Thermo-priming triggers species-specific physiological and transcriptome responses in Mediterranean seagrasses.

Heat-priming improves plants' tolerance to a recurring heat stress event. The underlying molecular mechanisms of heat-priming are largely unknown in seagrasses. Here, ad hoc mesocosm experiments were conducted with two Mediterranean seagrass species, Posidonia oceanica and Cymodocea nodosa. Plants were first exposed to heat-priming, followed by a heat-triggering event. A comprehensive assessment of plant stress response across different levels of biological organization was performed at the end of the triggering event. Morphological and physiological results showed an improved response of heat-primed P. oceanica plants while in C. nodosa both heat- and non-primed plants enhanced their growth rates at the end of the triggering event. As resulting from whole transcriptome sequencing, molecular functions related to several cellular compartments and processes were involved in the response to warming of non-primed plants, while the response of heat-primed plants involved a limited group of processes. Our results suggest that seagrasses acquire a primed state during the priming event, that eventually gives plants the ability to induce a more energy-effective response when the thermal stress event recurs. Different species may differ in their ability to perform an improved heat stress response after priming. This study provides pioneer molecular insights into the emerging topic of seagrass stress priming and may benefit future studies in the field.

Seagrass genomes reveal ancient polyploidy and adaptations to the marine environment.

We present chromosome-level genome assemblies from representative species of three independently evolved seagrass lineages: Posidonia oceanica, Cymodocea nodosa, Thalassia testudinum and Zostera marina. We also include a draft genome of Potamogeton acutifolius, belonging to a freshwater sister lineage to Zosteraceae. All seagrass species share an ancient whole-genome triplication, while additional whole-genome duplications were uncovered for C. nodosa, Z. marina and P. acutifolius. Comparative analysis of selected gene families suggests that the transition from submerged-freshwater to submerged-marine environments mainly involved fine-tuning of multiple processes (such as osmoregulation, salinity, light capture, carbon acquisition and temperature) that all had to happen in parallel, probably explaining why adaptation to a marine lifestyle has been exceedingly rare. Major gene losses related to stomata, volatiles, defence and lignification are probably a consequence of the return to the sea rather than the cause of it. These new genomes will accelerate functional studies and solutions, as continuing losses of the 'savannahs of the sea' are of major concern in times of climate change and loss of biodiversity.

Signs of local adaptation by genetic selection and isolation promoted by extreme temperature and salinity in the Mediterranean seagrass Posidonia oceanica.

Adaptation to local conditions is known to occur in seagrasses; however, knowledge of the genetic basis underlying this phenomenon remains scarce. Here, we analysed Posidonia oceanica from six sites within and around the Stagnone di Marsala, a semi-enclosed coastal lagoon where salinity and temperature exceed the generally described tolerance thresholds of the species. Sea surface temperatures (SSTs) were measured and plant samples were collected for the assessment of morphology, flowering rate and for screening genome-wide polymorphisms using double digest restriction-site-associated DNA sequencing. Results demonstrated more extreme SSTs and salinity levels inside the lagoon than the outer lagoon regions. Morphological results showed significantly fewer and shorter leaves and reduced rhizome growth of P. oceanica from the inner lagoon and past flowering events were recorded only for a meadow farthest away from the lagoon. Using an array of 51,329 single nucleotide polymorphisms, we revealed a clear genetic structure among the study sites and confirmed the genetic isolation and high clonality of the innermost site. In all, 14 outlier loci were identified and annotated with several proteins including those relate to plant stress response, protein transport and regulators of plant-specific developmental events. Especially, five outlier loci showed maximum allele frequency at the innermost site, likely reflecting adaptation to the extreme temperature and salinity regimes, possibly due to the selection of more resistant genotypes and the progressive restriction of gene flow. Overall, this study helps us to disentangle the genetic basis of seagrass adaptation to local environmental conditions and may support future works on assisted evolution in seagrasses.

Dim artificial light at night alters gene expression rhythms and growth in a key seagrass species (Posidonia oceanica).

