Breeding and selection of Brachiaria

Cacilda Borges do Valle 1 , Rosangela Maria Simeo Resende2, Liana Jank1, Valria
TPF FPT P P P P

Pacheco Batista Euclides1, Jos Raul Valrio1, Manuel Cludio Motta Macedo1,
P P P P P P

Rodrigo Amorim Barbosa1, and Celso Dornelas Fernandes1

P P P P

Introduction
Animal production in the tropics is commonly carried out on native and cultivated pastures. Presently, extensive production has given place to sustainable utilization of natural resources, optimizing inputs and income as a result of ecological, sociological, economical and market demands. Thus, cultivated pastures, previously upon marginal areas in the farms, now compete with crops for fertilizers, technology and integrate crop-livestock systems. Better, more productive pastures for more efficient systems require improved cultivars, thus justifying investment in breeding and selection programs. According to the last Brazilian census, cultivated pastures cover 50% of the pasture area in Brazil, or 100 million hectares (Censo Agropecurio, 1995-1996). This area has doubled in the last 20 years (~100 million hectares), due largely to the use of Brachiaria and Panicum grasses, which have placed Brazil as the largest exporter and second largest producer of beef in the world. The diversity of pastures, however, is dangerously restricted to a few cultivars of grasses that reproduce asexually through apomixis. Two cultivars of Brachiaria respond for 49% of the forage seed produced in Brazil and 85% of the seed commercialized in the savanna region. After about three decades of use, approximately 80% of the 49 million hectares of cultivated pastures in the Cerrados have varying degrees of resource degradation (soil/plant) and offer only marginal economic returns (Macedo, 2001). Renovation of these areas has become a major trend, either by alternating cycles of crops and pastures (also grass-legume pastures) or by altogether substitution of the pasture species. In response to this demand, research as well as extension programs are
Agronomist, Embrapa Beef Cattle, P.O. Box 154, Campo Grande, MS, 79002-970, CNPq scholarship. cacilda@cnpgc.embrapa.br; liana@cnpgc.embrapa.br; val@cnpgc.embrapa.br; macedo@cnpgc.embrapa.br; celsof@cnpgc.embrapa.br 2 Agronomist, Embrapa Beef Cattle, P.O. Box 154, Campo Grande, MS, 79002-970. jraul@cnpgc.embrapa.br 3 Biologist, Embrapa Beef Cattle, P.O. Box 154, Campo Grande, MS, 79002-970, rosangela@cnpgc.embrapa.br
TP PT P P P P

eagerly demanding specifically adapted forage cultivars. Forage cultivars for direct seeding are also needed and are not necessarily the same as for direct grazing or short cycle rotation with crops. The lack of choice of cultivars, in the case of grasses is due to the nature of the material utilized: exotic genera, mainly from Africa, such as Brachiaria and Panicum, with no natural variability amenable to selection in Brazil, and most of the genotypes exhibiting polyploidy and apomictic reproduction which requires more complex breeding and selection procedures. The introduction of large germplasm collections of Brachiaria, in the 1980s, by Embrapa Beef Cattle Center prompted efforts to select for new cultivars using two paths: direct selection from the natural diversity present in the germplasm and generation of new diversity through hybridization followed by selection for specific traits, as will be detailed later on. The breeding program has established the objectives of identifying genotypes adapted to soils of the savannas, with good productivity, resistance to insects, good nutritive value and high seed production.

Historical background on Brachiaria

The introduction of several African grasses to America happened during colonial times, probably as bedding on slave ships as describes by Parsons (1972). From those times are guineagrass (`colonio`), jaraguagrass (Hyparrhenia), molassesgrass (Melinis), pangola digitgrasss and two Brachiaria: B. mutica (paragrass), and B. plantaginea (marmeladegrass). The adaptation of these grasses to the soil and climate of tropical America was so good that all of them are frequently considered naturalized today throughout the continent. In Brazil, the first official introduction was of B. decumbens by the Instituto de Pesquisa Agropecuria do Norte (IPEAN) in Belm, in 1952, under the name of B. brizantha, coming from Suriname (Serro and Simo Neto, 1971). In 1965 there was another introduction from this same material, together with a true B. brizantha. As seed production was very poor, these genotypes were frequently distributed to other states by cuttings. The variety of B. decumbens became known as IPEAN, as has little economic importance nowadays. A second variety of B. decumbens, originally from Uganda, was introduced to So Paulo by the International Research Institute (IRI) in Mato in the beginning of the 60s (Sendulski, 1978), and became known as australiana. It was later registered under the name of Basilisk in 1973, in Australia (Mackay, 1982).

