Limonium Species Research Articles

Physiological and morphological characterisation of Limonium species in their natural habitats: Insights into their abiotic stress responses

Morphological and biochemical traits of four halophytes of the genus Limonium were analysed in plants sampled from salt marshes in SE Spain. This work aimed to explore the mechanism(s) behind the adaptation of these species to stressful habitats, with particular emphasis on responses to drought. Plants of each species together with soil samples were collected in summer, which is the most stressful season in the Mediterranean. Soil parameters and plant morphological traits were determined, and the levels of several biochemical stress markers in plants were measured using spectrophotometric assays. A multivariate analysis was performed to correlate soil and plant data. Morphological characteristics regarding the underground system topology and several biochemical traits (higher foliar Ca2+, sucrose and glucose, and lower proline, glycine-betaine and fructose) clearly separate L. santapolense individuals from plants of the other three species. Drought tolerance of L. santapolense in the field is mostly dependent on morphological adaptations: when growing in an arid location, plants of this species develop long taproots that can extract water from the deep, moist layers of the soil.

Insights on Salt Tolerance of Two Endemic Limonium Species from Spain.

We have analysed the salt tolerance of two endemic halophytes of the genus Limonium, with high conservation value. In the present study, seed germination and growth parameters as well as different biomarkers—photosynthetic pigments, mono and divalent ion contents—associated to salt stress were evaluated in response to high levels of NaCl. The study was completed with an untargeted metabolomics analysis of the primary compounds including carbohydrates, phosphoric and organic acids, and amino acids, identified by using a gas chromatography and mass spectrometry platform. Limonium albuferae proved to be more salt-tolerant than L. doufourii, both at the germination stage and during vegetative growth. The degradation of photosynthetic pigments and the increase of Na+/K+ ratio under salt stress were more accentuated in the less tolerant second species. The metabolomics analysis unravelled several differences between the two species. The higher salt tolerance of L. albuferae may rely on its specific accumulation of fructose and glucose under high salinity conditions, the first considered as a major osmolyte in this genus. In addition, L. albuferae showed steady levels of citric and malic acids, whereas the glutamate family pathway was strongly activated under stress in both species, leading to the accumulation of proline (Pro) and γ-aminobutyric acid (GABA).

Qualitative and Quantitative Differences in Osmolytes Accumulation and Antioxidant Activities in Response to Water Deficit in Four Mediterranean Limonium Species.

Limonium is a genus represented in the Iberian Peninsula by numerous halophytic species that are affected in nature by salinity, and often by prolonged drought episodes. Responses to water deficit have been studied in four Mediterranean Limonium species, previously investigated regarding salt tolerance mechanisms. The levels of biochemical markers, associated with specific responses—photosynthetic pigments, mono- and divalent ions, osmolytes, antioxidant compounds and enzymes—were determined in the control and water-stressed plants, and correlated with their relative degree of stress-induced growth inhibition. All the tested Limonium taxa are relatively resistant to drought on the basis of both the constitutive presence of high leaf ion levels that contribute to osmotic adjustment, and the stress-induced accumulation of osmolytes and increased activity of antioxidant enzymes, albeit with different qualitative and quantitative induction patterns. Limonium santapolense activated the strongest responses and clearly differed from Limonium virgatum, Limonium girardianum, and Limonium narbonense, as indicated by cluster and principal component analysis (PCA) analyses in agreement with its drier natural habitat, and compared to that of the other plants. Somewhat surprisingly, however, L. santapolense was the species most affected by water deficit in growth inhibition terms, which suggests the existence of additional mechanisms of defense operating in the field that cannot be mimicked in greenhouses.