Artificial light at night (ALAN) is a globally spreading anthropogenic stressor, affecting more than 20% of coastal habitats. The alteration of the natural light/darkness cycle is expected to impact the physiology of organisms by acting on the complex circuits termed as circadian rhythms. Our understanding of the impact of ALAN on marine organisms is lagging behind that of terrestrial ones, and effects on marine primary producers are almost unexplored. Here, we investigated the molecular and physiological response of the Mediterranean seagrass, Posidonia oceanica (L.) Delile, as model to evaluate the effect of ALAN on seagrass populations established in shallow waters, by taking advantage of a decreasing gradient of dim nocturnal light intensity (from < 0.01 to 4 lx) along the NW Mediterranean coastline. We first monitored the fluctuations of putative circadian-clock genes over a period of 24 h along the ALAN gradient. We then investigated whether key physiological processes, known to be synchronized with day length by the circadian rhythm, were also affected by ALAN. ALAN influenced the light signalling at dusk/night in P. oceanica, including that of shorter blue wavelengths, through the ELF3-LUX1-ZTL regulatory network, and suggested that the daily perturbation of internal clock orthologs in seagrass might have caused the recruitment of PoSEND33 and PoPSBS genes to mitigate the repercussions of a nocturnal stress on photosynthesis during the day. A long-lasting impairment of gene fluctuations in sites characterised by ALAN could explain the reduced growth of the seagrass leaves when these were transferred into controlled conditions and without lighting during the night. Our results highlight the potential contribution of ALAN to the global loss of seagrass meadows, posing questions about key interactions with a variety of other human-related stressors in urban areas, in order to develop more efficient strategies to globally preserve these coastal foundation species.

Chloroplast genomic comparison provides insights into the evolution of seagrasses.

BACKGROUND: Seagrasses are a polyphyletic group of monocotyledonous angiosperms that have evolved to live entirely submerged in marine waters. Thus, these species are ideal for studying plant adaptation to marine environments. Herein, we sequenced the chloroplast (cp) genomes of two seagrass species (Zostera muelleri and Halophila ovalis) and performed a comparative analysis of them with 10 previously published seagrasses, resulting in various novel findings. RESULTS: The cp genomes of the seagrasses ranged in size from 143,877 bp (Zostera marina) to 178,261 bp (Thalassia hemprichii), and also varied in size among different families in the following order: Hydrocharitaceae > Cymodoceaceae > Ruppiaceae > Zosteraceae. The length differences between families were mainly related to the expansion and contraction of the IR region. In addition, we screened out 2,751 simple sequence repeats and 1,757 long repeat sequence types in the cp genome sequences of the 12 seagrass species, ultimately finding seven hot spots in coding regions. Interestingly, we found nine genes with positive selection sites, including two ATP subunit genes (atpA and atpF), three ribosome subunit genes (rps4, rps7, and rpl20), one photosystem subunit gene (psbH), and the ycf2, accD, and rbcL genes. These gene regions may have played critical roles in the adaptation of seagrasses to diverse environments. In addition, phylogenetic analysis strongly supported the division of the 12 seagrass species into four previously recognized major clades. Finally, the divergence time of the seagrasses inferred from the cp genome sequences was generally consistent with previous studies. CONCLUSIONS: In this study, we compared chloroplast genomes from 12 seagrass species, covering the main phylogenetic clades. Our findings will provide valuable genetic data for research into the taxonomy, phylogeny, and species evolution of seagrasses.

DNA methylation dynamics in a coastal foundation seagrass species under abiotic stressors.

DNA methylation (DNAm) has been intensively studied in terrestrial plants in response to environmental changes, but its dynamic changes in a temporal scale remain unexplored in marine plants. The seagrass Posidonia oceanica ranks among the slowest-growing and longest-living plants on Earth, and is particularly vulnerable to sea warming and local anthropogenic pressures. Here, we analysed the dynamics of DNAm changes in plants collected from coastal areas differentially impacted by eutrophication (i.e. oligotrophic, Ol; eutrophic, Eu) and exposed to abiotic stressors (nutrients, temperature increase and their combination). Levels of global DNAm (% 5-mC) and the expression of key genes involved in DNAm were assessed after one, two and five weeks of exposure. Results revealed a clear differentiation between plants, depending on environmental stimuli, time of exposure and plants' origin. % 5-mC levels were higher during the initial stress exposure especially in Ol plants, which upregulated almost all genes involved in DNAm. Contrarily, Eu plants showed lower expression levels, which increased under chronic exposure to stressors, particularly to temperature. These findings show that DNAm is dynamic in P. oceanica during stress exposure and underlined that environmental epigenetic variations could be implicated in the regulation of acclimation and phenotypic differences depending on local conditions.

