH12, Barros J13,
Chandrasiri KSU1,2, Hu DM14,15,16, Hurdeal VG1,2, Rossi W17, Valle LG18,
Zhang H19, Figueroa M20, Raja HA21, Seena S13, Song HY14,22, Dong W6,
El-Elimat T23, Leonardi M17, Li Y10,11, Li YJ19, Luo ZL8, Ritter CD24,25,
Strongman DB26, Wei MJ19 and Balasuriya A27
1Center of Excellence in Fungal Research, Mae Fah Luang University, Chiang Rai 57100, Thailand
2School of Science, Mae Fah Luang University, Chiang Rai, 57100, Thailand
3Division of Biological Sciences, College of Arts and Sciences, University of the Philippines Visayas, Miagao, Iloilo,
5023, Philippines
4Research Center of Microbial Diversity and Sustainable Utilization, Faculty of Science, Chiang Mai University 50200,
Chiang Mai, Thailand
5Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand
6Innovative Institute for Plant Health, Zhongkai University of Agriculture and Engineering, Haizhu District,
Guangzhou 510225, China
7Department of Botany and Microbiology, College of Science, King Saud University, P.O Box 2455, Riyadh, 11451,
Kingdom of Saudi Arabia
8College of Agriculture and Biological Sciences, Dali University, Dali, 671003, China
9Department of Entomology and Plant Pathology, Faculty of Agriculture, Chiang Mai University, Chiang Mai, 50200,
Thailand
10Institute of Plant Protection, College of Agriculture, Jilin Agricultural University, Changchun, Jilin, P.R. China,
130118
11Engineering Research Center of Chinese Ministry of Education for Edible and Medicinal Fungi, Jilin Agricultural
University, Changchun, Jilin, P.R. China, 130118
12Faculty of Medicine, Jordan University of Science and Technology, Irbid 22110, Jordan. orcid.org/ 0000-0003-1311-
806X
13University of Coimbra, MARE - Marine and Environmental Sciences Centre, ARNET – Aquatic Research Network,
Department of Life Sciences, 3004-517 Coimbra, Portugal
14Bioengineering and Technological Research Centre for Edible and Medicinal Fungi, Jiangxi Agricultural University,
Nanchang 330045, China
15College of Bioscience and Bioengineering, Jiangxi Agricultural University, Nanchang 330045, China
16Jiangxi Key Laboratory for Conservation and Utilization of Fungal Resources, Jiangxi Agricultural University,
Nanchang 330045, China
17Department MeSVA, Section Environmental Sciences, University of L’Aquila, 67100, Coppito, AQ, Italy
18Unitat de Botànica. Departament de Biologia Animal, Biologia Vegetal i d’Ecologia. Facultat de Ciències.
Universitat Autònoma de Barcelona. 08193-Bellaterra (Barcelona). Spain
19Shandong Provincial Key Laboratory of Water and Soil Conservation and Environmental Protection, College of
Resources and Environment, Linyi University, West Side of North Section of Industrial Avenue, Linyi 276000, China
20Departamento de Farmacia, Facultad de Química, Universidad Nacional Autónoma de México, Ciudad de México
04510, Mexico
21Department of Chemistry and Biochemistry, the University of North Carolina at Greensboro, Greensboro, North
Carolina 27402, United States
22Key Laboratory of Crop Physiology, Ecology and Genetic Breeding (Jiangxi Agricultural University), Ministry of
Education of the P. R. China, Jiangxi Province 330045, China
23Department of Medicinal Chemistry and Pharmacognosy, Faculty of Pharmacy, Jordan University of Science and
Technology, Irbid, 22110, Jordan
24Instituto Nacional de Pesquisas da Amazônia. Avenida Efigênio Sales, 2239, Manaus, AM 69060-001, Brazil
25Instituto Juruá, Rua das Papoulas, 97, Aleixo, 69083-300 Manaus, AM, Brazil
Mycosphere 14(1): 195–413 (2023) www.mycosphere.org ISSN 2077 7019
Article
Doi 10.5943/mycosphere/14/1/4
196
26Halifax, Nova Scotia, Canada B3L 1S2
27Department of Plant Science, Faculty of Agriculture, Rajarata University of Sri Lanka, Anuradhapura, Sri Lanka
Calabon MS, Hyde KD, Jones EBG, Bao DF, Bhunjun CS, Phukhamsakda C, Shen HW,
Gentekaki E, Al Sharie AH, Barros J, Chandrasiri KSU, Hu DM, Hurdeal VG, Rossi W, Valle LG,
Zhang H, Figueroa M, Raja HA, Seena S, Song HY, Dong W, El-Elimat T, Leonardi M, Li Y,
Li YJ, Luo ZL, Ritter CD, Strongman DB, Wei MJ, Balasuriya A 2023 – Freshwater Fungal
Biology. Mycosphere 14(1), 195–413, Doi 10.5943/mycosphere/14/1/4
Abstract
Research into freshwater fungi has generated a wealth of information over the past decades
with various published articles, i.e., reviews, books, and monographs. With the advancement of
methodologies used in freshwater fungal research, and numerous mycologists working on this
ecological group, our knowledge progress and understanding of freshwater fungi, including novel
discoveries and new insights in the ecology of freshwater fungi, has advanced. With this enormous
progress, it is timely that an updated account of freshwater fungi be compiled in one volume. Thus,
this account is published to give a comprehensive overview of the different facets of freshwater
fungal biology. It includes an updated classification scheme based on the latest taxonomic and
phylogenetic analysis of freshwater fungal taxa, including their evolutionary history. The biology,
diversity, and geographical distribution of higher and basal freshwater fungi are also discussed in
the entries. A section on dispersal and adaptation of filamentous freshwater fungi is included in the
present work. The ecological importance and role of fungi in the breakdown of wood in freshwater
habitats, including their physiology, are discussed in detail. The biotechnological potential of
freshwater fungi as producers of bioactive metabolites are reviewed, with methodologies in
antimicrobial drug discovery. The present volume also provides an overview of different high
throughput sequencing (HTS) platforms for freshwater fungal research highlighting their
advantages and challenges, including recent studies of HTS in identification and quantification of
fungal communities in freshwater habitats. The present volume also identifies the knowledge gaps
and direction of future research in freshwater fungi.
Keywords – Aquatic mycology – biology of microfungi – ecosystem functions – fungal
classification – fungal ecology – taxonomy – systematics
Table of Contents
Introduction (contribution by Mark S. Calabon, E.B. Gareth Jones, Kevin D. Hyde) 197–203
Molecular Phylogeny of Freshwater Ascomycetes (contribution by Dan-Feng Bao, Hong-Wei
Shen, Wei Dong, Zong-Long Luo, Mark S. Calabon, Kevin D. Hyde) 203–217
Biology of Freshwater Basal Fungi (contribution by Vedprakash G. Hurdeal, Eleni Gentekaki)
217–233
The Trichomycetes, an Aquatic Group of Arthropod-Gut Endosymbionts (contribution by Laia
Guàrdia Valle, Douglas B. Strongman) 233–249
Freshwater Laboulbeniales (contribution by Walter Rossi, Marco Leonardi) 249–251
Wood Decay Fungi (contribution by Dian-Ming Hu, Kevin D. Hyde, Hai-Yan Song) 251–263
Evolution of Freshwater Fungi (contribution by Chayanard Phukhamsakda, Chitrabhanu S.
Bhunjun, Abhaya Balasuriya, Yu Li, Kevin D. Hyde) 263–266
197
Antimicrobials from Freshwater Ascomycota (contribution by Ahmed H. Al Sharie, Tamam El-
Elimat, Mario Figueroa, Huzefa A. Raja) 267–287
A Survey of Antimicrobials Produced by Different Taxonomic Groups of Freshwater Ascomycota
(contribution by Huang Zhang, Yong-Jiang Li, Ming-Jie Wei) 287–291
High Throughput Sequencing of Freshwater Fungi (contribution by Chitrabhanu S. Bhunjun,
Chayanard Phukhamsakda, Camila D. Ritter, Kevin D. Hyde) 291–309
Adaptation for Dispersal in Filamentous Freshwater Fungi (contribution by Sajini K.U.
Chandrasiri, Eleni Gentekaki, Kevin D. Hyde) 309–319
Impact of Metal Pollution on Freshwater Fungi: From Cellular Targets to Ecosystems (contribution
by Juliana Barros, Sahadevan Seena) 319–333
Recent Progress and Future Perspectives of Freshwater Fungal Research (contribution by Mark S.
Calabon, E.B. Gareth Jones, Kevin D. Hyde) 333–337
Introduction
Freshwater fungi are morphologically, phylogenetically, and ecologically a diverse group.
For species to be considered freshwater fungus, the life cycle, whole or part, must rely on free
freshwater or submerged substrates from lentic and lotic ecosystems, including artificial reservoirs
and extreme habitats (Thomas 1996, Jones et al. 2014a, Calabon et al. 2020a). Fungi from
freshwater habitats were reported as early as mid-19th century (Russell 1856, Saccardo 1880, de
Wildeman 1893, 1894, 1895) and fungal communities were documented with various studies on
their ecology, biology, biodiversity, and taxonomy (Tsui & Hyde 2003, Krauss et al. 2011, Jones et
al. 2014a, El-Elimat et al. 2021). In early research on freshwater fungi, identification relied mainly
on morphology, and in the case of yeasts, biochemical, fermentation, and assimilation tests.
Molecular data were later incorporated in taxonomic studies of freshwater fungi with early works
of Ranghoo (1998), Ranghoo et al. (1999), Nikolcheva & Bärlocher (2002), and Vijaykrishna et al.
(2006). Following barcoding of nuclear ribosomal regions (ITS, LSU, SSU), protein-coding genes
were added for a better resolution of the phylogenetic tree and resolving the evolutionary
relationships of closely related taxa (Luo et al. 2019, Bao et al. 2020, Hongsanan et al. 2020a, b,
Hyde et al. 2020c, 2021, Dong et al. 2020b). These resulted in a well-defined classification scheme
that is unceasingly changing as continuous exploration of various freshwater habitats in tropical
and temperate countries lead to discovery of novel taxa and recollection of taxa wherein
phylogenetic placement is unclear (Wijayawardene et al. 2020, 2022). Despite all the published
information, our knowledge of freshwater fungi is limited and research on the ecology of this group
is a neglected field. Almost 20 and 8 years after the publications of Tsui & Hyde (2003) and Jones
et al. (2014a), we review the recent information of freshwater fungi regarding their updated
taxonomic classification, numbers, ecological roles and functions, evolution, and adaptation to
changing environmental conditions.
The information on freshwater fungi has been compiled through reviews and books. Ingold
(1975) published an illustrated a guide to aquatic and water-borne hyphomycetes to encourage
mycologists at that time to study freshwater fungi, not only on taxonomic works but also their role
in freshwater habitats. A year later, Jones (1976), in his book Recent Advances in Aquatic
Mycology, brough together information on freshwater and marine fungi, and reviewed relevant
work over the past 12 years. Brlocher (1992c) provided a discussion on aquatic hyphomycetes and
their roles in nature. Almost a decade later, the first book Freshwater Mycology was published that
dealt with the ecology and biology of freshwater fungi, including methodology for physiological
and biodiversity studies (Tsui & Hyde 2003). Cai et al. (2006a) reviewed and compiled descriptions
of 100 freshwater fungal genera with comprehensive description, photographic plates, and notes.
198
Jones et al. (2014a), in the book, Freshwater Fungi and Fungal-like Organisms, reviewed the
recent information on molecular data and classification of freshwater fungi and fungus-like taxa,
with biodiversity and ecological reviews of the group. Several reviews of freshwater fungi
(Bärlocher 1992a, b, Goh & Hyde 1996a, Shearer et al. 2007, Sridhar 2009, Wurzbacher et al.
2010, Chauvet et al. 2016, Gulis et al. 2019, El-Elimat et al. 2021), and keys and monographs
(Gulis et al. 2005, Luo et al. 2019, Dong et al. 2020b) dealing with certain taxonomic and
ecological groups have also been published. In addition to the published literature, the online
databases, www.freshwaterfungi.org and http://fungi.life.illinois.edu/, have compiled all the
scattered data on taxonomic classification of freshwater fungi (Shearer & Raja 2013, Calabon et al.
2020a).
Classification and biodiversity of freshwater fungi
Calabon et al. (2022) provided the latest classification of freshwater fungi and listed 3,870
species reported from different substrates and geographical locations (Table 1). Among these,
2,968 species (in 1,018 genera) belong to Ascomycota, 333 species (in 97 genera) to
Chytridiomycota, 221 species (in 105 genera) to Rozellomycota, and 218 species (in 100 genera) to
Basidiomycota. Other phyla with less than 50 species include Blastocladiomycota,
Monoblepharomycota, Mucoromycota, Aphelidiomycota, Entomophthoromycota,
Mortierellomycota, Olpidiomycota, Zoopagomycota, and Sanchytriomycota. Most freshwater taxa
belong to Sordariomycetes (823 species, 298 genera) and Dothideomycetes (677 species, 229
genera). Pleosporales and Laboulbeniaceae are the largest freshwater fungal order (391 species) and
family (185 species), respectively. Calabon et al. (2022) provides a list of freshwater fungal basal
clades belonging to 11 phyla and 692 species (in 246 genera).
The freshwater fungal numbers in Calabon et al. (2022), 3,870 species, is within the estimated
range of Jones et al. (2014a) which suggested 3,069–4,145. The estimated number accounts for
0.26% of the conservative estimates of fungal species (Hawksworth 2001), and around 3–4% of the
extant fungal species (Kirk et al. 2008, Hyde et al. 2020b, c, Wijayawardene et al. 2020). Although
the list of Calabon et al. (2022) is not exhaustive and does not reflect the overall diversity of
freshwater fungi, the provided number gives an idea of species composition, distribution, and
habitat type to better understand their biology, biodiversity, and ecology.
Table 1 Classification and estimated number of freshwater fungi.
Taxa
Number of genera/species
Jones et al. (2014a)
Calabon et al. (2022)
Ascomycota
Arthoniomycetes
3/7
8/12
Candelariomycetes
–
2/4
Coniocybomycetes
–
1/4
Dothideomycetes
31/86
229/677
Eurotiomycetes
27/158
49/276
Laboulbeniomycetes
1/1
25/259
Lecanoromycetes
52/75
93/185
Leotiomycetes
19/28
82/260
Lichinomycetes
16/43
12/24
Orbiliomycetes
–
10/19
Pezizomycetes
9/9
9/13
Saccharomycetes
–
57/158
Sordariomycetes
61/142
298/823
Ascomycota incertae sedis
3/7
141/252
Pezizomycotina incertae sedis
–
2/2
Basidiomycota
Agaricomycetes
10/14
20/28
Agaricostilbomycetes
–
1/1
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Table 1 Continued.
