Chemical Composition of Myrtle (Myrtus communis L.) Berries Essential Oils as Observed in a Collection of Genotypes.

Myrtle (Myrtus communis L.) is a shrub spontaneously growing in the Mediterranean area. The leaf and fruit content of essential oils and phenolic compounds justify the wide use of the plant as medicinal and aromatic. Because of overexploitation of wild plants, a domestication process is in progress in different regions and the influence of the genotype variability on the chemical composition of fruit essential oils may be useful to breeding programs. Consequently, the analysis performed on a selected group of candidate clones growing in the same field collection in Sardinia is the object of this report. Forty-seven selections provided fully ripe fruits for essential oil extraction by hydrodistillation and Gas Chromatography-Mass Spectrometry (GC-MS) analysis. Only five candidate clones showed white fruits. The highest yield of essential oil was observed in the LAC31 genotype with 0.55 g·kg-1, while the samples BOS1, MON5, RUM4, RUM10, V4 and V8 showed values above 0.20 g·kg-1 and most of the genotypes under 0.10 g·kg-1. Geranyl acetate was the compound with the highest relative abundance. The second compound for relative abundance was the 1,8-cineole. Other compounds with high relative abundance were α-terpinyl acetate, methyleugenol, linalool, α-terpineol, ß-caryophyllene, α-humulene, Trans-caryophyllene oxide, and humulene epoxide II.

High Correlated Paternity Leads to Negative Effects on Progeny Performance in Two Mediterranean Shrub Species.

Anthropogenic habitat deterioration can promote changes in plant mating systems that subsequently may affect progeny performance, thereby conditioning plant recruitment for the next generation. However, very few studies yet tested mating system parameters other than outcrossing rates; and the direct effects of the genetic diversity of the pollen received by maternal plants (i.e. correlated paternity) has often been overlooked. In this study, we investigated the relation between correlated paternity and progeny performance in two common Mediterranean shrubs, Myrtus communis and Pistacia lentiscus. To do so, we collected open-pollinated progeny from selected maternal plants, calculated mating system parameters using microsatellite genotyping and conducted sowing experiments under greenhouse and field conditions. Our results showed that some progeny fitness components were negatively affected by the high correlated paternity of maternal plants. In Myrtus communis, high correlated paternity had a negative effect on the proportion and timing of seedling emergence in the natural field conditions and in the greenhouse sowing experiment, respectively. In Pistacia lentiscus, seedling emergence time under field conditions was also negatively influenced by high correlated paternity and a progeny survival analysis in the field experiment showed greater mortality of seedlings from maternal plants with high correlated paternity. Overall, we found effects of correlated paternity on the progeny performance of Myrtus communis, a self-compatible species. Further, we also detected effects of correlated paternity on the progeny emergence time and survival in Pistacia lentiscus, an obligate outcrossed species. This study represents one of the few existing empirical examples which highlight the influence that correlated paternity may exert on progeny performance in multiple stages during early seedling growth.

Antifungal and Herbicidal Effects of Fruit Essential Oils of Four Myrtus communis Genotypes.

The chemical composition of the essential oils isolated by hydrodistillation from the fruits of four selected Myrtus communis L. genotypes from Turkey was characterized by GC-FID and GC/MS analyses. 1,8-Cineole (29.20-31.40%), linalool (15.67-19.13%), α-terpineol (8.40-18.43%), α-pinene (6.04-20.71%), and geranyl acetate (3.98-7.54%) were found to be the major constituents of the fruit essential oils of all M. communis genotypes investigated. The oils were characterized by high amounts of oxygenated monoterpenes, representing 73.02-83.83% of the total oil compositions. The results of the fungal growth inhibition assays showed that the oils inhibited the growth of 19 phytopathogenic fungi. However, their antifungal activity was generally lower than that of the commercial pesticide benomyl. The herbicidal effects of the oils on the seed germination and seedling growth of Amaranthus retroflexus L., Chenopodium album L., Cirsium arvense (L.) Scop., Lactuca serriola L., and Rumex crispus L. were also determined. The oils completely or partly inhibited the seed germinations and seedling growths of the plants. The findings of the present study suggest that the M. communis essential oils might have potential to be used as natural herbicides as well as fungicides.

A curious case of resistance to a new encounter pathogen: myrtle rust in Australia.

