[Leuko-glycemic index in patients with ST elevation myocardial infarction]. / Índice leucoglucémico en pacientes con infarto miocárdico con elevación del ST.

Background: The myocardial infarction-associated (MI) mortality is not only due cardiovascular complications, but intrahospital non-cardiovascular complications (IHnCVCs). The leuko-glycemic index (LGI) has been used as a prognostic marker for the development of cardiovascular complications in MI. We focused this study on identifying the cut-off point of LGI for the IHnCVCs development in patients with ST-segment elevation myocardial infarction (STEMI). Material and methods: In this single-center and crosssectional design, we included patients with STEMI. The biochemical analysis included glucose and leucocytes; with them we calculated the LGI. Receiver operating characteristic curve, univariate and bivariate analysis, and multivariate analysis for IHnCVCs development were performed. A p < 0.05 was considered statistically significant. Results: We included 1294 patients, 79.8% were men and 20.2% women. The main comorbidities were hypertension, diabetes mellitus and dyslipidemia. Six hundred forty-four (49.8%) patients presented IHNCVCs. The LGI > 1200 (AUC 0.817) predict the IHNCVCs development in STEMI patients. The variables that increased the IHNCVCs development were LGI > 1200, creatinine > 0.91 mg/dL, diabetes mellitus and age > 65 years. Hospital acquired pneumonia and cardiovascular complications increase the risk of death among STEMI patients. Conclusion: A LGI > 1200 increased, just over nine times, the risk of IHnCVC development in STEMI patients.

Introducción: la mortalidad asociada a infarto del miocardio (IM) no solo se debe a complicaciones cardiovasculares, sino tambien a complicaciones intrahospitalarias no cardiovasculares (CIHNC). El índice leuco-glucémico (ILG) se ha utilizado como un marcador pronóstico para el desarrollo de complicaciones cardiovasculares en el IM. Centramos este estudio en identificar el punto de corte de ILG para el desarrollo de CIHNC en pacientes con infarto de miocardio con elevación del segmento ST (IAMCEST). Material y métodos: en este diseño de un solo centro y transversal, incluimos pacientes con IAMCEST. El análisis bioquímico incluyó glucosa y leucocitos; se calculó ILG. Se realizaron análisis univariados y bivariados, curva ROC y análisis multivariado para el desarrollo de IAMCEST. Resultados: incluimos 1294 pacientes, 79.8% hombres y 20.2% mujeres. Las principales comorbilidades fueron: hipertensión arterial sistémica, diabetes mellitus y dislipidemia. Seiscientos cuarenta y cuatro pacientes (49.8%) presentaron CIHNC. El ILG > 1200 con área bajo la curva (AUC) 0.817 predice el desarrollo de CIHNC en pacientes con IAMCEST. Las variables que aumentaron el desarrollo de CIHNC fueron: ILG > 1200, creatinina > 0.91 mg/dL, diabetes mellitus y edad > 65 años. La neumonía intrahospitalaria y las complicaciones cardiovasculares aumentaron el riesgo de muerte entre los pacientes con IAMCEST. Conclusión: un LGI > 1200 aumentó más de nueve veces el riesgo de desarrollo de CIHNC en pacientes con IAMCEST.

Índice leucoglucémico en pacientes con infarto miocárdico con elevación del ST / Leuko-glycemic index in patients with ST elevation myocardial infarction

