Epidemiology of Zucchini yellow mosaic virus in cucurbit crops in a remote tropical environment.

In the remote Ord River Irrigation Area (ORIA) in tropical northwest Australia, severe Zucchini yellow mosaic virus (ZYMV) epidemics threaten dry season (April-October) cucurbit crops. In 2016-2017, wet season (November-March) sampling studies found a low incidence ZYMV infection in wild Cucumis melo and Citrullus lanatus var. citroides plants, and both volunteer and garden crop cucurbits. Such infections enable its persistence in the wet season, and act as reservoirs for its spread to commercial cucurbit crops during the dry season. Tests on 1019 samples belonging to 55 species from 23 non-cucurbitaceous plant families failed to detect ZYMV. It was also absent from wild cucurbit weeds within sandalwood plantations. The transmission efficiencies of a local isolate by five aphid species found in the ORIA were: 10 % (Aphis craccivora), 7% (A. gossypii), 4% (A. nerii), and 0% (Rhopalosiphum maidis and Hysteroneura setariae). In 2016-2017, in all-year-round trapping at five representative sites, numbers of winged aphids caught were greatest in July-August (i.e. mid growing season) but varied widely between trap sites reflecting local aphid host abundance and year. Apart from one localised exception in 2017, flying aphid numbers caught and ZYMV spread in data collection blocks during 2015-2017 resembled what occurred commercial cucurbit crops. When ZYMV spread from external infection sources into melon blocks, its predominant spread pattern consisted of 1 or 2 plant infection foci often occurring at their margins. In addition, when plants of 29 cucurbit cultivars were inoculated with an ORIA isolate and two other ZYMV isolates and the phenotypes elicited were compared, they resembled each other in overall virulence. However, depending upon isolate-cultivar combination, differences in symptom expression and severity occurred, and one isolate caused a systemic hypersensitive phenotype in honeydew melon cvs Estilo and Whitehaven. When the new genomic RNA sequences of 19 Australian isolates were analysed, all seven ORIA isolates fitted within ZYMV phylogroup B, which also included two from southwest Australia, whereas the remaining 10 isolates were all within minor phylogroups A-I or A-II. Based on previous research and the additional knowledge of ZYMV epidemic drivers established here, an integrated disease management strategy targeting ZYMV spread was devised for the ORIA's cucurbit industry.

Cuticular Structure Proteomics in the Pea Aphid Acyrthosiphon pisum Reveals New Plant Virus Receptor Candidates at the Tip of Maxillary Stylets.

Aphids are phloem-feeding insects known as major pests in agriculture that are able to transmit hundreds of plant viruses. The majority of these viruses, classified as noncirculative, are retained and transported on the inner surface of the cuticle of the needle-like mouthparts while the aphids move from plant to plant. Identification of receptors of viruses within insect vectors is a key challenge because they are promising targets for alternative control strategies. The acrostyle, an organ discovered earlier within the common food/salivary canal at the tip of aphid maxillary stylets, displays proteins at the cuticle-fluid interface, some of which are receptors of noncirculative viruses. To assess the presence of stylet- and acrostyle-specific proteins and identify putative receptors, we have developed a comprehensive comparative analysis of the proteomes of four cuticular anatomical structures of the pea aphid, stylets, antennae, legs, and wings. In addition, we performed systematic immunolabeling detection of the cuticular proteins identified by mass spectrometry in dissected stylets. We thereby establish the first proteome of stylets of an insect and determine the minimal repertoire of the cuticular proteins composing the acrostyle. Most importantly, we propose a short list of plant virus receptor candidates, among which RR-1 proteins are remarkably predominant. The data are available via ProteomeXchange (PXD016517).

Identification of Plant Virus Receptor Candidates in the Stylets of Their Aphid Vectors.

