Isolation and characterization of microsatellite markers for Brachiaria brizantha (Hochst. ex A. Rich.) Stap

The first set of nuclear simple sequence repeat (SSR) loci for Brachiaria brizantha (Hochst. ex A. Rich.) Stap is described. A microsatellite-enriched library was constructed and 19 loci were characterized. About 13 SSR loci were found to be polymorphic and across-taxa amplification tests showed that six of them can be transferred to four other species of Brachiaria. This new SSR resource will be a powerful tool for population genetic studies of B. brizantha, for interspecific genetic studies within the genus Brachiaria, for mapping and for marker assisted selection in breeding.     

Genetic Diversity and Population Structure of the Brachiaria brizantha Germplasm

Brachiaria brizantha (Hochst. ex A. Rich.) Stapf. (syn. Urochloa brizantha (Hochst. ex A. Rich.) R.D. Webster) is a species used primarily as forage in tropical America and Southeast Asia. B. brizantha has been extensively researched since the 1980s with the initiation of the Tropical Forages Breeding Program conducted by the Brazilian Agricultural Research Corporation (Empresa Brasileira de Pesquisa Agropecuária; EMBRAPA), holding one of the largest germplasm collections in the world. This work has identified 15 new microsatellite markers for this species, which have been used in addition to five previously reported markers, to estimate the genetic similarities among 172 accessions and six cultivars of this species. Similarity index values ranged from 0.40 to 1.00. Two duplications were found in the germplasm. A Bayesian analysis performed using the STRUCTURE 2.3.3 program revealed the presence of three clusters with different allelic pools. This analysis is valuable for the performance of crosses to explore heterosis; however, the mode of reproduction of the accessions and ploidy barriers must be observed for effective exploration. A grouping analysis using the neighbor-joining method was consistent with the STRUCTURE analysis, and a combination approach suggested that this germplasm collection does not exhibit considerable genetic variability despite the presence of three distinct allelic pools. The lack of correlation between the genetic and geographic distances is also discussed.     

Silicon distribution and accumulation in shoot tissue of the tropical forage grass Brachiaria brizantha

Silicon (Si) accumulation in organs and cells is one of the most prominent characteristics of plants of the family Poaceae. Many species from this family are used as forage plants for animal feeding. The present study investigates in Brachiaria brizantha (Hochst. ex A. Rich.) Stapf. cv. Marandu: (1) the dry matter production and Si content in shoot due to soil Si fertilizations; (2) the Si distribution among shoot parts; and (3) the silica deposition and localization in leaves. Plants of B. brizantha cv. Marandu were grown under contrasting Si supplies in soil and nutrient solution. Silica deposition and distribution in grass leaf blades were observed using light microscope and scanning electron microscope equipped with an energy dispersive X-ray spectrometer (SEM-EDXS). Silicon concentration in the B. brizantha shoot increased according to the Si supply. Silicon in grass leaves decreased following the order: mature leaf blades > recently expanded leaf blades > non-expanded leaf blades. Silicon accumulates mainly on the upper (adaxial) epidermis of the grass leaf blades and, especially, on the bulliform cells. The Si distribution on adaxial leaf blade surface is non uniform and reflects a silica deposition exclusively on the cell wall of bulliform cells.     

Ethiopian mistletoes: new species and combinations

Preparatory to the publication of the Flora of Ethiopia, the following new species and combinations are made: Loranthaceae: Plicosepalus robustus Wiens & Polh., P. ogadenensis M. Gilbert, P. acaciae (Zucc.) Wiens & Polh., P. meridianus (Danser) Wiens & Polh.; Oncocalyx angularis M. Gilbert, O. fischeri (Engl.) M. Gilbert, O. glabratus (Engl.) M. Gilbert, O. ugogensis (Engl.) Wiens & Polh., 0. schimperi (Hochst. ex A. Rich.) M. Gilbert, O. ghikae (Volkens & Schweinfurth) M. Gilbert, O. kelleri (Engl.) M. Gilbert; Erianthemum aethiopicum Balle ex Wiens & Polh.; Englerina woodfordioides (Schweinfurth) Balle ex M. Gilbert; Phragmanthera ahhallensis (Engl.) M. Gilbert, P. erythraea (Sprague) M. Gilbert, P. macrosolen (Steud. ex A. Rich.) M. Gilbert, P. regularis (Steud. ex Sprague) M. Gilbert, P. sarertaensis (Hutch. & Bruce) M. Gilbert, P. usuiensis (Oliver) M. Gilbert. Viscaceae: Viscum triflorum DC. ssp. nervosum (A. Rich.) M. Gilbert.     