Taxa
Number of genera/species
Jones et al. (2014a)
Calabon et al. (2022)
Atractiellomycetes
1/1
1/1
Bartheletiomycetes
–
3/5
Classiculomycetes
2/2
2/2
Cystobasidiomycetes
2/6
7/14
Exobasidiomycetes
11/13
7/7
Microbotryomycetes
7/26
14/43
Moniliellomycetes
–
1/1
Tremellomycetes
12/41
28/81
Ustilaginomycetes
2/9
11/30
Agaricomycotina incertae sedis
–
1/1
Basidiomycota genera incertae sedis
11/13
4/4
Aphelidiomycota
Aphelidiomycetes
–
3/15
Blastocladiomycota
Blastocladiomycetes
–
10/47
Chytridiomycota
Chytridiomycetes
97/946
52/181
Cladochytriomycetes
–
7/47
Lobulomycetes
–
3/3
Mesochytriomycetes
–
2/2
Polychytriomycetes
–
5/8
Rhizophlyctidomycetes
–
2/4
Rhizophydiomycetes
–
19/72
Spizellomycetes
–
1/1
Synchytriomycetes
–
3/8
Chytridiomycota genera incertae sedis
–
2/3
Entomophthoromycota
Entomophthoromycetes
–
4/6
Kickxellomycota
Asellariomycetes
3/7
–
Harpellomycetes
44/176–212
–
Monoblepharomycota
Hyaloraphidiomycetes
–
1/1
Monoblepharidomycetes
6/50
5/28
Sanchytriomycetes
–
2/2
Mortierellomycota
Mortierellomycetes
–
3/5
Mucoromycota
Endogonomycetes
–
1/1
Mucoromycetes
–
9/18
Olpidiomycota
Olpidiomycetes
–
1/4
Rozellomycota
Microsporidea
–
105/221
Rozellomycota genera incertae sedis
–
3/20
Zoopagomycota
Zoopagomycetes
–
2/3
Freshwater Ascomycota
Since 1856, the number of novel taxa discovered from freshwater habitats has an increasing
trend with no sign of reaching a plateau (Fig. 1). It has the highest number of discoveries in the past
decade (2010–2019), wherein about 433 species have been discovered from 2010–2019 [data
200
extrapolated from Calabon et al. (2022), Figs 2, 3]. Most are Sordariomycetes represented by 489
freshwater species followed by Dothideomycetes (409 species), Laboulbeniomycetes (259 species),
Ascomycota incertae sedis (174 species), Leotiomycetes (133 species), and Eurotiomycetes (77)
species. The increase in the number of novel taxa discovered over the last years has occurred
because of continuous explorations of freshwater habitats in Asia. In fact, 169 and 129 novel
species were documented from 2015–2020 in China and Thailand, respectively (Bao et al. 2021,
Calabon et al. 2021a). Furthermore, there are more mycologists trained and presently working in
various research and training institutions in Asia focusing on taxonomy and phylogeny of
freshwater fungi. The published works of Luo et al. (2019) and Dong et al. (2020b) on freshwater
Sordariomycetes and Dothideomycetes, respectively, paved the way to the documentation of these
classes, and escalation of their species numbers.
Most studies on freshwater Ascomycota have focused on observation of unidentified
submerged decaying plant substrates (Hyde & Goh 1999, Ho et al. 2001, Tsui et al. 2003, Sivichai
& Boonyene 2004, Hyde et al. 2016a, Lu et al. 2018b). There are reports also of freshwater fungal
associates of specific hosts/habitats [e.g., peat swamp palms Eleiodoxa conferta, Licuala
longicalycata, Metroxylon sagu (Pinruan et al. 2007, 2014), grasses Phragmites, Typha, Scirpus,
Carex, Eriophorum (Webster & Lucas 1961, Pugh & Mulder 1971, Apinis et al. 1972a, b,
Cavaliere 1975, Magnes & Hafellner 1991), and wood (e.g., Alnus glutinosa, Calophyllum
brasiliense, Fagus sylvatica, Pinus roxburghii, Shorea obtusa, S. roxburghii, Wrightia tomentosa,
Xylia xylocarpa, Zollingeria dongnaiensis), shrubs (Beluba, Salix), and bushes (Roldan et al. 1992,
Czeczuga et al. 2005, Baschien et al. 2013, Fiuza et al. 2019)]. Wood test blocks (e.g.,
Dipterocarpus alatus, Erythrophleum teysmannii, Xylia dolabriformis) were also used to determine
species composition of freshwater fungi (Sivichai et al. 2000, 2002, Sivichai & Boonyene 2004,
Boonyuen et al. 2012). Other freshwater substrates include water (Yamaguchi et al. 2007,
Biedunkiewicz & Baranowska 2011, Raposeiro et al. 2018), wood in water cooling towers (Eaton
& Jones 1971a,b), glacial melt waters (Libkind et al. 2003, 2014, de García et al. 2007), foam
(Dixon 1959, Ingold 1967, Descals & Webster 1983, Bärlocher 1987, Harrington 1997, Descals et
al. 1998, Hosoya et al. 2019), rocks (Aptroot & Seaward 2003, Orange 2009, 2013, Shivarov et al.
2017), stemflow and throughfall of tree canopies (Gonczol & Revay 2004, Karamchand & Sridhar
2008, Ghate & Sridhar 2015), wastewaters and polluted freshwater habitats (Spencer et al. 1970,
Woollett & Hedrick 1970, Sláviková & Vadkertiová 1995, Sridhar et al. 2000, Raghu et al. 2001,
Luo et al. 2004, Pires et al. 2017)
Figure 1 – Number of new species discovered from freshwater habitats from 1856–2021.
Freshwater Basidiomycota
Calabon et al. (2022) listed and provided the latest number of freshwater Basidiomycota to be
around 218 species (in 100 genera, 43 families, 26 order, 11 classes). Fifty-six of these are unique
basidiomycetous taxa observed from freshwater habitats. Most of the taxa were under
Agaricomycetes (21 species), followed by Ustilaginomycetes (10 species), Microbotryomycetes (8
species), and Tremellomycetes (6 species) (Fig. 4) (Calabon et al. 2022). Almost 75% of these
basidiomycetous species are yeasts.
201
Filamentous Basidiomycota in freshwater habitats have mostly been documented in woody
and herbaceous substrates (Escobar et al. 1976, Desjardin et al. 1995, Hyde & Goh 1998,
Yamaguchi et al. 2009), foam (Nawawi et al. 1977, Marvanová & Barlocher 1998, Marvanová &
Bärlocher 2000), and sediments (Frank et al. 2010). Agaricomycetes (27 taxa) have mostly been
observed from streams and rivers, followed by Ustilaginomycetes (10 Doassansiopsis species) and
Bartheletiomycetes (5 species) (Calabon et al. 2022). Few taxa were recorded from
Exobasidiomycetes (Burrillia narasimhanii, Pseudodermatosorus alismatis-oligococci,
Rhamphospora nymphaeae), Microbotryomycetes (Camptobasidium hydrophilum, asexual morph
= Crucella subtilis), Atractiellomycetes (Helicogloea angustispora), Classiculomycetes (Classicula
fluitans, Jaculispora submersa), and Tremellomycetes (Xenolachne flagellifera), see Jones et al.
(2014b) and Calabon et al. (2022).
Figure 2 – Decadal data in the number of novel fungi from freshwater habitats.
Figure 3 – Number of freshwater fungal species under Ascomycota.
202
Freshwater basidiomycetous yeasts have also been reported also on a variety of substrates
(e.g., water, aquatic plants and animals, sediment) in a wide range of aquatic environments (e.g.,
polluted and unpolluted rivers, streams, artificial and natural lakes, wastewater, drinking and tap
water, acidic water, glacial meltwater) (Spencer et al. 1970, Libkind et al. 2003, 2009, 2014,
Yamaguchi et al. 2009, Morais et al. 2010, 2020, Brandão et al. 2011). Calabon et al (2022) listed
162 yeasts. Tremellomycetes constitutes 81 species (in 28 genera), followed by
Microbotryomycetes (43 species, 14 genera), Ustilaginomycetes (20 species, 11 genera),
Cystobasidiomycetes (14 species, 7 genera), Exobasidiomycetes (7 species, 7 genera),
Agaricostilbomycetes (Sterigmatomyces elviae), and Moniliellomycetes (Moniliella spathulata).
Figure 4 – Novel Basidiomycota taxa from freshwater habitats.
Freshwater fungal basal clades
The basal lineages of fungi found in freshwater habitats were distributed in 11 phyla with 684
species (Calabon et al. 2022). Chytridiomycota, distinguished by a posterior whiplash uniflagellate
zoospores, constitutes the largest phylum (333 species), followed by Rozellomycota (221 species),
Blastocladiomycota (47 species), Monoblepharomycota (29 species). Few taxa were recorded under
Mucoromycota (19 species), Aphelidiomycota (15 species), Entomophthoromycota (six species),
Mortierellomycota (five species), Olpidiomycota (four species), Zoopagomycota (three species),
and Sanchytriomycota (two species). Most of the basal taxa are parasites or saprobes of freshwater
phytoplankton, zooplankton, plants, fungi, and other invertebrates (Sparrow 1960, Gleason et al.
2008, Hurdeal et al. 2021).
Ecology and biodiversity of freshwater fungi
Fungi from freshwater habitats can be saprobes, mutualists, or parasites and have also been
isolated as endophytes (Brlocher 1992c, Wong et al. 1998, Ibelings et al. 2004, Bärlocher 2007,
Tsui et al. 2016). Saprobic taxa are key decomposers of a wide range of organic substrates, mostly
woody, leaf litter, and herbaceous debris. Freshwater ascomycetes and basidiomycetes are mostly
responsible for the degradation of woody debris (Shearer & Von Bodman 1983, Boonyuen et al.
2014), while hyphomycetes mainly break down leaf litter and herbaceous materials (Bärlocher &
Kendrick 1974, Gessner & Van Ryckegem 2003, Tsui et al. 2016). Freshwater fungi have
mutualistic relationships (Søndergaard & Laegaard 1977, Bärlocher 2007, Kohout et al. 2012) with
203
economically and ecologically important aquatic plants and animals. For instance, freshwater
trichomycetes are endosymbionts attached to the inner gut lining of the host (i.e., insects,
crustaceans, and millepedes) extracting nutrients from the food particles passing through the host’s
digestive system (Lichtwardt et al. 2003, 2014). Lichens are good example also wherein a fungus
has a symbiotic association with photosynthetically active algae or cyanobacteria (Thüs et al.
2014). Aquatic plants and animals are susceptible also to fungal parasites (Ibelings et al. 2003,
Gleason et al. 2014, Glockling et al. 2014). A large and interesting ascomycetous order,
Laboulbeniales, is obligate ectoparasitic on atrthopods, mainly insects. These endophytic, parasitic,
or competitive fungi in general, produce secondary metabolites with an array of biological activities
that enhance their functions contributing to the survival of the species in freshwater ecosystems.
The different molecular methods employed in fungal studies wherein from single or multi-
locus phylogenetic analysis for introduction of novel taxa or reassessment of certain groups,
additional analyses like ancestral state reconstruction methods and evolutionary analysis using
divergence time estimates, were incorporated. These revolutionized our knowledge on the origin,
early history, and evolutionary relationships of freshwater fungi (Vijaykrishna et al. 2006, Luo et
al. 2019, Calabon et al. 2020a, Dong et al. 2020b, Hyde et al. 2021). Ecological studies of
freshwater fungal communities using high-throughput sequencing (HTS) have advanced the field.
HTS are useful in determining the individual microbiome structure in freshwater ecosystems, and
even a community-wide analyses of fungal diversity, and interaction with the environment and
other organisms including forces that influence and shape these communities (Debroas et al. 2017,
Lepère et al. 2019). A vast diversity of Chytridiomycota-like sequences was uncovered in
freshwater habitats using HTS methods by Comeau et al. (2016).
Freshwater fungal communities are susceptible to various environmental changes and human
disturbances. Studies show that climate change can alter the structure of freshwater fungal
communities (e.g., species composition, abundance) in the future (Bärlocher et al. 2008, Dang et al.
2009, Větrovský et al. 2019). Anthropogenic disturbances (e.g., heavy metal loads, nutrient
enrichment, nanoparticles, xenobiotic concentrations) in freshwater habitats may also affect aquatic
fungal diversity and activity, and ecosystem functioning as a whole (Krauss et al. 2003a, Ferreira et
al. 2014).
Molecular Phylogeny of Freshwater Ascomycetes
Introduction
Fungi found in freshwater habitats can be classified into several morphological and
ecological groups: freshwater ascomycetes, freshwater hyphomycetes (i.e., Ingoldian fungi, aero-
aquatic hyphomycetes or asexual ascomycetes, terrestrial-aquatic hyphomycetes, submerged-
aquatic hyphomycetes), freshwater basidiomycetes, coelomycetes, zygomycetes, microsporidia,
and zoosporic fungi (Goh & Hyde 1996a, Shearer et al. 2007, Tsui et al. 2016, Schuster et al.
2022). Freshwater fungi are distributed in thirteen phyla: Aphelidiomycota, Ascomycota,
Basidiomycota, Blastocladiomycota, Chytridiomycota, Entomophthoromycota,
Monoblepharomycota, Mortierellomycota, Mucoromycota, Olpidiomycota, Rozellomycota,
Sanchytriomycota, and Zoopagomycota, with Ascomycota being the most speciose (Calabon et al.
2022).
In contrast to a taxonomic group that represents freshwater fungi as distinct lineages,
freshwater fungi constitute a phylogenetically varied Ascomycota group that may be conceived of
as an ecological group. Freshwater ascomycetes have been found in freshwater lentic (ponds, pools,
lakes, peat swamps) and lotic (creeks, streams, brooks, rivers) habitats and complete part or all of
their lifecycle within the freshwater environment (Shearer 1993, Wong et al. 1998, Tsui & Hyde
2003, Jones et al. 2014a). They also include sexual (teleomorphs) and asexual morphs (anamorphs)
of ascomycetes that grow on submerged wood and leaves (Cai et al. 2006b, Vijaykrishna et al.
2006). Freshwater fungi can also be found in artificial aquatic habitats in urban locations such as,
gutters, sewage, water-cooling towers, water pipes, and wastewater treatment plants (Jones & Eaton
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1969, Hosagoudar & Udaiyan 1993, Kane et al. 2002, Ghate & Sridhar 2018, Grossart et al. 2019),
as well as in ecologically extreme conditions, including, varied static and water movement,
pressure, temperature, nutrients, and salinity (Nakatsu & Hutchinson 1988, López-Archilla et al.
2001, Gadanho & Sampaio 2006, Gadanho et al. 2006, Vishniac 2006, Branda et al. 2010, Buzzini
et al. 2012, Libkind et al. 2004, 2014).
The study of fungi in freshwater habitats began in the 1880s. Pioneer mycologists were drawn
to the tetraradiate, sigmoid, and branching conidia because they were unusual. Heliscus
lugdunensis was initially reported as a hyphomycete species from freshwater by Saccardo (1880).
