Comparative analysis of genetic variation in kava (Piper methysticum) assessed by SSR and DArT reveals zygotic foundation and clonal diversification.

Kava (Piper methysticum) is a major cash crop in the Pacific. The aim of this study was to assess genetic variation among 103 accessions of kava using SSRs and DArTs. Genetic structure was determined using clustering analyses (WPGMA) and principal coordinate analyses (PCA). Thirteen SSR primers and 75 DArT markers were found polymorphic, and the two types of markers generated similar clustering patterns. Genetic distances ranged from 0 to 0.65 with an average of 0.24 using SSRs and from 0 to 0.64 with an average of 0.24 using DArT. Eleven genotypes were identified with SSR while 28 genotypes were identified with DArT markers. By combining the two sets of markers, a total of only 30 distinct genotypes were observed. In the Vanuatu archipelago, noble cultivars originating from different islands clustered together within a very narrow genetic base despite their diversity of morphotypes. SSR and DArT fingerprints allowed the identification of kava cultivars unsuitable for consumption, so called two-days, and clearly differentiated the wild types classified as P. methysticum var. wichmannii from the cultivars as var. methysticum. Molecular data reveals that all noble cultivars evolved by the predominance of clonal selection. Although they are represented by clearly distinct morphotypes, these cultivars are genetically vulnerable and their potential to adapt to forthcoming changes is limited. These newly developed markers provide high resolution and will be useful for kava diversity analyses and quality assessment.

Application of near infrared reflectance spectroscopy for the evaluation of yam (Dioscorea alata) germplasm and breeding lines.

BACKGROUND: Thousands of yam (Dioscorea spp.) accessions are maintained in germplasm collections. The physico-chemical characteristics of the tubers are rarely characterised. Unless a simple, low cost, screening tool is available, it is difficult to evaluate the quality of varieties and breeding lines. We investigated the potential of near infrared reflectance spectroscopy (NIRS) as an alternative method for predicting the major constituents of the yam tuber. RESULTS: Two hundred and sixty-five accessions, belonging to seven different Dioscorea spp., were analysed for starch, amylose, sugars, proteins, minerals and cellulose. The comparison of the NIR spectra and the chemical values allowed the establishment of equations of calibration for the prediction of starch, sugars and proteins (equivalent N). The r(2) pred values for starch, sugars and proteins (respectively 0.84, 0.86 and 0.88) are high enough to allow good estimates of their contents. Values for the ratio of performance to deviation (RPD) of 4.046 and 3.641 for the sugars and proteins models also allow good quantitative predictions to be made. Amylose, cellulose and minerals could not be predicted precisely. A second calibration conducted by adding the calibration and validation sets (260 accessions) revealed an improvement of the RPD values for starch, sugars and proteins, indicating that the models can be improved. Discriminant analysis conducted using 2151 wavelengths (in nanometres) as variables was applied to a set of 214 accessions of D. alata and the results were compared to the principal component analysis of chemical data. Accessions can be classified according to the amylaceous fraction of the chemotype. CONCLUSION: NIRS could be used in yam breeding programmes to characterise rapidly and at low cost the numerous accessions and breeding lines.

Understanding the genetic diversity and population structure of yam (Dioscorea alata L.) using microsatellite markers.

Yams (Dioscorea sp.) are staple food crops for millions of people in tropical and subtropical regions. Dioscorea alata, also known as greater yam, is one of the major cultivated species and most widely distributed throughout the tropics. Despite its economic and cultural importance, very little is known about its origin, diversity and genetics. As a consequence, breeding efforts for resistance to its main disease, anthracnose, have been fairly limited. The objective of this study was to contribute to the understanding of D. alata genetic diversity by genotyping 384 accessions from different geographical regions (South Pacific, Asia, Africa and the Caribbean), using 24 microsatellite markers. Diversity structuration was assessed via Principal Coordinate Analysis, UPGMA analysis and the Bayesian approach implemented in STRUCTURE. Our results revealed the existence of a wide genetic diversity and a significant structuring associated with geographic origin, ploidy levels and morpho-agronomic characteristics. Seventeen major groups of genetically close cultivars have been identified, including eleven groups of diploid cultivars, four groups of triploids and two groups of tetraploids. STRUCTURE revealed the existence of six populations in the diploid genetic pool and a few admixed cultivars. These results will be very useful for rationalizing D. alata genetic resources in breeding programs across different regions and for improving germplasm conservation methods.

New development and validation of 50 SSR markers in breadfruit (Artocarpus altilis, Moraceae) by next-generation sequencing.