Physiological and biochemical responses to salinity in two endemic Limonium species from Spain

Physiological and biochemical responses to salinity in two endemic Limonium species from Spain

Stomatal anatomy coordinates leaf size with Rubisco kinetics in the Balearic Limonium

Abstract Trait integration arises through both selection on functional coordination and shared developmental pathways. Different anatomical components must both work well and develop together to generate individuals with the appropriate physiology to survive and reproduce in their environment. In this study, we used a common garden experiment and Bayesian multilevel models to test whether stomatal anatomy coordinates leaf gas exchange, Rubisco kinetics, and leaf size across 10 closely related species of Limonium from the Balearic Islands. The results indicate that the anatomical determinants of maximum stomatal conductance, stomatal density and size, were functionally coordinated with Rubisco kinetics – species whose stomatal anatomy was correlated with low stomatal conductance have evolved Rubisco enzymes better adapted to low operational chloroplastic CO2 concentrations. Lower stomatal density was associated with greater leaf size, which can be explained by a greater proportion of pavement cells in large-leaved species. These results suggest that both selection for functional coordination (stomata and Rubisco kinetics) and shared development pathways (stomatal density and leaf area) likely shape patterns of trait integration between species.

Phytochemical and Biological Activities in Limonium Species Collected in Different Biotopes of Tunisia.

A particular interest is nowadays given to natural antioxidants occurring in foods which can reduce the risk of several diseases through their protective effect. The genus Limonium is widely distributed in different salt regions of Tunisia and known in traditional medicine for the presence of highly effective viral and bacterial replication inhibitors. Limonium leaves have possible beneficial effects on human health for their antioxidant activities and free radical scavenging abilities. To exploit the potential of plants from extreme environments as new sources of natural antioxidants, we studied the extracts from leaves of eight Limonium species growing in extreme environments in Tunisia. Antioxidant molecules (polyphenols, flavonoids, flavonols, ascorbate, tocopherols), in vitro (DPPH, ORAC) and ex vivo antioxidant potential on human erythrocytes, antioxidant enzymes activities (superoxide dismutase, peroxidases, glutathione reductase) were evaluated to identify the species with the best antioxidant capacity. The results showed variability among the species considered in function of the environmental conditions of their natural biotopes, as for the antioxidants measured. In particular, L. vulgare from Oued Rane biotope, characterized by dryness and high temperatures, was the species with the highest enzymatic activity and antioxidant capacity, making it interesting as possible edible halophyte plant or as food complement.

Leaf size as a key determinant of contrasting growth patterns in closely related Limonium (Plumbaginaceae) species

This study aims to analyze the importance of leaf size on plant growth capacity among an array of closely related Limonium species, and its impact on the underlying determinants of growth reduction under extreme water deficit conditions. To do so, thirteen Balearic Limonium species with contrasting leaf size were grown under long-term well-watered (WW) and severe water-deficit (WD) conditions in a common garden experiment. Fundamental growth traits were measured, including relative growth rate (RGR), net assimilation rate (NAR), leaf area ratio (LAR), leaf mass area (LMA) and leaf mass ratio (LMR). WD promoted small changes in leaf size, and species with larger leaves had higher RGR than species with smaller leaves, irrespective of the water treatment. Most RGR variation across species and treatments was explained by NAR, with comparatively much lower importance of LAR. The factorization of LAR underlying components denoted the importance of LMA in explaining RGR, whereas the impact of LMR on RGR was negligible in Limonium. Further, species with larger leaves had higher water consumption but also higher water use efficiency, especially under WD. Therefore, contrary to general trends in species from dry environments, increased leaf size is linked to increased growth capacity and also increased water use efficiency across closely related Limonium species.

Limonium dagmarae (Plumbaginaceae), a new species from Namaqualand coast, South Africa

A new species of Limonium from Namaqualand coast, South Africa is here described and named L. dagmarae. Shared morphological characters with related South African species place the new species within L. sect. Circinaria. Evidence is presented to segregate the new species from relatives based on discrete morphological and environmental characters. The holotype of the L. dagmarae is preserved at NBG.

