Objectives The partial or complete substitution of chemical fertilizers with livestock manure-derived organic fertilizers is a critical technical approach for the green production of vegetables. However, the differential effects of organic fertilizers from different sources and their substitution ratios on rhizosphere microecology and antibiotic resistance gene risks remain unclear. This study systematically explores the regulatory effects of equal-N substitution of chemical fertilizers by different proportions of chicken manure and pig manure on the structure and diversity of rhizosphere soil microbial communities and the abundance of key functional genes in greenhouse vegetable systems, so as to provide theoretical basis and technical support for the safe and rational application of organic fertilizers in vegetable planting systems, and offer scientific references for antibiotic resistance gene risk warning and ecological risk control in farmland soil.

Methods A pot experiment was conducted, using Chines cabbage as test materials and chicken manure (OF1) and pig manure (OF2) as organic fertilizers. The replace ratios of chemical fertilizer with OF1 and OF2 were established, including: 30%, 50%, 70%, and 100% (designated as OF1_30, OF1_50, OF1_70, OF1_100; OF2_30, OF2_50, OF2_70, and OF2_100, respectively), and a pure chemical fertilizer treatment was used as the control (CF), resulting in a total of nine fertilization treatments. After harvest of Chinese cabbage, rhizosphere soil samples were collected for determination of physicochemical properties, diversity of microbial communities, and abundances of functional genes.

Results Compared with CF treatment, organic fertilizer substitution had no significant effect on soil pH, EC, organic matter, and total P content. However, OF2_100 treatment significantly increased total N content, OF1_100 treatment significantly increased total Zn content. Organic fertilizer substitution effectively improved soluble sugar and vitamin C content while reducing nitrate, Cu, Zn, and Cd accumulation in vegetables (P<0.05). Organic fertilizer treatments significantly increased α diversity and altered β diversity of rhizosphere soil bacteria. Soil total N,P, Cu and Zn content were identified as key environmental factors driving the differentiation of microbial community structure. The type and substitution ratio of organic fertilizers were key factors driving functional differentiation in microbial communities: OF1treatments enriched more tolerant and metabolically specialized taxa such as Ohtaekwangia and Actinoplanes with increasing substitution ratios, whereas OF2_100 treatment primarily enriched Bacillus-related genera such as Virgibacillus and Gracilibacillus under high substitution ratios. These taxa contributed to enhancing soil stress tolerance and facilitating nutrient transformation. OF1_100 treatment significantly increased sul1 and sul2 abundances but decreased tetA abundance compared with CF (P<0.05). OF2 treatments did not significantly increase sul1 and sul2 abundances but decreased tetA abundance (P<0.05). No significant differences were observed in cbbL and nifH abundances among treatments.

Conclusions Replacing all chemical fertilizers with pig manure can significantly enhance the α-diversity of rhizosphere soil bacteria and alter the β-diversity. The enriched Bacillus taxa (such as Virgibacillus and Gracilibacillus) help strengthen soil stress resistance, promote organic matter decomposition and nutrient transformation, and further improve the stability of microbial functional communities. Pig manure organic fertilizer can notably reduce the abundance of the tetA gene and will not cause the enrichment of sulfonamide ARGs (sul1, sul2). The functional flora related to carbon and nitrogen cycling is also not significantly affected, showing relatively low environmental risk under the experimental conditions in this study. High-level chicken manure replacing chemical fertilizers will lead to the accumulation of soil Zn and the enrichment of sulfonamide resistance genes (sul1, sul2), but it shows a trend of reducing the abundance of the tetA gene. As the replacement ratio of chicken manure increases, the rhizosphere microbial community successively shifts from organic-degrading flora to tolerance and metabolic functional flora (such as Ohtaekwangia and Actinoplanes), which reflects the adaptive response of microorganisms to salt and heavy metal stress. Therefore, replacing all chemical fertilizers with chicken manure poses certain ecological risks.