First report of Groundnut ringspot virus (GRSV) infecting tobacco (Nicotiana tabacum L.) in South Africa.

Abstract

Tobacco (Nicotiana tabacum L.) is an important industrial crop from the Solanaceae family cultivated for its leaves in approximately 100 countries worldwide. In South Africa (SA), tobacco is grown in five of its nine provinces: Limpopo, North West, Western Cape, Eastern Cape, and Mpumalanga. The producer price for unmanufactured tobacco is approximately US$3570/ton annually (FAO, 2021). Emerging and recurrent plant viruses such as tomato spotted wilt orthortospovirus (TSWV) have seriously affected tobacco production yield and quality. Between November 2018 to April 2019 growing season, large thrips (order Thysanoptera) populations were observed in tobacco fields throughout South Africa's major tobacco-growing areas, and plants were showing severe necrosis, yellowing, stunted growth, ringspot, and leaf curling symptoms, which are usually associated with plant virus infections. The disease incidence was estimated at 30%, as per visual observations. Twenty-two symptomatic leaf tissue samples were collected from tobacco farms in Limpopo, Northwest, and Western cape provinces. Samples were initially screened using the double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with LOEWE®Fast Kit (LOEWE®, Germany), which detectsTSWV, groundnut ringspot orthotospovirus (GRSV), and tomato chlorotic spot orthotospovirus (TCSV). Nineteen of these samples tested positive for orthortospoviruses. Subsequently, a reverse transcription-polymerase chain reaction was performed on all ELISA-positive samples using orthotospovirus-specific primers gM410 & gM870c targeting the NSm gene (Chen et al., 2012). All samples were positive for orthotospoviruses, as indicated by the presence of a 500bp amplicon. Nineteen purified PCR products were sequenced in the forward and reverse directions using Sanger sequencing at the KwaZulu-Natal Research and Innovation Sequencing Platform (KRISP; South Africa). The sequences were aligned using Muscle in MEGA version X (Kumar et al., 2018) to generate the consensus sequences that were then subjected to the standard nucleotide basic local alignment search tool (blastn) on the NCBI website. Blastn analysis showed that 16 of the 19 samples matched TSWV nucleotide sequences, while three of the sequences; LP14 (accession number OL505552), LP18 (accession number OL505553), and NW4 (accession number OL505554), had a nucleotide sequence identity of 96,38%, 96,97% and 97,29% with the GSRV isolate SA-05 (accession number MH742957; Silva et al., 2019) from South Africa, respectively. This is the first report of GRSV infecting tobacco after TSWV was reported in SA. Previously, GSRV was reported on soybean and groundnut in South Africa by Petersen and Morris in (2002) and Silva et al. in (2019). The results from this study lay a foundation for more detailed studies on GRSV, which will result in the development of effective and sustainable control strategies to manage GRSV diseases on tobacco in SA.