De Wildeman (1893, 1894, 1895) discovered four new fungal species on various substrates in
ponds, ditches, and marshy areas, making a significant addition to aquatic hyphomycete research.
When Ingold (1942) discovered and introduced species from a typical freshwater habitat, growing
on submerged decaying leaves in well-aerated waters, he made a significant breakthrough. Later,
Ingold discovered the structure and details of 16 freshwater fungal species, ten of which were new
(Ingold 1942, 1953). However, it took almost 50 years after De Wildeman to observe the first
aquatic ascomycete where the sexual state was known, Aquanectria penicillioides (= Flagellospora
penicillioides) (Ranzoni 1956).
Ingold (Ingold 1951, 1954, 1955) noted the profusion of freshwater ascomycetes on reed
swamp plant stalks in the 1950s, with many of the ascospores having well developed appendages,
such as, Ceriospora caudaesuis and Loramyces macrospora. Following Ingold's first study, many
extensive investigations were published (Ranzoni 1953, Tubaki 1957, Petersen 1963, Ingestad &
Nilsson 1964, Jones & Eaton 1969, Eaton & Jones 1970, Webster & Descals 1981, Descals et al.
1981, Dudka 1985, Goh & Hyde 1996a, Chan et al. 2000, Pinruan et al. 2004a, b, Pinnoi et al.
2006). Numerous studies on sexual and asexual ascomycetes have been published from all over the
world (Tubaki et al. 1983, Hyde 1992a, Sridhar et al. 1992, Chang et al. 1998, Sivichai et al. 1998,
2011, Suh et al. 1999, Wong et al. 1999b, Hyde & Wong 2000, Tsui et al. 2001a, c, Tsui & Hyde
2003, Pinruan et al. 2004b, Shearer et al. 2004, 2007, 2014, Zhang et al. 2011, Liu et al. 2015b).
Whereas Ingold emphasized the prevalence of ascomycetes in temperate freshwater habitats (Ingold
1951, 1955), Hyde emphasized their presence in tropical locales such as Australia (Hyde 1992b),
Taiwan (Chang et al. 1998), the Philippines (Hyde & Wong 2000, Cai et al. 2003), and Hong Kong
(Tsui et al. 2001b, c, Dong et al. 2020b). Apart from that, Neubert et al. (2006) conducted a genetic
assessment of the phanerogam Phragmites australis' fungal diversity and discovered 350 different
operational taxonomic units (OTU). Many of the fungi were yet to be identified (Luo et al. 2004,
Šlapeta et al. 2005, Hyde et al. 2020b). In recent years, several new species, genera, families, and
orders of freshwater ascomycetes have been discovered (Zhang et al. 2017, Li et al. 2017, Bao et al.
2018, 2021, Luo et al. 2019, Calabon et al. 2020b, 2021a, b, Dong et al. 2020b, 2021a, b). Shearer
et al. (2014), Luo et al. (2019), Dong et al. (2020b), and Calabon et al. (2022) are the most recent
notable papers on freshwater ascomycetes.
Currently, there are 2,968 freshwater fungal species in Ascomycota, in 1,108 genera. Most
studies on freshwater ascomycetes have been morphological, with sequencing data utilized to aid in
the resolution of phylogenetic relationships. The earliest use of sequence data to resolve the
taxonomy of a freshwater ascomycetes was in the early 2000’s (Inderbitzin et al. 2001, Pang et al.
2002, Shearer et al. 2009). Freshwater ascomycetes are distributed in 13 classes: Arthoniomycetes,
Candelariomycetes, Coniocybomycetes, Dothideomycetes, Eurotiomycetes, Laboulbeniomycetes,
Lecanoromycetes, Leotiomycetes, Lichinomycetes, Orbiliomycetes, Pezizomycetes,
Saccharomycetes, and Sordariomycetes. Sordariomycetes is the largest of the thirteen fungal
classes (823 species, 28%), whereas Dothideomycetes account for 23% (677 species) (Calabon et
al. 2022).
The use of molecular sequence data has substantially enhanced the present classification
system of the Kingdom of Fungi (Hibbett et al. 2007, Maharachchikumbura et al. 2015, Spatafora
et al. 2017). Shearer et al. (2014) presented a phylogenetic tree of Dothideomycetes based on
molecular data and provided the phylogenetic placement of freshwater taxa. Four orders, i.e.,
Pleosporales, Jahnulales, Natipusillales, Tubeufiales, constitutes the major freshwater
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Dothideomycetes. Cai et al. (2014) used LSU sequence data to provide the phylogenetic placements
of freshwater Sordariomycetes which includes three subclasses: Sordariomycetidae,
Hypocreomycecetidae, and Xylariomycetidae, with 11 orders (Calosphaeriales, Coniochaetales,
Diaporthales, Hypocreales, Magnaporthales, Microascales, Phyllachorales, Savoryellales,
Sordariales, Trichosphaeriales, Xylariales). Luo et al. (2019) undertook a comprehensive classification
of freshwater sordariomycetous taxa, by a multi-locus phylogenetic tree, which are distributed in five
subclasses viz. Diaporthomycetidae, Hypocreomycetidae, (no freshwater taxa) Savoryellomycetidae,
Sordariomycetidae and Xylariomycetidae. Dong et al. (2020b) outlined the genera of freshwater
Dothideomycetes with comprehensive notes on taxa, and multi-locus phylogenetic analysis for
freshwater Dothideomycetes. Six orders, 43 families and 145 genera of Dothideomycetes include
freshwater taxa.
The use of DNA data has substantially expanded taxonomic research on freshwater fungi,
resulting in a rapid increase in fungal numbers (Zhang et al. 2017, Bao et al. 2019, 2020, Calabon
et al. 2021a) and recommendations to follow when introducing new species have been published by
Chethana et al. (2021) for fungi in general, Maharachchikumbura et al. (2021) for Sordariomycetes
and Pem et al. (2021) for Dothideomycetes. Dong et al. (2020b) also opined, that huge numbers of
fungi have yet to be reported in underexplored areas. Freshwater Sordariomycetes and
Dothideomycetes are well-studied with molecular data, while other classes of freshwater
ascomycetes are poorly studied, most of them were identified solely based on morphology, and if
coupled with phylogenetic analysis, only a single locus is available.
Herein, we review freshwater fungi in the main classes: Eurotiomycetes, Dothideomycetes,
Sordariomycetes, and conclude by considering other Ascomycota classes.
Freshwater Dothideomycetes
Dothideomycetes, are an intriguing ascomycetous class due to their incredible diversity of
lifestyles, habitats, and spores, as well as studies of their ecological, evolutionary, biological, and
taxonomic status (Suetrong et al. 2009, Hyde et al. 2013, Haridas et al. 2020, Hongsanan et al.
2020a, b, Saxena et al. 2021). It has become clear that Dothideomycetes is a single entity in
Ascomycota based on a divergence time and multi-locus phylogenetic study (Liu et al. 2017). With
more and more DNA sampling, Dothideomycetes was revealed to have evolved several lineages
with distinctive genetic variations to adapt to freshwater environments (Inderbitzin et al. 2001,
Shearer et al. 2009, Raja et al. 2012, 2015). In the past decade, many novel freshwater species have
been established in Dothideomycetes (Pang et al. 2002, Ferrer et al. 2011, Raja et al. 2011, 2013b,
Zhang et al. 2014a, Hyde et al. 2020a, b) and some higher taxa were also proposed (Fig. 5).
Natipusillales is the only order in Dothideomycetes with fusiform or clavate ascospores having
complex gelatinous sheaths and appendages (Hyde et al. 2013). Minutisphaeraceae
(Minutisphaerales) and Wicklowiaceae (Pleosporales) are another two distinct lineages with all
species from freshwater habitats (Dong et al. 2020b). Nevertheless, most freshwater species are
distributed throughout families of Pleosporales (the largest order) and Tubeufiales, and with
affinities to marine and terrestrial fungi (Lu et al. 2018b, Dong et al. 2020b, Calabon et al. 2022).
To better understand the fungal diversity of this class and its systematics, Dong et al. (2020b)
reviewed all freshwater Dothideomycetous species including their worldwide distribution,
taxonomic problems, phylogenetic relationships, and possible morphological traits adapted to
freshwater environments.
Molecular phylogeny of freshwater Dothideomycetes
The first major molecular phylogeny of freshwater Dothideomycetes was initially
investigated by Shearer et al. (2009). The results indicated that all freshwater taxa clustered in
Pleosporomycetidae as opposed to Dothideomycetidae. Four clades were revealed comprising only
freshwater taxa, with the Jahnulales clade as the largest of these, followed by Lindgomycetaceae,
Amniculicolaceae and Lentitheciaceae. However, further molecular studies showed that this
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Figure 5 – Freshwater Dothideomycetes. a Setoseptoria magniarundinacea (culture of KT 1174 =
CBS 139702). b Neotrematosphaeria biappendiculata (KT 1124, holotype, in black-blue ink).
c Wicklowia phuketensis (MFLU 20–0143). d Aquastroma magniostiolatum (MFLU 22–0121, in
Indian Ink). e Ascagilis submersa (MFLU 18–1527, holotype). f Jahnula appendiculata (PE0010).
g Aliquandostipite khaoyaiensis (MFLU 21-0125. h Aquimassariosphaeria kunmingensis (HKAS
102148, holotype). i Pseudojahnula potamophila (F111). j Mamillisphaeria dimorphospora (BRIP
22967a). k Byssothecium circinans (G-K 18367). l Caryospora aquatica (MFLU 18–1202).
m Neohelicascus elaterascus (MC0430–1). n Clohesyomyces aquaticus (MFLU 11–1112).
o Acrocalymma aquatica MFLU 22–0114. p Pseudoxylomyces elegans MFLU 20–0554. Scale
bars: a–p = 10 µm
ecological group was not monophyletic as some of these evolved together with terrestrial and
marine fungi (Raja et al. 2013a, Zhang et al. 2014b, Ariyawansa et al. 2015, Fournier et al. 2015,
Luo et al. 2016a, Huang et al. 2018). More recently, multi-locus phylogeny of Dothideomycetes
showed freshwater taxa were scattered in most pleosporalean families with many in Tubeufiaceae,
Tubeufiales (Dong et al. 2020b) (Fig. 6). There are 46 genera exclusively found in freshwater
habitats, with 14 freshwater species reported in Lindgomyces (Lindgomycetaceae, Pleosporales), 11
in Jahnula (Aliquandostipitaceae, Jahnulales) and eight in Neohelicascus (Morosphaeriaceae,
Pleosporales) (Dong et al. 2020b, Calabon et al. 2022). Based on a multi-locus phylogeny and
morphology, a taxonomic revision of Helicascus resulted in the transfer of some species to
Aquihelicascus and Neohelicascus, but marine species, including the type species, were retained in
the genus (Dong et al. 2020b).
Pang et al. (2002) introduced Jahnulales in Dothideomycetes with two families
Aliquandostipitaceae and Manglicolaceae, which contain approximately 80 species
(Wijayawardene et al. 2020). Aliquandostipitacea was introduced by Inderbitzin et al. (2001) and
typified with Aliquandostipite. The family has undergone significance changes with further
research resulting in nine genera: Aliquandostipite, Ascagilis, Brachiosphaera, Jahnula,
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Megalohypha, Neojahnula, Pseudojahnula, Speiropsis, and Xylomyces (Suetrong et al. 2011, Dong
et al. 2020b, Wijayawardene et al. 2020).
Figure 6 – Radial phylogenetic circular tree representation of freshwater Dothideomycetes.
Distribution and discussion of freshwater Dothideomycetes (Based on molecular data)
The distribution of freshwater Dothideomycetes is an eternal and unresolved topic as
distribution patterns are still largely based on the locations of researchers and this limits discussion.
Dothideomycetes is one of the largest classes in the phylum Ascomycota and it is also shown to be
highly adapted to freshwater environments (Dong et al. 2020b) as compared to Eurotiomycetes (Liu
et al. 2015b, Dong et al. 2018, 2020a, Wang et al. 2019). Therefore, freshwater Dothideomycetes
are distributed all over the world where suitable freshwater environments occur for fungal growth.
Freshwater dothideomycetous species have been extensively studied in some countries ranging
from temperate to tropical regions, such as Australia (Hyde 1995, Hyde & Wong 1999), Japan
(Tanaka et al. 2009, 2015), China (Hong Kong, Guangxi, Guizhou, Yunnan) (Tsui et al. 1999, Ho
et al. 2000, Su et al. 2016, Lu et al. 2018b), Thailand (Luo et al. 2016c, Zhang et al. 2016) and USA
(Raja et al. 2010, 2015). Dong et al. (2020b) concluded that freshwater Dothideomycetes are
distributed in 35 countries, with China having the most species, followed by Thailand. Mycologists
are questioning if global warming will affect fungal distribution (Hyde et al. 2016a) with studies
undertaken by Luo et al. (2019) and Dong et al. (2020b). However, because of the limited number
of studies, it is still untenable to speculate how climate change will exactly affect the distribution of
freshwater fungi, but they are very likely to be sensitive to environmental change and global
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warming (Hyde et al. 2016a). Based on the current data, many species occur worldwide, e.g.,
Aliquandostipite crystallinus, Aquihelicascus thalassioideus, Jahnula aquatica and Lindgomyces
ingoldianus (Dong et al. 2020b). It is still puzzling to understand the subtle connection between
these cosmopolitan species and freshwater ecology. Another challenge is that researchers are facing
a problem dealing with the fungal taxa discovered earlier because DNA sequence data was not
available, therefore the identification and geographical distribution must be treated with caution.
The worldwide distribution of freshwater Dothideomycetes, with or without molecular data, are
listed in Dong et al. (2020b).
Freshwater Sordariomycetes
Sordariomycetes is the second largest class of Ascomycota, after Dothideomycetes (Hyde et
al. 2020c). Research on Sordariomycetes started from the early morphological studies of Barr
(1983, 1987, 1990) and Eriksson & Hawksworth (1986, 1993) to the incorporation of molecular
data by Lumbsch & Huhndorf (2007, 2010), Maharachchikumbura et al. (2015, 2016), and
Hongsanan et al. (2017). The recent classification of Sordariomycetes is continuously updated and
recently compiled by Hyde et al. (2020c), with seven subclasses distributed in 45 orders and 167
families, as outlined by Wijayawardene et al. (2020). Sordariomycetous taxa are mainly
characterized by non-lichenized, perithecial ascomata and inoperculate unitunicate or non-
fissitunicate asci (Maharachchikumbura et al. 2016, Hyde et al. 2020c). Members of
Sordariomycetes have a cosmopolitan distribution and are mostly observed from terrestrial habitats.
In aquatic habitats, Sordariomycetes are mostly saprobic on submerged decaying wood (Luo et al.
2019, Calabon et al. 2022) (Fig. 7).
