Shewanella is a putative producer of polyunsaturated fatty acids in the gut soil of the composting earthworm Eisenia fetida.

Polyunsaturated fatty acids (PUFAs) play a crucial role in aiding bacteria to adapt to extreme and stressful environments. While there is a well-established understanding of their production, accrual, and transfer within marine ecosystems, knowledge about terrestrial environments remains limited. Investigation of the intestinal microbiome of earthworms has illuminated the presence of PUFAs presumably of microbial origin, which contrasts with the surrounding soil. To comprehensively study this phenomenon, a multi-faceted approach was employed, combining fatty acid analysis with amplicon sequencing of the PfaA-KS domain of the anaerobic fatty acid synthase gene (pfa), as well as the 16S rRNA and 18S rRNA genes. This methodology was applied to scrutinize the gut microbiome of Eisenia fetida, its compost-based dietary source, and the resultant castings. This study unveiled a distinct gut soil ecosystem from input compost and output castings in fatty acid profile as well as type and abundance of organisms. 16S sequencing provided insights into the microbial composition, showing increased relative abundance of certain Pseudomonadota, including Shewanellaceae, and Planctomycetota, including Gemmataceae within the gut microbiome compared to input bulk soil compost, while Actinomycetota and Bacillota were relatively enriched compared to the casted feces. Sequencing of the PfaA-KS domain revealed amplicon sequence variants (ASVs) belonging primarily to Shewanella. Intriguingly, the 20C PUFAs were identified only in gut soil samples, though PfaA-KS sequence abundance was highest in output castings, indicating a unique metabolism occurring only in the gut. Overall, the results indicate that Shewanella can explain PUFA enrichment in the gut environment because of the pfa gene presence detected via PfaA-KS sequence data.IMPORTANCEPrior research has demonstrated that earthworm microbiomes can potentially harbor polyunsaturated fatty acids (PUFAs) that are not found within their residing soil environment. Moreover, distinct indicator species have been pinpointed for various microbial genera in earthworm microbiomes. Nevertheless, none of these studies have integrated metataxonomic and fatty acid analyses to explore the origin of PUFA synthesis in any earthworm species, with the objective of identifying the specific organisms and locations responsible for this production. This study suggests that earthworms accumulate PUFAs produced from bacteria, especially Shewanella, activated through the gut ecosystem.