Advances in genomic characterization of Urochloa humidicola: exploring polyploid inheritance and apomixis.

KEY MESSAGE: We present the highest-density genetic map for the hexaploid Urochloa humidicola. SNP markers expose genetic organization, reproduction, and species origin, aiding polyploid and tropical forage research. Tropical forage grasses are an important food source for animal feeding, with Urochloa humidicola, also known as Koronivia grass, being one of the main pasture grasses for poorly drained soils in the tropics. However, genetic and genomic resources for this species are lacking due to its genomic complexity, including high heterozygosity, evidence of segmental allopolyploidy, and reproduction by apomixis. These complexities hinder the application of marker-assisted selection (MAS) in breeding programs. Here, we developed the highest-density linkage map currently available for the hexaploid tropical forage grass U. humidicola. This map was constructed using a biparental F1 population generated from a cross between the female parent H031 (CIAT 26146), the only known sexual genotype for the species, and the apomictic male parent H016 (BRS cv. Tupi). The linkage analysis included 4873 single nucleotide polymorphism (SNP) markers with allele dosage information. It allowed mapping of the ASGR locus and apospory phenotype to linkage group 3, in a region syntenic with chromosome 3 of Urochloa ruziziensis and chromosome 1 of Setaria italica. We also identified hexaploid haplotypes for all individuals, assessed the meiotic configuration, and estimated the level of preferential pairing in parents during the meiotic process, which revealed the autopolyploid origin of sexual H031 in contrast to apomictic H016, which presented allopolyploid behavior in preferential pairing analysis. These results provide new information regarding the genetic organization, mode of reproduction, and allopolyploid origin of U. humidicola, potential SNPs markers associated with apomixis for MAS and resources for research on polyploids and tropical forage grasses.

An exonuclease V homologue is expressed predominantly during early megasporogenesis in apomictic Brachiaria brizantha.

MAIN CONCLUSION: An exonuclease V homologue from apomictic Brachiaria brizantha is expressed and localized in nucellar cells at key moments when these cells differentiate to give rise to unreduced gametophytes. Brachiaria is a genus of forage grasses with economical and agricultural importance to Brazil. Brachiaria reproduces by aposporic apomixis, in which unreduced embryo sacs, derived from nucellar cells, other than the megaspore mother cell (MMC), are formed. The unreduced embryo sacs produce an embryo without fertilization resulting in clones of the mother plant. Comparative gene expression analysis in ovaries of sexual and apomictic Brachiaria spp. revealed a sequence from B. brizantha that showed a distinct pattern of expression in ovaries of sexual and apomictic plants. In this work, we describe a gene named BbrizExoV with strong identity to exonuclease V (Exo V) genes from other grasses. Sequence analysis in signal prediction tools showed that BbrizExoV might have dual localization, depending on the translation point. A longer form to the nucleus and a shorter form which would be directed to the chloroplast. This is also the case for monocot sequences analyzed from other species. The long form of BbrizExoV protein localizes to the nucleus of onion epidermal cells. Analysis of ExoV proteins from dicot species, with exception of Arabidopsis thaliana ExoVL protein, showed only one localization. Using a template-based AlphaFold 2 modelling approach the structure of BbrizExoV in complex with metal and ssDNA was predicted based on the holo structure of the human counterpart. Features predicted to define ssDNA binding but a lack of sequence specificity are shared between the human enzyme and BbrizExoV. Expression analyses indicated the precise site and timing of transcript accumulation during ovule development, which coincides with the differentiation of nucelar cells to form the typical aposporic four-celled unreduced gametophyte. A putative function for this protein is proposed based on its homology and expression pattern.

Reproductive and Agronomic Characterization of Novel Apomictic Hybrids of Paspalum (Poaceae).

