Comparison of the chloroplast genomes and phylogenomic analysis of Elaeocarpaceae.

Background: Elaeocarpaceae is a vital family in tropical and subtropical forests. Compared with the important position of Elaeocarpaceae species in forest ecosystem and the concern of medicinal value, the most research on Elaeocarpaceae are classification and taxonomy. Molecular systematics has corrected the morphological misjudgment, and it belongs to Oxalidales. Phylogenetic and divergence time estimates of Elaeocarpaceae is mostly constructed by using chloroplast gene fragments. At present, although there are reports on the chloroplast structure of Elaeocarpaceae, a comprehensive analysis of the chloroplast structure of Elaeocarpaceae is lacking. Methods: To understand the variation in chloroplast sequence size and structure in Elaeocarpaceae, the chloroplast genomes of nine species were sequenced using the Illumina HiSeq 2500 platform and further assembled and annotated with Elaeocarpus japonicus and Sloanea sinensis (family Elaeocarpaceae) as references. A phylogenomic tree was constructed based on the complete chloroplast genomes of the 11 species representing five genera of Elaeocarpaceae. Chloroplast genome characteristics were examined by using Circoletto and IRscope software. Results: The results revealed the following: (a) The 11 sequenced chloroplast genomes ranged in size from 157,546 to 159,400 bp. (b) The chloroplast genomes of Elaeocarpus, Sloanea, Crinodendron and Vallea lacked the rpl32 gene in the small single-copy (SSC) region. The large single-copy (LSC) region of the chloroplast genomes lacked the ndhK gene in Elaeocarpus, Vallea stipularis, and Aristotelia fruticosa. The LSC region of the chloroplast genomes lacked the infA gene in genus Elaeocarpus and Crinodendron patagua. (c) Through inverted repeat (IR) expansion and contraction analysis, a significant difference was found between the LSC/IRB and IRA/LSC boundaries among these species. Rps3 was detected in the neighboring regions of the LSC and IRb regions in Elaeocarpus. (d) Phylogenomic analysis revealed that the genus Elaeocarpus is closely related to Crinodendron patagua on an independent branch and Aristotelia fruticosa is closely related to Vallea stipularis, forming a clade with the genus Sloanea. Structural comparisons showed that Elaeocarpaceae diverged at 60 Mya, the genus Elaeocarpus diverged 53 Mya and that the genus Sloanea diverged 0.44 Mya. These results provide new insight into the evolution of the Elaeocarpaceae.

Phenotypic variation of fruit and ecophysiological traits among maqui (Aristotelia chilensis [Molina] Stuntz) provenances established in a common garden.

The domestication of forest species has traditionally relied on productivity issues. However, today there are concerns about the potential responses of natural populations and new cultivars to extreme climatic conditions derived from climate change and how to incorporate this knowledge into the domestication programs. Aristotelia chilensis (Molina) Stuntz ('Maqui') is a widely distributed native species in Chile. Its berry is considered a "super fruit" with an increasing interest in the food industry. This study investigated the phenotypic variation of growth, fruit, and ecophysiological traits of 20 A. chilensis clones originated from six provenances along the latitudinal gradient and established in a common-garden experiment in the Mediterranean zone of central Chile (center part of the species distribution). Differences among provenances were observed for most of the traits under study, especially between the northern and southernmost provenances (i.e., San Fernando versus Entre Lagos). Northern provenances showed higher development of vegetative tissue and fruit yield but lower intrinsic water use efficiency (WUEint) compared with southern ones. Clonal variation within provenances was found significant for the ripening index, WUEint, and fruit number and weight but not significant for traits related to the crown and leaf morphology. A genetic differentiation due to latitudinal cline was not evident in this study, but differences among provenances suggest local adaptation for some traits. The genotypic variation in productive traits must be considered in the outgoing domestication of the species and future selection programs.

Identification and Characterization of Microsatellite Loci in Maqui (Aristotelia chilensis [Molina] Stunz) Using Next-Generation Sequencing (NGS).

