Screening Of High Temperature Cellulose Degrading Bacteria In Compost Of Pig Carcasses

In composting treatment,cellulose has become a limiting factor for shortening the composting time and improving the efficiency of composting,due to its complex structure and poor degradation.Therefore,accelerating the decomposition of cellulose is one of the key factors to improve composting efficiency and accelerate the maturation of compost materials.During the high temperature period of composting,the temperature can reach above 50?,the development and utilization of high temperature bacteria has attracted wide attention.High temperature bacteria have the characteristics of fast metabolism,high activity,high thermal stability of enzymes,and the creation of high temperature conditions to kill pathogenic bacteria.They have an inestimable effect on the biological transformation of organic matter under high temperature environment.Therefore,screening high-efficiency and high-temperature fiber bacteria is an important means to improve the efficiency of composting.this study screened bacteria that can promote cellulose degradation during high-temperature composting by high-temperature enrichment culture,primary screening,double screening,ultraviolet mutagenesis,and enzyme activity determination,and identified them using whole-gene sequencing technology.The main findings are as follows:Collect pig carcasses and straw compost samples in the high temperature period,use beef extract peptone medium for enrichment culture at 60?,cellulose Congo red medium for separation and purification culture,screening medium for preliminary screening culture.Five dominant strains were screened by the cellulase activity method in the fermentation broth,namely CMC-1,CMC-2,CMC-3,CMC-4 and CMC-5.The carboxymethylcellulase(CMCase)activity was 20.81 U/m L,39.36 U/m L,54.65 U/m L,80.80 U/m L and 38.65 U/m L,respectively.The morphological characteristics,physiological and biochemical characteristics and 16 S r DNA gene sequence analysis of CMC-4 were preliminarily identified as Geobacillus thermodenitrificans.CMC-4 was irradiated with ultraviolet radiation for mutagenesis,and by Congo red staining and cellulase activity measurement by DNS method,three strains were obtained that could decompose more CMC-Na than the original strain under the same culture conditions.It is CMC-4-1,CMC-4-5,CMC-4-7.Among them,the CMC-4-1 strain had the largest enzyme activity of 94.00 U/m L,and the 16 S r DNA gene sequence identified the strain as a Parageobacillus thermoglucosidasius BGSC 95A1.The results of the growth characteristics and the effects of different factors on the enzyme production activity of the two strains before and after mutagenesis are shown.CMC-4 and CMC-4-1 optimal growth temperature are 55?,CMC-4 optimal growth p H is 7.CMC-4-1 optimal growth p H is 8,CMC-4 and CMC-4-1 optimal growth filling volume are 50 m L/250 m L.When the CMC-4 strain uses microcrystalline cellulose as the carbon source,and the casein is the nitrogen source,the enzyme production ability is strong;When the CMC-4-1 strain uses CMC-Na as the carbon source and ammonium nitrate is used as the nitrogen source,the enzyme-producing ability is strong;the optimal p H for the enzyme-producing CMC-4 and CMC-4-1 are 7,and the optimal temperature for the enzyme-producing CMC-4 and CMC-4-1 are 65?.CMC-4-1 zymogram analysis show that the molecular mass of extracellular cellulase was 45 k Da.CMC-4-1 HPLC analysis showed that degradation products of carboxymethyl cellulose are cellobiose and glucose,and the content of glucose in the degradation products is the majority,and cellobiose is less.The whole genome sequencing analysis results show that CMC-4-1 contains multiple endo-1,4-?-D-glucanase and ?-glucosidase protein coding genes.These coding genes indicate that CMC-4-1 has the ability to degrade cellulose.