Screening And Identification Of Lignin-degrading Microorganisms And Their Application In Tobacco Stem Degradation

China is the biggest tobacco market all of the world,and the plant area is 21.18 million mu.The annual produce a large number of discarded tobacco stem,which could not be applied.Also,a lot of tobacco stem have been accumulated in the fields,which might cause the soil pollution and disease.More seriously,some tobacco stem was burned directly,and it might cause the resource waste and air pollution.Through high temperature compost technology with the biological agents degradation method,convert waste tobacco stalk into biological organic fertilizer.To reduce the tobacco stalk discarded or burned cause waste of resources,environmental pollution and soil borne disease were spread.Recently,adding the cellulose degrading bacteria into the fermentation preparation of organic tobacco stem has entered the stage of industrialization.However,there were also a lot of lignin that could not degraded by this method,which not only affect the degree of compost,but also affect the speed of composting rotten.In this study,we have screened the high efficient degradation bacteria from the nature,isolated and purified the high-efficiency lignin-degrading microorgnism,then inoculated the lignin degrading microorganism in the tobacco stalk,and the determination of lignin degradation bacteria for tobacco stalk compost reinforcement effect.In order to further reduce tobacco stalk composting period,improve the quality of tobacco stem composting products laid the foundation.The aim of this study was to stalk lignin degradation,screening from tobacco rod pile,soil samples,tobacco stem,termites,vines and some other samples,and we have obtained 69 strains which can produce ligninase.To sieve effective strains again which were obtained at fiest sieve.The bacterias were inoculated into the liquid enzyme production culture medium and fungi were inoculated to the solid enzyme production culture medium.The activity of lignin enzymes were determnined to complete the fermentation curve of enzyme production.After screening for 14 strains synthesis enzyme activity is higher lignin degrading bacteria.Perform 14 high enzyme activity of degrading bacteria strains for antagonism experiment.To select no antagonism strains for the degradation effect of inhibiting experiment.Degradation effect experiment is divided into four groups,will strain according to alone strain inculated,two strains inoculated,three and four strains inoculated to turn to the rod solid fermentation medium.After 35 days of the static fermentation cultivation,extraction of crude enzyme fluid determination of enzyme activity,and the determination of each group of hemicellulose,cellulose and lignin degradation rate.Experimental results obtained,two combination of lignin degradation rate was 44%.These two degradation effect is obvious combination are B:Y38/P8/Y9,C:Y3/Y34/P9/Y9.To identifie obvious effect degradation strains,bacteria after gram staining,physiological and biochemical identification and 16S rDNA gene sequence analysis.The sequencing results indicted that Y3,Y34,Y38 were all Bacillus subtilis.Fungi by moorphological identification and ITS gene sequence analysis appraisal.The sequencing results indicted that P8 Colletotrichum gloeosporidides,and Y9,Y3,Y34,Y38 were Aspergullus versicolor.Tten,the strains were cultured in the tobacco stalk enzyme production medium,and the biggest lignin degradation enzyme activity are:lignin peroxidase 867.01 IU/mL,768.54 IU/mL,983.98 IU/mL,manganese peroxidase 643.97 IU/mL,581.97 IU/mL,270.77 IU/mL,laccase 11.73 IU/mL,30.24 IU/mL,26.89 IU/mL.P8,Y9 secretion in the tobacco stalk the biggest lignin degradation enzyme enzyme activity are:lignin peroxidase 977.45 IU/g,134.07 IU/g,manganese peroxidase 990.14 IU/g,194.98 IU/g,laccase 106.67 IU/g,34.99 IU/g.