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

Genetic fingerprinting and phylogenetic diversity of Staphylococcus aureus isolates from Nigeria

Genetic fingerprinting of 18 different isolates of Staphylococcus aureus from Nigeria using random amplified polymorphic DNA (RAPD) was carried out. Ten out of 100 Operon primers showed polymorphism among the isolates tested generating 88 bands, 51 of which were polymorphic with sizes ranging between 200 and 3,000 bp. All the isolates were classified completely into two major groups (Sa-1 and Sa-2) with twelve different subgroups. Sa-1 group originated from human while isolates from plant and animal origins formed the Sa-2 group. The twelve different subgroups suggest adaptation of S. aureus in the different host cells. This indicates possible relationship between host origin and genetic variation among S. aureus isolates. The DNA fingerprint defined for each race of S. aureus could be useful in epidemiological studies, medical diagnosis and the identification of new strains and their origins. (African Journal of Biotechnology: 2003 2(8): 246-250

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

Relationship between natural occurrence of banana streak badnavirus and symptom expression, relative concentration of viral antigen, and yield characteristics of some micropropagated Musa spp.

Micropropagated plants of 36 Musa genotypes with diverse genetic backgrounds, including 14 tetraploid plantain (TMPx) and banana (TMBx) hybrids, were evaluated for their response to banana streak badnavirus (BSV) infection under three environments from 1995 to 1997 in Nigeria. The characteristics evaluated were the natural incidence of BSV based on symptoms and virus indexing, relative concentration of BSV antigens in leaf tissues determined by ELISA, and some growth and yield descriptors. Virus occurrence and symptom expression, as well as the relative concentration of BSV antigens, fluctuated greatly between seasons during the cropping cycle, being high during the rainy season and low or negligible during the hot dry season. The natural incidence of plants with symptoms and BSV-infected plants varied between genotypes. Incidence of BSV on most International Institute of Tropical Agriculture (IITA) TMPx hybrids and three Fundación Hondureòa de Investigación Agrìcola (FHIA) hybrids was high in the three environments, with some variation. Most landraces and some FHIA or Empresa Brasileira de Pesquisa Agropecuaria (EMBRAPA) hybrids were not BSV-infected under either environment at Onne. However, a few expressed some foliar symptoms at Ibadan and indexed BSV positive. The relative concentration of BSV antigens in leaf samples was also high in most TMPx and some FHIA hybrids, but low in most landraces. While BSV infection had no significant effect on most growth characteristics, it had a highly variable effect on bunch weight loss among the genotypes. There was no relationship between the natural incidence of BSV, concentration of viral antigen and bunch weight loss among the 11 TMPx hybrids, three FHIA hybrids and three plantain landraces. Despite the high natural BSV incidence and the high relative antigen concentration in their leaf tissue, TMPx 548-9, TMPx 2637-49, TMPx 7002-1 and FHIA 21 suffered less than 15% bunch weight loss, and TMPx 548-4 and FHIA 22 suffered no loss. These results suggest that under the conditions specified in this study, these hybrids could be tentatively classified as ‘field tolerant’ to BS

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

Identification of okra mosaic virus from Indigofera spicata in Nigeria

Okra mosaic virus (OMV, tymovirus group) was isolated from Indigofera spicata plants growing at the International Institute of Tropical Agriculture (IITA) in Ibadan, Nigeria. Its identity was established on the basis of particle morphology, analysis of viral coat protein and nucleic acid and Indigofera spicata. In reciprocal agar gel diffusion tests, the virus isolate from I. spicata and an OMV isolate from okra in Ibadan (OMV-Ibadan isolate) were found to be serologically identical. However, because the isolates differ in symptom induction in various host plants, the name OMV-Indigofera isolate is suggested. This is the first report on the occurence of OMV in I