PREMISE OF THE STUDY: Using next-generation sequencing technology, new microsatellite loci were characterized in Artocarpus altilis (Moraceae) and two congeners to increase the number of available markers for genotyping breadfruit cultivars. METHODS AND RESULTS: A total of 47,607 simple sequence repeat loci were obtained by sequencing a library of breadfruit genomic DNA with an Illumina MiSeq system. Among them, 50 single-locus markers were selected and assessed using 41 samples (39 A. altilis, one A. camansi, and one A. heterophyllus). All loci were polymorphic in A. altilis, 44 in A. camansi, and 21 in A. heterophyllus. The number of alleles per locus ranged from two to 19. CONCLUSIONS: The new markers will be useful for assessing the identity and genetic diversity of breadfruit cultivars on a small geographical scale, gaining a better understanding of farmer management practices, and will help to optimize breadfruit genebank management.

Characterization of microsatellites in Xanthosoma sagittifolium (Araceae) and cross-amplification in related species.

PREMISE OF THE STUDY: To investigate the genetic diversity of a root crop, Xanthosoma sagittifolium, and to facilitate germplasm conservation, microsatellite loci were developed and characterized by genotyping 39 accessions from different geographic origins. • METHODS AND RESULTS: Using a microsatellite-enriched library approach, 17 polymorphic microsatellite markers were identified and characterized. The number of alleles for each locus ranged from two to six. Observed and expected heterozygosities ranged from 0.00 to 0.97 and from 0.09 to 0.78, respectively. Additionally, cross-amplification of these microsatellite markers was tested successfully in other species of Xanthosoma and Caladium, with rates varying from 23.5% to 100%. • CONCLUSIONS: These results indicate the effectiveness of microsatellite loci developed for the characterization of X. sagittifolium genetic diversity. They are crucial for the future investigation of population dynamics and clonal identification and, therefore, for prioritizing germplasm conservation. They should also enable research on other related species.

Rapid estimation of taro (Colocasia esculenta) quality by near-infrared reflectance spectroscopy.

The aim of the present study is to develop a methodology for the rapid estimation of taro (Colocasia esculenta) quality. Chemical analyses were conducted on 315 accessions for major constituents (starch, total sugars, cellulose, proteins, and minerals). NIRS calibration equations, developed on a calibration set composed of 243 accessions, showed high explained variances in cross-validation (r(2)(cv)) for starch (0.89), sugars (0.90), proteins (0.89), and minerals (0.90) but poor response for amylose (0.44) and cellulose (0.61). The predictions were tested on an independent set of 58 randomly selected accessions. The r(2)(pred) values for starch, sugars, proteins, and minerals were, respectively, of 0.76, 0.74, 0.85, and 0.85 with ratios of performance to deviation (RPD) of 3.41, 4.01, 3.78, and 3.64. New calibration equations developed on 303 accessions confirmed good RPD values for starch (3.30), sugars (4.13), proteins (3.61), and minerals (3.74). NIRS could be used to predict starch, sugars, proteins, and minerals contents in taro corms with reasonably high confidence.

NIR determination of major constituents in tropical root and tuber crop flours.

Tropical root and tuber crops (cassava, sweet potato, taro, and yam) are staples in developing countries where rapid urbanization is strengthening the demand for flour based foods. Quality control techniques are still under development, and when available, laboratory analyses are too expensive. The objectives of this study were to calibrate Near-infrared spectroscopy (NIRS) for routine analysis of flours and to test its reliability to determine their major constituents. Flours prepared from 472 accessions (traditional varieties and breeding lines) were analyzed for their starch, total sugars, cellulose, total nitrogen, and ash (total minerals) contents. The near-infrared (350-2500 nm) spectra of all samples were measured. Calibration equations with cross and independent validation for all analytical characteristics were computed using the partial least squares method. Models were developed separately for each of the four crop species and by combining data from all spp. to predict values within each of them. The quality of prediction was evaluated on a test set of 94 accessions (20%) by standard error of prediction (SEP) and r2 parameters between the measured and the predicted values from cross-validation. Starch, sugar, and total nitrogen content could be predicted, respectively, with 87%, 86%, and 93% confidence, whereas ash (minerals) could be predicted with 71%, and cellulose was not predictable (r2=0.31). The statistical parameters obtained for starch, sugars, and total nitrogen are of special interest for flour quality control. These constituents are quantitatively the most important in the chemical composition of flours, and starch content is negatively correlated with sugars and total nitrogen. NIRS is a low cost technique well adapted to the conditions in developing countries and can be used for the high-throughput screening of a great number of samples. Possible applications are discussed.