Investigates the ability of plant extracts from Lens culinaris to protect zucchini from the Zucchini yellow mosaic virus (ZYMV).

Evaluate the impact of extracts from the Lens culinaris plant on a number of physiological and biochemical parameters in squash leaves infected with ZYMV in this work. Compared to the untreated leaves, ZYMV infected leaves showed a range of symptoms, such as severe mosaic, size reduction, stunting, and deformation. Analysis of physiological data revealed that L. culinaris extract lectin therapies and viral infections had an impact on metabolism. Protein, carbohydrate, and pigment levels were all lowered by viral infection. However, phenolic compounds, total protein, total carbohydrates, total amino acids, proline, total chlorophyll and peroxidases levels are considerably elevated with all extract therapies. The other biochemical parameters also displayed a variety of changes. Moreover shoot length, number of leaves and number of flowers was significantly increased compared to viral control in all treatments. The L. culinaris extract treatment increases the plant's ZYMV resistance. This is detectable through reduction of the plants treated with lentil lectin pre and post virus inoculation, reduction in disease severity and viral concentration, and percentage of the infected plants has a virus. All findings demonstrate significant metabolic alterations brought by viral infections or L. culinaris extract treatments, and they also suggest that exogenous extract treatments is essential for activating the body's defences against ZYMV infection.

Investigations on annual spreading of viruses infecting cucurbit crops in Uttar Pradesh State, India.

During 2018 an intensive study was conducted to determine the viruses associated with cucurbitaceous crops in nine agroclimatic zones of the state of Uttar Pradesh, India. Total of 563 samples collected and analysed across 14 different cucurbitaceous crops. The results showed the dominance of Begomovirus (93%) followed by Potyvirus (46%), cucumber green mottle mosaic virus (CGMMV-39%), Polerovirus (9%), cucumber mosaic virus (CMV-2%) and Orthotospovirus (2%). Nearly 65% of samples were co-infected with more than one virus. Additionally, host range expansion of CMV, CGMMV and polerovirus was also observed on cucurbit crops. A new potyvirus species, zucchini tigre mosaic virus, earlier not documented from India has also been identified on five crops during the study. Risk map generated using ArcGIS for virus disease incidence predicted the virus severity in unexplored areas. The distribution pattern of different cucurbit viruses throughout Uttar Pradesh will help identify the hot spots for viruses and will facilitate to devise efficient and eco-friendly integrated management strategies for the mitigation of viruses infecting cucurbit crops. Molecular diversity and evolutionary relationship of the virus isolates infecting cucurbits in Uttar Pradesh with previously reported strains were understood from the phylogenetic analysis. Diverse virus infections observed in the Eastern Plain zone, Central zone and North-Eastern Plain zone indicate an alarming situation for the cultivation of cucurbits in the foreseeable future.

Analysis of the molecular and biological variability of Zucchini yellow mosaic virus isolates from Iran and Iraq.

During the growing season of 2018, several field-grown cucurbit plants in different parts of Iraq and Iran were surveyed for the presence of zucchini yellow mosaic virus (ZYMV), using two degenerate primer pairs (CIF/Rev and NIb2F/3R) targeting the two separated partial regions of the potyvirus genome (CI and NIb respectively). 7 out of 20 samples were confirmed to be infected with ZYMV. Phylogenetic analyses based on the CI gene grouped all Iranian and two Iraqi (ZYMV1 and ZYMV2) isolates together with isolates from the Middle East in the subgroup (AI), whereas the other Iraqi (ZYMV3 and ZYMV4) isolates were clustered in the subgroup (DI), which was only consisted of American isolates. The highest and lowest identity between the studied isolates and the GenBank isolates showed that the two genes (CI, NIb) of each isolate particularly the Iraqi isolates were more similar to a specific and geographically scattered mosaic of worldwide isolates, suggestive of mixed infection might have occurred between different worldwide isolates in Iraq. Furthermore, the first complete nucleotide sequence of an Iraqi ZYMV (ZYMV-Iq) isolate was done, using the Illumina sequencing technique. The complete nucleotide sequence of ZYMV-Iq isolate was 9650 nt, excluding the 3'poly (A) tail. ZYMV-Iq isolate shared the highest nt identity of 98.8% with an American (KC665630) isolate. Phylogenetic analysis based on the full genome sequence placed ZYMV-Iq in subgroup A of group I alongside 18 isolates from the US and two isolates from Australia. In addition, recombination analysis detected lone significant recombination between ZYMV-Iq and South Korean (AY279000) isolate. Moreover, the results showed that symptom intensity was varied across experimental host plants.

