Colletotrichum is a globally widespread fungal genus and the sole genus of the family Glomerellaceae within the order Glomerellales. It is listed among the world’s top ten most important phytopathogenic fungal genera, a distinction earned through its broad host range and strong pathogenicity. Members of this genus infect multiple plant organs, including leaves, fruits, and stems, triggering anthracnose diseases that frequently lead to sharp yield reductions or total harvest failure across diverse crops. The host-pathogen interactions of Colletotrichum are intricate: a single host plant may suffer co-infection by multiple Colletotrichum species, while one Colletotrichum species can parasitize a wide array of distinct hosts. Most species in this genus are important plant pathogens, while some can colonize hosts as endophytes or saprobes, making the ecological roles of individual species difficult to predict without precise identification.

The genus comprises multiple species complexes and harbors abundant cryptic species, which substantially impede accurate species identification. Morphological traits of Colletotrichum fluctuate drastically with environmental conditions and host substrates, resulting in low stability of diagnostic morphological markers and complex host-associated phenotypic variations. Traditional taxonomic frameworks for the genus were established primarily based on host association and morphological characters rather than natural evolutionary relationships, and such systems fail to reflect phylogenetic relatedness among taxa. With continuous refinement of fungal taxonomic frameworks, numerous cryptic species have been discovered and assigned to distinct species complexes. Based on a six-locus dataset, previous research resolved the genus Colletotrichum into 16 species complexes and 15 singleton species, among which the Colletotrichum bambusicola species complex was recognized as a distinct lineage associated with bamboo hosts.

Sichuan Province hosts the core wild giant panda habitats and concentrated distributions of their staple bamboos, where various exclusive food bamboo species for giant pandas, including Fargesia spathacea, Indocalamus latifolius, Qiongzhuea opienensis, Phyllostachys heteroclada, and Phyllostachys praecox, are widely distributed. The stable and healthy bamboo forest ecosystems provide favorable habitats for the colonization and proliferation of bamboo-inhabiting fungi. The health of bamboo communities directly determines the habitat stability and survival status of wild giant panda populations, and given their robust pathogenic potential, Colletotrichum fungi pose latent threats to the stability of vegetation communities and the ecological health of giant panda habitats. Reports on Colletotrichum infecting bamboos remain limited, with only a small number of records documenting bamboo-associated pathogens, highlighting an urgent need for research on early prevention and risk warning.

During fungal resource surveys carried out in the Giant Panda National Park in Ya’an City in April 2024 and in the Wolong Giant Panda National Park in the Aba Tibetan and Qiang Autonomous Prefecture in July 2024, researchers collected bamboo-derived fungal specimens from dead bamboo culms of Phyllostachys heteroclada. After preliminary processing, field-collected specimens were packaged in paper envelopes or resealable plastic bags, labeled with collection numbers and dates, and transported to the laboratory, where they were stored at 4 degrees Celsius until use. During sample processing, bamboo culms bearing distinct fungal structures were examined under a stereomicroscope, and a direct-picking method was employed to cross-section ascomata or pycnidia using a sterilized scalpel blade, after which the internal contents were precisely removed with a sterile needle and transferred into sterile water. The contents were thoroughly dispersed by agitation, and a sterile dropper was used to draw up the fungal-containing suspension and dispense it onto potato dextrose agar plates. The plates were incubated at 25 degrees Celsius or at room temperature for 12 to 24 hours, and after spore germination, individual germinated ascospores or conidia were sequentially transferred to fresh PDA plates using aseptic techniques.

To promote sporulation, mycelium was transferred to sterile bamboo leaf-bearing PDA plates and incubated under alternating conditions of 12 hours of near-ultraviolet light and 12 hours of darkness at 25 degrees Celsius. Once the target colonies produced spores, an appropriate amount of mycelium was scraped into sterile centrifuge tubes and stored at minus 20 degrees Celsius for subsequent DNA extraction and sequencing analysis. Genomic DNA was extracted from the ground samples following the operating instructions of a plant genomic DNA extraction kit, and polymerase chain reaction was employed to amplify six genetic loci, namely ITS, chs-1, tub2, act, his3, and gapdh. The newly generated sequence data were deposited in GenBank, and sequences of the Colletotrichum bambusicola species complex used for constructing the multigene dataset were downloaded from GenBank. The plant specimens collected in this study are deposited in the Mycological Herbarium of Sichuan Agricultural University, and the isolated strains are preserved in the Culture Collection Center of Sichuan Agricultural University.