Phylogenomic Analyses of Alismatales Shed Light into Adaptations to Aquatic Environments.

Land plants first evolved from freshwater algae, and flowering plants returned to water as early as the Cretaceous and multiple times subsequently. Alismatales is the largest clade of aquatic angiosperms including all marine angiosperms, as well as terrestrial plants. We used Alismatales to explore plant adaptations to aquatic environments by analyzing a data set that included 95 samples (89 Alismatales species) covering four genomes and 91 transcriptomes (59 generated in this study). To provide a basis for investigating adaptations, we assessed phylogenetic conflict and whole-genome duplication (WGD) events in Alismatales. We recovered a relationship for the three main clades in Alismatales as (Tofieldiaceae, Araceae) + core Alismatids. We also found phylogenetic conflict among the three main clades that was best explained by incomplete lineage sorting and introgression. Overall, we identified 18 putative WGD events across Alismatales. One of them occurred at the most recent common ancestor of core Alismatids, and three occurred at seagrass lineages. We also found that lineage and life-form were both important for different evolutionary patterns for the genes related to freshwater and marine adaptation. For example, several light- or ethylene-related genes were lost in the seagrass Zosteraceae, but are present in other seagrasses and freshwater species. Stomata-related genes were lost in both submersed freshwater species and seagrasses. Nicotianamine synthase genes, which are important in iron intake, expanded in both submersed freshwater species and seagrasses. Our results advance the understanding of the adaptation to aquatic environments and WGDs using phylogenomics.

Multidrug-resistant epi-endophytic bacterial community in Posidonia oceanica of Mahdia coast as biomonitoring factor for antibiotic contamination.

Faced with the significant disturbances, mainly of anthropogenic origin, which affect the Mediterranean coastal ecosystem, Posidonia oceanica (L.) Delile has often been used to assess the state of health of this environment. The present study aims to determine the multidrug resistance patterns among isolated and identified epi-endophytic bacterial strains in P. oceanica seagrass collected from Mahdia coastal seawater (Tunisia). To investigate the bacterial community structure and diversity from coastal seawater samples from Mahdia, total DNA extraction and 16S rRNA gene amplification were performed and analyzed by denaturing gradient gel electrophoresis (DGGE). The DGGE profiles showed that some bands were specific to a given site, while other bands were found to be common to more than one sample. In the other hand, bacterial strains were isolated from 1 mL of leaves and epiphytes suspension of P. oceanica seagrass in marine agar. Forty-three isolates were obtained, seven of them were selected and identified on the basis of 16S rRNA gene sequence analysis. These isolates belonged to the genus Bacillus, exhibiting 98-100% of identity with known sequences. Susceptibility patterns of these strains were studied toward commonly used antibiotics in Tunisia. All identified isolates were resistant to Aztreonam (72.1%), Ceftazidime (60.5%), Amoxicillin (56%) and Rifampicin (51.2%). S5-L13 strain had presented the highest multidrug resistance with a MAR index of 0.67.

Responses of the Mediterranean seagrass Cymodocea nodosa to combined temperature and salinity stress at the ionomic, transcriptomic, ultrastructural and photosynthetic levels.