Plant viruses transmitted by insects cause tremendous losses in most important crops around the world. The identification of receptors of plant viruses within their insect vectors is a key challenge to understanding the mechanisms of transmission and offers an avenue for future alternative control strategies to limit viral spread. We here report the identification of two cuticular proteins within aphid mouthparts, and we provide experimental support for the role of one of them in the transmission of a noncirculative virus. These two proteins, named Stylin-01 and Stylin-02, belong to the RR-1 cuticular protein subfamily and are highly conserved among aphid species. Using an immunolabeling approach, they were localized in the maxillary stylets of the pea aphid Acyrthosiphon pisum and the green peach aphid Myzus persicae, in the acrostyle, an organ earlier shown to harbor receptors of a noncirculative virus. A peptide motif present at the C termini of both Stylin-01 and Stylin-02 is readily accessible all over the surface of the acrostyle. Competition for in vitro binding to the acrostyle was observed between an antibody targeting this peptide and the helper component protein P2 of Cauliflower mosaic virus Furthermore, silencing the stylin-01 but not stylin-02 gene through RNA interference decreased the efficiency of Cauliflower mosaic virus transmission by Myzus persicae These results identify the first cuticular proteins ever reported within arthropod mouthparts and distinguish Stylin-01 as the best candidate receptor for the aphid transmission of noncirculative plant viruses.IMPORTANCE Most noncirculative plant viruses transmitted by insect vectors bind to their mouthparts. They are acquired and inoculated within seconds when insects hop from plant to plant. The receptors involved remain totally elusive due to a long-standing technical bottleneck in working with insect cuticle. Here we characterize the role of the two first cuticular proteins ever identified in arthropod mouthparts. A domain of these proteins is directly accessible at the surface of the cuticle of the acrostyle, an organ at the tip of aphid stylets. The acrostyle has been shown to bind a plant virus, and we consistently demonstrated that one of the identified proteins is involved in viral transmission. Our findings provide an approach to identify proteins in insect mouthparts and point at an unprecedented gene candidate for a plant virus receptor.

Fasting alters aphid probing behaviour but does not universally increase the transmission rate of non-circulative viruses.

A fasting period prior to non-circulative virus acquisition has been shown to increase the rate of transmission by aphids. However, this effect has only been studied for a few virus-vector combinations, and there are contradictory results in the literature as to the role of fasting on virus acquisition. We analysed the influence of fasting on the transmission of three non-circulative viruses, Cucumber mosaic virus, Zucchini yellow mosaic virus and Cauliflower mosaic virus, by two aphid vector species: Myzus persicae Sulzer (Hemiptera: Aphididae) and Aphis gossypii Glover (Hemiptera: Aphididae). All variables tested, including the virus species and isolate, and the species of aphid, influenced the effect of a fasting period on virus transmission efficiency. Furthermore, when aphids were subjected to an overnight feeding period on a sucrose solution, the fasting effect disappeared and the probing behaviour of these aphids was markedly different to plant-reared aphids. The electrical penetration graph (EPG) technique revealed that fasting altered the probing behaviour of M. persicae and A. gossypii, with fasted aphids beginning to feed sooner and having a significantly longer first intracellular puncture, measured as a potential drop. Significantly longer sub-phase II-3 of the potential drop and more archlets during this sub-phase were also observed for fasted aphids of both species. However, these behavioural changes were not predictive of increasing virus transmission following a fasting period. The impacts of pre-acquisition fasting on aphid probing behaviour and on the mechanisms of non-circulative virus transmission are discussed.

Development and Evaluation of ELISA and qRT-PCR for Identification of Squash vein yellowing virus in Cucurbits.

Squash vein yellowing virus (SqVYV) causes viral watermelon vine decline. To facilitate detection of SqVYV, enzyme linked-immunosorbent assay (ELISA) and quantitative reverse-transcription polymerase chain reaction (qRT-PCR) diagnostic methods were developed. Both methods were capable of detecting SqVYV in a wide range of cucurbit hosts. ELISA was able to detect virus in infected host tissue diluted to at least 1:2,560, which was sufficient for detection in symptomatic squash and watermelon plants. The qRT-PCR method was capable of reliably detecting as few as 3.4 copies of a cloned fragment of SqVYV genomic RNA with an average cycle threshold (Ct) value of 36.4. The sensitivities and specificities for each detection method were estimated by latent class analysis for a set of inoculated squash and watermelon plants at two sampling scales. The scales were hierarchical, with individual plants representing the upper scale and samples from the plant representing the lower scale. The number of samples per plant varied from 1 to 8, and a plant was diagnosed positive if any of its samples tested positive. For all analyses, a cutoff Ct of 35 was chosen for qRT-PCR, which is approximately 2.5 cycles lower than the lowest Ct value achieved for mock-inoculated plants (presumed to be a false positive). qRT-PCR showed high sensitivities (≥0.99) at both sampling scales for squash and watermelon, whereas the sensitivities for ELISA ranged from 0.58 to 0.76. The specificities for both tests were very similar (≥0.94), with ELISA sometimes outperforming qRT-PCR. These diagnostic methods provide additional tools for the identification of SqVYV and management of SqVYV-induced watermelon vine decline.

