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Given the 2,400-fold range of genome sizes (0.06–148.9 Gbp (gigabase pair)) of seed plants (angiosperms and gymnosperms) with a broadly similar gene content (amounting to approximately 0.03 Gbp), the repeat-sequence content of the genome might be expected to increase with genome size, resulting in the largest genomes consisting almost entirely of repetitive sequences. Here we test this prediction, using the same bioinformatic approach for 101 species to ensure consistency in what constitutes a repeat. We reveal a fundamental change in repeat turnover in genomes above around 10 Gbp, such that species with the largest genomes are only about 55% repetitive. Given that genome size influences many plant traits, habits and life strategies, this fundamental shift in repeat dynamics is likely to affect the evolutionary trajectory of species lineages., We thank Natural Environment Research Council (NE/G020256/1), the Czech Academy of Sciences (RVO:60077344) and Ramón y Cajal Fellowship (RYC-2017-2274) funded by the Ministerio de Ciencia y Tecnología (Gobierno de España) for support. We also thank Natural Environment Research Council for funding a studentship to S.D. and the China Scholarship Council for funding W.W., Main Methods Data availability Code availability References Acknowledgements Author information Ethics declarations Additional information Extended data Supplementary information Rights and permissions About this article Further reading

Societal Impact Statement For millennia, humans have used plants and fungi, as foods, fuels, fibers, and medicines; and have developed techniques for improving their usefulness to our species, mostly through selection of desirable traits. With human populations forecast to rise, the availability of arable land likely to fall amid climate change and increasing urbanization, and modern communications technologies accelerating the dispersal of pathogens, further improvement is urgently needed. However, ensuring long‐term resilience involves conservation of existing genetic diversity in addition to selection. New technologies, particularly those based on molecular biology, are increasingly driving conservation and improvement strategies. Summary Humans use plants and fungi for a wide range of purposes and, over millennia, have improved wild species by selecting for and combining genetic variation. Improvements in DNA sequencing technologies have enhanced our capacity to identify and manipulate genetic diversity, increasing the range of variation that can be utilized, and accelerating the breeding cycle to reduce the time taken to develop and put new varieties to use. Most recently, the CRISPR/Cas9 gene editing technology has greatly increased our capacity to directly introduce novel genetic variants without unwanted associated material. Moreover, increased knowledge of metabolic pathways resulting from genomic analysis can be used to design new varieties with desired properties with increased precision. Selecting for, or engineering, desirable variants has increased the usefulness of plants and fungi to humans, but at the cost of reducing their genetic diversity, decreasing their resilience and reducing the stock of variation available for future use. Conservation of genetic biodiversity is thus an essential counterpart of crop improvement and is essential to ensure that crop species retain resilience to emerging threats. Conservation efforts are focused on orphan crops, wild relatives of crop species, and landraces; in and exsitu efforts are complementary. Informatic approaches can inform use of these materials in breeding programmes even in the absence of genomic information. The application of some of these approaches may be restricted by ethical, legal, or organizational obstacles. If these can be overcome, there is great potential to unlock previously untapped reservoirs of biodiversity for human benefit.

Recent studies show that molecular convergence plays an unexpectedly common role in the evolution of convergent phenotypes. We exploited this phenomenon to find candidate loci underlying resistance to the emerald ash borer (EAB, Agrilus planipennis), the United States’ most costly invasive forest insect to date, within the pan-genome of ash trees (the genus Fraxinus). We show that EAB-resistant taxa occur within three independent phylogenetic lineages. In genomes from these resistant lineages, we detect 53 genes with evidence of convergent amino acid evolution. Gene-tree reconstruction indicates that, for 48 of these candidates, the convergent amino acids are more likely to have arisen via independent evolution than by another process such as hybridization or incomplete lineage sorting. Seven of the candidate genes have putative roles connected to the phenylpropanoid biosynthesis pathway and 17 relate to herbivore recognition, defence signalling or programmed cell death. Evidence for loss-of-function mutations among these candidates is more frequent in susceptible species than in resistant ones. Our results on evolutionary relationships, variability in resistance, and candidate genes for defence response within the ash genus could inform breeding for EAB resistance, facilitating ecological restoration in areas invaded by this beetle. By assembling the genomes of 22 Fraxinus species and conducting comparisons including further species, the authors identify candidate loci for emerald ash borer resistance that have evolved convergently.