The tetraploid germplasm of Paspalum contains a large diversity that can be used to generate better forages. The objective was to evaluate a group of Paspalum notatum and Paspalum simplex apomictic hybrids for a set of agronomic traits and apomixis expressivity. Forage yield, cold tolerance, winter regrowth, and seed yield were evaluated. The expressivity of apomixis was evaluated in P. simplex hybrids by flow cytometry. Progeny testing with molecular markers was used to determine the genotypic variability in the progeny. Differences within P. notatum and P. simplex hybrids were observed for all traits, and some of them were superior in comparison with the controls. The accumulated forage yield during three years was 988 g m-2 in the P. notatum hybrids, whereas, in P. simplex, the average forage yield per harvest (40 days of regrowth) was 180 g m-2. In P. simplex, the apomixis expressivity varied between 0 and 100%, and 65% of the hybrids showed high apomixis expressivity (superior to 70%). The genotypic mean homogeneity in the progeny was 76% and 85% in P. notatum and P. simplex, respectively. The generation of hybrids with high apomixis expressivity that combine good agronomic performance and homogeneity in the offspring is possible in tetraploid P. notatum and P. simplex.

Phylogeography of Eriotheca species complex: insights into the origin and range expansion of apomictic and polyploid trees in Neotropical Savannas.

Polyploidy and whole genome duplication are major evolutionary drivers in plants. Climate variations during the Pleistocene have influenced distribution and range expansion worldwide. Similar trends have been reported for Cerrado plants, but no attempt has been made to link phylogeography with ploidy and breeding changes. Thus, we aimed to (i) assess ploidy and genome size of Eriotheca estevesiae Carv.-Sobr., and compare it with E. pubescens (Mart.) Schott & Endl. (Both included into the Eriotheca Stellate Trichome Species Complex - ESTSC). (ii) Subsequently, we investigated their phylogeography to see whether genetic structure and range expansion trends were similar to those previously described for the Cerrado biome. Finally (iii), we discuss whether ESTSC phylogeographic patterns could be associated with geographic parthenogenesis processes. Common cytogenetic techniques and flow cytometry were used to confirm chromosome number and genome size of E. estevesiae. We used three cpDNA regions to analyse 14 ESTSC Cerrado populations, for which we also obtained ploidy level and breeding information. We investigated haplotype diversity, population structure and tested neutrality, aiming to reconstruct phylogeographic scenarios. We found three ploidy levels and eight cpDNA haplotypes in ESTSC, one shared by most populations. Haplotype and ploidy distribution corroborated that E. pubescens, the widely distributed polyploid and apomictic species, may have originated from northern diploid and probably sexual E. estevesiae. Matrilinear cpDNA links support the idea that apomixis and polyploidy in ESTSC may have allowed range expansion during the Pleistocene, in a process analogous to the geographic parthenogenesis described elsewhere.

Clone worth? Genetic diversity in obligate apomictic Miconia albicans (Melastomataceae).

Apomixis is the asexual production of seeds by plants and, in theory, would render low genetic diversity and even clonal lineages. However, recent studies have shown otherwise, although is not always clear where the genetic diversity of obligate apomicts comes from. We evaluated the genetic diversity among sister seedlings of M. albicans, an obligate apomictic species in Cerrado, Neotropical Savannas in Central Brazil. A total of 50 seedlings from five individuals were analysed using ISSR primers. We obtained 107 fragments, all with good resolution, consistently observed and replicable. The percentage of polymorphic loci ranged from 28.04% to 33.64% and Shannon's information index (I) averaged 0.173. The expected heterozygosity (He) averaged 0.117, similar to the observed for populations of M. albicans and other selfed species. Only two seedlings showed the same genotype (possible clones), but most differed at least for five loci. Most of variance was among progenies (62%), but we found that 38% was within progenies. Genetic distances separated the progenies in two groups, and analogous analyses between individuals reconstructed the original progenies clustering. The results confirmed a relatively high genetic diversity among sister seedling of this obligatory apomictic plant and clones were rare. This diversity can be generated during development, probably by restitutional meiosis or other recombination processes. These differences may accumulate into lineages and populations well adapted to heterogenous Cerrado environment.

A study of the heterochronic sense/antisense RNA representation in florets of sexual and apomictic Paspalum notatum.

