Under the “One Health” framework, untreated livestock waste serves as a critical vector for the cross-domain proliferation of antibiotic resistance genes (ARGs). While integrated anaerobic digestion-composting systems are increasingly adopted for agricultural waste treatment, the dynamic succession of ARGs and the regulatory roles of quorum sensing (QS) and viral communities during the drastic environmental transitions remain critically unexplored. We applied comprehensive viromic and metagenomic analyses to track the dynamic fate of ARGs, ARB (antibiotic resistance bacteria), QS, and viral communities across a single campaign of full-scale anaerobic digestion-composting system treating cattle slurry. The relative abundance of total ARG (RPKG) was higher in cattle slurry digestate than in raw cattle slurry and lower in mature compost than in compost feedstock. The relative abundance of VFG-carrying contigs (RPKM) was also lower in mature compost than in compost feedstock. However, specific high-risk ARGs, notably aac(3)-VIIa and sul1, exhibited extreme recalcitrance. At the maturation stage, we revealed that thermotolerant taxa, predominantly Bacillota and Actinomycetota, dominated the ultimate high-risk ARB. Crucially, the carriage of specific QS regulatory genes (LuxR and DGC) within ultimate high-risk ARB exhibited a highly significant positive correlation (R2 > 0.82, P < 0.001) with ARG and virulence factor gene (VFG) abundances. Furthermore, while lytic phages dominated the viral community, the thermophilic stage coincided with a marked decoupling of phage and ARB-host abundance trajectories, consistent with reduced viral abundance under thermal stress. Within this campaign, mean ARG abundance (RPKG) was higher in cattle slurry digestate than in raw cattle slurry. Mean ARG and VFG abundances (RPKG) were lower in mature compost than in compost feedstock. Nevertheless, aac(3)-VIIa, sul1, and tet(M), together with ARG-carrying MAGs affiliated with Bacillota and Actinomycetota, remained detectable in mature compost. QS (luxR and DGC) is associated mainly with persistence and potential horizontal gene transfer, whereas phages act chiefly as ecological predators and rarely carry ARGs, so that resistant-bacteria persistence coincided with weaker virus–host coupling at high temperature, rather than providing evidence for transduction. These observations suggest biologically targeted interventions (e.g., QS inhibitors or sequential phage inoculation) as candidate strategies whose feasibility and efficacy remain to be tested across different feedstocks, seasons, and full-scale operating conditions before any general management recommendation can be made.
Dimethyl sulfide (DMS) is a volatile organosulfur compound that contributes to global climate regulation through its role in atmospheric aerosol formation. The Southern Ocean is an important source of DMS and represents a key region for marine sulfur cycling. However, previous studies have primarily focused on pelagic ecosystems, and the microbial drivers of DMS cycling in Antarctic coastal sediments remain poorly understood. Here, we integrated 16S and 18S rRNA gene amplicon sequencing, metagenomics, and DMSP-amendment incubation to characterize the DMS cycling in intertidal sediments of the climate-sensitive Fildes Peninsula. Our results revealed notable potential for DMS production via the DMSP cleavage pathway, in contrast to the demethylation-dominated pelagic ocean. Furthermore, the incubation experiments confirmed rapid DMS generation upon the addition of DMSP. Granulosicoccus, Ilumatobacter, and Cocleimonas were the potential drivers of DMS cycling, with genomic reconstruction demonstrating their roles in the biogeochemical cycles of sulfur and nitrogen. Our findings identify Antarctic intertidal sediments as potential hotspots of DMS production mediated by the DMSP cleavage pathway and Granulosicoccus-dominated microbial communities, necessitating their inclusion in polar biogeochemical models under climate change.