Genome characterization of zucchini yellow mosaic virus infecting cucurbits reveals the presence of a new genotype in Trinidad and Tobago in the Caribbean region.

Zucchini yellow mosaic virus (ZYMV) is a member of the genus Potyvirus that is becoming a serious pathogen of pumpkin and other cucurbits in Trinidad and Tobago and the entire Caribbean region. In this study, four ZYMV isolates infecting pumpkin in Trinidad and Tobago were characterized by complete genome sequencing. Phylogenetic analysis showed 5.9-6.0% nt and 7.7-7.9% aa sequence divergence in comparison to the most closely related isolates NAT and AG from Israel and SE04T from Slovakia. Based on the variations in the complete genome sequence as well as individual gene sequences, a new genotype, designated ZYMV-Trini, is proposed for these isolates. Among the gene sequences of ZYMV-Trini isolates, the greatest variation was observed in the HC-Pro gene, with 20.8% aa sequence divergence from their closest relatives, whereas the least variation was observed in the NIb, P3, and CP genes, with 1.8-2.2% aa sequence divergence. This study also showed that transmission of ZYMV can occur through seeds, but this was less common than transmission via the aphid Aphis gossypii. The progression of ZYMV in pumpkin seedlings was quantified by RT-qPCR, which showed a rapid surge in viral load after 37 days. From recombination analysis, it could be concluded that the isolates SE04T from Slovakia, NAT from Israel, and AG from Israel have made major contributions to the genome architecture of ZYMV-Trini isolates.

Identification, molecular characterization and prevalence of a novel cytorhabdovirus infecting zucchini crops in Greece.

A new cytorhabdovirus was identified in zucchini (Cucurbita pepo) in Greece with the aid of high-throughput sequencing technology. The negative-sense, single-stranded genomic RNA of the new virus was determined and includes seven open reading frames in the order 3'-N-P-P3-P4-M-G-L-5' in the antigenomic orientation. Typical rhabdovirus-like particles were observed in infected leaf material. Comparative sequence analysis and phylogenetic reconstructions suggested that the described virus is a new member of the genus Cytorhabdovirus, and it was tentatively named cucurbit cytorhabdovirus 1 (CuCV1). To our knowledge CuCV1 is the first cytorhabdovirus infecting cucurbits in nature. Our surveys indicated that it occurs in a percentage of 36.7 % in zucchini crops in Greece.

Distribution and evolution of the major viruses infecting cucurbitaceous and solanaceous crops in the French Mediterranean area.

Plant viral diseases represent a significant burden to plant health, and their highest impact in Mediterranean agriculture is on vegetables grown under intensive horticultural practices. In order to understand better virus evolution and emergence, the most prevalent viruses were mapped in the main cucurbitaceous (melon, squashes) and solanaceous (tomato, pepper) crops and in some wild hosts in the French Mediterranean area, and virus diversity, evolution and population structure were studied through molecular epidemiology approaches. Surveys were performed in summer 2016 and 2017, representing a total of 1530 crop samples and 280 weed samples. The plant samples were analysed using serological and molecular approaches, including high-throughput sequencing (HTS). The viral species and their frequency in crops were quite similar to those of surveys conducted ten years before in the same areas. Contrary to other Mediterranean countries, aphid-transmitted viruses remain the most prevalent in France whereas whitefly-transmitted ones have not yet emerged. However, HTS analysis of viral evolution revealed the appearance of undescribed viral variants, especially for watermelon mosaic virus (WMV) in cucurbits, or variants not present in France before, as for cucumber mosaic virus (CMV) in solanaceous crops. Deep sequencing also revealed complex virus populations within individual plants with frequent recombination or reassortment. The spatial genetic structure of cucurbit aphid-borne yellows virus (CABYV) was related to the landscape structure, whereas in the case of WMV, the recurrence of introduction events and probable human exchanges of plant material resulted in complex spatial pattern of genetic variation.

