Repeat-based phylogenomics shed light on unclear relationships in the monocentric genus Juncus L. (Juncaceae).

The repetitive fraction (repeatome) of eukaryotic genomes is diverse and usually fast evolving, being an important tool for clarify plant systematics. The genus Juncus L. comprises 332 species, karyotypically recognized by having holocentric chromosomes. However, four species were recently described as monocentric, yet our understanding of their genome evolution is largely masked by unclear phylogenetic relationships. Here, we reassess the current Juncus systematics using low-coverage genome skimming data of 33 taxa to construct repeats, nuclear rDNA and plastome-based phylogenetic hypothesis. Furthermore, we characterize the repeatome and chromosomal distribution of Juncus-specific centromeric repeats/CENH3 protein to test the monocentricity reach in the genus. Repeat-base phylogenies revealed topologies congruent with the rDNA tree, but not with the plastome tree. The incongruence between nuclear and plastome chloroplast dataset suggest an ancient hybridization in the divergence of Juncotypus and Tenageia sections 40 Myr ago. The phylogenetic resolution at section level was better fitted with the rDNA/repeat-based approaches, with the recognition of two monophyletic sections (Stygiopsis and Tenageia). We found specific repeatome trends for the main lineages, such as the higher abundances of TEs in the Caespitosi and Iridifolii + Ozophyllum clades. CENH3 immunostaining confirmed the monocentricity of Juncus, which can be a generic synapomorphy for the genus. The heterogeneity of the repeatomes, with high phylogenetic informativeness, identified here may be correlated with their ancient origin (56 Mya) and reveals the potential of comparative genomic analyses for understanding plant systematics and evolution.

Plastome evolution in the Caesalpinia group (Leguminosae) and its application in phylogenomics and populations genetics.

MAIN CONCLUSION: The chloroplast genomes of Caesalpinia group species are structurally conserved, but sequence level variation is useful for both phylogenomic and population genetic analyses. Variation in chloroplast genomes (plastomes) has been an important source of information in plant biology. The Caesalpinia group has been used as a model in studies correlating ecological and genomic variables, yet its intergeneric and infrageneric relationships are not fully solved, despite densely sampled phylogenies including nuclear and plastid loci by Sanger sequencing. Here, we present the de novo assembly and characterization of plastomes from 13 species from the Caesalpinia group belonging to eight genera. A comparative analysis was carried out with 13 other plastomes previously available, totalizing 26 plastomes and representing 15 of the 26 known Caesalpinia group genera. All plastomes showed a conserved quadripartite structure and gene repertoire, except for the loss of four ndh genes in Erythrostemon gilliesii. Thirty polymorphic regions were identified for inter- or intrageneric analyses. The 26 aligned plastomes were used for phylogenetic reconstruction, revealing a well-resolved topology, and dividing the Caesalpinia group into two fully supported clades. Sixteen microsatellite (cpSSR) loci were selected from Cenostigma microphyllum for primer development and at least two were cross-amplified in different Leguminosae subfamilies by in vitro or in silico approaches. Four loci were used to assess the genetic diversity of C. microphyllum in the Brazilian Caatinga. Our results demonstrate the structural conservation of plastomes in the Caesalpinia group, offering insights into its systematics and evolution, and provides new genomic tools for future phylogenetic, population genetics, and phylogeographic studies.

Aiming off the target: recycling target capture sequencing reads for investigating repetitive DNA.

BACKGROUND AND AIMS: With the advance of high-throughput sequencing, reduced-representation methods such as target capture sequencing (TCS) emerged as cost-efficient ways of gathering genomic information, particularly from coding regions. As the off-target reads from such sequencing are expected to be similar to genome skimming (GS), we assessed the quality of repeat characterization in plant genomes using these data. METHODS: Repeat composition obtained from TCS datasets of five Rhynchospora (Cyperaceae) species were compared with GS data from the same taxa. In addition, a FISH probe was designed based on the most abundant satellite found in the TCS dataset of Rhynchospora cephalotes. Finally, repeat-based phylogenies of the five Rhynchospora species were constructed based on the GS and TCS datasets and the topologies were compared with a gene-alignment-based phylogenetic tree. KEY RESULTS: All the major repetitive DNA families were identified in TCS, including repeats that showed abundances as low as 0.01 % in the GS data. Rank correlations between GS and TCS repeat abundances were moderately high (r = 0.58-0.85), increasing after filtering out the targeted loci from the raw TCS reads (r = 0.66-0.92). Repeat data obtained by TCS were also reliable in developing a cytogenetic probe of a new variant of the holocentromeric satellite Tyba. Repeat-based phylogenies from TCS data were congruent with those obtained from GS data and the gene-alignment tree. CONCLUSIONS: Our results show that off-target TCS reads can be recycled to identify repeats for cyto- and phylogenomic investigations. Given the growing availability of TCS reads, driven by global phylogenomic projects, our strategy represents a way to recycle genomic data and contribute to a better characterization of plant biodiversity.

Draft Genome Sequence of "Candidatus Phytoplasma pruni" (X-Disease Group, Subgroup 16SrIII-B) Strain ChTDIII from Argentina.

Herein, we report the draft genome sequence of "Candidatus Phytoplasma pruni" strain ChTDIII (subgroup 16SrIII-B). The final assembly consists of 790,517 nucleotides organized in 67 contigs (minimal size, 1 kb), with a G+C content of 29.4% and encoding 672 proteins.

Evolutionary convergence or homology? Comparative cytogenomics of Caesalpinia group species (Leguminosae) reveals diversification in the pericentromeric heterochromatic composition.

