Diploid-dominant life cycles characterize the early evolution of Fungi.

Most of the described species in kingdom Fungi are contained in two phyla, the Ascomycota and the Basidiomycota (subkingdom Dikarya). As a result, our understanding of the biology of the kingdom is heavily influenced by traits observed in Dikarya, such as aerial spore dispersal and life cycles dominated by mitosis of haploid nuclei. We now appreciate that Fungi comprises numerous phylum-level lineages in addition to those of Dikarya, but the phylogeny and genetic characteristics of most of these lineages are poorly understood due to limited genome sampling. Here, we addressed major evolutionary trends in the non-Dikarya fungi by phylogenomic analysis of 69 newly generated draft genome sequences of the zoosporic (flagellated) lineages of true fungi. Our phylogeny indicated five lineages of zoosporic fungi and placed Blastocladiomycota, which has an alternation of haploid and diploid generations, as branching closer to the Dikarya than to the Chytridiomyceta. Our estimates of heterozygosity based on genome sequence data indicate that the zoosporic lineages plus the Zoopagomycota are frequently characterized by diploid-dominant life cycles. We mapped additional traits, such as ancestral cell-cycle regulators, cell-membrane- and cell-wall-associated genes, and the use of the amino acid selenocysteine on the phylogeny and found that these ancestral traits that are shared with Metazoa have been subject to extensive parallel loss across zoosporic lineages. Together, our results indicate a gradual transition in the genetics and cell biology of fungi from their ancestor and caution against assuming that traits measured in Dikarya are typical of other fungal lineages.

Sequence capture identifies fastidious chytrid fungi directly from host tissue.

The chytrid fungus Batrachochytrium dendrobatidis (Bd) was discovered in 1998 as the cause of chytridiomycosis, an emerging infectious disease causing mass declines in amphibian populations worldwide. The rapid population declines of the 1970s-1990s were likely caused by the spread of a highly virulent lineage belonging to the Bd-GPL clade that was introduced to naïve susceptible populations. Multiple genetically distinct and regional lineages of Bd have since been isolated and sequenced, greatly expanding the known biological diversity within this fungal pathogen. To date, most Bd research has been restricted to the limited number of samples that could be isolated using culturing techniques, potentially causing a selection bias for strains that can grow on media and missing other unculturable or fastidious strains that are also present on amphibians. We thus attempted to characterize potentially non-culturable genetic lineages of Bd from distinct amphibian taxa using sequence capture technology on DNA extracted from host tissue and swabs. We focused our efforts on host taxa from two different regions that likely harbored distinct Bd clades: (1) wild-caught leopard frogs (Rana) from North America, and (2) a Japanese Giant Salamander (Andrias japonicus) at the Smithsonian Institution's National Zoological Park that exhibited signs of disease and tested positive for Bd using qPCR, but multiple attempts failed to isolate and culture the strain for physiological and genetic characterization. We successfully enriched for and sequenced thousands of fungal genes from both host clades, and Bd load was positively associated with number of recovered Bd sequences. Phylogenetic reconstruction placed all the Rana-derived strains in the Bd-GPL clade. In contrast, the A. japonicus strain fell within the Bd-Asia3 clade, expanding the range of this clade and generating additional genomic data to confirm its placement. The retrieved ITS locus matched public barcoding data from wild A. japonicus and Bd infections found on other amphibians in India and China, suggesting that this uncultured clade is widespread across Asia. Our study underscores the importance of recognizing and characterizing the hidden diversity of fastidious strains in order to reconstruct the spatiotemporal and evolutionary history of Bd. The success of the sequence capture approach highlights the utility of directly sequencing pathogen DNA from host tissue to characterize cryptic diversity that is missed by culture-reliant approaches.

Integrative approach to species delimitation in Rhizophydiales: Novel species of Angulomyces, Gorgonomyces, and Terramyces from northern Thailand.

The Chytridiomycota is a phylum of zoosporic eufungi that inhabit terrestrial, freshwater, and oceanic habitats. Within the phylum, the Rhizophydiales contains several monotypic families theorized to hold a diverse assemblage of fungi yet to be discovered and properly described. Based on morphology alone, many species in this order are difficult or impossible to identify. In this study, we isolated three chytrids from northern Thailand. Phylogenetic analyses placed the isolates in three monotypic genera within Rhizophydiales. Intrageneric genetic distances in the internal transcribed spacer (ITS) ranged between 1.5 and 8.5%. Angulomyces solicola sp. nov. is characterized by larger sporangia, spores, and fewer discharge papilla than A.argentinensis; Gorgonomyces thailandicus sp. nov. has larger zoospores and fewer discharge papillae in culture compared to G. haynaldii; Terramyces chiangraiensis sp. nov. produces larger sporangia than T. subangulosum. We delimited species of Angulomyces, Gorgonomyces and Terramyces using a tripartite approach that employed phylogeny, ITS genetic distances and Poisson tree processes (PTP). Results of these approaches suggest more than one species in each genus. This study contributes to the knowledge of chytrids, an understudied group in Thailand and worldwide.

