Chromosome number and microsporogenesis of two accessions of Brachiaria dura Stapf (Poaceae) / Número de cromossomos e microsporogênese de dois acessos de Brachiaria dura Stapf (Poaceae)

The two accessions of B. dura analyzed (DU01 and DU02) are hexaploid (2n = 6x = 54), derived from x = 9. Meiotic abnormalities, such as precocious chromosome migration to the poles, laggards and micronuclei, were recorded in low frequency in both accessions. The few multivalent chromosome association at diakinesis and meiotic stability suggested that hexaploidy probably resulted from chromosome doubling. In DU02, chromosome transfer (cytomixis) among meiocytes, involving part or the entire genome was observed. The implication of these findings for the Brachiaria breeding is discussed.

Os dois acessos de B. dura analisados (DU01 e DU02) são hexaplóides (2n = 6x = 54), derivados de x = 9. Anormalidades meióticas como migração precoce de cromossomos para os polos, cromossomos retardatários e micronúcleos foram observados em baixa frequência em ambos os acessos. A presença de poucas associações cromossômicas em diacinese e a estabilidade meiótica sugere que a hexaploidia provavelmente resultou de duplicação cromossômica. No acesso DU02 observou-se transferência de cromossomos (citomixia) entre meiócitos, envolvendo parte ou todo o genoma. As implicações destes resultados para o melhoramento de Brachiaria são discutidas.

Desynapsis and precocious cytokinesis in Brachiaria humidicola (Poaceae) compromise meiotic division.

The forage grass species Brachiaria humidicola is native to African savannas. Owing to its good adaptation to poorly drained and infertile acid soils, it has achieved wide utilization for pastures in Brazilian farms. Among the 55 accessions of B. humidicola analysed from the Embrapa Beef Cattle collection, one (H022), presented desynapsis and an abnormal pattern of cytokinesis in the first meiotic division. Among 28 inflorescences analysed in this accession, 12 were affected by the anomaly. In affected meiocytes, the first cytokinesis occurred in metaphase I and was generally perpendicular to a wide-metaphase plate, dividing the genome into two parts with an equal or unequal number of chromosomes. The normal cytokinesis after telophase I did not occur, and the meiocytes entered metaphase II, progressing to the end of meiosis with the occurrence of the second cytokinesis. As the first cytokinesis occurred precociously, whereas the second was normal, tetrads were formed but with unbalanced chromosome numbers in microspores. Abnormal cytokinesis occurred only in those meiocytes that underwent desynapsis after diakinesis. The implications of this abnormality in the Brachiaria breeding programme are discussed.

Evaluation of microsporogenesis in an interspecific Brachiaria hybrid (Poaceae) collected in distinct years

Microsporogenesis in an interspecific Brachiaria hybrid, grown in the field under natural environmental conditions in Brazilian savannas, was analyzed in three distinct years of collection. Several types of meiotic abnormalities were recorded during those three years, but varied in type and frequency depending on the year. The average temperature and rainfall 15 days before collection was unusually high in those years. The percentage of abnormal meiocytes recorded was 62% in 2001, 73% in 2004, and 77% in 2005. The abnormalities observed during microsporogenesis compromised pollen viability by generating unbalanced gametes or affecting nucleolus organization. The environmental conditions under which the hybrid was growing could have affected the genetic control of meiosis. More detailed studies, under controlled conditions, are necessary to better understand the effects of environmental factors on Brachiaria microsporogenesis hybrids.

Mitotic behavior in root tips of Brachiaria genotypes with meiotic chromosome elimination during microsporogenesis

Three accessions of Brachiaria brizantha, three of B. humidicola, and two interspecific hybrids between B. ruziziensis and B. brizantha were analyzed with regard to their mitotic behavior in root tips. All these genotypes revealed chromosome elimination or lack of chromosome affinity in previous analyses of microsporogenesis. Analyses of root tips showed a normal mitotic division in all accessions and hybrids, reinforcing the notion that the genetic control of meiosis is totally independent of that of mitosis. The implications of these findings for the Brachiaria breeding program are discussed.

