BIOLOGICAL CONTROL OF CUCUMBER GREEN MOTTLE MOSAIC VIRUS

Field

[0001] The present application is directed to attenuated plant viruses. More specifically, the present application provides attenuated strains of cucumber green mottle mosaic virus (CGMMV), compositions thereof, and methods of using such virus strains and compositions for biological control of plant disease.

Background

[0002] Cucumber green mottle mosaic virus (CGMMV) is a member of the

Tobamovirus genus in the family Virgaviridae. CGMMV has a 6.4-kb single-stranded, positive-sense RNA genome with a 3’ tRNA-like structure instead of a poly(A) tail.

The genome contains three open reading frames (ORFs) that encode four defined proteins. The first ORF encodes a 129 kDa protein, including methyltransferase and helicase domains required for RNA replication, and a 186 kDa protein produced by readthrough translation of the ORF, including the methyltransferase and helicase domains and an additional RNA-dependent RNA polymerase (RdRp) domain. The remaining two ORFs encode a movement protein (MP) and a coat protein (CP), respectively.

[0003] CGMMV infection causes serious diseases in plants of the family

Cucurbitaceae (cucurbits), including cucumber, pumpkin, watermelon, melon, squash, zucchini, gourds, gherkins and others. CGMMV was first reported in 1935 in the UK and is found in Europe, Asia, the Middle East and, since 2013, in Canada and the United States. CGMMV infection is becoming a major limiting factor with regard to production of cucurbits worldwide, and has become an increasing threat to the commercial production of cucumber and other commercially-grown cucurbit crops.

[0004] CGMMV is a seed-borne virus and can be transmitted by root-to-root contact and by transfer from contaminated seeds, soil, gardening implements such as pruners and stakes, irrigation water, packing materials, or the clothing or hands of farmers, pickers or other persons handling the plants. The CGMMV virus is extremely stable and can remain infectious under relatively extreme conditions for long periods of time. Therefore, the presence of even a few infected plants in a cucumber greenhouse can eventually lead to the spread of CGMMV infection to the entire crop.

[0005] CGMMV is responsible for a wide range of symptoms on leaves and fruits of infected plants, depending on the CGMMV strain, stage of infection and plant species. The induced symptoms include vein clearing and crumpling in young leaves, light green mottling, mosaic patterns, necrotic lesions, fruit distortion or streak, change of sugar accumulation and flavor, and premature degradation of the pulp, making the fruit unmarketable and unfit for consumption. CGMMV infection can result in substantial yield losses up to 100%, although losses of 40-80% are more common in a commercial field or greenhouse production setting, in addition to having a major impact on fruit quality, leading to low market value.

[0006] There are no known effective chemical methods for controlling virus diseases of plants, and crop protection relies completely on aspects of sanitation that may include removal of infected plants, using only virus-free seeds and vegetative stocks, using resistant varieties, and controlling transmission by contact with insects, water and humans. Two new techniques that are now being investigated for managing plant virus diseases rely on cross protection by inoculation with attenuated/mild strains and by creating transgenic or engineered resistance based on expression of the viral coat protein of a specific viral pathogen.

[0007] Cross protection is an acquired immunity phenomenon which has been demonstrated in a number of systems. The technique makes use of the observation that plants infected with an attenuated virus isolate or strain causing mild or no symptoms can develop tolerance to further infection when challenged by an isolate or strain of the same or a related virus species which causes more severe symptoms. Cross protection is virus specific, occurring only between strains of the same virus or related virus species, and is seen as an acceptable method for crop protection in greenhouse systems as it does not rely on harmful materials or chemicals. This approach was first applied successfully in several countries in the 1970s to protect tomato plants against infection with tobacco mosaic virus. More recently, in 2015, the treatment PMV™-01 , which contains a mild isolate of the Chilean strain of pepino mosaic virus (PepMV), has been registered in Europe and commercially used to protect tomato plants from severe losses in quality and yield caused by aggressive infection with PepMV.

[0008] In order to develop cross protection, attenuated/mild strains of a plant virus are needed which cause no visible or only very mild symptoms but which prevent infection by strains causing more severe symptoms. Attenuated isolates

advantageously have little or no impact on plant symptoms, total yield and fruit quality, and effectively protect against more virulent isolates. The following properties have been proposed as desirable criteria for an attenuated virus strain for use in crop protection:

• no symptoms or very mild symptoms are induced in any of the cultivated hosts, and the quality and quantity of the crop products are not reduced; • most host tissues are fully infected systemically;

• the attenuated strains are highly stable genetically without mutating into a severe phenotype;

• there is no vector transmission to other crops;

• the attenuated strains protect against a wide range of viruses and strains; and

• the quality/quantity control of the inoculum and inoculations are easy and inexpensive.

[0009] An attenuated CGMMV strain SH33b is known (Motoyoshi, F., and

Nishiguchi, M. 1988. Control of virus diseases by attenuated virus strains:

comparison between attenuated strains of cucumber green mottle mosaic virus and tobacco mosaic virus. Gamma Field Symposia, 27: 91-107) which induces no or only mild systematic symptoms in leaves of muskmelon plants when a low concentration of attenuated virus was used to inoculate muskmelon seedlings. However, although this strain was effective in protecting muskmelon plants from outbreaks of severe symptoms, and in eliminating wild-type CGMMV from the greenhouse, inoculation of cotyledons with higher concentrations of the purified virus led to appearance of mosaic symptoms in the upper leaves.

[0010] In addition, an attenuated CGMMV strain VIROG-43M is known

(Slavokhotova, A.A., et al, American Journal of Plant Sciences (2016), 7: 724-732), which showed effectiveness in protecting inoculated cucumber plants from disease symptoms under greenhouse conditions. However, VIROG-43M itself could induce symptoms in inoculated cucumber plants after two months of inoculation. In addition, only 85% of the cucumber plants inoculated with VIROG-43M and challenged with the pathogenic CGMMV strain NC-1 were observed to be symptomless after 3 months of inoculation under laboratory greenhouse conditions.

[001 1] Recently, it was reported that several attenuated CGMMV strains were obtained from a pathogenic CGMMV strain by multi site-directed mutagenesis, based on sequence comparison of related tobamoviruses (Chen, Bin, "Molecular

Characterization of Viruses Infecting Greenhouse Vegetables in Ontario" (2016). Electronic Thesis and Dissertation Repository. 4222, University of Western Ontario. Available at https://ir.lib.uwo.ca/etd/4222). However, the reported mutants could develop visible mosaic symptoms in cucumber plants to various extents, although the symptoms were generally mild compared with those caused by the wild-type strain. Even the best attenuated CGMMV isolates induced symptoms in cucumber plants after 28 days post inoculation. [0012] It is therefore desirable to provide an attenuated strain of CGMMV for protecting plants against infection with CGMMV, and in particular, for protecting plants which are susceptible to infection with CGMMV such as species of the Cucurbitaceae family.

Summary

[0013] In one aspect, the present application provides an attenuated strain of cucumber green mottle mosaic virus (CGMMV) comprising a genome, wherein the genome is a polyribonucleotide having a sequence functionally equivalent to a variant of SEQ ID NO: 18, the sequence comprising at least one residue or group of residues selected from:

a) A at the position corresponding to position 4969 of SEQ ID NO: 18;

b) U at the position corresponding to position 3334 of SEQ ID NO: 18; and c) a group of at least six residues selected from:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

U at the position corresponding to position 1660 of SEQ ID NO: 18;

U at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

U at the position corresponding to position 4144 of SEQ ID NO: 18;

U at the position corresponding to position 4248 of SEQ ID NO: 18; and

U at the position corresponding to position 6228 of SEQ ID NO: 18.