Maqui (Aristotelia chilensis [Molina] Stunz) is a small dioecious tree native to South America with edible fruit characterized by very high antioxidant capacity and anthocyanin content. To preserve maqui as a genetic resource it is essential to study its genetic diversity. However, the complete genome is unknown and only a few gene sequences are available in databases. Simple sequence repeats (SSR) markers, which are neutral, co-dominant, reproducible and highly variable, are desirable to support genetic studies in maqui populations. By means of identification and characterization of microsatellite loci from a maqui genotype, using 454 sequencing technology, we develop a set of SSR for this species. Obtaining a total of 165,043 shotgun genome sequences, with an average read length of 387 bases, we covered 64 Mb of the maqui genome. Reads were assembled into 4,832 contigs, while 98,546 reads remained as singletons, generating a total of 103,378 consensus genomic sequences. A total of 24,494 SSR maqui markers were identified. Of them, 15,950 SSR maqui markers were classified as perfects. The most common SSR motifs were dinucleotide (31%), followed by tetranucleotide (26%) and trinucleotide motifs (24%). The motif AG/CT (28.4%) was the most abundant, while the motif AC (89 bp) was the largest. Eleven polymorphic SSRs were selected and used to analyze a population of 40 maqui genotypes. Polymorphism information content (PIC) ranged from 0.117 to 0.82, with an average of 0.58. Non-significant groups were observed in the maqui population, showing a panmictic genetic structure. In addition, we also predicted 11150 putative genes and 3 microRNAs (miRNAs) in maqui sequences. This results, including partial sequences of genes, some miRNAs and SSR markers from high throughput next generation sequencing (NGS) of maqui genomic DNA, constitute the first platform to undertake genetic and molecular studies of this important species.

Anthocyanin profiling of wild maqui berries (Aristotelia chilensis [Mol.] Stuntz) from different geographical regions in Chile.

BACKGROUND: Maqui (Aristotelia chilensis) is a Chilean species which produces small berries that are collected from the wild. Anthocyanins, because of their health benefits, are the major focus of interest in maqui fruit. For this study, we examined anthocyanin and phenolic content of maqui fruits from individuals that belonged to four geographical areas in Chile, and used DNA marker analysis to examine the genetic variability of maqui populations that had distinctly different fruit anthocyanin content. RESULTS: Twelve primers generated a total of 145 polymorphic inter simple sequence repeat-polymerase chain reaction (ISSR-PCR) bands. ISSR-PCR showed different banding patterns for the individuals evaluated, confirming that maqui populations belonged to different genotypes. Maqui fruit from four different geographical regions during two consecutive growing seasons showed high total anthocyanin (6.6-15.0 g cy-3-glu kg⁻¹ fresh weight (FW)) and phenolic (10.7-20.5 g GAE kg⁻¹ FW) contents and different anthocyanin profiles. CONCLUSION: Three maqui genotypes exhibited significantly higher anthocyanin content than the others, as measured by pH differential method and high-performance liquid chromatography-mass spectrometry. Significant genetic diversity was noted within each ecological population. ISSR-PCR analysis provided a fingerprinting approach applicable for differentiation of maqui genotypes.

Genetic differentiation in Elaeocarpus photiniifolia (Elaeocarpaceae) associated with geographic distribution and habitat variation in the Bonin (Ogasawara) Islands.

Gene flow between populations in different environmental conditions can be limited due to divergent natural selection, thus promoting genetic differentiation. Elaeocarpus photiniifolia, an endemic tree species in the Bonin Islands, is distributed in two types of habitats, dry scrubs and mesic forests. We aim to elucidate the genetic differentiation in E. photiniifolia within and between islands and between the habitat types. We investigated genotypes of 639 individuals from 19 populations of E. photiniifolia and its closely-related E. sylvestris at 24 microsatellite loci derived from expressed sequence tags. The data revealed genetic differentiation (1) between E. photiniifolia and E. sylvestris (0.307 ≤ F ST ≤ 0.470), (2) between the E. photiniifolia populations of the Chichijima and Hahajima Island Groups in the Bonin Islands (0.033 ≤ F ST ≤ 0.121) and (3) between E. photiniifolia populations associated with dry scrubs and mesic forests in the Chichijima Island Group (0.005 ≤ F ST ≤ 0.071). Principal coordinate analysis and Bayesian clustering analysis also showed that genetically distinct groups were associated with the habitat types, and isolation by distance was not responsible for the genetic differentiation. These findings suggest that E. photiniifolia is divided into genetically differentiated groups associated with different environmental conditions in the Bonin Islands.

Herbivores modify selection on plant functional traits in a temperate rainforest understory.

