Cytogenetic data for Paspalum notatum Flügge accessions

Dados citogenéticos para acessos de Paspalum notatum Flügge

GENETICS AND PLANT BREEDING

Cytogenetic data for Paspalum notatum Flügge accessions

Dados citogenéticos para acessos de Paspalum notatum Flügge

Nair Dahmer; Maria Teresa Schifino-Wittmann^*; Miguel Dall'Agnol; Biane de Castro

UFRGS/FA - Depto. de Plantas Forrageiras e Agrometeorologia, C.P. 15100 - 91501-970 - Porto Alegre, RS - Brasil

ABSTRACT

Several species of the genus Paspalum L. are important forages, due their to quality, productivity and tolerance to environmental stresses. Chromosome numbers, meiotic configurations and pollen fertility were evaluated in a collection of 85 accessions of Paspalum notatum Flügge and in seven accessions of Pensacola (P. notatum var saurae). All P. notatum accessions were tetraploid, with 2n = 4x = 40, except one diploid accession, considered as an escape of Pensacola. All Pensacola plants had 2n = 2x = 20. Meiotic configurations at diakinesis and metaphase I varied among tetraploid accessions, from plants with only bivalents to plants with high frequency of quadrivalents. Pollen fertility varied from 82.5 to 95.9% among diploid accessions and from 72.4 to 97.9% among the tetraploids. Due to the apomictic mode of reproduction of tetraploid P. notatum, meiotic irregularities can be maintained by plants without harming their propagation. At the same time, pollen fertility should be high enough to assure endosperm development, since the species is pseudogamous. Wild diploid P. notatum populations, apart from the endemic P. notatum var saurae, are very rare. From a plant breeding point of view, all the examined tetraploid accessions are potentially male-fertile and could be used as males in crosses.

Key words: apomixis, chromosome numbers, meiotic configurations, plant breeding, pollen fertility

RESUMO

Muitas espécies do genus Paspalum L. são importantes forragens, devido à qualidade, produtividade e tolerância para o stress ambiental. Números cromossômicos, configurações meióticas e fertilidade do pólen foram analisados em uma coleção de 85 acessos de Paspalum notatum Flügge e sete acessos de Pensacola (P. notatum var saurae). Todos os acessos de P. notatum eram tetraplóides, com 2n = 4x = 40, com exceção de um diplóide, considerado como escape de Pensacola. Todas as plantas de Pensacola examinadas tinham 2n = 2x = 20. Foi verificada variação entre os acessos tetraplóides quanto às configurações cromossômicas em diacinese e metáfase I, desde plantas com apenas bivalentes até aquelas com alta freqüência de quadrivalentes. A fertilidade do pólen variou de 82,5 a 95,9% entre os acessos diplóides e de 72,4 a 97,9% entre os tetraplóides. Devido ao modo de reprodução apomítico de P. notatum tetraplóide, irregularidades meióticas podem ser mantidas sem prejuízo da propagação. Ao mesmo tempo, a fertilidade do pólen deve ser suficientemente alta para assegurar a formação do endosperma, já que a espécie é pseudogâmica. Diplóides silvestres de P. notatum, além do endêmico P. notatum var saurae são muito raros. Do ponto de vista do melhoramento, todos os tetraplóides são potencialmente macho-férteis e poderiam ser utilizados como genitores masculinos em cruzamentos.

Palavras-chave: apomixia, números cromossômicos, configurações meióticas, melhoramento genético, fertilidade do pólen

INTRODUCTION

The genus Paspalum L. comprises around 400 species, most of them native to tropical and sub-tropical Americas (Moraes-Fernandes et al., 1968). Many of these species are important forages, due to their quality, productivity and tolerance to environmental stresses (Valls, 2000). In Brazil, where 75% of Paspalum species occur, they are part of several plant communities, in a broad range of ecological conditions (Barreto, 1974). Sexual reproduction and apomixis are common in the genus, generally related to diploid and polyploid levels (Quarín & Norrmann, 1990).