BACKGROUND: Apomixis, an asexual mode of plant reproduction, is a genetically heritable trait evolutionarily related to sexuality, which enables the fixation of heterozygous genetic combinations through the development of maternal seeds. Recently, reference floral transcriptomes were generated from sexual and apomictic biotypes of Paspalum notatum, one of the most well-known plant models for the study of apomixis. However, the transcriptome dynamics, the occurrence of apomixis vs. sexual expression heterochronicity across consecutive developmental steps and the orientation of transcription (sense/antisense) remain unexplored. RESULTS: We produced 24 Illumina TruSeq®/ Hiseq 1500 sense/antisense floral transcriptome libraries covering four developmental stages (premeiosis, meiosis, postmeiosis, and anthesis) in biological triplicates, from an obligate apomictic and a full sexual genotype. De novo assemblies with Trinity yielded 103,699 and 100,114 transcripts for the apomictic and sexual samples respectively. A global comparative analysis involving reads from all developmental stages revealed 19,352 differentially expressed sense transcripts, of which 13,205 (68%) and 6147 (32%) were up- and down-regulated in apomictic samples with respect to the sexual ones. Interestingly, 100 differentially expressed antisense transcripts were detected, 55 (55%) of them up- and 45 (45%) down-regulated in apomictic libraries. A stage-by-stage comparative analysis showed a higher number of differentially expressed candidates due to heterochronicity discrimination: the highest number of differential sense transcripts was detected at premeiosis (23,651), followed by meiosis (22,830), postmeiosis (19,100), and anthesis (17,962), while the highest number of differential antisense transcripts were detected at anthesis (495), followed by postmeiosis (164), meiosis (120) and premeiosis (115). Members of the AP2, ARF, MYB and WRKY transcription factor families, as well as the auxin, jasmonate and cytokinin plant hormone families appeared broadly deregulated. Moreover, the chronological expression profile of several well-characterized apomixis controllers was examined in detail. CONCLUSIONS: This work provides a quantitative sense/antisense gene expression catalogue covering several subsequent reproductive developmental stages from premeiosis to anthesis for apomictic and sexual P. notatum, with potential to reveal heterochronic expression between reproductive types and discover sense/antisense mediated regulation. We detected a contrasting transcriptional and hormonal control in apomixis and sexuality as well as specific sense/antisense modulation occurring at the onset of parthenogenesis.

How to Become an Apomixis Model: The Multifaceted Case of Paspalum.

In the past decades, the grasses of the Paspalum genus have emerged as a versatile model allowing evolutionary, genetic, molecular, and developmental studies on apomixis as well as successful breeding applications. The rise of such an archetypal system progressed through integrative phases, which were essential to draw conclusions based on solid standards. Here, we review the steps adopted in Paspalum to establish the current body of knowledge on apomixis and provide model breeding programs for other agronomically important apomictic crops. In particular, we discuss the need for previous detailed cytoembryological and cytogenetic germplasm characterization; the establishment of sexual and apomictic materials of identical ploidy level; the development of segregating populations useful for inheritance analysis, positional mapping, and epigenetic control studies; the development of omics data resources; the identification of key molecular pathways via comparative gene expression studies; the accurate molecular characterization of genomic loci governing apomixis; the in-depth functional analysis of selected candidate genes in apomictic and model species; the successful building of a sexual/apomictic combined breeding scheme.

Genes Modulating the Increase in Sexuality in the Facultative Diplosporous Grass Eragrostis curvula under Water Stress Conditions.

Eragrostis curvula presents mainly facultative genotypes that reproduce by diplosporous apomixis, retaining a percentage of sexual pistils that increase under drought and other stressful situations, indicating that some regulators activated by stress could be affecting the apomixis/sexual switch. Water stress experiments were performed in order to associate the increase in sexual embryo sacs with the differential expression of genes in a facultative apomictic cultivar using cytoembryology and RNA sequencing. The percentage of sexual embryo sacs increased from 4 to 24% and 501 out of the 201,011 transcripts were differentially expressed (DE) between control and stressed plants. DE transcripts were compared with previous transcriptomes where apomictic and sexual genotypes were contrasted. The results point as candidates to transcripts related to methylation, ubiquitination, hormone and signal transduction pathways, transcription regulation and cell wall biosynthesis, some acting as a general response to stress and some that are specific to the reproductive mode. We suggest that a DNA glycosylase EcROS1-like could be demethylating, thus de-repressing a gene or genes involved in the sexuality pathways. Many of the other DE transcripts could be part of a complex mechanism that regulates apomixis and sexuality in this grass, the ones in the intersection between control/stress and apo/sex being the strongest candidates.

Small RNA-seq reveals novel regulatory components for apomixis in Paspalum notatum.

