MOLECULAR TAXONOMY AND PHYLOGENOMICS OF CERAMIALES (RHODOPHYTA) HIGHLIGHT CHALLENGES AND ADVANCES IN UNDERSTANDING THE DIVERSITY AND SYSTEMATICS OF ALGAL TURFS

Algal turfs are ecosystem engineers receiving growing attention in relation to their expansion on temperate reefs as a consequence of global change. However, their species diversity and taxonomy are still poorly understood. Turfs are composed of densely entangled small seaweeds, in which the red algal order Ceramiales is commonly a major component. The advent of classical molecular taxonomy and -omic approaches has revolutionized investigations of seaweed diversity and systematics. These approaches have been applied to the study of the Ceramiales, facilitating the discovery of new species and resolving classification issues. A molecular diversity survey using the rbcL gene and involving the study of ca. 400 specimens of turf-forming rhodomelacean species in Macaronesia detected impressive amounts of newly discovered diversity. A total of 67 species were identified, of which half corresponded to undescribed species presumably endemic to this bioregion. Likewise, the application of molecular species delimitation methods to widely distributed records of Polysiphonia scopulorum (>150 samples collected in Australia, South Africa, southern Europe and Macaronesia) revealed that it is a complex in which 13 species were resolved. These 13 cryptic species range from endemics with narrow known distributions to a species found in all studied regions. At taxonomic levels above species, the use of plastid phylogenomics has produced well-resolved phylogenies that have been applied to the resolution of classification issues in the Ceramiales. These studies illustrate how classical and newer evolving molecular techniques facilitate the understanding of seaweed diversity and systematics, and provide new insights into the complexity of turf assemblages

Comparative Phylogeography of a Restricted and a Widespread Heather: Genetic Evidence of Multiple Independent Introductions of Erica Mackayana Into Ireland From Northern Spain

Financiado para publicación en acceso aberto: Universidade da Coruña/CISUG[Abstract] Species of flora and fauna occurring in the west of Ireland and south-west Europe, known as Lusitanian elements, constitute a puzzling case of isolated populations of uncertain origin. Here we studied the population genetic structure of the heather Erica mackayana in Ireland and northern Spain and compared it with its widespread close relative Erica tetralix using single nucleotide polymorphisms (SNPs). We reconstructed phylogenetic relationships using maximum likelihood (ML), inferred population genetic structure using cluster assignment and principal component analysis, and estimated population genetic diversity. The cluster analysis and ML phylogenetic tree showed a geographical pattern for E. tetralix supporting a post-glacial migration from Iberia to Ireland. In contrast, Irish populations of E. mackayana were supported in independent clades in the phylogenetic tree and shared clusters with Iberian populations in the structure analysis, and FST values were lower among Irish and Spanish populations than among Irish ones. This suggests that Irish populations of E. mackayana are the result of recent multiple independent introductions from its native area in northern Spain, probably assisted by humans. However, the origin of the largest Irish population at Roundstone Bog is unclear and should be further investigated. Post-glacial, long-distance dispersal is the most plausible explanation for Lusitanian species distribution in Ireland.This work was partially supported by Xunta de Galicia ‘Talento Senior’ (grant 03IN858A2019-1630129) to P.D-.T. and ‘Axudas para a consolidación e estruturación de unidades de investigación competitivas do SUG’ (grants ED431D 2017/20, ED431B 2018/49). Funding for open access charge: Universidade da Coruña/CISUGXunta de Galicia; 03_IN858A_2019_1630129Xutna de Galicia; ED431D 2017/20Xunta de Galicia; ED431B 2018/4

Phylogenetic analyses reveal a new old introduced red algal species in Europe

The detection of invasive species is commonly challenging in marine environments, often related with the insufficient taxonomic knowledge of the local diversity. Using DNA sequences in diversity surveys and their phylogenetic analysis can assist us to clarify the native or introduced origin of some species. We applied this approach to study the red algae Aphanocladia stichidiosa. It was originally described in the Mediterranean (1955), later recorded in Portugal (1960), the Canary Islands (1986), the northwestern Spain (1990) and the Azores (2012). Unexpectedly, our diversity surveys in Australia revealed its presence in Victoria in 2015, finding that European and Australian specimens had identical DNA sequences (rbcL gene) and suggesting that it is introduced in one of the regions. Phylogenetic analysis resolved A. stichidiosa in a highly supported clade with species restricted to the southern hemisphere, mainly from Australia. Accordingly, we conclude that A. stichidiosa is native from Australia, although it has never been recorded there before, and that it represents a relatively old introduction in Europe. In both native and introduced regions, it is frequent in algal turfs, a type of assemblage composed by a carpet of small, morphologically similar species whose identification is often difficult. The type of growth of this species explains the absence of previous records in Australia, where the number of taxonomists studying this assemblage is lower than in Europe. Sequential reports of A. stichidiosa in different European regions suggest that it is expanding its distribution, as well as its abundance is increasing. The invasive character of this species needs to be determined yet, and its small size should not lead to underestimate its potential harmfulness. In the current context, in which kelp forests are globally declining and algal turfs are expanding, this species might play a relevant role in the transformation of the European marine ecosystems

