Anti-SARS-CoV-2 Viral Activity of Sweet Potato Trypsin Inhibitor via Downregulation of TMPRSS2 Activity and ACE2 Expression In Vitro and In Vivo.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic. Known as COVID-19, it has affected billions of people worldwide, claiming millions of lives and posing a continuing threat to humanity. This is considered one of the most extensive pandemics ever recorded in human history, causing significant losses to both life and economies globally. However, the available evidence is currently insufficient to establish the effectiveness and safety of antiviral drugs or vaccines. The entry of the virus into host cells involves binding to angiotensin-converting enzyme 2 (ACE2), a cell surface receptor, via its spike protein. Meanwhile, transmembrane protease serine 2 (TMPRSS2), a host surface protease, cleaves and activates the virus's S protein, thus promoting viral infection. Plant protease inhibitors play a crucial role in protecting plants against insects and/or microorganisms. The major storage proteins in sweet potato roots include sweet potato trypsin inhibitor (SWTI), which accounts for approximately 60% of the total water-soluble protein and has been found to possess a variety of health-promoting properties, including antioxidant, anti-inflammatory, ACE-inhibitory, and anticancer functions. Our study found that SWTI caused a significant reduction in the expression of the ACE2 and TMPRSS2 proteins, without any adverse effects on cells. Therefore, our findings suggest that the ACE2 and TMPRSS2 axis can be targeted via SWTI to potentially inhibit SARS-CoV-2 infection.

Early-Stage Phenotyping of Sweet Potato Virus Disease Caused by Sweet Potato Chlorotic Stunt Virus and Sweet Potato Virus C to Support Breeding.

Sweet potato virus disease (SPVD) is a global constraint to sweetpotato (Ipomoea batatas) production, especially under intensive cultivation in the humid tropics such as East Africa. The objectives of this study were to develop a precision SPVD phenotyping protocol, to find new SPVD-resistant genotypes, and to standardize the first stages of screening for SPVD resistance. The first part of the protocol was based on enzyme-linked immunosorbent assay results for sweet potato chlorotic stunt virus (SPCSV) and sweet potato virus C (SPVC) with adjustments to a negative control (uninfected clone Tanzania) and was performed on a prebreeding population (VZ08) comprising 455 clones and 27 check clones graft inoculated under screenhouse conditions. The second part included field studies with 52 selected clones for SPCSV resistance from VZ08 and 8 checks. In screenhouse conditions, the resistant and susceptible check clones performed as expected; 63 clones from VZ08 exhibited lower relative absorbance values for SPCSV and SPVC than inoculated check Tanzania. Field experiments confirmed SPVD resistance of several clones selected by relative absorbance values (nine resistant clones in two locations; that is, 17.3% of the screenhouse selection), supporting the reliability of our method for SPVD-resistance selection. Two clones were promising, exhibiting high storage root yields of 28.7 to 34.9 t ha-1 and SPVD resistance, based on the proposed selection procedure. This modified serological analysis for SPVD-resistance phenotyping might lead to more efficient development of resistant varieties by reducing costs and time at early stages, and provide solid data for marker-assisted selection with a quantitative tool for classifying resistance.[Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

P1 of Sweet Potato Feathery Mottle Virus Shows Strong Adaptation Capacity, Replacing P1-HCPro in a Chimeric Plum Pox Virus.