BACKGROUND: Apomixis is considered an evolutionary deviation of the sexual reproductive pathway leading to the generation of clonal maternal progenies by seeds. Recent evidence from model and non-model species suggested that this trait could be modulated by epigenetic mechanisms involving small RNAs (sRNAs). Here we profiled floral sRNAs originated from apomictic and sexual Paspalum notatum genotypes in order to identify molecular pathways under epigenetic control that might be involved in the transition from sexuality to agamospermy. RESULTS: The mining of genes participating in sRNA-directed pathways from floral Paspalum transcriptomic resources showed these routes are functional during reproductive development, with several members differentially expressed in apomictic and sexual plants. Triplicate floral sRNA libraries derived from apomictic and a sexual genotypes were characterized by using high-throughput sequencing technology. EdgeR was apply to compare the number of sRNA reads between sexual and apomictic libraries that map over all Paspalum floral transcripts. A total of 1525 transcripts showed differential sRNA representation, including genes related to meiosis, plant hormone signaling, biomolecules transport, transcription control and cell cycle. Survey for miRNA precursors on transcriptome and genome references allowed the discovery of 124 entities, including 40 conserved and 8 novel ones. Fifty-six clusters were differentially represented in apomictic and sexual plants. All differentially expressed miRNAs were up-regulated in apomictic libraries but miR2275, which showed different family members with opposed representation. Examination of predicted miRNAs targets detected 374 potential candidates. Considering sRNA, miRNAs and target surveys together, 14 genes previously described as related with auxin metabolism, transport and signaling were detected, including AMINO ACID/AUXIN PERMEASE 15, IAA-AMIDO SYNTHETASE GH3-8, IAA30, miR160, miR167, miR164, miR319, ARF2, ARF8, ARF10, ARF12, AFB2, PROLIFERATING CELL FACTOR 6 and NITRATE TRANSPORTER 1.1. CONCLUSIONS: This work provides a comprehensive survey of the sRNA differential representation in flowers of sexual and apomictic Paspalum notatum plants. An integration of the small RNA profiling data presented here and previous transcriptomic information suggests that sRNA-mediated regulation of auxin pathways is pivotal in promoting apomixis. These results will underlie future functional characterization of the molecular components mediating the switch from sexuality to apomixis.

Translocation of a parthenogenesis gene candidate to an alternate carrier chromosome in apomictic Brachiaria humidicola.

BACKGROUND: The apomictic reproductive mode of Brachiaria (syn. Urochloa) forage species allows breeders to faithfully propagate heterozygous genotypes through seed over multiple generations. In Brachiaria, reproductive mode segregates as single dominant locus, the apospory-specific genomic region (ASGR). The AGSR has been mapped to an area of reduced recombination on Brachiaria decumbens chromosome 5. A primer pair designed within ASGR-BABY BOOM-like (BBML), the candidate gene for the parthenogenesis component of apomixis in Pennisetum squamulatum, was diagnostic for reproductive mode in the closely related species B. ruziziensis, B. brizantha, and B. decumbens. In this study, we used a mapping population of the distantly related commercial species B. humidicola to map the ASGR and test for conservation of ASGR-BBML sequences across Brachiaria species. RESULTS: Dense genetic maps were constructed for the maternal and paternal genomes of a hexaploid (2n = 6x = 36) B. humidicola F1 mapping population (n = 102) using genotyping-by-sequencing, simple sequence repeat, amplified fragment length polymorphism, and transcriptome derived single nucleotide polymorphism markers. Comparative genomics with Setaria italica provided confirmation for x = 6 as the base chromosome number of B. humidicola. High resolution molecular karyotyping indicated that the six homologous chromosomes of the sexual female parent paired at random, whereas preferential pairing of subgenomes was observed in the apomictic male parent. Furthermore, evidence for compensated aneuploidy was found in the apomictic parent, with only five homologous linkage groups identified for chromosome 5 and seven homologous linkage groups of chromosome 6. The ASGR mapped to B. humidicola chromosome 1, a region syntenic with chromosomes 1 and 7 of S. italica. The ASGR-BBML specific PCR product cosegregated with the ASGR in the F1 mapping population, despite its location on a different carrier chromosome than B. decumbens. CONCLUSIONS: The first dense molecular maps of B. humidicola provide strong support for cytogenetic evidence indicating a base chromosome number of six in this species. Furthermore, these results show conservation of the ASGR across the Paniceae in different chromosomal backgrounds and support postulation of the ASGR-BBML as candidate genes for the parthenogenesis component of apomixis.

