The decline in gut microbial diversity in modern humans is closely associated with the rising prevalence of various diseases. It is imperative to investigate the underlying causes of gut microbial loss and restoring methods. Although the impact of non-perinatal antibiotic use on gut microbiota has been recognized, its intergenerational effects remain unexplored. Our previous research has highlighted soil in the farm environment as a key factor for gut microbiome health by restoring gut microbial diversity and balance. In this study, we investigated the intergenerational consequences of antibiotic exposure and the therapeutic potential of sterile soil. We treated C57BL/6 mice with vancomycin and streptomycin for 2 weeks continuously, followed by a 4-8 week withdrawal period before breeding. The process was repeated across 3 generations. Half of the mice in each generation received an oral sterile soil intervention. We assessed gut microbial diversity, anxiety behavior, microglial reactivity, and gut barrier integrity across generations. Antibiotic exposure led to a decrease in gut microbial diversity over generations, along with aggravated anxiety behavior, microgliosis, and altered intestinal tight junction protein expression. Oral sterile soil intervention restored gut microbial diversity in adult mice across generations, concomitantly rescuing abnormalities in behavior, microgliosis, and intestinal barrier integrity. In conclusion, this study simulated an important process of the progressive loss of gut microbiota diversity in modern humans and demonstrated the potential of sterile soil to reverse this process. This study provides a theoretical and experimental basis for research and interventions targeting multiple modern chronic diseases related to intestinal microorganisms.

Soil has been considered the main microbial reservoir for plants, but the robustness of the plant microbiome when the soil resource is removed has not been greatly considered. In the present study, we tested the robustness of the microbiota recruited by Tartary buckwheat (Fagopyrum tataricum Gaertn.), grown on sterile humus soil and irrigated with sterile water. Our results showed that the microbiomes of the leaf, stem, root and next-generation seeds were comparable between treated (grown in sterile soil) and control plants (grown in non-sterile soil), indicating that the plants had alternative robust ways to shape their microbiome. Seed microbiota contributed greatly to endophyte communities in the phyllosphere, rhizosphere and next-generation seeds. The microbiome originated from the seeds conferred clear benefits to seedling growth because seedling height and the number of leaves were significantly increased when grown in sterilized soil. The overall microbiome of the plant was affected very little by the removal of the soil microbial resource. The microbial co-occurrence network exhibited more interactions, and Proteobacteria was enriched in the root of Tartary buckwheat planted in sterilized soil. Our research broadens the understanding of the general principles governing microbiome assembly and is widely applicable to both microbiome modeling and sustainable agriculture.

Exploring the assembly mechanism of microbiota is critical for understanding soil ecosystem functions. However, the relative importance of different biotic and abiotic factors in determining the bacterial community has not been properly clarified. In this study, the effects of inocula and recipients on the assembly of the soil community were investigated to evaluate their importance by inoculation experiments with sterile soil. Two distinct soils, conventional nitrogen-fertilized soil and aromatic-compound-contaminated soil, were sterilized, cross inoculated, and incubated for 2 months under different inoculation doses and oxygen conditions. The results showed that the greatest variation in community structure emerged in the samples inoculated with distinct inocula rather than in the samples of different soil recipients. The phylogenies in the two inocula were diverse and dissimilar, although there were many ecologically equivalent bacteria. When the inocula with dissimilar ecologically equivalent species were used, the assembled communities were primarily determined by the inocula as indicated by the beta diversity and variation partitioning analyses. In contrast, environmental selection dominated the process when ecologically equivalent species in the inocula were similar, as when only one type of inoculum was used, where the soil habitat selected the most adaptive bacteria from the defined inoculum pool. These results indicate that inoculated bacteria are dominant over environmental selection if they are sufficiently dissimilar, although the effect of environmental selection is more obvious when similar bacteria are inoculated in the soil for community assembly. Our findings suggest that the immigration of exotic bacteria could be a primary factor impacting community assembly.IMPORTANCE The soil microbiota conducts important biological ecosystem functions, but the mechanism underlying community-environment interactions for soil microbiota remains unclear. By using two distinct soils for cross inoculation, we successfully simulated the assembly of the bacterial community in sterile soil. Thus, the reasons why inoculum and recipient have dominated community assembly in previous investigations are investigated in this study. We found that inoculated bacteria presided over environmental selection for community assembly due to the varied difference of ecological equivalent bacteria, either divergent or convergent. The significance of neutrality for the ecologically equivalent bacterial species that immigrated into the recipients should be emphasized in exploring the mechanisms of community assembly. Our finding is helpful for understanding the community-environment interaction, a basic question in ecology, and it would shed light on this issue that has perplexed scientists for many years.