Pollen morphology of the coastal species of Turkish Limonium (Plumbaginaceae)

The pollen morphology of nine Limonium species (Plumbaginaceae), occurring along the coastal saline habitats of Turkey, are investigated. Pollen slides were prepared according to the methods of Wodehouse (W) and Erdtman (E) and examined using light microscope. Then pollen grains were placed directly to stubs and covered by gold-palladium mixture for scanning electron analyses. Pollen grains are usually tricolpate, rarely syncolpate, dicolpate, tetracolpate, and pentacolpate. Shape of pollen grains are varing from prolate-sphaeroidal to oblate. Medium to large sized pollen grains of Limonium has long and wide colpi with distinct margins, and acute terminals. Exine structure is semi-tectate and exine ornamentation is microreticulate-microechinate or reticulate-microechinate. Pollen grains of Limonium species have shown variation on colpus type, shape and exine ornamentation. Examined Limonium species can be seperated in three unformal groups according to the palynological features, and their pollen definitions have been done. Besides, a taxonomic key was proposed for quick and accurate identification based on different palyno-morphological characters.

An expanded molecular phylogeny of Plumbaginaceae, with emphasis on Limonium (sea lavenders): Taxonomic implications and biogeographic considerations.

Plumbaginaceae is characterized by a history of multiple taxonomic rearrangements and lacks a broad molecular phylogenetic framework. Limonium is the most species‐rich genus of the family with ca. 600 species and cosmopolitan distribution. Its center of diversity is the Mediterranean region, where ca. 70% of all Limonium species are endemic. In this study, we sample 201 Limonium species covering all described infrageneric entities and spanning its wide geographic range, along with 64 species of other Plumbaginaceae genera, representing 23 out of 29 genera of the family. Additionally, 20 species of the sister family Polygonaceae were used as outgroup. Sequences of three chloroplast (trnL‐F, matK, and rbcL) and one nuclear (ITS) loci were used to infer the molecular phylogeny employing maximum likelihood and Bayesian analyses. According to our results, within Plumbaginoideae, Plumbago forms a non‐monophyletic assemblage, with Plumbago europaea sister to Plumbagella, while the other Plumbago species form a clade sister to Dyerophytum. Within Limonioideae, Ikonnikovia is nested in Goniolimon, rejecting its former segregation as genus distinct from Goniolimon. Limonium is divided into two major clades: Limonium subg. Pteroclados s.l., including L. sect. Pteroclados and L. anthericoides, and L. subg. Limonium. The latter is divided into three well‐supported subclades: the monospecific L. sect. Limoniodendron sister to a clade comprising a mostly non‐Mediterranean subclade and a Mediterranean subclade. Our results set the foundation for taxonomic proposals on sections and subsections of Limonium, namely: (a) the newly described L. sect. Tenuiramosum, created to assign L. anthericoides at the sectional rank; (b) the more restricted circumscriptions of L. sect. Limonium (= L. sect. Limonium subsect. Genuinae) and L. sect. Sarcophyllum (for the Sudano‐Zambezian/Saharo‐Arabian clade); (c) the more expanded circumscription of L. sect. Nephrophyllum (including species of the L. bellidifolium complex); and (d) the new combinations for L. sect. Pruinosum and L. sect. Pteroclados subsect. Odontolepideae and subsect. Nobiles.

Limonium homoploid and heteroploid intra‐ and interspecific crosses unveil seed anomalies and neopolyploidy related to sexual and/or apomictic reproduction