Apomixis in flowering plants: Developmental and evolutionary considerations.

Apomixis refers to a set of reproductive mechanisms that invariably rely on avoiding meiotic reduction and fertilization of the egg cell to generate clonal seeds. After having long been considered a strictly asexual oddity leading to extinction, the integration of more than 100 years of embryological, genetic, molecular, and ecological research has revealed apomixis as a widely spread component of the dynamic processes that shape flowering plant evolution. Apomixis involves several flexible and versatile developmental pathways that can be combined within the ovule to produce offspring. Here we review the large body of classic and contemporaneous contributions that have addressed unreduced gamete formation, haploid induction, and parthenogenesis in flowering plants. We emphasize similarities and differences between sexual and apomictic reproduction, and highlight their implications for the evolutionary emergence of asexual reproduction through seeds. On the basis of these comparisons, we propose a model that associates the developmental origin of apomixis to a dynamic epigenetic landscape, in which environmental fluctuations reversibly influence female reproductive development through mechanisms of hybridization and polyploidization.

Heterochronic reproductive developmental processes between diploid and tetraploid cytotypes of Paspalum rufum.

BACKGROUND AND AIMS: Apomixis is an asexual reproductive mode via seeds that generate maternal clonal progenies. Although apomixis in grasses is mainly expressed at the polyploid level, some natural diploid genotypes of Paspalum rufum produce aposporous embryo sacs in relatively high proportions and are even able to complete apomixis under specific conditions. However, despite the potential for apomixis, sexuality prevails in diploids, and apomixis expression is repressed for an as yet undetermind reason. Apomixis is thought to derive from a deregulation of one or a few components of the sexual pathway that could be triggered by polyploidy and/or hybridization. The objectives of this work were to characterize and compare the reproductive development and the timing of apospory initial (AI) emergence between diploid genotypes with potential for apomixis and facultative apomictic tetraploid cytotypes of P. rufum. METHODS: Reproductive characterization was performed by cytoembryological observations of cleared ovaries and anthers during all reproductive development steps and by quantitative evaluation of the ovule growth parameters. KEY RESULTS: Cytoembryological observations showed that in diploids, both female and male reproductive development is equally synchronized, but in tetraploids, megasporogenesis and early megagametogenesis are delayed with respect to microsporogenesis and early microgametogenesis. This delay was also seen when ovary growth was taken as a reference parameter. The analysis of the onset of AIs revealed that they emerge during different developmental periods depending on the ploidy level. In diploids, the AIs appeared along with the tetrad (or triad) of female meiocytes, but in tetraploids they appeared earlier, at the time of the megaspore mother cell. In both cytotypes, AIs can be seen even during megagametogenesis. CONCLUSIONS: Overall observations reveal that female sexual reproductive development is delayed in tetraploids as compared with diploid genotypes, mainly at meiosis. In tetraploids, AIs appear at earlier sexual developmental stages than in diploids, and they accumulate up to the end of megasporogenesis. The longer extension of megasporogenesis in tetraploids could favour AI emergence and also apomixis success.

GID1 expression is associated with ovule development of sexual and apomictic plants.

KEY MESSAGE: BbrizGID1 is expressed in the nucellus of apomictic Brachiaria brizantha, previous to aposporous initial differentiation. AtGID1a overexpression triggers differentiation of Arabidopsis thaliana MMC-like cells, suggesting its involvement in ovule development. GIBBERELLIN-INSENSITIVE DWARF1 (GID1) is a gibberellin receptor previously identified in plants and associated with reproductive development, including ovule formation. In this work, we characterized the Brachiaria brizantha GID1 gene (BbrizGID1). BbrizGID1 showed up to 92% similarity to GID1-like gibberellin receptors of other plants of the Poaceae family and around 58% to GID1-like gibberellin receptors of Arabidopsis thaliana. BbrizGID1 was more expressed in ovaries at megasporogenesis than in ovaries at megagametogenesis of both sexual and apomictic plants. In ovules, BbrizGID1 transcripts were detected in the megaspore mother cell (MMC) of sexual and apomictic B. brizantha. Only in the apomictic plants, expression was also observed in the surrounding nucellar cells, a region in which aposporous initial cells differentiate to form the aposporic embryo sac. AtGID1a ectopic expression in Arabidopsis determines the formation of MMC-like cells in the nucellus, close to the MMC, that did not own MMC identity. Our results suggest that GID1 might be involved in the proper differentiation of a single MMC during ovule development and provide valuable information on the role of GID1 in sexual and apomictic reproduction.