Role of the Insect Supervectors Bemisia tabaci and Frankliniella occidentalis in the Emergence and Global Spread of Plant Viruses.

Emergence of insect-transmitted plant viruses over the past 10-20 years has been disproportionately driven by two so-called supervectors: the whitefly, Bemisia tabaci, and the Western flower thrips, Frankliniella occidentalis. High rates of reproduction and dispersal, extreme polyphagy, and development of insecticide resistance, together with human activities, have made these insects global pests. These supervectors transmit a diversity of plant viruses by different mechanisms and mediate virus emergence through local evolution, host shifts, mixed infections, and global spread. Associated virus evolution involves reassortment, recombination, and component capture. Emergence of B. tabaci-transmitted geminiviruses (begomoviruses), ipomoviruses, and torradoviruses has led to global disease outbreaks as well as multiple paradigm shifts. Similarly, F. occidentalis has mediated tospovirus host shifts and global dissemination and the emergence of pollen-transmitted ilarviruses. The plant virus-supervector interaction offers exciting opportunities for basic research and global implementation of generalized disease management strategies to reduce economic and environmental impacts.

Emergence of Groundnut ringspot virus and Tomato chlorotic spot virus in Vegetables in Florida and the Southeastern United States.

Groundnut ringspot virus (GRSV) and Tomato chlorotic spot virus (TCSV) are two emerging tospoviruses in Florida. In a survey of the southeastern United States, GRSV and TCSV were frequently detected in solanaceous crops and weeds with tospovirus-like symptoms in south Florida, and occurred sympatrically with Tomato spotted wilt virus (TSWV) in tomato and pepper in south Florida. TSWV was the only tospovirus detected in other survey locations, with the exceptions of GRSV from tomato (Solanum lycopersicum) in South Carolina and New York, both of which are first reports. Impatiens (Impatiens walleriana) and lettuce (Lactuca sativa) were the only non-solanaceous GRSV and/or TCSV hosts identified in experimental host range studies. Little genetic diversity was observed in GRSV and TCSV sequences, likely due to the recent introductions of both viruses. All GRSV isolates characterized were reassortants with the TCSV M RNA. In laboratory transmission studies, Frankliniella schultzei was a more efficient vector of GRSV than F. occidentalis. TCSV was acquired more efficiently than GRSV by F. occidentalis but upon acquisition, transmission frequencies were similar. Further spread of GRSV and TCSV in the United States is possible and detection of mixed infections highlights the opportunity for additional reassortment of tospovirus genomic RNAs.

Spatial and Temporal Analysis of Squash vein yellowing virus Infections in Watermelon.

Squash vein yellowing virus (SqVYV) is a whitefly-transmitted ipomovirus infecting watermelon and other cucurbits that was recently introduced to Florida. Effects on watermelon are devastating, with total vine collapse, often near harvest, and fruit rendered unmarketable by brown, discolored flesh. The epidemiology of SqVYV was studied in a 1-ha field of 'Fiesta' watermelon over six growing seasons (I to VI) to characterize the spatial patterning of disease and temporal rate of disease progress, as well as its association with Cucurbit leaf crumple virus (CuLCrV) and Cucurbit yellow stunting disorder virus (CYSDV), two additional whitefly-transmitted viruses that often occur with SqVYV. The field was scouted at regular intervals for the length of the season for incidence of virus and number of whiteflies. Incidence of SqVYV reached 100% during seasons I, II, and V and 20% during season III. SqVYV did not occur during seasons IV and VI. SqVYV progressed in a characteristic logistic fashion in seasons I, II, and V but less so in season III. The rate of disease progress was similar for the three seasons with high disease incidence, with an average value of 0.18. A positive correlation between the area under the disease progress curve and whitefly-days was found, where both progress curves were calculated as a function of thermal time (degree days, base 0°C). SqVYV displayed significant but variable levels of aggregation, as indicated by its fit to the ß-binomial distribution, the binary power law, and ordinary runs analysis. Association analysis indicated that the viruses were largely transmitted independently. Results of this study provide epidemiological information that will be useful in the development of management strategies for SqVYV-induced vine decline, and provide new information for CuLCrV and CYSDV.