Globally invasive genotypes of the amphibian chytrid outcompete an enzootic lineage in coinfections.

Competition between genotypes is likely to be a key driver of pathogen evolution, particularly following a geographical invasion by distant strains. Theory predicts that competition between disease strains will result in the most virulent strain persisting. Despite its evolutionary implications, the role of strain competition in shaping populations remains untested for most pathogens. We experimentally investigated the in vivo competitive differences between two divergent lineages of the amphibian-killing chytrid fungus ( Batrachochytrium dendrobatidis, Bd). These Bd lineages are hypothesized to have diverged in allopatry but been recently brought back into secondary contact by human introduction. Prior studies indicate that a panzootically-distributed, global lineage of Bd was recently introduced into southern Brazil, and is competitively excluding enzootic lineages in the southern Atlantic Forest. To test for differences in competitive ability between invasive and enzootic Brazilian Bd isolates, we coinfected a model host frog system which we developed for this study ( Hymenochirus curtipes). We tracked isolate-specific zoospore production over the course of the coinfection experiment with chip-based digital PCR (dPCR). The globally invasive panzootic lineage had a competitive advantage in spore production especially during the first one to four weeks of infection, and on frogs that eventually succumbed to Bd infection. Our study provides new evidence that competitive pressure resulting from the human movement of pathogen strains can rapidly alter the genetics, community dynamics and spatial epidemiology of pathogens in the wild.

Morphology, Ultrastructure, and Molecular Phylogeny of Rozella multimorpha, a New Species in Cryptomycota.

Increasing numbers of sequences of basal fungi from environmental DNA studies are being deposited in public databases. Many of these sequences remain unclassified below the phylum level because sequence information from identified species is sparse. Lack of basic biological knowledge due to a dearth of identified species is extreme in Cryptomycota, a new phylum widespread in the environment and phylogenetically basal within the fungal lineage. Consequently, we are attempting to fill gaps in the knowledge of Rozella, the best-known genus in this lineage. Rozella is a genus of unwalled, holocarpic, endobiotic parasites of hosts including Chytridiomycota, Blastocladiomycota, Oomycota, Basidiomycota, and a green alga, with most species descriptions based on morphology and host specificity. We found a Rozella parasitizing a Pythium host that was a saprobe on spruce pollen bait placed with an aquatic sample. We characterized the parasite with light microscopy, TEM of its zoospores and sporangia, and its 18S/28S rDNA. Comparison with other Rozella species indicates that the new isolate differs morphologically, ultrastructurally, and genetically from Rozella species for which we have data. Features of the zoospore also differ from those of previously studied species. Herein we describe the Rozella as a new species, R. multimorpha.

Complex history of the amphibian-killing chytrid fungus revealed with genome resequencing data.

Understanding the evolutionary history of microbial pathogens is critical for mitigating the impacts of emerging infectious diseases on economically and ecologically important host species. We used a genome resequencing approach to resolve the evolutionary history of an important microbial pathogen, the chytrid Batrachochytrium dendrobatidis (Bd), which has been implicated in amphibian declines worldwide. We sequenced the genomes of 29 isolates of Bd from around the world, with an emphasis on North, Central, and South America because of the devastating effect that Bd has had on amphibian populations in the New World. We found a substantial amount of evolutionary complexity in Bd with deep phylogenetic diversity that predates observed global amphibian declines. By investigating the entire genome, we found that even the most recently evolved Bd clade (termed the global panzootic lineage) contained more genetic variation than previously reported. We also found dramatic differences among isolates and among genomic regions in chromosomal copy number and patterns of heterozygosity, suggesting complex and heterogeneous genome dynamics. Finally, we report evidence for selection acting on the Bd genome, supporting the hypothesis that protease genes are important in evolutionary transitions in this group. Bd is considered an emerging pathogen because of its recent effects on amphibians, but our data indicate that it has a complex evolutionary history that predates recent disease outbreaks. Therefore, it is important to consider the contemporary effects of Bd in a broader evolutionary context and identify specific mechanisms that may have led to shifts in virulence in this system.