[0014] In another aspect, the present application provides a polydeoxyribonucleotide having a sequence functionally equivalent to a sequence of a cucumber green mottle mosaic virus (CGMMV) genome, wherein the sequence of the

polydeoxyribonucleotide is a variant of SEQ ID NO: 18 comprising at least one residue or group of residues selected from:

a) A at the position corresponding to position 4969 of SEQ ID NO: 18;

b) T at the position corresponding to position 3334 of SEQ ID NO: 18; and c) a group of at least six residues selected from:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

T at the position corresponding to position 1660 of SEQ ID NO: 18;

T at the position corresponding to position 3430 of SEQ ID NO: 18; G at the position corresponding to position 3528 of SEQ ID NO: 18;

T at the position corresponding to position 4144 of SEQ ID NO: 18;

T at the position corresponding to position 4248 of SEQ ID NO: 18; and

T at the position corresponding to position 6228 of SEQ ID NO: 18.

[0015] In at least one embodiment, the polydeoxyribonucleotide is configured for expression in a host cell. In at least one embodiment, the host cell is a

microorganism. In at least one embodiment, the host cell is a plant cell.

[0016] In another aspect, the present application provides a vector comprising a polydeoxyribonucleotide as described herein. In at least one embodiment, the vector is configured to genetically modify a host cell. In at least one embodiment, the host cell is a microorganism. In at least one embodiment, the host cell is a plant cell.

[0017] A further aspect of the present application provides a genetically modified cell comprising a polydeoxyribonucleotide as described herein. In at least one embodiment, the cell is a microorganism. In at least one embodiment, the cell is a plant cell.

[0018] Yet another aspect of the present application provides a composition for preventing symptoms associated with infection by wild-type CGMMV in a plant, where the composition comprises an attenuated strain of CGMMV or a genetically modified cell as described herein and an agriculturally acceptable carrier.

[0019] In an additional aspect, the present application provides a composition for increasing resistance of a plant to infection by wild-type CGMMV, where the composition comprises an attenuated strain of CGMMV or a genetically modified cell as described herein and an agriculturally acceptable carrier.

[0020] Another aspect of the present application provides a method for preventing symptoms associated with infection by wild-type CGMMV in a plant, where the method includes inoculating the plant with an attenuated strain of CGMMV or with a genetically modified cell or with a composition as described herein.

[0021] A further aspect of the present application provides a method for increasing resistance of a plant to infection by wild-type CGMMV, where the method includes inoculating the plant with an attenuated strain of CGMMV or with a genetically modified cell or with a composition as described herein.

[0022] In a further aspect, the present application provides a genetically modified plant comprising a genome which comprises a polydeoxyribonucleotide as described herein. In at least one embodiment, the plant is a cucurbit. In at least one

embodiment, the plant is a cucumber plant. Brief Description of the Drawings

[0023] Further features of the present invention will become apparent from the following written description and the accompanying figures, in which:

[0024] Figure 1A is a photograph showing leaves of a cucumber plant grown for two weeks after inoculation with a clone of a wild-type cucumber green mottle mosaic virus (CGMMV) Ontario strain;

[0025] Figure 1 B is a photograph showing leaves of a cucumber plant grown for two weeks after inoculation with a wild-type CGMMV Ontario strain isolate;

[0026] Figure 1 C is a photograph showing leaves of a cucumber plant grown for two weeks in the absence of CGMMV (negative control);

[0027] Figure 2A is a photograph showing leaves of a cucumber plant infected with wild-type CGMMV Ontario strain and grown under laboratory conditions;

[0028] Figure 2B is a photograph showing leaves of a cucumber plant infected with the mutant CGMMV Ontario strain ONB and grown under the laboratory conditions of Figure 2A;

[0029] Figure 2C is photograph showing leaves of a cucumber plant infected with the mutant CGMMV Ontario strain ONM and grown under the laboratory conditions of Figure 2A;

[0030] Figure 2D is a photograph showing leaves of a healthy cucumber plant without CGMMV infection grown as a control under the laboratory conditions of Figure 2A;

[0031] Figure 3A is a photograph showing leaves of a cucumber plant infected with wild-type CGMMV Ontario strain and grown under laboratory conditions;

[0032] Figure 3B is a photograph showing leaves of a healthy cucumber plant without CGMMV infection grown as a control under the laboratory conditions of Figure 3A;

[0033] Figure 3C is a photograph showing leaves of a cucumber plant exposed to an attenuated CGMMV strain according to the present invention (ONBM) and grown under the laboratory conditions of Figure 3A;

[0034] Figure 3D is a photograph showing leaves of a cucumber plant exposed to another attenuated CGMMV strain according to the present invention (ONBM-2) and grown under the laboratory conditions of Figure 3A; [0035] Figure 3E is a photograph showing leaves of a cucumber plant exposed to yet another attenuated CGMMV strain according to the present invention (ONBM-3) and grown under the laboratory conditions of Figure 3A;

[0036] Figure 4A is a photograph showing leaves of a cucumber plant inoculated with an attenuated CGMMV strain according to the present invention (ONBM), then challenged with a wild-type CGMMV and grown under laboratory conditions;

[0037] Figure 4B is a photograph showing leaves of a cucumber plant inoculated with another attenuated CGMMV strain according to the present invention (ONBM-2), then challenged with a wild-type CGMMV and grown under the laboratory conditions of Figure 4A;

[0038] Figure 4C is a photograph showing leaves of a cucumber plant inoculated with yet another attenuated CGMMV strain according to the present invention (ONBM-3), then challenged with a wild-type CGMMV and grown under the laboratory conditions of Figure 4A;

[0039] Figure 4D is a photograph showing leaves of a cucumber plant infected with wild-type CGMMV and grown under the laboratory conditions of Figure 4A;

[0040] Figure 4E is a photograph showing leaves of a healthy cucumber plant without CGMMV infection grown as a control under the laboratory conditions of Figure 4A;

[0041] Figure 5A is a photograph showing leaves of a cucumber plant grown in a commercial greenhouse and infected with a wild-type CGMMV Ontario strain;

[0042] Figure 5B is a photograph showing a mosaic pattern on fruit of a cucumber plant grown in the commercial greenhouse of Figure 5A and infected with a wild-type CGMMV Ontario strain;

[0043] Figure 5C is a photograph showing curling of fruit of a cucumber plant grown in the commercial greenhouse of Figure 5A and infected with a wild-type CGMMV Ontario strain;

[0044] Figure 5D is a photograph showing leaves and fruit of a cucumber plant exposed to an attenuated CGMMV strain according to the present invention (ONBM- 2) and grown in the commercial greenhouse of Figure 5A;

[0045] Figure 5E is a photograph showing leaves and fruit of a cucumber plant exposed to another attenuated CGMMV strain according to the present invention (ONBM-3) and grown in the commercial greenhouse of Figure 5A; [0046] Figure 6A is a photograph showing leaves of a cucumber plant exposed to another attenuated CGMMV strain according to the present invention (ONAL-1) and grown under laboratory conditions;

[0047] Figure 6B is a photograph showing leaves of a cucumber plant exposed to another attenuated CGMMV strain according to the present invention (ONAL-2) and grown under the laboratory conditions of Figure 6A;

[0048] Figure 6C is a photograph showing leaves of a cucumber plant exposed to another attenuated CGMMV strain according to the present invention (ONBM-32) and grown under the laboratory conditions of Figure 6A;

[0049] Figure 6D is a photograph showing leaves of a cucumber plant exposed to the mutant CGMMV Ontario strain ONB and grown under the laboratory conditions of Figure 6A;

[0050] Figure 6E is a photograph showing leaves of a healthy cucumber plant without CGMMV infection grown as a control under the laboratory conditions of Figure 6A; and

[0051] Figure 6F is a photograph showing leaves of a cucumber plant infected with wild-type CGMMV Ontario strain and grown under the laboratory conditions of Figure 6A.