Introducción: la mortalidad asociada a infarto del miocardio (IM) no solo se debe a complicaciones cardiovasculares, sino también a complicaciones intrahospitalarias no cardiovasculares (CIHNC). El índice leuco-glucémico (ILG) se ha utilizado como un marcador pronóstico para el desarrollo de complicaciones cardiovasculares en el IM. Centramos este estudio en identificar el punto de corte de ILG para el desarrollo de CIHNC en pacientes con infarto de miocardio con elevación del segmento ST (IAMCEST). Material y métodos: en este diseño de un solo centro y transversal, incluimos pacientes con IAMCEST. El análisis bioquímico incluyó glucosa y leucocitos; se calculó ILG. Se realizaron análisis univariados y bivariados, curva ROC y análisis multivariado para el desarrollo de IAMCEST. Resultados: incluimos 1294 pacientes, 79.8% hombres y 20.2% mujeres. Las principales comorbilidades fueron: hipertensión arterial sistémica, diabetes mellitus y dislipidemia. Seiscientos cuarenta y cuatro pacientes (49.8%) presentaron CIHNC. El ILG > 1200 con área bajo la curva (AUC) 0.817 predice el desarrollo de CIHNC en pacientes con IAMCEST. Las variables que aumentaron el desarrollo de CIHNC fueron: ILG > 1200, creatinina > 0.91 mg/dL, diabetes mellitus y edad > 65 años. La neumonía intrahospitalaria y las complicaciones cardiovasculares aumentaron el riesgo de muerte entre los pacientes con IAMCEST. Conclusión: un LGI > 1200 aumentó más de nueve veces el riesgo de desarrollo de CIHNC en pacientes con IAMCEST.

Background: The myocardial infarction-associated (MI) mortality is not only due cardiovascular complications, but intrahospital non-cardiovascular complications (IHnCVCs). The leuko-glycemic index (LGI) has been used as a prognostic marker for the development of cardiovascular complications in MI. We focused this study on identifying the cut-off point of LGI for the IHnCVCs development in patients with ST-segment elevation myocardial infarction (STEMI).Material and methods: In this single-center and cross-sectional design, we included patients with STEMI. The biochemical analysis included glucose and leucocytes; with them we calculated the LGI. Receiver operating characteristic curve, univariate and bivariate analysis, and multivariate analysis for IHnCVCs development were performed. A p < 0.05 was considered statistically significant. Results: We included 1294 patients, 79.8% were men and 20.2% women. The main comorbidities were hypertension, diabetes mellitus and dyslipidemia. Six hundred forty-four (49.8%) patients presented IHNCVCs. The LGI > 1200 (AUC 0.817) predict the IHNCVCs development in STEMI patients. The variables that increased the IHNCVCs development were LGI > 1200, creatinine > 0.91 mg/dL, diabetes mellitus and age > 65 years. Hospital acquired pneumonia and cardiovascular complications increase the risk of death among STEMI patients. Conclusion: A LGI > 1200 increased, just over nine times, the risk of IHnCVC development in STEMI patients.

A target enrichment probe set for resolving the flagellate land plant tree of life.

PREMISE: New sequencing technologies facilitate the generation of large-scale molecular data sets for constructing the plant tree of life. We describe a new probe set for target enrichment sequencing to generate nuclear sequence data to build phylogenetic trees with any flagellate land plants, including hornworts, liverworts, mosses, lycophytes, ferns, and all gymnosperms. METHODS: We leveraged existing transcriptome and genome sequence data to design the GoFlag 451 probes, a set of 56,989 probes for target enrichment sequencing of 451 exons that are found in 248 single-copy or low-copy nuclear genes across flagellate plant lineages. RESULTS: Our results indicate that target enrichment using the GoFlag451 probe set can provide large nuclear data sets that can be used to resolve relationships among both distantly and closely related taxa across the flagellate land plants. We also describe the GoFlag 408 probes, an optimized probe set covering 408 of the 451 exons from the GoFlag 451 probe set that is commercialized by RAPiD Genomics. CONCLUSIONS: A target enrichment approach using the new probe set provides a relatively low-cost solution to obtain large-scale nuclear sequence data for inferring phylogenetic relationships across flagellate land plants.

The hornworts: morphology, evolution and development.

Extant land plants consist of two deeply divergent groups, tracheophytes and bryophytes, which shared a common ancestor some 500 million years ago. While information about vascular plants and the two of the three lineages of bryophytes, the mosses and liverworts, is steadily accumulating, the biology of hornworts remains poorly explored. Yet, as the sister group to liverworts and mosses, hornworts are critical in understanding the evolution of key land plant traits. Until recently, there was no hornwort model species amenable to systematic experimental investigation, which hampered detailed insight into the molecular biology and genetics of this unique group of land plants. The emerging hornwort model species, Anthoceros agrestis, is instrumental in our efforts to better understand not only hornwort biology but also fundamental questions of land plant evolution. To this end, here we provide an overview of hornwort biology and current research on the model plant A. agrestis to highlight its potential in answering key questions of land plant biology and evolution.

