Humic acid reduces the dissemination of antibiotic resistance genes by reshaping the soil microbiome in farmland soils

초록

Humic acid (HA) is widely used as a soil amendment, but its role in modulating antibiotic resistance in agricultural soils is still not clear. Here, we conducted 60-day microcosm experiments using soils contaminated with sulfamethazine and tetracycline. We added HA at 0, 0.1, 1, 5, and 10 g·kg−1 and quantified its effects on culturable bacteria, antibiotic-resistant bacteria (ARB), antibiotic resistance genes (ARGs), and community assembly. In sulfamethazine-treated soils (S_HA), HA led to a strong decline in SRB and resistance frequency. At 5 g·kg−1, the richness of the sulfonamide-resistant community decreased by 74.5%. In tetracycline-treated soils (T_HA), HA addition produced a distinct pattern in which total culturable bacteria and ARB abundance increased, whereas resistance frequency decreased, suggesting that HA expanded the whole culturable bacterial pool rather than selectively enriching resistant bacteria. Across both antibiotics, HA strongly restructured resistant communities, with HA concentration emerging as the primary driver of β-diversity. Quantitative PCR revealed time- and dose-dependent suppression of tetM , tetR , sul1 , and sul2 , and the class 1 integron gene intI1 declined by up to 90.48% at the initial stage under HA amendment. Niche-breadth analyses further showed that a weakening of deterministic selection was observed under high HA in S_HA. The combined evidence from soil physicochemical properties and SEM indicates that the effect of HA on antibiotic resistance is largely indirect, and by reconfiguring the soil environment and microbiome that host and transmit ARGs. Together, these results show that HA regulates antibiotic resistance spread by altering antibiotic bioavailability, community structure, and mobile genetic elements.

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