Self-assembling a 1,4-dioxane-degrading consortium and identifying the key role of Shinella sp. through dilution-to-extinction and reculturing.

Assembling a functional consortium and identifying novel degraders from contaminated environments are still challenging due to the large diversity of microorganisms and the difficulty in isolating pure cultures. Here, we constructed a relatively simple functional consortium by enriching 1,4-dioxane-degrading-consortia using a culture-dependent dilution-to-extinction (DTE) method and reported a new key dioxane-degrader Shinella sp. Our results showed that serial dilution and reculture led to a divergence in the degradation ability of each consortium. Next-generation sequencing data revealed that the divergence in degradation performance was due to the reassembly of microbiota in the DTE process, which occurred most notably in 10 -8 and 10 -9 dilutions. The shift in community structure at 10 -9 prevented the recovery of 1,4-dioxane degradation capacity, and the newly dominant taxa, Xanthobacter and Acinetobacter , struggled to replace the original dominant genus Shinella for 1,4-dioxane biodegradation. Combining differential analysis of community structure and metabolic function, we confirmed that Shinella species have a stronger 1,4-dioxane degradation ability than Xanthobacter species in the enriched consortium. In addition, we verified our findings using our isolated dioxane-degrading bacteria, Shinella yambaruensis , resulting in the rapid recovery of degradation performance of a 10 -9 dilution consortium with Xanthobacter and Acinetobacter as the dominant microbiota. Taken together, this study provides a strategy for self-assembling functional consortiums and identifying the key degraders to explore the underlying biological mechanisms of enriched contaminant-degrading consortia.IMPORTANCEAssembling a functional microbial consortium and identifying key degraders involved in the degradation of 1,4-dioxane are crucial for the design of synergistic consortia used in enhancing the bioremediation of 1,4-dioxane-contaminated sites. However, due to the vast diversity of microbes, assembling a functional consortium and identifying novel degraders through a simple method remain a challenge. In this study, we reassembled 1,4-dioxane-degrading microbial consortia using a simple and easy-to-operate method by combining dilution-to-extinction and reculture techniques. We combined differential analysis of community structure and metabolic function and confirmed that Shinella species have a stronger 1,4-dioxane degradation ability than Xanthobacter species in the enriched consortium. In addition, a new dioxane-degrading bacterium was isolated, Shinella yambaruensis , which verified our findings. These results demonstrate that DTE and reculture techniques can be used beyond diversity reduction to assemble functional microbial communities, particularly to identify key degraders in contaminant-degrading consortia.