First Report of Maize yellow mosaic virus Infecting Maize (Zea mays) in Ethiopia

Abstract

The recent reports of the Polerovirus, Maize yellow mosaic virus (MaYMV) infecting sugarcane (Saccharum spp.) and itch grass (Rottboellia cochinchinensis) in Nigeria (Yahaya et al. 2017) and maize in Burkina Faso (Palanga et al. 2017) provided the motivation for this study. To determine the presence of MaYMV or other poleroviruses, we explored a collection of maize samples of local cultivars taken during virus surveys in August 2015 and July 2016 to assess the distribution of maize lethal necrosis disease (MLND) in three major maize cultivation regions of Ethiopia including Benishangul-Gumuz, Oromia, and South Nations, Nationalities, and People (SNNP). The collection comprised 47 leaf samples from symptomatic maize plants showing yellowing, whitish to yellowish stripes, and mosaic symptoms maintained as dried tissues over anhydrous calcium chloride. Total RNA was extracted using an RNA miniprep kit (Epoch Life Science) following the manufacturer’s instructions. RNA from each sample was screened by one-step RT-PCR with the universal polerovirus primer pair Gen 1 and Gen 2, comprising 322 nucleotides of the coat protein gene (Afouda et al. 2017). By RT-PCR, 78 and 71% of the samples collected in 2015 and 2016, respectively, tested positive for MaYMV. Direct sequencing of the amplified DNA fragments in both directions confirmed near identity (≥99% nt identity) of all Ethiopian sequences and shared 98 to 99% identity to the reference sequence of MaYMV (KU248489). The shared identity to another maize infecting polerovirus species, Maize yellow dwarf virus RMV (MYDV-RMV, NC_021484), ranged from 80 to 81%. To assess the genome variability of the Ethiopian MaYMV isolates, three isolates from three different provinces were selected to perform a deep sequencing analysis. Three independent Illumina libraries were prepared from the respective total RNA and were run on an Illumina MiSeq platform as described by Afouda et al. (2017). De novo and mapped based genome assembly was performed with Geneious v 10.2.3 (Biomatters LTD, NZ). The complete genome sequence of MaYMV isolate MV115 from Basha province was determined with 5,642 nt (MF684369), which was assembled from 36,673 of 2,003,892 reads with an average coverage of 1,141.7. Two near full-length genome sequences were determined, one for isolate MV32 from Wolenchiti Province (5,498 nt, MF684367, 1,820 reads of 2,517,898, mean coverage was 61.3) and for isolate MV90 from Dugda Bora Province (5,504 nt, MF684368, 878 reads of 1,925,886, mean coverage was 33.1). All three determined sequences of MaYMV shared 99.6% identity to each other. The complete MaYMV genome sequence (5,642 nt) of the Ethiopian isolate (MF684369) shared 96.8% nt identity with the Chinese isolate (KU248489), 98.2% with the Nigerian isolate (KY684356), and 95.4% with the isolate from Ecuador (KY052793). This first report of MaYMV in Ethiopia provides evidence for its nationwide distribution and the prevalence of a virus that may have occurred in Ethiopia for some time but was overlooked in targeted surveys focusing on MLND and other prominent maize diseases.