Maria Suely Pagliarini

Meiotic behavior of economically important plant species: the relationship between fertility and male sterility

Meiosis is an event of high evolutionary stability which culminates in a reduction of chromosome number. The normal and harmonious course of meiosis ensures gamete viability. The cytologic events of gametogenesis are controlled by a large number of genes that act from premeiotic to postmeiotic mitosis. Mutations in these genes cause anomalies that may impair fertility, and many abnormalities affecting plant fertility or causing total male sterility have been detected during the evaluation of meiotic behavior in some species. Some of these abnormalities have been frequently described in the literature, while others have not been previously reported. The most frequent abnormalities found in the species analyzed were irregular chromosome segregation, cytomixis, chromosome stickiness, mixoploidy, chromosome fragmentation, syncyte formation, abnormal spindles, and failure of cytokinesis. Uncommon abnormalities, such as chromosome elimination during microsporogenesis, were found in one species. Original meiotic mutations affecting different steps of meiosis were also observed in these species, especially in maize, Paspalum and soybean. Some mutants present characteristics that may be exploited successfully in breeding programs because they cause total male sterility.

Chromosome numbers and microsporogenesis in Brachiaria brizantha (Gramineae)

The genus Brachiaria, native to the African tropical savannas, has achieved significance as a pasture grass in many tropical and subtropical countries, including Brazil. Many species and accessions are polyploid and apomictic, which complicates the improvement of breeding stocks through hybridization. In support of breeding programs, cytogenetic characterization, including chromosome counts and evaluation of the meiotic behavior in the accessions of the Brachiaria has been undertaken at the Embrapa Beef Cattle Center. In this study, 22 accessions of B. brizantha were analyzed of which one was found to be diploid (2n = 2x = 18), 18 were tetraploid (2n = 4x = 36) and three were hexaploid (2n = 6x = 54). The meiotic chromosome behavior was slightly irregular in the diploid and in some tetraploid accessions, and highly irregular in most tetra- and hexaploid accessions. Meiotic abnormalities were those common to polyploidy, i.e., multivalent chromosome association at diakinesis and irregular chromosome segregation leading to micronuclei formation in the tetrad stage. Low frequencies of multivalent chromosome associations among polyploids suggest that they may be segmental allopolyploids.

Meiotic Behavior and Pollen Fertility in Aptenia Cordifolia (Aizoaceae)

SUMMARYMeiotic analysis of Aptenia cordifolia revealed that total chiasma terminalization in some bivalents at diakinesis led to the formation of univalent chromosomes which did not undergo regular segregation and reached the end of meiosis as micronuclei. As a consequence, final products of meiosis showing variation in microspore number and size were observed, causing high pollen sterility.

Meiotic behavior of several Brazilian soybean varieties.

Despite the importance of soybeans little cytogenetic work has traditionally been done, due to the small size and apparent similarity of the chromosomes. Fifteen soybean [Glycine max (L.) Merrill] varieties adapted for cultivation in two distinct regions of Brazil were analyzed cytogenetically. A low frequency of meiotic abnormalities was noted in all varieties, although they were not equally affected. Irregular chromosome segregation, chromosome stickiness, cytoplasmic connections between cells, cytomixis and irregular spindles were the main abnormalities observed, none of which had been described previously in soybeans. All of these abnormalities can affect pollen fertility. Pollen fertility was high in most varieties and was correlated with meiotic abnormalities. Although soybean is not a model system for cytological studies, we found that it is possible to conduct cytogenetic studies on this species, though some modifications in the standard methods for meiotic studies were necessary to obtain satisfactory results.

UNUSUAL CYTOLOGICAL PATTERNS OF MICROSPOROGENESIS IN BRACHIARIA DECUMBENS: ABNORMALITIES IN SPINDLE AND DEFECTIVE CYTOKINESIS CAUSING PRECOCIOUS CELLULARIZATION

Cytogenetic studies carried out in the tetraploid accession BRA001068 of Brachiaria decumbens, also known as cv. Basilisk, revealed an unusual pattern of microsporogenesis. The spindle in metaphase I and anaphase I became heavily stained with propionic carmine. In telophase I, the interzonal microtubules continued to be intensely stained, and during the phragmoplast formation the fibers were pushed to the cell wall, persisting until prophase II, even after cytokinesis. Due to its tetraploid condition, the accession presented many cells with precocious chromosome migration to the poles in metaphase I and laggards in anaphase I that gave rise to micronuclei in telophase I. While in other polyploid accessions of Brachiaria micronuclei remained in this condition until the second cytokinesis, the micronuclei in this accession organized their own spindle in the second division. In several microsporocytes, the micronuclei with their minispindle were divided further into microcytes by additional cytokinesis. Some curious planes of cytokinesis were found in some cells, with partitioning of cytoplasm into cells of irregular shape. The result consisted of a high frequency of abnormal products of meiosis. Quadrivalents were observed in diakinesis at low frequency, which suggests a segmental allotetraploid and the inability of both genomes to co‐ordinate their activities, leading to multiple spindle and precocious cellularization. In spite of abnormal meiotic products reducing pollen fertility, seed production was normal. Enough normal pollen was available to fertilize the central‐cell nucleus of the embryo sac and produce normal endosperm in this pseudogamous aposporous apomictic accession.