Economic Botany ofSphenostylis (Leguminosae)

Sphenostylis, a genus of seven species in tropical and southern Africa, includes three taxa that are used by humans. Flowers and seeds of S. schweinfurthii Harms are occasionally eaten in West Africa, and this species has potential value as a forage crop. Flowers and seeds of Sphenostylis erecta (E. G. Baker) E. G. Baker subsp. erecta are eaten in parts of Central Africa, while the roots are used medicinally and as a source of dye and fish poison. The edible tubers of S. stenocarpa (Hochst, ex A. Rich.) Harms, the African yam bean, are collected from the wild in Central and East Africa. This species is cultivated for its edible seeds in west tropical Africa and for its edible tubers in Zaire. Cultivated races of S. stenocarpa may be considered domesticated forms, since they differ from wild plants of the species in a number of morphological characteristics that are evidently the result of human selection. A list of common names for the three taxa, from throughout their ranges, is presented. Sphenostylis, un género con siete especies de las regiones sur y tropical de Africa, incluye tres taxa utilizados por humanos. Las flores y semillas deS. schweinfurthii Harms son alimento ocasional en el oeste de Africa, y la especie tiene, ademas, valor potencial como forraje. Las flores y semillas de Sphenostylis erecta (E. G. Baker) E. G. Baker subsp. erecta se comen en regiones del centro de Africa, mientras que las raíces tienen usos medicinales y son fuente de colorantes y embarbascantes. Los tubérculos comestibles de S. stenocarpa (Hochst, ex A. Rich.) Harms, la jícama Africana, se colectan de plantas silvestres en el centro y este de Africa. Esta especie se cultiva por sus semillas comestibles en el oeste de Africa tropical, y por sus tubérculos comestibles en Zaire. Las razas cultivadas deS. stenocarpa pueden ser consideradas como formas domesticadas, puesto que difieren de las plantas silvestres en muchas características morfológicas que son, evidentemente, el resultado de la selección humana. Se presenta una lista de los nombres comunes para los tres taxa a lo largo de sus rangos de distribución.     

Cytogenetic characterization and fae1 gene variation in progenies from asymmetric somatic hybrids between Brassica napus and Crambe abyssinica.

Sexual progenies of asymmetric somatic hybrids between Brassica napus and Crambe abyssinica were analyzed with respect to chromosomal behavior, fae1 gene introgression, fertility, and fatty-acid composition of the seed. Among 24 progeny plants investigated, 11 plants had 38 chromosomes and were characterized by the occurrence of normal meiosis with 19 bivalents. The other 13 plants had more than 38 chromosomes, constituting a complete chromosomal set from B. napus plus different numbers of additional chromosomes from C. abyssinica. The chromosomes of B. napus and C. abyssinica origin could be clearly discriminated by genomic in situ hybridization (GISH) in mitotic and meiotic cells. Furthermore, meiotic GISH enabled identification of intergenomic chromatin bridges and of asynchrony between the B. napus and C. abyssinca meiotic cycles. Lagging, bridging and late disjunction of univalents derived from C. abyssinica were observed. Analysis of cleaved amplified polymorphic sequence (CAPS) markers derived from the fae1 gene showed novel patterns different from the B. napus recipient in some hybrid offspring. Most of the progeny plants had a high pollen fertility and seed set, and some contained significantly greater amounts of seed erucic acid than the B. napus parent. This study demonstrates that a part of the C. abyssinica genome can be transferred into B. napus via asymmetric hybridization and maintained in sexual progenies of the hybrids. Furthermore, it confirms that UV irradiation improves the fertility of the hybrid and of its sexual progeny via chromosomal elimination and facilitates the introgression of exotic genetic material into crop species.     

