Biological and molecular diagnosis of seedborne viruses in cowpea germplasm of geographically diverse sub-Saharan origins

摘要

A total of 983 cowpea accessions obtained from the University of California, Riverside (UCR) Cowpea Repository were analysed for seedborne viruses. A majority of the accessions originated from 11 countries representing different agroclimatic zones in sub-Saharan Africa, and included landraces, local cultivars and breeding lines. Following the initial grow-out tests, 69 cowpea accessions, mostly with symptoms of virus infection, were selected for further evaluation using a combination of host range, reverse transcription-polymerase chain reaction (RT-PCR) and sequence analyses. The analyses revealed that samples from 46 (67%) accessions harboured one or more known seedborne viruses of cowpea. These included seed samples of accessions originating from Botswana (13 accessions), Ghana (6), Nigeria (6), Mali (1), Kenya (5), Cameroon (7), Niger (4), Cote d'lvoire (1), Benin (1), India (1) and China (1). Viruses were identified by RT-PCR analysis of total RNAs extracted from suspected virus-infected samples using virus species-specific primers, as well as the cloning and sequencing of RT-PCR products amplified using virus genus- and family-specific degenerate oligonucleotide primers. The viruses identified included Cowpea aphid-borne mosaic virus (CABMV), Cucumber mosaic virus (CMV) and Southern bean mosaic virus (SBMV). Phylogenetic analysis of the deduced coat protein (CP) amino acid sequences of selected CMV isolates recovered from five agroclimatically distinct locations confirmed their affiliations as new members of CMV subgroup IB. This is the first time that seedborne viruses of cowpea accessions in a major collection (UCR) have been identified using RT-PCR and sequencing approaches.