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The relic Criollo cacao in Belize – genetic diversity and relationship with Trinitario and other cacao clones held in the International Cocoa Genebank, Trinidad

  • Lambert A. Motilal (a1), Dapeng Zhang (a2), Pathmanathan Umaharan (a3), Sue Mischke (a2), Vishnarayan Mooleedhar (a4) and Lyndel W. Meinhardt (a2)...


Cacao (Theobroma cacao L.) was domesticated in Mesoamerica and is native to the South American rainforest. Belizean Criollo is a group of relic landraces that are thought to be similar to those used by the Olmecs and Mayans during early domestication. Knowledge of genetic diversity is essential for efficient conservation and use of these relic landraces. Using 30 microsatellite markers, we characterized genetic diversity in 77 Belize Criollo accessions collected from the Maya Mountains in Belize, and assessed their relationship with 62 cacao accessions including 25 Trinitario accessions. Genetic diversity and heterozygosity were low in Belizean Criollo germplasm. Eleven distinctive genotypes were identified among the Belizean germplasm. Results of ordination and cluster analysis supported their putative ancestral contribution to the Trinitario cacao. However, results of Bayesian assignment and parentage analysis both suggested that the contribution of Criollo cacao to the Imperial College Selections Trinitario is small. Our preferred hypothesis for the genesis of Trinitario cacao is that a limited population of Criollo × Forastero hybrids emanated from the introduced Forastero population of Trinidad. The present study provides new insights into the origin of the Trinitario cacao, which will be useful in the ex situ and in situ conservation of cacao landraces from Mesoamerica.


