一个嗜热分解纤维素的梭菌新种的分离和鉴定

ISOLATION AND CHARACTERIZATION OF CLOSTRIDIUM SUCCINOGENUM SP.NOV.,A CELLULOLYTIC THERMOPHILE

本文描述了一个严格厌氧,形成芽孢,嗜热,分解纤维素、糖和蛋白质的梭菌新种。这个种的两株菌(H11,H12)是从奶牛场的青贮饲料(山草)中分离出来的。在 PYC 液体培养基中细胞杆状,0.3—0.6×1.5—5.5μm,单生或成对,很少成链。革兰氏阴性,能运动,有周生鞭毛。芽孢卵至球形,端生和次端生,使细胞膨大。游离芽孢球形,有孢子外壁和附属丝。在 PYC 琼脂滚管中,深层菌落呈双凸镜状,湿润,浅黄色,直径1—2mm,边缘完整。生长的最适条件是50—53℃和 pH 7.5。所需要的生长因子是生物素、维生素 B_6、B_(12)和对氨基苯甲酸。发酵纤维素或纤维二糖产生琥珀酸、乙酸、乙醇、氢和二氧化碳。DNA 的 G+C 含量是38.8 mol%。参考这个种在非挥发酸中的主要产物是琥珀酸的特性,将它定名为产琥珀酸梭菌(Clostridium succinogenum Tan & Wang sp.nov.)。

A new species of obligately anaerobic,sporeforming,thermophilic,cellulolytic,saccharoly-tic,proteolytic Clostridium is described.Two strains(H11,H12)of this species are isolatedfrom silage(grass)on milk cow field.Cells in PY-cellobiose broth culture are Gram-negative rods,0.3 to 0.6 by 1.5 to 5.5 μm,motile and peritrichous,occur singly or in pairs,occasionally in short chains.Long filamentsmay be seen in older cultures.Spores are oval or round,terminal and subterminal,swell the cell.Sporulation occurs readily in PY-cellobiose broth or cellulose broth after incubation for 24 h.Free spores are round,have an exosporium and appendages.Deep colonies in PY-cellobiose agarroll tubes are lenticular,watery,yellowish color,1—2 mm in diameter,with entire margins.In PY-cellulose agar roll tubes they are pin-like,less than lmm in diameter,raised,colorless,translucent,with an irregular margins.After incubation for 5 days,colonies are surr(?)unded byclear zones,representing cellulose hydrolysis,that usually are 3 to 7 mm in diameter.Cultures inPY-cellobiose broth are moderate turbid with a stringy sediment and have a pH of 5.0 afterincubation for 5 days.The optimum temperature for growth is 50—53℃.There is little growthat 37℃,and none at 65℃.The optimum pH for growth is 7.5.No growth in media withoutfermentable carbohydrates,in a chemically defined medium,growth factors required are bio-tin,pyridoxine,vitamin B12,and p-aminobenzoic acid.Fermentation products(per 100 ml ofculture)are 1.7—2.4 meq of succinic acid,1.0—1.2 meq of acetic acid,less than 1 meq ofethanol,hydrogen,and carbon dioxide in 1% PY-cellobiose or PY-cellulose medium.The mol% G+C of the DNA is 38.8(Tm).Other characteristics of the species are given in Table 1.The above characteristics show that the species differ from the known close species of genusClostridium,sach as C.stercorarium,and other cellulolytic thermophile:C.thermocellum,C.thermolacticum,C.josui and C.thermocoprial.The name Clostridium succinogenumsp.nov.is proposed for this species,referring to chief product in nonvolatile acid of the spe-cies.The type strain(H11)has been deposited in the General Microbiological Center of ChinaCommittee of Culture Collection of Microorganism as AS 1.1782.