From the discovery of Lunulospora curvula by Ingold (1942) in submerged decaying leaves
of Alnus glutinosa and Salix, to the recent outline and monograph of freshwater Sordariomycetes
by Luo et al. (2019), knowledge on the classification of freshwater Sordariomycetes has improved
significantly. Annulatascaceous, distoseptisporaceous, pleurotheciaceous, and halosphaeriaceous
species are the most typical and common freshwater Sordariomycetes on submerged wood (Luo et
al. 2019, Calabon et al. 2022).
Molecular phylogeny of freshwater Sordariomycetes
The first major phylogenetic analysis of freshwater Sordariomycetes was by Cai et al. (2014)
and based on LSU sequence data, represented in three subclasses (Sordariomycetidae,
Hypocreomycecetidae, Xylariomycetidae) and 13 orders. The phylogenetic analysis of Cai et al.
(2014) resulted in a phylogenetically polyphyletic Annulatascaceae with members distributed in
five clades. Later, Maharachchikumbura et al. (2015) introduced Annulatascales to accommodate
the family Annulatascaceae and referred to the Diaporthomycetidae. The most comprehensive
phylogenetic study of combined LSU, SSU, RPB2 and TEF1α sequence data of freshwater
Sordariomycetes was conducted by Luo et al. (2019) wherein 356 freshwater fungal strains were
included in the study, with 129 fresh isolates. The 854 strains clustered in six Sordariomycetes
subclasses, Diaporthomycetidae, Hypocreomycetidae, Lulworthiomycetidae, Savoryellomycetidae,
Sordariomycetidae, and Xylariomycetidae (Lumbsch & Huhndorf 2010, Maharachchikumbura et
al. 2015, Hongsanan et al. 2017). Freshwater Sordariomycetes is well-distributed in 47 clades under
30 orders. Hypocreales contains the greatest number of genera (19 genera), followed by
Chaetosphaeriales (16 genera), Sordariales (13 genera), and Annulatascales (10 genera) (Fig. 8).
Hyde et al. (2021) studied the evolution of freshwater Diaporthomycetidae with a divergence time
of 238 MYA, with the introduction of novel taxa (orders: Barbatosphaeriales, Cancellidiales,
Ceratolentales, Conlariales, Rhamphoriales; families: Aquapteridosporaceae, Cancellidiaceae,
Ceratolentaceae, Bullimycetaceae, Phialemoniopsaceae, Pseudostanjehughesiaceae; species:
Cancellidium atrobrunneum, C. cinereum, C. griseonigrum.). Lately, newly introduced taxa are
supported with molecular data, and it gives a better understanding of the phylogeny and evolution
of freshwater Sordariomycetes. Unfortunately, the placements of many freshwater Sordariomycetes
remains unresolved and there are taxa without living cultures thus sequence data are unavailable.
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Distribution and discussion of freshwater Sordariomycetes (Based on molecular data)
Like freshwater Dothideomycetes, the distribution of freshwater Sordariomycetes is still
dependent on the locations of the laboratories wherein researchers are active in doing fungal
explorations on this group. There are taxa frequently isolated in tropical and temperate regions like,
Annulatascus velatisporus (Hyde 1992b, Hyde et al. 1998, Wong et al. 1999a, Ho et al. 2001, Tsui
et al. 2003, Hu et al. 2010, Sudheep & Sridhar 2011, Dayarathne et al. 2016); Ophioceras commune
(Shearer et al. 1999, Tsui et al. 2001c, 2003, Raja et al. 2009, Abdel-Aziz 2016); Aquanectria
penicillioides (Duarte et al. 2012, Ghate & Sridhar 2015, Lombard et al. 2015, Mun et al. 2016,
Luo et al. 2019); Neonectria lugdunensis (Gulis & Suberkropp 2003, Baschien et al. 2006, 2008,
Cornut et al. 2014, Raposeiro et al. 2018, Pietryczuk et al. 2018); Clavatospora longibrachiata
(Gulis & Suberkropp 2003, Menéndez et al. 2012, Cornut et al. 2014, Raposeiro et al. 2018,
Pietryczuk et al. 2018); and Lunulospora curvula (Schoenlein-Crusius et al. 2009, Duarte et al.
2012, Cornut et al. 2014, Pietryczuk et al. 2018, Raposeiro et al. 2018). Some genera are speciose
with various novel taxa introduced from Thailand and China: Pleurotheciella, Canalisporium,
Chaetosphaeria, Tainosphaeria; see Luo et al. (2019) and Calabon et al. (2022) for species and
distribution. Though most of the present taxonomic studies incorporate molecular data in the
introduction of taxa, there are still taxa with uncertain placements due to lack of living cultures and
type sequences. Furthermore, most of the geographical data of freshwater Sordariomycetes are
mainly based on biodiversity studies using morphology and it is worth noting to take cautions when
dealing with distributions of freshwater fungi.
Freshwater Eurotiomycetes
Eurotiomycetes is a morphologically diverse group of ascomycetes. The class was established
by Eriksson & Winka (1997) and presently consists of five subclasses: Chaetothyriomycetidae,
Coryneliomycetidae, Cryptocaliciomycetidae, Eurotiomycetidae, Mycocaliciomycetidae, and
Sclerococcomycetidae (Geiser et al. 2006, Hibbett et al. 2007, Wood et al. 2016, Réblová et al.
2017, Prieto et al. 2021). The microcolonies of Eurotiomycetes formed under natural conditions are
morphologically similar leading to difficulty in accurately distinguishing species from one another
The incorporation of molecular techniques in fungal taxonomy makes the classification of fungi
more objective, accurate, and comprehensive, and significantly improves our understanding of the
phylogeny and evolution of freshwater Eurotiomycetes (Wood et al. 2016, Réblová et al. 2017, Luo
et al. 2019, Wijayawardene et al. 2020; Dong et al. 2020a, Prieto et al. 2021). The new taxa
published in recent years are supported by molecular data which enables a better understanding of
phylogenetic relationship of Eurotiomycetes. Based on the phylogenetic analysis of molecular data,
the specific taxonomic status of some taxa of Eurotiomycetes has been clarified (Wood et al. 2016,
Réblová et al. 2017, Prieto et al. 2021). Wijayawardene et al. (2020) provides an outline of the
classification of the Fungi and fungus-like taxa and accepted 3,994 published species in the
Eurotiomycetes.
Compared with the classes Dothideomycetes and Sordariomycetes, the number of freshwater
Eurotiomycetes species was small (217 species), accounting for only 5.4% of the total number (Fig.
9). But these species still play an important role in freshwater environments as saprophytes on
submerged wood, decaying leaves, branches, and plant debris in lakes and streams (Liu et al.
2015b, Dong et al. 2018, Liu et al. 2018, Wang et al. 2019). Eurotiomycetes are also known
growing on inundated rocks and pebbles (e.g., Verrucaria spp.) and isolated from sediments and
freshwater (e.g., drinking water, groundwater, tap water) (Iwatsu et al. 1991, de Hoog et al. 2011,
Crous et al. 2013, Calabon et al. 2022). Some groups are parasitic on or in the body of aquatic
animals, causing disease or death of the animals e.g., Aspergillus and Penicillium infecting organs
of ornamental and aquacultures fishes (Iqbal et al. 2012, Chauhan et al. 2014, Chauhan &
Bankhede 2013). In addition, some freshwater Eurotiomycetes species are also potential producers
of biologically active substances (Yamazaki et al. 2016, Rotinsulu et al. 2017, Abdel-Wahab et al.
2018, Steenwyk et al. 2020), and they play important roles in basic research, industry, and public
health.
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Herein, we used the ITS, LSU, and β-tubulin (tub2) sequence data available from
Eurotiomycetes in freshwater and other environments to construct a phylogenetic tree (Fig. 10).
The phylogenetic position and distribution of freshwater Eurotiomycetes species in the main orders
and families are summarized.
Figure 7 – Freshwater Sordariomycetes. a, b Lepteutypa aquatica (MFLU 15–0077).
c, d Fluminicola thailandensis (MFLU 15–0085). e, f Tainosphaeria obclavata (MFLU 18–1455).
g Distoseptispora cangshanensis (MFLU 18-0474). h Acrodictys liputii (MFLU 21–0034).
i, j Aquapteridospora fusiformis (MFLU 18–1601). k, l Pseudodactylaria aquatica (MFLU 21–
0037). m, n Sporoschisma longicatenatum (MFLU 21–0033). o, p Sporoschisma chiangraiense
(MFLU 21–0036). q, r Neospadicoides thailandica (MFLU 21–0032). s, t Chloridium gonytrichii
(MFLU 21–0026). u Sporidesmium nujiangense (HKAS 115795). v Cancellidium atrobrunneum
(MFLU 20–0429). Scale bars: a, i, m, q, s, t, v = 50 µm, b = 5 µm, c = 40 µm, d, f, j, l = 10 µm, e =
30 µm, g = 60 µm, h, k, n, p, r, s, u, = 20 µm, o = 100 µm.
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Figure 8 – Radial phylogenetic circular tree representation of freshwater Sordariomycetes.
Discussion of Eurotiomycetes
Herein, we constructed a phylogenetic tree using the available molecular sequences of most
of the currently known freshwater Eurotiomycetes species (Fig. 10). Our analysis shows that
freshwater Eurotiomycetes are dispersed in three subclasses, Chaetothyriomycetidae,
Eurotiomycetidae and Sclerococcomycetidae. The main orders include Chaetothyriales, Eurotiales
and Verrucariales, and a small number are distributed in Sclerococcales. Although many species
have molecular sequence data, the specific taxonomic status of some species is still unclear. For
example, in our phylogenetic tree, the two species of Anthopsis, A. catenata (CBS 492.81),
A. deltoidea (CBS 263.77) was placed in Chaetothyriomycetidae and Sclerococcomycetidae,
respectively, and with good support.
Chaetothyriales
Chaetothyriales is a diverse group, is renowned for containing so-called black yeasts and their
filamentous relatives, among which are numerous opportunistic agents of disease in humans and
cold-blooded vertebrates (Quan et al. 2020), including saprobes, pathogens, lichenized taxa, and
epilithic fungi (Gueidan et al. 2008, Chomnunti et al. 2012a, b, Réblová et al. 2013, Hubka et al.
2014). Currently, five families are accepted in this order, viz. Chaetothyriaceae, Cyphellophoraceae,
Epibryaceae, Herpotrichiellaceae, and Trichomeriaceae (Réblová et al. 2013, Gueidan et al. 2014,
Chomnunti et al. 2012a, Barr 1976, 1987, Wijayawardene et al. 2020). Most of the freshwater
species are found in the Herpotrichiellaceae.
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Figure 9 – Freshwater Eurotiomycetes spp. a–c Colonies of Pseudobactrodesmium spp. on natural
substrates. d, e Conidia with sheath of Pseudobactrodesmium spp. f, g Colonies of
Minimelanolocus spp. on natural substrates. h–j Conidia of Minimelanolocus spp. k Colonies of
Thysanorea papuana on natural substrates. l Conidia of Thysanorea papuana. Scale bars: a, k =
150 μm, b, f, g = 100 μm, a = 50 μm, d, e, h–j = 20 μm, l = 10 μm.
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Figure 10 – Multi-gene phylogeny analysis was performed using the maximum likelihood analysis
method, and the online tool RAxML-HPC2 on XSEDE (8.2.12) under the CIPRES website was
used for analysis. All parameters in the analysis were set by default. The type species are indicated
with “T” after the strain/specimen number, the species found in freshwater are indicated by “F”
after the strain/specimen number. Lectera nordwiniana (CNUFC HRS5-3 and CNUFC HRS5-3-1)
as the outgroup.
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Figure 10 – Continued.
Herpotrichiellaceae is the best-known family in the order Chaetothyriales, which has been
well supported by molecular data, as most of the known species were described from cultures
(Quan et al. 2020). Herpotrichiellaceae comprise morphologically diverse dematiaceous fungi that
include some saprophytic and pathogenic taxa isolated from humans and animals (Haase et al.
1999, de Hoog et al. 2000, Liu et al. 2015b, Wang et al. 2019, Dong et al. 2018). The freshwater
species in this family are mainly distributed in the genera Exophiala, Minimelanolocus and
Thysanorea, isolated from submerged woody substrates and water bodies (Liu et al. 2015b, Fiuza et
al. 2017, Dong et al. 2018, Wang et al. 2019, Wan et al. 2021).
Verrucariales
The order Verrucariales is dominated by lichenized ascomycetes, most members have the
typical thallus morphologies, including crustose, squamulose, foliose and rarely subfruticose thalli
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(Muggia et al. 2017). Verrucariales species are widely distributed, from marine to freshwater to
terrestrial habitats, from wet intertidal stones to submerged stones and wood in streams to dry rocks
and tree trunks (Brodo et al. 1997, Harada & Wang 2004, Sanders et al. 2004, Orange et al. 2009,
2012, 2013, Thüs et al. 2015, Lucban et al. 2019). Freshwater Verrucariales is mainly distributed in
the genera Thelidium and Verrucaria. Since the early classification, species of Verrucariales has
been reported based on morphological characteristics, the phylogenetic relationships between
members are unclear. For this reason, many researchers use molecular sequence data obtained from
cultures to study the diversity of Verrucariales and the phylogenetic relationship of individual
members (Muggia et al. 2010, Gueidan et al. 2007).
Verrucariaceae is a group of mainly lichenized ascomycetes from widely diverse habitats.
Species classified within Verrucariaceae grow mainly on rocks, either epilithically or endolithically
within the superficial layer of the rock (Gueidan et al. 2007). They are can also colonize other types
of substrates in dry environments: soils (Breuss 1996), wood or bark (Orange 1989, Breuss 1998),
mosses (Dőbbeler 1997); in aquatic habitats: boulders located in rivers (Keller 2000, Thüs 2002), or
marine intertidal and supralittoral zones of rocky shores (Harada & Wang 2004, Sanders et al.
2004). Currently, 50 species of Verrucariaceae have been reported in freshwater environments,
mainly growing on rocky surfaces in freshwater streams and rivers (McCarthy 1995, Harada &
Wang 2008, Harada 2012, Orange 2013, Krzewicka et al. 2017).
Eurotiales
Eurotiales is a relatively large order of Eurotiomycetidae, which is widely distributed in the
world. Its members are found in various environments: soil, food, drinking water and human and
animal organisms (Yu et al. 2005, Hedayati et al. 2007, Engelhart et al. 2009, do Nascimento
Barbosa et al. 2016, Pangging et al. 2019, Samson et al. 2010, 2019), and have a positive and
negative impact on human activities. Aspergillaceae and Trichocomaceae are the two larger
families in the order Eurotiales, containing 15 and 8 genera, respectively (Wijayawardene et al.
