PHYLOGENETIC ANALYSIS OF 32 STRAINS OF VAUCHERIA (XANTHOPHYCEAE) USING THE rbcL GENE AND ITS TWO FLANKING SPACER REGIONS1

Abstract

The complete rbc L gene was sequenced for 21 spe-cies and 32 strains of Vaucheria and for five other Xan-thophyceae (Asterosiphon dichotomus (Kützing) Rieth, Botrydium becharianum Vischer, B. cystosum Vischer, B. stoloniferum Mitra, Tribonema intermixtum Pascher). The psb A-rbc L spacer, upstream of the rbc L gene, and the RUBISCO spacer between the rbc L and rbc S genes were also completely sequenced for the Vau-cheria strains and Asterosiphon. The psb A-rbc L spacer was the most variable region that was sequenced, and only the 3 Ј end of the spacer could be aligned. Phylogenetic analyses (maximum parsimony, neigh-bor joining, and maximum likelihood) were con-ducted using the DNA sequence and the amino acid sequence for the rbc L gene, and a second analysis was conducted using a portion of the psb A-rbc L spacer ϩ rbc L gene ϩ RUBISCO spacer. All analyses showed that Vaucheria species formed monophyletic clades that corresponded with morphologically based subgeneric sections, including the section Racemo-sae. Species producing a gametophore (ϭ fruiting branch, bearing both an antheridium and oogonium) formed a monophyletic clade in all analyses. The nongametophore species sometimes formed a mono-phyletic clade but other times formed a basal grade. Pair-wise comparisons of nucleotides and amino ac-ids showed that for some species, numerous nucle-otide changes resulted in relatively few amino acid changes. Consequently, phylogenetic analysis of the amino acids produced numerous trees, which in a strict consensus tree resulted in numerous polychot-omies. An original strain of V. terrestris that was de-posited in two culture collections over 25 years ago had identical sequences, suggesting no rapid change was occurring in the sequenced regions. Two strains of V. prona , isolated from Europe and North Amer-ica, had identical sequences. Other species, for which two or more strains were examined, had different se-quences. These results suggest that cryptic species complexes exist within Vaucheria because the rbc L gene is a conservative gene that is identical in other protists.