摘要
      抗生素抗性基因（ARGs）通过结合的方式传播给公众健康带来了巨大挑战。焦木脂酸（PA）作为土壤改良剂的应用已被证明是修复土壤中ARGs污染的一种实用策略。然而，人们对PA通过偶联对ARGs水平基因转移（HGT）的影响知之甚少。本研究研究了在450°C下制备的木质废物衍生PA及其三种蒸馏组分（F1、F2和F3）在不同温度（98、130和220°C）下对质粒RP4在大肠杆菌内偶联转移的影响。在30 mL交配系统中，相对较高量（40–100μL）的PA抑制了74–85%的结合，遵循PA>F3≈F2≈F1的顺序，证明了PA改良剂可以通过抑制HGT来减轻土壤ARG污染的假设。PA的抗菌成分，包括酸、酚和醇，以及其酸度（pH 2.81）引起的抑菌作用有助于抑制结合。然而，在同一交配系统中，相对较低量（10-20μL）的PA使ARG转移增强了26-47%，其顺序为PA>F3≈F2>F1。低剂量时的相反作用主要归因于细胞内活性氧产生的增加、细胞膜通透性的增强、细胞外聚合物含量的增加和细胞表面电荷的减少。我们的研究结果强调了PA修饰物对ARG结合的刺激作用（低量促进和高量抑制），并为选择适量的PA修饰物来控制土壤ARG的传播提供了证据。此外，促进的结合还引发了关于土壤改良剂（如PA）通过HGT传播ARGs的潜在风险的问题。
Abstract
Spread of antibiotic resistance genes (ARGs) by conjugation poses great challenges to public health. Application of pyroligneous acids (PA) as soil amendments has been evidenced as a practical strategy to remediate pollution of ARGs in soils. However, little is known about PA effects on horizontal gene transfer (HGT) of ARGs by conjugation. This study investigated the effects of a woody waste-derived PA prepared at 450°C and its three distillation components (F1, F2, and F3) at different temperatures (98, 130, and 220°C) on conjugative transfer of plasmid RP4 within Escherichia coli. PA at relatively high amount (40–100 μL) in a 30-mL mating system inhibited conjugation by 74–85%, following an order of PA > F3 ≈ F2 ≈ F1, proving the hypothesis that PA amendments may mitigate soil ARG pollution by inhibiting HGT. The bacteriostasis caused by antibacterial components of PA, including acids, phenols, and alcohols, as well as its acidity (pH 2.81) contributed to the inhibited conjugation. However, a relatively low amount (10–20 μL) of PA in the same mating system enhanced ARG transfer by 26–47%, following an order of PA > F3 ≈ F2 > F1. The opposite effect at low amount is mainly attributed to the increased intracellular reactive oxygen species production, enhanced cell membrane permeability, increased extracellular polymeric substance contents, and reduced cell surface charge. Our findings highlight the hormesis (low-amount promotion and high-amount inhibition) of PA amendments on ARG conjugation and provide evidence for selecting an appropriate amount of PA amendment to control the dissemination of soil ARGs. Moreover, the promoted conjugation also triggers questions regarding the potential risks of soil amendments (e.g., PA) in the spread of ARGs via HGT.

https://pubs.acs.org/doi/full/10.1021/acsenvironau.2c00056