Chromosome number and meiotic behaviour in Brachiaria jubata (Gramineae)

The production of new genetic variability through hybridization has been undertaken in Brachiaria, an important forage grass genus of African origin widely used in pastures in the Brazilian tropics. Although the genus comprises about 100 species, only a few of them present favourable agronomic attributes and are explored. In the genus Brachiaria, the majority of species and accessions are polyploid and apomictic, which makes breeding through hybridization more complex. Sexuality has been found in diploids and eventually among tetraploids with normal meiosis. Brachiaria jubata presents good forage value and adaptation to riverbanks and lake margins, and is therefore relevant to pasture systems i n the tropics. Furthermore, the most interesting form of resistance to spittlebugs—antibiosis—was identified in one a ccession of B. jubata. Thus its interest in the Brazilian breeding programme lies in the possibility of using hybridization to transfer high resistance to this insect to susceptible ac cessions. This paper reports chromosome numbers and meiotic behaviour in 21 accessions of this species as a tool in selecting promising ones for crossing. Only one accession was found to be diploid; the others were tetraploid. Microsporogenesis was normal in the diploid accession, while among the tetraploid ones, irregular chromosome segregation leading to unbalanced gamete formation was common but variable. Pollen fertility among tetraploid accessions was hig h. Cytogenetic studies were carried out on accessions of Brachiaria jubatafrom the Brachiaria germplasm collection at Embrapa Beef Cattle kept in the field in Campo Grande (state of Mato Grosso do Sul, Brazil), which comprises 475 accessions of 15 species collected in Africa in the mid-1980s by CIAT (Colombia). Inflorescences were collected and

Multiple spindles and cellularization during microsporogenesis in an artificially induced tetraploid accession of Brachiaria ruziziensis (Gramineae)

The genus Brachiaria is characterized by a majority of polyploid accessions—mainly tetraploid—and apomictic reproduction. Sexuality is found among diploids. To overcome incompatibility barriers, accessions with the same ploidy level are necessarily used in hybridization. Thus, sexual diploid accessions were tetraploidized to be used as female genitors. This paper reports microsporogenesis in an artificially induced tetraploid accession of Brachiaria ruziziensis. Chromosome pairing at diakinesis ranged from univalents to tetravalents, with predominance of bivalents. Irregular chromosome segregation was frequent in both meiotic divisions. During the first division, multiple spindles showing different arrangements were recorded. The spindle position determined the plane of first cytokinesis and the number of chromosomes determined the size of the cell. Meiotic products were characterized by polyads with spores of different sizes. Pollen sterility was estimated at 61.38%. The limitations of using this accession in the breeding program are discussed.

Meiotic behavior in interspecific hybrids between Brachiaria ruziziensis and Brachiaria brizantha (Poaceae)

The meiotic behavior of two half-sib interspecific tetraploid (2n = 4x = 36) promising hybrids, a sexual and an apomictic one, from crosses B. ruziziensis and B. brizantha, was evaluated. Although chromosome paired predominantly as bivalents, a few tri- and quadrivalents were recorded. Results suggest that B. brizantha and B. ruziziensis are closely related and genetic recombination is expected in hybrids. Introgression of specific target genes from B. ruziziensis into B. brizantha and vice-versa may be foreseen. However, abnormalities such as irregular chromosome segregation, chromosome stickiness and abnormal cytokinesis reported in these hybrids affect pollen fertility. More than 65% of pollen grains are sterile. Since the distinctive cytological feature of these hybrids is abnormal cytokinesis, this fact suggests that both parental genomes are unable to coordinate their activities with regard to this cytological phenomenon. Deployment of such hybrids in the process of developing varieties is discussed.

Elimination of micronuclei from microspores in a Brazilian oat (Avena sativa L.) variety

Chromosome elimination in interspecific hybrids is a powerful tool in breeding programs. Of 12 Brazilian oat varieties recommended for cultivation in southern Brazil, one variety, UFRGS 15, presented a new type of chromosome elimination never described in any other species. Chromosomes that produced micronuclei during meiosis were eliminated from microspores as microcytes. The micronucleus reached the microspore wall and formed a kind of bud, separated from the microspore. The eliminated microcytes gave origin to small and sterile pollen grains. The possible applications of this finding for oat breeding are discussed.

Abnormal timing of cytokinesis in microsporogenesis in Brachiaria humidicola (Poaceae: Paniceae)

Meiosis is controlled by a large number of genes, generally dominant, whose expression is stage-specific, site-specific and time-specific (Gottschalk and Kaul 1974, 1980a,b; Baker et al. 1976; Golubovskaya 1979, 1989). Among genes acting in the meiotic process, those responsible for the partitioning of the cytoplasm after nuclear division play a very important role in the formation of viable gametes. After two rounds of chromosome segregation (karyokinesis) and one simultaneous or two successive cytoplasmic divisions (cytokinesis), the final product of male meiosis in flowering plants emerges as a tetrad of haploid microspores enclosed in a callose wall. The timing of cytokinesis varies among angiosperms. In most monocot plants, cytokinesis is successive, i.e. one partitioning of the cytoplasm occurs after telophase I and another after telophase II, so that there is a distinct dyad stage. However, in most dicots, it is simultaneous and occurs after telophase II (Peirson et al. 1996). Many meiosis mutations affecting the pattern of cytokinesis have been reported in higher plants (Peirson et al. 1996). In Brachiaria, a promising African genus of forage grass for the Brazilian savannas, absence of cytokinesis leading to 2n microspores and binucleated or tetranucleated microspores have been reported in B. brizantha (Risso-Pascotto et al. 2003) and B. nigropedata (Utsunomiya et al. 2005). The present study details meiotic and postmeiotic abnormalities related to cytokinesis observed in one accession of B. humidicola.