Responses and assembly mechanisms of planktonic bacterial and fungal communities in the upper Chishui River during dry and wet seasons

Planktonic bacteria and fungi followed contrasting assembly pathways across the upper Chishui River, a karst headwater stream in Yunnan, China, according to a two-season field study. Bacterial communities were primarily structured by homogeneous selection in both the wet and dry seasons sampled, whereas fungal communities shifted from homogeneous selection in the wet season to predominantly stochastic processes, including ecological drift, in the dry season. The research, based on surface-water samples collected in January and May 2025 at three sites along a longitudinal disturbance gradient, found that the downstream nutrient pattern persisted across seasons even as community composition and assembly mechanisms changed.

Because discharge, temperature and water-residence time were not monitored continuously, the study could not separate their independent associations with microbial assembly. The researchers sampled three locations representing different levels of human disturbance: Chishuiyuan, an upstream forested reference site; Guozhu, a midstream site affected by agricultural and township runoff; and Erlongqiangbao, a downstream site receiving converged urban runoff. Each site had three biological replicates in each season, and 10 liters of surface water were collected per replicate from a depth of 0–50 centimeters.

High-throughput amplicon sequencing targeted the V3–V4 region of bacterial 16S rRNA genes and fungal ITS2 regions. Community composition, diversity and beta diversity were assessed using amplicon sequence variants, while the iCAMP framework partitioned assembly into homogeneous selection, heterogeneous selection, dispersal limitation, homogenizing dispersal and ecological drift. PICRUSt2 inferred bacterial metabolic potential, and FUNGuild assigned fungal trophic guilds.

Water chemistry established a consistent downstream gradient. Total nitrogen and total phosphorus increased from the forested upstream site through the agricultural midstream site to the urban-influenced downstream site, while dissolved oxygen declined. In May, total nitrogen ranged from 1.24 ± 0.08 milligrams per liter at Chishuiyuan to 2.87 ± 0.16 milligrams per liter at Erlongqiangbao; total phosphorus rose from 0.04 ± 0.01 to 0.12 ± 0.02 milligrams per liter.

January concentrations were lower but followed the same spatial order: total nitrogen increased from 0.84 ± 0.05 to 2.15 ± 0.12 milligrams per liter, and total phosphorus from 0.03 ± 0.01 to 0.09 ± 0.02 milligrams per liter. Water temperatures were 7.8–9.4 °C in January and 16.8–18.4 °C in May, while generally higher nutrient concentrations in the wet season were consistent with enhanced runoff inputs. The seasonal change altered concentration magnitudes without reversing the upstream-to-downstream pattern.

Application of China’s GB 3838-2002 surface-water standard identified the downstream reach as the most strongly nutrient-enriched. Total nitrogen at Erlongqiangbao was worse than Class V in May and remained above the 2.0-milligrams-per-liter Class V threshold in January. Downstream total phosphorus reached Class III in the wet season.

Dissolved oxygen, pH and ammonium nitrogen generally complied with Class II or better. However, the sampling design comprised only three fixed locations, and overlapping pollution sources at the midstream and downstream sites could not be fully separated. The observations therefore describe differences among sampled reaches rather than a continuous longitudinal response.

Bacterial alpha diversity was generally highest upstream, particularly in the dry season. The Ace richness index differed significantly among sites in January, with upstream values exceeding both midstream and downstream values after false-discovery-rate correction, while the two downstream sites did not differ significantly from each other. At Chishuiyuan, observed ASV richness, Chao1 richness and Shannon diversity were all significantly lower in winter than in spring.

Site also significantly affected bacterial community structure in both seasons: PERMANOVA reported p = 0.008 in spring and R² = 0.935 with p = 0.006 in winter. Non-metric multidimensional scaling separated upstream samples from the more compositionally similar midstream and downstream assemblages, with partial overlap between the latter two in the wet season and greater homogenization among them in the dry season. Seasonal turnover was pronounced among bacterial taxa.

Flavobacterium represented more than 95% of the wet-season upstream bacterial community but declined to approximately 30%–45% at the two downstream sites, where Limnohabitans accounted for about 20%–30%. In winter, Flavobacterium dominance at the upstream site receded. Limnohabitans exceeded 50% of the midstream community in the dry season, whereas Flavobacterium remained a dominant downstream genus at approximately 30%–35%.

The downstream winter community also included Clostridium_sensu_stricto_1 at about 15% and Malikia at about 10%. Linear discriminant analysis identified Flavobacterium as a wet-season indicator and Limnohabitans together with several Actinobacteriota lineages as dry-season indicators. These findings show that seasonal replacement was superimposed on the persistent spatial difference between the forested reference reach and the more disturbed downstream reaches.