2020). Freshwater Eurotiales species include the genera Aspergillus, Penicillium, and Trichoderma
(Aspergillaceae and Trichocomaceae), and are mainly reported in the sediments and water bodies
of lakes, rivers, and ponds (Gupta & Kushwaha 2012, Heo et al. 2019, Pangging et al. 2019,
Piontelli et al. 2019, Mun et al. 2020). Many Eurotiales reported in freshwater environments are
also often reported from terrestrial habitats. Therefore, some scholars believe these to be runoff
from terrestrial habitats and not true freshwater fungi. For example, Aspergillus, Penicillium, and
Trichoderma species reported in freshwater environments are not considered true aquatic species
(Daniel et al. 2007, Lind et al. 2017, Nielsen et al. 2017). It is because they are everywhere, being
washed into streams or lakes from terrestrial habitats. Therefore, it is challenging to determine if all
Eurotiales species isolated from freshwater are truly active in this environment.
Sclerococcales
Réblová et al. (2017) introduced Sclerococcomycetidae, Sclerococcales, and
Sclerococcaceae, based on five loci (nucSSU, ITS, nucLSU, mitSSU, rpb1 and rpb2) phylogeny.
The latest revision of the classification of lichenicolous fungi by Diederich et al. (2018)
synonymized Sclerococcaceae as Dactylosporaceae since Sclerococcum and Dactylospora
represent a monophyletic group based on a two-locus (nuLSU and mtSSU) phylogeny.
Sclerococcum (1821) predates Dactylospora (1855) so the former was set as the type genus.
The order Sclerococcales currently contains one family and five genera, including some
lichenicolous, marine, and lignicolous species. Freshwater Sclerococcales species are rare:
Pseudobactrodesmium and Cylindroconidiis, with a total of four species, C. aquaticus, P.
aquaticum, P. chiangmaiensis, and P. stilboideum (Yu et al. 2018, Dong et al. 2020a, Boonmee et
al. 2021).
Leotiomycetes
The class Leotiomycetes was introduced by Eriksson & Winka (1997) and is often referred as
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the “inoperculate discomycetes”, because the traditional concept of Leotiomycetes only includes
apothecial ascomycetes with inoperculate, unitunicate asci that open by apical perforation or pore
to release their ascospores (Nannfeldt 1932, Korf 1973, Pfister & Kimbrough 2001). In recent
years, more molecular sequences have been used for classification of their taxonomy, and some
previous groups have been removed to establish a more natural system (Baral et al. 2015).
Freshwater Leotiomycetes mostly grow on submerged or floating substrates such as scums, spume,
cryptogamic plants, herbaceous and woody substrates (Ingold 1954, 1974, Magnes & Hafellner
1991, Tsui et al. 2000, Wong & Hyde 2001, Baschien et al. 2013) in various lotic and lentic
freshwater environments, like ponds, rainfall, melting ice, lakes, springs, swamps, rivers, and water
distribution system (Ingold 1954, Shearer & Crane 1986, Czeczuga & Orłowska 1999, Luo et al.
2004, Czeczuga et al. 2007, Grabińska-Loniewska et al. 2007, Raja et al. 2008, Funck et al. 2015).
Previous discoveries of new taxa and sexual morph association of freshwater Leotiomycetes were
solely based on morphological and culture observation (Ingold 1942, Beaton & Weste 1977,
Shearer 1993, Webster 1993, Webster et al. 1995). In recent years, advances in sequencing
technology have introduced deeper morpho-phylogenetic insights on novel species and holomorph
revelation of freshwater Leotiomycetes (Belliveau & Bärlocher 2005, Campbell et al. 2006, 2009,
Baschien et al. 2013, Duarte et al. 2015, Sri-indrasutdhi et al. 2015, Baudy et al. 2019). Currently,
Leotiomycetes includes 14 orders and 52 families (Wijayawardene et al. 2020). Members of
freshwater Leotiomycetes are mainly distributed in Helotiales (188) and Leotiomycetes families
incertae sedis (54 species), and a few species are distributed in Thelebolales (8 species),
Rhytismatales (4 species), and Lauriomycetales (3 species) (Calabon et al. 2022).
Lichinomycetes
Freshwater Lichinomycetes is a group of lichen-forming fungi, and little is known about their
occurrence in freshwater habitats (Jørgensen et al. 2007). These species form gelatinous lichen-like
symbioses with cyanobacteria, relatively small and grow on rocks and soil in moist or dry but
temporarily wet localities (Egea & Rowe 1988, Gilbert 1996, Thüs et al. 2014, Kantvilas 2018,
Gumboski et al. 2019). Currently, Lichinomycetes contains one order and three families, and the
members found in freshwater environments are distributed in the families Lichinaceae (22 species)
and Peltulaceae (3 species) (Wijayawardene et al. 2020, Calabon et al. 2022).
Orbiliomycetes
The class Orbiliomycetes comprises a large group of inoperculate discomycetes previously
included in Helotiales. Based on morphological and molecular phylogenetic data, Orbiliaceae was
raised to Orbiliomycetes, and currently contains only a single order, a single family, and 12 genera
(Eriksson et al. 2003, Wijayawardene et al. 2020). Members of freshwater Orbiliomycetes mostly
grow on submerged decaying wood, leaves, and spores in foam in freshwater streams, rivers, and
swamps (Ingold 1944, Marvanová & Marvan 1969, Karamchand 2009, da Silva & Gusmão 2015,
Fiuza et al. 2019). Currently, 14 species in ten genera, viz. Arthrobotrys, Hyalorbilia, Dactylella,
Dicranidion, Helicoon, Monacrosporium, Orbilia, Orbiliella, Trinacrium, Vermispora, are found in
freshwater environments (Calabon et al. 2022).
Pezizomycetes
The class of Pezizomycetes is commonly known as cup-fungi or operculate discomycetes and
is one of the earliest diverging lineages of Pezizomycotina along with Orbiliomycetes (Spatafora et
al. 2006, Schoch et al. 2009). Pezizomycetes taxa are characterized by asci that usually open by
rupturing to form a terminal or eccentric lid or operculum, although some hypogeous and
cleistothecial forms lack an operculum (Lumbsch et al. 2005, Hansen & Pfister 2006), and they
occur on a variety of substrates, including soil, wood, dung (Abbott & Currah 1997, Kirk et al.
2008, Cheraghian 2016, Ekanayaka et al. 2016, 2017). Pezizomycetes are rarely reported from
freshwater habitats. Jones et al. (2014a) recorded eight species from freshwater. Currently, 13
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species in four families (Ascobolaceae, Pezizaceae, Pyronemataceae, Rhizinaceae) from freshwater
habitats, all of which are found on woody substrates (Calabon et al. 2022).
Saccharomycetes
Saccharomycetes is a monophyletic lineage, comprise more than 1,200 species of yeasts
(Wijayawardene et al. 2020). Saccharomycetes yeasts are found in nearly all regions of the world,
including hot deserts, polar areas, in freshwater, in salt water, and in the atmosphere (Kurtzman et
al. 2015). Their growth is mainly saprotrophic, often in association with plants, animals, but some
members are important pathogens of humans (Mortimer & Polsinelli 1999, Vega & Black 2005,
Martins et al. 2014, Erdogan & Rao 2015, Kurtzman et al. 2015). Freshwater Saccharomycetes
species can grow on various substrates (water bodies, mud, sediments, and stones) in various
freshwater environments (i.e., wetlands, lakes, ponds, canals, polluted water, tapwater) and some
species are also isolated on the surface of animal bodies (Sláviková & Vadkertiová 1995, Khan et
al. 2002, de García et al. 2007, Morais et al. 2010, 2020, Fell et al. 2011, Biedunkiewicz et al. 2013,
Moubasher et al. 2018). Based on morphological and analytical phylogenetic analysis, 14 families
of one order, Saccharomycetales, were accepted in Saccharomycetes (Wijayawardene et al. 2020).
Some 150 Saccharomycetes species have been reported from freshwater habitats, and are mostly
distributed in 11 families, with 50 reported from Saccharomycetales genera incertae sedis (Calabon
et al. 2022).
Conclusion
From the reviews of Cai et al. (2014) and Shearer et al. (2014) to the recent class-level
phylogenetic analysis of Luo et al. (2019) and Dong et al. (2020b), our understanding of the
classification and interrelationships of freshwater ascomycetes has increased significantly.
Freshwater fungal taxonomists recently introduced novel taxa with cultures and sequence data, and
integrated protein-coding loci in the phylogenies, leading towards a natural classification of
freshwater fungi. However, gaps remain in the molecular phylogeny of freshwater ascomycetes
mainly to the lack in availability of living cultures and molecular data, leading to uncertain
taxonomic placements and unknown evolutionary relationships, see Calabon et al. (2022) for list of
freshwater fungal taxa in the Ascomycota incertae sedis. Further exploration of freshwater habitats
with comprehensive sampling of various substrates, herbarium material observations, and
generation of nuclear and protein-coding sequence data will help in the further delineation of
freshwater ascomycetes and better resolution of their phylogenetic relationship.
Biology of Freshwater Basal Fungi
Introduction
Early branching fungi (basal fungi) are classified in several phyla including Aphelidiomycota,
Blastocladiomycota, Chytridiomycota, Monoblepharomycota, Mucoromycota, and Rozellomycota
(Hibbett et al. 2007, Tedersoo et al. 2018, Adl et al. 2019, Naranjo-Ortiz & Gabaldón 2019, James
et al. 2020, Voigt et al. 2021). They consist of two major lineages namely the zoosporic and
zygosporic fungi (Hibbett et al. 2007, Voigt et al. 2021). Zoosporic fungi refer to basal lineages that
produce motile spores called zoospores, while the spores of zygosporic fungi are non-motile
(O’Donnell et al. 2001, Powell & Letcher 2014a, Longcore & Simmons 2020, Voigt et al. 2021).
Basal fungi have been primarily discovered from freshwater, terrestrial, and marine ecosystems
(Abdel-Wahab et al. 2014, Longcore & Simmons 2020). The ecological roles of fungi in the
freshwater environment are pivotal and diverse, aiding in the overall function of that habitat.
Freshwater fungal species complete at least one part of their life cycle in freshwater, distribute
propagules in or above water, or use any resource of predominantly aquatic or semi-aquatic nature
as substratum (Tsui et al. 2016, Calabon et al. 2020a). Examples of freshwater habitats are streams,
ditches, canals, lakes, peats, and swamps (Tsui et al. 2016).
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Among basal fungal lineages, zoosporic taxa, especially Chytridiomycota, are the most
common in freshwater ecosystems. The currently reported diversity of zygosporic fungi (e.g.,
Mucoromycota and Mortierellomycota) in freshwater is significantly lower in terms of described
species or sequences generated from high throughput sequencing (Lepère et al. 2019). The roles of
basal fungi in freshwater ecology have been largely overlooked. Saprobic freshwater fungi assist in
the breakdown of allochthonous organic material, like leaves and twigs, which results in the
provision of 99% of total energy input in surface water (Ittner et al. 2018). Parasitic zoosporic
species infect numerous phytoplankton groups (diatoms, green algae, cyanobacteria, and
dinoflagellates) and invertebrates (opisthokonts), and they regulate the population density of their
hosts (Kagami et al. 2012, Ishida et al. 2015). Planktonic fungi are also indicators of water quality
(Ishida et al. 2015, Chen et al. 2018). The parasitic chytrid fungi, namely Batrachochytrium
dendrobatidis and B. salamandrivorans, have devastating effects on amphibians (Martel et al.
2013, Van Rooij et al. 2015). Batrachochytrids have caused mass declines and the near extinction
of several species of amphibians (Fisher & Garner 2020). Catenaria, Coelomomyces and Olpidium
are common parasites of freshwater and terrestrial invertebrates worldwide (Whisler 1985, Barron
2004). Most invertebrates that come into contact with freshwater at any stage of their life cycle,
whether as larvae or adults, can be parasitized by zoosporic fungi (Gleason et al. 2010a). Given the
importance of these groups of fungi, more studies on their diversity, and ecological significance in
freshwater habitats are needed.
This entry focuses on the biology of freshwater basal fungi. It aims to provide a brief
overview on the various aspects of basal fungi in freshwater habitats. However, it is not intended to
cover all aspects within this discipline or provide a full literary review.
Taxonomic classification of basal fungi
Fungi colonizing freshwater habitats are scattered across various phyla including the early
branching fungal groups. The taxonomic classification of early diverging lineages of fungi has been
revised in the last few years (James et al. 2006a, b, 2020, Hibbett et al. 2007, Spatafora et al. 2016,
Choi & Kim 2017, Tedersoo et al. 2018, Naranjo-Ortiz & Gabaldón 2019). Previously, all
zoosporic lineages were placed in the phylum Chytridiomycota, while zygosporic ones comprised
the now invalid phylum “Zygomycota” (Barr 2001, James et al. 2006b, Hibbett et al. 2007, James
et al. 2020). The advent and increased resolution of molecular tools has allowed the circumscription
of monophyletic groups, such as Blastocladiomycota, Chytridiomycota, and
Neocallimastigomycota. These groups are well supported not only by molecular evidence, but also
cellular ultrastructure such as the structure of the mitotic apparatus (James et al. 2006a, b, Hibbett
et al. 2007, Powell & Letcher 2014a, Longcore & Simmons 2020). The artificial phylum
Zygomycota was broken down to several phyla such as Mucoromycota, and Mortierellomycota
(James et al. 2006b, Hibbett et al. 2007, Spatafora et al. 2016). The classification of Zygomycota
was traditionally based on morphological characteristics, but molecular biology has revolutionized
the taxonomic classification of this group (Hibbett et al. 2007, Spatafora et al. 2016). Zygosporic
taxa have been grouped into new phyla, classes, orders, and families (Voigt et al. 2021). In
freshwater, members of Aphelidiomycota, Blastocladiomycota, Chytridiomycota,
Entomophthoromycota, Monoblepharomycota, Mortierellomycota, Mucoromycota, Olpidiomycota,
and Zoopagomycota have been found. Table 2 lists the genera that have been found in freshwater
habitats.
Chytridiomycota is the most species-rich zoosporic phylum (Blaalid & Khomich 2021).
Members of this phylum (or in some classification schemes, the class Chytridiomycetes) are
commonly referred to as chytrids or chytrid fungi (Powell & Letcher 2014a, Fisher & Garner
2020). Tedersoo et al. (2018a) proposed nine classes and ten orders, Naranjo-Ortiz & Gabaldón
(2019) accepted three classes and seven orders, and James et al. (2020) accepted 14 orders.
Visualization and proper identification of chytrids is difficult due to their small size and lack of
distinct morphological characters. Due to these reasons, chytrids are often misidentified as protists
(Blaalid & Khomich 2021). The zoospores (zoosporangiospores) of chytrids swim with a
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characteristic abrupt hopping and darting pattern assisted by a single posteriorly directed whiplash
flagellum (Powell & Letcher 2014a, Longcore & Simmons 2020). The motile spores are unwalled
but have a carbohydrate coat that protects them against desiccation (Longcore & Simmons 2020).