A rapid fluorescence-based real-time isothermal assay for the detection of Cucurbit yellow stunting disorder virus in squash and watermelon plants.

Cucurbit yellow stunting disorder virus (CYSDV) is a single-stranded positive-sense RNA virus that produces devastating disease in watermelon and squash. Foliar symptoms of CYSDV consist of interveinal yellowing, brittleness, and thickening of older leaves leading to reduced plant vigor. A rapid diagnostic method for CYSDV would facilitate early detection and implementation of best viral-based management practices. We developed a rapid isothermal reverse transcription-recombination polymerase amplification (exo RT-RPA) assay for the detection of CYSDV. The primers and a 6-fluorescein amidite (6-FAM) probe were developed to target the nucleocapsid gene. The real-time assay detected CYSDV at 2.5 pg purified total RNA extracted from CYSDV-infected leaf tissue and corresponded to 10 copies of the target molecule. The assay was specific and did not cross-react with other common cucurbit viruses found in Florida and Georgia. The performance of the exo RT-RPA was evaluated using crude extract from 21 cucurbit field samples and demonstrated that the exo RT-RPA is a rapid procedure, thus providing a promising novel alternative approach for the detection of CYSDV.

Genetic variability of watermelon mosaic virus isolates infecting cucurbit crops in Italy.

Watermelon mosaic virus (WMV; genus Potyvirus, family Potyviridae) is responsible for serious cucurbit yield losses worldwide. Different WMV genetic groups have been characterized so far. Among these, the "classical" (CL) group has been present in the Mediterranean basin for 40 years, whereas the "emergent" (EM) group includes isolates that are associated with more-severe symptoms observed since 2000. Information on the spatial and temporal evolution of WMV isolates in Italy is currently sparse. In this study, 39 WMV isolates samples collected in different regions over the last two decades were analysed at two different genomic regions that are known to be highly variable and contain recombination breakpoints. Most of the isolates collected between 2002 and 2009 were found to belong to the CL group, whereas the isolates from 2012 onwards were classified as EM, indicating that EM isolates have progressively displaced the CL population in Italy. Although genetic variability was observed within both CL and EM groups and recombinant isolates were detected, no positive selection or haplotype geographic structure were inferred. This suggest that the shift from CL to EM populations was likely due to multiple introductions of EM isolates in different regions of Italy rather than from genetic differentiation of local populations. The progressive increase in prevalence of the highly virulent EM populations is a serious concern because of their symptom severity, and the presence of multiple EM variants that include recombinants necessitates new efforts to develop durable control strategies.

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.

High-throughput sequencing of virus-infected Cucurbita pepo samples revealed the presence of Zucchini shoestring virus in Zimbabwe.

OBJECTIVES: Plant-infecting viruses remain a serious challenge towards achieving food security worldwide. Cucurbit virus surveys were conducted in Zimbabwe during the 2014 and 2015 growing seasons. Leaf samples displaying virus-like symptoms were collected and stored until analysis. Three baby marrow samples were subjected to next-generation sequencing and the data generated were analysed using genomics technologies. Zucchini shoestring virus (ZSSV), a cucurbit-infecting potyvirus previously described in South Africa was one of the viruses identified. The genomes of the three ZSSV isolates are described analysed in this note. RESULTS: The three ZSSV isolates had the same genome size of 10,297 bp excluding the polyA tail with a 43% GC content. The large open reading frame was found at positions 69 to 10,106 on the genome and encodes a 3345 amino acids long polyprotein which had the same cleavage site sequences as those described on the South African isolate except for the P1-pro site. Genome sequence comparisons of all the ZSSV isolates showed that the isolates F7-Art and S6-Prime had identical sequence across the entire genome while sharing 99.06% and 99.34% polyprotein nucleotide and amino acid sequence identities, respectively with the isolate S7-Prime.