Potyviridae is the largest family of plant RNA viruses. Their genomes are expressed through long polyproteins that are usually headed by the leader endopeptidase P1. This protein can be classified as type A or type B based on host proteolytic requirements and RNA silencing suppression (RSS) capacity. The main Potyviridae genus is Potyvirus, and a group of potyviruses infecting sweet potato presents an enlarged P1 protein with a polymerase slippage motif that produces an extra product termed P1N-PISPO. These two proteins display some RSS activity and are expressed followed by HCPro, which appears to be the main RNA silencing suppressor in these viruses. Here, we studied the behavior of the P1 protein of Sweet potato feathery mottle virus (SPFMV) using a viral system based on a canonical potyvirus, Plum pox virus (PPV), and discovered that this protein is able to replace both PPV P1 and HCPro. We also found that P1N-PISPO, produced after polymerase slippage, provides extra RNA silencing suppression capacity to SPFMV P1 in this viral context. In addition, the results showed that presence of two type A P1 proteins was detrimental for viral viability. The ample recombination spectrum that we found in the recovered viruses supports the strong adaptation capacity of P1 proteins and signals the N-terminal part of SPFMV P1 as essential for RSS activity. Further analyses provided data to add extra layers to the evolutionary history of sweet potato-infecting potyvirids. IMPORTANCE Plant viruses represent a major challenge for agriculture worldwide and Potyviridae, being the largest family of plant RNA viruses, is one of the primary players. P1, the leader endopeptidase, is a multifunctional protein that contributes to the successful spread of these viruses over a wide host range. Understanding how P1 proteins work, their dynamic interplay during viral infection, and their evolutionary path is critical for the development of strategic tools to fight the multiple diseases these viruses cause. We focused our efforts on the P1 protein of Sweet potato feathery mottle virus, which is coresponsible for the most devastating disease in sweet potato. The significance of our research is in understanding the capacity of this protein to perform several independent functions, using this knowledge to learn more about P1 proteins in general and the potyvirids infecting this host.

In Vitro Identification and In Vivo Confirmation of Inhibitors for Sweet Potato Chlorotic Stunt Virus RNA Silencing Suppressor, a Viral RNase III.

Sweet potato virus disease (SPVD), caused by synergistic infection of Sweet potato chlorotic stunt virus (SPCSV) and Sweet potato feathery mottle virus (SPFMV), is responsible for substantial yield losses all over the world. However, there are currently no approved treatments for this severe disease. The crucial role played by RNase III of SPCSV (CSR3) as an RNA silencing suppressor during the viruses' synergistic interaction in sweetpotato makes it an ideal drug target for developing antiviral treatment. In this study, high-throughput screening (HTS) of small molecular libraries targeting CSR3 was initiated by a virtual screen using Glide docking, allowing the selection of 6,400 compounds out of 136,353. We subsequently developed and carried out kinetic-based HTS using fluorescence resonance energy transfer technology, which isolated 112 compounds. These compounds were validated with dose-response assays including kinetic-based HTS and binding affinity assays using surface plasmon resonance and microscale thermophoresis. Finally, the interference of the selected compounds with viral accumulation was verified in planta In summary, we identified five compounds belonging to two structural classes that inhibited CSR3 activity and reduced viral accumulation in plants. These results provide the foundation for developing antiviral agents targeting CSR3 to provide new strategies for controlling sweetpotato virus diseases.IMPORTANCE We report here a high-throughput inhibitor identification method that targets a severe sweetpotato virus disease caused by coinfection with two viruses (SPCSV and SPFMV). The disease is responsible for up to 90% yield losses. Specifically, we targeted the RNase III enzyme encoded by SPCSV, which plays an important role in suppressing the RNA silencing defense system of sweetpotato plants. Based on virtual screening, laboratory assays, and confirmation in planta, we identified five compounds that could be used to develop antiviral drugs to combat the most severe sweetpotato virus disease.

Complete genome sequence of a divergent sweet potato chlorotic stunt virus isolate infecting Calystegia hederacea in China.

Sweet potato chlorotic stunt virus (SPCSV; genus Crinivirus, family Closteroviridae) is one of the most destructive viruses infecting sweet potatoes. In this study, we determined the complete genome sequence of an SPCSV-like isolate (CH) from Calystegia hederacea Wall. (Convolvulaceae), a weed species related to sweet potato, by combining next-generation sequencing and rapid amplification of cDNA ends. Comparisons of genome sequences and organization confirmed the classification of CH as SPCSV. However, the sequences and phylogenetic data revealed substantial genetic divergence between CH and all known SPCSV isolates. The amino acid sequence identity between the putative proteins in SPCSV-CH and the corresponding proteins in other known SPCSV isolates in each case was less than 85.0%. Phylogenetic analysis indicated that SPCSV-CH is separate from the groups of the known SPCSV isolates. Additionally, SPCSV-CH RNA1 lacks a p22 gene. A 10.1-kDa putative protein (p10) encoded by a sequence in the 5'-terminal region of RNA2 in SPCSV-CH is much larger than the corresponding protein in all known SPCSV isolates.

