The nitrogen-fixing fern Azolla has a complex microbiome characterized by varying degrees of cophylogenetic signal.

Abstract

Premise: Azolla is a genus of floating ferns that has closely evolved with a vertically transmitted obligate cyanobacterium endosymbiont-Anabaena azollae-that fixes nitrogen. There are also other lesser-known Azolla symbionts whose role and mode of transmission are unknown.

Methods: We sequenced 112 Azolla specimens collected across the state of California and characterized their metagenomes to identify the common bacterial endosymbionts and assess their patterns of interaction.

Results: Four genera were found across all samples, establishing that multiple Azolla endosymbionts were consistently present. We found varying degrees of cophylogenetic signal across these taxa as well as varying degrees of isolation by distance and of pseudogenation, which demonstrates that multiple processes underlie how this endosymbiotic community is constituted. We also characterized the entire Azolla leaf pocket microbiome.

Conclusions: These results show that the Azolla symbiotic community is complex and features members at potentially different stages of symbiosis evolution, further supporting the utility of the Azolla microcosm as a system for studying the evolution of symbioses.