Physical mapping of rDNA genes corroborates allopolyploid origin in apomictic Brachiaria brizantha

Brachiaria (Trin.) Griseb belongs to the family Poaceae, and within the genus, apomixis or sexuality is present in different accessions of the same species. The majority of Brachiaria species are polyploid and apomictic, making strategies for crop improvement by breeding very intricate. In spite of the high frequency of apomictic polyploids, the relationship of polyploidy and hybridization with apomixis in Brachiaria is still unclear. Further analysis requires detailed knowledge regarding the genomic composition of the polyploids. The present work introduces the use of fluorescent in situ hybridization (FISH) into cytogenetic analysis of Brachiaria. Physical mapping of heterologous rDNA sequences, associated with conventional karyotyping of the B. brizantha diploid sexual (BRA 002747) and the tetraploid apomictic (BRA000591) accessions, provided evidence of the latter being of allopolyploid origin. Based on our results and on previous knowledge on apomixis in B. brizantha, we suggest that the origin of apomixis was probably a consequence of hybridization.     

Selection of reference genes for quantitative real-time PCR expression studies in the apomictic and sexual grass Brachiaria brizantha

Background  

Brachiaria brizantha is an important forage grass. The occurrence of both apomictic and sexual reproduction within Brachiaria makes it an interesting system for understanding the molecular pathways involved in both modes of reproduction. Quantitative real time PCR (qRT-PCR) has emerged as an important technique to compare expression profile of target genes and, in order to obtain reliable results, it is important to have suitable reference genes. In this work, we evaluated eight potential reference genes for B. brizantha qRT-PCR experiments, isolated from cDNA ovary libraries. Vegetative and reproductive tissues of apomictic and sexual B. brizantha were tested to validate the reference genes, including the female gametophyte, where differences in the expression profile between sexual and apomictic plants must occur.     

The genus Urera (Urticaceae) in eastern tropical Africa

The species of Urera Gaud. occurring in eastern tropical Africa are revised. Three species are recognized: U. cameroonensis Wedd., occurring from Uganda to the South African Republic (Natal), U. sansibarica Engl., restricted to the coastal zone of Kenya and Tanzania, and U. hypselodendron (Hochst. ex A. Rich.) Wedd., occurring in montane forest from Ethiopia to Malawi. The original material of U. hypselodendron var. flamigniana Hauman (nom. inval.) and var. platyrrhachis Hauman (nom. inval.) is excluded from U. hypselodendron; the same applies to the other material of U. hypselodendron cited from central Zaire in Flore du Congo belge. Distribution maps of the eastern African species are provided. Also included is a preliminary study and discussion of the generic subdivision of Urera , with special reference to the species of this revision.     

Somatic embryogenesis and organogenesis in apomictic and sexual Brachiaria brizantha

Brachiaria brizantha (syn. Urochloa brizantha) is an important tropical forage grass widely cultivated in Brazil. In order to optimize tissue culture conditions for B. brizantha, in vitro culture of mature seeds, basal segments and leaf segments from in vitro plants of an apomictic and a sexual genotype of B. brizantha was performed. When cultured on different media, leaf segments yielded non-embryogenic calluses which formed several roots. Friable calluses from mature seeds and basal segments explants incubated on Murashige and Skoog medium supplemented with 2,4-dichlorophenoxyacetic acid and 6-benzyladenine yielded 80% compact and nodular embryogenic structures. Calluses with such compact embryogenic structures were highly regenerable upon transfer to medium supplemented with kinetin and naphthalene acetic acid. They produced isolated somatic embryos, multiple fused scutelli or isolated scutellum with polyembryos that germinated into isolated or multiple shoots. Green and morphologically normal plants were obtained for the two genotypes. Changing the media from pH 5.8 to pH 4.0 increased the number of explants that formed calluses as well as the number of shoots per explant. When embryogenic calluses from mature seeds were successively sub-cultured for 4 months, aiming at repetitive somatic embryogenesis, all the regenerated plants were albinos. The embryogenic nature of the compact structure was confirmed by scanning electron microscopy.     