Phylogenetic analyses were performed on the downloaded sequence fragments using maximum likelihood and Bayesian inference algorithms. All sequences were aligned using MAFFT version 7, with manual adjustments made when necessary, and the gene datasets were concatenated using Mesquite software to construct a combined multigene matrix. Maximum likelihood analyses were performed using RAxML-HPC2 under the GTRGAMMA model, with bootstrap support assessed from 1,000 pseudoreplicates, and Bayesian inference was implemented in MrBayes, with Bayesian posterior probabilities estimated via Markov chain Monte Carlo sampling. The optimal nucleotide substitution models for each locus were selected using MrModeltest, and the combined analyses included 32 strains and comprised 2,716 characters, including gaps. Phylogenetic trees generated from maximum likelihood and Bayesian inference analyses were similar in overall topology, and the best-scoring RAxML tree had a final likelihood value of minus 8,757.690199, with 771 distinct alignment patterns and 27.73 percent undetermined characters or gaps.

The phylogram generated from maximum likelihood analysis revealed that the newly collected strains formed two distinct clades within the Colletotrichum bambusicola species complex. Colletotrichum abaense was closely related to Colletotrichum parabambusicola and Colletotrichum bambusicola, while Colletotrichum phyllostachydis formed a clade with Colletotrichum hsienjenchang. Morphologically, both Colletotrichum parabambusicola and Colletotrichum bambusicola possess appressoria, and their ascospores lack guttules, whereas the strains of Colletotrichum abaense lacked appressoria and contained guttules within the ascospores. Colletotrichum hsienjenchang possesses distinct setae and appressoria, whereas the strains of Colletotrichum phyllostachydis lacked both setae and appressoria. Pairwise base comparisons showed that between the type strain of Colletotrichum abaense and Colletotrichum parabambusicola, the nucleotide differences for gapdh, chs-1, his3, and act gene sequences were 8.7 percent, 1.7 percent, 7 percent, and 2.2 percent respectively, and between the type strain of Colletotrichum abaense and Colletotrichum bambusicola, the nucleotide differences for gapdh, chs-1, and act gene sequences were 11.9 percent, 2.1 percent, and 3.6 percent respectively. Between the type strain of Colletotrichum phyllostachydis and Colletotrichum hsienjenchang, the nucleotide differences for chs-1 and his3 gene sequences were 2.5 percent and 3.2 percent respectively.

To evaluate whether recombination might obscure the phylogenetic relationships and species boundaries within the bamboo-associated Colletotrichum species complex, the pairwise homoplasy index test was performed on the concatenated six-locus dataset. When Colletotrichum abaense and Colletotrichum phyllostachydis were compared with their respective closely related taxa, the PHI test yielded values of 0.95 for both, substantially exceeding the 0.05 significance threshold. These results indicate no statistically significant recombination events between the two novel species and their phylogenetic neighbors, confirming that the concatenated dataset is free from reticulate evolutionary signals that could confound phylogenetic inference. The taxonomic status of the two species was further validated by the genealogical concordance phylogenetic species recognition approach, providing strong evidence for delimiting the two species as distinct taxa within the Colletotrichum bambusicola species complex. Both morphological characteristics and molecular phylogenetic analyses support the recognition of each fungus as a novel species.

The discovery of the two new species adds to the baseline data on fungi associated with giant panda staple bamboos and provides crucial references for species identification, diversity research, and habitat disease prevention of bamboo-inhabiting Colletotrichum. To date, more than 300 Colletotrichum species have been reported globally; however, complete multigene sequence data remain unavailable for certain species, and phylogenetic support for several closely related species complexes remains suboptimal. The findings lay a foundation for the exploration of bamboo fungal resources and the taxonomic study of the genus Colletotrichum. Current research on Colletotrichum has predominantly focused on species identification and disease surveys, whereas studies addressing their secondary metabolites, bioactivities, and potential applications remain comparatively underexplored. Previous investigations have indicated that certain Colletotrichum species are capable of producing diverse bioactive metabolites with promising potential for biological control and pharmaceutical development. Continued efforts in the exploration of Colletotrichum resources, species delineation, and functional characterization are expected to uncover additional taxa with potential applications in agriculture and biopharmaceutical development, thereby providing a theoretical foundation for the scientific management of plant anthracnose and the sustainable utilization of fungal resources.

Subject of Research: Two novel species of Colletotrichum ( Glomerellaceae , Glomerellales ) from Sichuan Province, China

Article Title: Two novel species of Colletotrichum ( Glomerellaceae , Glomerellales ) from Sichuan Province, China

Article References: Wang, F., Liu, F., Xu, X., Yang, C., & Liu, Y. (2026). Two novel species of Colletotrichum (Glomerellaceae, Glomerellales) from Sichuan Province, China. MycoKeys, 141, 77-93. https://doi.org/10.3897/mycokeys.141.209472

Image Credits: AI Generated

DOI: 10.3897/mycokeys.141.209472

Keywords: novel, species, Colletotrichum, Glomerellaceae, Glomerellales, Sichuan, Province, China, scientific research