Stable gene transformation in cowpea (Vigna unguiculata L. Walp.) using particle gun method

We investigated the possibility of transforming and obtaining transgenic cowpea (Vigna unguiculata L Walp) plants using the particle bombardment process. Meristematic explants that could give rise to whole fertile plants were used in transformation experiments with reporter and selectable marker genes driven by a 35S CaMV promoter. Conditions for optimal delivery of DNA to explants were established based on transient gus expression assays two days after bombardment. The size of microcarriers, microflight distance and helium pressure significantly affected transient expression of reporter genes. A total of 1692 explants were bombarded with DNA-coated particles and placed on 3 mg/l bialaphos selective medium. Only 12 regenerated shoots produced seeds eventually, and all were Gus negative even though 7 gave positive PCR signals with the bar primer. Eight out of 1400 seeds from To plants were GUS positive. DNA from eight of the GUS positive seedlings were amplified with both the gus and bar primers in PCR analysis but only two gave a positive Southern signal. Only two of the 3557 T2 seedlings obtained were GUS positive. However, 3 seedlings survived Basta spray. The two GUS positive and 3 Basta surviving seedlings gave positive Southern hybridisation signals. Twelve T3 seedlings from these were GUS positive and also gave positive Southern hybridisation signals. The positive reaction of T1, T2 and T3 seedlings under Southern analysis confirms the stable integration of introduced genes and the transfer of such genes to progenies. However, the level of expression of introduced genes in cowpea cells is very low and this accounted for the high mortality rate of progenies under Basta spray

Importance du virus de la marbrure de niébé et du virus de la mosaïque jaune du niébé au Togo

Deux virus du niébé (Vigna unguiculata), le virus de la marbrure du niébé et le virus de la mosaïque jaune du niébé, ont été étudiés. Les deux virus ont été identifiés et caractérisés à l'aide de la gamme de plantes indicatrices et de méthodes sérologiques. Les deux virus sont très répandus dans les principales régions productrices du niébé. Le virus de la mosaïque jaune du niébé a été retrouvé dans 65 % des échantillons virosés et le virus de la marbrure du niébé dans 35 % des échantillons. Des infections mixtes ont été souvent observées impliquant ces deux virus et d'autres virus du niébé. Le virus de la mosaïque du niébé transmis par le puceron, le virus de la marbrure peu sévère du niébé et le virus de la mosaïque du sud du haricot ont aussi été identifiés. L'étude des symptômes induits par ces deux virus sur une série de plantes indicatrices révèle que les Chenopodium amaranticolor, Phaseolus vulgaris cv. Saint-Fiacre, Vigna unguiculata cv. Locale Blanche et Glycine max cv. Jupiter pourraient être utilisés pour la séparation des deux virus lorsqu'ils sont en mélange. Les Crotalariajuncea, Phaseolus vulgaris cv. Saxa et Sesbania rostrata n'ont pas été infectés par ces deux virus. Vingt-trois cultivars du niébé ont été évalués en utilisant un isolat de chacun des deux virus et plusieurs sont résistants à l'un des deux virus, mais pas aux deux conjointement. Le cultivar du niébé TVxl850-01E ne démontre aucun symptôme face aux deux virus.Two cowpea (Vigna unguiculata) viruses, cowpea mottle virus and cowpea yellow mosaic virus were studied. The two viruses were identified and characterized by host range and serology. Both viruses were widespread in major cowpea producing areas. Cowpea yellow mosaic virus was found in 65 % and cowpea mottle virus in 35 % of infected samples. Mixed infections were often observed including both viruses and other cowpea viruses. Cowpea aphid-borne mosaic virus, cowpea mild mottle virus and southern bean mosaic virus were also identified. Host range studies showed that Chenopodium amaranticolor, Phaseolus vulgaris cv. Saint-Fiacre, Vigna unguiculata cv. Locale Blanche, and Glycine max cv. Jupiter could be used to separate both viruses from mixed infections. Crotalaria juncea, Phaseolus vulgaris cv. Saxa, and Sesbania rostrata were not host plants for both viruses. Twenty-three cowpea cultivars were screened using an isolate of each virus. Several cowpea cultivars were resistant to only one of the viruses but not to both. Cowpea cultivar TVxl850-01E was found to produce no symptoms with both viruses

Diversity of banana streak-inducing viruses in Nigeria and Ghana: Twice as many sources detected by immunoelectron microscopy (IEM) than by TAS-ELISA or IC-PCR