The use of latent class analysis to estimate the sensitivities and specificities of diagnostic tests for Squash vein yellowing virus in cucurbit species when there is no gold standard.

Squash vein yellowing virus (SqVYV) is the causal agent of viral watermelon vine decline, one of the most serious diseases in watermelon (Citrullus lanatus L.) production in the southeastern United States. At present, there is not a gold standard diagnostic test for determining the true status of SqVYV infection in plants. Current diagnostic methods for identification of SqVYV-infected plants or tissues are based on the reverse-transcription polymerase chain reaction (RT-PCR), tissue blot nucleic acid hybridization assays (TB), and expression of visual symptoms. A quantitative assessment of the performance of these diagnostic tests is lacking, which may lead to an incorrect interpretation of results. In this study, latent class analysis (LCA) was used to estimate the sensitivities and specificities of RT-PCR, TB, and visual assessment of symptoms as diagnostic tests for SqVYV. The LCA model assumes that the observed diagnostic test responses are linked to an underlying latent (nonobserved) disease status of the population, and can be used to estimate sensitivity and specificity of the individual tests, as well as to derive an estimate of the incidence of disease when a gold standard test does not exist. LCA can also be expanded to evaluate the effect of factors and was done here to determine whether diagnostic test performances varied among the type of plant tissue being tested (crown versus vine tissue), where plant samples were taken relative to the position of the crown (i.e., distance from the crown), host (i.e., genus), and habitat (field-grown versus greenhouse-grown plants). Results showed that RT-PCR had the highest sensitivity (0.94) and specificity (0.98) of the three tests. TB had better sensitivity than symptoms for detection of SqVYV infection (0.70 versus 0.32), while the visual assessment of symptoms was more specific than TB and, thus, a better indicator of noninfection (0.98 versus 0.65). With respect to the grouping variables, RT-PCR and TB had better sensitivity but poorer specificity for diagnosing SqVYV infection in crown tissue than it did in vine tissue, whereas symptoms had very poor sensitivity but excellent specificity in both tissues for all cucurbits analyzed in this study. Test performance also varied with habitat and genus but not with distance from the crown. The results given here provide quantitative measurements of test performance for a range of conditions and provide the information needed to interpret test results when tests are used in parallel or serial combination for a diagnosis.

Squash vein yellowing virus Infection of Vining Cucurbits and the Vine Decline Response.

The responses of a diverse group of vining cucurbits to inoculation with Squash vein yellowing virus (SqVYV) were determined. For the first time, Cucurbita maxima, Cucumis dipsaceus, and Cucumis metuliferus were observed to develop necrosis and plant death similar to the SqVYV-induced vine decline in watermelon (Citrullus lanatus var. lanatus). The majority of cucurbits inoculated, however, either exhibited no symptoms of infection, or developed relatively mild symptoms such as vein yellowing of upper, noninoculated leaves. All inoculated plants were sectioned and tested for the presence of SqVYV. The virus was widely distributed in mature, fruit-bearing cucurbits with over 72% of plant sections testing positive for SqVYV by tissue-blot and/or reverse transcription-polymerase chain reaction. Plants of several cucurbits, including a wild citron (Citrullus lanatus var. citroides), were symptomless and had a decreased frequency of virus infection of vine segments compared to susceptible vining cucurbits, indicating a higher level of resistance. However, no significant relationship between the frequency of infection or virus distribution within plants and the symptom response was observed. These results demonstrate that a diverse group of cucurbits may decline when infected with SqVYV, and suggest that widespread distribution of virus within the plant is not the sole cause of decline.

Physiological Effects of Squash vein yellowing virus Infection on Watermelon.

