Results of the study of invasive algae in National Parks published in Marine Pollution Bulletin

by Xavier Turon

We are happy to report that the results concerning the invaded vs non-invaded communities sampled in 2015 have now been published in the international journal Marine Pollution Bulletin: Wangensteen OS, Cebrian E, Palacín C, Turon X. 2018. Under the canopy: Community-wide effects of invasive algae in Marine Protected Areas revealed by metabarcoding. Marine Pollution Bulletin, 127:54-66.

Elsevier kindly provides a 50 days free access (starting 21 Dec) to this article at https://authors.elsevier.com/a/1WG02,ashmKxI. After this period you will need a subscription to be able to download it

In this paper we report on the effect that invasive algae have at the community level, targeting mainly the smaller size components (small macrofauna and meiofauna), using the 18S and the COI genes and the fractionation procedure and bioinformatic pipeline developed in our project. Here goes the abstract:

We analysed with multigene (18S and COI) metabarcoding the effects of the proliferation of invasive seaweeds on rocky littoral communities in two Spanish Marine Protected Areas. The invasive algae studied were Caulerpa cylindracea, Lophocladia lallemandii and Asparagopsis armata. They are canopy-forming, landscape-dominant seaweeds, and we were interested in their effects on the underlying communities of meiobenthos and macrobenthos, separated in two size fractions through sieving. A new semiquantitative treatment of metabarcoding data is introduced. The results for both markers showed that the presence of the invasive seaweed had a significant effect on the understory communities for Lophocladia lallemandii and Asparagopsis armata but not for Caulerpa cylindracea. Likewise, changes in MOTU richness and diversity with invasion status varied in magnitude and direction depending on the alga considered. Our results showed that metabarcoding allows monitoring of the less conspicuous, but not least important, effects of the presence of dominant invasive seaweeds.

Images of the communities analysed, with (left) and without (right) invasive algae. A,B: Cc (Caulerpa cylindracea) communities. C,D: Ll (Lophocladia lallemandii) communities. E,F: Aa (Asparagopsis armata) communities. Pictures: (A) from Eneko Aspillaga; (B,F) from the authors; (C,D) from Pol Capdevila; (E) from Enric Ballesteros.

Results from metabarcoding communities from the National Parks published in PeerJ Preprints

by Xavier Turon

While the article is being peer-reviewed, we have published the manuscript in Pre-print format in the repository PeerJ Preprints, where it is freely available at Wangensteen OS, Palacín C, Guardiola M, Turon X. 2017. Metabarcoding litoral hard-bottom communities: unexpected diversity and database gaps revealed by two molecular markers. PeerJ Preprints 5:e3429v1.

We report in this manuscript the astonishing diversity found in communities from Islas Cies and Cabrera. We address several technical aspects, compare the two genes studied (18S and COI), and perform ecological analyses. Here goes the abstract:

We developed a metabarcoding method for biodiversity characterization of structurally complex natural marine hard-bottom communities. Novel primer sets for two different molecular markers: the “Leray fragment” of mitochondrial cytochrome c oxidase, COI, and the V7 region of ribosomal RNA 18S were used to analyse eight different marine shallow benthic communities from two National Parks in Spain (one in the Atlantic Ocean and another in the Mediterranean Sea). Samples were sieved into three size fractions from where DNA was extracted separately. Bayesian clustering was used for delimiting molecular operational taxonomic units (MOTUs) and custom reference databases were constructed for taxonomic assignment. We found unexpectedly high values for MOTU richness, suggesting that these communities host a large amount of yet undescribed eukaryotic biodiversity. Significant gaps are still found in sequence reference databases, which currently prevent the complete taxonomic assignation of the detected sequences. Nevertheless, over 90% (in abundance) of the sequenced reads could be successfully assigned to phylum or lower taxonomical level. This identification rate might be significantly improved in the future, as reference databases are updated. Our results show that marine metabarcoding, currently applied mostly to plankton or sediments, can be adapted to structurally complex hard bottom samples, and emerges as a robust, fast, objective and affordable method for comprehensively characterizing the diversity of marine benthic communities dominated by macroscopic seaweeds and colonial or modular sessile metazoans, allowing for standardized biomonitoring of these ecologically important communities. The new universal primers for COI can potentially be used for biodiversity assessment with high taxonomic resolution in a wide array of marine, terrestrial or freshwater eukaryotic communities.