Development of novel detection system for sweet potato leaf curl virus using recombinant scFv.

Sweet potato leaf curl virus (SPLCV) causes yield losses in sweet potato cultivation. Diagnostic techniques such as serological detection have been developed because these plant viruses are difficult to treat. Serological assays have been used extensively with recombinant antibodies such as whole immunoglobulin or single-chain variable fragments (scFv). An scFv consists of variable heavy (VH) and variable light (VL) chains joined with a short, flexible peptide linker. An scFv can serve as a diagnostic application using various combinations of variable chains. Two SPLCV-specific scFv clones, F7 and G7, were screened by bio-panning process with a yeast cell which expressed coat protein (CP) of SPLCV. The scFv genes were subcloned and expressed in Escherichia coli. The binding affinity and characteristics of the expressed proteins were confirmed by enzyme-linked immunosorbent assay using SPLCV-infected plant leaves. Virus-specific scFv selection by a combination of yeast-surface display and scFv-phage display can be applied to detection of any virus.

Sweet potato viromes in eight different geographical regions in Korea and two different cultivars.

The sweet potato in the family Convolvulaceae is a dicotyledonous perennial plant. Here, we conducted a comprehensive sweet potato virome study using 10 different libraries from eight regions in Korea and two different sweet potato cultivars by RNA-Sequencing. Comprehensive bioinformatics analyses revealed 10 different virus species infecting sweet potato. Moreover, we identified two novel viruses infecting sweet potato referred to as Sweet potato virus E (SPVE) in the genus Potyvirus and Sweet potato virus F (SPVF) in the genus Carlavirus. Of the identified viruses, Sweet potato feathery mottle virus (SPFMV) was the dominant virus followed by Sweet potato virus C (SPVC) and SPVE in Korea. We obtained a total of 30 viral genomes for eight viruses. Our phylogenetic analyses showed many potyvirus isolates are highly correlated with geographical regions. However, two isolates of SPFMV and a single isolate of Sweet potato virus G (SPVG) were genetically distant from other known isolates. The mutation rate was the highest in SPFMV followed by SPVC and SPVG. Two different sweet potato cultivars, Beni Haruka and Hogammi, were infected by seven and five viruses, respectively. Taken together, we provide a complete list of viruses infecting sweet potato in Korea and diagnostic methods.

An isolate of sweet potato chlorotic stunt virus from Brazil with a distinct genome organization.

Sweet potato chlorotic stunt virus (SPCSV; genus Crinivirus, family Closteroviridae), is an economically important pathogen of sweet potato. In the present work, the nucleotide sequences of two RNA segments of SPCSV (isolate SPCSV-UNB-01) were determined by MiSeq Illumina sequencing of samples of sweet potato plants grafted onto Ipomoea setosa. A comparative analysis of the genome organization of SPCSV-UNB-01 and other SPCSV sequences showed that RNA1 was lacking p22, and p5.1 and that p5.2. was absent in RNA2, indicating a unique genomic pattern. SPCSV-UNB-01 contained longer p6 and p5 regions, with little similarity to orthologous sequences. Sequence comparison did not reveal any previously identified functional domains within these open reading frames (ORFs). No recombination or rearrangement events were detected. Phylogenetic analysis suggested the possibility of separate entries of SPCSV into South America based on the genetic distance between SPCSV-UNB-01 and the Peruvian isolate m2-47. Samples from northeastern Brazil (State of Pernambuco) were positive for SPCSV when tested using specific primers for the major coat protein (CP) gene. This is the first full-length genome sequence of SPCSV-UNB-01 from Brazil.

Comparative Transcriptome Analysis Reveals the Transcriptional Alterations in Growth- and Development-Related Genes in Sweet Potato Plants Infected and Non-Infected by SPFMV, SPV2, and SPVG.