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Bartley, BGD (2001) Refractario – an explanation of the meaning of the term and its relationship to the introductions from Ecuador in 1937. INGENIC Newsletter 6: 1015.
Bartley, BGD (2005) The Genetic Diversity of Cacao and its Utilization. Wallingford: CABI Publishing, 400 p.
Bekele, FL, Bekele, I, Butler, DR and Bidaisee, GG (2006) Patterns of morphological variation in a sample of cacao (Theobroma cacao L.) germplasm from the International Cocoa Genebank, Trinidad. Genetic Resources and Crop Evolution 53: 933948.
Cheesman, EE (1944) Notes on the nomenclature, classification and possible relationships of cacao populations. Tropical Agriculture (Trinidad) 21: 144159.
Ciferri, R and Ciferri, F (1957) The evolution of cultivated cacao. Evolution International Journal of Organic Evolution 9: 381397.
Cryer, NC, Fenn, MGE, Turnbull, CJ and Wilkinson, MJ (2006) Allelic size standards and reference genotypes to unify international cacao (Theobroma cacao L.) microsatellite data. Genetic Resources and Crop Evolution 53: 16431652. doi:10.1007/s10722-005-1286-9.
Dieringer, D and Schlötterer, C (2002) Microsatellite analyser (MSA): a platform independent analysis tool for large microsatellite data sets. Molecular Ecology Notes 3: 167169.
Eyre, C (2007) Cocoa demand on the up. Online documentation. Retrieved 2 August 2007.
Felsenstein, J (1989) PHYLIP – phylogeny inference package (Version 3.2). Cladistics 5: 164166. Software available at
Figueira, AVO and Cascardo, JCM (2001) Molecular markers in breeding. In: Dias, LAS (ed.) Genetic Improvement of Cacao. Translated by Abreu-Reichart CE, Viçosa MG, Dias LAS and Griffee P. Viçosa: FUNAPE, Brazil. EcoPort version, = earticleView&earticleId = 197&page = -1: FAO, pp. 120135.
Henderson, JS, Joyce, RA, Hall, GR, Hurst, WJ and McGovern, PE (2007) Chemical and archaeological evidence for the earliest cacao beverages. Proceedings of the National Academy of Sciences USA 104: 1893718940.
IBPGR (1981) Genetic Resources of Cocoa (AGP:IBPGR/80/56). Rome: IBPGR Working Group on Genetic Resources of Cocoa, IBPGR Secretariat, 25 p.
Iwaro, AD, Bekele, FL and Butler, DR (2003) Evaluation and utilisation of cacao (Theobroma cacao L.) germplasm at the International Cocoa Genebank, Trinidad. Euphytica 130: 207221.
Jakobsson, M and Rosenberg, NA (2007) CLUMPP: a cluster matching and permutation program for dealing with label switching and multimodality in analysis of population structure. Bioinformatics 23: 18011806. doi:10.1093/bioinformatics/btm233.
Johnson, E, Bekele, F and Schnell, R (2004) Field Guide to The ICS Clones of Trinidad. Serie Técnica, Manual Técnico No. 54, 32 p. Turrialba: CATIE.
Johnson, ES, Bekele, FB, Brown, SJ, Song, Q, Zhang, D, Meinhardt, LW and Schnell, RJ (2009) Population structure and genetic diversity of the Trinitario cacao (Theobroma cacao L.) from Trinidad and Tobago. Crop Science 49: 564572. doi:10.2135/cropsci2008.03.0128.
Kalinowski, ST, Taper, ML and Marshall, TC (2007) Revising how the computer program CERVUS accommodates genotyping error increases success in paternity assignment. Molecular Ecology 16: 10991106.
Kobayashi, N, Horikoshi, T, Katsuyama, H, Handa, T and Takayanagi, K (1998) A simple and efficient DNA extraction method for plants, especially woody plants. Plant Tissue Culture and Biotechnology 4: 7680.
Lanaud, C, Risterucci, AM, Pieretti, I, Falque, M, Bouet, A and Lagoda, PJL (1999) Isolation and characterization of microsatellites in Theobroma cacao L. Molecular Ecology 8: 21412152.
Lockwood, G and Gyamfi, MMO (1979) The CRIG cocoa germplasm collection with notes on codes used in the breeding programme at Tafo and elsewhere. Technical Bulletin No. 10. Tafo: Cocoa Research Institute.
Loor, RG, Risterucci, AM, Courtois, B, Fouet, O, Jeanneau, M, Rosenquist, E, Amores, F, Vasco, A, Medina, M and Lanaud, C (2009) Tracing the native ancestors of the modern Theobroma cacao L. population in Ecuador. Tree Genetics and Genomes 5: 421433.
Marshall, TC, Slate, J, Kruuk, LEB and Pemberton, JM (1998) Statistical confidence for likelihood-based paternity inference in natural populations. Molecular Ecology 7: 639655.
Mooleedhar, V (1998) A study of the morphological variation in a relic Criollo cacao population from Belize. Report for 1997. St. Augustine: Cocoa Research Unit, The University of the West Indies, pp. 514.
Mooleedhar, V (2000) Morphological characterisation and genetic evaluation of a relic Criollo cacao (Theobroma cacao L.) population in Belize. PhD thesis, The University of the West Indies, 272 p.
Mooleedhar, V, Bekele, F, Cassie, T, Warren, J and Spence, J (1991) Accessions held in the International Cocoa Genebank, Trinidad (ICG, T). Report for 1990. St Augustine: Cocoa Research Unit, The University of the West Indies, pp. 3133.
