不同倍性水稻亚种间杂种小孢子发生的细胞学观察

利用塑料半薄切片对水稻同源四倍体亚种间杂种F1及其对应的二倍体杂种F1小孢子母细胞减数分裂过程的细胞学变化进行观察研究.结果表明,同源四倍体水稻亚种间杂种造孢细胞期和小孢子母细胞期已经表现出较高频率的异常;减数分裂过程中,小孢子母细胞出现异常更加复杂,主要包括小孢子母细胞液泡化和退化两大类,这些异常是导致杂种花粉败育和花粉低育性的重要原因之一;此外,绒毡层异常也是导致杂种的花粉育性降低的因素.二倍体杂种小孢子母细胞在减数分裂过程的异常类型与水稻同源四倍体亚种间杂种基本相似,其绒毡层异常频率较低,对其花粉低育性影响不大.     

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 brizantha (Poaceae) as a selection tool for breeding

The genus Brachiaria comprises more than 100 species and is the single most important genus of forage grass in the tropics. Brachiaria brizantha, widely used in Brazilian pastures for beef and dairy production, is native to tropical Africa. As a subsidy to the breeding program underway in Brazil, cytological studies were employed to determine the chromosome number and to evaluate microsporogenesis in 46 accessions of this species available at Embrapa Beef Cattle (Brazil). Thirty-four accessions presented 2n = 36; seven had 2n = 45, and five had 2n = 54 chromosomes. Based on the higher level of chromosome association observed in diakinesis, in tetra-, penta-, and hexavalents, respectively, it was concluded that they are derived from x = 9; consequently, these accessions are tetra- (2n = 4x = 36), penta- (2n = 5x = 45), and hexaploids (2n = 6x = 54). The most common meiotic abnormalities were irregular chromosome segregation due to polyploidy. Chromosome stickiness, abnormal cytokinesis, non-congressed bivalents in metaphase I and chromosomes in metaphase II, and chromosome elimination were recorded at varying frequencies in several accessions. The mean percentage of meiotic abnormalities ranged from 0.36 to 95.76%. All the abnormalities had the potential to affect pollen viability by generating unbalanced gametes. Among the accessions, only the tetraploid ones with less than 40% of abnormalities are suitable as pollen donors in intra- and interspecific crosses. Currently, accessions with a high level of ploidy (5 and 6n) cannot be used as male genitors in crosses because of the lack of sexual female genitors with the same levels of ploidy.     

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

Microsporogenesis 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.     

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.     

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.     

Meiotic behavior in nonaploid accessions of Brachiaria humidicola (Poaceae) and implications for breeding

Brachiaria humidicola is a grass adapted to seasonally swampy grasslands in Africa; two cultivars, 'common' and Llanero, are widely used in Brazilian pastures. New cultivars are in great demand in order to diversify current production systems to achieve improved quality and yield. Cytological analyses of 55 accessions of this species available from the Embrapa Beef Cattle germplasm collection revealed that 27 are apomictic and have 2n = 54 chromosomes. Chromosome pairing as bi- to nonavalent associations at diakinesis indicated a basic chromosome number in this species of x = 6, as found in other closely related Brachiaria species. Thus, these 27 accessions are nonaploid (2n = 9x = 54). Abnormalities were found in the meiosis of these accessions, at variable frequencies. The most common abnormalities were those related to irregular chromosome segregation, which led to unbalanced gamete formation; but chromosome stickiness, cell fusion, and absence of cytokinesis were also recorded. Although some accessions have a low frequency of meiotic abnormalities, ensuring potentially good pollen viability, these cannot be used in hybridization due to a lack of sexual accessions with the same ploidy level.     

The cytological effects of the gametocides Ethrel and RH-531 on microsporogenesis in barley (Hordeum vulgare L)

Abstract. Barley plants ( Hordeum vulgare L. cv. Midas) raised under controlled environmental conditions were sprayed with either of the gametocides Ethrel and RH-531. At various times after spraying the anthers were fixed for light and electron microscopy. Abortion of sporogenous cells occurred in plants sprayed at both pre- and post-meiotic stages of microsporogenesis. In contrast, cells of the tapetum were insensitive to the immediate effects of gametocides. The cytological effects of the gametocides are similar to those induced by male sterile genes in a variety of plants. These range from the induction of additional mitotic divisions in the pollen mother cells to exine malformations on developing microspores. These observations are discussed in terms of the control mechanisms operating during microsporogenesis.     

