摘要

      质粒介导的抗生素抗性基因（ARGs）在细菌之间的水平转移促进了抗生素抗性的进化和传播。广泛的宿主质粒可以转移到土壤、植物根际和废水处理厂中的不同细菌宿主。尽管堆肥是将有机废物转化为肥料并减少某些ARG的有效方法，但很少有研究关注其对堆肥过程中携带ARG的质粒及其细菌宿主群落传播的影响。在本研究中，将携带携带三个ARG的宽宿主范围质粒RP4的荧光标记的恶臭假单胞菌（P.putida）接种到原料微宇宙中，并在不同的嗜热期持续时间下进行堆肥。研究了供体和RP4在堆肥过程中的命运。长期高温堆肥去除了95.1%的dsRed和98.0%的gfp，并在成熟阶段抑制了恶臭假单胞菌和RP4的反弹。堆肥后，RP4的传播潜力从每个受体的10−4降低到10−6转偶联物。此外，我们使用荧光激活细胞分选结合16S rRNA基因扩增子测序对RP4受体细菌的组成进行了分选和分析。RP4的受体细菌属于8个门，厚壁菌门占75.3%-90.1%，是跨偶联物中的优势门。RP4受体群落的多样性和丰富度因嗜热期延长而显著降低。总的来说，这些发现为评估堆肥在减缓质粒介导的ARGs传播方面的贡献提供了新的见解，并且堆肥的高温期延长会限制耐多药质粒的转移。The plasmid-mediated horizontal transfer of antibiotic resistance genes (ARGs) among bacteria facilitates the evolution and dissemination of antibiotic resistance. Broad-host-range plasmids can be transferred to different bacterial hosts in soil, plant rhizospheres, and wastewater treatment plants. Although composting is an effective way to convert organic waste into fertilizer and reduce some ARGs, few studies have focused on its effects on the spread of ARG-carrying plasmids and their bacterial host communities during composting. In this study, a fluorescently labeled Pseudomonas putida (P. putida) harboring a broad-host-range plasmid RP4 carrying three ARGs was inoculated into a raw material microcosm and composted with different durations of the thermophilic phase. The fate of the donor and RP4 in composting was investigated. The prolonged thermophilic composting removed 95.1% of dsRed and 98.0% of gfp, and it inhibited the rebound of P. putida and RP4 during the maturation phase. The spread potential of RP4 decreased from 10−4 to 10−6 transconjugants per recipient after composting. In addition, we sorted and analyzed the composition of RP4 recipient bacteria using fluorescence-activated cell sorting combined with 16S rRNA gene amplicon sequencing. The recipient bacteria of RP4 belonged to eight phyla, and Firmicutes, accounting for 75.3%–90.1%, was the dominant phylum in the transconjugants. The diversity and richness of the RP4 recipient community were significantly reduced by prolonged thermophilic periods. Overall, these findings provide new insights for assessing the contribution of composting in mitigating the dissemination of plasmid-mediated ARGs, and the prolonged thermophilic phase of composting can limit the transfer of multidrug-resistant plasmids.

https://www.frontiersin.org/articles/10.3389/fmicb.2022.989085/full