Quantification of Bt δ-endotoxins in leaf tissues of tropical Bt maize populations

In Kenya, stem borers destroy an estimated 13.5% of farmers' annual maize harvest. Maize transformed using Bacillus thuringiensis (Bt) derived genes controls stem borers without negative effects to humans, livestock or the environment. The effectiveness and sustainability of Bt transgenic technology in the control of stem borers depends on the levels of concentration of the Bt δ-endotoxins in plant tissues. Kenya introduced Bt maize events to test the efficacy of Bt maize in controlling stem borers, and to develop high-yielding and locally adapted Bt maize germplasm for farmers. The objective of this study was to assess under greenhouse conditions the concentration levels of Bt δ-endotoxins in the leaf tissues of the parents, the F1, and the F2:3 populations of tropical maize, as a measure of stability and sustainability. Kenya introduced Bt maize events to test the efficacy of Bt maize in controlling stem borers, and to develop high-yielding and locally adapted Bt maize germplasm for farmers. The objective of this study was to assess under greenhouse conditions the concentration levels of Bt δ-endotoxins in the leaf tissues of the parents, the F1, and the F2:3 populations of tropical maize, as a measure of stability and sustainability. Two public Bt maize lines (Event 216 and Event 223) containing the cry1Ab::ubi gene were crossed with two non-Bt maize inbred lines, CML144 and CML159, to assess how the concentrations of Bt δ-endotoxins are transmitted from parents to F1 and to F2 generations. The mean concentration of Bt δ-endotoxins (μg/g) was 4.93 and 4.63 in Events 216 and 223 respectively. As expected, F1 generations of all the crosses had similar concentrations of Bt δ-endotoxins. However, the F2 generations showed a spread of concentrations. These findings may imply that genotypes with a higher mean concentration of Bt δ-endotoxins also have a lower level of plant damage traits expressed. In addition, these observations indicate that the cry1Ab gene was dominant and was inherited following the Mendelian segregation and that Events 216 and 223 could be utilized as reliable sources of resistance to stem borers in maize breeding programmes.Keywords: Bt maize, stem borers, Bt δ-endotoxins, enzyme-linked immunosorbent assay (ELISA), dot blot analysis, cry1A

Genetic diversity and gene flow revealed by microsatellite DNA markers in some accessions of African Plum (Dacryodes edulis) in Cameroon

Dacryodes edulis is a multipurpose tree integrated in the cropping system of Central African region still dominated by subsistence agriculture. Some populations grown are wild which can provide information on the domestication process, and could also represent a potential source of gene flow. Leaves samples for DNA extraction were collected from wild forms in Mbakwa supe region and from cultivated forms in Yaounde and Santchou region. Six microsatellites DNA markers were employed in genotyping to analyze population structure and gene flow. Amplification rate was high and genotyping revealed high level of genetic variation. The overall polymorphic level at the six loci was also high with average expected heterozygosity of 0.53; polymorphism of 0.46; mean allelic diversity of 0.5 and mean allele number of 8.33. There were no clear differences with only 1% variation among the three populations and 6% variation among individuals within populations. In contrast, the rate of heterozygosis was high in all the three populations. Both the number of migrant per generation (Nm=20) and the Wright’s Fstatistics (FST=0.012) suggest that there was substantial gene flow among the populations. These findings indicate that D. edulis possess a great potential of pollen dispersal and dominant crosspollination within populations. Most of the loci with private alleles (45%) were found in wild individuals which could be a source of pollen for crossing their cultivated relatives.Key words: Dacryodes edulis, domestication, genetic diversity and structure

