Abstract: The innovative treatment of β-lactam residues has become a pressing concern for biopharmaceutical firms. This study determined if resource-based products of β-lactam residues meet the standards of Organic Fertilizer (NY 525-2021), and if the application will significantly affect crop inter-root soil microorganisms and lead to the enrichment of antibiotic resistance genes (ARGs). Study chose cephalosporin residue treated by hydrothermal spray-dried (HT+SD) and penicillin residue treated by multi-plate dryer (MD). A blank group and fertilizer treatment group served as controls. Two fertilizer dosing rates, a high application rate (1%) and a low application rate (0.5%), were used for potting experiments in each fertilizer treatment. Using 16S rRNA and HT-qPCR, the effects of mycorrhizal fertilization on lettuce inter-root soil bacterial diversity, species composition, ARGs, and mobile genetic elements (MGEs) were investigated. The results indicate that the residual antibiotics in the harmlessly treated residue are well below the detection limit of HPLC-MS (<6.78 μg/kg), the water content has been reduced by 95%, and the indexes of heavy metals and organic matter meet the requirements of Organic Fertilizer (NY 525-2021). Compared to the amount of fertilizer applied, the effect of fertilizer application type on soil bacterial community structure was more pronounced , as the Shannon-Wiener index was significantly higher (P < 0.05) in both residue treatment groups than in the chemical fertilizer treatment group. Both residues did not affect the bacterial composition of the soil, with Proteobacteria being the most abundant phylum (34.8%~48.1%) in all treatment groups. The number and total relative abundance of ARGs were significantly higher in the HP. Low application rate cephalosporin residue groups compared to the control group (P < 0.05). In contrast, the number and relative abundance of ARGs in the low application rate penicillin residue and HP treatment groups were not significantly different from the blank control group (P > 0.05), however, neither residue application altered the soil bacterial resistance mechanisms. The changes in ARGs were associated with introducing exogenous DNA and soil characteristic bacteria, with Pseudomonas spp. Exhibiting extremely strong correlations with three ARGs, mexE, ttgA and ttgB (correlation coefficients of 0.93, 0.90 and 0.84, respectively). Studies show that antibiotic residues treated with HT+SD and MD technologies can be repurposed as safe and effective alternative agricultural fertilizers.