Multi-omics provide mechanistic insight into the Pb-induced changes in tadpole fitness-related traits and environmental water quality.

Water pollution from lead/Pb2+ poses a significant threat to aquatic ecosystems, and its repercussions on aquatic animals have received considerable attention. Although Pb2+ has been found to affect numerous aspects of animals, including individual fitness, metabolic status, and symbiotic microbiota, few studies have focused on the associations between Pb2+-induced variations in fitness, metabolome, symbiotic microbiome, and environmental parameters in the same system, limiting a comprehensive understanding of ecotoxicological mechanisms from a holistic perspective. Moreover, most ecotoxicological studies neglected the potential contributions of anions to the consequences generated by inorganic lead compounds. We investigated the effects of Pb(NO3)2 at environmentally relevant concentrations on the Rana omeimontis tadpoles and the water quality around them, using blank and NaNO3-treated groups as control. Results showed that Pb(NO3)2 not only induced a rise in water nitrite level, but exposure to this chemical also impaired tadpole fitness-related traits (e.g., growth and development). The impacts on tadpoles were most likely a combination of Pb2+ and NO3-. Tissue metabolomics revealed that Pb(NO3)2 exposure influenced animal substrate (i.e., carbohydrate, lipid, and amino acid) and prostaglandin metabolism. Pb(NO3)2 produced profound shifts in gut microbiota, with increased Proteobacteria impairing Firmicutes, resulting in higher aerobic and possibly pathogenic bacteria. NaNO3 also influenced tadpole metabolome and gut microbiome, in a manner different to that of Pb(NO3)2. The presence of NO3- seemed to counteract some changes caused by Pb2+, particularly on the microbiota. Piecewise structural equation model and correlation analyses demonstrated connections between tissue metabolome and gut microbiome, and the variations in tadpole phenotypic traits and water quality were linked to changes in tissue metabolome and gut microbiome. These findings emphasized the important roles of gut microbiome in mediating the effects of toxin on aquatic ecosystem. Moreover, it is suggested to consider the influences of anions in the risk assessment of heavy metal pollutions.

Cascading effects of Pb on the environmental and symbiotic microbiota and tadpoles' physiology based on field data and laboratory validation.

Heavy metal pollution poses a serious threat to ecosystems. Currently, there is a lack of field data that would enable us to gain a systematic understanding of the influences of heavy metals on aquatic ecosystems, especially the interactions between environments and animals. We studied the relationships between the variations in heavy metal concentrations (10 species including Pb in sediments and surface water), the community structure of environmental and symbiotic microbiota, and the gut traits of Bufo gargarizans tadpoles across 16 sampling sites on the Chengdu Plain through rigorous statistical analysis and laboratory validation. The results show that heavy metal concentrations, especially the Pb concentration of the sediment, are linked to the variations in sediment and tadpoles' gut microbiomes but not to water microbiota. For the sediment microbiota, Pb causes a trade-off between the proportions of Burkholderiales and Verrucomicrobiae and affects the methane, sulfide, and nitrate metabolisms. For tadpoles, a high sediment Pb content leads to a low abundance of gut aerobic bacteria and a large relative gut weight under both field and laboratory conditions. In addition, Pb promotes the growth of B. gargarizans tadpoles under laboratory conditions. These effects seem to be beneficial to tadpoles. However, a high Pb content leads to a low abundance of probiotic bacteria (e.g., Verrucomicrobiae, Eubacteriaceae, and Cetobacterium) and a high abundance of pathogenic bacteria in the gut and environment, suggesting potential health risks posed by Pb. Interestingly, there is a causal relationship between Pb-induced variations in sediment and symbiotic microbiotas, and the latter is further linked to the variation in relative gut weight of tadpoles. This suggests a cascading effect of Pb on the ecosystem. In conclusion, our results indicate that among the heavy metals, the Pb in sediment is a critical factor affecting the aquatic ecosystem through an environment-gut-physiology pathway mediated by microbiota.