Anthoceros genomes illuminate the origin of land plants and the unique biology of hornworts.

Hornworts comprise a bryophyte lineage that diverged from other extant land plants >400 million years ago and bears unique biological features, including a distinct sporophyte architecture, cyanobacterial symbiosis and a pyrenoid-based carbon-concentrating mechanism (CCM). Here, we provide three high-quality genomes of Anthoceros hornworts. Phylogenomic analyses place hornworts as a sister clade to liverworts plus mosses with high support. The Anthoceros genomes lack repeat-dense centromeres as well as whole-genome duplication, and contain a limited transcription factor repertoire. Several genes involved in angiosperm meristem and stomatal function are conserved in Anthoceros and upregulated during sporophyte development, suggesting possible homologies at the genetic level. We identified candidate genes involved in cyanobacterial symbiosis and found that LCIB, a Chlamydomonas CCM gene, is present in hornworts but absent in other plant lineages, implying a possible conserved role in CCM function. We anticipate that these hornwort genomes will serve as essential references for future hornwort research and comparative studies across land plants.

Organellomic data sets confirm a cryptic consensus on (unrooted) land-plant relationships and provide new insights into bryophyte molecular evolution.

PREMISE: Phylogenetic trees of bryophytes provide important evolutionary context for land plants. However, published inferences of overall embryophyte relationships vary considerably. We performed phylogenomic analyses of bryophytes and relatives using both mitochondrial and plastid gene sets, and investigated bryophyte plastome evolution. METHODS: We employed diverse likelihood-based analyses to infer large-scale bryophyte phylogeny for mitochondrial and plastid data sets. We tested for changes in purifying selection in plastid genes of a mycoheterotrophic liverwort (Aneura mirabilis) and a putatively mycoheterotrophic moss (Buxbaumia), and compared 15 bryophyte plastomes for major structural rearrangements. RESULTS: Overall land-plant relationships conflict across analyses, generally weakly. However, an underlying (unrooted) four-taxon tree is consistent across most analyses and published studies. Despite gene coverage patchiness, relationships within mosses, liverworts, and hornworts are largely congruent with previous studies, with plastid results generally better supported. Exclusion of RNA edit sites restores cases of unexpected non-monophyly to monophyly for Takakia and two hornwort genera. Relaxed purifying selection affects multiple plastid genes in mycoheterotrophic Aneura but not Buxbaumia. Plastid genome structure is nearly invariant across bryophytes, but the tufA locus, presumed lost in embryophytes, is unexpectedly retained in several mosses. CONCLUSIONS: A common unrooted tree underlies embryophyte phylogeny, [(liverworts, mosses), (hornworts, vascular plants)]; rooting inconsistency across studies likely reflects substantial distance to algal outgroups. Analyses combining genomic and transcriptomic data may be misled locally for heavily RNA-edited taxa. The Buxbaumia plastome lacks hallmarks of relaxed selection found in mycoheterotrophic Aneura. Autotrophic bryophyte plastomes, including Buxbaumia, hardly vary in overall structure.

Complete Genomes of Symbiotic Cyanobacteria Clarify the Evolution of Vanadium-Nitrogenase.

Plant endosymbiosis with nitrogen-fixing cyanobacteria has independently evolved in diverse plant lineages, offering a unique window to study the evolution and genetics of plant-microbe interaction. However, very few complete genomes exist for plant cyanobionts, and therefore little is known about their genomic and functional diversity. Here, we present four complete genomes of cyanobacteria isolated from bryophytes. Nanopore long-read sequencing allowed us to obtain circular contigs for all the main chromosomes and most of the plasmids. We found that despite having a low 16S rRNA sequence divergence, the four isolates exhibit considerable genome reorganizations and variation in gene content. Furthermore, three of the four isolates possess genes encoding vanadium (V)-nitrogenase (vnf), which is uncommon among diazotrophs and has not been previously reported in plant cyanobionts. In two cases, the vnf genes were found on plasmids, implying possible plasmid-mediated horizontal gene transfers. Comparative genomic analysis of vnf-containing cyanobacteria further identified a conserved gene cluster. Many genes in this cluster have not been functionally characterized and would be promising candidates for future studies to elucidate V-nitrogenase function and regulation.