Structure, target-specificity and expression of PN_LNC_N13, a long non-coding RNA differentially expressed in apomictic and sexual Paspalum notatum.

KEY MESSAGE: ncRNA PN_LNC_N13 shows contrasting expression in reproductive organs of sexual and apomictic Paspalum notatum genotypes. Apomictic plants set genetically maternal seeds whose embryos derive by parthenogenesis from unreduced egg cells, giving rise to clonal offspring. Several Paspalum notatum apomixis related genes were identified in prior work by comparative transcriptome analyses. Here, one of these candidates (namely N13) was characterized. N13 belongs to a Paspalum gene family including 30-60 members, of which at least eight are expressed at moderate levels in florets. The sequences of these genes show no functional ORFs, but include segments of different protein coding genes. Particularly, N13 shows partial identity to maize gene BT068773 (RESPONSE REGULATOR 6). Secondary structure predictions as well as mature miRNA and target cleavage detection suggested that N13 is not a miRNA precursor. Moreover, N13 family members produce abundant 24-nucleotide small RNAs along extensive parts of their sequences. Surveys in the GREENC and CANTATA databases indicated similarity with plant long non-coding RNAs (lncRNAs) involved in splicing regulation; consequently, N13 was renamed as PN_LNC_N13. The Paspalum BT068773 predicted ortholog (N13TAR) originates floral transcript variants shorter than the canonical maize isoform and with possible structural differences between the apomictic and sexual types. PN_LNC_N13 is expressed only in apomictic plants and displays quantitative representation variation across reproductive developmental stages. However, PN_LNC_N13-like homologs and/or its derived sRNAs showed overall a higher representation in ovules of sexual plants at late premeiosis. Our results suggest the existence of a whole family of N13-like lncRNAs possibly involved in splicing regulation, with some members characterized by differential activity across reproductive types.

Temporal and spatial expression of genes involved in DNA methylation during reproductive development of sexual and apomictic Eragrostis curvula.

Recent reports in model plant species have highlighted a role for DNA methylation pathways in the regulation of the somatic-to-reproductive transition in the ovule, suggesting that apomixis (asexual reproduction through seeds) likely relies on RdDM downregulation. Our aim was therefore to explore this hypothesis by characterizing genes involved in DNA methylation in the apomictic grass Eragrostis curvula. We explored floral transcriptomes to identify homologs of three candidate genes, for which mutations in Arabidopsis and maize mimic apomixis (AtAGO9/ZmAGO104, AtCMT3/ZmDMT102/ZmDMT105, and AtDDM1/ZmCHR106), and compared both their spatial and temporal expression patterns during reproduction in sexual and apomictic genotypes. Quantitative expression analyses revealed contrasting expression patterns for the three genes in apomictic vs sexual plants. In situ hybridization corroborated these results for two candidates, EcAGO104 and EcDMT102, and revealed an unexpected ectopic pattern for the AGO gene during germ line differentiation in apomicts. Although our data partially support previous results obtained in sexual plant models, they suggest that rather than an RdDM breakdown in the ovule, altered localization of AtAGO9/ZmAGO104 expression is required for achieving diplospory in E. curvula. The differences in the RdDM machinery acquired during plant evolution might have promoted the emergence of the numerous apomictic paths observed in plants.