In the early 70s, results from agronomic evaluations already pointed to the potential of this ecotype. Through Government projects such as CONDEPE and PROPASTO, pasture establishment using seed imported from Australia and funds from international banks was promoted throughout central Brazil and large monocultures of B. decumbens were then planted (Pizarro et al., 1996). Until today cattlemen refer to an era before and another after the arrival of Brachiaria in the central plateau of Brazil. Sure enough, problems began to arise such as photosensitization in calves, young steers and heifers, burning of pastures due to massive spittlebugs attacks, especially in the Amazonian basin. Research groups began looking for substitute ecotypes and B. humidicola, more tolerant to the insect gained room in substitution to B. decumbens in the North. In 1967, IRI introduced a B. brizantha from Zimbabwe, which after further evaluation became cv. Marandu, released by Embrapa in 1984 (Nunes et al., 1984). This cultivar is by far the most extensively planted in the tropical world today, becoming the new monoculture. In the early 70s, the high cost of the imported seed (U$10/kg of only 10% pure live seed) prompted producers to experiment harvesting their own seed from the ground to start new pastures (Santos Filho, 1996). By the end of the decade, there was no need to import more seed and by the same token the seed industry began to develop its technology and machinery, responsible for making Brazil the largest exporter of forage seed to all of Latin America. A new era began in Brazil when a diverse germplasm bank of recently collected ecotypes was imported. The International Center for Agriculture Research (CIAT, 1985) gathered about 500 ecotypes in colleting trips in East Africa during 1984 and 1985. An agreement was signed between Embrapa and CIAT and the collection started arriving in Brasilia (Embrapa Genetic Resources and Biotechnology for quarantine and then Embrapa Cerrados for cultivation and evaluation) in 1986. It was later transferred to Embrapa Beef Cattle were it is being maintained live in plots, since many ecotypes of lesser known species (B. jubata, B. nigropedata, B. platynota, B. arrecta) are very poor seeders. A vigorous selection and breeding program based on this extraordinary diversity could thus begin.

The Brachiaria program

Brachiaria has shown remarkable plasticity in adapting to poor and acid soils of the Brazilian savannas, in a variety of climates and altitudes, competing well with weeds and displaying good animal performance. The species of economic importance to Brazil (B. decumbens, B. brizantha, B. ruziziensis and B. humidicola) are originally

from east and south of Africa (Renvoize et al., 1996). The rapid expansion of B. decumbens cv. Basilisk, through the tropics in the 1970s can be explained by its good adaptation to poor, acid soils, to a diversity of climate and latitudes, aggressive competition with weeds and specially, good animal performance (Bogdan, 1977). Until the diverse germplasm became available in 1986, selection was very restricted and new cultivars, impossible to obtain. The development of new cultivars is a long term process which may involve selection from natural diversity and generation of novel diversity through hybridization. It is necessarily a team work, where breeders, phytopathologists, entomologists, animal nutrition and soil fertility specialists, work together in order to identify those superior genotypes that will impact production systems with better animal performance and sustainable pasture production. Figure 1 illustrates t activities and stages of evaluation in the Brachiaria program at Embrapa Beef Cattle. Basic studies were initiated in 1986 at Embrapa Beef Cattle, and ranged from chromosome numbers and mode of reproduction to morphological and agronomical characterization (Valle and Savidan, 1996; Valle et al., 1993, 1999, 2001). Only one species of economic importance has obligate sexual reproduction and is diploid (B. ruziziensis, 2n=18), whereas the majority of the accessions in the other species are tetraploid and apomictic. Other sexual and diploid accessions in B. brizantha and B. decumbens have just recently been tetraploidized to allow intraspecific crosses. B. humidicola has just one sexual tetraploid accession and in 2005 the first hybrids were produced at this ploidy level. Pentaploid and higher ploidies have been identified (Penteado et al., 2000; Letteriello et al., 1999). The study of chromosome behavior in meiosis, carried out in the laboratories of the State University of Maring, has produced an amazing amount of information as was presented in another session in this event. Their results are invaluable in the selection of genitors and of hybrids with fewer abnormalities to continue in the program and also those that should be discarded due to infertility. They have detected irregularities and mutations that could impair the use of several accessions and hybrids in further breeding and cultivar development (Pagliarini et al., 2007 - this volume). Agronomic evaluation carried out since 1988 revealed wide variation among species and ecotypes within species: B. brizantha has the most promising accessions, with good total production (TDM), leaf dry matter production (LDM),