P. notatum Flügge is an apomictic tetraploid (2n = 4x = 40), common to native pastures along warm and temperate American regions, from Central Mexico to Uruguay (Burton, 1948; Burton & Forbes, 1960; Quarín et al., 1984) and it has been studied by botanists, geneticists, agronomists and plant breeders (Dahmer-Balbinot, 2007). P. notatum var. saurae is a sexual diploid (2n = 2x = 20), native to northeastern and central-eastern Argentina, introduced around 1926 to the Pensacola Bay region of Florida and later released as the Pensacola cultivar (Burton, 1967; 1974). It is widely cultivated in North and South America. Since the apomictic nature of P. notatum precludes its use as female in crosses (but not as male as there is no fertile pollen), the introduction of variability by sexual methods was possible by crossing the apomictic with artificially polyploidized P. notatum var. saurae (Burton & Forbes, 1960; Burton et al., 1970). If other wild diploids were identified, they could have been used to introduce novel variation in P. notatum, but, until now, no other diploids have been located outside the Argentinian region of P. notatum var. saurae (Pozzobon & Valls, 1997; Daurelio et al., 2004). The objective of the present work was to determine chromosome numbers, chromosome associations at diakinesis and metaphase I, and pollen viability in accessions of P. notatum which are part of a Brazilian breeding project.

MATERIAL AND METHODS

The original plants (accessions) were collected as tillers (propagated later vegetatively in pots) from populations in the Rio Grande do Sul State, Brazil. A few accessions came from the Santa Catarina and Paraná States (Brazil), Uruguay and Argentina (Table 1). Plants are kept in pots as part of a germplasm collection. The accessions are identified in the collection by species name and a number, for working purposes. In the collection list all accessions are registered by this number plus the collector´s number and place of collection.

Somatic chromosome numbers (2n) were determined in root-tip cells pre-treated with a saturated paradichlorobenzene solution for 24 h at 4°C, fixed in Carnoy 3:1 (ethanol:acetic acid) for 24 h at 4°C, hydrolyzed with 1 mol L^-1 HCl for 10 min at 60ºC, stained with Feulgen and squashed in propionic carmine. At least ten cells with good spreading and no chromosome overlapping were counted per plant. Gametic chromosome numbers (n) were determined in pollen-mother-cells undergoing meiosis, from young inflorescences fixed in Carnoy 3:1 at room temperature and squashed in propionic carmine. Most of the gametic chromosome counts were performed in cells at diakinesis and metaphase I stages, since at these stages also chromosome pairing configurations may clearly be observed.

Pollen fertility was estimated by stainability (in 2% propionic carmine, following Simioni et al., 2006) of 1500 mature pollen grains per plant. Full, well stained grains were classified as potentially viable, and unstained or poorly stained as sterile.

RESULTS AND DISCUSSION

Chromosome numbers were determined in a total of 92 accessions (Table 1), and, 83 of them were tetraploid, with 2n = 4x = 40 (or n = 20) (Figures 1A, B, C). One accession, originally collected as P. notatum had 2n = 2x = 20 (P. notatum 12) (Table 1, Figures 1G, H). The seven Pensacola accessions were diploid, with 2n = 2x = 20 (or n = 10), as expected (Table 1).

The P. notatum 12 accession with 2n = 20 is most probably an escape of the cultivated Pensacola. Despite having been collected among plants of P. notatum, Pensacola is cultivated in the region and most probably some plants of the cultivated variety have invaded natural fields.

Pozzobon & Valls (1997) reported 11 accessions with 2n = 20 among the 127 P. notatum accessions and considered these plants to be escapes from Pensacola cultivation. Moraes-Fernandes et al. (1973) found only the tetraploid level among 16 Brazilian (Rio Grande do Sul) and one introduced accession from P. notatum. Daurelio et al. (2004) reported that despite a search in other regions of Argentina, the diploids are all botanically P. notatum var. saurae and restricted to Northeastern and Central-Eastern Argentina. In this restricted region, but not further, these authors detected some triploid hybrids between P. notatum and P. notatum var. saurae.

Wild diploid cytotypes are most probably restricted to this specific Argentinian region. If "true" diploid P. notatum plants, apart from Pensacola and its escapes, exist in other geographical areas they are certainly extremely rare. To find other possible wild diploids in other regions would imply in collecting a high number of accessions along P. notatum distribution and would require other methods than traditional chromosome countings (that are laborious and time consuming) to estimate ploidy levels, such as flow cytometry that allows examination of a large number of plants in less time. However, once Pensacola is also widely cultivated, these studies should be coupled to a sound taxonomic determination, detailed morphological analysis and, if possible other approaches as molecular markers profiling, in order to avoid misidentifications and wrong assignments of Pensacola escapes as wild P. notatum.