Squash vein yellowing virus (SqVYV) is the cause of viral watermelon vine decline. The virus is whitefly-transmitted, induces a systemic wilt of watermelon plants, and causes necrosis and discoloration of the fruit rind. In the field, SqVYV is often detected in watermelon in mixed infections with other viruses including the aphid-transmitted Papaya ringspot virus type W (PRSV-W). In this study, watermelon plants of different ages were inoculated with SqVYV or SqVYV+PRSV-W in the greenhouse or SqVYV in the field to characterize the physiological response to infection. Symptoms of vine decline appeared about 12 to 16 days after inoculation with SqVYV regardless of plant age at time of inoculation, plant growth habit (trellised or nontrellised), and location (greenhouse or field). However, the presence of PRSV-W delayed the appearance of vine decline symptoms by 2 to 4 days, and vine decline did not develop on plants with no fruit. For all inoculation treatments, more severe symptoms were observed in younger watermelon plants. Physiological responses to SqVYV infection included reduction in plant and fruit weights, alterations in fruit rind and flesh color, reduction in fruit sucrose content, increase in fruit acid content, and changes in plant nutrient composition, particularly increases in Ca, Mg, B, Mn, and Zn and decreases in K and N. These results demonstrate wide-ranging physiological effects of SqVYV infection and provide new insights into watermelon vine decline.

Low genetic diversity of Squash vein yellowing virus in wild and cultivated cucurbits in the U.S. suggests a recent introduction.

Squash vein yellowing virus (SqVYV) isolates were collected from cultivated and weedy cucurbits representing major hosts and locations in the U.S. and analyzed to better understand the diversity and population structure. No differences in symptoms were observed in field-collected isolate source plants or subsequently inoculated greenhouse plants, and the complete genome of an SqVYV isolate from a wild cucurbit host (smellmelon, Cucumis melo var. dudaim) was highly similar (99.4% nucleotide identity, 99.3% amino acid identity) to the previously published type isolate from squash. Although analysis of the coat protein (CP) and two serine proteases (P1a and P1b) sequences for 41 isolates showed little diversity across seven years of sampling, it revealed two distinct groups of SqVYV isolates with low intra-group diversity. Our analyses also suggested that recombination had occurred between SqVYV isolates, similar to other ipomoviruses. Selection pressures on the genome regions analyzed were negative indicating purifying selection was occurring. The magnitude of negative selection in SqVYV was consistent with what has been reported for other ipomoviruses, and was greatest for the CP and least for the P1b. The observed genetic diversity was similar to that reported for Cucumber vein yellowing virus but less than that reported for Sweet potato mild mottle virus, Cassava brown streak virus and Ugandan cassava brown streak virus. Collectively, these results indicate that the current U.S. population of SqVYV has undergone a recent genetic bottleneck and was introduced from elsewhere.

Indigenous and introduced potyviruses of legumes and Passiflora spp. from Australia: biological properties and comparison of coat protein nucleotide sequences.

Five Australian potyviruses, passion fruit woodiness virus (PWV), passiflora mosaic virus (PaMV), passiflora virus Y, clitoria chlorosis virus (ClCV) and hardenbergia mosaic virus (HarMV), and two introduced potyviruses, bean common mosaic virus (BCMV) and cowpea aphid-borne mosaic virus (CAbMV), were detected in nine wild or cultivated Passiflora and legume species growing in tropical, subtropical or Mediterranean climatic regions of Western Australia. When ClCV (1), PaMV (1), PaVY (8) and PWV (5) isolates were inoculated to 15 plant species, PWV and two PaVY P. foetida isolates infected P. edulis and P. caerulea readily but legumes only occasionally. Another PaVY P. foetida isolate resembled five PaVY legume isolates in infecting legumes readily but not infecting P. edulis. PaMV resembled PaVY legume isolates in legumes but also infected P. edulis. ClCV did not infect P. edulis or P. caerulea and behaved differently from PaVY legume isolates and PaMV when inoculated to two legume species. When complete coat protein (CP) nucleotide (nt) sequences of 33 new isolates were compared with 41 others, PWV (8), HarMV (4), PaMV (1) and ClCV (1) were within a large group of Australian isolates, while PaVY (14), CAbMV (1) and BCMV (3) isolates were in three other groups. Variation among PWV and PaVY isolates was sufficient for division into four clades each (I-IV). A variable block of 56 amino acid residues at the N-terminal region of the CPs of PaMV and ClCV distinguished them from PWV. Comparison of PWV, PaMV and ClCV CP sequences showed that nt identities were both above and below the 76-77% potyvirus species threshold level. This research gives insights into invasion of new hosts by potyviruses at the natural vegetation and cultivated area interface, and illustrates the potential of indigenous viruses to emerge to infect introduced plants.

Ecology and management of whitefly-transmitted viruses of vegetable crops in Florida.

