# Assembly and co-occurrence networks of nitrogen-fixing bacteria associated with epiphyllous liverworts in fragmented tropical forests

## TL;DR

This study explores how habitat fragmentation affects nitrogen-fixing bacteria in liverworts in the Amazon, finding that smaller forest fragments disrupt microbial community stability.

## Contribution

The study reveals how habitat fragmentation influences the assembly and network structure of nitrogen-fixing bacterial communities in epiphyllous liverworts.

## Key findings

- Host-specific associations were observed, with C. surinamensis hosting Hassallia and R. flaccida hosting Fischerella.
- Smaller forest fragments showed more modular bacterial communities with fewer hubs and reduced robustness.
- Habitat size influences the stability and specificity of liverwort-associated diazotrophs.

## Abstract

Understanding the spatial dynamics of plant-associated microbial communities is increasingly urgent in the context of habitat loss and the biodiversity crisis. However, the influence of reduced habitat size and connectivity on the assembly mechanisms underlying microbial associations is fundamental to advancing microbial ecology and conservation. In the Brazilian Amazon, we investigated nitrogen-fixing (diazotrophic) bacterial communities associated with two epiphyllous liverworts, Cololejeunea surinamensis and Radula flaccida, across 11 forest sites within the Biological Dynamics of Forest Fragments Project landscape. Using amplicon sequencing targeting the nitrogenase gene (nifH), we characterized diazotroph community diversity, inferred assembly mechanisms through null models, and analyzed co-occurrence network structure. Host-specific associations were evident: C. surinamensis predominantly hosted Hassallia, while R. flaccida was primarily associated with Fischerella. Despite habitat fragmentation, diazotrophic richness and composition remained similar across habitats of different sizes, consistent with strong homogenizing dispersal. Network analyses revealed that smaller fragments harbored more modular communities with fewer module hubs, pronounced shifts in key species relative abundance, and reduced network robustness. Our findings underscore the influence of habitat size on the stability of liverwort-associated diazotrophs, with smaller fragments exhibiting lower key species specificity and disruption of microbe-microbe interactions. Our results emphasize the importance of conserving large, connected forest habitats to maintain the functional integrity of phyllosphere N-fixing microbiota.

Graphical Abstract