Viprey Manon, Guillou Laure, Ferréol Martial, Vaulot Daniel
Station Biologique de Roscoff, UMR7144, CNRS et Université Pierre et Marie Curie, BP74, 29682 Roscoff Cedex, France.
Environ Microbiol. 2008 Jul;10(7):1804-22. doi: 10.1111/j.1462-2920.2008.01602.x. Epub 2008 Apr 21.
The genetic diversity of picoplanktonic (i.e. cells that can pass through a 3 mum pore-size filter) green algae was investigated in the Mediterranean Sea in late summer by a culture-independent approach. Genetic libraries of the 18S rRNA gene were constructed using two different primer sets. The first set is commonly used to amplify the majority of eukaryotic lineages, while the second was composed of a general eukaryotic forward primer and a reverse primer biased towards the phylum Chloroplastida. A total of 3980 partial environmental sequences were obtained: 1668 using the general eukaryotic primer set and 2312 using the Chloroplastida-biased primer set. Of these sequences, 65 (4%) and 594 (26%) belonged to the Chloroplastida respectively. A 99.5% sequence similarity cut-off value allowed classification of these 659 Chloroplastida sequences into 74 different operational taxonomic units. A majority of the Chloroplastida sequences (99%) belonged to the prasinophytes. In addition to the seven independent prasinophyte lineages previously described, we discovered two new clades (clades VIII and IX), as well as a significant genetic diversity at the species and subspecies levels, notably among the genera Crustomastix, Dolichomastix and Mamiella (Mamiellales), but also within Pyramimonas and Halosphaera (Pyramimonadales). Such diversity within prasinophytes has not previously been observed by cloning approaches, illustrating the power of using targeted primers for clone library construction. Prasinophyte assemblages differed especially in relation to nutrient levels. Micromonas and Ostreococcus were mainly recovered from mesotrophic areas, whereas Mamiella, Crustomastix and Dolichomastix were mostly detected in oligotrophic surface waters. Within genera such as Ostreococcus or Crustomastix for which several clades were observed, depth seemed to be the main factor controlling differential distribution of genotypes.
夏末,采用不依赖培养的方法,在地中海对微微型浮游绿藻(即能够通过孔径为3微米的滤膜的细胞)的遗传多样性进行了研究。使用两套不同的引物构建了18S rRNA基因的遗传文库。第一套引物常用于扩增大多数真核生物谱系,而第二套引物由一个通用的真核生物正向引物和一个偏向叶绿体门的反向引物组成。总共获得了3980条部分环境序列:使用通用真核引物组获得1668条,使用偏向叶绿体门的引物组获得2312条。在这些序列中,分别有65条(4%)和594条(26%)属于叶绿体门。99.5%的序列相似性截止值允许将这659条叶绿体门序列分类为74个不同的操作分类单元。大多数叶绿体门序列(99%)属于绿藻纲。除了先前描述的七个独立的绿藻纲谱系外,我们还发现了两个新的分支(第八和第九分支),以及物种和亚种水平上显著的遗传多样性,特别是在壳状鞭毛藻属、长鞭毛藻属和玛氏藻属(玛氏藻目)中,在扁藻属和球石藻属(扁藻目)内也存在这种多样性。以前通过克隆方法尚未观察到绿藻纲内的这种多样性,这说明了使用靶向引物构建克隆文库的作用。绿藻纲组合尤其在营养水平方面存在差异。微小绿球藻和聚球藻主要从中等营养区域获得,而玛氏藻、壳状鞭毛藻和长鞭毛藻大多在贫营养表层水体中检测到。在观察到多个分支的诸如聚球藻属或壳状鞭毛藻属等属内,深度似乎是控制基因型差异分布的主要因素。