Four-locus phylogeny supports new freshwater fungus genus from Guizhou

A new genus and species of freshwater fungus, Hyalosympodium aquaticum, was identified from submerged, decaying wood collected in Guizhou Province, China. Researchers established the taxon through a combination of microscopic morphology and phylogenetic analyses of four gene regions, the large-subunit ribosomal RNA gene, the internal transcribed spacer, the small-subunit ribosomal RNA gene and part of the RNA polymerase beta-subunit gene. The study placed the fungus within the order Pararamichloridiales and in the class Sordariomycetes. Two isolates formed a previously unrecognized lineage that was sister to Tenebrosynnematica obclavata, with 94% maximum-likelihood support and a Bayesian posterior probability of 1.00. The work describes a new lineage from one freshwater locality, and the authors note that the broader relationships of Pararamichloridiales remain uncertain because several related taxa have limited sequence data and taxon sampling is incomplete.

The specimens were collected on 12 October 2025 from decaying wood in a freshwater habitat at Changpo Ling National Forest Park in Baiyun District, Guiyang City. The material was examined under a stereomicroscope, and the conidiophores, conidiogenous cells and conidia were photographed with a digital camera attached to a compound microscope. Measurements were made with image-analysis software. Single conidia were isolated and transferred to potato dextrose agar, where the resulting colonies were maintained at room temperature for 39 days. Their color, shape, dimensions, margins and elevation were documented. A dried specimen was deposited in the Herbarium of the Guizhou Academy of Agricultural Sciences, and two living cultures were placed in the Guizhou Culture Collection. The new species was also registered in MycoBank.

On its natural substrate, Hyalosympodium aquaticum produces superficial, effuse and gregarious colonies bearing masses of hyaline conidia. Its mycelium is partly superficial and consists of branched, septate, smooth-walled hyphae that range from hyaline to pale brown. The conidiophores are described as macronematous and arranged in groups as synnemata. They are cylindrical, straight or slightly flexuous, septate, and subhyaline to brown, with percurrent proliferation from their cut ends. The conidiogenous cells are integrated, cylindrical, smooth, polyblastic and sympodial, and measure 29–43 by 4–5 micrometres, with a reported mean of 34 by 4.5 micrometres based on 30 measurements. The conidiophores measure 163–211 by 4–6 micrometres, with a mean of 183.5 by 5 micrometres based on 30 measurements.

The conidia are acropleurogenous, ellipsoidal to oval and apically rounded, with smooth, hyaline walls. They measure 7–19 by 3–4.5 micrometres, with a mean of 10.5 by 4 micrometres based on 35 measurements. The spores are guttulate and have zero to two septa; when a septum is present, the spore is slightly constricted at it and carries a narrowed, frill-like structure. The genus name refers to the presence of hyaline conidia and sympodial conidiogenous cells, while the species epithet refers to the aquatic habitat from which it was collected. The sexual morph remains undetermined. In culture, conidia began germinating on potato dextrose agar within 14 hours, producing germ tubes from the conidial body. Colonies were white on both the upper and lower surfaces, had flat surfaces and undulate margins, and reached 33 millimetres in diameter after 39 days at 25 degrees Celsius.

For the molecular analysis, genomic DNA was extracted from mycelium scraped from living cultures. The four loci were amplified by PCR, and the products were purified and sequenced. Forward and reverse reads were checked and assembled before separate alignments were prepared with MAFFT and inspected in AliView. The LSU, ITS, SSU and rpb2 datasets were trimmed and merged into a 3,549-character matrix containing 75 taxa. Maximum-likelihood analysis was conducted through the IQ-TREE web server with substitution models selected automatically. Bayesian inference was performed in MrBayes on XSEDE. The selected models were GTR+I+G for LSU, ITS and rpb2, and GTR+G for SSU. Four simultaneous Markov chains were run for 10 to the seventh generations, trees were sampled every 1,000 generations, and a 25% burn-in was applied. The maximum-likelihood and Bayesian reconstructions produced consistent topologies for the sampled taxa.

