A global survey of the bacteria within earthworm nephridia

Earthworms comprise 16 described families in the Crassiclitellata plus a few other minor groups. Microscopy studies of the early 20th century detected bacteria within the excretory organs, the nephridia, of species within a few of these families. More recent evidence for the consistent and specific association of bacteria with nephridia within the Lumbricidae has been well documented, but the presence and identity of nephridial bacteria among the rest of the Crassiclitellata families had not been explored. The study presented here aimed to identify members of Crassiclitellata families that harbor bacteria in their nephridia, and identify these bacteria based on 16S rRNA gene sequences. Eleven earthworm families were surveyed from countries of six continents, and two island nations. The results revealed members of four bacterial orders commonly occurred within nephridia of genera within nine Crassiclitellata families. Members of the bacterial phyla Bacteroidetes (order Sphingobacteriales), Betaproteobacteria (order Burkholderiales; family Comamonadaceae), and Alphaproteobacteria (orders Rhodospirillales and Rhizobiales) were detected in the nephridia of basal Crassiclitellata, as well as in derived families. Earthworm genera with meronephridia, multiple small nephridia per segment, lacked bacteria, whereas bacteria were often detected in holonephridia, single pairs of large nephridia with a distinct morphology and external excretory pore. The Acanthodrilidae members, a large derived family of earthworms, did not appear to possess nephridial bacteria regardless of nephridial form. Although earthworms from a variety of habitat types were sampled, there were no clear correlations of lifestyle with symbiont types, with the exception of the aquatic earthworms that contained bacteria unrelated to those in any other earthworms. The findings support an evolutionarily long association of bacteria within the Crassiclitellata, and suggest a contribution to nitrogen conservation for the earthworms.