Phylogenetic challenges in a recently diversified and polyploid-rich Alyssum (Brassicaceae) lineage: low divergence, reticulation, and parallel polyploid speciation.

Elucidating the evolution of recently diverged and polyploid-rich plant lineages may be challenging even with high-throughput sequencing, both for biological reasons and bioinformatic difficulties. Here, we apply target enrichment with genome skimming (Hyb-Seq) to unravel the evolutionary history of the Alyssum montanum-A. repens species complex. Reconstruction of phylogenetic relationships in diploids supported recent and rapid diversification accompanied by reticulation events. Of the 4 main clades identified among the diploids, 3 clades included species from the Alps, Apennine, and Balkan peninsulas, indicating close biogeographic links between these regions. We further focused on the clade distributed from the Western Alps to the Iberian Peninsula, which comprises numerous polyploids as opposed to a few diploids. Using a recently developed PhyloSD (phylogenomic subgenome detection) pipeline, we successfully tracked the ancestry of all polyploids. We inferred multiple polyploidization events that involved 2 closely related diploid progenitors, resulting into several sibling polyploids: 2 autopolyploids and 6 allopolyploids. The skewed proportions of major homeolog-types and the occurrence of some minor homeolog-types, both exhibiting geographic patterns, suggest introgression with the progenitors and other related diploids. Our study highlights a unique case of parallel polyploid speciation that was enhanced by ecological and geographic separation and provides an excellent resource for future studies of polyploid evolution.

Multiple Drivers of High Species Diversity and Endemism Among Alyssum Annuals in the Mediterranean: The Evolutionary Significance of the Aegean Hotspot.

The Mediterranean Basin is a significant hotspot of species diversity and endemism, with various distribution patterns and speciation mechanisms observed in its flora. High species diversity in the Mediterranean is also manifested in the monophyletic lineage of Alyssum annuals (Brassicaceae), but little is known about its origin. These species include both diploids and polyploids that grow mainly in open and disturbed sites across a wide elevational span and show contrasting distribution patterns, ranging from broadly distributed Eurasian species to narrow island endemics. Here, we investigated the evolution of European representatives of this lineage, and aimed to reconstruct their phylogeny, polyploid and genome size evolution using flow cytometric analyses, chloroplast and nuclear high- and low-copy DNA markers. The origin and early diversification of the studied Alyssum lineage could be dated back to the Late Miocene/Pliocene and were likely promoted by the onset of the Mediterranean climate, whereas most of the extant species originated during the Pleistocene. The Aegean region represents a significant diversity center, as it hosts 12 out of 16 recognized European species and comprises several (sub)endemics placed in distinct phylogenetic clades. Because several species, including the closest relatives, occur here sympatrically without apparent niche differences, we can reject simple allopatric speciation via vicariance as well as ecological speciation for most cases. Instead, we suggest scenarios of more complex speciation processes that involved repeated range shifts in response to sea-level changes and recurrent land connections and disconnections since the Pliocene. In addition, multiple polyploidization events significantly contributed to species diversity across the entire distribution range. All seven polyploids, representing both widespread species and endemics to the western or eastern Mediterranean, were inferred to be allopolyploids. Finally, the current distribution patterns have likely been affected also by the human factor (farming and grazing). This study illustrates the complexity of evolutionary and speciation processes in the Mediterranean flora.

So Closely Related and Yet So Different: Strong Contrasts Between the Evolutionary Histories of Species of the Cardamine pratensis Polyploid Complex in Central Europe.

Recurrent polyploid formation and weak reproductive barriers between independent polyploid lineages generate intricate species complexes with high diversity and reticulate evolutionary history. Uncovering the evolutionary processes that formed their present-day cytotypic and genetic structure is a challenging task. We studied the species complex of Cardamine pratensis, composed of diploid endemics in the European Mediterranean and diploid-polyploid lineages more widely distributed across Europe, focusing on the poorly understood variation in Central Europe. To elucidate the evolution of Central European populations we analyzed ploidy level and genome size variation, genetic patterns inferred from microsatellite markers and target enrichment of low-copy nuclear genes (Hyb-Seq), and environmental niche differentiation. We observed almost continuous variation in chromosome numbers and genome size in C. pratensis s.str., which is caused by the co-occurrence of euploid and dysploid cytotypes, along with aneuploids, and is likely accompanied by inter-cytotype mating. We inferred that the polyploid cytotypes of C. pratensis s.str. are both of single and multiple, spatially and temporally recurrent origins. The tetraploid Cardamine majovskyi evolved at least twice in different regions by autopolyploidy from diploid Cardamine matthioli. The extensive genome size and genetic variation of Cardamine rivularis reflects differentiation induced by the geographic isolation of disjunct populations, establishment of triploids of different origins, and hybridization with sympatric C. matthioli. Geographically structured genetic lineages identified in the species under study, which are also ecologically divergent, are interpreted as descendants from different source populations in multiple glacial refugia. The postglacial range expansion was accompanied by substantial genetic admixture between the lineages of C. pratensis s.str., which is reflected by diffuse borders in their contact zones. In conclusion, we identified an interplay of diverse processes that have driven the evolution of the species studied, including allopatric and ecological divergence, hybridization, multiple polyploid origins, and genetic reshuffling caused by Pleistocene climate-induced range dynamics.

