秸秆木质素降解细菌的筛选及其降解条件优化

Isolation of Straw Lignin⁃Degrading Bacteria and Optimization of Their Degradation Conditions

本试验旨在分离和筛选靶向降解木质素能力强的细菌菌株,并优化其培养条件,应用于秸秆生物质中木质素的降解。利用碱性木质素为唯一碳源的无机盐培养基进行菌株初筛,通过苯胺蓝法进行复筛,对筛选出的菌株进行细菌形态学观察和16S rDNA序列测定,然后以碱性木质素降解率为评价指标,采用单因素和响应面法进行培养条件优化及产酶能力的研究。结果显示:1)筛选出的3株木质素降解细菌BH5、BH8和BJ5经鉴定分别为吉洛氏不动杆菌(Acinetobacter guillouiae)、嗜麦芽窄食单胞菌(Stenotrophomonas maltophilia)和变栖克雷伯氏菌(Klebsiella variicola)。2)菌株BH5在初始pH 6.8、培养温度30.3℃、接种量6.3%时木质素降解能力最佳,木质素降解率达到16.72%,所产漆酶(Lac)、木质素过氧化物酶(Lip)和锰过氧化物酶(Mnp)活性分别为1.53、15.65和43.85 U/mL;菌株BH8在初始pH 6.3、培养温度28.4℃、接种量4.7%时木质素降解能力最佳,木质素降解率达到18.01%,所产Lac、Lip和Mnp活性分别为4.17、21.66和72.26 U/mL;菌株BJ5在初始pH 6.4、培养温度29.7℃、接种量4.1%时木质素降解能力最佳,木质素降解率达到21.65%,所产Lac、Lip和Mnp活性分别为5.94、34.44和90.81 U/mL。综上所述,本试验筛选出的3株木质素降解细菌具有较高的木质素降解能力,这不仅丰富了木质素降解微生物库,同时也为后续应用微生物菌剂发酵秸秆,促进秸秆饲料化的研究提供了理论依据。

The purpose of this study was to isolate and screen bacterial strains with strong ability to target lignin degradation,and optimize their culture conditions for the degradation of lignin in straw biomass.The inorganic salt culture medium with alkaline lignin as the sole carbon source was used for the initial screening of the strains,and the aniline blue method was used for the secondary screening.The bacterial morphology and 16S rDNA sequence of the screened strains were observed,and then the degradation rate of alkaline lignin was used as the evaluation index.The single factor and response surface methods were used to optimize the culture condi⁃tions and study the enzyme production ability.The results showed as follows:1)three strains of lignin degrad⁃ing bacteria BH5,BH8 and BJ5 were identified as Acinetobacter guillouiae,Stenotrophomonas maltophilia and Klebsiella variicola,respectively.2)The strain BH5 had the best lignin degradation ability at initial pH 6.8,culture temperature 30.3℃and inoculation amount 6.3%,and the lignin degradation rate reached 16.72%,a⁃mong which laccase(Lac)activity was 1.53 U/mL,lignin peroxidase(Lip)activity was 15.65 U/mL,and manganese peroxidase(Mnp)activity was 43.85 U/mL;the strain BH8 had the best lignin degradation ability at initial pH 6.3,culture temperature 28.4℃and inoculation amount 4.7%,and the lignin degradation rate reached 18.01%,among which the Lac activity was 4.17 U/mL,Lip activity was 21.66 U/mL,and Mnp ac⁃tivity was 72.26 U/mL;the strain BJ5 had the best lignin degradation ability at initial pH 6.4,culture tempera⁃ture 29.7℃and inoculation amount 4.1%,and the lignin degradation rate reached 21.65%,among which Lac activity was 5.94 U/mL,Lip activity was 34.44 U/mL,and Mnp activity was 90.81 U/mL.To sum up,the three strains of lignin⁃degrading bacteria screened in this study have high lignin⁃degrading ability,which not only enriches the lignin⁃degrading microbial pool,but also provides theoretical basis for the subsequent research on the application of microbial agents to ferment straw and to promote the feed of straw.