The taxonomic position and derivation of Cerastium adnivale Chiov. (Caryophyllaceae)

Detailed studies of comprehensive herbarium material confirmed that the afroalpine Cerastium adnivale Chiov. is too indistinctly delimited from the earlier described C. octandrum Hochst. ex A. Rich, to be given more than varietal rank. The restricted distribution of var. adnivale on only some of the high mountains harbouring C. octandrum s. lat. as well as results of experimental cultivation show that the reduced pubescence of the former cannot be due to environmental influence alone but must result from genetic differences. The derivation of var. adnivale from C. octandrum s. lat. is discussed.     

Inheritance of transgenes in transgenic tall fescue (<Emphasis Type="Italic">Festuca arundinacea</Emphasis> schreb.)

Summary Tall fescue (Festuca arundinacea Schreb.) is the most important forage species worldwide of the Festuca genus. Single genotype-derived embryogenic suspension cultures were established from tall fescue cultivar Kentucky-31, and were used as target cells for biolistic transformation. A chimeric hygromycin phosphotransferase gene (hph) was used as the selectable marker, and a chimeric β-glucuronidase (gusA) gene was co-transformed with hph. Transgenic plants were recovered after microprojectile bombardment of suspension cells and subsequent selection in the presence of a high concentration of hygromycin. Fertile transgenic plants were obtained after vernalization under field conditions. T1 and T2 progenies were obtained after reciprocal crosses between transgenic and untransformed control plants. PCR and Southern hybridization analyses revealed a 1∶1 segregation ratio for both transgenes in the T1 and T2 generations. Southern hybridization patterns were identical for T0, T1, and T2 plants. The results demonstrated for the first time the stable meiotic transmission of transgenes following Mendelian rules in transgenic tall fescue.     

The fate of recombinant chromosomes and genome interaction in Nicotiana asymmetric somatic hybrids and their sexual progeny

Genomic in-situ hybridization (GISH) was used to monitor the behaviour of parental genomes, and the fate of intergenomic chromosome translocations, through meiosis of plants regenerated from asymmetric somatic hybrids between Nicotiana sylvestris and N. plumbaginifolia. Meiotic pairing in the regenerants was exclusively between chromosomes or chromosome segments derived from the same species. Translocation (recombinant) chromosomes contained chromosome segments from both parental species, and were detected at all stages of meiosis. They occasionally paired with respectively homologous segments of N. sylvestris or N. plumbaginifolia chromosomes. Within hybrid nuclei, the meiotic division of N. plumbaginifolia lagged behind that of N. sylvestris. However, normal and recombinant chromosomes were eventually incorporated into dyads and tetrads, and the regenerants were partially pollen fertile. Recombinant chromosomes were transmitted through either male or female gametes, and were detected by GISH in sexual progeny obtained on selfing or backcrossing the regenerants to N. sylvestris. A new recombinant chromosome in one plant of the first backcross generation provided evidence of further chromosome rearrangements occurring at, or following, meiosis in the original regenerants. This study demonstrates the stable incorporation of chromosome segments from one parental genome of an asymmetric somatic hybrid into another, via intergenomic translocation, and reveals their transmission to subsequent sexual progeny.     