Fayochytriomyces, a new genus within Chytridiales.

Chytriomyces is a complex genus in Chytridiales. The morphological concept of the genus expanded as new taxa were added, and studies of zoospore ultrastructure and molecular phylogenies have revealed the genus to be polyphyletic. One problematic taxon is C. spinosus Fay, a distinctive species characterized by whorls of spines on the zoosporangium and a large accumulation of vesicle material beneath the operculum. With light-, scanning-electron and transmission-electron microscopy, we examined a culture (WJD186) isolated from a muck sample collected from a temporary forest pond. We also analyzed the D1-D2 variable domains of the nuc 28S rDNA (28S) sequences to confirm the phylogenetic placement of the species relative to the type of Chytriomyces, C. hyalinus Karling. The morphology of culture WJD186 is consistent with features Fay described for C. spinosus, and the zoospore ultrastructure is consistent with the Group I-type zoospore characters of Chytriomycetaceae (Chytridiales). In our molecular phylogeny C. spinosus does not group with the type of Chytriomyces. Consequently, we erect a new genus in Chytriomycetaceae and present the new combination Fayochytriomyces spinosus.

Cytology and molecular phylogenetics of Monoblepharidomycetes provide evidence for multiple independent origins of the hyphal habit in the Fungi.

The evolution of filamentous hyphae underlies an astounding diversity of fungal form and function. We studied the cellular structure and evolutionary origins of the filamentous form in the Monoblepharidomycetes (Chytridiomycota), an early-diverging fungal lineage that displays an exceptional range of body types, from crescent-shaped single cells to sprawling hyphae. To do so, we combined light and transmission electron microscopic analyses of hyphal cytoplasm with molecular phylogenetic reconstructions. Hyphae of Monoblepharidomycetes lack a complex aggregation of secretory vesicles at the hyphal apex (i.e. Spitzenkörper), have centrosomes as primary microtubule organizing centers and have stacked Golgi cisternae instead of tubular/fenestrated Golgi equivalents. The cytoplasmic distribution of actin in Monoblepharidomycetes is comparable to the arrangement observed previously in other filamentous fungi. To discern the origins of Monoblepharidomycetes hyphae, we inferred a phylogeny of the fungi based on 18S and 28S ribosomal DNA sequence data with maximum likelihood and Bayesian inference methods. We focused sampling on Monoblepharidomycetes to infer intergeneric relationships within the class and determined 78 new sequences. Analyses showed class Monoblepharidomycetes to be monophyletic and nested within Chytridiomycota. Hyphal Monoblepharidomycetes formed a clade sister to the genera without hyphae, Harpochytrium and Oedogoniomyces. A likelihood ancestral state reconstruction indicated that hyphae arose independently within the Monoblepharidomycetes lineage and in at least two other lineages. Cytological differences among monoblepharidalean and other fungal hyphae are consistent with these convergent origins.

Dendrochytridium crassum gen. et sp. nov., a taxon in Chytridiales with unique zoospore ultrastructure.

A water culture of detritus collected from an Australian tree canopy yielded multiple isolates (designated JEL 352, JEL 353, JEL 354) of an unidentified chytrid that grew on pollen bait and encysted spores of a Dictyuchus sp. oomycete. Morphological information from JEL 352 and genetic information from JEL 354 of this unidentified chytrid have been in several publications but the organism has not been named. Because isolates JEL 352 and JEL 354 are no longer viable, we sequenced partial SSU and LSU rDNA of isolate JEL 353, documented its thallus morphology with light microscopy and determined its zoospore ultrastructure via transmission electron microscopy. DNA evidence placed JEL 353 in Chytridiaceae, and its genetic composition was identical to that of JEL 354. Thallus morphology of JEL 353 was similar to that of JEL 352. Its zoospore ultrastructure is less complex compared to other members of Chytridiaceae. In pure culture, the rhizoidal system differed from other members of the family in being unevenly broad and not tapering to fine tips. Based on genetic, morphological and ultrastructural evidence, we place this chytrid in a new genus in Chytridiaceae and describe it as the new species Dendrochytridium crassum.

Irineochytrium, a new genus in Chytridiales having zoospores and aplanospores.

