Spatial distribution and molecular characterization of persistently aphid-transmitted viruses causing yellowing and stunting in faba bean and chickpea crops in Tunisia

Abstract

Field surveys were conducted in the northeastern and northwestern regions of Tunisia between 2013/2014 and 2018/2019 growing seasons to identify viruses that infect faba bean and chickpea crops. Field observations showed that 18.8% of the faba bean and 21.0% of the chickpea fields surveyed had virus-like symptoms. These rates exceeded 20% during the growing seasons from 2013/2014 to 2018/2019, and were most common in the 2014/2015 growing season (28.6% of surveyed faba bean fields and 37.5% of chickpea fields showed virus-like symptoms in more than 20%). Totals of 1,538 faba bean and 1,511 chickpea plant samples showing yellowing and stunting symptoms were collected from 144 faba bean and 124 chickpea fields, . All collected samples were tested by tissue blot immunoassay (TBIA) using six monoclonal antibodies. These results showed that chickpea chlorotic stunt virus (CpCSV; Polerovirus CPCSV) was the most prevalent in faba bean and chickpea, with incidences of 20.2% and 37.6%, respectively, followed by beet western yellows virus (BWYV; Polerovirus BWYV) (13.5 and 9.7%), bean leafroll virus (BLRV; Luteovirus phaseoli) (3.7 and 3.3%), and faba bean necrotic yellows virus (FBNYV; Nanovirus necroflaviviciae) detected only in faba bean (6.6% of faba bean samples tested). In addition, TBIA results indicated that single virus infections were more prevalent than mixed infections in both crops. Mixed infections were predominantly co-infections involving viruses in Polerovirus (Solemoviridae), particularly CpCSV and BWYV (93% in faba bean and 69% in chickpea). Twenty-six samples that reacted positively with different monoclonal antibodies were assessed with reverse transcription-polymerase chain reaction (RT-PCR) using generic and specific primers followed by sequencing of the partial coat protein (CP) gene. The sequence analyses confirmed presence of CpCSV, BWYV, BLRV, brassica yellows virus (BrYV; Polerovirus TUYV), and turnip yellows virus (TuYV; Polerovirus TUYV). Comparative sequence analyses of the Tunisian isolates indicated that 13 CpCSV sequences had nucleotide sequence similarities of 95 to 99% with the reference isolate (EU541266) belonging to serotype I, and six BLRV isolates had similarities of 96 to 99% with BLRV reference isolate (PP333098). One sample (TuCp265-19) had a mixed infection with CpCSV and BLRV. Six isolates initially detected using BWYV-specific primers were sequenced and analyzed. BLASTn results showed that only three isolates were closely related (98 to 100%) to BWYV (OM419176), while the remaining four isolates were identified as Polerovirus TUYV and showed greatest similarity to BrYV (LC428361) and TuYV (OP699039), indicating co-occurrence of two distinct Polerovirus species within the analyzed samples. There is no information on the genetic variability of legume viruses in Tunisia, so this study shows that these viruses should be considered when developing disease management strategies to improve faba bean and chickpea production in Tunisia.