Resistance genes (R genes) in plants mediate a highly specific response to microbial pathogens, often culminating in localized cell death. Such resistance is generally pathogen race specific and believed to be the result of evolutionary selection pressure. Where a host and pathogen do not share an evolutionary history, specific resistance is expected to be absent or rare. Puccinia psidii, the causal agent of myrtle rust, was recently introduced to Australia, a continent rich in myrtaceous taxa. Responses within species to this new pathogen range from full susceptibility to resistance. Using the myrtle rust case study, we examine models to account for the presence of resistance to new encounter pathogens, such as the retention of ancient R genes through prolonged 'trench warfare', pairing of resistance gene products and the guarding of host integrity.

Genetic variation and structure in the Mediterranean shrubs Myrtus communis and Pistacia lentiscus in different landscape contexts.

Studies concerning different habitat configurations can provide insights into the complex interactions between species' life-history traits and the environment and can help to predict patterns in population genetics. In this study, we compared patterns of genetic variation in two Mediterranean shrub species (Myrtus communis and Pistacia lentiscus) that co-occur in populations within three contrasting landscape contexts: continuous, fragmented-connected and fragmented-isolated populations. Analysing variation at microsatellites loci, our results revealed weak responses to the landscape contexts. We rather found a population-specific response in both study species. However, despite both study species sharing similar levels of genetic diversity, Myrtus displayed higher levels of homozygosity and genetic differentiation among populations, stronger patterns of within-population spatial genetic structure, lower values of mutation-scaled effective population size and stronger evidence for recent genetic bottlenecks than Pistacia. This result highlights the influence of past events (e.g. historical connectivity, fluctuations in population size) and local factors (e.g. microhabitat availability for recruitment, habitat quality, plant density, native fauna) and that the landscape configuration per se (i.e. fragment size and/or isolation) might not completely determine the species' genetic patterns.

The genetic diversity of Sardinian myrtle (Myrtus communis L. ) populations

Background: The myrtle (Myrtus communis) is a common shrub widespread in the Mediterranean Basin. Its fruit and leaves exhibit antioxidant, antibacterial and antifungal properties, and are used for their content of essential oils and for their medicinal properties, but most commonly as an ingredient in locally made liquor. The uncontrolled exploitation of natural stands has reduced both the species' geographical coverage and the size of individual populations. The selection of genotypes for controlled cultivation requires a characterization of the genetic diversity present both within and between populations. Results: Genotypic variation was evaluated using ISSR profiling and genetic diversity characterized using standard population genetics approaches. Two major clusters were identified: one capturing all the candidate cultivars selected from various Sardinian localities, and the other wild individuals collected from Asinara, Corsica and Surigheddu. A moderate level of gene flow between the Sardinian and Corsican populations was identified. Discriminant analysis of principal components revealed a level of separation among the wild populations, confirming the population structure identified by the clustering methods. Conclusions: The wild accessions were well differentiated from the candidate cultivars. The level of genetic variability was high. The genetic data were compatible with the notion that myrtle has a mixed pollination system, including both out-pollination by insects and self-pollination. The candidate cultivars are suggested to represent an appropriate basis for directed breeding.

Surviving in mountain climate refugia: new insights from the genetic diversity and structure of the relict shrub Myrtus nivellei (Myrtaceae) in the Sahara Desert.

The identification of past glacial refugia has become a key topic for conservation under environmental change, since they contribute importantly to shaping current patterns of biodiversity. However, little attention has been paid so far to interglacial refugia despite their key role for the survival of relict species currently occurring in climate refugia. Here, we focus on the genetic consequences of range contraction on the relict populations of the evergreen shrub Myrtus nivellei, endemic in the Saharan mountains since at least the end of the last Green Sahara period, around 5.5 ka B.P. Multilocus genotypes (nuclear microsatellites and AFLP) were obtained from 215 individuals collected from 23 wadis (temporary rivers) in the three main mountain ranges in southern Algeria (the Hoggar, Tassili n'Ajjer and Tassili n'Immidir ranges). Identical genotypes were found in several plants growing far apart within the same wadis, a pattern taken as evidence of clonality. Multivariate analyses and Bayesian clustering revealed that genetic diversity was mainly structured among the mountain ranges, while low isolation by distance was observed within each mountain range. The range contraction induced by the last episode of aridification has likely increased the genetic isolation of the populations of M. nivellei, without greatly affecting the genetic diversity of the species as a whole. The pattern of genetic diversity observed here suggests that high connectivity may have prevailed during humid periods, which is consistent with recent paleoenvironmental reconstructions.

Studying genetic variability of pomegranate (Punica granatum L.) based on chloroplast DNA and barcode genes.

