Leaf Microbiome Differences Between Winter (Invasive) and Summer (Non-invasive) Annual Growth forms of Anthemis cotula L.: Insights into Invasiveness

Background Biological invasions are major drivers of global change, and plant-associated microbiomes are increasingly recognized as potential mediators of invasive success. While belowground microbial associations have been widely explored, the role of aboveground endophytes in shaping plant performance and ecological strategies remains poorly understood—particularly in seasonally dynamic mountain ecosystems. Anthemis cotula L., a widespread invader in the Kashmir Himalaya, occurs in two distinct growth forms: a winter annual that is highly invasive and a summer annual that is non-invasive. This unique intraspecific contrast offers an opportunity to assess how microbiome composition may vary with seasonal and ecological strategy, while controlling for phylogenetic differences. We hypothesized that seasonal shifts in leaf endophytic communities would parallel the contrasting ecological strategies of the two growth forms, thereby offering insights into possible relationships between microbiome structure and plant performance in a mountain environment. Results Sequencing revealed 420 bacterial species and 3,036 fungal OTUs spanning both winter and summer annuals. Alpha-diversity patterns were contrasting: the non-invasive summer annual harboured higher bacterial richness, Shannon diversity, and evenness, while invasive winter annuals showed notably greater fungal diversity. Beta-diversity analyses and ordination (NMDS, PERMANOVA) demonstrated strong seasonal and growth form-specific structuring, with clear separation and little overlap between bacterial communities of winter vs. summer forms, and more pronounced fungal diversity within winter. Differential abundance testing (ANCOM-BC2) identified enrichment of stress-tolerant bacteria (e.g., Massilia, Nocardioides) and cold-adapted or mutualistic fungi (e.g., Coniosporium, Cladosporium ) in winter annuals, while summer forms were associated with bacterial taxa supporting growth in milder conditions (e.g., Methylobacterium ). These contrasting patterns indicate that the invasive winter annual’s microbiome, though less diverse for bacteria and richer for fungi, may be compositionally structured in ways that could support nutrient uptake, abiotic stress tolerance, and competitive performance under harsher conditions. Conclusion This study shows that A. cotula harbors distinct, seasonally structured leaf microbiomes across its winter and summer growth forms. The invasive winter annual is associated with a functionally enriched bacterial and a more diverse, stress-tolerant fungal community, potentially supporting resilience and early competitive advantage. By relating microbial composition to plant life-history variation, our findings suggest that aboveground microbiome seasonality is an underexplored but potentially important aspect of plant–environment interactions in mountain ecosystems.