The Little Neptune grass Cymodocea nodosa is a key seagrass species in the Mediterranean Sea, forming extensive and patchy meadows in shallow coastal and transitional ecosystems. In such habitats, high temperatures and salinities, separately and in combination, can be significant stressors in the context of climate change, particularly during heatwave events, and seawater desalination plant effluents. Despite well-documented negative, macroscopic effects, the underlying cellular and molecular processes of the combined effects of increasing temperature and salinities have remained largely elusive in C. nodosa - which are addressed by the present study. High salinity and high temperature, alone and in combination, affected ion equilibrium in the plant cells. Non-synonymous mutations marked the transcriptomic response to salinity and temperature stress at loci related to osmotic stress. Cell structure, especially the nucleus, chloroplasts, mitochondria and organization of the MT cytoskeleton, was also altered. Both temperature and salinity stress negatively affected photosynthetic activity as evidenced by ΔF/Fm', following an antagonistic interaction type. Overall, this study showed that all biological levels investigated were strongly affected by temperature and salinity stress, however, with the latter having more severe effects. The results have implications for the operation of desalination plants and for assessing the impacts of marine heat waves.

Profiling the cell walls of seagrasses from A (Amphibolis) to Z (Zostera).

BACKGROUND: The polyphyletic group of seagrasses shows an evolutionary history from early monocotyledonous land plants to the marine environment. Seagrasses form important coastal ecosystems worldwide and large amounts of seagrass detritus washed on beaches might also be valuable bioeconomical resources. Despite this importance and potential, little is known about adaptation of these angiosperms to the marine environment and their cell walls. RESULTS: We investigated polysaccharide composition of nine seagrass species from the Mediterranean, Red Sea and eastern Indian Ocean. Sequential extraction revealed a similar seagrass cell wall polysaccharide composition to terrestrial angiosperms: arabinogalactans, pectins and different hemicelluloses, especially xylans and/or xyloglucans. However, the pectic fractions were characterized by the monosaccharide apiose, suggesting unusual apiogalacturonans are a common feature of seagrass cell walls. Detailed analyses of four representative species identified differences between organs and species in their constituent monosaccharide composition and lignin content and structure. Rhizomes were richer in glucosyl units compared to leaves and roots. Enhalus had high apiosyl and arabinosyl abundance, while two Australian species of Amphibolis and Posidonia, were characterized by high amounts of xylosyl residues. Interestingly, the latter two species contained appreciable amounts of lignin, especially in roots and rhizomes whereas Zostera and Enhalus were lignin-free. Lignin structure in Amphibolis was characterized by a higher syringyl content compared to that of Posidonia. CONCLUSIONS: Our investigations give a first comprehensive overview on cell wall composition across seagrass families, which will help understanding adaptation to a marine environment in the evolutionary context and evaluating the potential of seagrass in biorefinery incentives.

The Enzymatic and Non-Enzymatic Antioxidant System Response of the Seagrass Cymodocea nodosa to Bisphenol-A Toxicity.

The effects of environmentally relevant bisphenol A (BPA) concentrations (0.3, 1 and 3 µg L-1) were tested at 2, 4, 6 and 8 days, on intermediate leaves, of the seagrass Cymodocea nodosa. Hydrogen peroxide (H2O2) production, lipid peroxidation, protein, phenolic content and antioxidant enzyme activities were investigated. Increased H2O2 formation was detected even at the lowest BPA treatments from the beginning of the experiment and both the enzymatic and non-enzymatic antioxidant defense mechanisms were activated upon application of BPA. Elevated H2O2 levels that were detected as a response to increasing BPA concentrations and incubation time, led to the decrease of protein content on the 4th day even at the two lower BPA concentrations, and to the increase of the lipid peroxidation at the highest concentration. However, on the 6th day of BPA exposure, protein content did not differ from the control, indicating the ability of both the enzymatic and non-enzymatic mechanisms (such as superoxide dismutase (SOD) and phenolics) to counteract the BPA-derived oxidative stress. The early response of the protein content determined that the Low Effect Concentration (LOEC) of BPA is 0.3 µg L-1 and that the protein content meets the requirements to be considered as a possible early warning "biomarker" for C. nodosa against BPA toxicity.

RNA-Seq analysis reveals potential regulators of programmed cell death and leaf remodelling in lace plant (Aponogeton madagascariensis).