Many described chytrids exhibit distinct morphological features that permit positive identification by light microscopy. Chytriomyces annulatus is one such species. It has a flap-like operculum and its sporangial wall is ornamented with multiple collar-like annulations proximal to the rhizoidal axis, features that, in combination, do not occur in any other described chytrid. Recent molecular phylogenies placed C. annulatus in the Chytridiaceae (Chytridiales) lineage, which is characterized by a Group II zoospore. Here we use light microscopy and transmission electron microscopy to examine thallus morphology of an isolate (JEL 729) of C. annulatus to confirm its identity and transmission electron microscopy to examine zoospore ultrastructure to confirm its phylogenetic placement. Light microscopic examinations confirmed its identity, and transmission electron microscopy analysis revealed both motile spores (zoospores) and nonmotile spores (aplanospores). Zoospores had a unique suite of ultrastructural features characteristic of the Group II zoospore; aplanospores had similar ultrastructure minus a flagellum. Chytriomyces annulatus does not group with the Chytriomycetaceae (Chytridiales) lineage containing the type of Chytriomyces, C. hyalinus, nor does it have a zoospore typical of that lineage. These arguments support the recognition of a distinct genus in Chytridiaceae, including one species, Irineochytrium annulatum.

Diversity of Rhizophydiales (Chytridiomycota) in Thailand: unveiling the hidden gems of the Kingdom.

Chytrids, often overshadowed by their other fungal counterparts, take center stage as we unravel the mysteries surrounding new species within Rhizophydiales and explore their unique characteristics. In the broader spectrum of chytrids, their significance lies not only in their roles as decomposers but also as key players in nutrient cycling within aquatic ecosystems as parasites and saprobes. Baited soil and aquatic samples collected from various provinces of Thailand, yielded new species of the Rhizophydiales (Chytridiomycota), some of which expanded previously single species genera. Our investigation incorporated a combination of morphological and phylogenetic approaches, enabling us to identify these isolates as distinct taxa. The novel isolates possess distinguishing features, such as variations in size and shape of the sporangium and zoospores, that somewhat differentiate them from described taxa. To confirm the novelty of the species, we employed robust phylogenetic analyses using maximum likelihood and bayesian methods. The results provided strong support for the presence of eight distinct lineages within the Rhizophydiales, representing our newly discovered species. Furthermore, we employed Poisson Tree Processes to infer putative species boundaries and supplement evidence for the establishment of our new Rhizophydiales species. By meticulously exploring their morphological characteristics and genetic makeup, we expand the known catalogue of fungal diversity by describing Alphamyces thailandicus, Angulomyces ubonensis, Gorgonomyces aquaticus, G. chiangraiensis, G. limnicus, Pateramyces pingflumenensis, Terramyces aquatica, and T. flumenensis and also provide valuable insights into the intricacies of this order. This newfound knowledge not only enriches our understanding of Rhizophydiales but also contributes significantly to the broader field of mycology, addressing a critical gap in the documentation of fungal species. The identification and characterization of these eight novel species mark a noteworthy stride towards a more comprehensive comprehension of fungal ecosystems and their vital role.

Chytrid mycoparasitism of entomophthoralean azygospores.

Mycoparasitism - when one fungus parasitizes another - has been reported to affect Beauveria bassiana and mycorrhizal fungi in the field. However, mycoparasitism of any fungi in the Order Entomophthorales has never been reported before now. The majority of entomophthoralean species persist as resting spores (either zygospores or azygospores) in the environment and dormant entomophthoralean resting spores (whether formed as zygospores or azygospores) are thought to be especially well adapted for survival over long periods due to their thick double walls. Entomophthoralean resting spores can accumulate in the soil as large reservoirs of inoculum which can facilitate the onset and development of epizootics. We report parasitism of azygospores of the gypsy moth pathogen Entomophaga maimaiga caged in soil from southern Ohio by the chytrid fungus Gaertneriomyces semiglobifer. G. semiglobifer had previously been isolated from soil samples from North America, Europe and Australia or horse manure from Virginia. After isolation and identification of G. semiglobifer, azygospores of E. maimaiga exposed to zoospores of G. semiglobifer exhibited high levels of mycoparasitism and G. semiglobifer was subsequently reisolated from mycoparasitized resting spores. We discuss the importance of this finding to the epizootiology of insect diseases caused by entomophthoralean fungi.

Pseudorhizidium is a new genus with distinct zoospore ultrastructure in the order Chytridiales.