Characterization: ploidy, mode of reproduction, diversity (molecular markers, cytogenetics)

Selected Genitors
~ 30 Sex & Apo

Induction to flower, polyploidization, cytogenetics, hybridization & selection

Hybrid Population
~ 1000 genotypes

Germplasm
~ 500 accessions
Cuttings or seeds

Selected Hybrids

Ploidy determination, mode of reproduction, identity (RAPD, SSR), cytogenetics, physical, chemical, anatomical and biological characterization

Stage 1
100-200
Selection and seed multiplication

Plots: Production, Adaptation, Regrowth, Nutritive Value. Greenhouse and lab: Pests and Diseases, Seed production, Al tolerance, Flooding

Regional Trials 20-25 genotypes

Seed multiplication

Plots with selected genotypes: Production, Adaptation, Regrowth, Nutritive Value, Insects, Seeds

Stage 2
8 10 genotypes
Seed multiplication Paddocks under grazing: tolerance to grazing, palatability carrying capacity, regrowth after grazing

Stage 3
2 - 4 genotypes
seed multiplication

Pastures under grazing with elite genotypes: forage offer, animal performance, persistence, carrying capacity, stocking rate, insects, nutritive value, consumption

NEW

CULTIVAR

RELEASE & ADOPTION

Figure 1. Stages of cultivar development in the Brachiaria program

percentage production in the dry season (%DS), fast regrowth (R), leaf:culm proportion (L/C) and spittlebug resistance (INS). Results from Stage 1 trials showed that 22 selected accessions produced 8 to 10% more total and leaf dry matter yield respectively than the commercial cultivar cv. Marandu but 50 and 68% more than the average of the collection (Valle et al., 1993). Based on production, percentage of leaves and regrowth following cuts, 21 accessions were selected for evaluation in regional trials to assess ecotype x environment interaction (Valle et al., 1999). Eight were selected to be compared under grazing in Stage 2 (Euclides et al., 2001). Four were evaluated in Stage 3 - as regional grazing trials and two of those have been released: cv. Xaras in 2003 (Valle et al., 2004) and cv BRS Piat in 2007. Despite the significant advantages of selected accessions from the collection none gather all desirable characteristics, therefore the breeding program aims exactly at combining important traits into one cultivar through interspecific hybridization. This program started at Embrapa Beef Cattle Center in 1988 and has been using B. ruziziensis as sexual plant and B. decumbens and B. brizantha as apomictic pollen donors. While analyzing the inheritance of apomixis, hybrids were generated for both agronomic evaluation and as new sexual genitors for crossing schemes. Apomixis in Brachiaria is of the apospory Type Panicum - and is simply inherited (Valle and Savidan, 1996; Valle and Miles, 2001), thus apomictic and sexual (1:1) hybrids are generated at each sexual x apomictic cross. Fifty six of those (27 apo + 29 sex) were evaluated for performance under cuts, in plots (Valle et al., 2000): promising hybrids were identified and included in more advanced trials prior to grazing studies. An analysis was conducted to estimate genetic and phenotypic parameters as well as to predict genotypic values of hybrids and of the crosses (Resende et al., 2002). Heritability for yield was high and the strategy for new crosses will now be based on genotypic values predicted for the progeny of selected crosses, to exploit specific combining ability and short time gain. Superior apomictic hybrids thus obtained are immediate candidates for cultivar release whereas the sexual ones can be incorporated in the breeding programs. Rotational crosses exploiting the three species in this agamic complex allows for profiting from progress already obtained with only partial breakdown on heterotic patterns. Another benefit from rotational crosses as used in this program lies in the possibility of indirect crosses between apomictic accessions thus combining desirable traits of each. Progeny testing of sexual genitors are underway to determine next generation crosses only among the superior genotypes. Traits evaluated include yield, nutritive value, spittlebug presence, and regrowth.