There is limited evidence regarding the adaptive value of plant functional traits in contrasting light environments. It has been suggested that changes in these traits in response to light availability can increase herbivore susceptibility. We tested the adaptive value of plant functional traits linked with carbon gain in contrasting light environments and also evaluated whether herbivores can modify selection on these traits in each light environment. In a temperate rainforest, we examined phenotypic selection on functional traits in seedlings of the pioneer tree Aristotelia chilensis growing in sun (canopy gap) and shade (forest understory) and subjected to either natural herbivory or herbivore exclusion. We found differential selection on functional traits depending on light environment. In sun, there was positive directional selection on photosynthetic rate and relative growth rate (RGR), indicating that selection favors competitive ability in a high-resource environment. Seedlings with high specific leaf area (SLA) and intermediate RGR were selected in shade, suggesting that light capture and conservative resource use are favored in the understory. Herbivores reduced the strength of positive directional selection acting on SLA in shade. We provide the first demonstration that natural herbivory rates can change the strength of selection on plant ecophysiological traits, that is, attributes whose main function is resource uptake. Research addressing the evolution of shade tolerance should incorporate the selective role of herbivores.

Development of EST-SSR markers for Elaeocarpus photiniifolia (Elaeocarpaceae), an endemic taxon of the Bonin Islands.

PREMISE OF THE STUDY: Expressed sequence tag (EST)-derived microsatellite markers were developed for Elaeocarpus photiniifolia, an endemic taxon of the Bonin Islands. METHODS AND RESULTS: Initially, a complementary DNA (cDNA) library was constructed by de novo pyrosequencing of total RNA extracted from a seedling. A total of 267 primer pairs were designed from the library. Of the 48 tested loci, 25 loci were polymorphic among 41 individuals representing the entire geographical range of the species, with the number of alleles per locus and expected heterozygosity ranging from two to 14 and 0.09 to 0.86, respectively. Most loci were transferable to a related species, E. sylvestris. CONCLUSIONS: The developed markers will be useful for evaluating the genetic structure of E. photiniifolia.

The influence of environment and life-history traits on the distribution of genes and individuals: a comparative study of 11 rainforest trees.

This study investigates patterns of genetic connectivity among 11 co-distributed tropical rainforest tree species from the genus Elaeocarpus across a biogeographic barrier, the Black Mountain Corridor (BMC) in the Australian Wet Tropics (AWT). We analysed a combination of allelic and flanking region sequence data from microsatellite markers, and evaluated the relative influence of environmental preferences and functional traits on genetic diversity and gene flow. The results indicate that only in three species geographic structuring of haplotype distribution reflects a north vs. south of the BMC pattern. Environmental factors linked with altitude were recognized as affecting genetic trends, but the selective processes operating on upland species appear to be associated with competitiveness and regeneration opportunities on poor soil types rather than climate variables alone. In contrast to previous observations within southeastern Australian rainforests, genetic differentiation in the AWT appears to be associated with small-fruited rather than large-fruited species, highlighting how external factors can influence the dispersal dimension. Overall, this study emphasizes the importance of considering functional and environmental factors when attempting generalizations on landscape-level patterns of genetic variation. Understanding how plant functional groups respond to environmental and climatic heterogeneity can help us predict responses to future change.

Genetic effects of rainforest fragmentation in an early successional tree (Elaeocarpus grandis).

Rainforests in Australia and around the world have been extensively cleared and degraded. It is essential to recognize the changes in population diversity and dynamics that follow habitat fragmentation if better conservation and management strategies are to be developed. This study is an investigation of the medium term (over 100 years) effects of rainforest fragmentation on a long-lived, early successional tree species within a habitat matrix that includes various types of fragmented and undisturbed sites. Five microsatellite loci were used to assess the level and distribution of genetic variation across the southern range of Elaeocarpus grandis (Elaeocarpaceae). In all, 21 sites were sampled to provide a direct comparison between fragmented and undisturbed populations. Overall levels of diversity (A=3.4, He=0.568, f=0.094) were higher than those of closely related endemic species, but lower than those recorded across other rainforest trees. No significant genetic structure was detected across this species, suggesting the existence of efficient dispersal and colonization mechanisms responsible for the maintenance of gene flow. Rainforest fragments, and in particular those within the extensively cleared Big Scrub, show a trend for increased inbreeding levels caused by a loss of heterozygosity within juvenile cohorts. However, the overall rate of genetic decline within fragmented rainforests appears to be more subtle in E. grandis than across other species. A combination of ecological attributes and evolutionary history is likely to have contributed to this outcome and need to be considered in future rainforest restoration projects.