Societal Impact Statement The current ash dieback epidemic in Europe caused by Hymenoscyphus fraxineus poses a key question to policy makers: whether or not to commit time and resources to the initiation of a breeding programme for the development of more resistant ash, as a long‐term policy of adaptation to the epidemic. Here we review current evidence on the potential viability of such a programme, from a biological perspective. We conclude that a breeding programme for ash aimed at resistance to current strains of H. fraxineus in the British Isles is biologically feasible. Summary To evaluate the viability and feasibility of a future breeding programme to produce trees resistant to an emerging pest or pathogen, it is helpful to ask the following questions: How much variation in resistance exists in tree populations? To what extent is this resistance heritable? How many genetic loci are involved? What level of resistance is found in other species of the same genus? Here, we survey current knowledge of these issues in relation to the degree of resistance of European ash (Fraxinus excelsior) to H. fraxineus, the fungus causing ash dieback (ADB). Several studies have found a low frequency of heritable resistance in F. excelsior populations, which seems to be determined by many genetic loci. This suggests that a breeding programme is viable and that natural selection may also increase the mean resistance of populations over time. More research is needed on the genetic basis of resistance to ADB to understand how quickly natural selection can operate in woodlands and what acceleration may be possible in breeding programmes, including via use of genetic markers. Hybrid breeding programmes may also be a possibility, as some ash species appear to be more resistant to ADB than is F. excelsior, but more research is needed on this issue. We do not yet know if it will be possible to breed F. excelsior to have resistance to both ADB and the emerging threat of emerald ash borer. We recommend short‐term mitigation measures for the ADB epidemic and future research directions.

Numerous plant genera have a history including frequent hybridisation and polyploidisation (allopolyploidisation), which means that their phylogeny is a network of reticulate evolution that cannot be accurately depicted as a bifurcating tree with a single tip per species. The genus Betula, which contains many ecologically important tree species, is a case in point. We generated genome-wide sequence reads for 27 diploid and 36 polyploid Betula species or subspecies using restriction site associated DNA (RAD) sequences. These reads were assembled into contigs with a mean length of 675 bp. We reconstructed the evolutionary relationships among diploid Betula species using both supermatrix (concatenation) and species tree methods. We identified the closest diploid relatives of the polyploids according to the relative rates at which reads from polyploids mapped to contigs from different diploid species within a concatenated reference sequence. By mapping reads from allopolyploids to their different putative diploid relatives we assembled contigs from the putative sub-genomes of allopolyploid taxa. We used these to build new phylogenies that included allopolyploid sub-genomes as separate tips. This approach yielded a highly evidenced phylogenetic hypothesis for the genus Betula, including the complex reticulate origins of the majority of its polyploid taxa. Our phylogeny divides the genus into two well supported clades, which, interestingly, differ in their seed-wing morphology. We therefore propose to split Betula into two subgenera.

Recent advances have highlighted the ubiquity of whole-genome duplication (polyploidy) in angiosperms, although subsequent genome size change and diploidization (returning to a diploid-like condition) are poorly understood. An excellent system to assess these processes is provided by Nicotiana section Repandae, which arose via allopolyploidy (approximately 5 million years ago) involving relatives of Nicotiana sylvestris and Nicotiana obtusifolia. Subsequent speciation in Repandae has resulted in allotetraploids with divergent genome sizes, including Nicotiana repanda and Nicotiana nudicaulis studied here, which have an estimated 23.6% genome expansion and 19.2% genome contraction from the early polyploid, respectively. Graph-based clustering of next-generation sequence data enabled assessment of the global genome composition of these allotetraploids and their diploid progenitors. Unexpectedly, in both allotetraploids, over 85% of sequence clusters (repetitive DNA families) had a lower abundance than predicted from their diploid relatives; a trend seen particularly in low-copy repeats. The loss of high-copy sequences predominantly accounts for the genome downsizing in N. nudicaulis. In contrast, N. repanda shows expansion of clusters already inherited in high copy number (mostly chromovirus-like Ty3/Gypsy retroelements and some low-complexity sequences), leading to much of the genome upsizing predicted. We suggest that the differential dynamics of low- and high-copy sequences reveal two genomic processes that occur subsequent to allopolyploidy. The loss of low-copy sequences, common to both allopolyploids, may reflect genome diploidization, a process that also involves loss of duplicate copies of genes and upstream regulators. In contrast, genome size divergence between allopolyploids is manifested through differential accumulation and/or deletion of high-copy-number sequences.

Genome sequencing has been restricted to species with a small genome size. With the advent of second- and third-generation sequencing technologies, the potential to sequence genomes of all sizes is becoming a reality. As the field of whole genome sequencing has developed, there has been a growing appreciation of the need to better represent the major lineages of the plant tree of life, rather than just those that contain economically important taxa. We argue that as well as accounting for phylogenetic diversity when selecting species to analyse, in order to gain a comprehensive understanding of genome evolution, large-scale sequencing projects also need to reflect the diversity of genome sizes in plants. In this article we briefly outline evidence from the literature to support this view.

