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Multigene phylogenies of clonal Spumella-like strains, a cryptic heterotrophic nanoflagellate, isolated from different geographical regions

Thorsten Stoeck¹, Steffen Jost^1, and Jens Boenigk²

¹ School of Biology, University Kaiserslautern, Erwin-Schrödinger Str. 14, D-67663 Kaiserslautern, Germany
² Austrian Academy of Sciences, Institute for Limnology, Mondseestr. 9, A-5310 Mondsee, Austria

Correspondence
Thorsten Stoeck
stoeck{at}rhrk.uni-kl.de

Many nanoflagellate morphospecies comprise an enormous variation of genotypes, probably indicating cryptic species. One of the best-investigated morphospecies with respect to molecular and ecophysiological variation are flagellates of the Spumella morphotype. Here, we have phylogenetically analysed three protein-coding genes (actin, alpha-tubulin, beta-tubulin), internal transcribed spacers (ITS1, ITS2) and the 5.8S rDNA of 17 Spumella-like strains isolated from soil, freshwater and marine samples in order to (i) test the validity of the current Spumella-like phylogenetic classification system based exclusively on small subunit (SSU) rDNA, (ii) elucidate the phylogenetic associations of SSU rDNA-unresolved strains and (iii) evaluate the validity of the assignment of ecophysiological adaptations to previously identified SSU rDNA sequence clades. All single-gene analyses show different patterns of support, are incongruent and identify a number of conflicting nodes. Likewise, a concatenation of all protein genes fails to recover specific SSU rDNA clades. However, a combined analysis of all genes confidently resolved the conflicts of the single genes and the protein-gene concatenations and resulted in a tree topology that is identical to the SSU rDNA analysis, but with enhanced phylogenetic resolution and decisively greater support. We conclude that, depending on the genes concatenated, a ‘supergene’ analysis minimizes artefactual effects of single genes and may be superior in its performance in phylogenetically analysing cryptic species. We confirm the validity of the SSU rDNA Spumella-like phyloclades and support the suggestion that these clades indeed seem to reflect certain ecophysiological adaptations.

Present address: Austrian Academy of Sciences, Institute for Limnology, Mondseestr. 9, A-5310 Mondsee, Austria.

The GenBank/EMBL/DDBJ accession numbers for the SSU rDNA, ITS, alpha- and beta-tubulin and actin sequences reported in this paper are EF577164–EF577234, as detailed in Table 1 and Supplementary Table S1.

Details of GenBank accession numbers and further phylogenetic trees for the ITS locus, the actin gene, the alpha-tubulin gene and the beta-tubulin gene and concatenations of the actin, beta- and alpha-tubulin genes and the actin, beta-tubulin, alpha-tubulin and SSU rDNA genes are available as supplementary material with the online version of this paper.