The study, published in the journal IMA Fungus, tackles a problem that has long constrained the Chinese chaxingu industry: while cultivation has expanded dramatically since the 1980s, the relationship between the genetic makeup of individual strains and their agronomic performance has remained poorly characterised. Without this knowledge, breeders have lacked the tools to systematically select superior varieties, and the industry has had no formal Distinctness, Uniformity and Stability (DUS) test guidelines, the regulatory framework needed to protect new cultivars and ensure orderly commercial development.

To fill this gap, the research team assembled a panel of 100 strains drawn from three distinct sources: private mushroom farms, established culture collections and wild specimens. Geographically, the collection spanned 12 provinces and two municipalities across China, capturing the broad ecological and geographic range over which the species has adapted and been cultivated. All strains are maintained at the Jiangxi Agricultural University Culture Collection Center, creating a permanent reference resource for future breeding work.

Each strain was subjected to a standardised cultivation protocol designed to strip away environmental noise and reveal true genetic differences. Mycelia were first grown on potato dextrose agar at 25 degrees Celsius in darkness for seven days, then transferred to sterilised solid substrate packed in polypropylene filter bags. The substrate, a carefully balanced mixture of cottonseed hulls, coarse and fine sawdust, bagasse, wheat bran, soybean meal, cornmeal, gypsum and lime adjusted to roughly 65 percent moisture, was incubated at 20 to 25 degrees Celsius with 60 to 70 percent relative humidity. After approximately 50 days, once the bags were fully colonised, they were opened to trigger fruiting at 22 degrees Celsius, 95 percent humidity and diffuse light at around 600 lux. Six bags were prepared per strain, and from each bag ten representative fruiting bodies of standard shape and varying sizes were measured.

The team quantified ten agronomic traits on every strain, ranging from the number of days to first harvest and total yield per bag to the fine anatomy of the mushrooms themselves: stipe length, stipe diameter, cap diameter, cap thickness, gill width and a morphological index calculated as the ratio of cap diameter to stipe length. The results were striking. Every single trait showed statistically significant variation across strains, with one-way analysis of variance returning p-values below 0.01 across the board. The coefficient of variation, a measure of relative dispersion, was highest for the total number of fruiting bodies at 45.6 percent, followed by average fruiting-body weight at 35.0 percent. Days to first harvest varied the least, at just 7.6 percent, suggesting that timing of fruiting is a relatively stable characteristic across the species while the economics of yield, how many mushrooms and how big they are, is where the real breeding leverage lies.

To make sense of this multivariate data, the researchers turned to correlation analysis, hierarchical clustering and principal component analysis. The correlation heatmaps revealed a fundamental trade-off at the heart of chaxingu biology: total yield correlated positively with average fruiting-body weight, stipe diameter, cap diameter, cap thickness and gill width, but negatively with the total number of fruiting bodies. In other words, strains tend to follow one of two contrasting strategies, producing either fewer, heavier mushrooms or many, lighter ones. This trade-off, long suspected by growers, is now quantified with statistical rigour, giving breeders a clear target: increase cap and stipe size while moderating fruiting-body number to push yields higher.

Principal component analysis distilled the ten traits into three interpretable axes that together accounted for 83.98 percent of total variance. The first principal component, explaining 57.74 percent of the variation, loaded strongly and positively on cap thickness, average fruiting-body weight, stipe diameter, cap diameter and gill width, and negatively on fruiting-body number, effectively capturing a size-versus-count gradient. The second component, at 15.87 percent, was dominated by stipe length, describing an elongation axis. The third, at 10.37 percent, loaded on days to first harvest and total yield, capturing the timing of earliest harvest and yield onset. When strains were plotted along the first two components, the five colour-based groups, white, black, dark brown, brown and yellow, formed partially separated clusters, with yellow strains tending toward the lower-left, white toward the lower-right, brown toward the upper-left and black near the upper-right.

Hierarchical clustering using the unweighted pair group method with arithmetic mean, or UPGMA, grouped the 100 strains into four major phenotypic branches. Basidiome colour largely tracked these branches and served as a practical field discriminator, so the team partitioned strains into five colour-based groups. Between-group comparisons showed pervasive differences across all ten traits, with an overall p-value below 0.001. Black strains tended to have higher total yield and robust fruiting bodies, brown strains had the shortest time to first harvest, white strains had higher average fruiting-body weight but fewer fruiting bodies, and yellow strains produced the most fruiting bodies but the lightest ones with the lowest overall yield.

Alongside the phenotypic work, the researchers sequenced two gene fragments, the large subunit of the ribosomal RNA gene and the translation elongation factor 1-alpha gene, chosen for their universal primers, their established role in fungal phylogenetics and the known intraspecific variation in TEF1-alpha. Bayesian phylogenetic analysis of the concatenated sequences resolved four principal genetic clusters. When these genetic clusters were mapped against the phenotypic dendrogram, the picture was one of partial congruence: white and black strains frequently co-clustered genetically, and a subset of brown strains were mixed with dark brown, white and black lineages. Some agronomic patterns, therefore, appear to reflect genuine underlying genetic structure, while others likely arise from phenotypic plasticity or polygenic variation that two conserved loci cannot resolve.