Field co-infection of multiple viruses results in considerable losses in the yield and quality of storage roots in sweet potato. However, little is known about the molecular mechanisms underlying developmental disorders of sweet potato subjected to co-infection by multiple viruses. Here, a comparative transcriptomic analysis was performed to reveal the transcriptional alterations in sweet potato plants infected (VCSP) and non-infected (VFSP) by Sweet potato mild mottle virus (SPFMV), Sweet potato virus Y (SPV2) and Sweet potato virus G (SPVG). A total of 1580 and 12,566 differentially expressed genes (DEGs) were identified in leaves and storage roots of VFSP and VCSP plants, respectively. In leaves, 707 upregulated and 773 downregulated genes were identified, whereas 5653 upregulated and 6913 downregulated genes were identified in storage roots. Gene Ontology (GO) classification and pathway enrichment analysis showed that the expression of genes involved in chloroplast and photosynthesis and brassinosteroid (BR) biosynthesis in leaves and the vitamin biosynthetic process in storage roots was inhibited by co-infection of three viruses: SPFMV, SPV2, and SPVG. This was likely closely related to better photosynthesis and higher contents of Vitamin C (Vc) in storage roots of VFSP than that of VCSP. While some genes involved in ribosome and secondary metabolite-related pathways in leaves and alanine, aspartate, and glutamate metabolism in storage roots displayed higher expression in VCSP than in VFSP. Quantitative real-time PCR analysis demonstrated that the expression patterns of 26 DEGs, including 16 upregulated genes and 10 downregulated genes were consistent with the RNA-seq data from VFSP and VCSP. Taken together, this study integrates the results of morphology, physiology, and comparative transcriptome analyses in leaves and storage roots of VCSP and VFSP to reveal transcriptional alterations in growth- and development-related genes, providing new insight into the molecular mechanisms underlying developmental disorders of sweet potato subjected to co-infection by multiple viruses.

An atypical RNA silencing suppression strategy provides a snapshot of the evolution of sweet potato-infecting potyviruses.

Plant viruses usually encode proteins with RNA silencing suppression (RSS) activity to counteract plant defenses. In Potyvirus, the largest genus in the family Potyviridae, this role is taken over by the multifunctional HCPro, also involved in aphid transmission, polyprotein processing and virion formation. Recently, the large P1 of Sweet potato feathery mottle virus (SPFMV) was characterized finding an extra ORF produced after polymerase slippage, which originates the product P1N-PISPO. Transient expression assays showed that SPFMV P1 and P1N-PISPO presented RSS activity, while HCPro did not. In this work, we analyze possible differences between HCPro of SPFMV and other potyviruses, testing HCPro RSS activity in a transient expression assay, and using a Plum pox virus-based system to test the ability of SPFMV P1N-PISPO and HCPro to serve as RNA silencing suppressors in the context of a viral infection. Our results indicate that not only P1 and P1N-PISPO, but also HCPro display RSS activity when expressed in a suitable context, stressing the importance of the selected experimental system for testing anti-silencing capacity of proteins. The presence of multiple viral silencing suppressors in SPFMV adds complexity to an already intricate RSS system, and provides insight into the hypothetical evolution of sweet potato-infecting potyvirids.

Phylogeographic analysis of the full genome of Sweepovirus to trace virus dispersal and introduction to Korea.

Sweet potato is a vegetatively propagated crop that is produced for both growth in Korean fields and for export out of the country. The viruses that are present in introduced sweet potatoes can spread both domestically and to foreign countries. Determining the time and path of virus movement could help curtail its spread and prevent future dispersal of related viruses. Determining the consequences of past virus and sweet potato dispersal could provide insight into the ecological and economic risks associated with other sweet potato-infecting viral invasions. We therefore applied Bayesian phylogeographic inferences and recombination analyses of the available Sweepovirus sequences (including 25 Korean Sweepovirus genomes) and reconstructed a plausible history of Sweepovirus diversification and movement across the globe. The Mediterranean basin and Central America were found to be the launchpad of global Sweepovirus dispersal. Currently, China and Brazil are acting as convergence regions for Sweepoviruses. Recently reported Korean Sweepovirus isolates were introduced from China in a recent phase and the regions around China and Brazil continue to act as centers of Sweepovirus diversity and sites of ongoing Sweepovirus evolution. The evidence indicates that the region is an epidemiological hotspot, which suggests that novel Sweepovirus variants might be found.

New Isolates of Sweet potato feathery mottle virus and Sweet potato virus C: Biological and Molecular Properties, and Recombination Analysis Based on Complete Genomes.

