一株脱卤单胞菌属有机卤呼吸细菌的分离纯化与基础特征

Isolation and basic characterization of a novel organohalide-respiring bacterium within the genus Dehalogenimonas

摘要:

[目的] 分离鉴定新型有机卤呼吸细菌，拓展有机氯污染物降解菌种资源。[方法] 在限制性培养基中，基于特定脱卤微生物的能量代谢特点以及对抗生素的抗性特征，应用绝迹稀释法从脱氯富集培养物中分离纯化新型有机卤呼吸细菌。通过添加酵母提取物和聚合酶链反应-限制性片段长度多态性等方法鉴定菌株纯度。通过考察细胞形态、16S rRNA系统发育树以及对有机氯化物的利用能力来描述此新型菌株的基本特征。[结果] 菌株GP是一株来自脱卤单胞菌属（Dehalogenimonas）的新型有机卤呼吸细菌，可耐受1.0 g/L的氨苄青霉素和0.1 g/L的万古霉素，其细胞平面形态为直径0.4-0.8 μm的不规则圆形。在乙酸作碳源、氢气作电子供体的条件下，菌株GP可利用1，1-二氯乙烯和一氯乙烯作为电子受体进行有机卤呼吸，脱氯终产物为无毒害的乙烯。16S rRNA系统发育分析表明，菌株GP与Dehalogenimonas formicexedens菌株NSZ-14有高达99.5%的同源性，但两菌株对有机氯化物底物的利用范围不同。[结论] 从脱氯富集培养物中分离纯化出一株新型有机卤呼吸细菌，为深入研究脱卤单胞菌这一重要脱卤微生物的遗传学信息和生理生化性质提供了新材料。

Abstract:

[Objective] To isolate and characterize a novel organohalide-respiring bacterium and expand the diversity of dehalogenating microorganisms. [Methods] Based on the specialized energy metabolism and antibiotic resistance of certain dehalogenating microorganisms, we adapted the dilution-to-extinction strategy in defined mineral salts medium for the isolation of an anaerobic bacterium from a dehalogenating enrichment culture. Culture purity was investigated by yeast extract addition to growth medium and PCR-RFLP analysis on 16S rRNA gene amplicon. The basic characteristics of this novel microorganism were described in terms of cell morphology, 16S rRNA gene phylogeny and the ability to utilize chlorinated compounds as electron acceptors. [Results] A novel organohalide-respiring bacterial isolate, strain GP, belonging to Dehalogenimonas genus, was obtained. Strain GP is able to dechlorinate 1,1-dichloroethene (1,1-DCE) and grow at the presence of 1 g/L ampicillin and 0.1 g/L vancomycin. Cell morphology of strain GP is irregular disc-shaped coccus with a 0.4-0.8 μm diameter. Strain GP couples growth with reductive dechlorination of 1,1-DCE and vinyl chloride to nontoxic ethene using acetate as carbon source and hydrogen as electron donor. Phylogenetic analysis indicated that the 16S rRNA gene of strain GP shares 99.5% sequence similarity to that of Dehalogenimonas formicexedens NSZ-14, which grows with different types of chlorinated compounds. [Conclusion] A novel organohalide-respiring bacterium strain GP was isolated from a dehalogenating enrichment culture. This work expands the Dehalogenimonas pangenome and provides new biomaterial for detailed biochemical and physiological characterizations of Dehalogenimonas microorganisms.