Percent of MOTUs found for the main categories of organisms in the three fractions studied with 18S (left) and COI (right) in all communities.

New paper published: Genetic diversity and demography of the red starfish

A beautiful specimen of the red starfish Echinaster sepositus.

by Alex Garcia-Cisneros

One of the most wonderful starfish of the Mediterranean suffered a large demographic expansion since the last maximum glacial period. However, its demographic expansion does not prevent it to have a weakness point: a low genetic diversity that might indicate the vulnerability of the species. These and more results were recently published by the ChallenGen team in a manuscript entitled “Low genetic diversity and demographic expansion in the red starfish Echinaster sepositus (Retzius 1816)” at the Scientific Reports.

The authors used both mitochondrial and nuclear markers to resolve the phylogeography and population genetics of the commonly named “red starfish” (Echinaster sepositus). The authors analysed samples from most of the distribution range of the species, with 15 localities distributed between both Mediterranean basins and the Atlantic Ocean.

Besides to demonstrate the low genetic diversity of the species compared with other echinoderms, the species showed a weak genetic structure within marine basins despite the a priori low dispersal potential of its lecithotrophic larva. The lecithotrophic larva in this species does not live more than few days before the settlement and therefore, it makes it difficult to connect distant or isolated populations. Furthermore, we found sharp differences in two Mediterranean localities, Cartagena and Livorno, that are located close to large marine harbours and coastal areas affected by industry, although our experimental design does not allow us to assess the effects of pollutants on the genetic structure.

Bar plots of the Bayesian clustering analysis obtained using STRUCTURE for different K values and based on the combination of both mitochondrial sequences (COI) and microsatellite loci.

Crossing Borders: Temporal genetic differentiation over oceanographic fronts

by Ferran Palero

A new paper from the CHALLENGEN team entitled “Temporal and spatial genetic differentiation in the crab Liocarcinus depurator across the Atlantic-Mediterranean transition” has been recently published in the JCR journal Scientific Reports.

Median joining haplotype network. Each circle represents a haplotype and its size is proportional to its frequency. Location acronyms are DELT (Delta de l’Ebre), VALE (València), ALAC (Alacant), WALB (Málaga) CADI (Cádiz).

Proportion of individuals assigned to the MED haplogroup in each location and sampled year from 1000 pseudoreplicates.

Spatial genetic studies often require sampling broadly separated areas, difficult to access simultaneously. Although comparing localities surveyed at different time periods might result in spurious genetic differentiation, there is a general believe on the stability of genetic structure through time, particularly if sampled localities are isolated or very distant. However, should we really expect stable genetic patterns in marine species? To test this important question, Marta Pascual and collaborators have assessed the time-variation of phylogeography patterns of the portunid crab Liocarcinus depurator across the Atlantic-Mediterranean transition. A partial fragment of the cytochrome oxidase I gene was sequenced in 366 individuals collected during three time periods from localities at both sides of each of the three main oceanographic discontinuities in the area: Gibraltar Strait, Almeria-Oran Front and Ibiza Channel. Although localities showed genetic fluctuations through time, a significant gradient was detected along the coast for all sampling periods. Significant inter-annual differences identified within the Alicante area, north of the Almeria-Oran Front, were associated with shifts in the relative contribution of Atlantic and Mediterranean water masses. The authors conclude that the persistence of a clinal pattern in the Atlantic-Mediterranean transition area despite local fluctuations suggests a complex balance of dispersal and selection.

Despite the Alicante population from 2007 had a larger proportion of Atlantic haplotypes, an overall genetic cline remains stable across the 3-year sampling period.

New paper published: Spatio-temporal patterns of genetic diversity of Styela plicata in harbour populations

by Xavier Turon

We add another paper to the Styela plicata history: “Stochasticity in space, persistence in time: genetic heterogeneity in harbour populations of the introduced ascidian Styela plicata”, by Mari-Carmen Pineda, Beatriz Lorente, Susanna López-Legentil, Creu Palacín and Xavier Turon, which has been published today (23rd June) in PeerJ.

