Antonio Lopez-Montes

The prevalence of badnaviruses in West African yams (Dioscorea cayenensis-rotundata) and evidence of endogenous pararetrovirus sequences in their genomes.

Yam (Dioscorea spp.) is an important vegetatively-propagated staple crop in West Africa. Viruses are pervasive in yam worldwide, decreasing growth and yield, as well as hindering the international movement of germplasm. Badnaviruses have been reported to be the most prevalent in yam, and genomes of some other badnaviruses are known to be integrated in their host plant species. However, it was not clear if a similar scenario occurs in Dioscorea yam. This study was conducted to verify the prevalence of badnaviruses, and determine if badnavirus genomes are integrated in the yam genome. Leaf samples (n=58) representing eight species of yam from global yam collections kept at CIRAD, France, and 127 samples of D. rotundata breeding lines (n=112) and landraces (n=15) at IITA, Nigeria, were screened using generic badnavirus PCR primers. Positive amplification of an expected ca. 579bp fragment, corresponding to a partial RT-RNaseH region, was detected in 47 (81%) of 58 samples analysed from CIRAD collections, and 100% of the 127 IITA D. rotundata samples. All the D. cayenensis and D. rotundata samples from the CIRAD and IITA collections tested PCR-positive, and sequencing of a selection of the PCR products confirmed they were typical of the genus Badnavirus. A comparison of serological and nucleic acid techniques was used to investigate whether the PCR-positives were sequences amplified from badnavirus particles or putative endogenous badnavirus sequences in the yam genome. Protein A sandwich-enzyme-linked immunosorbent assay (PAS-ELISA) with badnavirus polyclonal antisera detected cross-reacting viral particles in only 60% (92 of 153) of the CIRAD collection samples analysed, in contrast to the aforementioned 81% by PCR. Immunosorbent electron microscopy (ISEM) of virus preparations of a select set of 16 samples, representing different combinations of positive and negative PCR and PAS-ELISA results, identified bacilliform particles in 11 of these samples. Three PCR-positive yam samples from Burkina Faso (cv. Pilimpikou) were identified in which no viral particles were detected by either PAS-ELISA or ISEM. Southern hybridisation results using a yam badnavirus RT-RNaseH sequence (Gn155Dr) as probe, supported a lack of badnavirus particles in the cv. Pilimpikou and identified their equivalent sequences to be of plant genome origin. Probe Gn155Dr, however, hybridised to viral particles and plant genomic DNA in three D. rotundata samples from Guinea. These results represent the first data demonstrating the presence of integrated sequences of badnaviruses in yam. The implications of this for virus-indexing, breeding and multiplication of seed yams are discussed.

Genome sequencing of the staple food crop white Guinea yam enables the development of a molecular marker for sex determination

BackgroundRoot and tuber crops are a major food source in tropical Africa. Among these crops are several species in the monocotyledonous genus Dioscorea collectively known as yam, a staple tuber crop that contributes enormously to the subsistence and socio-cultural lives of millions of people, principally in West and Central Africa. Yam cultivation is constrained by several factors, and yam can be considered a neglected “orphan” crop that would benefit from crop improvement efforts. However, the lack of genetic and genomic tools has impeded the improvement of this staple crop.ResultsTo accelerate marker-assisted breeding of yam, we performed genome analysis of white Guinea yam (Dioscorea rotundata) and assembled a 594-Mb genome, 76.4% of which was distributed among 21 linkage groups. In total, we predicted 26,198 genes. Phylogenetic analyses with 2381 conserved genes revealed that Dioscorea is a unique lineage of monocotyledons distinct from the Poales (rice), Arecales (palm), and Zingiberales (banana). The entire Dioscorea genus is characterized by the occurrence of separate male and female plants (dioecy), a feature that has limited efficient yam breeding. To infer the genetics of sex determination, we performed whole-genome resequencing of bulked segregants (quantitative trait locus sequencing [QTL-seq]) in F1 progeny segregating for male and female plants and identified a genomic region associated with female heterogametic (male = ZZ, female = ZW) sex determination. We further delineated the W locus and used it to develop a molecular marker for sex identification of Guinea yam plants at the seedling stage.ConclusionsGuinea yam belongs to a unique and highly differentiated clade of monocotyledons. The genome analyses and sex-linked marker development performed in this study should greatly accelerate marker-assisted breeding of Guinea yam. In addition, our QTL-seq approach can be utilized in genetic studies of other outcrossing crops and organisms with highly heterozygous genomes. Genomic analysis of orphan crops such as yam promotes efforts to improve food security and the sustainability of tropical agriculture.

Integration of ploidy level, secondary metabolite profile and morphological traits analyses to define a breeding strategy for trifoliate yam (Dioscorea dumetorum (Kunth) Pax)

The literature in recent times lacks adequate report on the utilization and genetic improvement programmes on Dioscorea dumetorum . Despite the wide application of this yam species in agriculture and medicine, it suffers neglect while other species are becoming increasingly popular. Therefore, it is pertinent to focus on research that will bring this species to the limelight. The aim of the present study was to evaluate the ploidy levels, morphological traits and secondary metabolite profile of 53 accessions of D. dumetorum from six countries in West and Central Africa. Ploidy levels were determined using flow cytometry. Overall, 18 morphological traits were recorded from the above- and underground parts of the plant. The 53 accessions were subjected to statistical analyses using the data on ploidy levels, morphological traits and qualitative phytochemical screening. A total of 15 accessions from the generated clusters were selected for thin layer chromatographic and quantitative phytochemical analyses. The analyses revealed diploid (2 x ) and triploid (3 x ) levels in these accessions. The pruned dendrogram derived from agglomerative hierarchical clustering based on the distance matrix revealed three main groups, showing a relationship between sex and ploidy level in the accessions and exhibiting sufficient cluster variability that may be important in designing breeding programmes. The crop was also shown to possess metabolites such as alkaloids, saponins and flavonoids, which are known to be useful in the application of phytomedicine. Genetic variability observed among the yam accessions in this study can be used for breeding purposes and to broaden the genetic basis of the crop for efficient utilization of the genetic potential possessed by this species.