Within that phylogeny, the two Hyalosympodium isolates formed a distinct branch within Pararamichloridiales. Hyalosympodium aquaticum, represented by cultures GZCC 27-27596 and GZCC 27-27597, was positioned as the sister lineage of Tenebrosynnematica obclavata. Direct sequence comparisons also separated the new species from its closest named relative. Hyalosympodium aquaticum and T. obclavata differed at 70 of 473 base pairs in ITS, corresponding to 14.8% with 15 gaps, and at 12 of 813 base pairs in LSU, corresponding to 1.5% with one gap. BLASTn searches found 85.79% ITS similarity across 98% of the query sequence coverage and 96.27% LSU similarity across 100% coverage with T. obclavata. These molecular comparisons accompanied a clear morphological contrast: H. aquaticum has hyaline, ellipsoidal to oval conidia, while T. obclavata has brown, fusiform to obclavate conidia.

The new genus could also be distinguished from Brunneoloconidia. The two share macronematous, cylindrical and septate conidiophores; polyblastic, integrated, sympodial and cylindrical conidiogenous cells; and acropleurogenous, guttulate, ellipsoidal, hyaline conidia. Hyalosympodium nevertheless forms gregarious colonies and has synnematous conidiophores, whereas Brunneoloconidia has mononematous conidiophores and conidiogenous cells that are recurved at the apex. Brunneoloconidia also produces fusiform conidia that are subhyaline to pale brown rather than the hyaline oval conidia observed in Hyalosympodium. Sequence comparisons between the Hyalosympodium type isolate and Brunneoloconidia aquatica showed 77 of 541 base-pair differences in ITS, or 14.2% with 14 gaps, and 32 of 858 differences in LSU, or 3.7% with two gaps. The combined molecular and structural evidence therefore supported establishment of Hyalosympodium as a separate genus with H. aquaticum as its type species.

Three genera now form a well-supported cluster in the sampled phylogeny. Tenebrosynnematica was introduced from lignicolous terrestrial substrates in Guizhou and initially treated as a Diaporthomycetidae genus of uncertain family placement. Brunneoloconidia was subsequently described from lignicolous freshwater substrates in Guizhou, and both genera were placed in Pararamichloridiales. Hyalosympodium was likewise recovered from freshwater wood and occupies a distinct branch alongside those taxa. All three possess elongated, septate conidiophores that may be pale brown to dark brown and grouped in synnemata, as well as septate conidia ranging from hyaline to pale brown. The study’s authors said this combination of molecular and morphological evidence helps clarify relationships among hyphomycetous fungi from freshwater and terrestrial substrates.

The analysis also raised a preliminary question about the order as a whole. Cyanoannulus petersenii was not recovered within Pararamichloridiales in the reconstructed tree and instead appeared as a separate lineage distant from other members. That result raises the possibility that the order, as currently circumscribed, is polyphyletic. The authors did not treat the placement as conclusive, citing the shortage of sequence data for several related taxa, insufficient sampling across the group and the possible influence of morphological convergence. Additional molecular data and more comprehensive phylogenetic studies would be required to reassess the evolutionary relationships and boundaries of Pararamichloridiales. Within those stated limits, the evidence from two isolates, detailed morphology and four-locus phylogeny supports Hyalosympodium aquaticum as a new genus and species associated with decaying freshwater wood in Guizhou.

Subject of Research: Taxonomic and phylogenetic characterization of a new lignicolous freshwater fungus from Guizhou, China

Article Title: Hyalosympodium gen. et sp. nov. ( Pararamichloridiales , Diaporthomycetidae ) from freshwater habitats in Guizhou, China

Article References: Zhang, W., Gao, Z., Song, X., Zhou, X., Xie, W., Yang, Y., Lu, J., & Feng, Q. (2026). Hyalosympodium gen. et sp. nov. (Pararamichloridiales, Diaporthomycetidae) from freshwater habitats in Guizhou, China. MycoKeys, 136, 79-98. https://doi.org/10.3897/mycokeys.136.195917

Image Credits: AI Generated

DOI: 10.3897/mycokeys.136.195917

Keywords: freshwater fungi, Hyalosympodium aquaticum, fungal taxonomy, four-locus phylogeny, Diaporthomycetidae, Pararamichloridiales, Guizhou, decaying wood, hyphomycetes, molecular evolution, Hyalosympodium, freshwater