基于高通量测序技术筛选低温秸秆降解菌群的研究

Screening low temperature straw degradation bacteria using high-throughput sequencing technology

[目的]本文旨在筛选低温(10℃)高效的玉米秸秆降解菌群。[方法]采集不同低温菌源的样品,进行低温(10℃)连续富集继代(10代)培养,利用第二代高通量测序方法分析不同处理的细菌群落结构和数量,并测定不同世代的纤维素酶、秸秆失重率等数据。[结果]筛选获得一组高效稳定分解玉米秸秆的复合菌群SLX。该菌群在15 d内产生复合纤维素酶系,对玉米秸秆的分解率达到58.97%。第二代高通量测序技术结果表明不同菌源来源是菌群分类的主要依据,复合菌群SLX主要含有酸杆菌门(Acidobacteria)、放线菌门(Actinobacteria)和芽单胞菌门(Gemmatimonadetes)的微生物菌株,种分类上,uncultured Acidobacteriaceae,uncultured Sphingomonas等为优势菌株。外切β-葡聚糖酶(C1)与α-变形菌纲(Deltaproteobacteria)、β-葡萄糖苷酶(BG)与β-变形菌纲(Betaproteobacteria)表现正相关;内切β-葡聚糖酶(CMCase)和滤纸酶(FPA)位于第二象限,与酸杆菌门(Acidobacteria)表现正相关,可以解释微生物群落结构总变异的34.15%,菌群通过菌种之间的协同作用,共同维持了体系的稳定。固体发酵培养实验结果显示:麦麸∶秸秆粉的比例为7∶3时为最佳配比,水料比的最佳比例为1∶1时,接种量的最佳值为5%。[结论]研究结果为明确低温秸秆降解菌群降解机理和提高纤维素降解效率提供了理论基础。

[Objective]The present research was aimed at screening and identifying corn stalk degradation complex flora which were efficient at the low temperature(10℃).[Methods]Experimental samples were collected from different low-temperature microbial sources,and cultured at low temperature(10℃)for performing continuous enrichment for 10 generations.The second-generation high-throughput sequencing method was used to analyze the bacterial community structure and number of different treatments,and to determine the cellulase,straw weight loss rate and other data of different generations.[Results]The main results showed that a group of complex flora SLX which efficiently and stably decomposed corn stalks were selected.The flora produced a composite cellulase system within 15 days,and the decomposition rate of corn stover reached 58.97%.The results produced by the second-generation high-throughput sequencing technology showed that different sources of bacteria were the main basis for the classification of flora.The complex flora SLX mainly contained Acidobacteria,Actinobacteria and Bacillus phylum(Gemmatimonadetes);in terms of species classification,uncultured Acidobacteriaceae,uncultured Sphingomonas,etc.were the dominant strains.Exo-β-glucanase(C1)andα-Proteobacteria,β-glucosidase(BG)andβ-Proteobacteria were positively correlated with their performance;endo-β-glucan Enzymes(CMCase)and filter paper enzymes(FPA)were located in the second quadrant,and were positively correlated with Acidobacteria,which contributed for 34.15%of the total variation of the microbial community structure.The bacterial community maintained the stability of the system through the synergic function of the bacterial species.The results of solid fermentation culture experiments showed that the optimal ratio of wheat bran:straw powder was 7∶3,and when the optimal ratio of water∶material was 1∶1,the optimal inoculation amount was 5%.[Conclusion]The research results provided a theoretical basis for clarifying the degradation mechanism of low-temperature straw degradation bacteria and improving the efficiency of cellulose degradation.