Environmental DNA surveys have revealed a great deal of hidden diversity within the Cercozoa. An investigation into the biodiversity of heterotrophic flagellates in marine benthic habitats of British Columbia, Canada, demonstrated the presence of several undescribed taxa with morphological features that resemble the cercozoan genera Cryothecomonas and Protaspis. Nine novel species of marine interstitial cercozoans are described that are distributed into five genera, four of which are new. Phylogenetic analyses of small subunit rDNA sequences derived from two uncultured isolates of Protaspis obliqua and nine novel cercozoan species (within four novel genera) provided organismal anchors that helped establish the cellular identities of several different environmental sequence clades. These data, however, also showed that the rarity of distinctive morphological features in cryomonads, and other groups of cercozoans, makes the identification and systematics of the group very difficult. Therefore, a DNA barcoding approach was applied as a diagnostic tool for species delimitation that used a 618 bp region at the 5′ end of the SSU rDNA sequence. Nucleotide sequence analysis of this region showed high intergeneric sequence divergences of about 7 % and very low intraspecific sequence divergences of 0–0.5 %; phylogenetic analyses inferred from this barcoding region showed very similar tree topologies to those inferred from the full-length sequence of the gene. Overall, this study indicated that the 618 bp barcoding region of SSU rDNA sequences is a useful molecular signature for understanding the biodiversity and interrelationships of marine benthic cercozoans.
Al QassabS.,
LeeW. J.,
MurrayS.,
PattersonD. J.2002; Flagellates from stromatolites and surrounding sediments in Shark Bay, Western Australia. Acta Protozool 41:91–144
AuerB.,
ArndtH.2001; Taxonomic composition and biomass of heterotrophic flagellates in relation to lake trophy and season. Freshw Biol 46:959–972[CrossRef]
BarthD.,
KrenekS.,
FokinS. I.,
BerendonkT. U.2006; Intraspecific genetic variation in Paramecium revealed by mitochondrial cytochrome c oxidase I sequences. J Eukaryot Microbiol 53:20–25[CrossRef]
BassD.,
Cavalier-SmithT.2004; Phylum-specific environmental DNA analysis reveals remarkably high global biodiversity of Cercozoa (Protozoa). Int J Syst Evol Microbiol 54:2393–2404[CrossRef]
BerneyC.,
FahrniJ.,
PawlowskiJ.2004; How many novel eukaryotic ‘kingdoms’? Pitfalls and limitations of environmental DNA surveys. BMC Biol 2: 13 [CrossRef]
Cavalier-SmithT.1998b; Neomonada and the origin of animals and fungi
. In Evolutionary Relationships Among Protozoa pp 375–407 Edited by
CoombsG. H.,
VickermanK.,
SleighM. A.,
WarrenA.
London: Kluwer Academic Publishers;
ChantangsiC.,
LynnD. H.,
BrandlM. T.,
ColeJ. C.,
HetrickN.,
IkonomiP.2007; Barcoding ciliates: a comprehensive study of 75 isolates of genus Tetrahymena
. Int J Syst Evol Microbiol 57:2412–2425[CrossRef]
ChantangsiC.,
EssonH. J.,
LeanderB. S.2008; Morphology and molecular phylogeny of a marine interstitial tetraflagellate with putative endosymbionts: Auranticordis quadriverberis n. gen. et sp. (Cercozoa). BMC Microbiol 8: 123 [CrossRef]
GuindonS.,
GascuelO.2003; PhyML - A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol 52:696–704[CrossRef]
HondeveldB. J. M.,
BakR. P. M.,
van DuylF. C.1992; Bacterivory by heterotrophic nanoflagellates in marine sediments measured by uptake of fluorescently labeled bacteria. Mar Ecol Prog Ser 89:63–71[CrossRef]
HoppenrathM.,
LeanderB. S.2006a; Dinoflagellate, euglenid or cercomonad? The ultrastructure and molecular phylogenetic position of Protaspis grandis n. sp. J Eukaryot Microbiol 53:327–342[CrossRef]
KimuraM.1980; A simple method of estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120[CrossRef]
KühnS. F.,
LangeM.,
MedlinL. K.2000; Phylogenetic position of Cryothecomonas inferred from nuclear-encoded small subunit ribosomal RNA. Protist 151:337–345[CrossRef]
LeeW. J.2008; Free-living heterotrophic euglenids from marine sediments of the Gippsland Basin, southeastern Australia. Mar Biol Res 4:333–349[CrossRef]
LeeW. J.,
SimpsonA. G. B.,
PattersonD. J.2005; Free-living heterotrophic flagellates from freshwater sites in Tasmania (Australia), a field survey. Acta Protozool 44:321–350
LynnD. H.,
Strüder-KypkeM. C.2006; Species of Tetrahymena identical by small subunit rRNA gene sequences are discriminated by mitochondrial cytochrome c oxidase I gene sequences. J Eukaryot Microbiol 53:385–387[CrossRef]
NormanJ. E.,
GrayM. W.1997; The cytochrome oxidase subunit 1 gene ( cox1 ) from the dinoflagellate, Crypthecodinium cohnii
. FEBS Lett 413:333–338[CrossRef]
ParkS. J.,
ParkB. J.,
PhamV. H.,
YoonD. N.,
KimS. K.,
RheeS. K.2008; Microeukaryotic diversity in marine environments, an analysis of surface layer sediments from the East Sea. J Microbiol 46:244–249[CrossRef]
SaundersG. W.2005; Applying DNA barcoding to red macroalgae: a preliminary appraisal holds promise for future applications. Philos Trans R Soc Lond B Biol Sci 360:1879–1888[CrossRef]
SchnepfE.,
KühnS. F.2000; Food uptake and fine structure of Cryothecomonas longipes sp. nov., a marine nanoflagellate incertae sedis feeding phagotrophically on large diatoms. Helgol Mar Res 54:18–32[CrossRef]
ŠlapetaJ.,
MoreiraD.,
López-GarcíaP.2005; The extent of protist diversity: insights from molecular ecology of freshwater eukaryotes. Proc Biol Sci 272:2073–2081[CrossRef]
ThompsonJ. D.,
HigginsD. G.,
GibsonT. J.1994; clustal w: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res 22:4673–4680[CrossRef]
ThomsenH. A.,
BuckK. R.,
BoltP. A.,
GarrisonD. L.1991; Fine structure and biology of Cryothecomonas gen. nov. (Protista incertae sedis ) from the ice biota. Can J Zool 69:1048–1070[CrossRef]
VørsN.1993; Heterotrophic amoebae, flagellates and heliozoa, from Arctic marine waters (North West Territories. Canada and West Greenland). Polar Biol 13:113–126