Stress consistently disrupts the vertebrate gut microbiome, leading to negative health outcomes. However, how systemic stress translates into reproducible microbial dysbiosis remains poorly understood. While current research primarily focuses on late-stage compositional shifts, the early biochemical signals and microenvironmental alterations that precede and initiate this structural breakdown represent a critical knowledge gap. Here we show that stress destabilizes the gut microbiome through a cascade initiated by intestinal reactive oxygen species (ROS). In zebrafish, stress rapidly elicits intestinal ROS accumulation in the distal intestine, emerging within hours and preceding any detectable changes in microbial composition. This early oxidative response does not immediately restructure the community but instead induces a functional collapse: anaerobic fermentation is selectively suppressed and short-chain fatty acids (SCFAs) production sharply declines while community structure remains largely intact. The resulting SCFAs deficit weakens PPARγ-dependent epithelial metabolic programs, diminishes epithelial oxygen consumption, and erodes the physiological hypoxic barrier. Loss of this barrier re-engineers the luminal niche, enabling the relative enrichment of facultative pathobionts. Restoring epithelial bioenergetics or neutralizing intestinal ROS in zebrafish preserves hypoxia and prevents dysbiosis despite continued stress, identifying intestinal ROS as an early initiating signal of stress-induced dysbiosis. In restraint-stressed mice, the same ordered sequence—early ROS elevation, SCFAs decline, hypoxia loss, and the relative enrichment of facultative taxa—is likewise observed, supporting phenotypic and temporal conservation of this cascade. Our findings reveal that stress-induced dysbiosis is not a stochastic microbial drift, but a highly reproducible ecological cascade that is triggered by an early intestinal ROS burst in zebrafish and phenotypically conserved in mice. By shifting the paradigm from late-stage structural dysbiosis to the initial functional and microenvironmental collapse, we identify the maintenance of the intestinal hypoxic barrier as a critical therapeutic target for stress-related microbiome disorders.
The conventional ‘isolate-and-screen’ strategy for probiotic discovery often lacks ecological context, resulting in strains with uncertain safety profiles and unpredictable functional performance. Indigenous pig breeds, particularly those thriving under low-input husbandry conditions, represent underexplored ‘living libraries’ that offer host- and environment-shaped gut niches. Tunchang (TC) pigs, an indigenous breed from Hainan Province in southeastern China, provide a useful model for top-down, ecology-guided mining of host-derived probiotics from native gut niches. By combining cross-sectional age-gradient metagenomics with cross-breed comparisons, we defined a TC-pig gut niche with a lower overall burden of annotated VF/ARG features than that of Duroc × Landrace × Yorkshire (DLY) pigs. Notably, Limosilactobacillus reuteri was consistently detected within a lactobacilli-enriched core microbiota across developmental-stage groups and showed limited associations with annotated VF/ARG features at the community level. This niche was associated with higher relative levels of several host–microbe co-metabolites, including indoleacrylic acid and N-decanoylglycine. Guided by this framework, we isolated a TC pig-derived L. reuteri strain (TC-Lr) and a co-isolated Lactobacillus johnsonii strain (TC-Lj). Targeted profiling of cell-free culture supernatants revealed a distinct tryptophan-associated metabolic profile of these two isolates, with TC-Lr showing higher concentrations of indole-3-lactic acid and tryptophol than TC-Lj. In a mouse model of dextran sulphate sodium (DSS)-induced colitis, TC-Lr, but not TC-Lj, alleviated weight loss and disease activity, attenuated colon shortening and reduced inflammation-associated haematological alterations. TC-Lr also partially restored DSS-disrupted colonic metabolism, increasing adenosine while reducing creatinine and histamine levels. These findings support an ecology-guided, top-down framework for prioritising host-derived probiotics from indigenous livestock breeds. They further suggest that the TC pig gut, characterised by a lactobacilli-enriched community and a lower annotated VF/ARG burden at the community level, may serve as an ecological source for prioritising host-derived probiotic candidates.
Most antibiotic resistance genes (ARGs) evolved in environmental microbes long before humanity's antibiotic breakthrough, and widespread antibiotic use accelerated the dissemination of ARGs among clinical pathogens. Although environmental ARG classes, such as β-lactamases, are widely discussed, their presence and distribution remain underexplored, with existing studies often lacking the scalability and taxonomic resolution required for comprehensive analysis. In this study, we present a comprehensive map of the β-lactamase resistome using an integrated bioinformatics and experimental workflow. We identify a vast reservoir of putative β-lactamase sequences spanning all four β-lactamase classes—more than 12-fold the number listed in current ARG databases. We generated a β-lactamase taxonomy-environment map by identifying 108,000 putative β-lactamases across diverse bacterial phyla and environmental ecosystems. Notably, the relative frequency of putative β-lactamases was only 3.9-fold lower than that of the essential recA gene across diverse environmental ecosystems, with signals of relative enrichment observed in wastewater and plant-associated samples. The enrichment in plant-associated and wastewater samples suggests that these environments may represent important reservoirs of putative β-lactamases. We identify taxonomic and environmental contexts that may represent candidate reservoirs of close homologs of clinically relevant β-lactamases, extending beyond wastewater to plant-associated, terrestrial, and aquatic settings. Overall, we present a comprehensive workflow and reusable mapping tool to identify and classify the β-lactamase resistome, revealing its diversity, distribution, and a largely unexplored reservoir with potential for mobilization into clinical pathogens.