Chytrids, and zoosporic fungi in general, were referred to as aquatic fungi, however these have also
been reported in terrestrial habitats (Powell 2017a). To date, very few basal taxa have been isolated
from marine habitats (Abdel-Wahab et al. 2014, Jones et al. 2019, Hassett et al. 2020b). High
throughput sequencing (HTS) studies have shown a high diversity of some zoosporic lineages
including chytrids in oceanic networks (Abdel-Wahab et al. 2014, Comeau et al. 2016). However,
their diversity in marine habitats is poorly understood despite their predicted ecological
significance and function. The various distinct methodologies that are required to isolate zoosporic
organisms as opposed to those conventionally used for filamentous fungi, poses a constraint for
their study. In environmental surveys, chytrid sequences are seemingly difficult to amplify possibly
due to primer bias, which might partly explain their underreporting (Grossart et al. 2019, Blaalid &
Khomich 2021). These issues highlight that the taxonomy and systematics of these fungi is far from
resolved.
Rozellomycota (also known as Cryptomycota) and Aphelidiomycota consist of endoparasitic
taxa placing at a basal position in the fungal tree of life. Both form the Opisthosporidia with
ongoing debates on whether they are indeed fungi (Karpov et al. 2014b, Tedersoo et al. 2018,
Wijayawardene et al. 2018, 2020, Adl et al. 2019, Naranjo-Ortiz & Gabaldón 2019, James et al.
2020). Aphelids and rozellids have several similarities. Both are endoparasites that feed by
phagocytosis (Letcher & Powell 2019). Aphelids parasitize various green algae and diatoms and
currently comprise a single class, order, and family with four genera (Letcher & Powell 2019,
Wijayawardene et al. 2020). Rozellomycota species are endoparasites of oomycetes and
opisthokonts, such as chytrids, Bryozoa, fish and invertebrates, such as Daphnia species, and
amphipods. This group (including Microsporidia) accounts for 105 genera, 221 species in four
orders and 21 families of freshwater taxa, thus a significant group of basal fungi (Calabon et al.
2022). All described species can produce a chitinous cell wall, yet they grow as naked protoplasts
inside their host. They vary greatly in morphology where some species are fungus-like (Rozella)
and infect their host during their motile stage (flagellated), while others lack flagella and infection
occurs from spores through a polar filament (Weiss 2001, Franzen 2004, Corsaro et al. 2014,
Quandt et al. 2017). Data on the distribution of these members comes mostly from environmental
surveys. Most aphelids and rozellids have been isolated or described from freshwater habitats such
as streams, ditches, and lakes (Calabon et al. 2022). High throughput sequencing has also unveiled
a plethora of diversity from freshwater ecosystems (Rojas-Jimenez et al. 2017).
Microsporidia comprises a distinct lineage of unicellular eukaryotes, which groups as sister
to, or within Rozellomycota (Tedersoo et al. 2018, Naranjo-Ortiz & Gabaldón 2019, James et al.
2020, Wijayawardene et al. 2020). Microsporidia are obligate intracellular eukaryotic parasites that
use the polar tube, a unique invasion apparatus, to infect hosts (Weiss 2001, Franzen 2004). These
organisms inhabit many ecosystems including freshwater. They infect both vertebrates and
invertebrates and have been reported from a broad range of hosts from protists to mammals,
including humans, to arthropods (Desportes et al. 1985, Call et al. 1998, Meissner et al. 2012, Han
et al. 2020). In freshwater, microsporidia infect fish, crustaceans, amphipods, Bryozoa and other
fauna (Jones et al. 2019, Drozdova et al. 2020, Liu et al. 2020, Weng et al. 2022). The intracellular
lifestyle of microsporidia has led to compact genomes, highly reduced mitochondria, which are
referred to as mitosomes, and presence of many types of transporters that enhance uptake of
compounds from the host (Tsaousis et al. 2008, Williams et al. 2008, Nakjang et al. 2013, Bass et
al. 2018, Park & Poulin 2021).
Zygosporic fungi are considered as the basal terrestrial lineages that most likely evolved from
flagellate, aquatic ancestors (Hibbett et al. 2007, Hoffmann et al. 2011). Currently, scarce data are
available on the distribution of these fungi in aquatic habitats. Only few species from the phyla
Entomophthoromycota, Monoblepharomycota, Mortierellomycota, Mucoromycota, and
Zoopagomycota have been found in freshwater. This could be due to several factors such as low
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number of studies targeting these fungi, difficulty in isolating them and primer bias during HTS.
Alternatively, it is possible that freshwater might not be the ideal habitat for these fungi. Among the
zygosporic fungi, Mucoromycota seems to be more prevalent in freshwater than other zygosporic
taxa.
Diversity of basal fungi in freshwater ecosystems
Freshwater accounts for less than 1% of the Earth’s surface and comprises 2.5% of all water
on the planet. Fungal taxa in freshwater habitats are important components of microbial
communities of water columns and sediments in both lentic and lotic systems (Sutcliffe et al.
2018). Zoosporic fungi, especially chytrids, are the most speciose representatives in aquatic
ecosystems (Gleason et al. 2017). The worldwide distribution of zygosporic fungi in freshwater
habitats is comparatively poorly documented. The study of freshwater zoosporic fungi dates to
1960s, but studies on their diversity, quantitative abundance and especially their interaction with
other microorganisms are scarce (Gleason et al. 2017, Grossart et al. 2019).
High throughput sequencing studies have revealed a high biodiversity and predominance of
unexplored zoosporic taxa in freshwater habitats. In ice-covered lakes of Antarctica, Rozellomycota
and Chytridiomycota are the most abundant fungal phyla (Comeau et al. 2016). In the temperate
Lake Tahoe, in the United States and freshwater Arctic habitats, Chytridiomycota-like sequences
seemingly representing novel lineages dominated the fungal diversity (Comeau et al. 2016, Gleason
et al. 2017). Pelagic zones of lakes also have high diversity of undescribed zoosporic fungi
(Lefèvre et al. 2010). However, morphological studies involving light microscopy have revealed
the opposite (Sime-Ngando et al. 2011). Furthermore, not all environmental surveys of freshwater
habitats show a dominance of zoosporic fungi, several indicate that Dikarya is the predominant
group (Shearer et al. 2007, Lepère et al. 2019). For example, in a study of 25 lakes and four rivers,
Dikarya fungi (Ascomycota and Basidiomycota) represented the most OTU-rich groups (Lepère et
al. 2019). Nonetheless, even though Dikarya were the most abundant, basal groups such as
Rozellomycota and Chytridiomycota were also found. The results of these studies indicate the
presence of undocumented diversity of these fungi in various habitats.
An increasing number of freshwater zygosporic species are being described with several
being novel taxa. Previously, known species from freshwater habitats included members of Erynia
and Acaulopage, which are parasites of aquatic insects (Goh & Hyde 1996a). The distribution and
diversity studies on these taxa have now expanded. Aquatic zygosporic species include
Aquamortierella, Mortierella, Gryganskiella, Endogone, Gilbertella, Cunninghamella, Absidia,
Gongronella, Rhizomucor, Actinomucor and Mucor (Table 2) (Schell et al. 2011, Nguyen & Lee
2016, Nguyen et al. 2017a, b, 2019, Moubasher et al. 2018, Crous et al. 2020, Vandepol et al.
2020).
Ecology of freshwater basal fungi
Basal fungi contribute to various processes in freshwater habitats, but only scant studies
exist. They are mainly involved in water purification and recycling of organic matter that is used by
other organisms (such as invertebrates) (Voronin 2008). Furthermore, basal fungi aid in
mineralization of organic matter, and regulation of insect population abundance. In freshwater
ecosystems, these fungi have various life modes such as parasites or saprobes. Saprobic freshwater
basal fungi survive on dead organic matter found in waterbodies. Zoosporic fungi are usually
isolated by means of baits, such as snake skin, cellophane, exoskeleton of shrimp, and pollen.
Examples of freshwater saprobes are Homolaphlyctis polyrhiza, and Synchytrium microbalum
(Longcore et al. 2012, 2016). Parasites depend on their hosts for food, such as Collimyces mutans
and Staurastromyces oculus, and parasitize Microglena coccifera and Staurastrum sp. (Van den
Wyngaert et al. 2017, Seto & Degawa 2018a). Being heterotrophs, these fungi consume live
organic matter, but they primarily process dead organic matter hence are involved in the formation
of the structural-functional organization of aquatic biocenoses (Voronin 2008). Basal fungi are
ecological competitors, can tolerate stress and are ruderals (Dix & Webster 1995). These fungi
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thrive in environments that have a continuous supply of substrates/nutrients and with optimal
conditions, such as temperature (Sparrow 1960). They have commonly been isolated from
terrestrial environments such as soil and sometimes their diversity is picked up in hydrothermal
vents (Le Calvez et al. 2009). Basal fungi can survive harsh habitats due to the production of
resistant structures, such as zoospore cysts and resistant sporangia. Depending on the environment,
these fungi use optimal strategies to occupy several ecological niches (Gleason et al. 2010a, b,
2012).
Parasites of freshwater phytoplankton
Recently, there has been a surge of research in phytoplankton parasitism by zoosporic fungi.
Chytrids and aphelids comprise common parasites of phytoplankton (Kagami et al. 2007, Sime-
Ngando et al. 2011, Lepère et al. 2019, Song et al. 2021). Studies on chytrids parasitizing algae
have been the main focus, while aphelids research is still lagging. Phytoplankton is responsible for
a large proportion of primary production and crucial for the survival of food webs (Winder &
Schindler 2004). Due to the importance of phytoplankton in aquatic ecosystems, the ecological
roles of both groups of fungal parasites are of interest (Jephcott et al. 2017). The increased
encroachment of freshwater habitats and surrounding environment may lead to disturbances in the
relationship and dynamics between organisms. For instance, invasive species may move afield into
a new ecosystem rapidly and disturb the delicate balance between a host and its parasite. Hence, an
understanding of host parasite relationships is crucial to recognize ecosystems under stress
(Jephcott et al. 2017). In this context, basal freshwater fungal parasites may be used as such
indicators.
In freshwater ecosystems, where phytoplankton and zooplankton abound, fungal parasitism
has an important role (Voronin 2008). Fungal parasitism affects and controls the population density
of planktonic species and influences competitive interactions between hosts and other species.
Selective parasitism is a decisive factor in the seasonal succession and food web relationships
(Canter & Lund 1951). For example, fungal parasitism of one algal species favorably affects the
development of other phytoplankters; when parasites infect one algal species, its resources may
become available to other phytoplankton species thereby increasing their population density
(Paterson 1960).
Chytrid parasitism has been documented in several species of diatoms, dinoflagellates and
cyanobacteria in various habitats. Chytrids are involved in nutrient cycling and comprise vital
components of the mycoloop. The mycoloop is defined as a process that alters carbon flow in
aquatic habitats. Specifically, chytrids infect inedible phytoplankton, such as Asterionella, or
cyanobacteria that are poorly edible to other organisms, use the nutrients of their host to grow and
produce zoospores that have high content of fatty acids and sterol (Voigt et al. 2021). When
zooplankton (e.g., Daphnia) grazes on the nutrient-rich zoospores, carbon that would have
otherwise been inaccessible is recycled (Kagami et al. 2007).
The extent of host-specificity of chytrids has not been fully explored to date. Chytrids are
usually considered as highly host specific parasites, however this is only speculative. Chytrids with
narrow and broader host ranges have been reported (Gromov et al. 1999). Parasites can be
facultative or obligate. Among chytrids, facultative parasitism has been observed in Dinochytrium
kinnereticum. This fungus is parasitic on weakened cells of the dinoflagellate Peridinium
gatunense, but is also saprobic on pollen. In this type of parasitism, the parasite can infect and
reproduce on the living host but is also able to exploit other nutrient sources or hosts (Leshem et al.
2016). One example of obligate parasitism among chytrids is Rhizophydium planktonicum, which
parasitizes the diatom Asterionella formosa (Canter & Jaworski 1978).
Infection by chytrids encompasses several phases (Frenken et al. 2017). The initial attraction of
zoospores to the host is most likely mediated by chemotaxis. Upon encounter, the zoospores encyst,
retract their flagellum, produce a cell wall, and a germ tube, which is used to penetrate the host cell.
However, depending on the host, chytrids use a different feature of the host cell to invade (Powell
1994, Gromov et al. 1999).
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Table 2 Classification of freshwater basal fungi.
Phylum
Class
Order
Family
Genus
References
Aphelidiomycota
Aphelidiomycetes
Aphelidiales
Aphelidiaceae
Amoeboaphelidium
Letcher et al. (2015b), Letcher &
Powell (2019)
Aphelidium
Gromov & Mamkaeva (1975),
Tcvetkova et al. (2019)
Paraphelidium
Karpov et al. (2017b, c)
Blastocladiomycota
Blastocladiomycetes
Blastocladiales
Blastocladiaceae
Allomyces
Sparrow (1964), Ali & Abdel-
Raheem (2003), Powell (2017a)
Blastocladia
Dasgupta & John (1988a), El-
Hissy et al. (1996), Steciow &
Marano (2006)
Blastocladiopsis
Hsiao (1969), Waterhouse (1942),
Czeczuga et al. (1990)
Catenariaceae
Catenophlyctis
Karling (1968b)
Nematoceromyces
Martin (1978)
Paraphysodermataceae
Paraphysoderma
James et al. (2012)
Catenomycetales
Catenomycetaceae
Catenomyces
Hanson (1944b)
Coelomomycetaceae
Coelomomyces
Porter et al. (2011)
Coelomycidium
Weiser (1984)
Microallomyces
Emerson & Robertson (1974)
Chytridiomycota
Chytridiomycetes
Chytridiales
Asterophlyctaceae
Asterophlyctis
Letcher et al. (2018)
Wheelerophlyctis
Letcher et al. (2018)
Chytridiaceae
Chytridium
Nowakowski (1876), Rooney &
McKnight (1972), Dasgupta &
John (1988b)
Dendrochytridium
Powell & Letcher (2014b)
Dinochytrium
Leshem et al. (2016), Hassett et al.
(2020a)
Irineochytrium
Dogma (1969)
Polyphlyctis
Letcher & Powell (2018)
Zopfochytrium
Powell et al. (2018)
Chytriomycetaceae
Avachytrium
Vélez et al. (2013)
Chytriomyces
Karling (1945), Willoughby &
Townley (1961), Letcher & Powell
(2002), Davis et al. (2019)
223
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Entophlyctis
Sparrow (1943), Haskins (1946),
Karling (1968a), Dasgupta & John
(1988b), Shin et al. (2001)
Fayochytriomyces
Davis et al. (2015)
Obelidium
Karling (1967), Blackwell et al.
(2012)
Odontochytrium
Vélez et al. (2013)
Pendulichytrium
Seto & Degawa (2018b)
Physocladia
Davis et al. (2019)
Podochytrium
Sparrow (1951), Willoughby &
Townley (1961)
Rhizidium
Karling (1944), Dayal & Kirin
(1981)
Rhizoclosmatium
Paterson (1967), Davis et al.