A chytrid isolate (JEL 221) we identified as the rarely reported species, Rhizidium endosporangiatum Karling, was cultured axenically for the first time. The purposes of this study are to characterize the developmental morphology of isolate JEL 221 and to elucidate its zoospore ultrastructural features. Thallus development and morphology of isolate JEL 221 are characteristic of R. endosporangiatum as it was originally described. However, thallus morphology of R. endosporangiatum is not entirely typical of the genus Rhizidium, especially that of the type R. mycophilum. The presence of an endosporangium, a layer of material encapsulating the edges of the protoplast protruding through multiple discharge pores, makes this a distinctive species. Consistent with its published molecular-based phylogenetic placement, we found that isolate JEL 221 shared ultrastructural features with the two major zoospore types described for the Chytridiales but had distinct zoospore architecture. A new genus, Pseudorhizidium, is erected for this chytrid based on its thallus morphology, molecular phylogenetic placement and unique zoospore ultrastructure. This new genus does not fit into either of the described families (Chytridiaceae or Chytriomycetaceae) in the Chytridiales because of its unique zoospore ultrastructure, especially the two-layered nature of the electron-opaque plug in the base of the flagellum.

Novel, panzootic and hybrid genotypes of amphibian chytridiomycosis associated with the bullfrog trade.

Global amphibian declines are linked with the presence of specific, highly virulent genotypes of the emerging fungal disease chytridiomycosis caused by Batrachochytrium dendrobatidis (Bd) known as the global panzootic lineage (Bd-GPL). The global trade in amphibians for human consumption is suspected to have facilitated emergence of the disease, but evidence to support this is largely lacking. Here, we investigated the role the Lithobates catesbeianus (North American bullfrog) trade in spreading Bd genotypes by comparing strains associated with L. catesbeianus to a global panel using 36 sequenced loci from multiple chromosomal regions. Most bullfrogs were infected with Bd-GPL genotypes, but we also detected novel, highly divergent Bd genotypes (Bd-Brazil) from a live bullfrog in a US market and from native Brazilian anurans in the Atlantic Forest where bullfrogs are widely farmed. Sexual reproduction was also detected for the first time in Bd in the form of a hybrid genotype between the Bd-GPL and Bd-Brazil lineages in the Atlantic Forest. Despite the demonstration that ribosomal RNA types in Bd fail to undergo concerted evolution (over 20 sequence types may be found in a single strain), the Bd-GPL and Bd-Brazil lineages form largely separate clusters of related internal transcribed spacer (ITS) RNA sequences. Using ITS sequences, we then demonstrate the presence of Bd-Brazil in Japan, primarily on invasive L. catesbeianus. The finding that Bd is capable of sexual reproduction between panzootic and endemic genotypes emphasizes the risk of international wildlife trade as a source of additional Bd epizootics owing to hybridization.

Varying responses of northeastern North American amphibians to the chytrid pathogen Batrachochytrium dendrobatidis.

Chytridiomycosis, caused by Batrachochytrium dendrobatidis (Bd), is widespread among amphibians in northeastern North America. It is unknown, however, whether Bd has the potential to cause extensive amphibian mortalities in northeastern North America as have occurred elsewhere. In the laboratory, we exposed seven common northeastern North American amphibian species to Bd to assess the likelihood of population-level effects from the disease. We exposed larval wood frogs (Lithobates sylvaticus) and postmetamorphic frogs of six other species to two different strains of Bd, a northeastern strain (JEL404) and a strain that caused die-offs of amphibians in Panama (JEL423), under ideal in vitro growth conditions for Bd. Exposed American toads (Anaxyrus americanus) all died; thus, this species may be the most likely to die from Bd-caused disease in the wild. Both Bd strains were associated with mortalities of wood frogs, although half the metamorphs survived. The Bd strain from Panama killed metamorphic green frogs (L. clamitans), whereas the northeastern strain did not, which means novel strains of Bd may lead to death even when local strains may not. No mortality was observed in four species (bullfrogs [L. catesbeianus], northern leopard frogs [L. pipiens], spring peepers [Pseudacris crucifer], and blue-spotted salamanders [Ambystoma laterale]) and in some individuals of green frogs and wood frogs that we exposed. This finding suggests these six species may be Bd vectors. Our results show that systematic exposures of amphibian species to Bd in the laboratory may be a good first step in the identification of species susceptible to Bd-caused declines and in directing regional conservation efforts aimed at susceptible species.

Reconstructing the early evolution of Fungi using a six-gene phylogeny.