The practical implications are considerable. Based on the variance structure and between-strain contrasts, the team proposes stipe diameter, stipe length and gill width as the primary indicator traits for DUS testing in C. chaxingu, with size-related traits such as average fruiting-body weight, cap diameter, cap thickness and fruiting-body number serving as complementary selection targets. Under current Chinese practice, distinctness can be established when a candidate variety shows a repeatable, significant difference in at least one trait versus a reference variety, making these three anatomical measurements a pragmatic and reproducible starting point for formal variety protection.

The study also identified individual strains with superior performance in one or multiple economic traits, including short cultivation periods and high overall productivity, flagging them as candidate parental stocks for future breeding programmes. The researchers are careful to note the limitations of their approach: the fruiting assessment was conducted at a single site under a single set of conditions, so multi-site, multi-season trials will be needed to quantify genotype-by-environment interactions and trait repeatability. And with only two loci sequenced, both highly conserved within the species, denser genetic markers such as multilocus sequence typing, amplified fragment length polymorphisms or whole-genome sequencing will be needed to draw tighter links between genomic regions and the traits that matter most to growers.

What makes this work resonate beyond the chaxingu industry is its methodological clarity. Rather than treating breeding as an art guided by intuition, the study demonstrates how a disciplined pipeline of standardised cultivation, quantitative trait measurement, multivariate statistics and molecular phylogenetics can transform a traditionally managed crop into one amenable to modern genetic improvement. The tetrapolar heterothallic mating system of C. chaxingu, in which compatibility is governed by two unlinked mating-type loci, is particularly well suited to such investigations and offers a powerful experimental platform for dissecting the genetic architecture of yield, size and earliness.

For a mushroom that has been cultivated in China for decades yet has never before been subjected to a systematic diversity assessment at this scale, the study marks a turning point. It provides the quantitative foundation for germplasm collection, parental selection and the development of DUS guidelines, and it offers growers a practical field guide: colour is a useful first cue, but the real story of each strain is written in millimetres of stipe diameter, gill width and the subtle trade-off between how many mushrooms a bag produces and how heavy each one becomes.

Subject of Research: Genetic and phenotypic diversity assessment for multi-trait selection in the cultivated edible mushroom Cyclocybe chaxingu

Article Title: Assessing genetic and phenotypic diversity to advance multi-trait selection in Cyclocybe chaxingu

Article References: Yi, J., Su, G., Yang, Q., Song, H., Xu, J., Chen, M., Zhou, J., Yin, H., Gafforov, Y., & Hu, D. (2026). Assessing genetic and phenotypic diversity to advance multi-trait selection in Cyclocybe chaxingu. IMA Fungus, 17, Article e194629. https://doi.org/10.3897/imafungus.17.194629

Image Credits: AI Generated

DOI: 10.3897/imafungus.17.194629

Keywords: Cyclocybe chaxingu, edible mushrooms, genetic diversity, phenotypic diversity, DUS testing, mushroom breeding, principal component analysis, phylogenetics, agronomic traits, China, strain selection, yield trade-off

The study’s methodology was deliberately comprehensive. Floral traits were measured with a vernier caliper, allowing the team to quantify features such as corolla length, petal number, stamen number, pistil number, and ovule number across the collected accessions. Pollen morphology, a character set that has historically proven valuable in camellia systematics, was examined with scanning electron microscopy and then quantified using ImageJ software, yielding measurements of polar axis length, equatorial axis length, colpus length, and the ratio between the polar and equatorial dimensions. To evaluate how much variation existed and how it was structured, the researchers applied the coefficient of variation, correlation analysis, the Shannon-Wiener diversity index, principal component analysis, cluster analysis, and a supervised method known as orthogonal partial least squares-discriminant analysis, or OPLS-DA, which was used specifically to identify the traits that best separate groups of accessions from one another.

The results revealed striking variability in the flowers. The Shannon-Wiener diversity index for floral traits ranged from 0.22 to 2.76, with corolla length showing the highest diversity and pistil number the lowest. Coefficients of variation for floral characters spanned 7.21 percent to 23.80 percent, with pistil number again the most stable trait and stamen number the most variable. In practical terms, this means that within a single species growing in a single mountain region, individual plants differ dramatically in the size and composition of their blossoms. For breeders, such variation is raw material: corolla length and stamen number, the traits that vary most, are precisely the kinds of characters that ornamental horticulture prizes when selecting plants with showier or more unusual flowers.

The pollen data added a second, complementary layer of evidence. Under the scanning electron microscope, the pollen grains were oblong in equatorial view and trilobate-circular in polar view, and their exine ornamentation was granulate-foveolate, a surface texture of granules interspersed with small pits that the authors highlight as a distinct palynological signature of these accessions. Pollen traits were generally less diverse than floral traits, with Shannon-Wiener indices between 1.42 and 1.91, but the coefficients of variation ranged widely, from 4.76 percent up to 45.09 percent, indicating that certain pollen dimensions are far more labile than others. Because pollen morphology often reflects deep evolutionary constraints, the consistency of the granulate-foveolate pattern across accessions supports the placement of C. rosthorniana within the genus while the quantitative variation hints at local differentiation.