In this study, we analysed genetic structure of the populations of Styela plicata in 9 harbours along the Iberian Mediterranean coast and adjacent Atlantic waters (>1,200 km range) at two time points 5 years apart (2009 and 2014). Using COI sequence data of ca. 400 specimens, we found strong spatial genetic structure, with significant differences among many populations, but no significant differences among years. Our results revealed spatial genetic heterogeneity and temporal homogeneity in S. plicata, suggesting a limited role of recurrent, vessel-mediated transport of organisms among small to medium-size harbours. Our study area is representative of many highly urbanized coasts with dense harbour networks. In these environments, the episodic chance arrival of colonisers appears to determine the genetic structure of populations and the genetic composition of these early colonising individuals persists in the respective harbours, at least over moderate time frames (five years) that encompass ca. 20 generations of S. plicata.

This article is yet another instance of the importance of the temporal component in studies of introduced species to unravel processes occurring during secondary dispersal of non-indigenous species.

Map of the Iberian Peninsula (NW Mediterranean) showing the sampling sites of Styela plicata. Pie charts represent haplotype frequencies for the COI gene in each population analysed in 2009 and 2014.

ChallenGen at the workshop “Application of genomic tools for benthic monitoring”

by Xavier Turon

The 4-5 April 2016, the Natural History Museum of Geneva (Switzerland) hosted the workshop “Application of genomic tools for benthic monitoring of marine environment: from technology to legal and socio-economic aspects”, organised by Prof. Jan Pawlowski of the University of Geneva.

The workshop was intended as a forum for the exchange of ideas about the potential of eDNA- related techniques for assessing biodiversity and the impact of human activities on it. It was attended by environmental professionals, members of regulating bodies, and scientists. The presentations and discussion were very lively, and the overarching idea is that DNA-derived indicators may be effectively used in the field of environmental monitoring (as they are in other applied sciences). However, more development and standardization is necessary before they can be integrated with- or substitute altogether- current techniques using morphology-based indicators, which are slow, time-consuming, and reliant on a worlwide dwindling taxonomic expertise.

Xavier Turon and Owen Wangensteen contributed to the workshop with the talk “Issues in metabarcoding of marine benthos”, where they pinpointed some conflicting issues, such as the use of DNA vs RNA, the choice between 18S rDNA or COI markers, or the use of a fixed vs a variable threshold for clustering sequences into MOTUs. The talk was illustrated with results obtained in the projects CHALLENGEN and METABARPARK.

Xavier Turon and Tom Wilding, of the Scottish Association for Marine Science, at the entrance of the Natural History Museum, with a lovely pet!

Temporal genetic structure in the introduced ascidian Styela plicata

by M.Carmen Pineda

It’s been almost three years since the last paper on Styela plicata was published by our group, but the saga is not over yet!! The article entitled “Stable populations in unstable habitats: temporal genetic structure of the introduced ascidian Styela plicata in North Carolina”, by M.Carmen Pineda, Xavier Turon, Rocio Perez-Portela and Susanna Lopez-Legentil, has been just published online in the journal Marine Biology.

The solitary ascidian Styela plicata.

Styela plicata is a solitary ascidian, which has been introduced by ship fouling to harbours and marinas around the globe. This species has been extensively studied during the last years, based on its potential to become a plague. However, this is the first time that its genetic composition is studied on a temporal scale. On this recent study, a combination of nuclear and mitochondrial markers (7 microsatellites and a fragment of the gene Cytochrome Oxidase subunit I, COI) were used to analyse the genetic composition of one population subjected to environmental fluctuations and periodical die-offs, every 2 months and over 2.5 years. Overall, our results indicate that the investigated population is stable over time and relies on a periodic arrival of larvae from other populations, maintaining high genetic diversity and a complex interplay of allele gains and losses.

Temporal analyses exploring genetic trends over time are a great tool to predict the likelihood of long time survival of an introduced population in a new habitat and its invasive potential. This kind of information is particularly relevant when deciding which introduced species are more detrimental, and should help resource managers to focus their control and eradication efforts.