垫料对异位发酵床中四环素抗性基因及Ⅰ类整合子丰度的影响

Effect of Bio-Bed Litters on the Abundance of Tetracycline Resistance Genes and Class Ⅰ Integron in Ectopic Fermentation Bed

【目的】研究垫料对异位发酵床中四环素抗性基因(TC-ARGs)及Ⅰ类整合子(intI1)丰度的影响并进一步探究各基因主要宿主。【方法】采用实时荧光定量PCR技术和16S rRNA微生物测序技术测定了两种不同发酵床垫料(锯末与谷壳混合垫料和单独酒糟垫料)的TC-ARGs(tetA、tetC、tetM、tetO、tetQ、tetW和tetX)丰度及微生物群落结构,并分析了垫料理化性质和微生物群落与TC-ARGs及intI1丰度的关系。【结果】锯末和谷壳发酵床和酒糟发酵床的TC-ARGs和intI1的丰度范围分别为6.96×108~1.51×109和9.07×108~2.29×109 copies/g。其中,tetW、tetM、tetQ和tetX在酒糟发酵床中的丰度总体上高于锯末+谷壳发酵床,而锯末+谷壳发酵床中tetO、tetA、tetC和intI1的丰度更高。另外,研究发现温度、C/N和有效Zn含量可能是造成异位发酵床中TC-ARGs和intI1丰度差异的主要原因,Firmicutes可能是tetW、tetM、tetQ和tetX基因的主要携带菌门,Bacteroidetes和Actinobacteria可能是tetO基因的主要携带菌门,而Proteobacteria和Chloroflexi可能是tetA、tetC和intI1基因的主要携带菌门。【结论】不同异位发酵床垫料中TC-ARGs和intI1的丰度存在明显差异,而且各TC-ARGs的主要宿主也不同,温度、C/N和重金属含量等因素会影响ARGs和intI1的丰度,研究为异位发酵床去除ARG和降低ARGs环境风险提供了一定的理论依据。

【Objective】This study aimed to explore the effect of two different bio-bed litters(one is a mixture of sawdust and husk,and the other one is lees alone) on the abundance of tetracycline antibiotic resistances(TC-ARGs) and class Ⅰ integron(intI1) in industrial manure fermentation beds as well as their potential hosts.【Method】The qQT-PCR and high-throughput sequencingwere used to determine the abundance of TC-ARGs(including tetA,tetC,tetM,tetO,tetQ,tetW,and tetX) and intI1 as well as the bacterial communities. Furthermore,the abundance of TC-ARGs and intI1 was correlated with the properties of fermentation litters and bacterial communities to figure out the factors affecting the spreading of TC-ARGs and intI1 and the potential hosts.【Result】The abundance of TC-ARGs and intI1 in the fermentation bed of sawdust and husk and lees was in the range of 6.96×108-1.51×10~9 and 9.07×108-2.29×10~9 copies/g,respectively. The abundance of tetW,tetM,tetQ,and tetX in the fermentation bed of lees was higher,while the abundance of tetO,tetA,tetC,and intI1 was lower than that in the fermentation bed of sawdust and husk. The temperature,C/N ratio,and the available Zn might largely explain the differences of TCARGs and intI1 abundance between two different fermentation beds. Besides,it is found that the possible hosts of tetW,tetM,tetQ,and tetX might be Firmicutes,and tetO might be mainly hosted in Bacteroidetes and Actinobacteria,with Proteobacteria and Chloroflexi might primarily hosting tetA,tetC,and intI1.【Conclusion】This study found that the abundance of TC-ARGs and intI1 in two different bio-bed litters are significantly different,and the main host of each gene is also various. The environmental factors such as temperature,C/N ratio,the content of heavy metals could influence the abundance of the above mentioned genes. It is expected that this study could provide a theoretical basis for using ectopic fermentation bed technology to remove the antibiotic resistance gene and reduce its environmental risk.