Correlation analysis exposed an interesting asymmetry in how the traits covary. Associations among floral traits were stronger than associations between floral and pollen traits, suggesting that the two character sets respond to partly independent developmental and evolutionary pressures. Two negative correlations stood out: ovule number was negatively correlated with colpus length, and petal number was negatively correlated with equatorial axis length. These relationships imply trade-offs within the reproductive system, where investment in one component of the flower or pollen grain appears to come at the expense of another. Such trade-offs are of interest to evolutionary biologists because they can reveal the functional architecture underlying reproductive strategies in wild populations.

Multivariate statistics then turned this sea of measurements into a coherent structure. Principal component analysis identified six principal components that together accounted for 72.973 percent of the total variance, a substantial share indicating that a handful of composite axes capture most of the morphological information in the dataset. Cluster analysis, using Euclidean distance with a linkage distance of L = 11, grouped the germplasm accessions into three major clusters. OPLS-DA, the supervised discriminant technique, pinpointed six key discriminatory variables that drive the separation between these groups: ovule number, equatorial axis length, corolla length, the polar-to-equatorial ratio, polar axis length, and petal number. Box plot analyses confirmed that the three clusters differ significantly across eight floral and pollen traits, meaning the groupings are not statistical artifacts but reflect genuine morphological divergence among accessions.

Perhaps the most intriguing finding concerns the stamens. Frequency distribution patterns of specific stamen traits revealed a numerical shift that the authors interpret as a potential phenotypic tendency toward stamen-to-petal transformation. This phenomenon, known as homeosis, is the developmental process by which one organ type assumes the identity of another, and it is famously responsible for the double flowers that make ornamental camellias so prized. A natural population showing a statistical tendency in this direction offers a rare window on how the raw variation for such transformations might arise in the wild, before any human selection has acted. If validated genetically, this tendency could make Fanjing Mountain germplasm especially valuable for breeding camellias with fuller, more petal-rich blooms.

The study also assessed the practical quality of the germplasm through in vitro germination tests. Clusters I and II both maintained high average germination rates exceeding 70 percent, with no significant difference between them, indicating vigorous, viable material across much of the collection. Cluster II, however, exhibited greater germination stability and a more consistent distribution than Clusters I and III, suggesting that accessions in that group may be more reliable candidates for propagation and conservation. Germination performance is a critical filter in germplasm utilization, because morphological novelty is of little use if the material cannot be propagated dependably in nurseries or breeding programs.

The systematic implications of the work are equally noteworthy. Pollen morphology has long served as a taxonomic marker in Theaceae, and the granulate-foveolate exine ornamentation documented here provides a palynological character that distinguishes these accessions and contributes to understanding their position within the genus Camellia. At the same time, the clear morphological differentiation among three clusters within a single species raises questions about how local populations of C. rosthorniana have diverged across the heterogeneous terrain of the Fanjing Mountain region, a UNESCO-recognized biodiversity hotspot in Guizhou. The authors are careful to frame their results as preliminary morphological evidence: the observed differentiation will require further genetic and field validation before it can be translated into formal varietal classification or taxonomic revisions.

For conservationists and breeders alike, the message is that this wild camellia deserves closer attention. C. rosthorniana is currently listed as Not Evaluated on the IUCN Red List and is not covered by CITES appendices, although Guizhou Province categorizes it as Least Concern owing to its stable local populations. That stable status, combined with the substantial phenotypic diversity now documented, makes the Fanjing Mountain region a potentially important reservoir of genetic resources for ornamental Camellia improvement. The study, conducted with specimens from the C. rosthorniana Germplasm Nursery at Tongren University within the Kaima Forest Farm and supported by provincial science and education funding programs, demonstrates how a rigorous morphological pipeline, from caliper and microscope to PCA and OPLS-DA, can transform a little-known wild shrub into a candidate resource for horticulture, while reminding researchers that the mountains of southern China still hold unexplored variation in even their most familiar plants.

Subject of Research: Floral and pollen morphological diversity of Camellia rosthorniana germplasm from the Fanjing Mountain region

Article Title: Floral and pollen diversity of Camellia rosthorniana germplasm accessions from Fanjing Mountain region: systematic significance and germplasm utilization

Article References: Xu, B., Zhao, J., Zhu, Z., Wu, J., Ming, Y., Yang, H., Yang, C., & Li, J. (2026). Floral and pollen diversity of Camellia rosthorniana germplasm accessions from Fanjing Mountain region: systematic significance and germplasm utilization. BMC Plant Biology. https://doi.org/10.1186/s12870-026-09994-6

Image Credits: AI Generated

DOI: 10.1186/s12870-026-09994-6

Keywords: Camellia rosthorniana, Fanjing Mountain, pollen morphology, floral traits, germplasm, phenotypic diversity, scanning electron microscopy, principal component analysis, OPLS-DA, cluster analysis, plant breeding, Guizhou