Calystegia hederacea: a new natural host of Sweet potato latent virus in China.

Abstract

Sweet potato is a global root crop, with a worldwide production of 91.5 million tons in 2019 (FAOSTAT, 2019). However, virus diseases cause significant yield losses and quality decline in sweet potato. Up to now, over 30 different viruses have been identified in sweet potato (Clark et al. 2012). Expanding knowledge of the host range of sweet potato viruses will provide a benefit for the understanding of virus occurrence and designing appropriate virus control measures. In August 2019, ten Calystegia hederacea and two Convolvulus arvensis (Convolvulaceae) weed plants with or without symptoms of leaf yellowing symptoms were collected from various virus disease-affected sweet potato fields in four cities (Jiaozuo, Xinxiang, Zhengzhou and Kaifeng) of Henan Province for virus detection. The leaves of these plants were harvested and pooled for total RNA extraction using a Plant Total RNA Purification Kit (GMbiolab, Taichung, Taiwan). A library for high-throughput sequencing (HTS) was constructed and sequenced using the Illumina HiSeq 2000 platform by BGI Tech (Shenzhen, China). Clean reads (n = 100,570,346), each 150 bp in length, were de novo assembled using CLC Genomics Workbench 9.5 (Qiagen, USA). The assembled contigs were analyzed against the viral reference genome database in GenBank using the BLASTN and BLASTX searches. Three contigs related to sweet potato chlorotic stunt virus (SPCSV, genus Crinivirus, family Closteroviridae) were identified (Liu et al. 2021). In addition, a total of 20 contigs, ranging from 1,019 to 9,859 bp in length with an average depth of coverage of 1439.26, showed 74.80-87.59% nucleotide (nt) sequence identities with corresponding sequences of sweet potato latent virus (SPLV, genus Potyvirus, family Potyviridae). The sequence of the 9,859-bp contig covering nearly complete genome sequence for SPLV, was deposited in GenBank (accession no.OL625609). These results demonstrated the presence of genetically diverse isolates of SPLV in the pooled samples. To further confirm the HTS result, each of the 12 samples were tested by RT-PCR using SPLV primers (SPLV-F1: 5'-AATGCCAAGGCTACAAGGAGT-3' and SPLV-R1: 5'-CAAGTAGTGTGTGTATGTTCC-3') that targets a partial conserved region of the coat protein gene in SPLV and SPCSV primers designed based on three contigs (ctg1-F1/R1, ctg2-F1/R1, and ctg3-F1/R1) (Liu et al. 2021), respectively. As a result, four symptomless C. hederacea samples tested positive for SPLV, yielding the expected approximately 500 bp PCR fragment, and one leaf yellowing C. hederacea sample tested positive for SPCSV (Liu et al. 2021). The sequences obtained from two of the four amplicons of SPLV (MZ089700 and OM056706) showed 90.2 and 89.8% nt (100 and 99.4% amino acid) identities with the corresponding sequences of the SPLV isolate Shaanxi1 from sweet potato (HQ844148). In 2021, a further 45 C. hederacea plants collected from Shangqiu (n = 6), Xinxiang (n =30) and Pingdingshan (n = 9) cities in Henan Province, were screened by RT-PCR with SPLV-F1/R1 primers, giving an incidence of 33.33%. SPLV is an important potyvirus infecting sweet potato. SPLV is asymptomatic in most sweet potato cultivars in single infection but is able to mediate synergistic viral disease in co-infection with SPCSV (Untiveros et al. 2007). To the best of our knowledge, this is the first report of SPLV in C. hederacea. The finding reported here indicated that C. hederacea may act as a reservoir of SPLV and possible infection source for the sweet potato crop.