Molecular characterization using RAPD (Random Amplified Polymorphic DNA) is being applied to estimate genetic diversity in accessions of Brachiaria as well as for detecting selfed plants among the progenies from artificial crossings in the greenhouse. It has been particularly useful in the intraspecific hybrids of B. humidicola since selfs cannot be readily identified phenotipically. The search for molecular markers closely linked to the apomixis gene in the B. ruziziensis - B. brizantha complex are also being sought in order to speed selection of sexual versus apomictic hybrids in the progenies. This line of work is being pursued in cooperation with the team on apomixis at Embrapa Genetic Resources and Biotechnology. Another line of work in cooperation with Embrapa Maize and Sorghum includes the search for Aluminum tolerance genes, a trait highly expressed by B. decumbens, in order to isolate and transfer to other grasses. Microsatellite markers are being produced for five Brachiaria species in cooperation with UNICAMP, first to study diversity but later preparing for mapping and later to try and locate genes of agronomic importance in marker assisted selection for continuous gain in the breeding of these species.

Final comments
Intensification of animal production presupposes the existence of forage cultivars with improved performance and efficiency in utilizing available resources (biotic and abiotic), thus the large demand for improved varieties adapted to different production systems in Brazil. Particularly in the Cerrados, cultivated pasture over millions of hectares still depend on very few cultivars, basically of Brachiaria decumbens and B. brizantha, characterizing a monoculture of low genetic variability. Despite their widespread use, they exhibit several qualitative and quantitative deficiencies amenable to be ameliorated through breeding. For the development of cultivars there are important pre-requisites: germplasm collections, information on genetic diversity available for the problem(s) at hand, study of mode of reproduction/breeding behavior, ploidy level, and inheritance of traits of importance. Despite years of research, information is still much needed for efficient breeding in tropical forages. The Brachiaria breeding program at Embrapa Beef Cattle Center has identified superior genotypes which resulted in the release of cvs. Marandu, Xaras and BRS Piat of Brachiaria. Intra and interspecific hybrids have been produced and are at different stages of evaluation in the process of cultivar development. Important traits considered are production (total, dry season), resistance to spittlebug, nutritive value, adaptation to different ecosystems (soil, climate), and

good seed production. The cultivar development process is a long term investment: it involves evaluation in plots, regional trials of selected lines for estimation of genotype x environment interaction, and grazing trials for animal performance results.

Multidisciplinary teams are essential for successful adoption of new cultivars. The activities and team involvement in the Brachiaria program have produced knowledge and methods to improve efficiency of selection and with that reduce the time lag to release new cultivars in order to diversify pastures in Brazil and elsewhere in tropical America. Now with recently obtained artificial tetraploidized B. decumbens and B. brizantha, it should be possible to tackle this important and vast germplasm through intraspecific hybridization to try and avoid the seed sterility problems encountered in interspecific hybrids. Traits of interest in this pool are the high aluminum tolerance, good adaptation to poor soils, vigorous growth even with less rainfall and good seed production. There are joint efforts at different research centers from Embrapa studying this gene pool to identify gene(s) associated to Al tolerance. Hybrids of B. humidicola were obtained for the first time and this population is being used to validate the inheritance of apomixis using embryological analysis, but also molecular markers associated to apomixis. RAPD markers were developed to identify self from hybrids. The 50 best full-sib hybrids were planted in plots to evaluate agronomic performance and determine genetic, phenotypical and environmental components using intrapopulational variability analysis. The apomictic elite hybrids are cultivar candidates and the sexual will be used in further crossing to broaden the sexual tetraploid base of this population. The early and trustworthy identification of superior genotypes as predicted by this program should shorten the time for new releases as well as contribute to the knowledge on how to improve apomictic grasses. The adoption of improved cultivars should enhance productivity per animal and per hectare to avoid the need to open new areas for pastures as well as to diversify the existing areas. Very important in this program has been the association with the private sector through UNIPASTO Associao de Melhoramento para o Fomento a Pesquisa de Forrageiras Tropicais - since 2002, which has invested in research and acted as the extension arm in transferring of the technological package (cultivar +

recommendations) through the commercialization of the seed. Brachiaria has changed the scenery of Central Brazil and the impact not in agribusiness alone was felt in several cities and villages. A lot more can be expected from this genus, now that techniques, personnel and methodologies are being

channeled to rationally explore its potential. Genetic breeding is a reality and hybrids are being evaluated to identify those that attend specific an general demands of the different production systems.

