Seasonal dynamics of free-living tintinnid ciliate communities revealed by environmental sequences from the North-West Mediterranean Sea.

The species-rich order Tintinnida is a group of planktonic ciliates ubiquitous in coastal marine waters, which can be well described using molecular estimates of diversity. We studied temporal changes of tintinnid diversity over 1 year in a coastal Mediterranean Sea location (Villefranche-sur-Mer, France) at five different depths (5, 25, 50, 100, and 160 m) and one additional year at 50 m depth by combining denaturing gel gradient electrophoresis (DGGE) community fingerprinting with direct PCR amplification, cloning and sequencing of small subunit (SSU), 5.8S, and large-subunit (LSU) rRNA genes together with the corresponding internal transcribed spacers (ITS). We amplified tintinnid sequences in all samples. All identified phylotypes were closely related to described species, showing that there is a good phylogenetic reference sequence data set allowing accurate estimation of tintinnid diversity in these waters. Tintinnid community composition exhibited marked seasonal changes. Surprisingly, the tintinnid SSU rDNA-based species composition did not show any clear relationship to measured environmental parameters (temperature, salinity, light, phytoplankton biomass). Nonetheless, the comparison of tintinnid community composition using UniFrac revealed three significant clusters of sequences grouping, respectively, samples collected in winter, autumn, and summer, leading to the hypothesis that seasonal effects on tintinnid community composition might be related to biotic parameters. In addition, phylogenetic trees based on the concatenated SSU + LSU rDNA and ITS sequences showed a better resolution than SSU rDNA alone to discriminate closely related species.