Coupling multifactor dominated the biochemical response and the alterations of intestinal microflora of earthworm Pheretima guillelmi due to typical herbicides.

The excessive application of herbicides on farmlands can substantially reduce labor costs and increase crop yields, but can also have undesirable effects on terrestrial ecosystems. To evaluate the ecological toxicity of herbicides, metolachlor and fomesafen, two typical herbicides that are extensively used worldwide were chosen as target pollutants, and the endogeic earthworm Pheretima guillelmi, which is widely distributed in China, was selected as the test organism. A laboratory-scale microcosmic experiment was set, and energy resources, enzymes, and the composition and connections of intestinal microorganisms in earthworms were determined. Both herbicides depleted the energy resources of the earthworms, especially glycogen contents; increased the levels of antioxidant enzymes; and inhibited acetylcholinesterase. Moreover, the richness and diversity of the intestinal bacterial community of the earthworms were suppressed. Additionally, the bacterial composition at the genus level changed greatly and the connections between dominant bacteria increased dramatically. Most interactions among the bacterial genera belonging to the same and different phyla showed mutualism and competition, respectively. Importantly, metolachlor with higher toxicity had a transitory effect on these indicators in earthworms, whereas fomesafen, with lower toxicity but stronger bioaccumulation potential, exerted a sustaining impact on earthworms. Collectively, these results indicate that the toxic effects of herbicides on terrestrial organisms should be comprehensively considered in combination with biological toxicity, persistence, bioaccumulation potential, and other factors.