Hornwort Stomata: Architecture and Fate Shared with 400-Million-Year-Old Fossil Plants without Leaves.

As one of the earliest plant groups to evolve stomata, hornworts are key to understanding the origin and function of stomata. Hornwort stomata are large and scattered on sporangia that grow from their bases and release spores at their tips. We present data from development and immunocytochemistry that identify a role for hornwort stomata that is correlated with sporangial and spore maturation. We measured guard cells across the genera with stomata to assess developmental changes in size and to analyze any correlation with genome size. Stomata form at the base of the sporophyte in the green region, where they develop differential wall thickenings, form a pore, and die. Guard cells collapse inwardly, increase in surface area, and remain perched over a substomatal cavity and network of intercellular spaces that is initially fluid filled. Following pore formation, the sporophyte dries from the outside inwardly and continues to do so after guard cells die and collapse. Spore tetrads develop in spore mother cell walls within a mucilaginous matrix, both of which progressively dry before sporophyte dehiscence. A lack of correlation between guard cell size and DNA content, lack of arabinans in cell walls, and perpetually open pores are consistent with the inactivity of hornwort stomata. Stomata are expendable in hornworts, as they have been lost twice in derived taxa. Guard cells and epidermal cells of hornworts show striking similarities with the earliest plant fossils. Our findings identify an architecture and fate of stomata in hornworts that is ancient and common to plants without sporophytic leaves.

Hornworts: An Overlooked Window into Carbon-Concentrating Mechanisms.

Hornworts are the only land plant lineage harboring a biophysical carbon-concentrating mechanism (CCM). Here, we argue that hornworts are a promising, yet currently overlooked, model system to study the evolution and genetic basis of CCMs. The results of such studies could have translational values toward engineering a CCM in crop plants.

World checklist of hornworts and liverworts.

A working checklist of accepted taxa worldwide is vital in achieving the goal of developing an online flora of all known plants by 2020 as part of the Global Strategy for Plant Conservation. We here present the first-ever worldwide checklist for liverworts (Marchantiophyta) and hornworts (Anthocerotophyta) that includes 7486 species in 398 genera representing 92 families from the two phyla. The checklist has far reaching implications and applications, including providing a valuable tool for taxonomists and systematists, analyzing phytogeographic and diversity patterns, aiding in the assessment of floristic and taxonomic knowledge, and identifying geographical gaps in our understanding of the global liverwort and hornwort flora. The checklist is derived from a working data set centralizing nomenclature, taxonomy and geography on a global scale. Prior to this effort a lack of centralization has been a major impediment for the study and analysis of species richness, conservation and systematic research at both regional and global scales. The success of this checklist, initiated in 2008, has been underpinned by its community approach involving taxonomic specialists working towards a consensus on taxonomy, nomenclature and distribution.

Phytochrome diversity in green plants and the origin of canonical plant phytochromes.

Phytochromes are red/far-red photoreceptors that play essential roles in diverse plant morphogenetic and physiological responses to light. Despite their functional significance, phytochrome diversity and evolution across photosynthetic eukaryotes remain poorly understood. Using newly available transcriptomic and genomic data we show that canonical plant phytochromes originated in a common ancestor of streptophytes (charophyte algae and land plants). Phytochromes in charophyte algae are structurally diverse, including canonical and non-canonical forms, whereas in land plants, phytochrome structure is highly conserved. Liverworts, hornworts and Selaginella apparently possess a single phytochrome, whereas independent gene duplications occurred within mosses, lycopods, ferns and seed plants, leading to diverse phytochrome families in these clades. Surprisingly, the phytochrome portions of algal and land plant neochromes, a chimera of phytochrome and phototropin, appear to share a common origin. Our results reveal novel phytochrome clades and establish the basis for understanding phytochrome functional evolution in land plants and their algal relatives.