MAIN CONCLUSION: We demonstrated by cytogenomic analysis that the proximal heterochromatin of the Northeast Brazilian species of Caesalpinia group is enriched with phylogenetically conserved Ty3/Gypsy-Tekay RT, but diverge in the presence of Ty3/Gypsy-Athila RT and satDNA. The Caesalpinia Group includes 225 species and 27 monophyletic genera of which four occur in Northeastern Brazil: Erythrostemon (1 sp.), Cenostigma (7 spp.), Libidibia (1 sp.), and Paubrasilia (1 sp.). The last three genera are placed in different clades in the Caesalpinia Group phylogeny, and yet they are characterized by having a numerically stable karyotype 2n = 24 (16 M+8A) and GC-rich heterochromatic bands (chromomycin A3 positive/CMA+ bands) in the proximal chromosome regions. To characterize the composition of their heterochromatin and test for the homology of these chromosomal regions, genomic DNA was extracted from Cenostigma microphyllum, Libidibia ferrea, and Paubrasilia echinata, and sequenced at low coverage using the Illumina platform. The genomic repetitive fractions were characterized using a Galaxy/RepeatExplorer-Elixir platform. The most abundant elements of each genome were chromosomally located by fluorescent in situ hybridization (FISH) and compared to the CMA+ heterochromatin distribution. The repetitive fraction of the genomes of C. microphyllum, L. ferrea, and P. echinata were estimated to be 41.70%, 38.44%, and 72.51%, respectively. Ty3/Gypsy retrotransposons (RT), specifically the Tekay lineage, were the most abundant repeats in each of the three genomes. FISH mapping revealed species-specific patterns for the Tekay elements in the proximal regions of the chromosomes, co-localized with CMA+ bands. Other species-specific patterns were observed, e.g., for the Ty3/Gypsy RT Athila elements which were found in all the proximal heterochromatin of L. ferrea or restricted to the acrocentric chromosomes of C. microphyllum. This Athila labeling co-localized with satellite DNAs (satDNAs). Although the Caesalpinia Group diverged around 55 Mya, our results suggest an ancestral colonization of Tekay RT in the proximal heterochromatin. Thus, the present-day composition of the pericentromeric heterochromatin in these Northeast Brazilian species is a combination of the maintenance of an ancestral Tekay distribution with a species-specific accumulation of other repeats.

Development of ten microsatellite markers for Alibertia edulis (Rubiaceae), a Brazilian savanna tree species.

Ten microsatellite markers were developed using next-generation sequencing data for Alibertia edulis (Rubiaceae), a widely distributed species typical of Cerrado (Brazilian savanna) vegetation. The markers were polymorphic in the two populations analyzed. The numbers of alleles, and observed (HO) and expected (HE) heterozygosities per polymorphic locus ranged from 2 to 11, 0.091 to 1.0, and 0.100 to 0.937 respectively. The SSR loci demonstrated moderate to high polymorphism values in both populations analyzed, with PIC values ranging from 0.26 to 0.91, and total allele numbers ranging from three to 16. The inbreeding coefficient values were generally higher in the Piauí population (ranging from - 0.593 to 0.762) than in the Mato Grosso population (ranging from - 1 to 0.575). The differences observed between those disjunct populations suggest they harbor different alleles, which has implications for Cerrado conservation strategies. Those loci will be useful for population studies of A. edulis.

Development and characterization of eleven microsatellite loci for the tropical understory tree Paypayrola blanchetiana Tul. (Violaceae).

Microsatellites markers were developed for Paypayrola blanchetiana (Violaceae), a near-dispersing forest tree forming aggregated populations, to investigate genetic diversity and gene flow among subpopulations in a fragmented environment. Next generation sequencing (Illumina platform) was used to develop ten nuclear microsatellite loci and one plastid microsatellite locus that amplify in P. blanchetiana. Polymorphism was tested in two subpopulations separated by a distance of approximately 11 km. The identified loci contained between two and five alleles per locus. Observed heterozygosity ranged between 0.063 and 0.563 in both subpopulations, while expected heterozygosity ranged from 0.063 to 0.567 in the first, and 0.063-0.627 in the second subpopulation. The microsatellites are among the first in the family Violaceae and will be useful for population genetic studies in this species. Amplification was successful in one further Paypayrola species from Amazonia, which suggest a wider usefulness of the present markers.

A set of plastid microsatellite loci for the genus Dyckia (Bromeliaceae) derived from 454 pyrosequencing.

PREMISE OF THE STUDY: Phylogeographical analyses of Dyckia (Bromeliaceae) suffer from low levels of sequence variation. Plastid microsatellite markers were developed to achieve a better-resolved genus-wide plastid genealogy of Dyckia. • METHODS AND RESULTS: Approximately 84% of the D. marnier-lapostollei plastome was sequenced using 454 technology. Flanking primer pairs were designed for 34 out of 36 chloroplast simple sequence repeats (cpSSRs) detected, and 12 loci were further characterized by genotyping Dyckia samples at the level of populations and species. Three, five, and six cpSSRs were polymorphic among four individuals of D. limae, 12 individuals of D. dissitiflora, and 12 of D. pernambucana, respectively, with two to three alleles per locus and species. All loci were polymorphic among 19 different Dyckia species, with three to 10 alleles per locus. Ten primer pairs cross-amplified with bromeliad genera from five subfamilies. • CONCLUSIONS: The set of 12 cpSSR markers provides a toolbox to analyze phylogeographical patterns of Dyckia species.