Detailed Description

[0052] The present application provides an attenuated strain of the cucumber green mottle mosaic virus (CGMMV). In at least one embodiment, the attenuated CGMMV strain may be useful for protecting one or more plants against the deleterious effects of infection by a wild-type strain of CGMMV. In at least one embodiment, plants which may be protected against infection by wild-type CGMMV include but are not limited to any plant susceptible to infection by CGMMV, including but not limited to plants of the family Cucurbitaceae (cucurbits), including but not limited to varieties of cucumber (Cucumis sativus), pumpkin, watermelon, melon, squash, zucchini, gourds, gherkins and others well known in the art. In at least one embodiment, curcubits such as cucumber plants inoculated with at least one embodiment of the attenuated CGMMV strain may not show significant visible symptoms for a period of up to two months or longer after inoculation with the attenuated CGMMV strain.

[0053] In at least one embodiment, the attenuated strain of CGMMV has a genome which is a polyribonucleotide having a sequence which is functionally equivalent to a variant of SEQ ID NO: 18 including one or more variations thereof. [0054] In at least one embodiment, the polyribonucleotide has a sequence including A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0055] In at least one embodiment, the polyribonucleotide has a sequence including U at the position corresponding to position 3334 of SEQ ID NO: 18.

[0056] In at least one embodiment, the polyribonucleotide has a sequence including A at the position corresponding to position 4969 of SEQ ID NO: 18 and U at the position corresponding to position 3334 of SEQ ID NO: 18.

[0057] In at least one embodiment, the polyribonucleotide has a sequence including at least six of the following residues:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

U at the position corresponding to position 1660 of SEQ ID NO: 18;

U at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

U at the position corresponding to position 4144 of SEQ ID NO: 18;

U at the position corresponding to position 4248 of SEQ ID NO: 18; and

U at the position corresponding to position 6228 of SEQ ID NO: 18.

[0058] In at least one such embodiment, the polyribonucleotide sequence also includes one or both of:

U at the position corresponding to position 3334 of SEQ ID NO: 18; and A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0059] In at least one embodiment, the polyribonucleotide has a sequence including the following residues:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

U at the position corresponding to position 1660 of SEQ ID NO: 18;

U at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

U at the position corresponding to position 4144 of SEQ ID NO: 18;

U at the position corresponding to position 4248 of SEQ ID NO: 18; and

U at the position corresponding to position 6228 of SEQ ID NO: 18.

[0060] In at least one such embodiment, the polyribonucleotide sequence also includes one or both of: U at the position corresponding to position 3334 of SEQ ID NO: 18; and A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0061] As used herein, the term“polynucleotide” is intended to mean a polymeric molecule comprising two or more nucleosides linked through covalent bonds to phosphate groups, such that the 5’-hydroxyl group of a nucleoside and the

3’-hydroxyl group of an adjacent nucleoside are both covalently bonded to the same phosphate group. As understood in the art, when covalently linked together to form the polynucleotide molecule, each individual nucleoside unit is also known as a “residue”. As used herein, the term“nucleoside” is intended to mean a molecule in which a sugar moiety selected from ribose and deoxyribose is bonded to a purine or pyrimidine base moiety selected from adenine (A), guanine (G), cytosine (C), thymine (T) or uracil (U). A polynucleotide includes polynucleotides of any length, including but not limited to dinucleotides, trinucleotides, tetranucleotides, oligonucleotides and nucleic acids.

[0062] When the sugar moiety is ribose, the base is selected from A, G, C and U, and the nucleoside is referred to as a ribonucleoside. A polynucleotide comprising two or more such ribonucleosides is referred to as a polyribonucleotide or ribonucleic acid (RNA). When the sugar moiety is deoxyribose, the base is selected from A, G, C and T, and the nucleoside is referred to as a deoxyribonucleoside. A polynucleotide comprising two or more such deoxyribonucleosides is referred to as a

polydeoxyribonucleotide or deoxyribonucleic acid (DNA).

[0063] In at least one embodiment, the attenuated strains can be obtained by mutation of wild type CGMMV to introduce one or more variations or mutations into the genome sequence of the wild type CGMMV. As used herein interchangeably with respect to a polynucleotide sequence, the term“variation” or“mutation” is intended to refer to a difference in the polynucleotide sequence with respect to a reference polynucleotide sequence. Variations or mutations can include substitution of one or more nucleotide residues with different nucleotide residues, insertion of additional nucleotide residues or deletion of nucleotide residues. A variation or mutation may or may not alter the open reading frame(s) of the polynucleotide or the amino acid sequence of any protein(s) encoded by the polynucleotide. In at least one embodiment, the variation or mutation is a naturally occurring variation or mutation arising without artificial intervention. In at least one embodiment, the variation or mutation is introduced intentionally by methods well known in the art, including but not limited to random mutagenesis or directed mutagenesis. [0064] An RNA virus such as CGMMV, including but not limited to attenuated strains thereof, contains an RNA genome, in which the genetic information of the virus is stored in the form of RNA. As will be understood by a person of skill in the art, it is possible to express viral proteins encoded by such an RNA genome in a host cell which expresses genetic information from DNA by preparing a complementary DNA (cDNA) molecule carrying the same genetic information as is encoded by the RNA genome, and transforming the host cell with the cDNA such that the transformed host cell can express viral proteins encoded by the cDNA. In at least one embodiment, the virus can be assembled and/or replicated within the transformed host cell.

[0065] Therefore, another aspect of the present invention provides a

polydeoxyribonucleotide having a sequence functionally equivalent to a sequence of a cucumber green mottle mosaic virus (CGMMV) genome, wherein the

polydeoxyribonucleotide sequence comprises a variant of SEQ ID NO: 18 including one or more variations from SEQ ID NO:18.

[0066] In at least one embodiment, the polydeoxyribonucleotide sequence comprises a sequence wherein the one or more variations from SEQ ID NO:18 include A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0067] In at least one embodiment, the polydeoxyribonucleotide sequence comprises a sequence wherein the one or more variations from SEQ ID NO:18 include T at the position corresponding to position 3334 of SEQ ID NO: 18.

[0068] In at least one embodiment, the polydeoxyribonucleotide sequence comprises a sequence wherein the one or more variations from SEQ ID NO:18 include A at the position corresponding to position 4969 of SEQ ID NO: 18 and T at the position corresponding to position 3334 of SEQ ID NO: 18.

[0069] In at least one embodiment, the polydeoxyribonucleotide sequence comprises a sequence wherein the one or more variations from SEQ ID NO:18 include at least six of the following residues:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

T at the position corresponding to position 1660 of SEQ ID NO: 18;

T at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

T at the position corresponding to position 4144 of SEQ ID NO: 18;

T at the position corresponding to position 4248 of SEQ ID NO: 18; and

T at the position corresponding to position 6228 of SEQ ID NO: 18. [0070] In at least one such embodiment, the one or more variations from SEQ ID NO: 18 further include one or both of:

T at the position corresponding to position 3334 of SEQ ID NO: 18; and A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0071] In at least one embodiment, the polydeoxyribonucleotide sequence comprises a sequence wherein the one or more variations from SEQ ID NO:18 include the following residues:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

T at the position corresponding to position 1660 of SEQ ID NO: 18;

T at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

T at the position corresponding to position 4144 of SEQ ID NO: 18;

T at the position corresponding to position 4248 of SEQ ID NO: 18; and

T at the position corresponding to position 6228 of SEQ ID NO: 18.