BACKGROUND: The lace plant (Aponogeton madagascariensis) is an aquatic monocot that develops leaves with uniquely formed perforations through the use of a developmentally regulated process called programmed cell death (PCD). The process of perforation formation in lace plant leaves is subdivided into several developmental stages: pre-perforation, window, perforation formation, perforation expansion and mature. The first three emerging "imperforate leaves" do not form perforations, while all subsequent leaves form perforations via developmentally regulated PCD. PCD is active in cells called "PCD cells" that do not retain the antioxidant anthocyanin in spaces called areoles framed by the leaf veins of window stage leaves. Cells near the veins called "NPCD cells" retain a red pigmentation from anthocyanin and do not undergo PCD. While the cellular changes that occur during PCD are well studied, the gene expression patterns underlying these changes and driving PCD during leaf morphogenesis are mostly unknown. We sought to characterize differentially expressed genes (DEGs) that mediate lace plant leaf remodelling and PCD. This was achieved performing gene expression analysis using transcriptomics and comparing DEGs among different stages of leaf development, and between NPCD and PCD cells isolated by laser capture microdissection. RESULTS: Transcriptomes were sequenced from imperforate, pre-perforation, window, and mature leaf stages, as well as PCD and NPCD cells isolated from window stage leaves. Differential expression analysis of the data revealed distinct gene expression profiles: pre-perforation and window stage leaves were characterized by higher expression of genes involved in anthocyanin biosynthesis, plant proteases, expansins, and autophagy-related genes. Mature and imperforate leaves upregulated genes associated with chlorophyll development, photosynthesis, and negative regulators of PCD. PCD cells were found to have a higher expression of genes involved with ethylene biosynthesis, brassinosteroid biosynthesis, and hydrolase activity whereas NPCD cells possessed higher expression of auxin transport, auxin signalling, aspartyl proteases, cysteine protease, Bag5, and anthocyanin biosynthesis enzymes. CONCLUSIONS: RNA sequencing was used to generate a de novo transcriptome for A. madagascariensis leaves and revealed numerous DEGs potentially involved in PCD and leaf remodelling. The data generated from this investigation will be useful for future experiments on lace plant leaf development and PCD in planta.

Gene body DNA methylation in seagrasses: inter- and intraspecific differences and interaction with transcriptome plasticity under heat stress.

The role of DNA methylation and its interaction with gene expression and transcriptome plasticity is poorly understood, and current insight comes mainly from studies in very few model plant species. Here, we study gene body DNA methylation (gbM) and gene expression patterns in ecotypes from contrasting thermal environments of two marine plants with contrasting life history strategies in order to explore the potential role epigenetic mechanisms could play in gene plasticity and responsiveness to heat stress. In silico transcriptome analysis of CpGO/E ratios suggested that the bulk of Posidonia oceanica and Cymodocea nodosa genes possess high levels of intragenic methylation. We also observed a correlation between gbM and gene expression flexibility: genes with low DNA methylation tend to show flexible gene expression and plasticity under changing conditions. Furthermore, the empirical determination of global DNA methylation (5-mC) showed patterns of intra and inter-specific divergence that suggests a link between methylation level and the plants' latitude of origin and life history. Although we cannot discern whether gbM regulates gene expression or vice versa, or if other molecular mechanisms play a role in facilitating transcriptome responsiveness, our findings point to the existence of a relationship between gene responsiveness and gbM patterns in marine plants.

Leaf proteome modulation and cytological features of seagrass Cymodocea nodosa in response to long-term high CO2 exposure in volcanic vents.

Seagrass Cymodocea nodosa was sampled off the Vulcano island, in the vicinity of a submarine volcanic vent. Leaf samples were collected from plants growing in a naturally acidified site, influenced by the long-term exposure to high CO2 emissions, and compared with others collected in a nearby meadow living at normal pCO2 conditions. The differential accumulated proteins in leaves growing in the two contrasting pCO2 environments was investigated. Acidified leaf tissues had less total protein content and the semi-quantitative proteomic comparison revealed a strong general depletion of proteins belonging to the carbon metabolism and protein metabolism. A very large accumulation of proteins related to the cell respiration and to light harvesting process was found in acidified leaves in comparison with those growing in the normal pCO2 site. The metabolic pathways linked to cytoskeleton turnover also seemed affected by the acidified condition, since a strong reduction in the concentration of cytoskeleton structural proteins was found in comparison with the normal pCO2 leaves. Results coming from the comparative proteomics were validated by the histological and cytological measurements, suggesting that the long lasting exposure and acclimation of C. nodosa to the vents involved phenotypic adjustments that can offer physiological and structural tools to survive the suboptimal conditions at the vents vicinity.