Chloroplast DNA has been used extensively to analyze plant phylogenies at different taxonomic levels because of its size, organization and sequence conservation. In the present research, two chloroplastic regions, petA­psaJ, trnC­trnD and four DNA barcodes (trnH­psbA, ITS, rbcL, matK), were used to introduce suitable regions for the assessment of genetic diversity among P. granatum L. genotypes. Analysis of psbE­petL in petA­psaJ region revealed 1,300 nucleotides with 4.29 % genetic diversity among genotypes, while trnC­petN in trnC­trnD region showed 1.8 % genetic diversity. Therefore, despite the results obtained from the study of other plants, the trnC­trnD region had a low potential for the evaluation of diversity among pomegranate genotypes. Analysis of DNA barcodes in pomegranate showed that trnH­psbA (genetic diversity 2.91 %) provides the highest intra-species variation, followed by ITS (genetic diversity 0.44 %). Eighteen genotypes from different geographical origins of Iran were used to investigate psbE­petL and trnH­psbA potential as novel barcodes to determine genetic polymorphism and characterize pomegranate genotypes. The results suggested that two regions, psbE­petL and trnH­psbA, were more suitable for determining intra-species relationships of pomegranate.

Contrasting heterozygosity-fitness correlations between populations of a self-compatible shrub in a fragmented landscape.

The mechanisms underlying heterozygosity-fitness correlations (HFCs) are subject of intense debates, especially about how important population features such as size or degree of isolation influence HFCs. Here, we report variation in HFCs between Large and Small populations of a self-compatible shrub (Myrtus communis) occurring within an extremely fragmented landscape. In each of the five study populations, we obtained data on both heterozygosity and fitness for 9-12 maternal families (i.e. offspring from the same mother plant). Whereas heterozygosity explained most of the variance (60-86 %) in growth rate of seedling families within Large populations, this relationship was absent within Small populations. Our results suggest that inbreeding may explain the observed HFCs within Large populations, and that different genetic processes (such as genetic drift and/or selection) could have overridden HFCs within Small populations. While it is difficult to draw general conclusions from five populations, we think our results open new research perspectives on how different genetic processes underlie variation in HFCs under different population contexts. Our study also points to a need for further attention on the complex relationships between heterozygosity in self-compatible plants and their progeny in relation to mating system variation. Finally, our results provide interesting new insights into how population genetic diversity is maintained or lost in a highly fragmented landscape.

Isolation of Ty1-copia retrotransposon in myrtle genome and development of S-SAP molecular marker.

Long terminal repeat (LTR)-retrotransposons are mobile genetic elements that are ubiquitous in plants and constitute a major portion of their nuclear genomes. LTR- retrotransposons possess unique properties that make them appropriate for investigating relationships between populations, varieties and closely related species. Myrtus communis L. is an evergreen shrub growing spontaneously throughout the Mediterranean area. Accessions show significant variations for agriculturally important traits, so the development of specific molecular markers for conservation and characterization of myrtle germplasm is desirable to conserve biodiversity. In this study, we isolated the first retrotransposon Ty1-copia-like element (Tmc1) in Myrtus communis L. genome and used this as a molecular marker. We successfully employed the S-SAP marker system to specifically characterize four myrtle accessions belonging to different areas in the province of Caserta (Italy). The high level of polymorphism detected in isolated LTRs, make Tmc1 a good molecular marker for this species. Our findings confirm that retrotransposon-based molecular markers are particularly valuable tools for plant molecular characterization studies.

Molecular cloning and characterization of a linalool synthase from lemon myrtle.

Using a homology-based PCR strategy, we identified a cDNA with sequence similarity to linalool synthase from lemon myrtle. Functional expression of the cDNA (designated BcLS) gene in Escherichia coli yielded an active enzyme capable of catalyzing the conversion of geranyl diphosphate to (-)-linalool, i.e., an acyclic monoterpene alcohol, and to lesser amounts of cyclic monoterpenes. The kinetic parameters of BcLS were similar to those of synthases producing cyclic monoterpenes. PCR analysis revealed that the BcLS gene transcript was ubiquitously expressed in lemon myrtle and was upregulated in response to jasmonic acid treatment. Although the physiological role of neryl diphosphate (NPP) dependency of BcLS remains unclear, the cyclization activity of BcLS was enhanced when NPP was used as substrate, resulting in predominant production of cyclic monoterpenes. These findings indicate that BcLS has novel specificity and kinetic parameters, but its physiological responses to stresses such as insect damage appear to be similar to known linalool synthases.