Cytogenetic analysis of Lycopersicon esculentum (+) Solanum etuberosum somatic hybrids and their androgenetic regenerants

The aim of the study was to characterize genomic relationships among cultivated tomato (Lycopersicon esculentum Mill.) (2n=2x=24) and diploid (2n=2x=24) non-tuberous wild Solanum species (S. etuberosum Lindl.). Using genomic in situ hybridization (GISH) of mitotic and meiotic chromosomes, we analyzed intergeneric somatic hybrids between tomato and S. etuberosum. Of the five somatic hybrids, two plants were amphidiploids (2n=4x=48) mostly forming intragenomic bivalents in their microsporocytes, with a very low frequency of multivalents involving the chromosomes of tomato and S. etuberosum (less than 0.2 per meiocyte). Tomato chromosomes showed preferential elimination during subsequent meiotic divisions of the amphidiploids. Transmission of the parental chromosomes into microspores was also evaluated by GISH analysis of androgenic plants produced by direct embryogenesis from the amphidiploid somatic hybrids. Of the four androgenic regenerants, three were diploids (2n=2x=24 or 2n=2x+1=25) derived from reduced male gametes of the somatic hybrids, and one plant was a hypertetraploid (2n=4x+4=52). GISH revealed that each anther-derived plant had a unique chromosome composition. The prospects for introgression of desirable traits from S. etuberosum into the gene pool of cultivated tomato are discussed. Received: 2 August 2000 / Accepted: 4 December 2000     

A taxonomic revision of Crambe, sect. Leptocrambe (Brassicaceae)

As part of a revision of the genus Crambe based on the morphological study of herbaria and cultivated material, the systematics of sect. Leptocrombe DC. is presented here. Section Leptocrambe is considered to comprise five species: C. kilimandscharica O. E. Schulz, C. sinuatodentata Hochst. ex Petri, C. hispanica L., C. filiformis Jacq. and C. kralikii Coss. C. hispanica includes three subspecies, subsp. hispanica , subsp. glabrata (DC.) Cout. and subsp. abyssnica (Hochst. ex R. E. Fr.) stat. nov. which includes var. dyssinica and var. meyeri (O. E. Schulz) comb. nov. C. kralikii includes two subspecies, subsp. kralikii and subsp. garamas (Maire) Podlech.     

Meiosis-driven genome variation in plants

Meiosis includes two successive divisions of the nucleus with one round of DNA replication and leads to the formation of gametes with half of the chromosomes of the mother cell during sexual reproduction. It provides a cytological basis for gametogenesis and nheritance in eukaryotes. Meiotic cell division is a complex and dynamic process that involves a number of molecular and cellular events, such as DNA and chromosome replication, chromosome pairing, synapsis and recombination, chromosome segregation, and cytokinesis. Meiosis maintains genome stability and integrity over sexual life cycles. On the other hand, meiosis generates genome variations in several ways. Variant meiotic recombination resulting from specific genome structures induces deletions, duplications, and other rearrangements within the genic and non-genic genomic regions and has been considered a major driving force for gene and genome evolution in nature. Meiotic abnormalities in chromosome segregation lead to chromosomally imbalanced gametes and aneuploidy. Meiotic restitution due to failure of the first or second meiotic division gives rise to unreduced gametes, which triggers polyploidization and genome expansion. This paper reviews research regarding meiosis-driven genome variation, including deletion and duplication of genomic regions, aneuploidy, and polyploidization, and discusses the effect of related meiotic events on genome variation and evolution in plants. Knowledge of various meiosis-driven genome variations provides insight into genome evolution and genetic variability in plants and facilitates plant genome research.     

Host plant selection behaviour of Chilo partellus and its implication for effectiveness of a trap crop