Our previous study had shown that some Musa leaf samples with Banana streak symptoms tested negative for Banana streak virus (BSV) in triple antibody-sandwich ELISA (TAS-ELISA). Therefore, in this study 63 additional Musa leaf samples were tested for BSV by TAS-ELISA, immunoelectron microscopy (IEM) and immunocapture polymerase chain reaction (IC-PCR). Sensitivity tests by sap dilution end-point analyses indicated that IC-PCR was considerably more sensitive than IEM fordetecting typical BSV, while IEM proved to be of similar sensitivity as TAS-ELISA. However, when leaf samples of Musa plants, obtained from different farmers’ fields in Nigeria and Ghana and some Nigeriansources maintained in the greenhouse were screened for BSV, more than twice as many samples revealed BSV-like particles by IEM than were detected by TAS-ELISA or IC-PCR. Of the 51 leaf samplesthat were BSV positive in all tests taken together, 48 were positive by IEM, 25 by IC-PCR and only 19 by TAS-ELISA. Upon IEM examination, typical bacilliform BSV-like particles were clearly recognized although in very diverse concentrations. Bacilliform particles deviating in length from the main particle populations or showing an angularly bent morphology were found. Occasionally, in certain samples and with certain antisera the IEM decoration tests revealed mixtures of strongly decorated and weaklydecorated BSV-like particles or bacilliform particles which did not at all react with the antibodies available. This proved, the occurrence, besides the presence of typical BSV, of diverse populations of BSV-like viruses in West Afric

Historical Contingencies Modulate the Adaptability of Rice Yellow Mottle Virus

The rymv1-2 and rymv1-3 alleles of the RYMV1 resistance to Rice yellow mottle virus (RYMV), coded by an eIF(iso)4G1 gene, occur in a few cultivars of the Asiatic (Oryza sativa) and African (O. glaberrima) rice species, respectively. The most salient feature of the resistance breaking (RB) process is the converse genetic barrier to rymv1-2 and rymv1-3 resistance breakdown. This specificity is modulated by the amino acid (glutamic acid vs. threonine) at codon 49 of the Viral Protein genome-linked (VPg), a position which is adjacent to the virulence codons 48 and 52. Isolates with a glutamic acid (E) do not overcome rymv1-3 whereas those with a threonine (T) rarely overcome rymv1-2. We found that isolates with T49 had a strong selective advantage over isolates with E49 in O. glaberrima susceptible cultivars. This explains the fixation of the mutation T49 during RYMV evolution and accounts for the diversifying selection estimated at codon 49. Better adapted to O. glaberrima, isolates with T49 are also more prone than isolates with E49 to fix rymv1-3 RB mutations at codon 52 in resistant O. glaberrima cultivars. However, subsequent genetic constraints impaired the ability of isolates with T49 to fix rymv1-2 RB mutations at codons 48 and 52 in resistant O. sativa cultivars. The origin and role of the amino acid at codon 49 of the VPg exemplifies the importance of historical contingencies in the ability of RYMV to overcome RYMV1 resistance

Digital Gene Expression Analysis Based on Integrated De Novo Transcriptome Assembly of Sweet Potato [Ipomoea batatas (L.) Lam.]

Background: Sweet potato (Ipomoea batatas L. [Lam.]) ranks among the top six most important food crops in the world. It is widely grown throughout the world with high and stable yield, strong adaptability, rich nutrient content, and multiple uses. However, little is known about the molecular biology of this important non-model organism due to lack of genomic resources. Hence, studies based on high-throughput sequencing technologies are needed to get a comprehensive and integrated genomic resource and better understanding of gene expression patterns in different tissues and at various developmental stages. Methodology/Principal Findings: Illumina paired-end (PE) RNA-Sequencing was performed, and generated 48.7 million of 75 bp PE reads. These reads were de novo assembled into 128,052 transcripts ($100 bp), which correspond to 41.1 million base pairs, by using a combined assembly strategy. Transcripts were annotated by Blast2GO and 51,763 transcripts got BLASTX hits, in which 39,677 transcripts have GO terms and 14,117 have ECs that are associated with 147 KEGG pathways. Furthermore, transcriptome differences of seven tissues were analyzed by using Illumina digital gene expression (DGE) tag profiling and numerous differentially and specifically expressed transcripts were identified. Moreover, the expression characteristics of genes involved in viral genomes, starch metabolism and potential stress tolerance and insect resistance were also identified

Virus diseases of cowpea in Africa: IITA Research Guide, No. 53

Eight viruses are reported from cowpeas in Africa. Cowpea viruses are described from lowland, humid and sub-humid West Africa, as well as from mid-altitude ecological zones mainly found in eastern and southern Africa. Wherever facilities are limited and detailed characterization is not possible, a simple procedure using a combination of serology (agar-gel diffusion test), inoculation to test plants, and vector transmission studies, is proposed for the identification of the cowpea viruses reported so far from Africa. The possibilities of controlling these viruses are briefly discussed