[0072] In at least one such embodiment, the one or more variations from SEQ ID NO: 18 further include one or both of:

T at the position corresponding to position 3334 of SEQ ID NO: 18; and A at the position corresponding to position 4969 of SEQ ID NO: 18.

[0073] As used herein with reference to a polynucleotide sequence, the term “functionally equivalent to” is intended to mean that the polynucleotide sequence contains the same genetic information, including but not limited to coding information, as the genetic information contained in the reference polynucleotide sequence. In at least one embodiment, a functionally equivalent polynucleotide sequence will encode the same protein or proteins as are encoded by the reference polynucleotide sequence. In at least one embodiment, a given position in the polynucleotide sequence will bear a base equivalent to the base borne by the corresponding position of the reference polynucleotide sequence. As used herein with reference to bases in functionally equivalent polynucleotide sequences, the term“equivalent” is intended to mean that the bases are either identical or provide the same coding information. Thus, the base T (which is found in polydeoxyribonucleotides) and the base U (which is found in polyribonucleotides) are considered herein to be equivalent to each other. In other words, when a polydeoxyribonucleotide sequence is functionally equivalent to a polyribonucleotide sequence, positions in the polydeoxyribonucleotide sequence which bear the base T are considered to correspond to positions in the polyribonucleotide sequence which bear the base U.

[0074] In at least one embodiment, the sequence of the polydeoxyribonucleotide is selected from SEQ ID NO:42, SEQ ID NO:45, SEQ ID NO:48, SEQ ID NO:53, SEQ ID NO:58, SEQ ID NO:60, and variants thereof which include at least one residue or group of residues selected from:

a) A at the position corresponding to position 4969 of SEQ ID NO: 18;

b) T at the position corresponding to position 3334 of SEQ ID NO: 18; and c) a group of at least six residues selected from:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

T at the position corresponding to position 1660 of SEQ ID NO: 18;

T at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

T at the position corresponding to position 4144 of SEQ ID NO: 18;

T at the position corresponding to position 4248 of SEQ ID NO: 18; and

T at the position corresponding to position 6228 of SEQ ID NO: 18.

[0075] In at least one embodiment, the sequence of the variant can have at least 89%, at least 90%, at least 95%, at least 99% or at least 99.9% identity to a sequence selected from SEQ ID NO:42, SEQ ID NO:45, SEQ ID NO:48, SEQ ID NO:53, SEQ ID NO:58 or SEQ ID NO:60, wherein the variant sequence comprises at least one residue or group of residues selected from:

a) A at the position corresponding to position 4969 of SEQ ID NO: 18;

b) T at the position corresponding to position 3334 of SEQ ID NO: 18; and c) a group of at least six residues selected from:

A at the position corresponding to position 315 of SEQ ID NO: 18;

G at the position corresponding to position 1498 of SEQ ID NO: 18;

T at the position corresponding to position 1660 of SEQ ID NO: 18;

T at the position corresponding to position 3430 of SEQ ID NO: 18;

G at the position corresponding to position 3528 of SEQ ID NO: 18;

T at the position corresponding to position 4144 of SEQ ID NO: 18;

T at the position corresponding to position 4248 of SEQ ID NO: 18; and

T at the position corresponding to position 6228 of SEQ ID NO: 18. [0076] In at least one embodiment, the sequence of the polydeoxyribonucleotide is selected from SEQ ID NO:42, SEQ ID NO:45, SEQ ID NO:48, SEQ ID NO:53, SEQ ID NO:58, SEQ ID NO:60, and variants thereof which encode one or more of the proteins encoded by the polydeoxyribonucleotide having the sequence selected from SEQ ID NO:42, SEQ ID NO:45, SEQ ID NO:48, SEQ ID NO:53, SEQ ID NO:58 and SEQ ID NO:60.

[0077] As used herein, the term“variant” when used in reference to a polynucleotide is intended to refer to a polynucleotide which differs in its nucleotide sequence from the sequence of a reference polynucleotide to which the variant is being compared by one or more nucleotide residues. The differences between the sequence of the variant and the sequence of the reference polynucleotide, also referred to herein as variations or mutations, can include substitution of one or more nucleotide residues with different nucleotide residues, insertion of additional nucleotide residues or deletion of nucleotide residues. In certain embodiments, a variant can differ from a reference polynucleotide by substitution of one or more nucleotide residues with replacement nucleotide residues which do not alter the open reading frame(s) of the polynucleotide or the amino acid sequence of any protein(s) encoded by the polynucleotide.

[0078] As used herein, the term“variant” when used in reference to a polypeptide is intended to refer to a polypeptide which differs in its amino acid sequence from the sequence of a reference polypeptide to which the variant is being compared by one or more amino acid residues. The differences between the sequence of the variant and the sequence of the reference polypeptide can include substitution of one or more amino acid residues with different amino acid residues, insertion of additional amino acid residues or deletion of amino acid residues. In certain embodiments, a variant can differ from a reference polypeptide by conservative substitution of one or more amino acid residues with replacement amino acid residues which may have similar properties, including but not limited to charge, size and hydrophilicity, to the amino acid residues which the new residues replace. In certain embodiments, variants may completely or partially retain one or more biological functions of the reference polypeptide. In certain embodiments, variants may not retain one or more biological functions of the reference polypeptide.

[0079] As used herein, the term“percent identity” or“% identity” when used in reference to the sequence of a polypeptide or a polynucleotide is intended to mean the percentage of the total number of amino acid or nucleotide residues, respectively, in the sequence which are identical to those at the corresponding position of a reference polypeptide or polynucleotide sequence. In at least one embodiment, when the length of the variant sequence and the length of the reference sequence are not identical, percent identity can be calculated based on the total number of residues in the variant sequence or based on the total number or residues in the reference sequence. Percent identity can be measured by various local or global sequence alignment algorithms well known in the art, including but not limited to the Smith- Waterman algorithm and the Needleman-Wunsch algorithm. Tools using these or other suitable algorithms include but are not limited to BLAST (Basic Local Alignment Search Tool) and other such tools well known in the art.

[0080] In at least one embodiment, a variant polynucleotide sequence can hybridize to a polyribonucleotide or polydeoxyribonucleotide as described herein under at least moderately stringent conditions. By "at least moderately stringent hybridization conditions" it is meant that conditions are selected which promote selective hybridization between two complementary nucleic acid molecules in solution.

Hybridization may occur to all or a portion of a nucleic acid sequence molecule. The hybridizing portion is typically at least 15 (e.g. 20, 25, 30, 40 or 50) nucleotides in length. Those skilled in the art will recognize that the stability of a nucleic acid duplex, or hybrid, is determined by the melting temperature (T _m ), which in sodium-containing buffers is a function of the sodium ion concentration ([Na ⁺ ]) and temperature (T _m = 81.5°C - 16.6 (Log-io [Na ⁺ ]) + 0.41 (%(G+C) - 600/I), where %G+C is the percentage of cytosine and guanine nucleotides in the nucleic acid and I is the length of the nucleic acid in base pairs, or similar equation). Accordingly, the parameters in the wash conditions that determine hybrid stability are sodium ion concentration and temperature. In order to identify molecules that are similar, but not identical, to a known nucleic acid molecule, a 1 % mismatch may be assumed to result in about a 1 °C decrease in T _m . For example, if nucleic acid molecules are sought that have a >95% identity, the final wash temperature may be reduced by about 5°C. Based on these considerations those skilled in the art will be able to readily select appropriate hybridization conditions.