Microsporogenesis in inbred line of popcorn (Zea mays L.)

Endogamy places genes for several characteristics in homozygosis, which include those related to meiosis causing abnormalities that may impair gamete viability. An original population (S0) of popcorn (CMS-43) produced by Embrapa Maize and Sorghum was self-pollinated for seven years, generating inbred lines (S1 to S7). Conventional studies of microsporogenesis revealed that meiotic abnormalities did not increase with endogamy. Univalent chromosomes, irregular chromosome segregation, abnormal cell shape, partial asynapsis, cell fusion, absence of cytokinesis, abnormal spindle orientation, and chromosome stickiness were recorded in low frequency in meiocytes. Since the frequency of abnormalities was low, mainly in S7, inbred lines from CMS-43 have a high potential for hybridization.     

Meiotic behaviour in three interspecific three-way hybrids between Brachiaria ruziziensis and B. brizantha (Poaceae: Paniceae)

The meiotic behaviour of three three-way interspecific promising hybrids (H17, H27, and H34) was evaluated. These hybrids resulted from the crosses between B. ruziziensis X B. brizantha and crossed to another B. brizantha. Two half-sib hybrids (H27 and H34) presented an aneuploid chromosome number (2n = 4x = 33), whereas hybrid H17 was a tetraploid (2n = 4x = 36), as expected. Chromosome paired predominantly as multivalents suggesting that genetic recombination and introgression of specific target genes from B. brizantha into B. ruziziensis can be expected. Arrangement of parental genomes in distinct metaphase plates was observed in H27 and H34, which have different male genitors. Hybrids H17 and H34 have the same male genitor, but did not display this abnormality. In H17, abnormalities were more frequent from anaphase II, when many laggard chromosomes appeared, suggesting that each genome presented a different genetic control for meiotic phase timing. Despite the phylogenetic proximity among these two species, these three hybrids presented a high frequency of meiotic abnormalities, mainly those related to irregular chromosome segregation typical of polyploids, H34, 69.1%; H27, 56.1% and H17, 44.9%. From the accumulated results obtained through cytological studies in Brachiaria hybrids, it is evident that cytogenetical analysis is of prime importance in determining which genotypes can continue in the process of cultivar development and which can be successfully used in the breeding. Hybrids with high frequency of meiotic abnormalities can seriously compromise seed production, a key trait in assuring adoption of a new apomictic cultivar of Brachiaria for pasture formation.     

A rise of ploidy level influences the rate of cytomixis in tobacco male meiosis

The effect of plant ploidy level on the rate of cytomixis in microsporogenesis has been analyzed with the help of a unique model, the collection of tobacco plants of different ploidies (2n?=?2x?=?24, 4x?=?48, 6x?=?72, and 8x?=?96). As has been shown, the rate of cytomixis proportionally increases in 6x and 8x cytotypes, being rather similar in 2x and 4x plants. The rate of cytomixis is highly variable, differing even in the genetically identical plants grown under the same conditions. The cytological pattern of cytomixis in the microsporogenesis of control 4x plants has been compared with the corresponding patterns of 2x, 6x, and 8x plants. Involvement of cytomixis in production of unreduced gametes and stabilization of the newly formed hybrid and polyploidy genomes is discussed.     

EMS-induced cytomictic variability in safflower (Carthamus tinctorius L.)

Seeds of safflower (Carthamus tinctorius L.) were subjected to three treatment durations (3h, 5h and 7h) of 0.5 % Ethyl Methane Sulphonate (EMS). Microsporogenesis was carried out in the control as well as in the treated materials. EMS treated plants showed interesting feature of partial inter-meiocyte chromatin migration through channel formation, beak formation or direct cell fusion. Another interesting feature noticed during the study was the fusion among tetrads due to wall dissolution. The phenomenon of cytomixis was recorded at nearly all the stages of microsporogenesis connecting from a few to several meiocytes. Other abnormalities such as laggards, precocious movement, bridge and non-disjunction of chromosomes were also recorded but in very low frequencies. The phenomenon of cytomixis increased along with the increase in treatment duration of EMS. Cells with these types of cytomictic disturbances may probably result in uneven formation of gametes or zygote, heterogenous sized pollen grains or even loss of fertility in future.     

