摘要

       采用高通量定量PCR和16srRNA测序技术，研究了猪粪厌氧消化过程中温度和残留抗生素对猪粪中抗生素耐药基因（ARGs）和微生物群落动态的影响。四种处理（25℃、37℃、37℃、50mg体重湿重抗生素和55℃）的总ARGs和16s rRNA基因丰度均显著降低。大多数ARG基因的丰度与16srrna基因和转座子基因的丰度显著相关（P<0.01）。但总ARGs在55℃时的丰度远高于其它处理。同时，在55℃时，链球菌病原菌仍保持较高的丰度，纤维素降解剂和产氢剂如乙醇酸和黄连球菌增加，与其他处理相比差异显著。冗余度分析表明，温度、pH值和链球菌属分别是实验因子、化学性质和代表属中ARG变化的最高解释。网络分析进一步表明，链球菌属对55℃时较高的ARG丰度有很大的贡献，中度的抗生素残留只对微生物多样性种群产生轻微的暂时性抑制作用，并促进ARG丰度的提高，这可能是由于抗生素的降解和微生物的适应性。我们的研究结果阐明了基因转移相关项目对ARG变异的协同作用，并进一步证明，在厌氧消化过程中，除非对病原菌和基因转移元件进行更有效的抑制，否则较高的温度往往不能达到更好的ARG去除效果施肥土壤丰度高。厌氧消化是一种广泛应用的动物粪便处理工艺。因此，了解厌氧消化对ARGs的抑制作用是非常重要的。尽管之前的一些研究建议采用高温消化法去除ARGs，但他们没有获得足够的证据支持这一观点。应用于动物的抗生素，由于代谢不完全，大多通过粪便和尿液排出。必须了解粪便中残留的抗生素是否会阻碍厌氧消化器中精氨酸的衰减。本研究的意义在于全面了解猪粪厌氧消化过程中受温度和残留抗生素影响的抗性基因（ARGs）的演变和作用机理，为其在释放到环境中之前的消除策略的制定提供依据。

       This study employed high-throughput quantitative PCR and 16S rRNA sequencing to evaluate the effect of temperature and residual antibiotics on the dynamics of antibiotic resistance genes (ARGs) and microbial communities during anaerobic digestion of swine manure. The abundances of total ARGs and 16S rRNA genes significantly decreased in all of four treatments (25°C, 37°C, and 37°C with 50 mg of wet weight antibiotics of body weight, and 55°C). The abundances of most ARG types were significantly correlated with those of the 16S rRNA gene and transposase gene (P < 0.01). However, the abundances of total ARGs at 55°C were much higher than those of other treatments. Meanwhile, the microbial communities at 55°C, where the Streptococcus pathogen remained at a relatively high abundance and cellulose degraders and hydrogen producers, such as Ethanoligenens and Coprococcus bacteria, increased, were markedly different from those of other treatments. Redundancy analysis indicates that temperature, pH, and the genus Streptococcus had the highest explanation for ARG variation among experimental factors, chemical properties, and representative genera, respectively. Network analysis further showed that the genus Streptococcus contributed greatly to the higher ARG abundance at 55°C. The moderate antibiotic residue only caused a slight and transitory inhibition for microbially diverse populations and promotion for ARG abundance, probably due to the degradation of antibiotics and microbial adaptability. Our results clarify the cooperativity of gene transfer-related items on ARG variation and intensively prove that higher temperature cannot always achieve better ARG removal in anaerobic digestion unless pathogens and gene transfer elements are more efficiently inhibited.IMPORTANCE Antibiotic resistance genes (ARGs) are frequently detected with high abundance in manure-applied soils. Anaerobic digestion is one of widely used processes for animal waste treatment. Thus, it is critical to understand the potential of anaerobic digestion to attenuate ARGs. Although some previous studies recommended thermophilic digestion for ARG removal, they did not get sufficient evidence to support this view. The antibiotics applied to animals are mostly excreted through feces and urine because of incomplete metabolism. It is indispensable to know whether residual antibiotics in manure will hinder ARG attenuation in anaerobic digesters. The significance of our research is in comprehensively understanding the evolution and mechanism of ARGs in anaerobic digestion of swine manure affected by temperature and residual antibiotics, which will allow the development of an ARG elimination strategy before their release into the environment.

       https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6585508/