The ancestors of fungi are believed to be simple aquatic forms with flagellated spores, similar to members of the extant phylum Chytridiomycota (chytrids). Current classifications assume that chytrids form an early-diverging clade within the kingdom Fungi and imply a single loss of the spore flagellum, leading to the diversification of terrestrial fungi. Here we develop phylogenetic hypotheses for Fungi using data from six gene regions and nearly 200 species. Our results indicate that there may have been at least four independent losses of the flagellum in the kingdom Fungi. These losses of swimming spores coincided with the evolution of new mechanisms of spore dispersal, such as aerial dispersal in mycelial groups and polar tube eversion in the microsporidia (unicellular forms that lack mitochondria). The enigmatic microsporidia seem to be derived from an endoparasitic chytrid ancestor similar to Rozella allomycis, on the earliest diverging branch of the fungal phylogenetic tree.

Host invasion by Batrachochytrium dendrobatidis: fungal and epidermal ultrastructure in model anurans.

The chytridiomycete fungus Batrachochytrium dendrobatidis (Bd) colonizes mouthparts of amphibian larvae and superficial epidermis of post-metamorphic amphibians, causing the disease chytridiomycosis. Fungal growth within host cells has been documented by light and transmission electron microscopy; however, entry of the fungus into host cells has not. Our objective was to document how Bd enters host cells in the wood frog Lithobates sylvaticus, a species at high mortality risk for chytridiomycosis, and the bullfrog L. catesbeianus, a species at low mortality risk for chytridiomycosis. We inoculated frogs and documented infection with transmission electron microscopy. Zoospores encysted on the skin surface and produced morphologically similar germination tubes in both host species that penetrated host cell membranes and enabled transfer of zoospore contents into host cells. Documenting fungal and epidermal ultrastructure during host invasion furthers our understanding of Bd development and the pathogenesis of chytridiomycosis.

Thoreauomyces gen. nov., Fimicolochytrium gen. nov. and additional species in Geranomyces.

Powellomycetaceae (Spizellomycetales) contain a diverse group of exogenously developing chytrids found by baiting water preparations of soils and manure with pollen. A previous molecular phylogenetic study indicated that some lineages within this family represent undescribed genera and species. Description of genera within the Spizellomycetales traditionally has relied on ultrastructural characters of zoospores, whereas species have been based on thallus development and morphology. We analyzed Powellomycetaceae chytrids that had not yet had ultrastructural and thallus morphologies determined. Because these chytrids vary little morphologically, we used a linear discriminant function analysis of thallus characters to determine features most useful for separating species when grown in pure culture on identical media. Zoosporic ultrastructures of two groups of isolates differed from those of the two described genera in the family, and we describe the new genera Thoreauomyces with one new species and Fimicolochytrium with two new species. Also, we describe three new species within Geranomyces. Linear discriminant function analysis, although helpful for determining more stable morphological characters, was not completely accurate in assigning chytrids to the correct genus or species, thus emphasizing the importance of molecular characters for identifying these taxa.

The Polychytriales ord. nov. contains chitinophilic members of the rhizophlyctoid alliance.

During the past 5 y the Rhizophydiales, Cladochytriales and Lobulomycetales have been segregated from the formerly recognized Chytridiales. Descriptions of new chytridiomycete orders are based on molecular and ultrastructural characters, which have been phylogenetically mutually supportive. The Polychytrium clade has consisted of a few chitinophilic, soil and aquatic chytrids that clustered in phylogenetic hypotheses but have not been placed in a new order. We isolated additional putative members of this clade, sequenced their nucSSU and nucLSU rDNA and examined zoospores of some of the isolates with TEM. Our isolates are in a well supported clade with previous Polychytrium clade members, but zoospore ultrastructural types vary within the clade, with characters that often are conserved within other orders (e.g. flagellar plug, rumposome) being either present or absent. Based on the isolates in culture we describe the Polychytrium clade as the Polychytriales. This order contains Polychytrium, Lacustromyces, Karlingiomyces, two new genera (Arkaya and Neokarlingia) and additional undescribed taxa.

Alogomyces tanneri gen. et sp. nov., a chytrid in Lobulomycetales from horse manure.

The order Lobulomycetales contains chytrids from soil, freshwater and marine habitats; environmental DNA sampling has indicated that representatives of this order might be found in deep ocean localities. We describe Alogomyces tanneri as the first lobulomycetalean chytrid isolated from horse manure; A. tanneri is also the first species in the order to possess a rumposome in its zoospore. This species widens the range of habitats, ultrastructural variation and thallus morphology for Lobulomycetales.