Low phosphorus tolerance mechanisms: phosphorus recycling and photosynthate partitioning in the tropical forage grass, Brachiaria hybrid cultivar Mulato compared with rice

The Brachiaria hybrid cv. Mulato is well adapted to low-fertility acid soils deficient in phosphorus (P). To study the grassy forage's mechanisms for tolerating low P supply, we compared it with rice (Oryza sativa L. cv. Kitaake). We tested by using nutrient solution cultures, and quantified the effects of P deficiency on the enzymatic activities of phosphohydrolases and on carbon metabolism in P-deficient leaves. While P deficiency markedly induced activity of phosphohydrolases in both crops, the ratio of inorganic phosphorus to total P in leaves was greater in Brachiaria hybrid. Phosphorus deficiency in leaves also markedly influenced the partitioning of carbon in both crops. In the Brachiaria hybrid, compared with rice, the smaller proportion of (14)C partitioned into sugars and the larger proportion into amino acids and organic acids in leaves coincided with decreased levels of sucrose and starch. Hence, in P-deficient leaves of the Brachiaria hybrid, triose-P was metabolized into amino acids or organic acids. Results thus indicate that the Brachiaria hybrid, compared with rice, tolerates low P supply to leaves by enhancing sugar catabolism and by inducing the activity of several phosphohydrolases. This apparently causes rapid P turnover and enables the Brachiaria hybrid to use P more efficiently.     

Impaction onto a Glass Slide or Agar versus Impingement into a Liquid for the Collection and Recovery of Airborne Microorganisms

To study impaction versus impingement for the collection and recovery of viable airborne microorganisms, three new bioaerosol samplers have been designed and built. They differ from each other by the medium onto which the bioaerosol particles are collected (glass, agar, and liquid) but have the same inlet and collection geometries and the same sampling flow rate. The bioaerosol concentrations recorded by three different collection techniques have been compared with each other: impaction onto a glass slide, impaction onto an agar medium, and impingement into a liquid. It was found that the particle collection efficiency of agar slide impaction depends on the concentration of agar in the collection medium and on the sampling time, when samples are collected on a nonmoving agar slide. Impingement into a liquid showed anomalous behavior with respect to the sampling flow rate. Optimal sampling conditions in which all three new samplers exhibit the same overall sampling efficiency for nonbiological particles have been established. Inlet and collection efficiencies of about 100% have been achieved for all three devices at a sampling flow rate of 10 liters/min. The new agar slide impactor and the new impinger were then used to study the biological factors affecting the overall sampling efficiency. Laboratory experiments on the total recovery of a typical environmental microorganism, Pseudomonas fluorescens ATCC 13525, showed that both sampling methods, impaction and impingement, provided essentially the same total recovery when relatively nonstressed microorganisms were sampled under optimal sampling conditions. Comparison tests of the newly developed bioaerosol samplers with those commercially available showed that the incorporation of our research findings into the design of the new samplers yields better performance data than data from currently available samplers.     

Abnormal male meiosis explains pollen sterility in the polyploid medicinal plant Pinellia ternata (Araceae)

Pinellia ternata is an important traditional Chinese medicinal plant. Its different populations in China have various ploidy levels, based on x = 13, as well as extensive aneuploid series. The microsporogenesis process was observed in specimens from three populations from three regions of Hubei Province; they were characterized by normal and abnormal meiotic divisions in pollen mother cells (PMCs) at all stages simultaneously. Meiotic abnormalities including univalents/multivalents, chromosomal laggards/bridges and micronuclei appeared in about 50% of the PMCs, together with abnormal cytokinesis. Chromatin/chromosome transfer between meiocytes occurred only during the first division, at low frequency; this might contribute to these meiotic abnormalities. Although the remaining 50% of the PMCs presented normal cytological behavior, pollen fertility was only about 2%. These results provide cytological explanations for its low seed-set and the general use of asexual reproduction through tubers and bulbils; it also explains the wide variations in chromosome number.     

Genome elimination during microsporogenesis in two pentaploid accessions of Brachiaria decumbens (Poaceae)

Polyploidy is a prominent and significant force in plant evolution, taking place since ancient times and continuing until today. Recent cytogenetic studies in the genus Brachiaria using germplasm collected from wild African savannas in the 1980s revealed that most species and accessions within species are polyploid. Diploid, tetraploid, and pentaploid accessions have been found. We found asynchronous meiosis during microsporogenesis, followed by genome elimination, in two pentaploid (2n = 5x = 45) accessions (D53 and D71) of a hardy, invasive pasture grass, introduced from Africa to Brazil, Brachiaria decumbens. In these accessions, chromosomes paired as 18 bivalents and nine univalents during diakinesis, suggesting that these accessions resulted from a recent event of natural hybridization. The lack of chromosome associations in the genomes suggests that these accessions resulted from hybridization between two genotypes that are not closely related, with low genome affinity and with different meiotic rhythms. This supposition is reinforced by the meiotic behavior of the nine univalents, which were always laggard in relation to the other chromosomes and eliminated as micronuclei in microspores. The behavior of these accessions, which have an odd level of ploidy and confirmed genome elimination, supports the general assumption that a polyploid accession can undergo a new event of polyploidization by natural hybridization (neopolyploidyzation). This evidence for natural hybridization in Brachiaria shows that this is a wild genus in an ongoing evolutionary process.     