Female lepidopterans can display a hierarchy of preference among potential host species, a trait thought to arise from the balance between attractants and deterrents to which the insects respond. Host plant ranking by moths and larvae of Chilo partellus Swinhoe (Lepidoptera: Crambidae), an important pest of cereals in Africa, was investigated, and whether eggs deposited on specific host plants yield larvae of particular host preferences. Trap plants are used in management of this pest. However, any ‘disagreement’ in host ranking between moths and larvae could potentially reduce effectiveness of trap crops as larvae emigrate to the main crop from the parent’s preferred trap plant. We also investigated whether host plant preference is influenced by the diet upon which larvae fed as part of an integrated assessment of the relationship between host plant selection and learning in C. partellus. Five host plants (all Poaceae) were used: maize (Zea mays L.), sorghum (Sorghum bicolor Moench), Napier grass (Pennisetum purpureum Schumach), and two varieties of signal grass [Brachiaria brizantha (A. Rich.) Stapf], viz., local (henceforth signal grass) and improved (‘Mulato’). In multiple choice tests, C. partellus female moths preferentially oviposited on Napier grass, followed by sorghum, maize, and signal grass, and least preferred ‘Mulato’. Larvae however equally orientated and settled on leaf cuts of maize, sorghum, signal grass, and Napier grass, but least preferred ‘Mulato’. Moreover, eggs from specific host plants did not yield larvae of particular host preferences. Furthermore, oviposition preference was not altered by the larval food. These results imply only a slight ‘disagreement’ in host ranking behaviour between moths and larvae, which is beneficial for trap cropping as larvae would not ‘reject’ the trap plant and appreciably disperse to the neighboring plants. Moreover, absence of larval learning behaviour indicates that regardless of the larval food C. partellus moths would still be attracted to the selected trap plant.     

FORMATION OF 2N MEGAGAMETOPHYTES IN DIPLOID TUBER-BEARING SOLANUMS

Development of megaspores and megagametophytes was analyzed for several diploid potato clones (Solanum spp.) that exhibit either high (HI) or low (LO) seed set when crossed as female with the tetraploid cultivated potato S. tuberosum Group Tuberosa. The objectives were to determine the relationship between ploidy and diam of nuclei and nucleoli, and to determine the mechanism(s) and frequencies of 2n megagametophyte formation. Sizes of nuclei and nucleoli were found to depend on ploidy. For HI clones, the distributions of sizes indicated that doubling occurred during meiosis, and that 30 to 50% of the megaspores and megagametophytes were 2n rather than haploid. Omission of the second meiotic division led to formation of second division restitution (SDR) 2n megagametophytes. Only one HI clone had abnormal meiosis I, in addition to omission of meiosis II in some meiocytes; this clone seemed to produce not only 1n and 2n, but also 4n megagametophytes. The results indicated that high crossability of the HI clones as female with tetraploids largely was due to formation of SDR 2n megagametophytes, a finding strongly supporting the hypothesis that sexual polyploidization is the driving force behind polyploidization of Solanums. The results contribute to increasing evidence that meiotic mutants and abnormalities play an important role in angiosperm evolution.     

What are the spermatocyte's requirements for successful meiotic division?

The consequences of error during meiotic division in spermatogenesis can be serious: aneuploid spermatozoa, embryonic lethality, and developmental abnormalities. Recombination between homologs is essential to ensure normal segregation; thus the spermatocyte must time division precisely so that it occurs after recombination between chromosomes and accumulation of the cell-cycle machinery necessary to ensure an accurate segregation of chromosomes. We use two systems to investigate meiotic division during spermatogenesis in the mouse: pharmacological induction of meiotic metaphase in cultured spermatocytes and transillumination-mediated dissection of stage XII seminiferous tubule segments to monitor progress through the division phase. By these approaches we can assess timing of acquisition of competence for the meiotic division phase and the temporal order of events as division proceeds. Competence for the meiotic division arises in the mid-pachytene stage of meiotic prophase, after chromosomes have synapsed and coincident with the accumulation of the cell-cycle regulatory protein CDC25C. The activity of both MPF and topoisomerase II are required. The earliest hallmarks of the division phase are nuclear envelope breakdown, followed by phosphorylation of histone H3 and chromosome condensation. These events are likely to be monitored by checkpoint mechanisms since checkpoint proteins can be localized in nuclei and DNA-damaging agents delay entry into the meiotic division phase. Understanding how the spermatocyte regulates its entry into the meiotic division phase can help clarify the natural mechanisms ensuring accurate chromosome segregation and preventing aneuploidy. J. Exp. Zool. (Mol. Dev. Evol.) 285:243-250, 1999.