泰乐菌素对转基因棉秆堆肥过程中微生物群落多样性及抗性基因的影响

发布时间：2018-06-28 21:46

本文选题：转基因棉秆 + 堆肥　；参考：《西北农林科技大学》2017年硕士论文

【摘要】：中国是世界上最大的棉花生产国,其中80%是转基因棉花,因而转基因棉秆的产量非常大,据统计,仅新疆自治区的棉秆年产量就高达600~750万吨(湿重)。棉秆中含有大量的木质纤维素、粗纤维、氮和磷等,通常被焚烧处理,不仅浪费了宝贵资源而且造成环境污染。好氧堆肥被认为是处理农业固体废弃物以及畜禽粪便最有前景的方法之一,并且畜禽粪便中残留的抗生素也得到广泛的关注。本研究以转基因和非转基因棉杆为原材料,和猪粪进行好氧堆肥,并添加养殖业大量使用的抗生素(泰乐菌素),采用Biolog法和实时定量PCR技术等,研究堆肥过程中的理化性质、微生物群落多样性、抗生素抗性基因以及Ⅰ、Ⅱ类整合子的变化。主要结果如下:1.转基因棉杆对堆肥过程中抗生素抗性基因(ARGs)和移动基因元件的影响转基因棉杆和非转基因棉杆用作为堆肥原料,经过40天的堆肥,均能达到腐熟标准。在堆肥各个时期的样品中检出了5种大环内酯抗性基因(ermB、ermF、ermQ、ermT、ermX)、2种四环素类抗性基因(tetC、tetX)、2种磺酰胺抗性基因(sul1、sul2)、I类和II类整合子基因(intI1,int I2)。堆肥结束时,转基因和非转基因处理中ARGs,intI1和intI2的绝对丰度分别减少了5.12log和4.45log,且ARGs的变化受温度和铵态氮影响最为显著。此外,转基因棉秆和非转基因棉秆对大多数ARGs,intI1和intI2的影响没有差异。因此,同非转基因棉秆一样,转基因棉秆也可用于同畜禽粪便的好氧堆肥,并通过堆肥减少ARGs的丰度。2.泰乐菌素对堆肥过程中理化性质和微生物群落多样性的影响经过40天的堆肥,四个处理的温度均在50℃以上持续约6天,达到无害化处理要求。在堆肥结束时,四个处理的pH均在7.5-8.5之间,C/N值小于20,达到了腐熟标准。Biolog结果显示,添加泰乐菌素能抑制堆肥初期微生物活性,但是随着堆肥的进行,堆肥中样品中泰乐菌素逐渐降解,抑制用作逐渐消失。多样性指数表明,泰乐菌素影响了微生物群落功能多样性和物种的优势度,在高温期低浓度的泰乐菌素影响作用更加显著,中浓度处理能提高微生物群落的均一性。并且添加泰乐菌素能提高堆肥末期微生物对六大类糖原的利用。3.泰乐菌素对堆肥过程中抗性基因和遗传基因元件的影响经过好氧堆肥,ARGs,intI1和int I2的总绝对丰度在CK、L、M和H处理中分别减少了8.61log、9.23log、9.27log和8.17log,说明通过好氧堆肥能有效减少抗性基因。16s rDNA定量结果表明,添加泰乐菌素能增加堆肥末期的细菌丰度。通过RDA分析发现,环境变量能解释总基因丰度变化的54.4%,且ARGs,intI1和int I2的变化受pH值影响最显著,占总解释量的19.1%。好氧堆肥虽然是一种有效去除抗生素的方法,但是不能完全去除抗性基因,因此还需要进一步研究来降低抗性基因对环境带来的风险。转基因棉秆和非转基因棉杆均可作为堆肥底料进行对肥,且泰乐菌素能抑制堆肥初期微生物活性,并提高堆肥末期微生物对六大类糖原的利用,本文为评价含抗生素畜禽粪便堆肥的生态风险提供了理论参考依据。
[Abstract]:China is the largest cotton producing country in the world, 80% of which are genetically modified cotton, so the yield of transgenic cotton stalk is very large. According to statistics, the annual output of cotton stalk in Xinjiang autonomous region is up to 600~750 ten thousand tons (wet weight). The cotton stalk contains a large amount of lignocellulose, coarse fiber, nitrogen and phosphorus, which is usually burned and wasted the precious capital. Source and environmental pollution. Aerobic composting is considered as one of the most promising ways to deal with agricultural solid waste and livestock manure, and antibiotics residues in livestock and poultry manure have been widely concerned. The use of antibiotics (tylosin), Biolog and real-time quantitative PCR techniques were used to study the physical and chemical properties of the composting process, microbial community diversity, antibiotic resistance genes and the changes of class I and class II integrons. The main results are as follows: 1. the effects of transgenic cotton rods on the antibiotic resistance gene (ARGs) and mobile gene elements in the composting process Transgenic cotton rods and non transgenic cotton rods were used as compost materials, and after 40 days of composting, they all reached the standard of decayed maturity. 5 kinds of macrolide resistance genes (ermB, ermF, ermQ, ermT, ermX), 2 tetracycline resistance genes (tetC, tetX), 2 sulfonamide resistance genes (sul1, sul2), I class and II class were detected in the samples of each period of composting. The subgenes (intI1, int I2). At the end of the composting, the absolute abundance of ARGs, intI1 and intI2 in the transgenic and non transgenic treatments decreased 5.12log and 4.45log, respectively, and the variation of ARGs was most significantly affected by the temperature and ammonium nitrogen. In addition, the effects of transgenic cotton stalk and non transgenic cotton stalk on most ARGs, intI1 and intI2 were not different. As with non transgenic cotton stalks, transgenic cotton stalks can also be used in aerobic composting with livestock and poultry manure, and the effects of ARGs abundance.2. tylosin on the physical and chemical properties and microbial community diversity during the composting process after 40 days of composting, the temperature of the four treatments lasted for about 6 days above 50 degrees centigrade, reaching harmless treatment. At the end of the composting, the four treated pH were between 7.5-8.5 and C/N less than 20, and the result showed that the addition of tylosin could inhibit the early microbial activity of the composting, but with the composting, tylosin gradually degraded and the inhibition was gradually disappearing. The diversity index indicated tylosin. Factors affecting the functional diversity of microbial community and the dominance of species, the effect of tylosin at low concentration in high temperature is more significant, and medium concentration treatment can improve the homogenization of microbial community. And adding tylosin can improve the use of telophore to six major types of glycogen at the end of the composting, and.3. tylosin resistance to the composting process. The total absolute abundance of gene and genetic elements through aerobic composting, ARGs, intI1 and int I2 reduced 8.61log, 9.23log, 9.27log and 8.17log, respectively in CK, L, M and H treatments, indicating that aerobic composting could effectively reduce the resistance genes, indicating that the addition of tylosin could increase the abundance of bacteria at the end of the composting. RDA analysis showed that environmental variables could explain 54.4% of the changes in total gene abundance, and the changes in ARGs, intI1 and int I2 were most significantly affected by pH, while 19.1%. aerobic composting, which accounted for the total explanation, was an effective method to remove antibiotics, but it could not completely remove the resistance base, so further study was needed to reduce the resistance genes. Both transgenic cotton stalk and non transgenic cotton stalk can be used as fertilizer on the base of compost, and tylosin can inhibit the initial microbial activity in the early composting, and improve the use of microbes at the end of composting at the end of the composting. This paper provides a theoretical reference for evaluating the ecological risk of livestock manure composting with antibiotics.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S141.4

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