[0081] In some embodiments, stringent hybridization conditions are selected. By way of example the following conditions may be employed to achieve stringent hybridization: hybridization at 5x sodium chloride/sodium citrate (SSC)/5x Denhardt's solution/1.0% sodium dodecylsulfate (SDS) at T _m - 5°C based on the above equation, followed by a wash of 0.2x SSC/0.1 % SDS at 60°C. Moderately stringent

hybridization conditions include a washing step in 3x SSC at 42°C. It is understood, however, that equivalent stringencies may be achieved using alternative buffers, salts and temperatures. Additional guidance regarding hybridization conditions may be found in: Current Protocols in Molecular Biology, John Wiley & Sons, N.Y., 2002, and in: Sambrook et al., Molecular Cloning: a Laboratory Manual, Cold Spring Harbor Laboratory Press, 2001.

[0082] In at least one embodiment, the polyribonucleotide CGMMV genome and the functionally equivalent polydeoxyribonucleotide can each encode one or more viral proteins including but not limited to a 129 kDa protein including methyltransferase and helicase domains required for RNA replication, a 186 kDa protein including the methyltransferase and helicase domains and an additional RNA-dependent RNA polymerase (RdRp) domain, a movement protein (MP) and a coat protein (CP).

[0083] In at least one embodiment, the 129 kDa protein has an amino acid sequence comprising valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0084] In at least one embodiment, the 129 kDa protein sequence includes at least two residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63.

[0085] In at least one such embodiment, the 129 kDa protein sequence further includes valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0086] In at least one embodiment, the 129 kDa protein sequence includes at least three residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63. [0087] In at least one such embodiment, the 129 kDa protein sequence further includes valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0088] In at least one embodiment, the 129 kDa protein sequence includes the residues:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63.

[0089] In at least one such embodiment, the 129 kDa protein sequence further includes valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0090] In at least one embodiment, the sequence of the 129 kDa protein is selected from SEQ ID NO:43, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:54, SEQ ID NO:61 and variants thereof which include valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0091] In at least one embodiment, the sequence of the 129 kDa protein is selected from SEQ ID NO:43, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:54, SEQ ID NO:61 and variants thereof which include at least two residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63.

[0092] In at least one such embodiment, the variants of the 129 kDa protein sequence further include valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63. [0093] In at least one embodiment, the sequence of the 129 kDa protein is selected from SEQ ID NO:43, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:54, SEQ ID NO:61 and variants thereof which include at least three residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63.

[0094] In at least one such embodiment, the variants of the 129 kDa protein sequence further include valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0095] In at least one embodiment, the sequence of the 129 kDa protein is selected from SEQ ID NO:43, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:54, SEQ ID NO:61 and variants thereof which include the residues:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:63; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:63;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:63; and

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:63.

[0096] In at least one such embodiment, the variants of the 129 kDa protein sequence further include valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:63.

[0097] In at least one embodiment, the 186 kDa protein has an amino acid sequence comprising histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[0098] In at least one embodiment, the 186 kDa protein sequence includes valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64.

[0099] In at least one embodiment, the 186 kDa protein sequence includes histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64 and valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64. [0100] In at least one embodiment, the 186 kDa protein sequence includes at least five residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and

serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[0101] In at least one such embodiment, the 186 kDa protein sequence further includes one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[0102] In at least one embodiment, the 186 kDa protein sequence includes at least six residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[0103] In at least one such embodiment, the 186 kDa protein sequence further includes one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[0104] In at least one embodiment, the 186 kDa protein sequence includes the residues:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and

serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[0105] In at least one such embodiment, the 186 kDa protein sequence further includes one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[0106] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64. [0107] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64.

[0108] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64 and valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64.

[0109] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include at least five residues selected from:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and

serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[01 10] In at least one embodiment, the variants of the 186 kDa protein sequence further include one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[01 1 1] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include at least six residues selected from: serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and

serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[01 12] In at least one embodiment, the variants of the 186 kDa protein sequence further include one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[01 13] In at least one embodiment, the sequence of the 186 kDa protein is selected from SEQ ID NO:44, SEQ ID NO:47, SEQ ID NO:50, SEQ ID NO:55, SEQ ID NO:59, SEQ ID NO:62 and variants thereof which include the residues:

serine (S, Ser) at the position corresponding to position 86 of SEQ ID NO:64; glycine (G, Gly) at the position corresponding to position 480 of SEQ ID NO:64;

phenylalanine (F, Phe) at the position corresponding to position 534 of SEQ ID NO:64;

valine (V, Val) at the position corresponding to position 1 124 of SEQ ID NO:64;

aspartic acid (D, Asp) at the position corresponding to position 1 157 of SEQ ID NO:64;

leucine (L, Leu) at the position corresponding to position 1362 of SEQ ID NO:64; and serine (S, Ser) at the position corresponding to position 1397 of SEQ ID NO:64.

[01 14] In at least one embodiment, the variants of the 186 kDa protein sequence further include one or both of:

valine (V, Val) at the position corresponding to position 1092 of SEQ ID NO:64; and

histidine (H, His) at the position corresponding to position 1637 of SEQ ID NO:64.

[01 15] Because the 186 kDa protein is encoded by an open reading frame in the CGMMV genome which includes the open reading frame for the 129 kDa protein, it will be clear to the person of skill in the art that a polyribonucleotide or a

polydeoxyribonucleotide as described herein which encodes a 186 kDa protein as described herein will also encode a 129 kDa protein as described herein.

[01 16] In at least one embodiment, the coat protein has an amino acid sequence comprising valine (V, Val) at the position corresponding to position 156 of SEQ ID NO:65. In at least one embodiment, the sequence of the coat protein is selected from SEQ ID NO:32 and variants thereof comprising valine (V, Val) at the position corresponding to position 156 of SEQ ID NO:65.

[01 17] As will be understood in the art, the degeneracy of the genetic code allows for some amino acids to be encoded by more than one codon or group of three nucleoside residues. Therefore, it is contemplated that the sequences of the present polyribonucleotide or polydeoxyribonucleotide can also include mutations other than those specifically described herein such that the polyribonucleotide or

polydeoxyribonucleotide will encode proteins having amino acid sequences as described herein.

[01 18] In at least one embodiment, the polydeoxyribonucleotide is configured for expression in a host cell so as to permit expression of viral proteins and/or assembly and/or replication of infectious virus in the host cell, as will be understood by those skilled in the art, who will be capable of configuring the polydeoxyribonucleotide for expression in such a host cell without undue experimentation in light of the teaching herein. In at least one embodiment, the host cell is a microorganism. In at least one embodiment, the host cell is a plant cell.

[01 19] In another aspect, the present application provides a vector comprising a polydeoxyribonucleotide as described herein. In at least one embodiment, the vector is configured for use to genetically modify a cell. Thus, a further aspect of the present application provides a genetically modified cell comprising a polydeoxyribonucleotide as described herein. In at least one embodiment, the cell is a microorganism. In at least one embodiment, the cell is a plant cell. Those skilled in the art would be aware of methods for preparing such vectors and using them to genetically modify such cells.

[0120] Another aspect of the present application provides a composition for preventing symptoms associated with infection by wild-type CGMMV in a plant, where the composition comprises an attenuated strain of CGMMV or a genetically modified cell as described herein and an agriculturally acceptable carrier. In at least one embodiment, the composition comprises two or more attenuated strains of CGMMV or genetically modified cells as described herein.