Phylotranscriptomic analysis of the origin and early diversification of land plants.

Reconstructing the origin and evolution of land plants and their algal relatives is a fundamental problem in plant phylogenetics, and is essential for understanding how critical adaptations arose, including the embryo, vascular tissue, seeds, and flowers. Despite advances in molecular systematics, some hypotheses of relationships remain weakly resolved. Inferring deep phylogenies with bouts of rapid diversification can be problematic; however, genome-scale data should significantly increase the number of informative characters for analyses. Recent phylogenomic reconstructions focused on the major divergences of plants have resulted in promising but inconsistent results. One limitation is sparse taxon sampling, likely resulting from the difficulty and cost of data generation. To address this limitation, transcriptome data for 92 streptophyte taxa were generated and analyzed along with 11 published plant genome sequences. Phylogenetic reconstructions were conducted using up to 852 nuclear genes and 1,701,170 aligned sites. Sixty-nine analyses were performed to test the robustness of phylogenetic inferences to permutations of the data matrix or to phylogenetic method, including supermatrix, supertree, and coalescent-based approaches, maximum-likelihood and Bayesian methods, partitioned and unpartitioned analyses, and amino acid versus DNA alignments. Among other results, we find robust support for a sister-group relationship between land plants and one group of streptophyte green algae, the Zygnematophyceae. Strong and robust support for a clade comprising liverworts and mosses is inconsistent with a widely accepted view of early land plant evolution, and suggests that phylogenetic hypotheses used to understand the evolution of fundamental plant traits should be reevaluated.

A review of molecular-clock calibrations and substitution rates in liverworts, mosses, and hornworts, and a timeframe for a taxonomically cleaned-up genus Nothoceros.

Absolute times from calibrated DNA phylogenies can be used to infer lineage diversification, the origin of new ecological niches, or the role of long distance dispersal in shaping current distribution patterns. Molecular-clock dating of non-vascular plants, however, has lagged behind flowering plant and animal dating. Here, we review dating studies that have focused on bryophytes with several goals in mind, (i) to facilitate cross-validation by comparing rates and times obtained so far; (ii) to summarize rates that have yielded plausible results and that could be used in future studies; and (iii) to calibrate a species-level phylogeny for Nothoceros, a model for plastid genome evolution in hornworts. Including the present work, there have been 18 molecular clock studies of liverworts, mosses, or hornworts, the majority with fossil calibrations, a few with geological calibrations or dated with previously published plastid substitution rates. Over half the studies cross-validated inferred divergence times by using alternative calibration approaches. Plastid substitution rates inferred for "bryophytes" are in line with those found in angiosperm studies, implying that bryophyte clock models can be calibrated either with published substitution rates or with fossils, with the two approaches testing and cross-validating each other. Our phylogeny of Nothoceros is based on 44 accessions representing all suspected species and a matrix of six markers of nuclear, plastid, and mitochondrial DNA. The results show that Nothoceros comprises 10 species, nine in the Americas and one in New Zealand (N. giganteus), with the divergence between the New Zealand species and its Chilean sister species dated to the Miocene and therefore due to long-distance dispersal. Based on the new tree, we formally transfer two species of Megaceros into Nothoceros, resulting in the new combinations N. minarum (Nees) J.C. Villarreal and N. schizophyllus (Gottsche ex Steph.) J.C. Villarreal, and we also newly synonymize eight names described in Megaceros.

Horizontal transfer of an adaptive chimeric photoreceptor from bryophytes to ferns.

Ferns are well known for their shade-dwelling habits. Their ability to thrive under low-light conditions has been linked to the evolution of a novel chimeric photoreceptor--neochrome--that fuses red-sensing phytochrome and blue-sensing phototropin modules into a single gene, thereby optimizing phototropic responses. Despite being implicated in facilitating the diversification of modern ferns, the origin of neochrome has remained a mystery. We present evidence for neochrome in hornworts (a bryophyte lineage) and demonstrate that ferns acquired neochrome from hornworts via horizontal gene transfer (HGT). Fern neochromes are nested within hornwort neochromes in our large-scale phylogenetic reconstructions of phototropin and phytochrome gene families. Divergence date estimates further support the HGT hypothesis, with fern and hornwort neochromes diverging 179 Mya, long after the split between the two plant lineages (at least 400 Mya). By analyzing the draft genome of the hornwort Anthoceros punctatus, we also discovered a previously unidentified phototropin gene that likely represents the ancestral lineage of the neochrome phototropin module. Thus, a neochrome originating in hornworts was transferred horizontally to ferns, where it may have played a significant role in the diversification of modern ferns.