Diversification and independent polyploid origins in the disjunct species Alyssum repens from the Southeastern Alps and the Carpathians.

PREMISE: Disjunct distributions have been commonly observed in mountain plant species and have stimulated phylogeographic and phylogenetic research. Here we studied Alyssum repens, a member of the polyploid species complex A. montanum-A. repens, which exhibits SE Alpine-Carpathian disjunctions with a large elevational span and consists of diploid and tetraploid populations. We aimed to investigate the species' genetic and cytotype structure in the context of its distribution patterns, to elucidate the polyploid origins and to propose an appropriate taxonomic treatment. METHODS: We combined AFLP fingerprinting markers, sequence variation of the highly repetitive ITS region of rDNA and the low-copy DET1 nuclear gene, genome size, and morphometric data. RESULTS: We identified four geographically structured genetic lineages. One consisted of diploid populations from the foothills of the Southeastern Alps and neighboring regions, and the three others were allopatric montane to alpine groups comprising diploids and tetraploids growing in the Southeastern Carpathians and the Apuseni Mts. in Romania. CONCLUSIONS: We inferred a vicariance scenario associated with Quaternary climatic oscillations, accompanied by one auto- and two allopolyploidization events most likely involving a northern Balkan relative. Whereas genetic differentiation and allopatric distribution would favor the taxonomic splitting of this species, the genetic lineages largely lack morphological distinguishability, and their ecological, cytotype and genome size divergence is only partial. Even though we probably face here a case of incipient speciation, we propose to maintain the current taxonomic treatment of Alyssum repens as a single, albeit variable, species.

Low-residue breakfast during the preparation for colonoscopy using a polyethylene glycol electrolyte solution: a randomised non-inferiority trial.

GOALS: To test the hypothesis that the use of a low-residue breakfast (LRB) the day prior to colonoscopy was not inferior to consuming clear fluids alone (CFD) in patients undergoing outpatient colonoscopy with a polyethylene glycol (PEG) bowel preparation. BACKGROUND: Optimal colon cleansing is essential for complete visualisation of the mucosa during colonoscopy. Few studies have examined the effect of diet on the quality of bowel cleansing or tolerance in patients using a PEG bowel preparation for colonoscopy. METHODS: Randomised, single-blinded non-inferiority trial. Adult patients scheduled for outpatient colonoscopy with PEG solution were randomised to an LRB followed by clear fluids or CFD using either a traditional or split-dose PEG solution for bowel preparation. The primary outcome was colon cleansing based on the Ottawa Bowel Preparation Score (OBPS). RESULTS: On an intention-to-treat (ITT) basis, a total of 109 and 105 patients were included in the CFD and LRB arms, respectively, with 116 and 98 patients, respectively, for the per-protocol (PP) analysis. Although there was no difference in the mean total OBPS between the CFD or LRB arms in either the ITT or PP analysis, the threshold for non-inferiority was not met. Patient acceptance of the regimens was higher in the LRB arm than in the CFD arm in the ITT and PP analyses. CONCLUSIONS: This study failed to show the non-inferiority of an LRB in patients receiving bowel preparation with a PEG-based solution. A CFD should be prescribed when using a PEG bowel preparation. TRIAL REGISTRATION NUMBER: This trial is registered at ClinicalTrials.gov (NCT01454388).

A low-residue breakfast improves patient tolerance without impacting quality of low-volume colon cleansing prior to colonoscopy: a randomized trial.

OBJECTIVES: We compared the efficacy and patient tolerance of two dietary regimens used as an adjunct to standard bowel cleansing in preparation for elective colonoscopy. METHODS: Our prospective, randomized, single-blind, controlled, noninferiority trial compared two dietary regimens administered the day before colonoscopy. Subjects received low-volume bowel preparation. The primary outcome was efficacy of colon cleansing, rated by the Ottawa bowel preparation scale (OBPS). Patient acceptance of the assigned dietary regimen was assessed as a secondary outcome. RESULTS: The mean OBPS score for the standard clear-fluid diet group was 4.47, compared with 4.62 for the low-residue breakfast group, meeting criteria for noninferiority. The groups reported similar mean intensity of hunger on a visual analog hunger scale. However, overall satisfaction was higher with the low-residue diet than with the clear-fluid diet. CONCLUSIONS: A low-residue breakfast on the day before elective colonoscopy does not compromise the overall quality of bowel preparation as compared with a standard clear-fluid diet and is likely to be preferred by patients.