Longfei Shu

Understanding how interspecific interactions shape ecosystem functioning is a fundamental and longstanding question in ecology, especially along an evolutionary trajectory. Here, we used experimental evolution to evaluate how microbial interactions and adaptive mutations shaped community-level functions in synthetic denitrifying communities (SDCs) along a species richness gradient (one, two, and six species). Our results showed that competitive microbial interactions were observed and significantly ( P < 0.05) enhanced denitrification rates in the six-species SDCs during experimental evolution. Genome and metagenome re-sequencing analysis revealed the highest fixed non-synonymous and frameshift mutations within the six-species SDCs, among genes involved in nutrient transport, DNA repair, and secretion pathways. AlphaFold2 protein structure prediction, molecular docking, and Evo 2 host fitness prediction indicated that the fixed mutations could improve host strain fitness, and partial least squares path modeling (PLS-PM) further revealed that interspecific competition accelerated adaptive mutations, thereby reinforcing community-level functions, indicating potential Red Queen dynamics. This study offers novel insights into our understanding of eco-evolutionary mechanisms linking species richness, microbial interactions, adaptive mutations, and community functions, with important implications for active nitrogen removal and greenhouse gas mitigation in diverse ecosystems. IMPORTANCE: Microbial diversity is widely recognized as a key driver of ecosystem functioning, yet how species interactions shape ecosystem functions through evolutionary processes remains poorly understood. Using synthetic denitrifying communities with different levels of species richness, we show that interspecific competition can accelerate adaptive mutations and enhance community-level denitrification over evolutionary timescales. Communities with higher species richness accumulated more adaptive mutations, particularly in genes associated with nutrient transport, DNA repair, and secretion systems, ultimately improving microbial fitness and denitrification performance. Our findings provide direct experimental evidence that microbial competition can promote functional adaptation, consistent with the Red Queen hypothesis. This study advances our understanding of the eco-evolutionary mechanisms linking species richness, microbial interactions, adaptive evolution, and ecosystem functioning.

Impact statement: Protists are an underexplored but functionally important component of aerobic‐activated granular sludge under pollution stress. Using metagenomics, we profiled protistan responses to ciprofloxacin, triclosan, and Cu ²⁺ (alone or in combination). Protists remained a stable 6.35%–7.88% of the bacterial community, and the consumers were the most abundant groups. Ciprofloxacin showed little effect on protist abundance, while Cu ²⁺ increased protist abundance, especially consumers. Stress conditions also strengthened predominantly positive protist–bacteria associations, suggesting cross‐domain interactions that may enhance community resilience. These results demonstrate that protists are key determinants in stabilizing microbial communities under multiple stressors.

Soil amoebae function as both keystone microbial predators and sensitive indicators of soil ecosystem health. Although polylactic acid (PLA) biodegradable plastics are increasingly adopted to reduce persistent plastic pollution, their ecological impacts, particularly during the active degradation phase, remain poorly understood. In this study, we established multiple co-culture interfaces to investigate the interactions and effects between soil amoebae and polylactic acid microplastics (PLA MPs). Our results show that PLA MPs fundamentally reshape amoeboid behavioral strategies and restructure soil microbial networks. Specifically, PLA MPs induce particle-size-dependent shifts in microbial community composition and functional potential, trigger strong avoidance behavior in amoebae that impairs its chemotactic capacity to locate bacterial prey, and disrupt the development of the PLA-associated plastisphere by suppressing microbial interactions. PLA MPs also impose significant fitness costs on amoebae, reducing spore production and altering developmental morphology. This study establishes soil amoebae as powerful bioindicators for evaluating the ecological impact of biodegradable plastics and demonstrates that PLA-driven disruption arises not only from its persistence but also from its degradation intermediates. These findings uncover hidden ecological trade-offs in the adoption of biodegradable plastics and offer key mechanistic insights to support environmental risk assessment.

Dictyostelium discoideum is a social amoeba that transitions from unicellular to multicellular forms in response to environmental signals, making it an intriguing model for studying cell aggregation and differentiation. Although previous studies have demonstrated that Dictyostelium slugs exhibit phototaxis, the mechanisms behind light-induced developmental changes remain unclear. In this study, we investigated how light triggers the transition to multicellularity and its associated metabolites and genes. Our findings revealed that spore yield depends on light exposure, with slower multicellular development under dark incubation. Transcriptomics analysis on QS9 amoebae identified upregulation of small GTPases such as rasD and racL in response to light, which likely promote cell movement, phagocytosis and actin protrusions. Light also enhanced cAMP production, driving the aggregation, post-aggregation and development of single cells. Additionally, c-di-GMP was essential for cell differentiation during multicellular growth and was upregulated by light. Metabolomic analysis on QS9 amoebae revealed that the downregulation of LPC (lysophosphatidylcholine) was detected under both unicellular and multicellular phases. Moreover, reduced levels of GSH (glutathione) in dark may impede multicellular structures of D. discoideum . These findings provide insights into light-triggered cell differentiation and pattern formation, offering a better understanding of molecular mechanisms underlying the transition to multicellularity in Dictyostelium cells.