A variety of fresh market vegetables, including watermelon and tomato are economically important crops in Florida. Whitefly-transmitted Squash vein yellowing virus (SqVYV) was first identified in squash and watermelon in Florida in 2005 and shown to cause a severe decline of watermelon vines as crops approach harvest. Florida is most economically impacted by SqVYV, although the virus has been detected more recently in Indiana and South Carolina. The origin and evolutionary history of SqVYV, one of the few members of the genus Ipomovirus within the family Potyviridae, are not known. Sequence diversity of SqVYV isolates collected at different times, from different locations and from different plant species is being analyzed for insights into the origin of the virus. More recently, Cucurbit leaf crumple virus (CuLCrV) and Cucurbit yellow stunting disorder virus (CYSDV), also whitefly-transmitted, have been detected in watermelon in Florida. Tomato yellow leaf curl virus (TYLCV) was first detected in south Florida tomato crops in 1997. Several surveys have been conducted in the region to identify alternative hosts for these four viruses. Cucurbit weeds including Balsam-apple (Momordica charantia), creeping cucumber (Melothria pendula) and smellmelon (Cucumis melo var. dudaim) provide reservoirs for SqVYV, CuLCrV and/or CYSDV. Green bean (Phaseolus vulgaris) also can be a reservoir for CuLCrV. No wild hosts of TYLCV have been reported in Florida. The effectiveness of insecticides and silver plastic mulch to manage whiteflies and mitigate TYLCV has been demonstrated and is currently being evaluated for SqVYV, CuLCrV and CYSDV. In addition, potential sources of SqVYV resistance have been identified in greenhouse and field screening of watermelon germplasm. Further studies to refine these sources of resistance are underway. Lastly, a comprehensive map of 33,560 hectares (82,928 acres) of vegetable fields in the three counties comprising the majority of the southwest Florida vegetable production area has been developed to identify 'hot spots' and reservoir crops for viruses and whiteflies, and will be useful in evaluation of management strategies to decrease virus incidence in commercial fields.

A natural M RNA reassortant arising from two species of plant- and insect-infecting bunyaviruses and comparison of its sequence and biological properties to parental species.

Reassortment allows multicomponent viruses to exchange genome segments, a process well-documented in the vertebrate- and arthropod-infecting members of the family Bunyaviridae but not between distinct species of the plant- and insect-infecting members of the genus Tospovirus. Genome sequence comparisons of a virus causing severe tospovirus-like symptoms in Florida tomato with Groundnut ringspot virus (GRSV) and Tomato chlorotic spot virus (TCSV) demonstrated that reassortment has occurred, with the large (L) and small (S) RNAs coming from GRSV and the medium (M) RNA coming from TCSV (i.e. L(G)M(T)S(G)). Neither parental genotype is known to occur in the U.S. suggesting that L(G)M(T)S(G) was introduced as a reassortant. L(G)M(T)S(G) was transmitted by western flower thrips (Frankliniella occidentalis [Pergande]), and was not able to overcome the Sw5 resistance gene of tomato. Our demonstration of reassortment between GRSV and TCSV suggests caution in defining species within the family Bunyaviridae based on their ability to reassort.

Widespread Occurrence and Low Genetic Diversity of Colombian datura virus in Brugmansia Suggest an Anthropogenic Role in Virus Selection and Spread.

Brugmansia (Brugmansia spp.) is a perennial shrub in the Solanaceae, originating from South America, that is a popular landscape plant in the tropics and subtropics and container plant in temperate regions. Virus-like symptoms including mosaic, rugosity, and faint chlorotic spots were first observed on leaves of Brugmansia plants in a south Florida nursery in November 2003. Colombian datura virus (CDV) was identified in these initial plants and subsequent Brugmansia and Datura metel (a Brugmansia relative also grown as an ornamental) plants obtained from Florida, Connecticut, Wisconsin, and California. Overall, 77.5% of Brugmansia and two of four D. metel plants tested were infected with CDV. Partial NIb/CP sequences of 28 Brugmansia CDV isolates from this study were compared with all 16 CDV isolates in GenBank and found to share high levels of nucleotide and amino acid identity, with negative selection estimated to be occurring. A single Brugmansia plant was also infected with a recently described tobamovirus. The low genetic diversity of CDV observed, along with negative selection pressure on NIb/CP, suggests a recent ancestry (<400 years) of the worldwide population of CDV, coinciding with anthropogenic collection and dissemination of Brugmansia plants from their center of origin.