微生物学报Acta Microbiologica Sinica
48(4):421~425; 4 April 2008
徐春光1,2, 梁秀梅2 , 黄英1*, 王黎明1, 崔庆峰1
(1 中国科学院微生物研究所, 微生物资源前期开发国家重点实验室, 北京 100101)
(2 呼伦贝尔学院生命科学与化学学院, 呼伦贝尔 021008)
摘要:【目的】了解酸性土壤环境里中度嗜酸链霉菌的多样性, 调查其物种资源。【方法】用分散和差速离心法及选择性分离培养基从14 份云南酸性土壤样品中分离到367 株具有链霉菌培养特征的放线菌, 并进行了颜色分群。从各颜色类群中选取代表菌株共97 株, 通过显微形态观察和pH 梯度生长实验确定其中的中度嗜酸链霉菌。进一步从中筛选出16 株中度嗜酸链霉菌代表菌株, 进行16SrRNA 基因序列的相似性和系统发育分析, 并结合基因组DNA-DNA 相关性数据。【结果】分离菌株归为12 同的颜色类群, 其中80%属于中度嗜酸链霉菌, 其代表菌株在系统发育树上形成了8 个距离较远且与已知种不同的进化分枝, 可能代表链霉菌属内至少8 个不同的新基因种。【结论】用以上方法筛选出的中度嗜酸链霉菌可归为8 个不同于已知种的进化群, 说明云南酸性土壤含有丰富多样的中度嗜酸链霉菌新物种。
关键词:中度嗜酸链霉菌; 16S rRNA 基因; 系统发育分析; 多样性; 进化群
中图分类号:Q939 文献标识码:A 文章编号:0001-6209 (2008) 04-0421-05
在酸性和偏酸性自然环境特别是土壤中, 中度嗜酸放线菌(pH 生长范围4.5~7.5, 最适pH5.0~5.5)的数量远远超过严格嗜酸放线菌(pH 生长范围3.5~6.5,最适pH4.5), 而链霉菌是其中的绝对优势菌群, 人们推断中度嗜酸链霉菌在酸性土壤有机物质的降解和转化过程中扮演着重要角色[1~4]。近年来国内外的研究[4, 5]表明, 酸性土壤中嗜酸稀有放线菌具有较丰富的种属多样性。而对酸性环境中常见放线菌—链霉菌多样性的研究, 更有可能发现新的抗真菌活性物质和在低pH 下有高活性的酶类, 并且对阐明它们的生态作用具有普遍意义[6, 7]。1991 年, Park 等[8]用5S rRNA 基因序列对专性嗜酸链霉菌、中度嗜酸链霉菌和嗜中性链霉菌进行了系统发育学研究, 发现它们分别在进化树上形成独立分枝。2003 年, Kim 等[9]利用分散和差速离心法从酸性土壤中分离到3 株专性嗜酸链霉菌, 并建立了链嗜酸菌新属(Streptacidiphilus gen. nov.)。2004 年, Kim等[10]对一群中度嗜酸链霉菌的代表菌株KCTC 9926T进行了多相分类研究, 发现此菌株在链霉菌属内形成一个单独的分枝, 并将它所代表的菌群定为链霉菌属的一个新种(Streptomyces yeochonensis sp. nov.)。我们从云南酸性土壤中分离纯化到一批嗜酸链霉菌,通过表型特征和分子系统学分析, 对其中的中度嗜酸链霉菌的物种多样性进行了研究。
Isolation and Identification of the Temperate Bacteriophage from Isolated Strains of Streptococcus suis Serotype 2
Yuling Ma, Chengping Lu*, Hongjie Fan
(College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210095, China)
Abstract: A PCR assay was developed to study the distributional characteristics of phage integrase gene in Streptococcus suis serotype 2 (SS2). A 323bp distinct DNA target can be amplified in 25 strains of virulent SS2, while can not be amplified in avirulent strain T15, 5 strains of other serotypes(SS1, SS7, SS9) and strains of group C Streptococcus strains from pigs, which suggested that the phage integrase gene may be related to the pathogenicity of SS2 and can be consider as a detection factor of the virulent gene of SS2. The sequencing and restriction endonuclease analysis of the PCR products were also done. Comparisons between the sequences of phage integrase gene with that of SS2 strain, showed a high homology with SS2 China strains 98HAH33, 05ZYH33 and North American strain 89.1591. Complete cell lysis was observed with SS2 virulent strains but not with avirulent strain T15 after the induction by mitomycin C. Electron microscopy analysis of the lysate from SS2 virulent strains HA9801 and ZY05719 revealed the presence of phage particles. The induced phage, named SS2-HA and SS2-ZY, both have a small isometric nucleocapsid approximately 50 nm in diameter and have no tail and is therefore a member of the Tectiviridae family. The phage integrase gene sequence of phage SS2-HA and SS2-ZY shared high homologue identities with virulent SS2 strains, which suggested that the phage integrase gene of SS2 has high specify. The temperate phage and phage integrase gene can only detected from SS2 virulent strains but not from avirulent strain, and the detection of phage integrase gene was related to the virulence-associate factors of SS2, such as the muramidase-released protein gene (mrp), which suggested that the temperate phage of SS2 may be related to the pathogenicity of SS2.
Keywords: Streptococcus suis type 2; temperate bacteriophage; induce
基金项目: 国家自然科学基金(30770002); 国家“973 项目”(2004CB719601)
*通讯作者。Tel/Fax: +86-10-64807311; E-mail: huangy@im.ac.cn
作者简介: 徐春光(1978. ), 女, 内蒙古呼伦贝尔人, 呼伦贝尔学院生命科学与化学学院助教, 主要从事放线菌系统分类学研究。
收稿日期: 2007-10-29; 修回日期: 2008-01-03
ISSN 0001-6209; CN11-1995/Q
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