AbstractApomixis is a form of asexual reproduction that consists in cloning through seeds. In Limonium (Plumbaginaceae) species present a pollen–stigma dimorphism linked to a sporophytic self‐incompatibility system associated with sexual and/or apomictic reproductive modes. Previous work in other genera suggests that the emergence of apomixis is associated with hybridization and/or polyploidy. In this study, our goal was to test the ability of diploid and tetraploid species to hybridize and to evaluate the variate outcomes from these crosses. To achieve this, sexual diploid (L. nydeggeri, L. ovalifolium) and facultative apomict tetraploid (L. binervosum, L. dodartii) plants from cultivated material, previously cytogenetically and reproductively characterized, were used for experimental intra‐ and interspecific crosses. Genome sizes, ploidy levels and morphology were examined in the resulting progenies. Results showed a high production of viable seeds in particular in plants from tetraploid × diploid (heteroploid) crosses. In these crosses, some seedlings exhibited pleiocotyly (tricotyl, tetracotyl), while others showed polyembryony. In both homoploid (diploid × diploid) and heteroploid (tetraploid × diploid) crosses, most of the offspring plants were morphologically and in their ploidy similar to the female receiver, although some morphological abnormalities were found. Molecular progeny tests using the nrDNA ITS1‐ITS2 sequence demonstrated an astounding range of diploid offspring plants originated from diploid × diploid crosses that were either genetically similar or distinct from parental plants. Although in intraspecific crosses most of the resulting progeny was diploid, one triploid plant was formed. Moreover, in homoploid interspecific crosses, neopolyploids (two tetraploid plants) were produced. Progeny plants from heteroploid crosses always showed nrDNA ITS1‐ITS2 sequences identical to the parental plant used as female receiver. In conclusion, diploid homoploid crosses presented genetically diverse offspring arising from sexual reproduction. By contrast, heteroploid crosses generated clonal, maternal (apomictic) offspring.

Edaphic characterization of coastal Western Mediterranean Limonium (Plumbaginaceae)

Introduction – Edaphic factors influence the structure and composition of plant communities. The main objective is to identify soil properties associated with the presence of different Limonium species. Methods – We conducted a systematic phytocoenological and edaphic survey over 37 locations across the Balearic archipelago. Canonical Correspondence Analysis was applied to the physicochemical characteristics of soils in which 29 species of Limonium grow. Results – The distribution of Limonium species has a high correlation to specific edaphic factors. The species can be clustered into four major groups. The first group represents species with a high edaphic selectivity – soils characterized by having a high proportion of sand, SO42- and Ca2+. The second group is composed of species with high levels of SAR, OM and SO42- /Cl- ratio. The third group includes species present in soils with a loamy texture, low Ca2+ /Mg2+ ratio and high levels of CO32- and Mg2+. The fourth group includes plants which colonize soils that have a sandy texture, low salinity and high proportion of CO32-. Conclusions – A strong correlation between the distribution of Limonium species and soil characteristics exist. The study contributes to the establishment of foundations for habitat conservation, cultivation and recovery projects for endangered species of Limonium.

Genome sizes and phylogenetic relationships suggest recent divergence of closely related species of the Limonium vulgare complex (Plumbaginaceae)

Limonium vulgare and related species form a complex group, but until now cytological and genetic studies have been based on single species and specific geographical areas. We investigated genome size, karyological and genetic diversity in samples from Western Mediterranean and evaluated the phylogenetic relationships among the species of this complex. Genome size was assessed using flow cytometry on samples from natural populations of L. vulgare, L. maritimum and L. narbonense. Chromosome counts were conducted in plants obtained from seeds collected in the field. The internal transcribed spacer ITS1 of the nuclear rDNAs was used to assess ITS polymorphisms as well as the phylogenetic relationships within the L. vulgare complex. Our analyses showed that all species were tetraploid, with the chromosome number of L. maritimum being presented here for the first time. Significant differences were observed in genome size, with L. narbonense having lower genome sizes than the other two species, and possible aneuploids being detected. Ten new ITS sequences from L. vulgare, L. narbonense and L. maritimum were provided. Most species’ populations showed unique ribotypes, and L. narbonense has the highest ribotype diversity. One of the L. maritimum populations presented a closer genetic relationship with L. vulgare, whereas the other two seemed to be more related with L. narbonense. Phylogenetic analyses confirmed that L. vulgare and L. narbonense form a monophyletic group, sister to the remaining Limonium species. Our results put into evidence that the studied species may represent a relatively early stage of divergence.

