| Consortium strains GF-20 has stable decomposition activity of corn stover at low temperature and complex composition of microorganisms,and it is not clear that how consortium strains GF-20 worked under low temperature,which is not conducive to microbial inoculum.Therefore,this study used metagenomic sequencing technology to carry out high-throughput sequencing on the consortium strains GF-20.Based on the sequencing data,the flora structure and species composition of the consortium strains were analyzed,and the microbial composition types,dominant functional bacteria,functional analysis and metabolic mode were deeply discussed.On this basis,the key components of GF-20 strains were isolated and recombined,and the functional characteristics of consortium strains GF-20 bacteria were clarified to simplify the composition of consortium strains GF-20 bacteria and improve the degradation efficiency.Main results for this study are as follows:1.Consortium strains GF-20 diversity of consortium bacteria:metagenomics was used to identify the microbial community structure,functional genes and species diversity of consortium strains GF-20,and to explore the relationship between microbial community and straw degradation.The results showed that consortium strains GF-20 was mainly composed of bacteria,with Proteobacteria(62.84%)and Bacteroidetes(10.24%)at the phylum level,and was mainly distributed in Pseudomonas(50.84%),Dysgonomonas(5.86%),Achromobacter(4.94%),Stenotrophomonas(3.67%),Flavobacterium(2.04%)at the genus level.Metabolic ways mainly for carbohydrate metabolism and amino acid metabolism,and at the same time had a way of signal transduction,cell membrane,and other metabolic.The function of the encoding genes were mainly distributed in the glycoside hydrolase.The functional enzyme mainly for beta glycosidase enzymes,acetyl coenzyme A,pyruvate dehydrogenase,and galactose glucoside enzyme.Moreover consortium strains GF-20 encoding the genes of amylase(Amy),glycogen phosphorylase(Pyg),glycogen debranching enzyme(Agl),cut inside glycosidase enzymes(bglX)and glucanase.the complete the pentose phosphate pathway and glycolytic pathway,has such as starch,cellulose and sucrose degradation function.2.Degradation mode of corn stover of GF-20:The endoglucanases and β-glucosidase produced by Dysgonomonas,Sphingobacterium,Flavobacterium,Parabacteroides,Pleomorphomonas,A rcticibacter,Elizabethkingia,Neisseria,Mycobacterium,Stenotrophomonas,Paenibacillus and Cellulomonas were the mainly enzyme that degraded cellulose of corn stover by GF-20.The xylanase,xyloglucanase,manganase and branched enzyme produced Pedobacter,Pleomophomonas,Cellulomonas,Prevotella,Dysgonomonas,Parabacteroides,Pusillimonas,Nocardiopsis,Rhizobium,Chitinophaga,Agrobacterium,Rikenella,Alistipes,Draconibacterium and Proteiniphiluby were the mainly enzyme that degraded hemicellulocellulose of corn stover by GF-20.The polysaccharides or monosaccharides produced by degradation are completely degraded into water,CO2 and organic acids through glycolysis,pentose phosphate pathway and tricarboxylic acid cycle3.Isolation and construction of composite strains:29 strains were isolated and purified,13 single strains with obvious hydrolyzation circle.According to the antagonistic test,11 groups of consortium strains were obtained.The degradation effect of corn straw of ZH was optimal with the FPA 11.12 U/ml,xylanase activity 2.65 U/ml,laccase activity 27.68 U/L,and degradation rate of 28.72%,22.94%better than CK.ZH was composed of only A3 and A4 strains.The 16S rDNA molecule identified A3 as Achromobacter deleyi and A4 as Pseudomonas plecoglossicida.In order to further improve the efficiency of the degradation of the mixed bacteria useful,add the Aspergillus terreus and Phanerochaete Chrysosporium with A3 and A4 are combined to optimize the distribution of bacteria in Z.Compared with ZH the Z useful maize straw degradation rate,FPA,Xylanase,Laccase activity significantly increased by 11.71%,2.58 U/ml,3.21 U/ml,36.76 U/L.4.The degradation characteristics of the consortium strains Z:By studying the enzyme activity characteristics,growth characteristics,VFA and content the degradation characteristics of the consortium strains Z were investigated.The consortium strains were cultured at 15℃ for 72 hours into the logarithmic growth period.The pH of the fermentation broth fluctuated between 6.7 and 8.6.The enzyme activity,xylanase activity,laccase activity of the consortium strains filter paper and the degradation rate of corn straw all maintained above 80%efficiency within the temperature range of 0.02%-2.5%salt concentration,initial pH 4.0-9.0 and 13℃-25℃.With the increase of culture algebra,there were no significant differences in the degradation rate,lignocellulosic degradation rate,filter paper enzyme activity,xylanase activity,laccase activity,VFA content and pH(sterilization).Under the condition of carbon source of unsterilized corn straw,the degradation rate of corn straw,lignocellulosic degradation rate,filter paper enzyme activity,xylanase activity,laccase activity and VFA content were all significantly increased(p<0.05)and pH value significantly decreased(p<0.05).Outdoor soil culture application test showed that the consortium strains Z could promote the degradation of lignin cellulose and enhance the activities of sucrase,catalase,cellulase and urease in soil.5.Analysis of synergistic promotion effect of Z:Using one by one by means of culling method to explore the synergistic promoting relationship between the component bacteria of Z.It can be known by the culling method A3,A4,A and P all played an important role in improving the enzyme activity of filter paper in the consortium strains system.The increase rate of A3,A4,A and P were 18.70%,18.06%,17.44%,18.06%;A3 increased the activity of compound xylanase by more than 80%and played an important role in increasing the xylanase activity.The activity of laccase in consortium strains Z was increased by more than 70%by P.With the addition of single strain P,the degradation rate of lignin in corn stover was doubled increased.The A3,A4,A and P increase degradation rate of cellulose in corn straw was 16.87%,20.73%,15.17%,20.57%.The A3,A4,A and P worked in degradation of hemicellulose in corn straw.The improvement rate of degradation rate of A4 was 53.91%.The overall trend of relative gene expression increased with the addition of A3,A4,A and P.The addition of fungi can increase the microbial population of bacteria,A3,A4,A and P degradated different components of corn straw,Promote the forward reaction,synergistic degradation of corn straw,improve the degradation efficiency. |
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