Despite the absence of chromosome number variability among the P. notatum accessions, when examining meiotic configurations (Table 1), the 36 tetraploid accessions presented several chromosome configurations, with the occurrence of univalents (I), bivalents (II), trivalents (III), and quadrivalents (IV) (Table 1, Figures 1E, F). The frequencies of different chromosome configurations varied widely among different accessions. In P. notatum 06, 07, 13, 17 and 49 all the cells presented only bivalents. On the other hand, in P. notatum-14, 75% of the cells presented other configurations than bivalents, and these percentages were 93.1% for P. notatum-33, 80.9% for P. notatum-Bagual and 85.0% for P. notatum-André da Rocha. In general, the most common configurations among cells were 20 II or 18 II and 1 IV (Table 1). Therefore, the absence of chromosome number variability is accompanied by the variability in meiotic configurations among the tetraploid accessions. Conversely, the diploid accessions presented always 10 II at diakinesis and metaphase I (7% of the cells of accession 12) (Table 1, Figures 1H, I). Moraes-Fernandes et al. (1973) also found varying chromosome configurations among the examined accessions of P. notatum, with predominance of quadrivalents and bivalents, while trivalents and univalents were less frequent. One accession presented a predominance of cells with just bivalents (Moraes-Fernandes et al., 1973). Variability in chromosome configurations in other Paspalum species have been reported by other authors such as Moraes-Fernandes et al. (1968), Pagliarini et al. (2001) and Adamowski et al. (2005).

Pollen viability was mostly over 80% in 48 accessions and ranged from 82.47 to 95.93% among diploid accessions and from 72.40 to 97.93% among tetraploids (Table 1). Moraes-Fernandes et al. (1973) reported pollen fertility (ascertained with acetic carmine) ranging from 0 to 84.3% in the tetraploid P. notatum.

The variability observed among the P. notatum accessions may be explained by the polyploid nature of the species and by its mode of reproduction. Tetraploid P. notatum is apomictic and also propagates vegetatively, therefore the quadrivalents remaining from the polyploidization evolutionary process could be maintained (without need for chromosome "diploidization", a process known to occur widely among sexual tetraploids). Moreover, the eventual chromosomal rearrangements that lead to quadrivalents, such as translocations, or other configurations like trivalents and unpaired chromosomes (univalents) could be maintained by the accessions without harming their assexual reproduction. At the same time, since P. notatum is a pseudogamous apomictic, that is, needs pollination to form the endosperm, a certain degree of meiotic regularity leading to at least some pollen fertility is necessary and that would explain the relatively high pollen fertility that was observed. It could be speculated that this kind of situation could represent a genetic and evolutionary conflict: apomictic reproduction with the possibility of accumulating meiotic irregularities versus the need of fertile pollen.

From a plant breeding point of view, all the examined accessions are potentially male-fertile and could be used as male genitors in crosses. However, as no other diploids apart from the endemic P. notatum var. saurae have been identified, improvement of tetraploid P. notatum should rely on the selection of well adapted apomictic populations. To introduce variation through sexual crosses would imply in looking for wild P. notatum diploid cytotypes, to use other plants of P. notatum var. saurae from Argentina or even to employ Pensacola plants. Before looking for new diploids, a better option for the plant breeder would be first to see if the desirable characteristics could be found in known diploids.

CONCLUSIONS

Wild diploid cytotypes, apart from P. notatum var. saurae are extremely rare. Despite no variability was found in chromosome numbers among tetraploid P. notatum, there is a variability in meiotic chromosome associations among accessions, explained by the apomictic kind of reproduction. Pollen fertility among apomicts is relatively high, assuring endosperm formation and allowing the potential use of these plants as males in crosses.

ACKNOWLEDGMENTS

To the Bazilian institutions CNPq, FAPERGS and CAPES for financial support and fellowships. To Dr. José Francisco Montenegro Valls (EMBRAPA-CENARGEN), Dr. Carlos Nabinger and MSc Marcelo Steiner (UFRGS) for providing some of the accessions.

