Relationship between hydrocarbons and molecular phylogeny of Botryococcus braunii

The relationship between hydrocarbons and the molecular phylogeny of Botryococcus braunii was investigated using 31 axenic strains isolated in Japan. The resulting 18S rRNA tree, which includes additional sequences from the gene bank, revealed the existence of three major pylogenetic clades (I, II, and III) and four subclades (II1, II2, III1, and III2) with high bootstrap values. By gas chromatography/mass spectrometry analysis, nine hydrocarbon species were detected and categorized as 4 types: the three known races A, B, and L and the tentatively named race S comprising epoxy-n-alkane and saturated n-alkane chains with carbon numbers 18 and 20, respectively. Strains of clade I were coincident with race A. Clade II contained race B, and clade III contained race L and its derivatives, and race S. Clade III was further categorized into subclade III1 containing the typical race L strains and subclade III2 containing race S with high bootstrap values. The phylogenetic relationship of Botryococcus strains appeared to be in considerable agreement with unique hydrocarbon synthesis pathways. Maximum divergence values of 18S rRNA indicated large speciation among the clades at almost the species level, suggesting the utility of biochemical properties, e.g., hydrocarbon types, in determining the taxonomy of Botryococcus.