Distribution pattern and salt excretion rate of salt glands in two recretohalophyte species of Limonium (Plumbaginaceae)

Salt glands are the unique salt-excreting structures of recretohalophytes. The most important strategy by which recretohalophytes tolerate salinity is to excrete excess salt via salt glands. However, little is known about how the salt gland distribution pattern and salt excretion rate of dicotyledon recretohalophytes are correlated with salinity. This work focuses on the salt gland distribution patterns and excretion rates in two Limonium species of Plumbaginaceae under control conditions and NaCl stress. In the two dicotyledonous recretohalophytes, Limonium bicolor and Limonium gmelinii, the majority of the salt glands and veins were separated by no more than one epidermal cell, and the percentage of salt glands near veins of L. bicolor was greater than that of L. gmelinii. No salt glands were located alongside one another in either of the Limonium species and the salt glands and stomata were usually separated by more than one epidermal cell. The salt gland density on the adaxial epidermis of L. bicolor was greater than that of the abaxial epidermis whereas L. gmelinii showed the opposite trend. The optimal NaCl concentrations for maximal biomass were 100mM for L. bicolor and 50mM for L. gmelinii. Salt gland and stomata density increased when the NaCl concentration was higher than the optimal concentration. The Na+ excretion rate of a single salt gland in both Limonium species increased with increasing NaCl concentrations. These results suggest that salt gland distribution patterns and excretion rates are positively correlated with salt tolerance.

Assessing Effects of Temperature Change on Four Limonium Species from Threatened Mediterranean Salt-Affected Habitats

Evaluation of tolerance to temperatures in the germination stage is an important tool to foresee the possible effect of global warming. Moreover, establishing germination protocols for endemic and threatened species is most interesting for restoration and management plans that focus on habitat conservation. Seed germination was analysed in four species of the genus Limonium (L. virgatum, L. narbonense, L. girardianum and L. santapolense) growing in two saline habitats on the eastern coast of the Iberian Peninsula. The study aim was to compare responses of seed germination to different temperature regimes. Seeds were collected in two protected areas, ‘L´ Albufera de Valencia’ (province of Valencia) and ‘Clot de Galvany’ (province of Alicante). Responses to temperature were checked within a constant range, 10-30 °C at 5 °C intervals, by considering the environmental conditions at both sampling sites. The final germination percentage and mean germination time (MGT) were calculated after 30 days. In order to compare thermal responses, base temperature (Tb) and thermal time (S) were determined for each species. The possible correlations of soil parameters with the species´ germination pattern were also analysed. To identify distinct behavioural groups, a non-linear principal component analysis was performed. Significant differences between species were found in the velocity of germination. A fast response as opportunistic germination was observed in all the species. The base temperature and thermal time showed significant differences in competitiveness between species.

Limonium ilergabonum (Plumbaginaceae), a new species from northeastern Iberian Peninsula

A new species of Limonium (Plumbaginaceae), L. ilergabonum sp. nov., is described from the inland gypsum soils of the northeastern Iberian Peninsula (Flix, Tarragona province, Spain). The new species is closely related to the Iberian L. hibericum on morphological traits as assessed by multivariate ordination analysis (PCA), but it can be distinguished by the narrower leaves, inflorescence shape, and longer calyx. Limonium ilergabonum shows mixoploidy (2n = 18, 2n = 27), irregular pollen shapes, low levels of pollen stainability, and a single pollen-stigma combination morph (B type). All evidences suggest that L. ilergabonum is a polyploid relict species with an apomictic breeding system that has been originated from 2n = 18 ancestors, but to date any known extant diploid species may be suggested as a likely parental species.

Phylogenetic relationships of Limonium (Plumbaginaceae) inferred from multiple chloroplast and nuclear loci