Caracteres forrageiros de diferentes espécies do gênero Paspalum no Rio Grande do Sul: uma meta-análise / Forage characters of different Paspalum species in Rio Grande do Sul: a meta-analysis

The objective of this study was to evaluate, through meta-analysis, the forage characteristics of various species of the genus Paspalum and to use them to select the best ecotypes that can be used in artificial hybridization as parents and hybrids for pasture production and natural pasture recovery systems. Data were obtained from studies conducted by the Department of Forage Plants and Agrometeorology of the Universidade Federal do Rio Grande do Sul. Database comprised tests conducted with ecotypes and/or hybrids of Paspalum spp. in plots for evaluating total dry mass production, leaf dry mass production, and stem dry mass production by means of cuts. Total dry mass production, which included leaves and stems, differed between the ecotypes and hybrids. Hybrid H12 was the most divergent of all evaluated accessions. The greatest genetic divergence occurred due to dry mass production. Hybrids showed high total dry mass production, comprised mainly of leaves. Hybrid H12 and the accession of Paspalum lepton 28E were identified as the most dissimilar based on the generalized Mahalanobis distance using Tochers method. Total dry mass production is the characteristic that most contributed to the detection of genetic variability.(AU)

O objetivo deste trabalho foi avaliar, por meio de meta-análise, a variabilidade dos caracteres forrageiros de espécies do gênero Paspalum e utilizá-los para selecionar os melhores ecótipos para serem utilizados em hibridações artificiais como genitores e híbridos para serem empregados em sistemas de produção a pasto e recuperação de pastagens naturais. Os dados foram obtidos a partir ensaios, do Departamento de Plantas Forrageiras e Agrometeorologia da Universidade Federal do Rio Grande do Sul. A base de dados foi composta por ensaios conduzidos com ecótipos e/ou híbridos do gênero Paspalum, em parcelas, avaliando por meio de cortes a produção de massa seca total, de folhas e colmos. Houve diferença entre ecótipos/híbridos para produção de matéria seca total, de folhas e colmos. O híbrido H12 foi o mais divergente dos acessos avaliados. A maior divergência genética ocorreu devido à produção de massa seca. Os híbridos apresentam elevada produção de massa seca total, sendo esta composta principalmente por folhas. O método de Tocher utilizando a distância generalizada de Mahalanobis identifica o híbrido H12 e o acesso de Paspalum lepton 28E como os mais dissimilares. A produção de massa seca total é o caractere que mais contribui para a detecção da variabilidade genética.(AU)

Caracteres forrageiros de diferentes espécies do gênero Paspalum no Rio Grande do Sul: uma meta-análise / Forage characters of different Paspalum species in Rio Grande do Sul: a meta-analysis

The objective of this study was to evaluate, through meta-analysis, the forage characteristics of various species of the genus Paspalum and to use them to select the best ecotypes that can be used in artificial hybridization as parents and hybrids for pasture production and natural pasture recovery systems. Data were obtained from studies conducted by the Department of Forage Plants and Agrometeorology of the Universidade Federal do Rio Grande do Sul. Database comprised tests conducted with ecotypes and/or hybrids of Paspalum spp. in plots for evaluating total dry mass production, leaf dry mass production, and stem dry mass production by means of cuts. Total dry mass production, which included leaves and stems, differed between the ecotypes and hybrids. Hybrid H12 was the most divergent of all evaluated accessions. The greatest genetic divergence occurred due to dry mass production. Hybrids showed high total dry mass production, comprised mainly of leaves. Hybrid H12 and the accession of Paspalum lepton 28E were identified as the most dissimilar based on the generalized Mahalanobis distance using Tochers method. Total dry mass production is the characteristic that most contributed to the detection of genetic variability.

O objetivo deste trabalho foi avaliar, por meio de meta-análise, a variabilidade dos caracteres forrageiros de espécies do gênero Paspalum e utilizá-los para selecionar os melhores ecótipos para serem utilizados em hibridações artificiais como genitores e híbridos para serem empregados em sistemas de produção a pasto e recuperação de pastagens naturais. Os dados foram obtidos a partir ensaios, do Departamento de Plantas Forrageiras e Agrometeorologia da Universidade Federal do Rio Grande do Sul. A base de dados foi composta por ensaios conduzidos com ecótipos e/ou híbridos do gênero Paspalum, em parcelas, avaliando por meio de cortes a produção de massa seca total, de folhas e colmos. Houve diferença entre ecótipos/híbridos para produção de matéria seca total, de folhas e colmos. O híbrido H12 foi o mais divergente dos acessos avaliados. A maior divergência genética ocorreu devido à produção de massa seca. Os híbridos apresentam elevada produção de massa seca total, sendo esta composta principalmente por folhas. O método de Tocher utilizando a distância generalizada de Mahalanobis identifica o híbrido H12 e o acesso de Paspalum lepton 28E como os mais dissimilares. A produção de massa seca total é o caractere que mais contribui para a detecção da variabilidade genética.

