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Aleppo pine ( Pinus halepensis Mill.) is an important component of Mediterranean forests. However, the scarcity of molecular resources limits population genetics studies shedding light on biogeographical patterns and/or within-population dynamics in this species. This is due to the complexity of conifers' genome, which includes many repetitive elements, and to the low genetic variation found in large parts of Aleppo pine distribution. The use of Next-Generation Sequencing (NGS) allows the development of molecular resources even for non-model species with complex genomes (i.e., complete genome and transcriptome sequencing, high numbers of neutral and putative functional markers, EST-SSRs, and SNPs). In this study, we developed a set of 14 polymorphic microsatellites (EST-SSRs) derived from transcriptome data, and combined them in three highly informative multiplexes for cost-effective genetic studies in P. halepensis. The three multiplexes were designed to allow addition of previously available SSRs of adequate size. Loci included in the EST-SSR multiplexes generally showed no null alleles and replicable, easily scorable amplification patterns. Furthermore, the cross-species transferability to two other Mediterranean pines ( Pinus pinaster Aiton and Pinus heldreichii H. Christ 1863) was tested, revealing that these EST-SSRs are highly transferable and polymorphic across Mediterranean Pinus species. Finally, preliminary genetic analyses on multiple populations showed some potential applications of the newly developed EST-SSRs to population genetics studies within and among Mediterranean pine populations. [ABSTRACT FROM AUTHOR]

The strong association observed between fire regimes and variation in plant adaptations to fire suggests a rapid response to fire as an agent of selection. It also suggests that fire-related traits are heritable, a precondition for evolutionary change. One example is serotiny, the accumulation of seeds in unopened fruits or cones until the next fire, an important strategy for plant population persistence in fire-prone ecosystems. Here, we evaluate the potential of this trait to respond to natural selection in its natural setting. For this, we use a SNP marker approach to estimate genetic variance and heritability of serotiny directly in the field for two Mediterranean pine species. Study populations were large and heterogeneous in climatic conditions and fire regime. We first estimated the realized relatedness among trees from genotypes, and then partitioned the phenotypic variance in serotiny using Bayesian animal models that incorporated environmental predictors. As expected, field heritability was smaller (around 0.10 for both species) than previous estimates under common garden conditions (0.20). An estimate on a subset of stands with more homogeneous environmental conditions was not different from that in the complete set of stands, suggesting that our models correctly captured the environmental variation at the spatial scale of the study. Our results highlight the importance of measuring quantitative genetic parameters in natural populations, where environmental heterogeneity is a critical aspect. The heritability of serotiny, although not high, combined with high phenotypic variance within populations, confirms the potential of this fire-related trait for evolutionary change in the wild. [ABSTRACT FROM AUTHOR]

Abstract: Campylobacter jejuni and Campylobacter coli are important animal-related waterborne pathogens that are distributed worldwide. To further understand Campylobacter populations in water from the Mediterranean area, the genetic diversity of environmental strains was analyzed using multilocus sequence typing (MLST). MLST was also used to determine the potential geographical differences between these bacterial strains and other campylobacters isolated worldwide. The typing study was conducted using 58 strains isolated from the Llobregat river and other water sources, such as urban sewage, animal wastewater and clinical samples. Thirty-nine different sequence types were obtained; eight of these sequences were described for the first time in this study, suggesting the presence of local strains. The identified C. jejuni strains were the most diverse population, whereas the identified C. coli strains showed a high clonal structure, which clustered most of the sequence types into a few clonal complexes. The strains were not exclusively related to specific water sources. However, comparing the identified strains with an international database showed that most of the Mediterranean strains that were exclusively isolated from environmental waters have previously been isolated from similar sources, particularly those obtained from river water. Additional studies, including those in different geographical areas using a wide range of Campylobacter sources, are required to improve the global knowledge concerning Campylobacter dissemination in the environment. [Copyright &y& Elsevier]