Microsporogenesis in sexual Brachiaria hybrids (Poaceae)

Three sexual interspecific hybrids of Brachiaria (HBGC076, HBGC009, and HBGC014) resulting from crosses between B. ruziziensis (female genitor) and B. decumbens and B. brizantha (male genitors) produced by Embrapa Beef Cattle in the 1980s were cytologically analyzed by conventional methods for meiotic studies. The cytogenetic analysis showed the occurrence of common meiotic abnormalities among them. The most frequent abnormalities were those related to irregular chromosome segregation due to polyploidy. Other abnormalities, such as chromosome stickiness, absence of cytokinesis, irregular cytokinesis, abnormal spindle orientation, and abnormal nucleolus disintegration, were found in the three hybrids, while, chromosome disintegration was detected only in HBGC014. All the abnormalities, except for abnormal nucleolus disintegration, can cause unbalanced gamete formation, leading to pollen sterility. Multivalent chromosome association at diakinesis revealed genome affinity between the two parental species in the hybrids, suggesting some possibility for gene introgression. Presently, the Brachiaria breeding program has the objective of releasing, primarily, apomictic hybrids as new cultivars since they do not segregate but preserve the genetic makeup indefinitely. Besides, they result in homogeneous pastures which are easier to manage. The sexual hybrids, however, are paramount in the breeding program: they work as ‘bridges’ to introgress traits of interest into the apomictic genotypes. The cytogenetic analyses of these three hybrids substantiate their maintenance in the breeding program due to low frequency of meiotic abnormalities, complemented by interesting agronomic traits. They may be used in crosses to generate new cultivars in the future.

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

Microsporogenesis was evaluated in the Brachiaria humidicola collection of the Embrapa Beef Cattle Center, represented by 60 accessions. One accession (H121) presented an abnormal pattern of cytokinesis that had never been reported in this genus. Among 900 meiocytes analyzed in the first division, 10.7% underwent precocious and multiple cytokinesis in metaphase I, fractionating the genome and the cytoplasm into two or more parts. The expected cytokinesis after telophase I did not occur. The abnormal meiocytes from the first division entered the second division but the second cytokinesis after telophase II was also abnormal. Among the 857 meiocytes analyzed in the second division, 10.9% presented abnormal, incomplete or total absence of cytokinesis. Dyads and binucleated microspores were recorded among the meiotic products. The use of this accession in the Embrapa breeding program is compromised.

Evidence of programmed cell death during microsporogenesis in an interspecific Brachiaria (Poaceae: Panicoideae: Paniceae) hybrid

Morphological changes have been investigated during plant programmed cell death (PCD) in the last few years due to the new interest in a possible apoptotic-like phenomenon existing in plants. Although PCD has been reported in several tissues and specialized cells in plants, there have been few reports of its occurrence during microsporogenesis. The present study reports a typical process of PCD during meiosis in an interspecific Brachiaria hybrid leading to male sterility. In this hybrid, some inflorescences initiated meiosis but it was arrested in zygotene/pachytene. From this stage, meiocytes underwent a severe alteration in shape showing substantial membrane blebbing; the cytoplasm became denser at the periphery; the cell nucleus entered a progressive stage of chromatin disintegration, and then the nucleolus disintegrated, and the cytoplasm condensed and shrunk. The oldest flowers of the raceme showed only the callose wall in the anthers showing obvious signs of complete sterility.

Microsporogenesis in Brachiaria dictyoneura (Fig. & De Not.) Stapf (Poaceae: Paniceae)

icrosporogenesis was analyzed in five accessions of Brachiaria dictyoneura presenting x = 6 as the basic chromosome number. All accessions were tetraploid (2n = 4x = 24) with chromosome pairing in bi-, tri-, and quadrivalents. The recorded meiotic abnormalities were those typical of polyploids, including precocious chromosome migration to the poles, laggard chromosomes, and micronucleus formation. The frequency of these abnormalities, however, was lower than those reported for other polyploid accessions previously analyzed for other Brachiaria species. Cell fusion and absence of cytokinesis were also recorded in some accessions, leading to restitutional nucleus formation in some cells. Genetically unbalanced microspores, binucleate, and 2n microspores were found among normal meiotic products as results from these abnormalities. The limitation in using these accessions as pollen donor in interspecific crosses with sexual species with x = 7 or x = 9 in breeding programs is discussed.