Se evaluaron seis regiones de ADN como posibles códigos de barras de ADN para Hongos, el segundo reino más grande de vida eucariota, por un consorcio multinacional y de laboratorios múltiples. La región de la mitocondria se utilizó la subunidad 1 de la citocromo c oxidasa como código de barras animal excluido como un marcador potencial, porque es difícil de amplificar en hongos, a menudo incluye intrones grandes y puede ser insuficientemente variable. Tres subunidades del cistrón de ARN ribosómico nuclear fueron en comparación con las regiones de tres genes de codificación proteica representativos (la subunidad más grande de la ARN polimerasa II, la segunda más grande subunidad de la ARN polimerasa II y mantenimiento de minicromosomas proteína). Aunque las regiones de genes que codifican proteínas a menudo tenían un porcentaje más alto de identificación correcta en comparación con ribosomal marcadores, baja amplificación por PCR y éxito de secuenciación eliminados ellos como candidatos para un código de barras universal para hongos. Entre la regiones del cistrón ribosómico, el espaciador transcrito interno (ITS) tiene la mayor probabilidad de identificación exitosa para la gama más amplia de hongos, con la brecha de código de barras más claramente definida entre la variación inter e intraespecífica. El nuclear subunidad ribosómica grande, un marcador filogenético popular en ciertos grupos, tuvieron una resolución de especies superior en algunos grupos taxonómicos, como los primeros linajes divergentes y las levaduras ascomicetas, pero por lo demás era ligeramente inferior al ITS. El ribosomal nuclear La subunidad pequeña tiene una resolución pobre a nivel de especie en los hongos. SU será propuesto formalmente para su adopción como el código de barras fúngico principal marcador al Consorcio para el Código de Barras de la Vida, con la posibilidad de que se desarrollen códigos de barras suplementarios para grupos taxonómicos estrechamente circunscritos., Six DNA regions were evaluated as potential DNA barcodes for Fungi, the second largest kingdom of eukaryotic life, by a multinational, multilaboratory consortium. The region of the mitochondrial cytochrome c oxidase subunit 1 used as the animal barcode was excluded as a potential marker, because it is difficult to amplify in fungi, often includes large introns, and can be insufficiently variable. Three subunits from the nuclear ribosomal RNA cistron were compared together with regions of three representative proteincoding genes (largest subunit of RNA polymerase II, second largest subunit of RNA polymerase II, and minichromosome maintenance protein). Although the protein-coding gene regions often had a higher percent of correct identification compared with ribosomal markers, low PCR amplification and sequencing success eliminated them as candidates for a universal fungal barcode. Among the regions of the ribosomal cistron, the internal transcribed spacer (ITS) region has the highest probability of successful identification for the broadest range of fungi, with the most clearly defined barcode gap between inter- and intraspecific variation. The nuclear ribosomal large subunit, a popular phylogenetic marker in certain groups, had superior species resolution in some taxonomic groups, such as the early diverging lineages and the ascomycete yeasts, but was otherwise slightly inferior to the ITS. The nuclear ribosomal small subunit has poor species-level resolution in fungi. ITS will be formally proposed for adoption as the primary fungal barcode marker to the Consortium for the Barcode of Life, with the possibility that supplementary barcodes may be developed for particular narrowly circumscribed taxonomic groups.

Genome size in plants is characterised by its extraordinary range. Although it appears that the majority of plants have small genomes, in several lineages genome size has reached giant proportions. The recent advent of next-generation sequencing (NGS) methods has for the first time made detailed analysis of even the largest of plant genomes a possibility. In this review, we highlight investigations that have utilised NGS for the study of plants with large genomes, as well as describing ongoing work that aims to harness the power of these technologies to gain insights into their evolution. In addition, we emphasise some areas of research where the use of NGS has the potential to generate significant advances in our current understanding of how plant genomes evolve. Finally, we discuss some of the future developments in sequencing technology that may further improve our ability to explore the content and evolutionary dynamics of the very largest genomes.

DNA barcoding has the possibility for application in many fields, ranging from biological inventory and species discov ery to forensics and quality control. Barcoding could have great utility within the aquatic angiosperm family Podostemaceae, as these plants can be extremely difficult to identify on the basis of morphological characters alone, but are important components of tropical rivers and face threats from increased water pollution. Several potential barcodes for land plants have been suggested, including two 3-locus combinations proposed by the Plant Working Group of the Consortium for the Barcode of Life. These loci, matK, rpoB, rpoCl, and trnH-psbA, were tested for their potential utility in African members of the Podostemaceae. Our results show that although the rpoCl region performs best in terms of universality of amplification and sequencing, the superior resolving power of matK makes it the most promising of the regions for use in barcoding. In the light of these results, we discuss the potential to apply barcoding to Podostemaceae of Africa and make suggestions for the directions of future work to exploit the full potential of this new technology in this group of plants.

