Background: Corn stover is a globally abundant agricultural byproduct with significant potential as livestock feed. Ensiling is a crucial method for its preservation, relying on microbial fermentation. However, a deep understanding of the complex bacterial community structure, their interactions, functional roles, and associated pathogenic risks during corn stover ensiling remains limited. This study aimed to elucidate these microbial dynamics to improve fermentation efficiency and ensure feed safety.

Methods: Corn stover was ensiled in laboratory-scale silos for up to 60 days. Fermentation quality was assessed by measuring pH, organic acids, and ammonia nitrogen. The bacterial community dynamics were profiled using high-throughput 16S rRNA gene sequencing. Bioinformatic analyses were employed to determine microbial diversity and succession, construct co-occurrence networks to identify keystone taxa, and predict the functional potential of the microbiome using PICRUSt2 based on the KEGG database.

Results: The ensiling process was successful, characterized by a rapid pH drop to below 4.0 and a high concentration of lactic acid. Microbial community analysis revealed a distinct temporal succession, with initial epiphytic Proteobacteria being replaced by a dominant Firmicutes population. Specifically, Lactiplantibacillus (formerly Lactobacillus) emerged as the most abundant genus in mature silage. Co-occurrence network analysis identified several keystone taxa that likely drive community stability. Functional predictions showed a significant enrichment of pathways related to carbohydrate metabolism, consistent with vigorous lactic acid fermentation. Notably, genera containing opportunistic pathogens, such as Pseudomonas and Clostridium, were detected, though their relative abundance decreased significantly in the well-fermented silage.

Conclusion: This study provides a comprehensive ecological perspective on the corn stover silage microbiome. Integrating community structure, network interactions, and functional predictions offers a deeper understanding of the fermentation process. The findings highlight the importance of promoting LAB dominance to not only ensure preservation but also to mitigate the risks associated with potential pathogens, thereby enhancing the overall safety and quality of silage. 

Corn stover, Silage, Microbial community, 16S rRNA sequencing

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