Apomixis frequency under stress conditions in weeping lovegrass (Eragrostis curvula).

To overcome environmental stress, plants develop physiological responses that are triggered by genetic or epigenetic changes, some of which involve DNA methylation. It has been proposed that apomixis, the formation of asexual seeds without meiosis, occurs through the temporal or spatial deregulation of the sexual process mediated by genetic and epigenetic factors influenced by the environment. Here, we explored whether there was a link between the occurrence of apomixis and various factors that generate stress, including drought stress, in vitro culture, and intraspecific hybridization. For this purpose, we monitored the embryo sacs of different weeping lovegrass (Eragrostis curvula [Schrad.] Nees) genotypes after the plants were subjected to these stress conditions. Progeny tests based on molecular markers and genome methylation status were analyzed following the stress treatment. When grown in the greenhouse, the cultivar Tanganyika INTA generated less than 2% of its progeny by sexual reproduction. Plants of this cultivar subjected to different stresses showed an increase of sexual embryo sacs, demonstrating an increased expression of sexuality compared to control plants. Plants of the cv. Tanganyika USDA did not demonstrate the ability to generate sexual embryo sacs under any conditions and is therefore classified as a fully apomictic cultivar. We found that this change in the prevalence of sexuality was correlated with genetic and epigenetic changes analyzed by MSAP and AFLPs profiles. Our results demonstrate that different stress conditions can alter the expression of sexual reproduction in facultative tetraploid apomictic cultivars and when the stress stops the reproductive mode shift back to the apomixis original level. These data together with previous observations allow us to generate a hypothetical model of the regulation of apomixis in weeping lovegrass in which the genetic/s region/s that condition apomixis, is/are affected by ploidy, and is/are subjected to epigenetic control.

A Parthenogenesis Gene Candidate and Evidence for Segmental Allopolyploidy in Apomictic Brachiaria decumbens.

Apomixis, asexual reproduction through seed, enables breeders to identify and faithfully propagate superior heterozygous genotypes by seed without the disadvantages of vegetative propagation or the expense and complexity of hybrid seed production. The availability of new tools such as genotyping by sequencing and bioinformatics pipelines for species lacking reference genomes now makes the construction of dense maps possible in apomictic species, despite complications including polyploidy, multisomic inheritance, self-incompatibility, and high levels of heterozygosity. In this study, we developed saturated linkage maps for the maternal and paternal genomes of an interspecific Brachiaria ruziziensis (R. Germ. and C. M. Evrard) × B. decumbens Stapf. F1 mapping population in order to identify markers linked to apomixis. High-resolution molecular karyotyping and comparative genomics with Setaria italica (L.) P. Beauv provided conclusive evidence for segmental allopolyploidy in B. decumbens, with strong preferential pairing of homologs across the genome and multisomic segregation relatively more common in chromosome 8. The apospory-specific genomic region (ASGR) was mapped to a region of reduced recombination on B. decumbens chromosome 5. The Pennisetum squamulatum (L.) R.Br. PsASGR-BABY BOOM-like (psASGR-BBML)-specific primer pair p779/p780 was in perfect linkage with the ASGR in the F1 mapping population and diagnostic for reproductive mode in a diversity panel of known sexual and apomict Brachiaria (Trin.) Griseb. and P. maximum Jacq. germplasm accessions and cultivars. These findings indicate that ASGR-BBML gene sequences are highly conserved across the Paniceae and add further support for the postulation of the ASGR-BBML as candidate genes for the apomictic function of parthenogenesis.

Somatic Embryogenesis and Plant Regeneration of Brachiaria brizantha.

The genus Brachiaria (Trin.) Griseb. belongs to the family Poaceae, order Poales, class Monocotyledonae. In Brachiaria brizantha (Hochst. ex A. Rich.) Stapf., embryogenic callus can be induced from seeds from apomictic plants, which results in high frequency somatic embryo development and plant regeneration. We report here a detailed protocol for callus induction from apomictic seed; followed by in vitro morphogenesis (somatic embryo and bud differentiation), plant regeneration, and acclimatization in the greenhouse. Important details regarding the positioning of seeds for callus induction and precautions to avoid endophytic contamination and the occurrence of albino plants are presented.