Meiotic studies of the F1 hybrid between rice bean (Vigna umbellata) and its wild relative V. minima

The interspecific F₁ hybrid V. umbellata x V. minima, together with the parental species was investigated cytogenetically. Both parents showed normal meiosis, with 11 bivalents at diakinesis. The hybrid was completely pollen sterile. The cytological causes underlying sterility in the hybrid include an array of meiotic abnormalities ranging from non-pairing of chromosomes through the presence of univalents, chain and ring multivalents and anaphase bridges to the formation of micronuclei, during microsporogenesis. The presence of chain and ring multivalents at diakinesis not only demonstrates the structural hybridity of the F₁ hybrid, but also suggests that small interchanges and inversions played a significant role in the speciation within the genus Vigna.     

Abnormalities induced by agricultural pesticides in the microsporogenesis of olive tree (Olea europaea L.) cultivars

The study evaluated the microsporogenesis of olive trees subjected to different agricultural pesticide applications during flowering. Inflorescences of cultivars Arbequina and MGS GRAP541 were subjected to agricultural pesticides: mineral oil, neem oil, dimethoate and deltamethrin. The floral buds were fixed in Carnoy for the microsporogenesis analysis and in Karnovsky for scanning electron microscopy. The slides were prepared by squash technique and staining with propionic carmine. The pollen viability was determined by Alexander’s stain and in vitro germination. Results show that the quantification of abnormalities in meiosis in the two cultivars caused significant effect among the treatments, being that all differed statistically from the control group. Both methods showed a higher percentage of viable pollens in the control treatment and lower percentage of viability with the agricultural pesticides. The method of pollen viability by staining presented the highest averages of viable pollens, but when compared together, both methods presented a strongly related positive linear correlation. It was concluded that the used chemical products increased the percentage of chromosomal abnormalities during microsporogenesis, which interfered in the pollen viability of the two analyzed cultivars. The product deltamethrin caused the strongest effect on meiosis and on pollen viability.     

EMS-induced cytomictic variability in safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.)

Seeds of safflower (Carthamus tinctorius L.) were subjected to three treatment durations (3, 5 and 7 h) of 0.5% Ethyl Methane Sulphonate (EMS). Microsporogenesis was carried out in the control as well as in the treated materials. EMS treated plants showed interesting feature of partial inter-meiocyte chromatin migration through channel formation, beak formation or direct cell fusion. Another interesting feature noticed during the study was the fusion among tetrads due to wall dissolution. The phenomenon of cytomixis was recorded at nearly all the stages of microsporogenesis connecting from a few to several meiocytes. Other abnormalities such as laggards, precocious movement, bridge and non-disjunction of chromosomes were also recorded but in very low frequencies. The phenomenon of cytomixis increased along with the increase in treatment duration of EMS. Cells with these types of cytomictic disturbances may probably result in uneven formation of gametes or zygote, heterogenous sized pollen grains or even loss of fertility in future.     

Postpollination drop production in hybrid larch is not related to the diurnal pattern of xylem water potential

The hourly production of postpollination drops in the ovules of three hybrid larch trees (Larix x. marschlinsii Coaz) was examined and compared with the measured diurnal pattern of xylem water potential of the same trees under non-stressed conditions during two reproductive seasons. There was no consistent relationship between xylem water potential and ovular drop production in hybrid larch. Individual trees that showed a diurnal drop production in one year did not follow a similar pattern the other year. One tree produced drops in over half of its ovules hourly, regardless of the measured xylem water potential. Variation in number of drops produced per ovule and drop mass was observed between the three trees both years, and between years for two of the trees. This variation was not in response to xylem water potential or environmental conditions. Here we report the first simultaneous measurements of branch water status and the production of ovular secretions for any conifer. We conclude that the postpollination drop production of hybrid larch is not regulated by a tree's overall water status, but is under the control of localized structures such as the cone or the ovule.