DIATOMS AND DINOFLAGELLATES DIVERSITY INHABITING A COASTAL UPWELLING SYSTEM: A METABARCODING APPROACH

Small eukaryotic plankton has been traditionally characterized using conventional microscopy techniques. Current advances in sequencing technologies allow the cost-effective study the diversity within microbial plankton based on DNA sequences. This technique has been implemented in the last 15 years for investigating prokaryotic diversity, but its application to unravel the diversity and ecology of eukaryotic organisms is still incipient. In this study we analyze the diversity of the diatoms and dinoflagellates in the marine communities using rDNA sequencing techniques, as well as the taxonomic resolution provided by the V4 region of the 18S rRNA gene. Twenty monthly 6L seawater samples for DNA metabarcoding were collected and filtered through 3 μm polycarbonate filter in a station off the Ría de A Coruña (NW Iberian Peninsula). DNA was extracted, the V4 region of the 18S rRNA was PCR amplified and subsequently sequenced using the High Throughput Sequencing (HTS) platform Illumina. Amplicon sequence variants (ASVs) were differentiated using DADA2 implemented in R. Sequences were aligned against PR2 v4.12.0 and SILVA 132 18S rRNA databases as references, as well as studied using BLAST and phylogenetic trees. Sequence-based taxonomic approach found 128 and 416 metabarcodes corresponding to diatoms and dinoflagellates, respectively. Sequences had limited resolution at species or even genus level, and overall the taxonomic resolution of diatoms was substantially higher than those of dinoflagellates

Nano- and microplankton diversity inhabiting a coastal upwelling system: a metabarcoding approach

Small eukaryotic plankton has been traditionally characterized using conventional microscopy techniques. Current advances in sequencing technologies allow the costeffective study the diversity within microbial plankton based on DNA sequences. This technique has been implemented in the last 15 years for investigating prokaryotic diversity, but its application to unravel the diversity and distribution of eukaryotic organisms is still incipient. Alleged advantages of this approach include the ability of detecting the smaller fraction of the community that trend to be overlooked in microscopy studies, as well as improving the resolution of taxonomic identification for groups whose morphological study is challenging due the paucity of morphological characters. In this study we characterize the diversity of the nano- and microplankton marine communities using microscopy and rDNA sequencing techniques. The aim of this work is to compare results of both techniques, assessing whether rDNA data can provide new insights into the study of the diversity of eukaryotic planktonic communities. Twenty monthly samples were collected in a station off the Ría de A Coruña (NW Iberian Peninsula). Planktonic samples for DNA studies were collected by filtering seawater samples through a 3 μm pore size polycarbonate filter. DNA was extracted, the V4 region of the 18S rRNA was PCR amplified and subsequently sequenced using the High Throughput Sequencing (HTS) platform Illumina. Amplicon sequence variants (ASVs) were differentiated using DADA2 implemented in R. Sequences were aligned against PR2 v4.12.0 and SILVA 132 18S rRNA databases as references. Microscopic identification of plankton taxa (phytoplankton and protozoa) was made using the Uthermöhl technique. In total 1182 ASVs were identified based on rDNA data, which contrasts with only 65 species identified by microscopy. Morphological identifications of eukaryotic plankton was restricted mainly to diatoms, dinoflagellates and ciliates, while rDNA data allowed the detection of additional taxonomic groups (i.e. protist). Some of them were very abundant, such as the algae Cryptophyceae and Mamiellophyceae or the heterotrophs Ascomycota or parasitic Stramenopiles. Therefore, our study further evidenced that DNA metabarcoding is a valuable tool to uncover part of the great diversity within microbial plankton, improving the detection of small eukaryotes

"Alsidium oliveiranum" sp. nov. (Rhodomelaceae, Rhodophyta), an overlooked species from the southwestern Atlantic based on morphology and DNA sequence data

[Abstract] The exploration of seaweed diversity in poorly studied habitats has often led to the discovery of new species. Sand-covered rocks are an example, as they received less attention than sand-free rocky intertidal habitats during seaweed diversity surveys in Brazil. In sand-covered rocks from Espírito Santo and Rio de Janeiro we found an alga whose morphology was unique among rhodomelacean species previously reported in Brazil. With the aim to clarify the taxonomic identity of this species we studied its morphology, as well as its phylogenetic relationships. Molecular analyses resolved this species in the genus Alsidium (tribe Alsidieae) and differed from sequenced congeners with divergences ≥2.5 and 4.2% in the rbcL and cox1 genes, respectively. Morphological characters were in agreement with the genus Alsidium, and differed from other species currently recognized in the genus. The species consisted of a basal crust and scarcely branched erect axes with seven pericentral cells covered by a continuous layer of cortical cells. Reproductive structures were formed on clusters of short determinate branches. Therefore, the new species A. oliveiranum is proposed based on morphological and molecular evidence. Our findings contribute to better understand the diversity of the tribe Alsidieae, which is particularly diverse in the Americas.Xunta de Galicia; GPC2015/025Brasil. Conselho Nacional de Desenvolvimento Científico e Tecnológico; 304899/2017–8Fundação de Amparo à Pesquisa do Estado de São Paulo; FAPESP 2016/50370-