[0121] As used herein, the term "carrier" is intended to refer to a diluent, adjuvant, excipient, or vehicle with which an attenuated strain of CGMMV or a genetically modified cell can be applied or administered to a plant or crop. As used herein, the term "agriculturally acceptable" is intended to refer to carriers and compositions containing such carriers that are tolerable and do not typically produce untoward reactions to a plant or crop being treated with such carriers and compositions, or to a worker applying such carriers and compositions to a plant or crop under normal agricultural conditions. Preferably, as used herein, the term "agriculturally acceptable" means approved by a regulatory agency of the federal or a state government for use in agricultural applications. Such agriculturally acceptable carriers are well known in the art.

[0122] In an additional aspect, the present application provides a composition for increasing resistance of a plant to infection by wild-type CGMMV, where the composition comprises an attenuated strain of CGMMV or a genetically modified cell as described herein and an agriculturally acceptable carrier. In at least one embodiment, the composition comprises two or more attenuated strains of CGMMV or genetically modified cells as described herein.

[0123] Another aspect of the present application provides a method for preventing symptoms associated with infection by wild-type CGMMV in a plant, where the method includes inoculating the plant with an attenuated strain of CGMMV or with a genetically modified cell as described herein. Methods of inoculating plants with viruses, including but not limited to attenuated strains thereof, and/or with cells, including but not limited to microorganisms, genetically modified to express such viruses or associated viral proteins, and/or with compositions thereof are well known in the art, and well within the capability of the skilled person in light of the teaching of the present application.

[0124] A further aspect of the present application provides a method for increasing resistance of a plant to infection by wild-type CGMMV, where the method includes inoculating the plant with an attenuated strain of CGMMV or with a genetically modified cell as described herein.

[0125] In a further aspect, the present application provides a genetically modified plant comprising a genome which comprises a polydeoxyribonucleotide as described herein. It is contemplated that such a genetically modified plant may have increased resistance to infection by wild type strains of CGMMV. In at least one embodiment, the plant is a cucurbit. In at least one embodiment, the plant is a cucumber plant.

[0126] As used herein, the terms“about” or“approximately” as applied to a numerical value or range of values are intended to mean that the recited values can vary within an acceptable degree of error for the quantity measured given the nature or precision of the measurements, such that the variation is considered in the art as equivalent to the recited values and provides the same function or result. For example, the degree of error can be indicated by the number of significant figures provided for the measurement, as is understood in the art, and includes but is not limited to a variation of ±1 in the most precise significant figure reported for the measurement. Typical exemplary degrees of error are within 20 percent (%), preferably within 10%, and more preferably within 5% of a given value or range of values. Alternatively, and particularly in biological systems, the terms "about" and "approximately" can mean values that are within an order of magnitude, preferably within 5-fold and more preferably within 2-fold of a given value. Numerical quantities given herein are approximate unless stated otherwise, meaning that the term "about" or "approximately" can be inferred when not expressly stated.

[0127] As used herein, the term“substantially” refers to the complete or nearly complete extent or degree of an action, characteristic, property, state, structure, item, or result. For example, an object that is“substantially” in a given position including but not limited to vertical, horizontal, or adjacent to or aligned with another object, would mean that the object is either completely in that position or nearly completely in that position. The exact allowable degree of deviation from absolute completeness may in some cases depend on the specific context. However, generally speaking, the nearness of completion will be so as to have the same overall result as if absolute and total completion were obtained. [0128] The use of“substantially” is equally applicable when used in a negative connotation to refer to the complete or near complete lack of an action, characteristic, property, state, structure, item, or result. For example, a composition that is “substantially free of an ingredient or element would either completely lack that ingredient or element, or so nearly completely lack that ingredient or element that the effect would be the same as if it completely lacked that ingredient or element. In other words, a composition that is“substantially free of an ingredient or element may still actually contain such item as long as there is no measurable or significant effect thereof.

EXAMPLES

[0129] Other features of the present invention will become apparent from the following non-limiting examples which illustrate, by way of example, the principles of the invention.

Example 1 : Cucumber green mottle mosaic virus (CGMMV) Ontario strain

Isolation, cloning and sequencing of a wild-type Ontario strain

[0130] Cucumber green mottle mosaic virus (CGMMV) Ontario strain (also referred to herein as the“wild-type” strain) was extracted from cucumber plants showing green mottle and mosaic symptoms collected from a commercial greenhouse in Ontario. cDNA was synthesized from the CGMMV RNA genome using Agilent AccuScript™ High-Fidelity Reverse Transcriptase (Agilent) and an oligonucleotide primer having the sequence

CGGCTCGAGCCCGTTTCGTCCTTTAGGGACTCGTCAGTGTACTGATATAAGTAC AGACTGGGCCCCTACCCGGGGAAAGGGGGGATT (SEQ ID NO: 1). The full-length genome of CGMMV Ontario strain was amplified from cDNA by polymerase chain reaction (PCR) using Q5™ High-fidelity 2X Master Mix (New England Biolabs) and the primers G TTTT AATTTTT AAAATT AAAC AAAC AAC AAC AAC AAC AAC AAAC (SEQ ID NO:2) and CCCCGGCTCGAGCCCGTTTCGTCCTTTAGGGACTCGT (SEQ ID NO:3). The PCR product was digested with Xho\ and cloned into a binary vector pKW8 between the Stu\ and Xho\ sites in 10-beta Escherichia coli (New England Biolabs) by electroporation. The whole cDNA genome was sequenced using primers which were designed based on analysis of genome sequences published in

GenBank for other CGMMV strains. The sequences of the primers are listed in Table 1 below. Table 1 : Primers used to sequence the cDNA genome of CGMMV wild-type Ontario strain

