First Report of Southern bean mosaic virus infecting common bean in Zambia

Southern bean mosaic virus (SBMV, genus Sobemovirus) is one of the causative agents of mosaic and mottle diseases in common bean (Phaseolus vulgaris) (Tremaine and Hamilton 1983). The ∼4.2 kb (+)ssRNA genome of SBMV is encapsidated in isometric virions of about 30 nm in diameter (Tremaine and Hamilton 1983). During surveys conducted in the Eastern Province of Zambia in April 2018, several common bean plants were observed with virus-like symptoms such as vein clearing, leaf rolling, mosaic, and leaf narrowing. Incidence of symptomatic plants across 20 fields ranged from 23 to 100%, averaging 67.5%. A total of 121 symptomatic leaf samples were collected from 20 farmers’ fields and preserved dry on silica gel. Sixteen samples were randomly selected and pooled into one sample (Mse-1) for total nucleic acid (TNA) extraction as described (Djami-Tchatchou and Straker 2012). The TNA was preserved in RNAStable (Biomatrica, Japan) and shipped to Inqaba Biotechnical Industries (Pretoria, South Africa) for high-throughput sequencing (HTS). HTS was achieved via cDNA synthesis and library construction with the Illumina TruSeq RNA Library Prep Kit (Illumina, San Diego, CA). The cDNA library was sequenced on the Illumina MiSeq platform, generating 4.1 million single raw reads of ∼300 nucleotides (nt) each. De novo assembly of 188,160 virus-aligned reads and their subsequent use to query GenBank with BLASTn resulted in the detection of SBMV, bean common mosaic necrosis virus, bean common mosaic virus, cowpea aphid-borne mosaic virus, and cucumber mosaic virus in the sample. To confirm SBMV occurrence and also validate the HTS results, a pair of virus-specific primers (SBMV-1F, 5′-AGCTGGATTTCCTACCTTTGTG; SBMV-1R, 5′-GGCGTCATCTCCGTTTATCTT) was designed to amplify a 873-bp fragment encompassing 32 bp of the 5′ untranslated region, a 492-bp movement protein (ORF1), and a 353-bp partial P2a gene (ORF2) of SBMV by reverse transcription PCR. Each of the 121 samples was screened for SBMV along with the other four viruses (Ha et al. 2008; Niimi et al. 2003). Twenty-five of the 121 samples (20.7%) tested positive only for SBMV, whereas the remaining 96 samples had different combinations of the other four viruses (data not shown). The SBMV-positive samples had mosaic and vein clearing symptoms. The purified PCR products from three samples (EP-88, Mse-13, and Mse-17) were directly Sanger sequenced and deposited in GenBank (MN653952 to MN653954). Pairwise distance analysis revealed that the three Sanger-derived sequences shared 81 to 100% nt identities with each other. The 4,100-bp-long near-complete genome of SBMV was independently Sanger sequenced from sample Mse-13 (MN326873) and determined to encode all the four predicted ORFs of the virus. In pairwise comparisons, the Mse-13 sequence shared the highest (97.2%) and lowest (89.3%) nt sequence identities with the resistance-breaking strain SBMV-B(ARK) (AF055887) from Arkansas and isolate TZ (MG344643) from Tanzania, respectively. SBMV has been previously reported in common bean in many countries across several continents (Verhoeven et al. 2003). However, to the best of our knowledge, this is the first report of SBMV infecting common bean in Zambia. The result adds to the repertoire of viruses circulating in farmers’ fields in Zambia and further confirms the complexity of probable viruses contributing to suboptimal common bean yields in the country.