UNDERSTOREY CHANGES COMPOSITION AFTER TEMPERATE KELP FOREST COLLAPSE BUT KEEPS RICHNESS AND DIVERSITY

Kelps are foundation species that provide important ecosystem services in temperate rocky shores worldwide. Similarly to terrestrial forests, healthy kelp forests are structurally complex as they are often arranged as patches composed by a multi-layered understorey of algae aggregations with different canopy adaptations. Over the last decades, several studies have reported a global kelp forest degradation, turning seascapes dominated by complex forest into structurally simpler mats of low-laying seaweeds. In NW Spain, golden kelp (Laminaria ochroleuca) canopy forests have recently receded within the limits of a MPA. This paradoxical loss inside a MPA allowed us to investigate the consequences of kelp forest collapse for other members of the biotic community, using nearby healthy kelp forest outside the MPA as a control. To assess these changes, four degraded sites within the MPA and four healthy kelp forest in nearby areas were sampled year round to assess the seasonal dynamics of understorey algae. Healthy and degraded kelp reefs had significantly different understorey assemblage compositions. However, unlike our expectations, these differences had little to none impact on the richness and diversity of the understorey assemblage. Moreover, understory differences were only perceptible when the assemblages were compared at the lowest taxonomic resolution (species). Unlike other studies, comparisons based on functional groups (canopy, sub-canopy, turf and crust) failed to detect any significant difference between healthy and degraded kelp forest

Seasonal niche of planktonic prokaryotes inhabiting surface waters of the upwelling region off NW Iberia

Prokaryotes play an important role in biogeochemical cycling in marine ecosystems, but little is known about their diversity and composition, and it’s even less understood how they may contribute to the ecological functioning of highly variable coastal areas affected by upwelling. Between May 2016 and May 2018, we carried out 26 one-day samplings in the temperate northwestern Iberian upwelling system to investigate the temporal patterns of variability for prokaryotic abundance, diversity and community composition by combining flow cytometry and 16S RNA high- throughput sequencing. A marked seasonality was found for prokaryotic abundance, peaking during summer upwelling and relaxation season (≈May to September), when extracellular release of organic matter from phytoplanktonic blooms is a significant process, and decreasing in downwelling events (≈October to April). Those downwelling conditions, characterized by deeper mixed layers and a homogeneous water column, favored a higher abundance of the archaeal groups, Marine Group II (Euryarchaeota) and Nitrosopelagicus (Thaumarchaeota), as well as of Marinimicrobia bacterium (SAR406 clade) and the group Bacteria_Others. By contrast, upwelling and relaxation conditions characterized by enhanced vertical stratification and hydrographic variability, included a community generally less diverse with core-phylotypes (occurring > 75% of the samples) proliferating, i.e. Flavobacteriaceae (Bacteroidetes), Chloroplast (Cyanobacteria) and Amylibacter (Proteobacteria). Overall, the environmental conditions explained 60% (R2=0.60, AIC=125.64) of the prokaryotic community composition, being temperature, inorganic nutrients, chlorophyll and particulate organic nitrogen the variables that best explained the variation in the prokaryotic community composition (r=0.40). Additional efforts are currently underway on a fine-tuning composition assessment (oligotypes composition from particular core-phylotypes) to study if this variability along the temporal environmental gradient could be associated with the differentiation of ecotypes (oligotype ́s seasonality within particular phylotypes). Overall, the present study provides new insights into the barely explored seasonal niche partitioning of surface prokaryotic community driven by hydrodynamic forcing in an upwelling system, which may support further investigations on the role of bacterioplankton in the biogeochemistry of these ecosystems

Seasonal niche of planktonic prokaryotes inhabiting surface waters of the upwelling region off NW Iberia

Oral communicationProkaryotes play an important role in biogeochemical cycling in marine ecosystems, but little is known about their diversity and composition, and it’s even less understood how they may contribute to the ecological functioning of highly variable coastal areas affected by upwelling. Between May 2016 and May 2018, we carried out 26 one-day samplings in the temperate northwestern Iberian upwelling system to investigate the temporal patterns of variability for prokaryotic abundance, diversity and community composition by combining flow cytometry and 16S RNA high- throughput sequencing. A marked seasonality was found for prokaryotic abundance, peaking during summer upwelling and relaxation season (≈May to September), when extracellular release of organic matter from phytoplanktonic blooms is a significant process, and decreasing in downwelling events (≈October to April). Those downwelling conditions, characterized by deeper mixed layers and a homogeneous water column, favored a higher abundance of the archaeal groups, Marine Group II (Euryarchaeota) and Nitrosopelagicus (Thaumarchaeota), as well as of Marinimicrobia bacterium (SAR406 clade) and the group Bacteria_Others. By contrast, upwelling and relaxation conditions characterized by enhanced vertical stratification and hydrographic variability, included a community generally less diverse with core-phylotypes (occurring > 75% of the samples) proliferating, i.e. Flavobacteriaceae (Bacteroidetes), Chloroplast (