Received April 24, 2007

Accepted December 07, 2007

• ADAMOWSKI, E.V.; PAGLIARINI, M.S.; BONATO, A.B.; BATISTA, L.A.R.; VALLS, J.F.M. Chromosome numbers and meiotic behavior of some Paspalum accessions. Genetics and Molecular Biology, v.28, p.773-780, 2005.
• BARRETO, I.L. O gênero Paspalum (Gramineae) no Rio Grande do Sul. Porto Alegre: UFRGS, 1974. 258 p. Tese (Livre Docência).
• BURTON, G.W. A search for the origin of Pensacola Bahiagrass. Economic Botany, v.21, p.319-328, 1967.
• BURTON, G.W. Recurrent restricted phenotypic selection increases forage yield of Pensacola Bahiagrass. Crop Science, v.14, p.831-835, 1974.
• BURTON, G.W. The method of reproduction of common Bahiagrass, Paspalum notatum. Journal of the American Society of Agronomy, v.40, p.443-452, 1948.
• BURTON, G.W.; FORBES JR., I.; JACKSON, J. Effect of ploidy on fertility and heterosis in Pensacola Bahiagrass. Crop Science, v.10, p.63-65, 1970.
• BURTON, G.W.; FORBES, I. Genetics and manipulation of obligate apomixis in common Bahia grass (Paspalum notatum Flügge). In: INTERNATIONAL GRASSLAND CONGRESS, 8., Reading, 1960. Proceedings Reading: International Grassland Society, 1960. p.66-71.
• DAHMER-BALBINOT, N. Variabilidade citogenética em uma coleção de acessos de Paspalum notatum Flügge. Porto Alegre: UFRGS, 2007. 71p. Dissertação (Mestrado).
• DAURELIO, L.D.; ESPINOZA, F.; QUARÍN, C.L.; PESSINO, S.C. Genetic diversity in sexual diploid and apomictic tetraploid populations of Paspalum notatum situated in sympatry or allopatry. Plant Systematics and Evolution, v.244, p.189-199, 2004.
• MORAES-FERNANDES, M.I.B.; BARRETO, I.L.; SALZANO, F.M. Cytogenetic, ecologic and morphologic studies in Brazilian forms of Paspalum dilatatum Canadian Journal of Genetics and Cytology, v.10, p.131-138, 1968.
• MORAES-FERNANDES, M.I.B.; BARRETO, I.L.; SALZANO, F.M. Cytogenetic, ecologic and morphologic studies in Brazilian forms of Paspalum notatum Canadian Journal of Genetics and Cytology, v.15, p.523-531, 1973.
• PAGLIARINI, M.S.; CARRARO, L.R.; FREITAS, P.M.; ADAMOWSKI, E.V.; BATISTA, L.A.R.; VALLS, J.F.M. Cytogenetic characterization of Brazilian Paspalum accessions. Hereditas, v.135, p.27-34, 2001.
• POZZOBON, M.T.; VALLS, J.F.M. Chromosome number in germplasm accessions of Paspalum notatum (Gramineae). Brazilian Journal of Genetics, v.20, p.29-34, 1997.
• QUARÍN, C.L.; NORRMANN, G.A. Interspecific hybrids between five Paspalum species. Botanical Gazette, v.151, p.366-369, 1990.
• QUARÍN, C.L.; BURSON, B.L.; BURTON, G.W. Cytology of intra- and interspecific hybrids between two cytotypes of Paspalum notatum and P. cromyorrhizon Botanical Gazette, v.145, p.420-426, 1984.
• SIMIONI, C.; SCHIFINO-WITTMANN, M.T.; DALL´AGNOL, M. Sexual polyploidization in red clover. Scientia Agricola, v.63, p.26-31, 2006.
• VALLS, J.F.M. Impacto do conhecimento citogenético na taxonomia de Paspalum e Axonopus (Gramineae). In: CAVALCANTI, T.B.; WALTER, B.M.T. (Ed.) Tópicos atuais em botânica. Brasília: EMBRAPA-CENARGEN, 2000. p.15-21.

Publication Dates

• Publication in this collection
  21 July 2008
• Date of issue
  2008

History

• Accepted
  07 Dec 2007
• Received
  24 Apr 2007