(2019)
Siphonaria
Karling (1967), Dogma (1976)
Phlyctochytriaceae
Phlyctochytrium
Ajello (1945), Johnson (1969)
Phlyctorhizaceae
Phlyctorhiza
Hanson (1946)
Pseudorhizidiaceae
Pseudorhizidium
Davis et al. (2019)
Scherffeliomycetaceae
Scherffeliomyces
Sparrow (1938)
Zygorhizidiales
Zygorhizidiaceae
Zygorhizidium
Canter (1963), Seto et al. (2020)
Zygophlyctidales
Zygophlyctidaceae
Zygophlyctis
Paterson (1958), Seto et al. (2020)
Delfinachytrium
Vélez et al. (2013)
Polyphagales
Polyphagaceae
Polyphagus
Bartsch (1945), Johns (1964),
Doweld (2014)
Sparrowiaceae
Sparrowia
Papa & Cruz-Papa (2020)
Chytridiomycetes
genera incertae sedis
Bertramia
Weiser & McCauley (1974)
Blyttiomyces
Sparrow & Barr (1955), Dasgupta
& John (1988b), Blackwell et al.
(2011)
Canteria
Karling (1971)
Dangeardia
Canter (1946), Geitler (1963),
Batko (1970)
224
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Dangeardiana
Johns (1956), Valkanov (1964)
Dictyomorpha
Mullins (1961), Sarkar & Dayal
(1988)
Ichthyochytrium
Korting (1983)
Loborhiza
Hanson (1944a)
Macrochytrium
Crasemann (1954)
Mitochytridium
Dangeard (1911), Hassan (1982)
Mucophilus
Červinka et al. (1974)
Perolpidium
Canter (1949b)
Pseudopileum
Canter (1963)
Rhizosiphon
Canter & Lund (1968), Gleason et
al. (2014)
Rhopalophlyctis
Karling (1945), Davis et al. (2019)
Saccomyces
Demchenko (2019)
Septosperma
Seymour (1971), Dogma (1974)
Sorokinocystis
Saccardo (1888)
Sporophlyctidium
Sparrow (1978)
Sporophlyctis
Sparrow (1960)
Truittella
Karling (1949)
Volvorax
Van den Wyngaert et al. (2018)
Zygochytrium
Sorokin (1874)
Cladochytriomycetes
Cladochytriales
Catenochytridiaceae
Catenochytridium
Willoughby & Townley (1961)
Cladochytriaceae
Cladochytrium
Czeczuga et al. (2005)
Endochytriaceae
Diplophlyctis
Willoughby & Townley (1961),
Dogma (1976)
Endochytrium
Karling (1941)
Nowakowskiellaceae
Nowakowskiella
Jerônimo et al. (2019)
Septochytriaceae
Septochytrium
Karling (1942), Johanson (1943)
Lobulomycetes
Lobulomycetales
Lobulomycetaceae
Algomyces
Van den Wyngaert et al. (2018)
Clydaea
Simmons et al. (2009)
Lobulomyces
Simmons et al. (2009), Davis et al.
(2018)
Mesochytriomycetes
Gromochytriales
Gromochytriaceae
Gromochytrium
Karpov et al. (2014a)
Mesochytriales
Mesochytriaceae
Mesochytrium
Gromov et al. (2000)
225
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Polychytriomycetes
Polychytriales
Arkayaceae
Arkaya
Longcore & Simmons (2012)
Polychytriaceae
Karlingiomyces
Blackwell et al. (2004)
Lacustromyces
Longcore (1993)
Neokarlingia
Longcore & Simmons (2012)
Polychytrium
Ajello (1945)
Rhizophydiomycetes
Rhizophydiales
Alphamycetaceae
Alphamyces
Canter (1961), Letcher et al.
(2012), Davis et al. (2018)
Betamyces
Letcher et al. (2012)
Angulomycetaceae
Angulomyces
Letcher et al. (2008b)
Aquamycetaceae
Aquamyces
Letcher et al. (2008b)
Collimycetaceae
Collimyces
Seto & Degawa (2018a)
Coralloidiomycetaceae
Coralloidiomyces
Letcher et al. (2008a)
Globomycetaceae
Globomyces
Sparrow (1952), Letcher et al.
(2008b)
Urceomyces
Letcher et al. (2008b)
Gorgonomycetaceae
Gorgonomyces
Letcher et al. (2008b)
Halomycetaceae
Halomyces
Letcher et al. (2015a)
Paranamyces
Letcher et al. (2015a)
Ulkenomyces
Letcher et al. (2015a)
Kappamycetaceae
Kappamyces
Letcher & Powell (2005)
Pateramycetaceae
Pateramyces
Letcher et al. (2008b)
Protrudomycetaceae
Protrudomyces
Letcher et al. (2008b)
Rhizophydiaceae
Rhizophydium
Dayal & Kirin (1981),
Dasgupta & John (1988b),
Gromov et al. (1999),
Davis et al. (2018, 2019)
Staurastromycetaceae
Staurastromyces
Van den Wyngaert et al. (2017)
Homolaphlyctis
Longcore et al. (2012)
Terramycetaceae
Boothiomyces
Davis et al. (2016), Jerônimo &
Pires-Zottarelli (2020)
Rhizophlyctidomycetes
Rhizophlyctidales
Borealophlyctidaceae
Borealophlyctis
Davis et al. (2016)
Rhizophlyctidaceae
Rhizophlyctis
Sparrow (1952), Willoughby &
Townley (1961), Dasgupta & John
(1988b)
226
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Spizellomycetes
Spizellomycetales
Spizellomycetaceae
Karlingia
Hassan (1983), Czeczuga et al.
(1990)
Spizellomyces
Wakefield et al. (2010)
Synchytriomycetes
Synchytriales
Synchytriaceae
Endodesmidium
Canter (1949b)
Synchytrium
Canter (1949b), Longcore et al.
(2016)
Micromyces
Roberts (1953), Davis et al. (2019)
Chytridiomycota genera
incertae sedis
Achlyella
Czeczuga & Muszynska (2001)
Achlyogeton
Czeczuga & Muszyńska (1993)
Entomophthoromycota
Entomophthoromycetes
Entomophthorales
Ancylistaceae
Ancylistes
Davis et al. (2019)
Conidiobolus
Sparrow (1952)
Entomophthoraceae
Erynia
Sridhar & Kaveriappa (1992),
Voglmayr (1996)
Zoophthora
Davis et al. (2019)
Monoblepharomycota
Hyaloraphidiomycetes
Hyaloraphidiales
Hyaloraphidiaceae
Hyaloraphidium
Ustinova et al. (2000)
Monoblepharidomycetes
Monoblepharidales
Gonapodyaceae
Gonapodya
Thaxter (1895), Waterhouse
(1942)
Harpochytriaceae
Harpochytrium
Jane (1946), Schumacher &
Whitford (1961)
Monoblepharidaceae
Monoblepharis
Lagerheim (1900), Perrott (1957),
Sparrow (1960)
Oedogoniomycetaceae
Oedogoniomyces
Kobayasi & Ôkubo (1954)
Telasphaerulaceae
Telasphaerula
Karpov et al. (2017a)
Sanchytriomycetes
Sanchytriales
Sanchytriaceae
Amoeboradix
Karpov et al. (2018)
Sanchytrium
Karpov et al. (2017a)
Mortierellomycota
Mortierellomycetes
Mortierellales
Mortierellaceae
Aquamortierella
Vandepol et al. (2020)
Gryganskiella
Vandepol et al. (2020)
Mortierella
Hyde et al. (2016b), Nguyen &
Lee (2016), Nguyen et al. (2019)
Mucoromycota
Endogonomycetes
Endogonales
Endogonaceae
Endogone
Sparrow (1952)
Mucoromycetes
Mucorales
Choanephoraceae
Gilbertella
Lee et al. (2018)
Cunninghamellaceae
Absidia
Moubasher et al. (2018)
Cunninghamella
Nguyen et al. (2017a)
227
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Gongronella
Crous et al. (2020)
Lichtheimiaceae
Rhizomucor
Schell et al. (2011)
Mucoraceae
Actinomucor
Nguyen et al. (2017b)
Mucor
Nguyen et al. (2020), Magray et al.
(2020)
Rhizopodaceae
Rhizopus
Gonçalves et al. (2006)
Syncephalastraceae
Syncephalastrum
El-Morsy et al. (2013)
Olpidiomycota
Olpidiomycetes
Olpidiales
Olpidiaceae
Olpidium
Canter (1949a), Sparrow (1957),
Czeczuga et al. (1990)
Zoopagomycota
Zoopagomycetes
Zoopagales
Zoopagaceae
Acaulopage
Voglmayr (1996)
Zoophagus
Karling (1966), Davis et al. (2019)
Rozellomycota
Microsporidea
Amblyosporida
Amblyosporidae
Amblyospora
Andreadis et al. (2012, 2018)
Becnelia
Tonka & Weiser (2000)
Crepidulospora
Simakova et al. (2004)
Culicospora
Weiser & Prasertphon (1982)
Culicosporella
Hazard et al. (1984)
Dimeiospora
Simakova et al. (2003)
Hyalinocysta
Andreadis & Vossbrinck (2002)
Intrapredatorus
Chen et al. (1998)
Novothelohania
Andreadis et al. (2012)
Parathelohania
Andreadis et al. (2012)
Trichoctosporea
Andreadis et al. (2012)
Tricornia
Pell & Canning (1992)
Caudosporidae
Binucleospora
Bronnvall & Larsson (1995)
Caudospora
Vávra & Undeen (1981)
Flabelliforma
Bronnvall & Larsson (2001)
Neoflabelliforma
Morris & Freeman (2010)
Scipionospora
Bylén & Larsson (1996)
Gurleyidae
Agglomerata
Sokolova et al. (2016), Weng et al.
(2020)
Binucleata
Larsson & Voronin (2000)
Conglomerata
Vávra et al. (2018)
Episeptum
Hyliš et al. (2007)
Gurleya
Friedrich et al. (1996)
228
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Lanatospora
Vávra et al. (2016b)
Larssonia
Vidtmann (1993)
Marssoniella
Vávra et al. (2005)
Norlevinea
Vávra (1984)
Paraepiseptum
Hyliš et al. (2007)
Pseudoberwaldia
Vávra et al. (2019)
Senoma
Simakova et al. (2005)
Zelenkaia
Hyliš et al. (2013)
Amblyosporida genera
incertae sedis
Alfvenia
Sokolova et al. (2016)
Takaokaspora
Andreadis et al. (2013)
Trichotuzetia
Vávra et al. (1997)
Glugeida
Glugeidae
Alloglugea
Paperna & Lainson (1995)
Amazonspora
Azevedo & Matos (2003)
Cambaraspora
Bojko et al. (2020b)
Glugea
Ward et al. (2005), Minter (2019)
Loma
Casal et al. (2009)
Pseudoloma
Ramsay et al. (2010)
Spragueidae
Apotaspora
Sokolova & Overstreet (2018)
Microgemma
Ralphs & Matthews (1986)
Potaspora
Ding et al. (2016)
Pseudokabatana
Liu et al. (2019)
Thelohaniidae
Cucumispora
Bojko et al. (2015)
Napamichum
Larsson (1990a)
Nudispora
Larsson (1990b)
Thelohania
Pretto et al. (2018)
Unikaryonidae
Dictyocoela
Terry et al. (2004)
Unikaryon
Voronin (1977, 1999)
Glugeida genus
incertae sedis
Triwangia
Wang et al. (2013a)
Neopereziida
Berwaldiidae
Berwaldia
Simakova et al. (2018)
Fibrillanosema
Galbreath Slothouber et al. (2004)
Neopereziidae
Bacillidium
Morris et al. (2005b)
Bryonosema
Canning et al. (2002)
229
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Neoperezia
Issi et al. (2012)
Pseudonosema
Canning et al. (2002)
Schroedera
Morris & Adams (2002), Morris et
al. (2005a)
Trichonosema
Desser et al. (2004)
Neopereziida genera
incertae sedis
Janacekia
Weng et al. (2021)
Systenostrema
Sokolova et al. (2006)
Nosematida
Enterocytozoonidae
Paranucleospora
Nylund et al. (2010)
Mrazekiidae
Helmichia
Tokarev et al. (2012)
Jirovecia
Liu et al. (2020)
Mrazekia
Larsson et al. (1993)
Nosematidae
Nosema
Terry et al. (2004)
Vairimorpha
Pretto et al. (2018)
Ordosporidae
Ordospora
Larsson et al. (1997)
Nosematida genera
incertae sedis
Anisofilariata
Tokarev et al. (2010b)
Crispospora
Tokarev et al. (2010a)
Enterocytospora
Jiang et al. (2020)
Glugoides
Larsson et al. (1996)
Microsporidea
families incertae
sedis
Cougourdellidae
Cougourdella
Larsson (1989)
Duboscqiidae
Duboscqia
Larsson & Yan (1988)
Tardivesicula
Larsson & Bylén (1992)
Trichoduboscqia
Batson (1982)
Fibrillasporidae
Fibrillaspora
Simakova et al. (2018)
Golbergiidae
Krishtalia
Kilochitskii (1997)
Microfilidae
Microfilum
Matos & Azevedo (2004)
Neonosemoidiidae
Neonosemoides
Faye et al. (1991)
Pleistophoridae
Heterosporis
Phelps et al. (2015), Tomamichel
et al. (2018)
Ovipleistophora
Lovy & Friend (2017), Bojko et al.
(2020a)
230
Table 2 Continued.
Phylum
Class
Order
Family
Genus
References
Pleistophora
Casal et al. (2016)
Toxoglugeidae
Toxospora
Voronin (1993)
Tuzetiidae
Pankovaia
Simakova et al. (2009)
Paratuzetia
Poddubnaya et al. (2006)
Tuzetia
Voronin (1986)
Microsporidea genera
insertae sedis
Baculea
Loubes & Akbarieh (1978)
Caullerya
Wolinska et al. (2004)
Evlachovaia
Issi (1986)
Globulispora
Vávra et al. (2016a)
Gurleyides
Voronin (1986)
Hamiltosporidium
Haag et al. (2011)
Holobispora
Voronin (1986)
Kabataia
Casal et al. (2010)
Kabatana
Lom et al. (2001)
Microsporidium
Jones et al. (2017, 2020)
Myosporidium
Jones et al. (2020)
Stempellia
Voronin (1996)
Rozellomycota genera
incertae sedis
Mitosporidium
Haag et al. (2014)
Paramicrosporidium
Corsaro et al. (2014)
Rozella
Canter (1969)
Rozellomycota orders
incertae sedis
Chytridiopsida
Chytridiopsidae
Chytridiopsis
Larsson (1993)
Chytrids that infect diatoms enter the host cell through the girdle region of the frustule using a germ tube. Infection in other algal hosts is via the
mucilage surrounding the host or directly through the cell wall if a mucilage layer is absent (Frenken et al. 2017). Rhizoids are produced, which
expand through the host cell to enable nutrient gathering. Chytrid parasites use the host resources to mature and produce zoospores, which are released
to the environment (Canter 1950, Canter & Lund 1951).