A Plant SMALL RNA-BINDING PROTEIN 1 Family Mediates Cell-to-Cell Trafficking of RNAi Signals.

In plants, RNA interference (RNAi) plays a pivotal role in growth and development, and responses to environmental inputs, including pathogen attack. The intercellular and systemic trafficking of small interfering RNA (siRNA)/microRNA (miRNA) is a central component in this regulatory pathway. Currently, little is known with regards to the molecular agents involved in the movement of these si/miRNAs. To address this situation, we employed a biochemical approach to identify and characterize a conserved SMALL RNA-BINDING PROTEIN 1 (SRBP1) family that mediates non-cell-autonomous small RNA (sRNA) trafficking. In Arabidopsis, AtSRBP1 is a glycine-rich (GR) RNA-binding protein, also known as AtGRP7, which we show binds single-stranded siRNA. A viral vector, Zucchini yellow mosaic virus (ZYMV), was employed to functionally characterized the AtSRBP1-4 (AtGRP7/2/4/8) RNA recognition motif and GR domains. Cellular-based studies revealed the GR domain as being necessary and sufficient for SRBP1 cell-to-cell movement. Taken together, our findings provide a foundation for future research into the mechanism and function of mobile sRNA signaling agents in plants.

Phylogenetic classification of a group of self-replicating RNAs that are common in co-infections with poleroviruses.

Tombusvirus-like associated RNAs (tlaRNAs) are positive-sense single-stranded RNAs found in plants co-infected with viruses of the genus Polerovirus. TlaRNAs depend upon capsid proteins supplied in trans by the co-infecting polerovirus vector for transmission and intra-host systemic movement. Here, the full-length genomes of five tlaRNAs were determined using a combination of RT-PCR and next-generation sequencing, and evidence is provided for an additional tlaRNA associated with potato leafroll virus. Phylogenetic analyses based on conserved domains of the RdRp placed tlaRNAs as a monophyletic clade clustering with members of the family Tombusviridae and comprising three different subclades. Full-length clones of tlaRNAs from two of three subclades were confirmed to replicate autonomously, and each produces a subgenomic RNA during infection.

Simultaneous detection of three common potyviruses infecting cucurbits by multiplex reverse transcription polymerase chain reaction assay.

Watermelon mosaic virus (WMV), Zucchini yellow mosaic virus (ZYMV), and Papaya ring spot virus (PRSV-W) are the three most common potyviruses infecting cucurbits in the United States of America. In this study, a Multiplex reverse transcription polymerase chain reaction (RT-PCR) assay was developed for the simultaneous detection and differentiation of WMV, ZYMV, and PRSV-W. A mixture of specific primers set for each virus successfully amplified a distinct PCR product of 980 bp from the coat protein (CP) gene of WMV, 708 bp from the cylindrical inclusion (CI) gene of ZYMV and 496 bp from the helper component (Hc-Pro) gene of PRSV-W. Sanger sequencing confirmed that the amplified PCR products for each virus are specific. The sensitivity and specificity of these primers were tested by serial dilution assay of total RNA extracted from virus-infected samples for all three viruses. The detection limit of the Multiplex RT-PCR assay was between 10-5-10-6-fold dilution. The Multiplex RT-PCR assay was successfully applied for the detection of these viruses on 54 field samples, which were collected from two counties of Oklahoma during the 2018 growing season. Based on these results, this Multiplex RT-PCR assay is specific, rapid, and economical for the detection of three common potyviruses of cucurbits and has the potential to screen a large number of field samples of cucurbits against these viruses.

Whitefly Control Strategies against Tomato Leaf Curl New Delhi Virus in Greenhouse Zucchini.

