浮霉菌门严格厌氧产氢细菌(Thermopirellula anaerolimosa)的分离及其生理特性

Isolation and characterization of Thermopirellula anaerolimosa gen. nov., sp. nov., an obligate anaerobic hydrogen-producing bacterium of the phylum Planctomycetes

【目的】厌氧颗粒污泥中含有大量未知微生物资源,利用低浓度底物及添加抗生素的培养基进行厌氧发酵细菌的筛选,并对分离菌株进行生理生化特性研究。【方法】利用系列稀释法及亨盖特厌氧滚管技术从制糖废水厌氧处理反应器的颗粒污泥中分离到一株高温厌氧产氢细菌VM20-7T,通过16S rRNA基因序列同源性确定其系统发育地位。【结果】菌株VM20-7T为高温、严格厌氧、革兰氏阴性梨形细菌,细胞大小为(0.7-2.0)μm×(0.7-2.0)μm,不运动,不产芽胞。其生长温度范围为35℃-50℃(最适温度45℃),pH范围为6.0-8.3(最适pH 7.0-7.5),NaCl耐受范围为0%-0.5%(w/v,最适浓度0%)。菌株VM20-7T可利用葡萄糖、麦芽糖、核糖等多种糖类为唯一碳源生长,葡萄糖发酵终产物是乙酸和H2。该菌株不利用硝酸盐、硫酸盐等作为电子受体生长。G+C含量为60.9 mol%,16S rRNA基因序列同源性显示菌株属于浮霉菌门,但与已培养菌株的同源性较低,与梨形菌属-红小梨形菌属-芽殖小小梨形菌属(Pirellula-Rhodopirellula-Blastopirellula,PRB)分支的亲缘关系最近,但序列相似性也仅为82.7%-84.3%。【结论】利用低浓度糖类并添加抗生素分离厌氧颗粒污泥中的微生物,获得了浮霉菌门首例严格厌氧细菌VM20-7T。生理生化特性和系统发育分析显示,菌株VM20-7T为浮霉菌目的新属新种,命名为Thermopirellula anaerolimosa。该菌株的菌种保藏号为CGMCC 1.5169T=JCM 17478T=DSM 24165T。

[Objective]To cultivate various yet-to-be cultured heterotrophs from anaerobic granule sludge,we used a selective culture medium with low concentrations of substrates supplemented a variety of antibiotics.[Methods] An obligate anaerobic,thermophilic,hydrogen-producing bacterium,strainVM20-7T,was isolated from an upflow anaerobic sludge blanket（UASB） reactor treating high-strength organic wastewater from isomerized sugar production processes.[Results] Cells of strain VM20-7T are non-motile,spherical,pear or teardrop shaped,occurring singly or as aggregates（0.7-2.0 μm×0.7-2.0 μm）.Spore formation was not observed.Growth temperature ranges from 35-50℃（optimum 45℃）,pH ranges from 6.0-8.3（optimum 7.0-7.5）,NaCl tolerant concentration ranges from 0%-0.5%（w/v,optimum 0%）.Nitrate,sulfate,thiosulfate,sulfite,elemental sulfur and Fe（III）-NTA were not used as terminal electron acceptors.Strain VM20-7T utilizes a wide range of carbohydrates,including glucose,maltose,ribose,xylose,sucrose,galactose,mannose,raffinose,pectin,yeast extract and xylan.Acetate and H2 are the main end products of glucose fermentation.The G＋C content of the genomic DNA was 60.9 mol%.16S rRNA gene sequence analysis revealed that it is related to the Pirellula-Rhodopirellula-Blastopirellula（PRB） clade within the order Planctomycetales（82.7-84.3% similarity with 16S rRNA genes of other known related species）.[Conclusion] The first obligate anaerobic bacterium within the phylum Planctomycetes was isolated with low concentration of carbohydrates and antibiotics.On the basis of the physiological and phylogenetic data,the name Thermopirellula anaerolimosa gen.nov.,sp.nov.is proposed for strain VM20-7T（=CGMCC 1.5169T= JCM 17478T = DSM 24165T）.