Background and Aims The study of local adaptation in plant reproductive traits has received substantial attention in short-lived species, but studies conducted on forest trees are scarce. This lack of research on long-lived species represents an important gap in our knowledge, because inferences about selection on the reproduction and life history of short-lived species cannot necessarily be extrapolated to trees. This study considers whether the size for first reproduction is locally adapted across a broad geographical range of the Mediterranean conifer species Pinus pinaster. In particular, the study investigates whether this monoecious species varies genetically among populations in terms of whether individuals start to reproduce through their male function, their female function or both sexual functions simultaneously. Whether differences among populations could be attributed to local adaptation across a climatic gradient is then considered. Methods Male and female reproduction and growth were measured during early stages of sexual maturity of a P. pinaster common garden comprising 23 populations sampled across the species range. Generalized linear mixed models were used to assess genetic variability of early reproductive life-history traits. Environmental correlations with reproductive life-history traits were tested after controlling for neutral genetic structure provided by 12 nuclear simple sequence repeat markers. Key Results Trees tended to reproduce first through their male function, at a size (height) that varied little among source populations. The transition to female reproduction was slower, showed higher levels of variability and was negatively correlated with vegetative growth traits. Several female reproductive traits were correlated with a gradient of growth conditions, even after accounting for neutral genetic structure, with populations from more unfavourable sites tending to commence female reproduction at a lower individual size. Conclusions The study represents the first report of genetic variability among populations for differences in the threshold size for first reproduction between male and female sexual functions in a tree species. The relatively uniform size at which individuals begin reproducing through their male function probably represents the fact that pollen dispersal is also relatively invariant among sites. However, the genetic variability in the timing of female reproduction probably reflects environment-dependent costs of cone production. The results also suggest that early sex allocation in this species might evolve under constraints that do not apply to other conifers. [ABSTRACT FROM PUBLISHER]

We have developed a set of eight polymorphic nuclear microsatellite markers for the Mediterranean shrub Pistacia lentiscus by means of an enriched library method. Characterization for the eight loci was carried out on 42 individuals from two populations sampled in southern Spain. The overall number of alleles detected was 59, ranging from three to 13 per locus. Expected heterozygosity per locus and population ranged from 0.139 to 0.895. Two loci albeit only in one population (Seville) departed significantly from Hardy–Weinberg equilibrium expectations and no linkage disequilibrium between pairs of loci was detected. These markers will be used in studies of gene flow across a fragmented landscape. [ABSTRACT FROM AUTHOR]

Aleppo pine (Pinus halepensis Mill.) is a relevant conifer species for studying adaptive responses to drought and fire regimes in the Mediterranean region. In this study, we performed Illumina next-generation sequencing of two phenotypically divergent Aleppo pine accessions with the aims of (i) characterizing the transcriptome through Illumina RNA-Seq on trees phenotypically divergent for adaptive traits linked to fire adaptation and drought, (ii) performing a functional annotation of the assembled transcriptome, (iii) identifying genes with accelerated evolutionary rates, (iv) studying the expression levels of the annotated genes and (v) developing gene-based markers for population genomic and association genetic studies. The assembled transcriptome consisted of 48,629 contigs and covered about 54.6 Mbp. The comparison of Aleppo pine transcripts to Picea sitchensis protein-coding sequences resulted in the detection of 34,014 SNPs across species, with a Ka /Ks average value of 0.216, suggesting that the majority of the assembled genes are under negative selection. Several genes were differentially expressed across the two pine accessions with contrasted phenotypes, including a glutathione-s-transferase, a cellulose synthase and a cobra-like protein. A large number of new markers (3334 amplifiable SSRs and 28,236 SNPs) have been identified which should facilitate future population genomics and association genetics in this species. A 384-SNP Oligo Pool Assay for genotyping with the Illumina VeraCode technology has been designed which showed an high overall SNP conversion rate (76.6%). Our results showed that Illumina next-generation sequencing is a valuable technology to obtain an extensive overview on whole transcriptomes of nonmodel species with large genomes., (© 2014 John Wiley & Sons Ltd.)