References
BOGDAN, A.V. Tropical pasture and fodder plants. Longman, New York. 455 p. 1977.
U U

CENSO AGROPECURIO 1995-1996, Brasil. Rio de Janeiro: IBGE, n.1, p.1-366, 1995-1996. (CIAT). Centro Internacional de Agricultura Tropical. Germoplasma, p.5-28. Informe
U

Anual, 1985, Pastos Tropicales. Cali. 1986.

U

EUCLIDES, V. P. B., VALLE, C. B. DO, MACEDO, M. C. M., OLIVEIRA, M. P. 2001. Evaluation of Brachiaria brizantha ecotypes under grazing in small plots. In: INTERNATIONAL GRASSLAND CONGRESS, 19, 2001, Proceedings... So Pedro:
U U

FEALQ. CD-ROM. ID#13-13. LETTERIELLO, G., VALLE, C.B. DO, CHRISTIANE, D, PENTEADO, M.I. DE O. 1999. Citologia e modo de reproduo de acessos pentaplides de B. brizantha. In: REUNIO ANUAL DA SOCIEDADE BRASILEIRA DE ZOOTECNIA, 36., Anais...1999,
U U

Porto Alegre, SBZ. 1CD-ROM.140. Forragicultura- FOR- Avaliao de Forrageiras. MACEDO, M. C. 2001. Sustainability of pasture production in the savannas of tropical America. In: INTERNATIONAL GRASSLAND CONGRESS, 19, Proceedings...
U U

Piracicaba: FEALQ. CD-ROM. ID#13-14. pp. 391-395. MACKAY, J. H. E. 1982. Register of Australian herbage plant cultivars. Commonwealth Scientific and Industrial Research Organization (CSIRO), Canberra, A.C.T, Australia. 122p NUNES, S. G.; BOOCK, A.; PENTEADO, M. I. DE O.; GOMES, D. T. 1984. Brachiaria brizantha cv. Marandu. Documentos Embrapa, 21. Embrapa/CNPGC, Campo Grande, MS, Brazil. PARSONS, J. J. 1972. Spread of African grasses to the American tropics. Journal of Range Management. 25:12-17.

PENTEADO, M. I. DE O, SANTOS, A. C. M. DOS, RODRIGUES, I. F., VALLE, C. B.DO., SEIXAS, M. A. C, ESTEVES, A. Determinao de ploidia e avaliao da quantidade de DNA total em diferentes espcies do gnero Brachiaria. Campo Grande: Embrapa Gado de Corte, 2000. 32p. Boletim de Pesquisa: 11. EMBRAPA Gado de Corte. PIZARRO, E. A.; VALLE, C. B. DO; KELLER-GREIN, G.; SCHULZE-KRAFT, R.; ZIMMER, A. H. Regional Experiences with Brachiaria: Tropical America - Savannas. In: Miles, J.W.; Maass, B.L.; Valle C.B., ed. Brachiaria: Biology, Agronomy, and Improvement. Cali: CIAT/Braslia:EMBRAPA-CNPGC, 1996.p. 225-246. 1996. RENVOIZE,S. A.,CLAYTON, W. D.,KABUYE, C. H. S. 1996. Morphology, taxonomy and natural distribution of Brachiaria (Trin.)Griseb. In: Brachiaria: Biology, Agronomy, and Improvement. Miles, J.W., Maass. B.L. e Valle, C.B.do (eds). Centro Internacional de Agricultura Tropical - CIAT/Empresa Brasileira de Pesquisa Agropecuria EMBRAPA . CIAT Publication N 259. pp-1-15. RESENDE, R. M. S.; VALLE, C. B. DO; BONATO, A. L. V.; JANK, L.; CALIXTO, S.; CARVALHO, J. DE. Estimao de parmetros genticos e predio de valores genotpicos de cruzamentos interespecficos em Brachiaria. In: REUNIO ANUAL DA SOCIEDADE BRASILEIRA DE Brasileira de Zootecnia, 2002. v. 39. SANTOS FILHO, L. F. 1996. Seed production: perspective from the Brazilian private sector. p. 141-146. In J.W. Miles et al. (ed.) Brachiaria: Biology, agronomy, and improvement. CIAT, Cali, Colombia, and CNPGC/Embrapa, Campo Grande, MS, Brazil. SENDULSKY, T. Brachiaria: taxonomy of cultivated and native species in Brazil. Hoehnea, v.7, p.99-139. 1978. SERRO, E. A. S.; SIMO NETO, M. Informaes sobre duas espcies de gramneas forrageiras do gnero Brachiaria na Amaznia: B. decumbens Stapf e B. ruziziensis Germain et Evrard. IPEAN srie: Estudos sobre forrageiras na Amaznia, vol. 2, no. 1. 1971.Instituto de Pesquisa Agropecuria do Norte (IPEAN), Belm, PA, Brazil. 3 p VALLE C. B. DO, EUCLIDES, V. P. B., MACEDO, M. C. M., VALRIO, J. R., CALIXTO, S. 2001. Selecting new Brachiaria for
U