Ford, C. S., Ayres, K. L., Toomey, N., Haider, N., Stahl, J. V., Kelly, L. J., Wikstrom, N., Hollingsworth, P. M., Duff, R. J., Hoot, S. B., Cowan, R. S., Chase, M. W., Wilkinson, M. J. (2009). Selection of candidate coding DNA barcoding regions for use on land plants. Botanical Journal of the Linnean Society, 159, (1), 1-11. Sponsorship: Alfred P. Sloan Foundation Gordon and Betty Moore Foundation IMPF: 00.98 RONO: 00

Chase, M. W., Cowan, R. S., Hollingsworth, P. M., van den Berg, C., Madrinan, S., Petersen, G., Seberg, O., Jorgsensen, T., Cameron, K. M., Carine, M., Pedersen, N., Hedderson, T. A. J., Conrad, F., Salazar, G. A., Richardson, J. E., Hollingsworth, M. L., Barraclough, T. G., Kelly, L., Wilkinson, M. (2007). A proposal for a standardised protocol to barcode all land plants. Taxon, 56, (2), 295-299.

Endogenous pararetroviral sequences are the most commonly found virus sequences integrated into angiosperm genomes. We describe an endogenous pararetrovirus (EPRV) repeat in Fritillaria imperialis, a species that is under study as a result of its exceptionally large genome (1C = 42 096 Mbp, approximately 240 times bigger than Arabidopsis thaliana). The repeat (FriEPRV) was identified from Illumina reads using the RepeatExplorer pipeline, and exists in a complex genomic organization at the centromere of most, or all, chromosomes. The repeat was reconstructed into three consensus sequences that formed three interconnected loops, one of which carries sequence motifs expected of an EPRV (including the gag and pol domains). FriEPRV shows sequence similarity to members of the Caulimoviridae pararetrovirus family, with phylogenetic analysis indicating a close relationship to Petuvirus. It is possible that no complete EPRV sequence exists, although our data suggest an abundance that exceeds the genome size of Arabidopsis. Analysis of single nucleotide polymorphisms revealed elevated levels of C→T and G→A transitions, consistent with deamination of methylated cytosine. Bisulphite sequencing revealed high levels of methylation at CG and CHG motifs (up to 100%), and 15-20% methylation, on average, at CHH motifs. FriEPRV's centromeric location may suggest targeted insertion, perhaps associated with meiotic drive. We observed an abundance of 24 nt small RNAs that specifically target FriEPRV, potentially providing a signature of RNA-dependent DNA methylation. Such signatures of epigenetic regulation suggest that the huge genome of F. imperialis has not arisen as a consequence of a catastrophic breakdown in the regulation of repeat amplification.

Nicotiana (Solanaceae) provides an ideal system for understanding polyploidization, a pervasive and powerful evolutionary force in plants, as this genus contains several groups of allotetraploids that formed at different times from different diploid progenitors. However, the parental lineages of the largest group of allotetraploids, Nicotiana section Suaveolentes, have been problematic to identify. Using data from four regions of three low-copy nuclear genes, nuclear ribosomal DNA, and regions of the plastid genome, we have reconstructed the evolutionary origin of sect. Suaveolentes and identified the most likely diploid progenitors by using a combination of gene trees and network approaches to uncover the most strongly supported evidence of species relationships. Our analyses best support a scenario where a member of the sect. Sylvestres lineage acted as the paternal progenitor and a member of either sect. Petunioides or sect. Noctiflorae that also contained introgressed DNA from the other, or a hypothetical hybrid species between these two sections, was the maternal progenitor. Nicotiana exemplifies many of the factors that can complicate the reconstruction of polyploid evolutionary history and highlights how reticulate evolution at the diploid level can add even greater complexity to allopolyploid genomes.

The number of older illiterate adults seeking therapy for acquired communication disorders is increasing. Communication bias may place health‐care professionals at risk for providing appropriate services to these patients. There is a need to identify illiterate patients, and to study the impact of current diagnostic and rehabilitative procedures on them. Possible effects of illiteracy on hearing‐impaired, aphasic, and laryngectomy patients are discussed, and preliminary suggestions for providing comprehensive rehabilitative services to them are provided.