Data access for the 1,000 Plants (1KP) project.

The 1,000 plants (1KP) project is an international multi-disciplinary consortium that has generated transcriptome data from over 1,000 plant species, with exemplars for all of the major lineages across the Viridiplantae (green plants) clade. Here, we describe how to access the data used in a phylogenomics analysis of the first 85 species, and how to visualize our gene and species trees. Users can develop computational pipelines to analyse these data, in conjunction with data of their own that they can upload. Computationally estimated protein-protein interactions and biochemical pathways can be visualized at another site. Finally, we comment on our future plans and how they fit within this scalable system for the dissemination, visualization, and analysis of large multi-species data sets.

Correlates of monoicy and dioicy in hornworts, the apparent sister group to vascular plants.

BACKGROUND: Whether male and female gametes are produced by single or separate individuals shapes plant mating and hence patterns of genetic diversity among and within populations. Haploid-dominant plants ("bryophytes": liverworts, mosses and hornworts) can have unisexual (dioicous) or bisexual (monoicous) gametophytes, and today, 68% of liverwort species, 57% of moss species, and 40% of hornwort species are dioicous. The transitions between the two sexual systems and possible correlations with other traits have been studied in liverworts and mosses, but not hornworts. Here we use a phylogeny for 98 of the 200 species of hornworts, the sister group to vascular plants, representing roughly equal proportions of all monoicous and all dioicous species, to test whether transitions in sexual systems are predominantly from monoicy to dioicy as might be expected based on studies of mosses. We further investigate possible correlations between sexual system and spore size, antheridium number, ploidy level, and diversification rate, with character selection partly based on findings in mosses and liverworts. RESULTS: Hornworts underwent numerous transitions between monoicy and dioicy. The transition rate from dioicy to monoicy was 2× higher than in the opposite direction, but monoicous groups have higher extinction rates; diversification rates do not correlate with sexual system. A correlation important in mosses, that between monoicy and polyploidy, apparently plays a small role: of 20 species with chromosome counts, only one is polyploid, the monoicous Anthoceros punctatus. A contingency test revealed that transitions to dioicy were more likely in species with small spores, supporting the hypothesis that small but numerous spores may be advantageous for dioicous species that depend on dense carpets of gametophytes for reproductive assurance. However, we found no evidence for increased antheridium-per-chamber numbers in dioicous species. CONCLUSIONS: Sexual systems in hornworts are labile, and the higher number of extant monoicous species (60%) may be largely due to frequent transitions to monoicy.

Genome size increases in recently diverged hornwort clades.

As our knowledge of plant genome size estimates continues to grow, one group has continually been neglected: the hornworts. Hornworts (Anthocerotophyta) have been traditionally grouped with liverworts and mosses because they share a haploid dominant life cycle; however, recent molecular studies place hornworts as the sister lineage to extant tracheophytes. Given the scarcity of information regarding the DNA content of hornworts, our objective was to estimate the 1C-value for a range of hornwort species within a phylogenetic context. Using flow cytometry, we estimated genome size for 36 samples representing 24 species. This accounts for roughly 10% of known hornwort species. Haploid genome sizes (1C-value) ranged from 160 Mbp or 0.16 pg (Leiosporoceros dussii) to 719 Mbp or 0.73 pg (Nothoceros endiviifolius). The average 1C-value was 261 ± 104 Mbp (0.27 ± 0.11 pg). Ancestral reconstruction of genome size on a hornwort phylogeny suggests a small ancestral genome size and revealed increases in genome size in the most recently divergent clades. Much more work is needed to understand DNA content variation in this phylogenetically important group, but this work has significantly increased our knowledge of genome size variation in hornworts.