(1) Background: Tomato leaf curl New Delhi virus (ToLCNDV), transmitted by tobacco whitefly (Bemisia tabaci Gennadius) (Hemiptera: Aleyrodidae), is of major concern in the cultivation of zucchini. The threat of this virus motivates reliance on chemical vector control but European consumers' demands for vegetables grown free of pesticides provides an important incentive for alternative pest management; (2) Methods: Different whitefly management strategies and ToLCNDV incidences were surveyed in commercial zucchini greenhouses in south-east Spain. In an experimental greenhouse, three different whitefly control strategies, biological, chemical, and integrated (IPM), were evaluated in a replicated trial to determine the most effective strategy for virus suppression (3) Results: Whitefly was present in all commercial zucchini crops surveyed, whereas fewer crops had Amblyseius swirskii or other natural enemies. During three consecutive years, pest management was increasingly based on chemical treatments. Yet, ToLCNDV was widespread in zucchini greenhouses. Experimental results showed that the order of best strategy for virus suppressing was integrated management (73%) > biological control (58%) > chemical control (44%); and (4) Conclusions: IPM was the best strategy for virus suppression. The results can assist in the design of appropriate control strategies for chemical pesticide reduction and decision-making in pest management.

Characterization and in silico proteomic analysis of C2 and C3 proteins of squash leaf curl China virus associated with pumpkin leaf curl disease in Assam, India.

The pumpkin leaf curl disease is an emerging disease of pumpkin in Assam, India. Symptomatic pumpkin leaf samples from different locations were immunologically tested using Begomovirus specific antibody. PCR with the ELISA-positive samples, using Geminivirus universal primers amplified 1.4 kb virus-specific fragments. Sequence of these amplicons showed around 95% identity with squash leaf curl China virus-[Pumpkin: Varanasi] (SLCCV-Pumpkin: Varanasi EU573715). To investigate the possible functions of the viral proteins present in the fragment, the full-length C2 and C3 genes were conceptually translated and were subjected to in silico proteomic analyses. The phylogenetic analysis of both the proteins divulged the relationship of our isolate with related viruses and isolates. Multiple sequence alignment (MSA) of the proteins revealed the presence of the known viral conserved motifs, viz., zinc-finger (ZNF) motif [36CXCX(7)CX(6)H53], the arginine-rich nuclear localization signal (NLS) motif (28RRRR31) as well as the minimal activation domain in C2 protein. In the C3 protein, the 91LKYLD95 and the replication enhancer motif (30YFK32) were found to be conserved. Finally, 3-D models of the two proteins were predicted via ab initio approach and subsequently, the models were validated. To our knowledge, this study is a pioneering attempt to construct the ab initio 3-D models of two begomoviral proteins taking a SLCCV isolate as a model. Keywords: begomovirus; ELISA; ZNF motif; NLS motif; ab initio modelling.

RNA phloem transport mediated by pre-miRNA and viral tRNA-like structures.

Phloem-mobile mRNAs are assumed to contain sequence elements directing RNA to the phloem translocation pathway. One of such elements is represented by tRNA sequences embedded in untranslated regions of many mRNAs, including those proved to be mobile. Genomic RNAs of a number of plant viruses possess a 3'-terminal tRNA-like structures (TLSs) only distantly related to genuine tRNAs, but nevertheless aminoacylated and capable of interaction with some tRNA-binding proteins. Here, we elaborated an experimental system for analysis of RNA phloem transport based on an engineered RNA of Potato virus X capable of replication, but not encapsidation and movement in plants. The TLSs of Brome mosaic virus, Tobacco mosaic virus and Turnip yellow mosaic virus were demonstrated to enable the phloem transport of foreign RNA. A miRNA precursor, pre-miR390b, was also found to render RNA competent for the phloem transport. In line with this, sequences of miRNA precursors were identified in a Cucurbita maxima phloem transcriptome, supporting the hypothesis that, at least in some cases, miRNA phloem signaling can involve miRNA precursors. Collectively, the data presented here suggest that RNA molecules can be directed into the phloem translocation pathway by structured RNA elements such as those of viral TLSs and miRNA precursors.

Detection and genome characterization of a novel member of the genus Polerovirus from zucchini (Cucurbita pepo) in China.

Here, we report a novel member of the genus Polerovirus, zucchini aphid-borne yellows virus (ZABYV), which was identified in zucchini grown for seed production in the Xinjiang Uygur Autonomous Region, China. The complete nucleotide sequence of the ZABYV genome was determined and found to be 5,792 nucleotides in length, and like those of other poleroviruses, to contain seven open reading frames (ORFs). Multiple sequence alignments and phylogenetic analysis indicated that ZABYV is a new member of the genus Polerovirus, although several regions of its genome are closely related to chickpea chlorotic stunt virus (CpCSV). Further comparative analysis suggested that ZABYV originated from a recombination event between CpCSV and another unknown virus in the genus Polerovirus.