Mooleedhar, V, Maharaj, W and O'Brien, HO (1995) The collection of Criollo cocoa germplasm in Belize. Cocoa Growers' Bulletin 49: 2640.
Motamayor, JC, Lachneaud, P, da Silva e Mota, JW, Loor, R, Kuhn, DN, Brown, JS and Schnell, RJ (2008) Geographic and genetic population differentiation of the Amazonian chocolate tree (Theobroma cacao L.). PLoS ONE 3: e3311. doi:10.1371/journal.pone.0003311.
Motamayor, JC, Risterucci, AM, Heath, M and Lanaud, C (2003) Cacao domestication II: progenitor germplasm of the Trinitario cacao cultivar. Heredity 91: 322330.
Motamayor, JC, Risterucci, AM, Lopez, PA, Ortiz, CF, Moreno, A and Lanaud, C (2002) Cacao domestication I: the origin of the cacao cultivated by the Mayas. Heredity 89: 380386.
Motilal, L and Butler, D (2003) Verification of identities in global cacao germplasm collections. Genetic Resources and Crop Evolution 50: 799807.
Motilal, LA, Zhang, D, Umaharan, P, Mischke, S, Boccara, M and Pinney, S (2008) Increasing accuracy and throughput in large-scale microsatellite fingerprinting of cacao field germplasm collections. Tropical Plant Biology 2: 2337. doi:10.1007/s12042-008-9016-z.
Nei, M, Tajima, F and Tateno, Y (1983) Accuracy of estimated phylogenetic trees from molecular data II. Gene frequency data. Journal of Molecular Evolution 19: 153170.
Page, RDM (1996) TREEVIEW: an application to display phylogenetic trees on personal computers. Computer Applications in the Biosciences 12: 357358. TREEVIEW v1.6.6 available at
Peakall, R and Smouse, PE (2006) Genalex 6: genetic analysis in Excel. Population genetic software for teaching and research. Molecular Ecology Notes 6: 288295.
Pound, FJ (1934) The progress of selection. In: Third Annual Report on Cacao Research 1933. Port-of-Spain: Trinidad Government Printing Office, pp. 2528.
Pound, FJ (1935) The progress of selection. In: Fourth Annual Report on Cacao Research 1934. Port-of-Spain: Trinidad Government Printing Office, pp. 711.
Pound, FJ (1936) The completion of selection. In: Fifth Annual Report on Cacao Research 1935. Port-of-Spain: Trinidad Government Printing Office, pp. 716.
Pound, FJ (1938) Cacao and Witches' Broom Disease (Marasmius perniciosus) of South America with Notes on Other Species of Theobroma. Port-of-Spain: Yuille's Printery.
Pritchard, JK, Stephens, M and Donnelly, P (2000) Inference of population structure from multilocus genotype data. Genetics 155: 945959.
Risterucci, AM, Grivet, L, N'Goran, JAK, Pieretti, I, Flament, MH and Lanaud, C (2000) A high-density linkage map of Theobroma cacao L. Theoretical and Applied Genetics 101: 948955.
Saitou, N and Nei, M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution 4: 406425.
Saunders, JA, Mischke, S, Leamy, EA and Hemeida, AA (2004) Selection of international molecular standards for DNA fingerprinting of Theobroma cacao. Theoretical and Applied Genetics 110: 4147.
Takrama, JF, Cervantes-Martinez, C, Philips-Mora, W, Brown, JS, Motamayor, JC and Schnell, RJ (2005) Determination of off-types in a cacao breeding program using microsatellites. INGENIC Newsletter 10: 27.
Turnbull, CJ, Cryer, NC, Ford, CS, Daymond, AJ, Wadsworth, RM and Hadley, P (2004) ICGD. Available at Euronext.liffe/University of Reading.
Valière, N (2002) GIMLET: a computer program for analysing genetic individual identification data. Molecular Ecology Notes 2: 377379. doi:10.1046/j.1471-8286.2002.00228.x.
Waits, LP, Luikart, G and Taberlet, P (2001) Estimating the probability of identity among genotypes in natural populations: cautions and guidelines. Molecular Ecology 10: 249256.
Wood, GAR and Lass, RA (1985) Cocoa. 4th edn., Essex: Longman Group Ltd, 620 p.
Zhang, D, Boccara, M, Motilal, L, Butler, DR, Umaharan, P, Mischke, S and Meinhardt, L (2008) Microsatellite variation and population structure in the ‘Refractario’ cacao of Ecuador. Conservation Genetics 9: 327337. doi:10.1007/s10592-007-9345-8.
Zhang, D, Boccara, M, Motilal, L, Mischke, S, Johnson, ES, Butler, DR, Bailey, B and Meinhardt, L (2009) Molecular characterization of an earliest cacao (Theobroma cacao L.) collection from Upper Amazon using microsatellite DNA markers. Tree Genetics and Genomes 5: 595607. doi:10.1007/s11295-009-0212-2.
Zhang, D, Mischke, S, Goenaga, R, Hemeida, AA and Saunders, JA (2006) Accuracy and reliability of high-throughput microsatellite genotyping for cacao clone identification. Crop Science 46: 20842092.


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The relic Criollo cacao in Belize – genetic diversity and relationship with Trinitario and other cacao clones held in the International Cocoa Genebank, Trinidad

  • Lambert A. Motilal (a1), Dapeng Zhang (a2), Pathmanathan Umaharan (a3), Sue Mischke (a2), Vishnarayan Mooleedhar (a4) and Lyndel W. Meinhardt (a2)...


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