Null-model analysis indicated that bacterial assembly was consistently dominated by homogeneous selection. In the wet season, homogeneous selection accounted for 35%–46% of assembly events and homogenizing dispersal contributed 25%–30%; dispersal limitation and drift together explained 21%–23%, while heterogeneous selection remained below 1%. In the dry season, homogeneous selection increased to 40%–65% and was the leading process at every site, with its largest contribution—about 65%—recorded upstream.

The authors associated the stronger upstream signal with relatively uniform forest inputs, limited pollution and carbonate-buffered chemistry, whereas variable agricultural and urban inputs at the disturbed sites corresponded with a smaller relative contribution from homogeneous filtering. These are associations inferred from the site comparisons rather than experimental demonstrations of the underlying filters, and unmeasured local variables could also have contributed. Fungi underwent substantial spatial and seasonal reorganization.

Wet-season upstream assemblages were enriched in terrestrial saprotrophic and plant-pathogenic Ascomycota, including Didymella, Cercospora, Cladosporium and Boeremia. At the midstream site, unclassified fungi exceeded 50% of wet-season reads, while dry-season samples contained approximately 20% Cladosporium. Rozellomycota were consistently enriched at the downstream site in both seasons.

ANOSIM detected significant site effects, with global R = 0.944 and p = 0.003 in the wet season and R = 0.972 and p = 0.004 in the dry season. The upstream fungal assemblage remained distinct, while midstream and downstream communities converged more closely. The study therefore identified the disturbance gradient as a stronger spatial influence than season on fungal beta diversity, while seasonal shifts chiefly altered taxon abundances and evenness.

Fungal assembly changed more sharply than its taxonomic composition. Homogeneous selection contributed 44%–46% of wet-season assembly events. In the dry season, its share fell to 10%–14%, while ecological drift increased to 46%–50% and became the dominant process; homogenizing dispersal also contributed more than 30% in some comparisons.

The authors proposed that low discharge and cold conditions may restrict the supply or establishment of fungal propagules, allowing random drift to become relatively more influential. They explicitly noted, however, that continuous discharge and temperature measurements and residence-time estimates were unavailable, preventing the independent effects of these covarying seasonal conditions from being disentangled. The result applies to the two sampled seasons and three sites, and the authors cautioned that the contrast with bacterial assembly may be specific to this karst system rather than directly transferable to non-karst rivers.

Environmental ordination linked both microbial communities to pH and total phosphorus. For bacteria, the first two redundancy-analysis axes explained 37.66% of constrained variation in the wet season and 53.96% in the dry season; total phosphorus and pH were significant variables in both analyses. For fungi, the corresponding totals were 42.07% and 51.36%, with total phosphorus and pH again associated with spatial separation.

Upstream samples consistently occupied low-pH, low-phosphorus regions, while midstream and downstream samples grouped toward higher values. PICRUSt2 predicted relatively greater representation of environmental-adaptation, cellular-stress and virulence-associated pathways in the wet season and relatively greater carbohydrate-, amino-acid- and energy-metabolism pathways in the dry season. These are sequence-based estimates of genomic capacity, not direct measurements of metabolic rates or realized activity in the river.

Fungal guild richness was also season dependent. Wet-season communities had limited resolved functional diversity, with unknown trophic assignments, plant pathogens and saprotrophs with unspecified substrate affinity accounting for roughly 80% of reads. In the dry season, guild richness expanded at the midstream and downstream sites, including generalist saprotrophs, undefined saprotrophs, animal pathogens and endophytes; the downstream site had the greatest dry-season functional-group richness.

The authors interpreted this pattern as possibly associated with accumulated allochthonous organic matter, although dissolved and particulate carbon were not quantified. Rozellomycota and the dry-season expansion of fungal guilds were proposed as preliminary candidate indicators of chronic downstream disturbance, but neither was validated against independent rivers or other seasons. The study supports considering both bacterial and fungal assemblages when assessing karst headwaters, while continuous hydrological monitoring, denser site sampling, direct organic-matter measurements, metatranscriptomics and controlled experiments will be needed to test the proposed mechanisms.

Subject of Research: Seasonal and spatial assembly of planktonic bacterial and fungal communities along a karst-river disturbance gradient

Article Title: Responses and assembly mechanisms of planktonic bacterial and fungal communities in the upper Chishui River during dry and wet seasons

Article References: Liu, D., Zheng, W., Wen, H., Yang, C., Qu, D., Zhang, K., Xie, Y., Ji, G., & Feng, Z. (2026). Responses and assembly mechanisms of planktonic bacterial and fungal communities in the upper Chishui River during dry and wet seasons. Frontiers in Microbiology, 17, Article 1949002. https://doi.org/10.3389/fmicb.2026.1949002

Image Credits: AI Generated

DOI: 10.3389/fmicb.2026.1949002

Keywords: Chishui River, karst headwaters, planktonic bacteria, planktonic fungi, community assembly, seasonal dynamics, nutrient gradient, homogeneous selection, ecological drift, microbial bioindicators, Responses, assembly