Molecular characterisation of a putative new polerovirus infecting pumpkin (Cucurbita pepo) in Kenya.

Next-generation sequencing of RNA extracted from a pumpkin plant with mosaic symptoms in Kenya identified the presence of a polerovirus sequence closely related to pepo aphid-borne yellows virus (PABYV). The near-complete polerovirus sequence comprised 5,810 nucleotides and contained seven putative open reading frames (ORFs) with a genome organisation typical of poleroviruses. BLASTp analysis of the translated sequences of ORFs 0, 1 and 2 revealed that their amino acid sequences differed by more than 10% from the corresponding protein sequences of other poleroviruses. These results suggest that this virus is a putative novel member of the genus Polerovirus, which has been provisionally named "pumpkin polerovirus" (PuPV).

Cross protection against the watermelon strain of Papaya ringspot virus through modification of viral RNA silencing suppressor.

Papaya ringspot virus watermelon strain (PRSV-W) causes huge economic losses to cucurbits production. Here, we constructed an infectious clone of PRSV-W, pCamPRSV-W, which can induce similar symptoms and accumulate to same levels as wild type virus in plants of Cucurbita pepo, Cucumis melo, Citrullus lanatus and Cucumis sativus. The green fluorescence protein gene gfp was cloned into pCamPRSV-W to produce pCamPRSV-W-GFP, which produced strong green fluorescence in systemic leaves of inoculated Cucurbita pepo, Cucumis melo, Citrullus lanatus and Cucumis sativus plants, indicating that pCamPRSV-W can be used to express foreign genes. Ten mutants of PRSV-W, obtained by site-directed mutagenesis in the RNA silencing suppressor helper-component proteinase encoding region, produced dramatically attenuated symptoms in plants of Cucumis melo. The Cucumis melo plants pre-infected with mutants K125D and G317 K showed effective protection against the challenge inoculation of wild type PRSV-W. The attenuated mutants generated in this study will be helpful for the eco-friendly control of PRSV-W.

Impact of Cultivated Hosts on the Recombination of Cucumber Mosaic Virus.

Cucumber mosaic virus (CMV) is one of the most successful viruses known, infecting over 1,200 species of plants. Like other single-stranded RNA viruses, CMV is known to have a high potential for population diversity due to error-prone replication and short generation times. Recombination is also a mechanism that allows viruses to adapt to new hosts. Host genes have been identified that impact the recombination of RNA viruses by using single-cell yeast systems. To determine the impact that the natural plant host has on virus recombination, we used a high-recombination-frequency strain of CMV, LS-CMV, which belongs to subgroup II, in three different cultivated hosts: Capsicum annuum cv. Marengo (pepper), Nicotiana tabacum cv. Xanthi nc (tobacco), and Cucurbita pepo cv. Black Beauty (zucchini). The recombination frequency was calculated by using an RNA 3 reporter carrying restriction enzyme sites created by introducing silent mutations. Our results show that the recombination frequency of LS-CMV is correlated with the infected host. The recombination events in pepper were 1.8-fold higher than those in tobacco and 5-fold higher than those in zucchini. Furthermore, we observed the generation of defective RNAs in inoculated pepper plants, but not in tobacco or zucchini. These results indicate that the host is involved in both intra- and intermolecular recombination events and that hosts like pepper could foster more rapid evolution of the virus. In addition, we report for the first time the production of defective RNAs in a CMV subgroup II isolate.IMPORTANCE Recombination is an important mechanism used by viruses for their diversification and to adapt to diverse hosts. Understanding the host role in the mechanisms of evolution is important for virus disease management and controlling the emergence of new strains. This study shows the impact that cultivated hosts are playing in the evolution of CMV. Furthermore, our results and previous studies show how some specific hosts could be an ideal environment for the emergence of new viral strains.