Synergistic interactions of begomoviruses with Sweet potato chlorotic stunt virus (genus Crinivirus) in sweet potato (Ipomoea batatas L.)

Three hundred and ninety‐four sweet potato accessions from Latin America and East Africa were screened by polymerase chain reaction (PCR) for the presence of begomoviruses, and 46 were found to be positive. All were symptomless in sweet potato and generated leaf curling and/or chlorosis in Ipomoea setosa. The five most divergent isolates, based on complete genome sequences, were used to study interactions with Sweet potato chlorotic stunt virus (SPCSV), known to cause synergistic diseases with other viruses. Co‐infections led to increased titres of begomoviruses and decreased titres of SPCSV in all cases, although the extent of the changes varied notably between begomovirus isolates. Symptoms of leaf curling only developed temporarily in combination with isolate StV1 and coincided with the presence of the highest begomovirus concentrations in the plant. Small interfering RNA (siRNA) sequence analysis revealed that co‐infection of SPCSV with isolate StV1 led to relatively increased siRNA targeting of the central part of the SPCSV genome and a reduction in targeting of the genomic ends, but no changes to the targeting of StV1 relative to single infection of either virus. These changes were not observed in the interaction between SPCSV and the RNA virus Sweet potato feathery mottle virus (genus Potyvirus), implying specific effects of begomoviruses on RNA silencing of SPCSV in dually infected plants. Infection in RNase3‐expressing transgenic plants showed that this protein was sufficient to mediate this synergistic interaction with DNA viruses, similar to RNA viruses, but exposed distinct effects on RNA silencing when RNase3 was expressed from its native virus, or constitutively from a transgene, despite a similar pathogenic outcome.     

The incidence of sweet potato virus disease and virus resistance of sweet potato grown in Uganda

Sweet potato virus disease (SPVD) was common (25–30% average incidences), and farmers recognised it as an important disease, in sweet potato crops in southern Mpigi, Masaka and Rakai Districts in Uganda, but SPVD was rare in Soroti and Tororo Districts. Whiteflies, which are the vector of sweet potato chlorotic stunt crinivirus (SPCSV) a component cause of SPVD, were correspondingly common on sweet potato crops in Mpigi and rare on crops in Tororo. However, aphids, which are the vectors of sweet potato feathery mottle potyvirus (SPFMV), the other component cause of SPVD, were not found colonising sweet potato crops, and itinerant alate aphids may be the means of transmission. Different sweet potato cultivars were predominant in the different districts surveyed and four local cultivars obtained from Kanoni in S. Mpigi, where whiteflies and SPVD were common, were more resistant to SPVD than four cultivars from Busia in Tororo District, where whiteflies and SPVD were rare. However, nationally released cultivars were even more resistant than the local cultivars from Kanoni. Yield results and interviews with farmers indicated that farmers in S. Mpigi were making compromises in their choice of cultivars to grow, some key factors being SPVD susceptibility, and the yield, taste, and marketability, duration of harvest and in-ground storability of the storage roots. These compromises need to be included in an assessment of yield losses attributable to SPVD.     

Aspects of resistance to sweet potato virus disease in sweet potato

In field trials during the first and the second rainy season of 1996 in Uganda, whiteflies were similarly abundant and aphids were absent on three clones of sweet potato (NIS-93–63, cv. Tanzania and cv. New Kawogo) although the three clones differed considerably in their resistance to sweet potato virus disease (SPVD), a complex disease resulting from infection by both the aphid-borne sweet potato feathery mottle virus (SPFMV) and the whitefly-borne sweet potato chlorotic stunt virus (SPCSV). This suggests that vector resistance does not determine the relative SPVD resistance of these genotypes. SPFMV alone had only a low virus titre in sweet potato cvs Tanzania and New Kawogo, became increasingly difficult to detect in plants of these cultivars and was seldom acquired by aphids. However, this resistance to SPFMV was not apparent in plants which were also infected with SPCSV. Plants then had a high SPFMV titre, appeared unable to eliminate SPFMV and provided good sources for aphids to acquire it.     

Molecular genetic analysis of virus isolates from wild and cultivated plants demonstrates that East Africa is a hotspot for the evolution and diversification of Sweet potato feathery mottle virus

Sweet potato feathery mottle virus (SPFMV, genus Potyvirus) is globally the most common pathogen of cultivated sweet potatoes (Ipomoea batatas; Convolvulaceae). Although more than 150 SPFMV isolates have been sequence‐characterized from cultivated sweet potatos across the world, little is known about SPFMV isolates from wild hosts and the evolutionary forces shaping SPFMV population structures. In this study, 46 SPFMV isolates from 14 wild species of genera Ipomoea, Hewittia and Lepistemon (barcoded for the matK gene in this study) and 13 isolates from cultivated sweet potatoes were partially sequenced. Wild plants were infected with the EA, C or O strain, or co‐infected with the EA and C strains of SPFMV. In East Africa, SPFMV populations in wild species and sweet potato were genetically undifferentiated, suggesting inter‐host transmission of SPFMV. Globally, spatial diversification of the 178 isolates analysed was observed, strain EA being largely geographically restricted to East Africa. Recombination was frequently detected in the 6K2‐VPg‐NIaPro region of the EA strain, demonstrating a recombination ‘hotspot’. Recombination between strains EA and C was rare, despite their frequent co‐infections in wild plants, suggesting purifying selection against strain EA/C recombinants. Positive selection was predicted on 17 amino acids distributed over the entire coat protein in the globally distributed strain C, as compared to only four amino acids in the coat protein N‐terminus of the EA strain. This selection implies a more recent introduction of the C strain and a higher adaptation of the EA strain to the local ecosystem. Thus, East Africa appears as a hotspot for evolution and diversification of SPFMV.     

A critical domain of Sweet potato chlorotic fleck virus nucleotide‐binding protein (NaBp) for RNA silencing suppression,nuclear localization and viral pathogenesis

RNA silencing is an important mechanism of antiviral defence in plants. To counteract this resistance mechanism, many viruses have evolved RNA silencing suppressors. In this study, we analysed five proteins encoded by Sweet potato chlorotic fleck virus (SPCFV) for their abilities to suppress RNA silencing using a green fluorescent protein (GFP)‐based transient expression assay in Nicotiana benthamiana line 16c plants. Our results showed that a putative nucleotide‐binding protein (NaBp), but not other proteins encoded by the virus, could efficiently suppress local and systemic RNA silencing induced by either sense or double‐stranded RNA (dsRNA) molecules. Deletion mutation analysis of NaBp demonstrated that the basic motif (an arginine‐rich region) was critical for its RNA silencing suppression activity. Using confocal laser scanning microscopy imaging of transfected protoplasts expressing NaBp fused to GFP, we showed that NaBp accumulated predominantly in the nucleus. Mutational analysis of NaBp demonstrated that the basic motif represented part of the nuclear localization signal. In addition, we demonstrated that the basic motif in NaBp was a pathogenicity determinant in the Potato virus X (PVX) heterogeneous system. Overall, our results demonstrate that the basic motif of SPCFV NaBp plays a critical role in RNA silencing suppression, nuclear localization and viral pathogenesis.