[0131 ] The full-length cDNA genome sequence of CGMMV Ontario strain is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAAT GG CAAACATT AAT G AACAAAT CAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACT G ATGCGTAT CCGG AGTTTT CG A TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT GCGGGTGGTCT GAG G C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATGGTCACACAACATGTACAGAGGCACAAGGGATCTGGCGGGTGCAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAGG AGGTACG AT AATT CTCCCTGTG CG GT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTT GTTATG CAG CCTTT CACTT CTCG G AG G CATT G CTTTT AG GTT CACCTGTAG GT AATTT AAAT AGTATT G G GG CTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATT AATTGT G ATGCGT ACTT ATTTTCCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AGGTTGGT CAG AG CAGACACACAT AT GCTT CAT AAAT CTGTGGGG CA CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAG CCACGTTTT CT GTGTGGTTT CCT G AG GCG AAG CGT AAG GT GTTG ATA CCCAAGTTT G AACTTT CAAG ATT CCTTT CT G GG AAT GT G AAAAT CT CT AGG AT G C TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGTAT GAT AAT AAG G CCTT AGTGT G G AAG AAT GTT CAGTCCTTT GTG G AAT CT AT ACG CTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT T CG CAG ACAGT CTT AAAT CCGCCGTAG AAG G ACT AGGT GATT GCGTCTAT GAT G CTCTAGTTCAAACCGG CTG GTTTG ATACCTCTAG CG ACG AACTG AAAGTTTTG C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAGG G AT GT ACG AGG CAG ATGCAAAG AT CG AG AG AG AG AGTGTCT CT G AGTTGCT CGCTT CCGGT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GATACATGTGAAGATATGGATGTAACTGAAGATATGGAGGATATAGTGTTGATG GCGGACAAGAGTCATTCTTACATGTCCCCAGAAATGGCGAGATGGGCTGATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTTACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT GAG GTT AT CAAACAT G AATTT CTTT AAGGT G AG CG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACC ATT CG AG AG AGG CAAAT G AAG AATT AT ATT GATT ACTT ATCG GCCTCT CTTGGTT CTACG CTG GGT AAT CT G GAG AG AATT G TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTG GTT CT CCCAT AATT GAT AAG AG AAACT G G AAG CG ATTT GCT GTTT GC T CT G AG ACCAAAGT CT AT AG CGTAATT CGT AGTTT AG AGGT ACT AAAT AAG G AAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG GAAAG ACCG CCG AAATT AT AG CG AG GGTC AATT G G AAAACCG AT CT AGT ATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGTCACCTGTGGCAACCAGTGACAACGTTAGAACTTTCGATTCTTTTGTGATG AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGTG ATG CT AAG CAAAT CCCGTTT AT AAACAG AGT CAT G AATTTT GATT AT CCTAAGGAGTTAAGAACTTTAATAGTCGATAATGTAGAGCGTAGGTATGTTACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG GAAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CG ACT CCG AT AG GTTT AATT GCCCGT GATT CACCACAT GT ACT AGT GG CCTT AA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTATAATAGCGGATGTGGAAAAGGTCGACCAGTCGATCTTGACTATGTTTGCTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT AA T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G CAG G AGTT CT AT GAT AG AT G CCTT CCT GG G AATT CCTT CGTG CT G AAT G ATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT GGTT AAG AGTG AAG CG CAG AATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCT GAT CT AG CT G GG ACTGTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AGG G CT AGTT CCATT CAAAGTTT TT CT G ATTGGTTTT CGTGT CAG CCAACCT CGGCGGTTGGT CAATT AG CT AATTT C AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G CACAT GATT AAGCG G CAG C CCAAG AGT CGGTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TT GTTT AT CACCCT AAAGT GGT AAAT G CAGTTTT CG GT CCG GTTTTT AAGT ATTT G ACCACCAAGTTT CTT AG CAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AG G AAAAAACCAG AAG AT CT G CAG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT GAG ATT CTT GAG CTG GAT GTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCA TTTCTCTATTGAGATGGCAATTTGGGAAAAATTGGGGCTGGACGATATTTTGGCT TGGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGG AT AAAG ACG CT CATTT ACT AT CAACG G AAGTCT GGT GAT GT AACT ACTTT CAT AG GT AAT ACCTTT ATT AT CG CAG CGTGT GT AGCT AGT ATGTTG CCGTT AG ACAAGT G TTTT AAAG CT AGTTTTTGTG GT GAT GATT CG CT GAT CT ACCTT CCT AAG GGTTT G G AGTATCCTG ATATACAG GCTACTG CCAACTTG GTTTG G AATTTTG AG GCG AAA CTTTT CCG AAAG AAGTAT G GTT ACTT CT GTGG G AAGT AT AT AATT CACCAT G CCA ACGG CTGTATT GTTT ACCCT G ACCCTTT AAAATT AATT AGT AAATT AG GT AAT AAG AGT CTTGTAG GGT AT GAG CAT GTT G AGG AGTTT CGTATATCTCTCCTCG ACGTC G CT CAT AGTTT GTTT AAT GGTG CTT ATTT CCATTT ACT CG ACG ATG CAAT CCACG AATT ATTT CCT AACG CT G GG G GTT GCAGTTTTGTAATT AATT GTTT GTG CAAGT A TTT G AGTG AT AAG CG CCTTTT CCGT AGT CTTT AT AT AG AT GTCTCT AAGT AAG GT GTCGGTCGAGAACTCATTGAAACCCGAGAAGTTTGTTAAAATCTCTTGGGTCGA T AAGTT G CTCCCT AACT ATTTTT CCATT CTT AAGT ATTT AT CT AT AACT G ACTTT AG CGTAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGG TGTTGATCTTACAAAACACCTTTATGTCACATTGTTGGGCGTTGTGGTTTCTGGT GTATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGA CACAAG G AT GC ATT CTGTTG CAG AGG G AACT AT AT G CAAATTTT C AGCT CCCG C CACCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGC GGATGCCCTTCGCGATCCTTGGTCTTTATTTGTGAGACTCTCTAATGTAGGGATT AAAG ATGGTTT CCAT CCTTT GACCTT AGAGGT CGCTTGTTT AGT CGCTACAACTA ACT CT ATT AT CAAAAAGGGTCTTAG AGCTT CTGTAGTCG AGT CT GT CGT CT CTT C CG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAGTT G AACCTTT CTTT GAT AAAGTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGG TCGCAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAG GTT G G ACT CT G CTT CT G AAG AGT CCAGTT CTGTTT CTTT CG AAG AT G G CTTACAA TCCGATCACACCTAGCAAACTTATTGCGTTTAGTGCTTCTTATGTTCCCGTCAGG ACTTT ACTT AATTTT CT AGTT G CTT CACAAGGT ACCG CCTT CC AG ACT CAAG CG G GAAGAGATTCTTTCCGCGAGTCCCTGTCTGCGTTACCCTCGTCTGTCGTAGATA TTAATTCTAGGTTCCCAAATG CG G GTTTTTACG CTTTCCTCAACG GTCCTGTGTT GAGGCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCAT TGAGGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAA GCGTACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGA GTCTATTTCTAAGGGTTTTGATGTTTATGATAGGGCTTCATTTGAAGCCGCGTTT TCGGTAGTCTGGTCAGAGGCTACCACCTCGAAAGCTTAGCTTCGAGGGTCTTCT GATGGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACG GTTTGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGG GCAGTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA

(SEQ ID NO: 18).

[0132] The 129 kDa protein encoded by the wild type CGMMV Ontario strain has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTK (SEQ ID NO:63).

[0133] The 186 kDa protein encoded by the wild type CGMMV Ontario strain has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRNIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHPKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKPEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:64).

[0134] The coat protein encoded by the wild type CGMMV Ontario strain has the following sequence.

MAYNPITPSKLIAFSASYVPVRTLLNFLVASQGTAFQTQAGRDSFRESLSALPSSW DINSRFPNAGFYAFLNGPVLRPIFVSLLSSTDTRNRVIEWDPSNPTTAESLNAVKRT DDASTAARAEIDNLIESISKGFDVYDRASFEAAFSWWSEATTSKA (SEQ ID NO:65).

Infection of cucumber plants with the wild-type Ontario strain

[0135] The CGMMV Ontario strain clone was transformed into Agrobacterium tumefaciens strain EHA105 by electroporation. The Agrobacterium transformants were selected on LB medium plates containing 50 pg/ml of kanamycin and 20 pg/ml of rifampicin. After confirmation by colony PCR, the Agrobacterium transformants carrying CGMMV Ontario strain were cultured overnight at 30°C with shaking at 200 rpm in LB medium (lysogeny broth, also known as Luria-Bertani medium) containing 50 pg/ml of kanamycin and 20 pg/ml of rifampicin. The overnight culture was used to inoculate fresh LB medium containing 50 pg/ml of kanamycin, 20 pg/ml of rifampicin, 10 mM MES (2-(N-morpholino)ethanesulfonic acid), and 200 pM acetosyringone (3’,5’-dimethoxy-4’-hydroxyacetophenone) and the culture was incubated with shaking at 30°C until the optical density at 600 nm (OD ₆ oo) reached between 0.5 and 1.0.