AbstractLimonium (Plumbaginaceae) is a nearly cosmopolitan halophytic genus with ca. 350 species, a large number of which occurs in the Mediterranean region. We have generated a sequence dataset of several plastid (trnK intron including the matK gene, petD intron with the petB‐petD spacer, trnL intron with the trnL‐trnF spacer) and one nuclear region (nrITS), spanning the major lineages within the genus (102 accessions representing 76 species of Limonium) and related genera. Bayesian inference, maximum likelihood and maximum parsimony analyses converge on a Limonium clade that includes the genera Afrolimon, Eremolimon and Linczevskia. Plastid and nuclear trees congruently depict a number of major clades, most of them with a characteristic vegetative morphology and growing in specific habitats. Although the position of Afrolimon is incongruently resolved as the sister of a subclade comprising Limonium vulgareand relatives in the plastid trees versus being the sister to a Limonium sogdianum–L. bellidifolium–L. aureum clade in the nrITS tree, both genomic compartments show that Afrolimon is deeply nested in Limonium. Eremolimon is resolved as a close relative of L. sogdianum and L. ferganense based on plastid and nuclear genomic compartments. Our phylogenetic analysis resulted in an improved picture of internal relationships. Apart from the monophyletic subgenera Limonium subg. Limonium and L.subg.Pteroclados , nine well‐supported clades of L. subg. Limonium are evident, several of which are predominantly constituted by species of a certain geographic range. A subclade of Irano‐Turanian species (L. iranicum, L. suffruticosum and relatives) is shown to be unrelated to a L. axillare subclade, indicating an independent origin of the woody habit in these two lineages. Limonium sect. Iranolimonsect. nov. is described for the Irano‐Turanian woody lineage. The new combination L. sect. Circinaria comb. nov., is established for taxa sometimes included in Afrolimon.

Coordination between leaf CO2 diffusion and Rubisco properties allows maximizing photosynthetic efficiency in Limonium species.

High photosynthetic efficiency intrinsically demands tight coordination between traits related to CO2 diffusion capacity and leaf biochemistry. Although this coordination constitutes the basis of existing mathematical models of leaf photosynthesis, it has been barely explored among closely related species, which could reveal rapid adaptation clues in the recent past. With this aim, we characterized the photosynthetic capacity of 12 species of Limonium, possessing contrasting Rubisco catalytic properties, grown under optimal (WW) and extreme drought conditions (WD). The availability of CO2 at the site of carboxylation (Cc ) determined the photosynthetic capacity of Limonium under WD, while both diffusional and biochemical components governed the photosynthetic performance under WW. The variation in the in vivo caboxylation efficiency correlated with both the concentration of active Rubisco sites and the in vitro-based properties of Rubisco, such as the maximum carboxylase turnover rate (kcatc ) and the Michaelis-Menten constant for CO2 (Kc ). Notably, the results confirmed the hypothesis of coordination between the CO2 offer and demand functions of photosynthesis: those Limonium species with high total leaf conductance to CO2 have evolved towards increased velocity (i.e. higher kcatc ), at the penalty of lower affinity for CO2 (i.e. lower specificity factor, Sc/o ).

Phytochemical characterization and biological activity of secondary metabolites from three Limonium species

The comparative phytochemical constituents of three Limonium species Limonium myrianthum, Limonium leptophyllum, and Limonium gmelinii afforded a new compound (2R,3S)-2,3,4-trihydroxy-2-methylbutyl gallate (1) and twenty known compounds (2–21). Each species displayed different profiles in their phytochemical constituents. The isolated compounds (1–21) were evaluated for antifungal, antimalarial, and antitrypanosomal activities. Compound 1 showed good activity against chloroquine-resistant and chloroquine-sensitive strains of malaria, while compound 5 displayed moderate antimalarial activity. Compound 14 showed a significant activity against Trypanosoma brucei.

Nomenclatural and distributional remarks on Limonium tenoreanum (Plumbaginaceae), a narrow endemic from southern Italy

A rare Limonium species, which is strictly endemic to a short stretch of the coast of the Sorrento Peninsula (Gulf of Naples, Campania region, southern Italy), has been known as Statice tenoreana since the first half of 19th century. Although this name was published without a diagnosis or description by Gussone, it can be considered as validly published by an indirect reference to a previous work of Tenore (‘Flora Napolitana’). Therefore, the combination Limonium proposed more than a century later by Pignatti is correct. The name is lectotypified here by a specimen preserved at NAP in the Tenore collection.