Novel diversity within marine Mamiellophyceae (Chlorophyta) unveiled by metabarcoding

Abstract

The Ocean Sampling Day (OSD) project provided metabarcoding data for the V4 hyper-variable regions of the 18S rRNA gene from 157 samples collected at 143 mostly coastal stations. In this paper we focus on the class Mamiellophyceae, which was found at nearly all OSD stations and represented 55 % of the green microalgae (Chlorophyta) reads in the 2014 OSD dataset. We performed phylogenetic analyses of unique OSD metabarcodes (ASV, amplicon single variants) and reference GenBank sequences from cultures and from the environment, focusing on the four most represented genera: Ostreococcus (45 % of the Mamiellophyceae reads), Micromonas (34 %), Bathycoccus (10 %) and Mantoniella (8.7 %).These analyses uncovered novel diversity within each genus except Bathycoccus. In Ostreococcus, a new clade (E) with 2 very clear base pair differences compared to the oceanic clade B in the V4 region was the second most represented clade after the coastal Ostreococcus "lucimarinus". Within Micromonas, ten clades were found exceeding the 4 species and 2 candidate species already described. Finally, we found 2 new environmental clades of Mantoniella. Each Mamiellophyceae clade had a specific distribution in the OSD dataset suggesting that they are adapted to different ecological niches. Monomastigales 10 . Monomastigales are confined to freshwater environments while Dolichomasti-20 gales, although quite diversified in marine waters 12, 13 , are a minor component of Mamiel-21 lophyceae in coastal waters. In contrast, Mamiellales and Bathycoccales host some of the most 22 common Chlorophyta microalgae such as the ubiquitous Micromonas, the smallest known eu-23 karyote Ostreococcus or the coccoid Bathycoccus 10 . Within Mamiellales, Micromonas pusilla 14 24 was recently split into four species: Micromonas bravo (previously clade B.E.3), Micromonas com-25 moda (previous clade A.ABC.1-2), Micromonas polaris (previously clade B arctic), Micromonas 26 pusilla (previously clade C.D.5) and two clades mentioned as candidate species 1 (clade B. .4) 27 2/34 and candidate species 2 (clade B warm) 15 . Within the genus Mantoniella, only two species 28 have been described: the ubiquitous Mantoniella squamata 3, 16 , first described as Micromonas 29 squamata 17 , and Mantoniella antarctica 18 . Within Bathycoccales, four Ostreococcus clades have 30 been delineated 19 : Ostreococcus tauri 20 , Ostreococcus mediterraneus 21 , both of which have 31 been formerly described, Ostreococcus "lucimarinus" (clade A) and clade B, which both lack 32 formal taxonomic description. Analyses of pigment content and response to light levels allowed 33 to distinguish two broad ecotypes: strains adapted to high (O. tauri,O. mediterraneus, O. "luci-34 marinus" ) and low (Ostreococcus clade B) light 22 . The second genus within Bathycoccales hosts 35 a single species, Bathycoccus prasinos 23 . No clades can be delineated inside this species based 36 on 18S rRNA gene sequences from cultures and the environment. However, divergence in ITS 37 sequences suggest that Bathycoccus p...

Do inferences about freshwater phytoplankton communities change when based on microscopy or high‐throughput sequencing data?

“…While Chlorophyta showed relatively high % values with microscopy, with HTS they were rarely detected and with a low % sequence abundance. The discrepancy in the detection of Chlorophyta in a freshwater lake (this study) with respect to marine waters (Tragin et al, 2018; Tragin & Vaulot, 2019) might indicate that the efficiency of HTS for Chlorophyta can be habitat dependent. Dinophyta and Eustigmatophyta showed the odd situation of a higher diversity but a lower % abundance with HTS compared to microscopy.…”

“…Despite the higher richness of Charophyta reported with microscopy, microscopy and HTS data showed similarly low % values (generally <2%), corroborating that this phylum was a rare component of the phytoplankton community. For Chlorophyta, tufA (Vieira et al, 2016) or a combination of different primers should be used (Marcelino & Verbruggen, 2016); however, the V4 region of the 18S rRNA reveals a vast diversity of Chlorophyta in marine waters (Tragin, Zingone, & Vaulot, 2018; Tragin & Vaulot, 2019). While Chlorophyta showed relatively high % values with microscopy, with HTS they were rarely detected and with a low % sequence abundance.…”

Investigating temperature effects on coastal microbial populations and trophic interactions with 16S and 18S rRNA metabarcoding

“…Eukaryotes in the estuary were dominated (in terms of relative abundance) by Bacillariophyta, Mamiellophyceae, and Dinophyceae, consistent with other coastal 18S barcoding studies (Massana et al, 2015;Tragin et al, 2018;Gong et al, 2020). These protist groups (and many others) are functionally and phylogenetically diverse (Caron et al, 2012;Tragin and Vaulot, 2019), which may explain why we observed little fluctuation in class level relative abundance with temperature. Diatoms are known to exhibit species-specific temperature-growth relationships and occupy thermally driven environmental niches (Canesi and Rynearson, 2016;Rynearson et al, 2020).…”

Seasonal dynamics of natural Ostreococcus viral infection at the single cell level using VirusFISH

“…However, metatranscriptomic data from surface waters unveiled that the dominating species was O. lucimarinus. A previous study has shown that this species inhabits waters from the surface to the DCM (Rodríguez et al, 2005) and it is the most widely distributed, whereas O. tauri and O. mediterraneus are mostly restricted to the surface layer of coastal waters and lagoons (Rodríguez et al, 2005;Tragin and Vaulot, 2019). Thus, it is likely that most Ostreococcus cells found in our samples belonged to O. lucimarinus.…”

“…Ostreococcus (Mamiellaceae) is the world's smallest free-living eukaryote known, with a cell diameter of $1 μm (Courties et al, 1994;Derelle et al, 2006;Sanchez et al, 2019). The genus comprises several species that can be ubiquitously found from the coast to the open ocean, and from mesotrophic to oligotrophic waters (Demir-Hilton et al, 2011;Tragin and Vaulot, 2019). The number of sequenced Ostreococcus viruses is constantly increasing, and nowadays the complete genomes of many Ostreococcus viruses are available (e.g.…”