Molecular characterization of an earliest cacao (Theobroma cacao L.) collection from Upper Amazon using microsatellite DNA markers

Abstract

Cacao (Theobroma cacao L.) is indigenous to the Amazon region of South America. The river basins in the Upper Amazon harbor a large number of diverse cacao populations. Since the 1930s, several numbers of populations have been collected from the present-day Peruvian Amazon and maintained as ex situ germplasm repositories in various countries, with the largest held in the International Cacao Genebank in Trinidad. The lack of information on population structure and pedigree relationship and the incorrect labeling of accessions are major concerns for efficient conservation and use of cacao germplasm. In the present study, we assessed the individual identity, sibship, and population structure in cacao populations collected from the present-day Loreto Region, Peru in the 1930-1940s. Using a capillary electrophoresis genotyping system, we analyzed the simple sequence repeat variation of 612 cacao accessions collected from the Marañon, Nanay, and Ucayali river systems. A total of 180 cases of mislabeling were identified using a Bayesian clustering method for admixture detection. Using maximum likelihood-based methods, we reconstructed 78 full-sib families nested in 48 half-sib families, indicating that the pods collected in the 1930s were from 48 mother trees, maximum. Likelihood simulation also identified eight probable parents that are responsible for 117 pairs of mother-offspring relationships in this collection. Principal coordinate analysis (PCoA) and the Bayesian clustering method cohesively demonstrated a pronounced structure of genetic diversity, stratified by the river systems of the Peruvian Amazon. Our results also show that, in spite of the high level of allelic diversity in this collection, it was composed of a large number of related family members collected from a relatively small area, including a couple of sites in the Ucayali and Nanay rivers, as well as the lower Marañon river near Iquitos. The vast majority of the Peruvian Amazon, especially the upper Marañon River and its tributaries, have not been sampled by collecting expeditions. The improved understanding of the individual identities, genealogical relationships, and geographical origin of cacao germplasm in this collection will contribute to more efficient conservation and utilization of these germplasm. Additionally, this study also provides more baseline information to help guide future collecting expeditions in the Peruvian Amazon.

Verification of genetic identity of introduced cacao germplasm in Ghana using single nucleotide polymorphism (SNP) markers

“…The 39 tested cacao accessions from the Ghana cacao collection, as well as the 100 reference accessions, were stratified as germplasm groups of Amelonado, IMC, SCA/Ucayali, Morona, Nanay and Parinari, respectively (Figure 2). The assignment result largely agreed with the previously classified germplasm groups (Figure 2; Zhang et al, 2009b;Motamayor et al, 2008) except that the germplasm groups of SCA/Ucayali and Morona were not separated. The assigned memberships for all the tested trees from Ghana were compatible with their known parentage germplasm groups (Figure 2).…”

“…The SCA/Ucayali and Morona accessions represent two distinct geographical regions and were clustered as two different genetic groups when SSR markers were used (Zhang et al, 2009b;Motamayor et al, 2008). However, in the present study, the Bayesian clustering analysis based on 53 SNP markers did not significantly differrentiate these two germplasm groups (Figure 2).…”

“…For the identification of mislabeling (off-types), SNP profiles of 100 reference trees maintained in the International Cacao Genebank, Trinidad (ICG,T) were used in the analysis. The genetic identity of the 100 reference trees has been characterized by both SNP (D. Zhang, USDA/ARS, Beltsville, personal communication) and SSR fingerprinting (Zhang et al, 2009b;Motilal et al, 2010;Johnson et al, 2009). Pairwise multilocus matching was applied among each pair of individual trees, including the reference trees from the international germplasm collections, using the same program.…”

“…These 100 accessions were taken from six known Forastero germplasm groups, including Amelonado, Scavina (SCA) and Ucayali, Iquitos Mixed Calabacillo (IMC), Morona (MO), Nanay (NA) and Parinari (PA). Classification of these accessions have been reported by Zhang et al, 2009b). The number of clusters (K-value, which indicated the number of sub-populations of the program attempted to find) was set from two to ten, and the analysis was carried out without assuming any prior information about the genetic group or geographic origin of the samples.…”

Theobroma

“…The result of the assignment test is a posterior probability value that can be interpreted directly as the probability of origin of each individual from each population sampled (assuming the true population of origin has been sampled). Since the majority of the cacao germplasm was collected from different geographical regions and belongs to different populations, the assignment test in combination with SSR markers is a very powerful tool in detecting mislabeled accessions for cacao (Zhang et al 2006b(Zhang et al , 2009bMotamayor et al 2008). …”