ZOOTECNIA, 2002, Recife.. Anais.. Sociedade

Brazilian
U

pastures.

In:

INTERNATIONAL GRASSLAND CONGRESS, 19, 2001, Proceedings... So Pedro: FEALQ. CD-ROM. ID#13-14.

VALLE, C. B. DO, CALIXTO, S., AMZQUITA, M. C. 1993. Agronomic evaluation of Brachiaria germplasm in Brazil. In: INTERNATIONAL GRASSLAND CONGRESS, 17. Proceedings... Palmerston North, New Zealand, Rockampton, Australia. p.511-512.
U U

VALLE, C. B. DO, SAVIDAN, Y. H. 1996. Genetics, Cytogenetics, and Reproductive Biology of Brachiaria. In: Brachiaria: Biology, Agronomy, and Improvement. Miles, J.W.,
U U

Maass. B.L. e Valle, C.B.do (eds). Centro Internacional de Agricultura Tropical CIAT/Empresa Brasileira de Pesquisa Agropecuria - EMBRAPA . CIAT Publication N 259. pp-147- 163. VALLE, C. B. DO; EUCLIDES, V. P. B.; PEREIRA, J. M.; VALRIO, J. R.; PAGLIARINI, M. S.; MACEDO, M. C. M.; LEITE, G. G.; LOURENO, A. J.; FERNANDES, C. D.; DIAS-FILHO, M. B.; LEMPP, B.; POTT, A.; SOUZA, M. A. O capim-xaras (Brachiaria brizantha cv. Xaras) na diversificao das pastagens de braquiria. Campo Grande: Embrapa Gado de Corte, 2004. 36p. (Documentos, 149): Embrapa Gado de Corte. VALLE, C. B. DO; MILES, J. W. Breeding of apomictic species. In: SAVIDAN, Y.H; CARMAN, J.G.; DRESSELHAUS, T. (eds).. (Org.). The flowering of apomixis: from mechanisms to genetic engineering. Mexico: CIMMYT, IRD, European Commission, 2001, p. 137-152. VALLE, C. B. DO; M. C. M. MACEDO; CALIXTO, S. 2000. Avaliao agronmica de hbridos de Brachiaria. p. FOR 383. In Anais Reunio Anual da Sociedade Brasileira de Zootecnia, 37th, Viosa, MG, Brazil. 24-28 July 2000, SBZ/Videolar, So Paulo, SP, Brazil. VALLE, C. B. DO, VALRIO, J. R., CALIXTO, S., BARCELLOS, A. O. 1999. Characteristics of selected genotypes of Brachiaria for Brazilian pastures. In: INTERNATIONAL GRASSLAND CONGRESS, 18., 1997, Winnipeg. Proceedings...
U U

[S.l.]: Canadian Forage Council, p.1/81-1/82. CD-ROM. v.1, seccion 1. ID n.1358.