New insights into the population genetics of partially clonal organisms: When seagrass data meet theoretical expectations.

Seagrass meadows are among the most important coastal ecosystems in terms of both spatial extent and ecosystem services, but they are also declining worldwide. Understanding the drivers of seagrass meadow dynamics is essential for designing sound management, conservation and restoration strategies. However, poor knowledge of the effect of clonality on the population genetics of natural populations severely limits our understanding of the dynamics and connectivity of meadows. Recent modelling approaches have described the expected distributions of genotypic and genetic descriptors under increasing clonal rates, which may help us better understand and interpret population genetics data obtained for partial asexuals. Here, in the light of these recent theoretical developments, we revisited population genetics data for 165 meadows of four seagrass species. Contrasting shoot lifespan and rhizome turnover led to the prediction that the influence of asexual reproduction would increase along a gradient from Zostera noltii to Zostera marina, Cymodocea nodosa and Posidonia oceanica, with increasing departure from Hardy-Weinberg equilibrium (Fis ), mostly towards heterozygote excess, and decreasing genotypic richness (R). This meta-analysis provides a nested validation of this hypothesis at both the species and meadow scales through a significant relationship between Fis and R within each species. By empirically demonstrating the theoretical expectations derived from recent modelling approaches, this work calls for the use of Hardy-Weinberg equilibrium (Fis ) rather than only the strongly sampling-sensitive R to assess the importance of clonal reproduction (c), at least when the impact of selfing on Fis can be neglected. The results also emphasize the need to revise our appraisal of the extent of clonality and its influence on the dynamics, connectivity and evolutionary trajectory of partial asexuals in general, including in seagrass meadows, to develop the most accurate management strategies.

Convergent Loss of an EDS1/PAD4 Signaling Pathway in Several Plant Lineages Reveals Coevolved Components of Plant Immunity and Drought Response.

Plant innate immunity relies on nucleotide binding leucine-rich repeat receptors (NLRs) that recognize pathogen-derived molecules and activate downstream signaling pathways. We analyzed the variation in NLR gene copy number and identified plants with a low number of NLR genes relative to sister species. We specifically focused on four plants from two distinct lineages, one monocot lineage (Alismatales) and one eudicot lineage (Lentibulariaceae). In these lineages, the loss of NLR genes coincides with loss of the well-known downstream immune signaling complex ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1)/PHYTOALEXIN DEFICIENT 4 (PAD4). We expanded our analysis across whole proteomes and found that other characterized immune genes were absent only in Lentibulariaceae and Alismatales. Additionally, we identified genes of unknown function that were convergently lost together with EDS1/PAD4 in five plant species. Gene expression analyses in Arabidopsis (Arabidopsis thaliana) and Oryza sativa revealed that several homologs of the candidates are differentially expressed during pathogen infection, drought, and abscisic acid treatment. Our analysis provides evolutionary evidence for the rewiring of plant immunity in some plant lineages, as well as the coevolution of the EDS1/PAD4 pathway and drought responses.

Adaptive responses along a depth and a latitudinal gradient in the endemic seagrass Posidonia oceanica.

Seagrass meadows provide important ecosystem services and are critical for the survival of the associated invertebrate community. However, they are threatened worldwide by human-driven environmental change. Understanding the seagrasses' potential for adaptation is critical to assess not only their ability to persist under future global change scenarios, but also to assess the persistence of the associated communities. Here we screened a wild population of Posidonia oceanica, an endemic long-lived seagrass in the Mediterranean Sea, for genes that may be target of environmental selection, using an outlier and a genome-wide transcriptome analysis. We identified loci where polymorphism or differential expression was associated with either a latitudinal or a bathymetric gradient, as well as with both gradients in an effort to identify loci associated with temperature and light. We found the candidate genes underlying growth and immunity to be divergent between populations adapted to different latitudes and/or depths, providing evidence for local adaptation. Furthermore, we found evidence of reduced gene flow among populations including adjacent populations. Reduced gene flow, combined with low sexual recombination, small effective population size, and long generation time of P. oceanica raises concerns for the long-term persistence of this species, especially in the face of rapid environmental change driven by human activities.