231
All aphelids described to date are obligate parasitoids (biotrophs) and can only be cultured
with their hosts (Held 1981, Gleason et al. 2012, Letcher & Powell 2019). Aphelids appear to be
very common parasitoids in many aquatic ecosystems, for example the genera Aphelidium and
Amoeboaphelidium (Schweikert & Schnepf 1996, Letcher et al. 2013, Ilicic & Grossart 2022).
Green algae and diatoms are common hosts of aphelids (Gleason et al. 2014, Jephcott et al. 2017).
For example, several species of Aphelidium and Amoeboaphelidium infect unicellular freshwater
green algae (Jephcott et al. 2017). Frequently, aphelid infection is observed in chlorococcous algae
and Tribonema gayanum and hence these hosts are commonly used to maintain gross cultures of
these parasites. Host specificity of aphelids has been previously investigated (Gromov &
Mamkaeva 1968a, b, Letcher et al. 2017). Most often these host-parasite relationships are genus
specific (Karpov et al. 2014b). However, scarce data is available to fully comprehend the extent of
host-specificity of these organisms and to understand how aphelids detect and differentiate between
host cells (Höger et al. 2021).
The life cycles of Aphelidium, Amoeboaphelidium and Pseudaphelidium are to some degree
comparable to each other (Letcher et al. 2013, Karpov et al. 2017b, Hurdeal et al. 2021, Ilicic &
Grossart 2022). The life cycle of aphelids is complex with different stages such as cyst, trophont,
plasmodium and zoospore stage (Karpov et al. 2014a, Letcher & Powell 2019). Briefly, the
parasitoid zoospore attaches to the host, encysts, and sheds the flagellum. The cyst germinates,
producing an infection tube through which it penetrates the host cell. A vacuole is produced within
the cyst, which enlarges and eventually pushes the cyst contents into the host cell (Karpov et al.
2017c). The parasitoid undergoes an intracellular phagotrophic amoebic life stage during which it
engulfs the cytoplasmic contents of the host and transports the nutrients into a central digestive
vacuole. As the parasitoid grows, it forms an endobiotic plasmodium that consumes the cytoplasm
of the host cell. The plasmodium then divides into uninucleate cells (zoospores) (Letcher & Powell
2019). Once the uniflagellate zoospores mature, they are released from the empty host cell through
the hole previously made by the infection tube. The cycle starts again, with the new zoospores
infecting other host cells (Letcher et al. 2013, Karpov et al. 2014a).
The endobiotic nature of aphelids may decrease the likelihood of observing them in culture-
based studies (Jephcott et al. 2017). The plasmodial stage is the most common phase observed in
culture due to its longevity and consists of a large vacuole containing the residual body of this stage
(Karpov et al. 2014a). Hence, studies on the ecological roles of the different life stages might
provide more insight on their importance in freshwater habitats.
Adaptations of basal fungi to aquatic habitats
Zoosporic fungi produce and propagate motile spores that require free water for dispersal
(Longcore & Simmons 2020). Presence of water is necessary, even for the culture and baiting of
zoosporic fungi from substrates such as soil (Gleason et al. 2012). Fungal zoospores are usually
uniflagellated (Sparrow 1960, Gleason & Lilje 2009). Some genera, especially parasites of algae,
invertebrates and fungi produce amoeboid zoospores with pseudopodia (Gleason & Lilje 2009).
The role of pseudopodia is speculative and needs to be further investigated. A proposed role is that
they might be assisting in spore movement over solid surfaces of the hosts. For short dispersal of
the zoospores, two mechanisms for active movement are known: flagellar and amoeboid. Flagellar
movement is best suited and common in larger volumes of water, while amoeboid movement is
best adapted for wet surfaces. For long range dispersal in water, zoospores may also use passive
mechanisms. Structures, such as the zoospore cyst, mature and/or resistant sporangium, and the
entire thallus function as asexual propagules. In freshwater habitats, these are carried horizontally
by currents and vertically in water columns (Gleason et al. 2008, Gleason & Lilje 2009).
Zoospores use chemotaxis to scour fresh substrates (Gleason et al. 2017). They are attracted
to specific sugar and amino acid exudates released as photosynthetic by-products by the host or
substrate. These spores swim for minutes to hours (or even days in rare cases) seeking new
substrates and encyst when favorable conditions are met (Powell & Letcher 2014a). The zoospore
relies exclusively on endogenous reserves, primarily lipids, and glycogen for energy (Powell &
232
Letcher 2014b). Once spores find a suitable substrate, they adhere to it and encyst. During
encystment, depending on the group of zoosporic fungi, the zoospores retract the flagellum or shed
it prior to assuming a spherical shape. Zoospores can then adhere firmly to the surface of the
substrates upon contact to encyst (Sparrow 1960). This behavior is of ecological significance for
both saprobic and parasitic taxa as this establishes a permanent contact between the fungus and its
potential food source. Zoospores become adhesive only during the initial stages of encystment
before a cyst wall is made and the adhesiveness only lasts for 30–60s (Tsui et al. 2016). The timing
of the adhesive phase coincides with the change from motile to sessile form, which is ecologically
beneficial to the fungus. Once the cell wall forms and the cyst matures the adhesive properties are
lost. Then, the cyst germinates, followed by penetration and colonization of the substrate (Tsui et
al. 2016).
Microsporidian species are mostly unable to grow or divide outside of their host cells
(Keeling 2009). They can only survive without their hosts as environmentally resistant, chitin-
containing spores, which also comprise their infective stage (Wadi & Reinke 2020). The intimate
relationship of microsporidia with their hosts, has led to a heavy dependence on the host for
resources. Hence, microsporidia have undergone extensive genomic reduction (Corradi 2015, Wadi
& Reinke 2020). The invasion apparatus of microsporidia is distinct from other intracellular
eukaryotic pathogens. During infection and upon spore germination, the polar tube pierces the host
cell. The parasite cytoplasm is delivered inside the host cytoplasm through the polar tube. The
parasite then proliferates and eventually produces spores (Keeling 2009, Wadi & Reinke 2020).
Abiotic and biotic factors affecting basal lineages of fungi
Spatial distribution, and seasonal fluctuations affect the community structure of zoosporic
fungi (Nascimento et al. 2011). Freshwater habitats are subject to variations in physical conditions.
The pH, temperature, concentration of soluble metals and salinity may fluctuate and have
repercussions on the freshwater microbial communities (Gleason et al. 2017). Temperature is an
important factor that affects the fungal community. Research based on saprobic isolates from soil
indicates the maximum temperature at which most of these zoosporic species grew was 30 °C.
While some grew at 35 °C, 37 °C, few at 40 °C, none survived at 45 °C or over (Gleason et al.
2017). Blastocladia species grew well in temperatures varying from 11 °C to 14 °C and the
maximum temperature at which they produce zoospores varied between 5 °C and 7 °C (Sparrow
1968). Batrachochytrium dendrobatidis, an important pathogenic chytrid, usually grows between
4–25 °C and does not survive long exposures at high temperatures (Powell 2017b). Some basal taxa
can tolerate a wide range of temperatures. Examples are the parasites Rhizophydium planktonicum
and Rhizosiphon anabaenae (Canter & Lund 1948, Paterson 1960).
Oxygen content in water is an integral part of freshwater habitats and dictates the survival of
basal fungi in those environments. It regulates the abundance of zoosporic fungi at differing depths
of the aquatic habitat by affecting their growth, distribution, and development (Voronin 2008).
Dissolved oxygen content has been correlated with parasitism by Rhizosiphon anabaenae (Paterson
1960). Specifically, the parasite maximum occurred when the oxygen content was around 6.4–8.0
ppm indicating that these values were optimal for growth, development, and reproduction of the
parasite in that study. However, whether infection rate is indeed affected by these values of oxygen
content remains undetermined. The amount of dissolved oxygen content in water depends on
factors such as the temperature of the water and concentration of salts. Temperature is inversely
proportional to the quantity of dissolved oxygen in water; high temperature decreases the solubility
of water oxygen content. Some basal fungal lineages grow at high oxygen content, while others
grow at low concentration (Lund 1934, Paterson 1960, Sparrow 1968, Voronin 2008). Zoospores
are usually concentrated in the oxygen rich water layers and the number of spores positively
correlates with the water oxygen content (Lund 1934, Paterson 1960). Among zygosporic fungi,
Mucor species may have higher resistance to the lack of oxygen than other freshwater fungal
species (Collins & Willoughby 1962).
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Acid rainfall, runoff, and fertilizers cause pH fluctuations in aquatic ecosystems. Some
species grow in acidic water, some at neutral-alkaline, and others in alkaline waters. Most fungi
grow at a pH between 4.0 and 9.0. Species diversity of zoosporic fungi decreases at acidotrophic
conditions (Sparrow 1968). However, some species have been found in environments of extreme
pH (Starkey & Waksman 1943). Zoosporic fungi have been found in bogs with pH as low as 3.6
(Mullen et al. 2000, Gleason et al. 2010a). In a fungal-based molecular survey of the acidic Rio
Tinto River in Spain (pH: 2.0), sequences corresponding to chytrids were recovered (Zettler et al.
2002). Several species of chytrids, including Rhizophlyctis rosea, have been cultured in a broad pH
range (Gleason et al. 2010a). In the case of zoosporic fungi, the dispersal stage might be the most
affected by variation in the environment as the zoospores are unwalled.
The concentration of dissolved organic matter (DOM) strongly influences zoosporic fungi.
An increase in DOM, increased the number of fungal species if the temperature and salt
concentration were relatively low. Nonetheless, a significant increase in DOM with varying
pollution decreased species diversity and occurrence, both of which could potentially lead to
extinction (Harvey 1952, Tan & Lim 1984). Toxic metals have both beneficial and detrimental
effects on basal fungi especially on the zoospores. The type of toxic metal and the life stage of the
organism dictates the effects. Presence of some of these metals can promote encystment and
germination of the zoospores (Gleason et al. 2017). For example, low levels of copper, lead and
zinc have a stimulatory effect on zoospore release (Henderson et al. 2015). The effects of abiotic
factors on fungal parasitism are still scarce and largely unknown. Canter & Lund (1948) noticed
that all but one epidemic by Rhizophydium on Asterionella occurred when the concentrations of
dissolved nitrate, silica and phosphate were high or rising. These epidemics were also noted to
happen when the concentration of inorganic matter was low. This study was based on Esthwaite
Water, and the south and north basins of Lake Windermere. The correlation of abiotic factors such
as available silica in water, duration and intensity of light to degree of parasitism is difficult (Lund
1934, Canter & Lund 1951, 1953). Some researchers have speculated on the effects of factors such
as light in parasitism. Bruning (1991) investigated the effects of light and phosphorus limitation on
the parasite Rhizophydium. The effects of these two variables on the growth of the parasite in the
study were comparable. Phosphorus limitations, as well as light limitation, reduced zoospore
production, and slightly delayed the development of the sporangia. Ultimately, they could reduce
epidemic threshold to a certain degree.
Biotic factors significantly affect growth and proliferation of basal fungi. A high diversity of
Rhizophydium and Nowakowskiella was observed in the mesotrophic Lake Bourget in eastern
France, when green algae and diatoms were abundant (Lepère et al. 2008). Moreover, a decline of
rotifer population was linked to the Olpidiomycota species Olpidium gregarium in the Rio Grande
Reservoir, São Bernardo do Campo, São Paulo State, Brazil (Meirinho et al. 2013). Studies have
also shown a positive correlation between either total chlorophyll concentration or proportions of
diatom sequences and zoosporic fungi, which may indicate the likelihood to parasitize algae
(Gleason et al. 2017).
The Trichomycetes, an Aquatic Group of Arthropod-Gut Endosymbionts
Introduction and overview of the group
The Trichomycetes were once a fungal Class within the Zygomycota (Lichtwardt 1986), but
actually the trichomycetes (with lower t, to indicate that this is not a natural, monophyletic clade)
are recognized as a polyphyletic group of fungal and protozoan microorganisms sharing the
ecological characteristic of living as obligate symbionts associated with diverse arthropod groups
(Lichtwardt et al. 2001a). Most trichomycetes are endosymbionts, living attached to the inner gut
lining of their hosts and extract nutrients from the food particles passing through the digestive
system. Only one known genus contains ectosymbiont species (Amoebidium spp) living on the
host’s chitinous exoskeleton (Lichtwardt et al. 2001a). Most trichomycetes are aquatic, associated
with immature stages of insects in freshwater and crustaceans in freshwater and marine ecosystems,
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but some species have terrestrial hosts such as collembolans, coleoptera, miriapoda and isopoda
(Lichtwardt et al. 2001a).
This ecological group includes three fungal orders: Harpellales, Orphellales and Asellariales,
placed within the Zoopagomycota, subphylum Kickxellomycotina (Hibbett et al. 2007, Tretter et al.
2014, Spatafora et al. 2016, White et al. 2018), which are the subject of the present entry. On the
other hand, the Eccrinales and Amoebidiales (“protistan trichomycetes”) are now placed within the
protist class Mesomycetozoea (= Ichthyosporea), Order Eccrinida (Cafaro 2005), but were once
classified as fungi due to their thallial and spore structure and their ecology. This is the historical
and practical reason why mycologists studying aquatic arthropod gut endobionts
(trichomycetologists) also study this group of protists. In fact, species of Paramoebidium
(Amoebidiales) are frequently found sharing the insect host hindgut with their fungal counterparts
(Harpellales and Orphellales), while the Asellariales and Eccrinales are mostly associated with non-
insect hosts such as Crustacea and Diplopoda, with a few Eccrinid species found in terrestrial
beetles (Coleoptera).
The ecology of Trichomycetes
Ecologically, the trichomycetes may have a role in the integrity of the system since they take
part in the macroinvertebrates’ biology. As often, unexpected changes in inland water systems are
due to the alterations in the complex connections among macroinvertebrates and associated trophic
webs (Goedkoop & Johnson 1996, Lodge et al. 1998, Stockley et al. 1998), the study of arthropod-
associated microorganisms becomes interesting indeed. It is now known that aquatic
macroinvertebrates harbor a wide diversity of gut endosymbionts serving different functions, some
of which are not yet well understood, especially regarding fungi. The existence of gut bacteria has
long been recognized in freshwater and marine environments (Sochard et al. 1979, Gowing &
Silver 1983, Sinsabaugh et al. 1985, Harris 1993). The metabolic activity of these microorganisms
provides essential amino acids and vitamins (Fong & Mann 1980) and enhances the digestibility of
plant food by providing enzymes such as cellulases into the gut of their host (Sochard et al. 1979).
Although the bacterial microbiome is generally well recognized, fungal gut associates are less
familiar, but they are not rare, even in the gut of aquatic arthropods, as demonstrated by the
ubiquitous and common presence of t