一株中温厌氧纤维素降解细菌的分离、鉴定及其系统发育分析  

Isolation,Characterization and Phylogenetic Analysis of A Mesophilic,Cellulytic,Anaerobic Bacterium

作　　者：罗辉[1] 仇天雷[2] 马诗淳[1] 刘来雁[1] 张辉[1] 尹小波[1] 邓宇[1] 

机构地区：[1]农业部沼气科学研究所,成都610041 [2]北京农业生物技术研究中心,北京100089

出　　处：《中国沼气》2008年第3期3-7,14,共6页China Biogas

基　　金：国家自然科技资源平台项目(2004PKA30560);国家"863"项目沼气发酵过程微生物调控技术研究项目(2006AA10Z420);农业科技成果转化资金项目(2006GB23260391)

摘　　要：以树中心湿木为样品,富集分离获得一株具有较强纤维素分解能力的中温厌氧纤维素降解细菌CSZM-6。分离菌株革兰氏染色阴性,直杆或稍弯曲杆状,多数单生,少数成对或成串,部分菌体产芽孢。菌体大小为0.5μm×1.7-2.6μm,严格厌氧,最适生长温度40℃,最适pH6.5,以葡萄糖为底物时倍增时间8.0 h。分离菌株在纤维素粉固体培养基中培养3周后,菌落为圆形或不规则形状,产黄色素,直径为0.5-4.0 mm,水解圈直径为2.0-18.0mm。分离菌株不仅能利用纤维素、纤维二糖、葡萄糖等碳水化合物,还能利用未经处理的报纸、水稻秸秆、麦秆。发酵纤维素产乙醇、乙酸、H2、CO2、甲酸、丙酸、乳酸。在菌株CSZM-6的纤维素酶系中,只存在外切-β-1,4-葡聚糖酶（C1酶）和内切-β-1,4-葡聚糖酶（CX酶）两种组分,最适温度分别为40℃和45℃。通过16S rDNA序列分析表明,菌株CSZM-6为梭菌属Clostridium,与C.papyrosolvens具有99.6%的相似性。Through selective enrichment culture, an obligately anaerobic, mesophilic, cellulytic bacterium, strain CSZM-6 was isolated from wetwood. The cells were straight or slightly curved rods that measured between 1.7μm and 2.6μm length and approximately 0.5 μm in diameter , non-motile, most appearing singly, a few in pairs or filament, Gram-negative. Spores were observed in a few bacteria. The optimum temperature was 40℃ and an optimum pH 6.5. The specific growth rate of strain CSZM-6 was 0.12 h^-1 and the doubling time on glucose of 2 g · L^-1 was 8.0 h at 40℃ and pH 6. 5 without NaCl. After the isolated bacteria grew on solid agar medium with cellulose powder for 3 weeks, the colonies appeared round or irregular with diameter of 0.5 - 4.0 mm, and were opaque, non-glossy, moisture and yellow pigmented with complete margin. Clear zones which showed cellulose hydrolysis were generally 2.0- 18.0 mm in diameter after an incubation of 3 weeks. Not only cellulose, cellobiose and glucose could be utilized by the bacteria, but also unpretreated paper, rice straw, wheat straw, etc. The major end products from cellulose were ethanol, acetate, carbon dioxide and hydrogen. And formate, propionate and lactate were the minor products. In cellulases of strain CSZM-6, only exoglucanase and endoglucanase existed, and the optimum temperature was 40℃ and 45℃ respectively. β-Glucosidases was not found in the cellulases. The phylogenetic analysis based on 16S rDNA suggests that strain CSZM-6 is closely related to C. papyrosolven with 99.6% sequence similarity. Therefore, strain CSZM-6 was identified as a species Clostrdium.

关 键 词：中温厌氧纤维素降解细菌 鉴定 16SrDNA 系统发育分析 

分 类 号：S216.4[农业科学—农业机械化工程]