Fungal symbioses in hornworts: a chequered history.

Hornworts are considered the sister group to vascular plants, but their fungal associations remain largely unexplored. The ancestral symbiotic condition for all plants is, nonetheless, widely assumed to be arbuscular mycorrhizal with Glomeromycota fungi. Owing to a recent report of other fungi in some non-vascular plants, here we investigate the fungi associated with diverse hornworts worldwide, using electron microscopy and molecular phylogenetics. We found that both Glomeromycota and Mucoromycotina fungi can form symbioses with most hornworts, often simultaneously. This discovery indicates that ancient terrestrial plants relied on a wider and more versatile symbiotic repertoire than previously thought, and it highlights the so far unappreciated ecological and evolutionary role of Mucoromycotina fungi.

The plastid genome of the hornwort Nothoceros aenigmaticus (Dendrocerotaceae): phylogenetic signal in inverted repeat expansion, pseudogenization, and intron gain.

PREMISE OF THE STUDY: The previously sequenced plastome of the hornwort Anthoceros angustus differs from that of other bryophytes by an expanded inverted repeat (IR) and the presence of a type I intron in the 23S ribosomal RNA (rrn23) gene. We assembled the plastome of the hornwort Nothoceros aenigmaticus, contrasted its architecture to that of other bryophytes, and assessed the phylogenetic significance of genomic characters in hornwort evolution. • METHODS: The Nothoceros plastome was reconstructed from shotgun sequencing of genomic DNA. Comparison with the Anthoceros plastome revealed three structural differences. We sequenced these regions in taxa spanning the hornwort phylogeny. • KEY RESULTS: The Nothoceros plastome is colinear with other bryophyte plastomes, but differs from the Anthoceros plastome by several gene regions located within the IR in Anthoceros being in the large single-copy region in Nothoceros, by the rrn23 gene lacking an intron, and by the rpl2 being a pseudogene. Comparisons across the hornwort phylogeny indicate that the first two characters are restricted to Anthocerotaceae, while rpl2 pseudogenization diagnoses the sister lineage to Anthocerotaceae. • CONCLUSIONS: The Nothoceros plastome is structurally similar to that of most bryophytes. However, we identified more structural differences within hornworts than have been described within either the mosses or the liverworts. The distribution of the gene duplication involving the IR and an intron in the rrn23 gene are restricted to Anthocerotaceae. Occurrence of the intron and the conserved intron sequence between Anthoceros and distantly related chlorophyte algae may be due to horizontal gene transfer.

Evaluating methods for isolating total RNA and predicting the success of sequencing phylogenetically diverse plant transcriptomes.

Next-generation sequencing plays a central role in the characterization and quantification of transcriptomes. Although numerous metrics are purported to quantify the quality of RNA, there have been no large-scale empirical evaluations of the major determinants of sequencing success. We used a combination of existing and newly developed methods to isolate total RNA from 1115 samples from 695 plant species in 324 families, which represents >900 million years of phylogenetic diversity from green algae through flowering plants, including many plants of economic importance. We then sequenced 629 of these samples on Illumina GAIIx and HiSeq platforms and performed a large comparative analysis to identify predictors of RNA quality and the diversity of putative genes (scaffolds) expressed within samples. Tissue types (e.g., leaf vs. flower) varied in RNA quality, sequencing depth and the number of scaffolds. Tissue age also influenced RNA quality but not the number of scaffolds ≥ 1000 bp. Overall, 36% of the variation in the number of scaffolds was explained by metrics of RNA integrity (RIN score), RNA purity (OD 260/230), sequencing platform (GAIIx vs HiSeq) and the amount of total RNA used for sequencing. However, our results show that the most commonly used measures of RNA quality (e.g., RIN) are weak predictors of the number of scaffolds because Illumina sequencing is robust to variation in RNA quality. These results provide novel insight into the methods that are most important in isolating high quality RNA for sequencing and assembling plant transcriptomes. The methods and recommendations provided here could increase the efficiency and decrease the cost of RNA sequencing for individual labs and genome centers.