Karyological features of wild and cultivated forms of myrtle (Myrtus communis, Myrtaceae).

Myrtle is an evergreen shrub or small tree widespread throughout the Mediterranean region. In Turkey, both cultivated and wild forms, differing in plant and fruit size and fruit composition, can be found. These differences may have resulted from the domestication of the cultivated form over a long period of time. We investigated whether wild and cultivated forms of myrtle differ in karyological features (i.e., number of somatic chromosomes and relative genome size). We sampled two wild forms and six cultivated types of myrtle. All the samples had the same chromosome number (2n = 2x = 22). The results were confirmed by 4',6-diamidino-2-phenylindole (DAPI) flow cytometry. Only negligible variation (approximately 3%) in relative fluorescence intensity was observed among the different myrtle accessions, with wild genotypes having the smallest values. We concluded that despite considerable morphological differentiation, cultivated and wild myrtle genotypes in Turkey have similar karyological features.

In vitro propagation of ornamental myrtus (Myrtus communis).

The Myrtle (Myrtus communis L.) is an evergreen shrub typical of the Mediterranean area; it is an interesting plant with multipurpose use. The ornamental use takes into account the production of green cut branches for indoor decoration and production of pot plants for gardening. In this species, there is a great variability in the natural germplasm around the Mediterranean coasts for type and size of fruit, plant architecture, leaf size and internode length. Selected genotypes have been successfully sterilized and cultured in vitro. The shoots were multiplied on MS (16) salts and vitamins, with 0.5 mg/L BA and 0.2 mg/L IAA. Clones showed variation of multiplication rate and rooting percentage. IAA or IBA at 0.5 mg/L increased the rooting percentage and noticed differences in root number and length. The sucrose concentration can affect rooting, such as light intensity during the in vitro rooting phase can modulate biomass production and chlorophyll content. The combination of these factors enhanced the frequency rate of acclimatization.

Networks of spatial genetic variation across species.

Spatial patterns of genetic variation provide information central to many ecological, evolutionary, and conservation questions. This spatial variability has traditionally been analyzed through summary statistics between pairs of populations, therefore missing the simultaneous influence of all populations. More recently, a network approach has been advocated to overcome these limitations. This network approach has been applied to a few cases limited to a single species at a time. The question remains whether similar patterns of spatial genetic variation and similar functional roles for specific patches are obtained for different species. Here we study the networks of genetic variation of four Mediterranean woody plant species inhabiting the same habitat patches in a highly fragmented forest mosaic in Southern Spain. Three of the four species show a similar pattern of genetic variation with well-defined modules or groups of patches holding genetically similar populations. These modules can be thought of as the long-sought-after, evolutionarily significant units or management units. The importance of each patch for the cohesion of the entire network, though, is quite different across species. This variation creates a tremendous challenge for the prioritization of patches to conserve the genetic variation of multispecies assemblages.

Population genetic structure in Myrtus communis L. in a chronically fragmented landscape in the Mediterranean: can gene flow counteract habitat perturbation?

Ancient managed landscapes provide ideal opportunities to assess the consequences of habitat fragmentation on the patterns of genetic diversity and gene flow in long-lived plant species. Using amplified fragment length polymorphism (AFLP) and allozyme markers, we quantified seed-mediated gene flow and population genetic diversity and structure in 14 populations of Myrtus communis (myrtle), a common endozoochorous shrub species of forest patches in lowland agricultural Mediterranean areas. Overall, allozyme diversity for myrtle was low (P(95) = 25%; A = 1.411; H(e) = 0.085) compared to other known populations, and a significant portion of populations (57%) had lower levels of allelic diversity and/or heterozygosity than expected at random, as shown by simulated resampling of the whole diversity of the landscape. We found significant correlations between allozyme variability and population size and patch isolation, but no significant inbreeding in any population. Genetic differentiation among populations for both allozyme and AFLP markers was significant (Phi(ST) = 0.144 and Phi(ST) = 0.142, respectively) but an isolation-by-distance pattern was not detected. Assignment tests on AFLP data indicated a high immigration rate in the populations (ca. 20-22%), likely through effective seed dispersal across the landscape by birds and mammals. Our results suggest that genetic isolation is not the automatic outcome of habitat destruction since substantial levels of seed-mediated gene flow are currently detectable. However, even moderate rates of gene flow seem insufficient in this long-lived species to counteract the genetic erosion and differentiation imposed by chronic habitat destruction.