Metagenomic analysis of viruses associated with maize lethal necrosis in Kenya

Background: Maize lethal necrosis is caused by a synergistic co-infection of Maize chlorotic mottle virus (MCMV) and a specific member of the Potyviridae, such as Sugarcane mosaic virus (SCMV), Wheat streak mosaic virus (WSMV) or Johnson grass mosaic virus (JGMV). Typical maize lethal necrosis symptoms include severe yellowing and leaf drying from the edges. In Kenya, we detected plants showing typical and atypical symptoms. Both groups of plants often tested negative for SCMV by ELISA. Methods: We used next-generation sequencing to identify viruses associated to maize lethal necrosis in Kenya through a metagenomics analysis. Symptomatic and asymptomatic leaf samples were collected from maize and sorghum representing sixteen counties. Results: Complete and partial genomes were assembled for MCMV, SCMV, Maize streak virus (MSV) and Maize yellow dwarf virus-RMV (MYDV-RMV). These four viruses (MCMV, SCMV, MSV and MYDV-RMV) were found together in 30 of 68 samples. A geographic analysis showed that these viruses are widely distributed in Kenya. Phylogenetic analyses of nucleotide sequences showed that MCMV, MYDV-RMV and MSV are similar to isolates from East Africa and other parts of the world. Single nucleotide polymorphism, nucleotide and polyprotein sequence alignments identified three genetically distinct groups of SCMV in Kenya. Variation mapped to sequences at the border of NIb and the coat protein. Partial genome sequences were obtained for other four potyviruses and one polerovirus. Conclusion: Our results uncover the complexity of the maize lethal necrosis epidemic in Kenya. MCMV, SCMV, MSV and MYDV-RMV are widely distributed and infect both maize and sorghum. SCMV population in Kenya is diverse and consists of numerous strains that are genetically different to isolates from other parts of the world. Several potyviruses, and possibly poleroviruses, are also involved

Analysis of the impact of domestication of Warburgia ugandensis (Sprague) on its genetic diversity based on amplified fragment length polymorphism

Warburgia ugandensis Sprague (Canellaceae) occurs in East and Central Africa and is an important multipurpose tree species. Over-exploitation of natural forests for medicinal purposes and clearance for farming threaten the species survival.  Cultivation of the tree species would ensure sustainable medicinal source and its conservation. However, on-farm genetic diversity of the species is currently unknown. The genetic diversity of the on-farm W. ugandensis populations and their proximate natural populations were analyzed using the amplified fragment length polymorphism (AFLP). Four primer combinations produced a total of 223 polymorphic bands. Both the natural and on-farm populations had high genetic diversity ranging from H = 0.2892 to H = 0.1278. Principal co-ordinates analysis and dendrogram separated the ten populations into two major groups corresponding to Kenyan and Tanzanian populations, respectively. Ugandan populations were shared between the two major groups; this is probably because Uganda is believed to be the centre of diversity for W. ugandensis. Close genetic relationships between the on-farm and their proximate natural population were revealed. Analysis of molecular variance (AMOVA) revealed that a total of 54% AFLP variation resided within populations with 46% reside among populations. The high genetic diversity of W. ugandensis on-farm populations could be useful in germplasm collection and conservation strategies.Key words: Warburgia ugandensis, amplified fragment length polymorphism (AFLP), domestication, genetic diversity, on-farm, natural

Yield potential of sandponically produced sweetpotato (Ipomoea batatas (L.) Lam) pre-basic seed for selected genotypes

Sufficient sweetpotato (Ipomoea batatas (L.) Lam) pre-basic seed at the start of the “seed” value chain is critical and often a bottleneck in the production of sweetpotato in Sub-Saharan Africa. Predominantly, pre-basic seed is multiplied in screenhouse using the conventional soil substrate method which is costly, is untenable, and achieves sub-optimal yields. The sandponics system is a better alternative for sweetpotato pre-basic seed multiplication in the screenhouse attributed to increased yields and cost-effectiveness. This experiment compared sweetpotato yield- and growth-related traits of planting materials sourced from the sandponics system with conventional soil substrate planting materials for four genotypes when grown in the field. A randomized complete block design was used with three replicates. Results showed a significant difference (p ≤ 0.05) between sources of planting materials and among genotypes for the measured traits. The interaction of source of planting materials and genotype was significant (p ≤ 0.05) for harvest index. Vine survival, storage root yield, number of storage roots per plant, and vine yield were higher by 4.1%, 24%, 27%, and 24%, respectively, in favor of planting materials sourced from the sandponics system. Sandponically multiplied planting materials showed superior performance for yield and most of the measured growth-related traits to planting materials multiplied by the conventional soil substrate method