Sweet potato feathery mottle virus (SPFMV) and Sweet potato virus C (SPVC) isolates were obtained from sweetpotato shoot or tuberous root samples from three widely separated locations in Australia's tropical north (Cairns, Darwin, and Kununurra). The samples were planted in the glasshouse and scions obtained from the plants were graft inoculated to Ipomoea setosa plants. Virus symptoms were recorded in the field in Kununurra and in glasshouse-grown sweetpotato and I. setosa plants. RNA extracts from I. setosa leaf samples were subjected to high-throughput sequencing. New complete SPFMV (n = 17) and SPVC (n = 6) genomic sequences were obtained and compared with 47 sequences from GenBank. Phylogenetic analysis revealed that the 17 new SPFMV genomes all fitted within either major phylogroup A, minor phylogroup II, formerly O; or major phylogroup B, formerly RC. Major phylogroup A's minor phylogroup I, formerly EA, only appeared when recombinants were included. Numbers of SPVC genomes were insufficient to subdivide it into phylogroups. Within phylogroup A's minor phylogroup II, the closest genetic match between an Australian and a Southeast Asian SPFMV sequence was the 97.4% nucleotide identity with an East Timorese sequence. Recombination analysis of the 43 SPFMV and 27 SPVC sequences revealed evidence of 44 recombination events, 16 of which involved interspecies sequence transfers between SPFMV and SPVC and 28 intraspecies transfers, 17 in SPFMV and 11 in SPVC. Within SPFMV, 11 intraspecies recombination events were between different major phylogroups and 6 were between members of the same major phylogroup. Phylogenetic analysis accounting for the detected recombination events within SPFMV sequences yielded evidence of minor phylogroup II and phylogroup B but the five sequences from minor phylogroup I were distributed in two separate groups among the sequences of minor phylogroup II. For the SPVC sequences, phylogenetic analysis accounting for the detected recombination events revealed three major phylogroups (A, B, and C), with major phylogroup A being further subdivided into two minor phylogroups. Within the recombinant genomes of both viruses, their PI, NIa-Pro, NIb, and CP genes contained the highest numbers of recombination breakpoints. The high frequency of interspecies and interphylogroup recombination events reflects the widespread occurrence of mixed SPVC and SPFMV infections within sweetpotato plants. The prevalence of infection in northern Australian sweetpotato samples reinforces the need for improved virus testing in healthy sweetpotato stock programs. Furthermore, evidence of genetic connectivity between Australian and East Timorese SPFMV genomes emphasizes the need for improved biosecurity measures to protect against potentially damaging international virus movements.

Simultaneous detection and differentiation of three Potyviridae viruses in sweet potato by a multiplex TaqMan real time RT-PCR assay.

A multiplex TaqMan real time RT-PCR was developed for detection and differentiation of Sweet potato virus G, Sweet potato latent virus and Sweet potato mild mottle virus in one tube. Amplification and detection of a fluorogenic cytochrome oxidase gene was included as an internal control. The assay was compared with a multiplex RT-PCR developed in the initial study for the detection and differentiation of the three viruses and host 18S rRNA. Primers and/or probes of the two assays were designed from conserved regions of each virus. The two assays were optimized for primers/probes and primer concentrations and thermal cycling conditions. Sensitivity and specificity of the assays were compared each other and with other assay. Both assays were evaluated by 74 field samples original from five different provinces of China. RESULTS: showed that the TaqMan real time RT-PCR offered rapid, sensitive, effective and reliable for the simultaneous detection and differentiation of the three viruses in sweet potato plants. The assay will be useful to quarantine and certification programs and virus surveys when large numbers of samples are tested.

Sweet potato feathery mottle virus and Sweet potato virus C from East Timorese and Australian Sweetpotato: Biological and Molecular Properties, and Biosecurity Implications.

