# Multi-guild microbial cooperation sustains long-term anaerobic toluene degradation through sulfur cycling

## TL;DR

A microbial community uses sulfur cycling to sustain long-term toluene breakdown in oxygen-free environments.

## Contribution

Discovery of a sulfur redox loop enabling stable anaerobic toluene degradation through microbial cooperation.

## Key findings

- A microbial consortium co-dominated by Desulfoprunum and Sulfurovum enables sustained toluene degradation.
- Genome-resolved metagenomics reveals a distributed sulfur redox loop involving multiple taxa.
- Sulfur cycling interactions buffer sulfide toxicity and stabilize redox dynamics for long-term degradation.

## Abstract

Anaerobic degradation of aromatic hydrocarbons such as toluene plays a critical role in the natural and engineered attenuation of contaminated environments. Here, we developed and characterized a microbial consortium enriched under strictly anoxic conditions, capable of sustained toluene degradation through sulfate reduction. By integrating biodegradation kinetics, long-read 16S rRNA profiling, and genome-resolved metagenomics, we elucidated the structure and function of a multi-guild community. The consortium was co-dominated by Desulfoprunum, a sulfate-reducing bacterium (SRB), and Sulfurovum-affiliated sulfur oxidizers (~34% each), with additional members including Stenotrophomonas, Achromobacter, and Stutzerimonas. Such co-dominance appears uncommon, as sulfate-reducing enrichments are often characterized by low diversity and the predominance of a single lineage, such as Desulfobacula or Desulfosarcina in marine systems. Genome-resolved analyses recovered seven metagenome-assembled genomes (MAGs) with distinct but complementary metabolic roles. Desulfoprunum encoded the fumarate-addition pathway (bss/bbs) for anaerobic toluene activation and dissimilatory sulfate reduction (aprAB, dsrAB). In contrast, Sulfurovum and several Gammaproteobacteria encoded sulfide:quinone oxidoreductase (sqr), coupling H2S detoxification to energy conservation, while a Moranbacterales MAG carried a putative sulfhydrogenase (hydAB) potentially catalyzing elemental sulfur (S°) reduction. Additional MAGs encoded assimilatory sulfate reduction (cys), suggesting integration of sulfur into biosynthetic pathways. Together, these features are consistent with the presence of a putative distributed sulfur redox loop, in which biogenic H2S may be recycled via oxidation and reduction reactions mediated by co-occurring taxa. This sulfur loop is hypothesized to contribute to buffering sulfide toxicity and stabilize redox dynamics, thereby potentially supporting long-term toluene degradation under sulfidic conditions. Our findings highlight anaerobic degradation as a community-driven process enabled by sulfur-cycling interactions. By revealing the role of cryptic sulfur cycling in stabilizing hydrocarbon degradation, this work offers a new framework for designing bioremediation strategies in contaminated anoxic environments.