Degeneration of cleaned-up, virus-tested sweetpotato seed vines in Tanzania

Viruses pose a major challenge to sweetpotato production in Tanzania. Use of cleaned-up, virus-tested seed vines distributed through a formal seed system is among the proposed strategies to address this challenge. However, virus-tested seed vines can get infected once in the field and it is not known how they will perform following several seasons of on farm propagation. We assessed the performance of virus-tested seed vines and farmer-sourced seed vines of a susceptible variety, Ejumula, and a relatively tolerant variety, Kabode, over five seasons to understand the trend in root yields, vine yields and virus incidences. The experiments were done in high and low virus pressure areas. The most prevalent viruses were sweet potato chlorotic stunt virus (SPCSV) followed by sweet potato feathery mottle virus (SPFMV) and sweet potato leaf curl virus (SPLCV), respectively. Both farmer-sourced and cleaned-up, virus-tested seed of cv. Ejumula were rapidly infected with SPCSV. The incidence of this virus on Ejumula's farmer-sourced material at the high-virus-pressure area reached 100% by the second season. The incidences for all three viruses remained stable for cv. Kabode across the five seasons. Plants generated from cleaned-up, virus-tested seed had lower incidences for all viruses compared to those from farmer-sourced planting material. Virus-tested seed produced significantly higher root yields for cv. Ejumula in the high-virus-pressure site, with a gradual drop across the seasons. The findings show that regular replenishment of clean, virus-tested seed is more economical in high-virus-pressure areas and for more susceptible varieties like cv. Ejumula. They also indicate that farmers may be reluctant to invest in cleaned-up, virus-tested seed in cases where they have virus-tolerant varieties such as cv. Kabode due to lack of obvious virus effect on yields

Genetic diversity in Napier grass (Pennisetum purpureum) cultivars: Implications for breeding and conservation

Napier grass is an important forage crop for dairy production in the tropics; as such, its existing genetic diversity needs to be assessed for conservation. The current study assessed the genetic variation of Napier grass collections from selected regions in Eastern Africa and the International Livestock Research Institute Forage Germplasm-Ethiopia. The diversity of 281 cultivars was investigated using five selective amplified fragment length polymorphism (AFLP) markers and classical population genetic parameters analysed using various software. The number of bands generated was 216 with fragments per primer set ranging from 50 to 115. Mean percentage polymorphic loci was 63.40. Genetic diversity coefficients based on Nei's genetic diversity ranged from 0.0783 to 0.2142 and Shannon's information index ranged from 0.1293 to 0.3445. The Fst value obtained was moderately significant (Fst = 0.1688). Neighbour-joining analysis gave two distinct clusters which did not reflect geographical locations. Analysis of molecular variance showed all variance components to be highly significant (P < 0.001), indicating more variation within (91 %) than between populations (9 %). Results suggested moderate genetic differentiation among Napier grass populations sampled, which could imply a high germplasm exchange within the region. The AFLP markers used in this study efficiently discriminate among cultivars and could be useful in identification and germplasm conservation

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Additional file 9: of Metagenomic analysis of viruses associated with maize lethal necrosis in Kenya

MYDV-RMV contigs used for phylogenetic analysis. (FASTA 27 kb

Additional file 6: of Metagenomic analysis of viruses associated with maize lethal necrosis in Kenya

SCMV contigs used for phylogenetic analysis. (FASTA 193 kb