Gut diazotrophs in lagomorphs are associated with season but not altitude and host phylogeny.

Invertebrates such as termites feeding on nutrient-poor substrate receive essential nitrogen by biological nitrogen fixation of gut diazotrophs. However, the diversity and composition of gut diazotrophs of vertebrates such as Plateau pikas living in nutrient-poor Qinghai-Tibet Plateau remain unknown. To fill this knowledge gap, we studied gut diazotrophs of Plateau pikas (Ochotona curzoniae) and its related species, Daurian pikas (Ochotona daurica), Hares (Lepus europaeus) and Rabbits (Oryctolagus cuniculus) by high-throughput amplicon sequencing methods. We analyzed whether the gut diazotrophs of Plateau pikas are affected by season, altitude, and species, and explored the relationship between gut diazotrophs and whole gut microbiomes. Our study showed that Firmicutes, Spirochaetes, and Euryarchaeota were the dominant gut diazotrophs of Plateau pikas. The beta diversity of gut diazotrophs of Plateau pikas was significantly different from the other three lagomorphs, but the alpha diversity did not show a significant difference among the four lagomorphs. The gut diazotrophs of Plateau pikas were the most similarly to that of Rabbits, followed by Daurian pikas and Hares, which was inconsistent with gut microbiomes or animal phylogeny. The dominant gut diazotrophs of the four lagomorphs may reflect their living environment and dietary habits. Season significantly affected the alpha diversity and abundance of dominant gut diazotrophs. Altitude had no significant effect on the gut diazotrophs of Plateau pikas. In addition, the congruence between gut microbiomes and gut diazotrophs was low. Our results proved that the gut of Plateau pikas was rich in gut diazotrophs, which is of great significance for the study of ecology and evolution of lagomorphs.