Seeds in motion: Genetic assignment and hydrodynamic models demonstrate concordant patterns of seagrass dispersal.

Movement is fundamental to the ecology and evolutionary dynamics within species. Understanding movement through seed dispersal in the marine environment can be difficult due to the high spatial and temporal variability of ocean currents. We employed a mutually enriching approach of population genetic assignment procedures and dispersal predictions from a hydrodynamic model to overcome this difficulty and quantify the movement of dispersing floating fruit of the temperate seagrass Posidonia australis Hook.f. across coastal waters in south-western Australia. Dispersing fruit cohorts were collected from the water surface over two consecutive years, and seeds were genotyped using microsatellite DNA markers. Likelihood-based genetic assignment tests were used to infer the meadow of origin for seed cohorts and individuals. A three-dimensional hydrodynamic model was coupled with a particle transport model to simulate the movement of fruit at the water surface. Floating fruit cohorts were mainly assigned genetically to the nearest meadow, but significant genetic differentiation between cohort and most likely meadow of origin suggested a mixed origin. This was confirmed by genetic assignment of individual seeds from the same cohort to multiple meadows. The hydrodynamic model predicted 60% of fruit dispersed within 20 km, but that fruit was physically capable of dispersing beyond the study region. Concordance between these two independent measures of dispersal provides insight into the role of physical transport for long distance dispersal of fruit and the consequences for spatial genetic structuring of seagrass meadows.

Frond transformation system mediated by Agrobacterium tumefaciens for Lemna minor.

The Lemnaceae, known as duckweed, the smallest flowering aquatic plant, shows promise as a plant bioreactor. For applying this potential plant bioreactor, establishing a stable and efficient genetic transformation system is necessary. The currently favored callus-based method for duckweed transformation is time consuming and genotype limited, as it requires callus culture and regeneration, which is inapplicable to many elite duckweed strains suitable for bioreactor exploitation. In this study, we attempted to establish a simple frond transformation system mediated by Agrobacterium tumefaciens for Lemna minor, one of the most widespread duckweed species in the world. To evaluate the feasibility of the new transformation system, the gene CYP710A11 was overexpressed to improve the yield of stigmasterol, which has multiple medicinal purposes. Three L. minor strains, ZH0055, D0158 and M0165, were transformed by both a conventional callus transformation system (CTS) and the simple frond transformation system (FTS). GUS staining, PCR, quantitative PCR and stigmasterol content detection showed that FTS can produce stable transgenic lines as well as CTS. Moreover, compared to CTS, FTS can avoid the genotype constraints of callus induction, thus saving at least half of the required processing time (CTS took 8-9 months while FTS took approximately 3 months in this study). Therefore, this transformation system is feasible in producing stable transgenic lines for a wide range of L. minor genotypes.

Dramatic loss of seagrass habitat under projected climate change in the Mediterranean Sea.

Although climate warming is affecting most marine ecosystems, the Mediterranean is showing earlier impacts. Foundation seagrasses are already experiencing a well-documented regression in the Mediterranean which could be aggravated by climate change. Here, we forecast distributions of two seagrasses and contrast predicted loss with discrete regions identified on the basis of extant genetic diversity. Under the worst-case scenario, Posidonia oceanica might lose 75% of suitable habitat by 2050 and is at risk of functional extinction by 2100, whereas Cymodocea nodosa would lose only 46.5% in that scenario as losses are compensated with gained and stable areas in the Atlantic. Besides, we predict that erosion of present genetic diversity and vicariant processes can happen, as all Mediterranean genetic regions could decrease considerably in extension in future warming scenarios. The functional extinction of Posidonia oceanica would have important ecological impacts and may also lead to the release of the massive carbon stocks these ecosystems stored over millennia.