Evidence of allopolyploidy in Brachiaria brizantha (Poaceae: Paniceae) through chromosome arrangement at metaphase plate during microsporogenesis

In the hexaploid (2n = 6x = 54) accession B176 of Brachiaria brizantha, one cytological characteristic differentiated it from the other accessions previously analyzed with the same ploidy level. Nearly 40% of meiocytes displayed the chromosome set arranged at two metaphase plates at the poles of the cell, close to the membrane. In these cells, both metaphase plates were arranged in an angle to form a typical tripolar spindle. Therefore, cells did not show normal chromosome segregation at anaphase I. Only nine univalent chromosomes migrated from each plate to the opposite pole with the remainder staying immobile on the plate. As a result of such spindle orientation and chromosome behavior, trinucleate telophases I were recorded. After telophase, cytokinesis eliminated the small nuclei into a microcyte. The second division proceeded normally, with the presence of microcytes in all phases. The origin of such an abnormality was explained on the hexaploid level of the accession which could have resulted by chromosome doubling of a triploid derived from species that did not display the same behavior for spindle organization. The high percentage of meiotic abnormalities recorded in this accession compromises fertility and renders it inadequate for the breeding program.

Microsporogenesis in tetraploid accessions of Brachiaria nigropedata (Ficalho & Hiern) Stapf (Gramineae)

The genus Brachiaria (Trin.) Griseb. has achieved considerable importance to cattle production systems, as a result of the good production and adaptation of a few cultivars to poor and acid soils of the Brazilian savannas. Many of its species and accessions are polyploid and apomictic, which limits direct hybridization. To assist the breeding program, cytogenetic characterization has been undertaken on the accessions of Brachiaria collection at the Embrapa Beef Cattle Research Center. In this study, chromosome number and meiotic behavior are reported for the Brachiaria nigropedata (Ficalho & Hiern) Stapf collection. The 20 available accessions are tetraploid (2n = 4x = 36). Chromosomes paired preferentially as bivalents, but quadrivalents were found in high frequencies in some cells. Meiotic behavior was, in general, irregular, and varied among accessions. Most accessions presented more than 20% of abnormal tetrads. The most common meiotic abnormalities were those related to irregular chromosome segregation due to polyploidy, leading to micronuclei formation in the tetrad stage. A low frequency of other meiotic abnormalities such as the absence of cytokinesis, chromosome stickiness, cell fusion, anaphase bridges, and chromosome transfer among microsporocytes were also recorded in some accessions. Limitations of these accessions for use in hybridization programs are discussed.

Symmetric pollen mitosis I and suppression of pollen mitosis II prevent pollen development in Brachiaria jubata (Gramineae)

Microsporogenesis and pollen development were analyzed in a tetraploid (2n = 4x = 36) accession of the forage grass Brachiaria jubata (BRA 007820) from the Embrapa Beef Cattle Brachiaria collection that showed partial male sterility. Microsporocytes and pollen grains were prepared by squashing and staining with 0.5 percent propionic carmine. The meiotic process was typical of polyploids, with precocious chromosome migration to the poles and laggards in both meiosis I and II, resulting in tetrads with micronuclei in some microspores. After callose dissolution, microspores were released into the anther locule and appeared to be normal. Although each microspore initiated its differentiation into a pollen grain, in 11.1 percent of them nucleus polarization was not observed, i.e., pollen mitosis I was symmetric and the typical hemispherical cell plate was not detected. After a central cytokinesis, two equal-sized cells showing equal chromatin condensation and the same nuclear shape and size were formed. Generative cells and vegetative cells could not be distinguished. These cells did not undergo the second pollen mitosis and after completion of pollen wall synthesis each gave rise to a sterile and uninucleate pollen grain. The frequency of abnormal pollen mitosis varied among flowers and also among inflorescences. All plants were equally affected. The absence of fertile sperm cells in a considerable amount of pollen grains in this accession of B. jubata may compromise its use in breeding and could explain, at least in part, why seed production is low when compared with the amount of flowers per raceme.