Sweet potato feathery mottle virus (SPFMV) and Sweet potato virus C (SPVC) isolates from sweetpotato were studied to examine genetic connectivity between viruses from Australia and Southeast Asia. East Timorese samples from sweetpotato were sent to Australia on FTA cards. Shoot and tuberous root samples were collected in Australia and planted in the glasshouse, and scions were graft inoculated to Ipomoea setosa plants. Symptoms in infected sweetpotato and I. setosa plants were recorded. RNA extracts from FTA cards and I. setosa leaf samples were subjected to high-throughput sequencing (HTS). Complete genomic sequences (CS) of SPFMV and SPVC (11 each) were obtained by HTS, and coat protein (CP) genes from them were compared with others from GenBank. SPFMV sequences clustered into two major phylogroups (A and B = RC) and two minor phylogroups (EA[I] and O[II]) within A; East Timorese sequences were in EA(I) and O(II), whereas Australian sequences were in O(II) and B(RC). With SPVC, CP trees provided sufficient diversity to distinguish major phylogroups A and B and six minor phylogroups within A (I to VI); East Timorese sequences were in minor phylogroup I, whereas Australian sequences were in minor phylogroups II and VI and in major phylogroup B. With SPFMV, Aus13B grouped with East Timorese sequence TM64B within minor phylogroup O, giving nucleotide sequence identities of 97.4% (CS) and 98.3% (CP). However, the closest match with an Australian sequence was the 97.6% (CS) and 98.7% (CP) nucleotide identity between Aus13B and an Argentinian sequence. With SPVC, closest nucleotide identity matches between Australian and East Timorese sequences were 94.1% with Aus6a and TM68A (CS) and 96.3% with Aus55-4C and TM64A (CP); however neither pair member belonged to the same minor phylogroup. Also, the closest Australian match was 99.1% (CP) nucleotide identity between Aus4C and New Zealand isolate NZ4-4. These first complete genome sequences of SPFMV and SPVC from sweetpotato plantings in the Australian continent and neighboring Southeast Asia suggest at least two (SPFMV) and three (SPVC) separate introductions to Australia since agriculture commenced more than two centuries ago. These findings have major implications for both healthy stock programs and biosecurity management in relation to pathogen entry into Australia and elsewhere.

Genome characterization of sweet potato symptomless virus 1: a mastrevirus with an unusual nonanucleotide sequence.

Complete genomic sequences of nine isolates of sweet potato symptomless virus 1 (SPSMV-1), a virus of the genus Mastrevirus in the family Geminiviridae, were determined from sweet potato accessions from different countries and found to be 2,559-2,602 nucleotides in length. These isolates shared 97-100% genome sequence identity and had an unusual nonanucleotide sequence (TAAGATTCC) in a large intergenic region as well as an additional open reading frame, C3, which is conserved in dicot-infecting mastreviruses.

Diversity of Sweepoviruses Infecting Sweet Potato in China.

Sweepoviruses (a group of begomoviruses that infect plants in the family Convolvulaceae) have monopartite genomes that consist of a circular, single-stranded DNA molecule. Seventy-three complete genomic sequences of sweepoviruses were characterized from the sweet potato samples collected in China. Eight sweepovirus species, including two novel species with proposed names of Sweet potato leaf curl China virus 2 and Sweet potato leaf curl Sichuan virus 2, were identified among these samples. One species, Sweet potato leaf curl Canary virus, was first identified in China. Among the 13 identified strains of Chinese sweepoviruses, 4 were newly discovered. Sweet potato leaf curl virus had the highest frequency (53.4%) of occurrence in the sweet potato samples from China. The similarities among the 73 sweepovirus genomic sequences were between 77.6 and 100.0%. Multiple recombination events were identified, and 16 recombinant sequences were determined. Recombination was observed between different species and between different strains of the same species. Recombination breakpoints were mainly localized on the intergenic region and in three open reading frames (AC1, AV1, and AV2). This study is the first comprehensive report on the genetic diversity of sweepoviruses in China.

Mixed Infections of Four Viruses, the Incidence and Phylogenetic Relationships of Sweet Potato Chlorotic Fleck Virus (Betaflexiviridae) Isolates in Wild Species and Sweetpotatoes in Uganda and Evidence of Distinct Isolates in East Africa.