Carotenoid profiling of yams: Clarity, comparisons and diversity

Screening carotenoids of elite accessions of yam (Dioscorea spp.) used in the global yam breeding program has been conducted to quantitatively determine the carotenoid composition of the crop. Comparisons to previous data reporting cerotenoid levels in yam has been made, in order to deduce greater perspectives across multiple studies. Characterisation of complex species and accession -specific profiles have shown a rich base of diversity that can inform breeding strategies. Key findings include; (i) the identification of accessions rich in β-carotene which can aid provitamin A biofortification, (ii) Data disputing the commonly held belief that yellow Guinea yam (D. cayennensis) has higher β-carotene content than that of white Guinea yam (D. rotundata), and (iii) the tentative identification of C25-epoxy-apocarotenoid persicaxanthin with potential implications for tuber dormancy.

Re-defining the yam (Dioscorea spp.) core collection using morphological traits

Development of core collection representing the diversity in the entire germplasm creates a better access and enhanced utilization of the main collection thus allowing rapid evaluation in crop improvement programs. Core collections are dynamic in nature and needs revisiting when additional germplasm and information becomes available. In the current study, an attempt was made to re-define the previously developed yam ( Dioscorea spp) core collection using 56 morphological traits. Information on additional acquired germplasm and presence of duplicates or mislabelled accessions in the entire collection was also used. The re-defined core collection consisted of 843 accessions and represented about 20% of the entire collection. It included six Dioscorea species, of which accessions of Dioscorea rotundata are in the majority (73.54%) followed by Dioscorea alata (21.35%), Dioscorea bulbifera (1.66%), Dioscorea cayenensis (1.42%), Dioscorea dumetorum (1.42%) and Dioscorea esculenta (0.59%). The Shannon weaver diversity index and principal component analysis revealed the maximum diversity captured in the core from the base collection. This re-defined core collection is more valuable than the original core since it represents true-to-type accessions ensuring reliability for enhanced utilization of germplasm in yam improvement programs.

Comparative Reliability of Screening Parameters for Anthracnose Resistance in Water Yam (Dioscorea alata)

Anthracnose, caused by Colletotrichum gloeosporioides, is one of the major constraints limiting water yam (Dioscorea alata) production in the tropics. In this region, yam anthracnose control is mostly achieved by the deployment of moderately resistant yam genotypes. Therefore, screening for new sources of anthracnose resistance is an important aspect of yam research in the tropics. The reliability and applicability of different yam anthracnose rating parameters has not been fully examined. Disease severity on detached leaves in the laboratory and leaf severity, lesion size, and spore production on whole plants in the greenhouse were used to screen an F1 yam population and correlate screening results with field evaluations. Anthracnose lesion size had the smallest predicted residual means but whole-plant severity and detached-leaf severity had the best variance homogeneity and relatively small predicted residual means. The concordance correlation coefficient (rc) and κ statistic were used to determine the agreement between anthracnose rating parameters and field evaluations. Detached-leaf (rc = 0.95, κ = 0.81) and whole-plant (rc = 0.96, κ = 0.86) evaluations had high positive agreement with field evaluation but spore production (κ = 0.69) and lesion size (κ = 0.57) had moderate positive agreement. These results suggest that all the evaluated rating parameters can be used to successfully screen yam germplasm for anthracnose resistance but lesion size and spore production data may need to be transformed.

An EST-SSR based genetic linkage map and identification of QTLs for anthracnose disease resistance in water yam (Dioscorea alata L.)

Water yam (Dioscorea alata L.) is one of the most important food yams with wide geographical distribution in the tropics. One of the major constraints to water yam production is anthracnose disease caused by a fungus, Colletotrichum gloeosporioides (Penz.). There are no economically feasible solutions as chemical sprays or cultural practices, such as crop rotation are seldom convenient for smallholder farmers for sustainable control of the disease. Breeding for development of durable genetic resistant varieties is known to offer lasting solution to control endemic disease threats to crop production. However, breeding for resistance to anthracnose has been slow considering the biological constraints related to the heterozygous and vegetative propagation of the crop. The development of saturated linkage maps with high marker density, such as SSRs, followed by identification of QTLs can accelerate the speed and precision of resistance breeding in water yam. In a previous study, a total of 1,152 EST-SSRs were developed from >40,000 EST-sequences generated from two D. alata genotypes. A set of 380 EST-SSRs were validated as polymorphic when tested on two diverse parents targeted for anthracnose disease and were used to generate a saturated linkage map. Majority of the SSRs (60.2%) showed Mendelian segregation pattern and had no effect on the construction of linkage map. All 380 EST-SSRs were mapped into 20 linkage groups, and covered a total length of 3229.5 cM. Majority of the markers were mapped on linkage group 1 (LG 1) comprising of 97 EST-SSRs. This is the first genetic linkage map of water yam constructed using EST-SSRs. QTL localization was based on phenotypic data collected over a 3-year period of inoculating the mapping population with the most virulent strain of C. gloeosporioides from West Africa. Based on threshold LOD scores, one QTL was consistently observed on LG 14 in all the three years and average score data. This QTL was found at position interval of 71.1–84.8 cM explaining 68.5% of the total phenotypic variation in the average score data. The high marker density allowed identification of QTLs and association for anthracnose disease, which could be validated in other mapping populations and used in marker-assisted breeding in D. alata improvement programmes.