Viruses infecting wild flora may have a significant negative impact on nearby crops, and vice-versa. Only limited information is available on wild species able to host economically important viruses that infect sweetpotatoes (Ipomoea batatas). In this study, Sweet potato chlorotic fleck virus (SPCFV; Carlavirus, Betaflexiviridae) and Sweet potato chlorotic stunt virus (SPCSV; Crinivirus, Closteroviridae) were surveyed in wild plants of family Convolvulaceae (genera Astripomoea, Ipomoea, Hewittia and Lepistemon) in Uganda. Plants belonging to 26 wild species, including annuals, biannuals and perennials from four agro-ecological zones, were observed for virus-like symptoms in 2004 and 2007 and sampled for virus testing. SPCFV was detected in 84 (2.9%) of 2864 plants tested from 17 species. SPCSV was detected in 66 (5.4%) of the 1224 plants from 12 species sampled in 2007. Some SPCSV-infected plants were also infected with Sweet potato feathery mottle virus (SPFMV; Potyvirus, Potyviridae; 1.3%), Sweet potato mild mottle virus (SPMMV; Ipomovirus, Potyviridae; 0.5%) or both (0.4%), but none of these three viruses were detected in SPCFV-infected plants. Co-infection of SPFMV with SPMMV was detected in 1.2% of plants sampled. Virus-like symptoms were observed in 367 wild plants (12.8%), of which 42 plants (11.4%) were negative for the viruses tested. Almost all (92.4%) the 419 sweetpotato plants sampled from fields close to the tested wild plants displayed virus-like symptoms, and 87.1% were infected with one or more of the four viruses. Phylogenetic and evolutionary analyses of the 3'-proximal genomic region of SPCFV, including the silencing suppressor (NaBP)- and coat protein (CP)-coding regions implicated strong purifying selection on the CP and NaBP, and that the SPCFV strains from East Africa are distinguishable from those from other continents. However, the strains from wild species and sweetpotato were indistinguishable, suggesting reciprocal movement of SPCFV between wild and cultivated Convolvulaceae plants in the field.

Infectivity, effects on helper viruses and whitefly transmission of the deltasatellites associated with sweepoviruses (genus Begomovirus, family Geminiviridae).

Begomoviruses (family Geminiviridae) are whitefly-transmitted viruses with single-stranded DNA genomes that are frequently associated with DNA satellites. These satellites include non-coding satellites, for which the name deltasatellites has been proposed. Although the first deltasatellite was identified in the late 1990s, little is known about the effects they have on infections of their helper begomoviruses. Recently a group of deltasatellites were identified associated with sweepoviruses, a group of phylogenetically distinct begomoviruses that infect plants of the family Convolvulaceae including sweet potato. In this work, the deltasatellites associated with sweepoviruses are shown to be transreplicated and maintained in plants by the virus with which they were identified, sweet potato leaf curl virus (SPLCV). These deltasatellites were shown generally to reduce symptom severity of the virus infection by reducing virus DNA levels. Additionally they were shown to be maintained in plants, and reduce the symptoms induced by two Old World monopartite begomoviruses, tomato yellow leaf curl virus and tomato yellow leaf curl Sardinia virus. Finally one of the satellites was shown to be transmitted plant-to-plant in the presence of SPLCV by the whitefly vector of the virus, Bemisia tabaci, being the first time a deltasatellite has been shown to be insect transmitted.

A novel sweet potato potyvirus open reading frame (ORF) is expressed via polymerase slippage and suppresses RNA silencing.

The single-stranded, positive-sense RNA genome of viruses in the genus Potyvirus encodes a large polyprotein that is cleaved to yield 10 mature proteins. The first three cleavage products are P1, HCpro and P3. An additional short open reading frame (ORF), called pipo, overlaps the P3 region of the polyprotein ORF. Four related potyviruses infecting sweet potato (Ipomoea batatas) are predicted to contain a third ORF, called pispo, which overlaps the 3' third of the P1 region. Recently, pipo has been shown to be expressed via polymerase slippage at a conserved GA6 sequence. Here, we show that pispo is also expressed via polymerase slippage at a GA6 sequence, with higher slippage efficiency (∼5%) than at the pipo site (∼1%). Transient expression of recombinant P1 or the 'transframe' product, P1N-PISPO, in Nicotiana benthamiana suppressed local RNA silencing (RNAi), but only P1N-PISPO inhibited short-distance movement of the silencing signal. These results reveal that polymerase slippage in potyviruses is not limited to pipo expression, but can be co-opted for the evolution and expression of further novel gene products.