[0136] Bacterial cells were harvested by centrifugation at 4000g for 10 minutes and resuspended in the same volume of Agrobacterium induction buffer (10 mM MES, 200 pM acetosyringone). The suspended cells were incubated at room temperature with gentle shaking (50 rpm) for 3-4 hours. The Agrobacterium cells were inoculated into the cotyledon of 1-2 week-old cucumber plants by leaf infiltration using a 1 ml needleless syringe. After incubation for a further two weeks under laboratory greenhouse conditions (16 h daylight at 22°C, 8 h darkness at 20°C), cucumber plant leaves were sampled and tested for the presence of CGMMV by ELISA using a commercial ELISA kit for detecting CGMMV (Agdia) and following the manufacturer’s directions. The results shown in Table 2 demonstrated that the clone of CGMMV Ontario strain was fully infectious.

Table 2: ELISA results of cucumber plants inoculated with the CGMMV clone and wild-type CGMMV isolate after 2 weeks inoculation.

[0137] In addition, as seen in Figures 1A to C, the CGMMV Ontario strain clone expressed in Agrobacterium tumefaciens produced similar symptoms in infected leaves (Figure 1A) as those produced by infection with the wild-type CGMMV Ontario strain isolate (Figure 1 B). Figure 1 C shows uninfected leaves as a negative control.

Example 2: Attenuated CGMMV strains

Mutant CGMMV Ontario strain ONB

[0138] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out to introduce mutations (c.1498A>G; c.3430C>T;

c.3528A>G; c.4248C>T; and c.6228C>T) corresponding to those observed in the attenuated SH33b strain of CGMMV (Tan et al, Ann. Phytopathoi. Soc. Jpn (1997), 63(6): 470-474). These mutations resulted in amino acid substitutions in the encoded viral proteins (E480G and A1 124V in the 129 kDa protein; E480G, A1 124V, N1 157D, and P1397S in the 186 kDa protein; and A156V in the coat protein).

[0139] Mutations were introduced using the QuikChange™ Lightning Multi Site- Directed Mutagenesis kit (Agilent Technologies), following the manufacturer’s instructions, and using the mutagenic primers listed in Table 3. Nucleotide residues indicated in bold indicate sites of mutation. The resulting mutant CGMMV strain was designated Ontario strain ONB.

Table 3: Primers used to produce mutant CGMMV Ontario strain ONB

[0140] The cDNA genome sequence of CGMMV strain ONB is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAAT GG CAAACATT AAT G AACAAAT CAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACTG ATG CGTATCCG G AGTTTTCG A TTAG CTTTACCG CCACCAAG AACTCTGTACACTCCCTTG CG GGTG GTCTG AGG C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATG GT CACACAACATGTACAG AG G CACAAGG G ATCTG G CG G GTG CAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAG G AGGTACG AT AATT CTCCCTGTGCGGT CACCTGTT CAGACGTTTT CCAAG AGT GTT CCTAT G ATTTT GGG AGCGGT AGGG A T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG GAGGCATTGCTTTTAGGTTCACCTGTAGGTAATTTAAATAGTATTGGGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATT AATTGTG AT G CGT ACTT ATTTT CCTG CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AG G TTG GT CAG AG CAG AC AC AC AT ATG CTT CAT AAAT CTGTGGGG C A CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCG AT CTT CCAA G ACAAAG CCACGTTTT CT GTGTGGTTT CCT G AG GCG AAG CGT AAG GT GTTG ATA CCCAAGTTTGAACTTTCAAGATTCCTTTCTGGGAATGTGAAAATCTCTAGGATGC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAGGCCTTAGTGTGGAAGAATGTTCAGTCCTTTGTGGAATCTATACGCTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AG G AAG GT ACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGGAGGACTAGGTGATTGCGTCTATGATG CT CT AGTT CAAACCGG CT GGTTT GAT ACCT CT AG CG ACG AACT G AAAGTTTT G C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAGG G AT GT ACG AGG CAG AT G CAAAG AT CG AG AG AG AG AGTGTCT CT G AGTT G CT CG CTT CCGGT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACG G GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GATACATGTGAAGATATGGATGTAACTGAAGATATGGAGGATATAGTGTTGATG GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTTACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG GAAAG ACCG CCG AAATT AT AG CG AGG GT CAATT G G AAAACCG AT CT AGT ATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTG ACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGTGATGCT AAG CAAAT CCCGTTTATAAACAGAGT CAT G AATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG GAAAG AT AAT AACGTTT AC T CAAT CT GAT AAG CAGT CCTT GAT CAAG AGT G GGTACAAT G ACGTG AACACT GT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CGACTCCGATAGGTTTAATTGCCCGTGATTCACCACATGTACTAGTGGCCTTAA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTAT AAT AGTG G AT GT G G AAAAGGTCG ACC AGT CG AT CTT G ACT AT GTTT G CTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT G A T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G CAG G AGTT CTAT GAT AG ATGCCTT CCT GGG AATT CCTT CGTGCT G AAT GATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT GGTT AAG AGTG AAG CG CAG AATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCT GAT CT AG CT G GG ACTGTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AGG G CT AGTT CCATT CAAAG TTT TTCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTC AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G CACAT GATT AAG CG G CAG C CCAAG AGT CGGTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TT GTTT AT CACCCT AAAGTG GT AAAT G CAGTTTT CGGTCCG GTTTTT AAGTATTT G ACCACCAAGTTT CTT AG CAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AG G AAAAAAT CAG AAG AT CT GC AG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT A T GAG ATT CTT GAG CTGG AT GTTT CT AAAT AT G ACAAGTCACAAT CCG ATTT CCAT TT CT CT ATT GAG AT GG CAATTT GGG AAAAATT G G GG CTG G ACG AT ATTTT G G CTT GGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGGA T AAAG ACG CT C ATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AG G T AAT ACCTTT ATT AT CG CAG CGT GTGTAG CT AGT ATGTTG CCGTT AG ACAAGT GT TTT AAAG CT AGTTTTT GTGGTG AT GATT CG CTG ATCT ACCTT CCT AAGG GTTT GG AGTATCCTG ATATACAG G CT ACT G CCAACTT G GTTT G G AATTTT GAG G CG AAACT TTT CCG AAAG AAGT ATG GTT ACTT CTGT GGG AAGT AT AT AATT CACCAT G CCAAC GGCTGTATTGTTTACCCTGACCCTTTAAAATTAATTAGTAAATTAGGTAATAAGAG TCTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGC T CAT AG TTT GTTT AAT G GT GCTT ATTT CCATTT ACT CG ACG ATG CAAT CCACG AAT T ATTT CCT AACG CTGGG G GTTG CAGTTTTGTAATTAATTGTTTGTG C AAGT ATTT G AGTG AT AAG CG CCTTTT CCGT AGTCTTT AT AT AG AT GTCTCT AAGT AAG GT GTC GGT CG AG AACT CATT GAAACCCGAG AAGTTT GTT AAAAT CT CTT GGGT CG AT AA GTTG CTCCCT AACTATTTTTCCATTCTT AAGTATTT ATCT AT AACTG ACTTT AG CG TAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTG TTG ATCTT AC AAAAC ACCTTT ATGT C AC ATT GTTGGGCGTTGTGGTTTCTGGTGT ATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACA CAAG G ATG CATTCTGTTG CAG AG GG AACTATATGC AAATTTTCAGCTCCCG CCA CCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGG AT G CCCTT CG CG AT CCTT GGT CTTT ATTT GT GAG ACT CT CT AAT GT AG GG ATT AA AG AT GGTTT CCAT CCTTT G ACCTT AG AG GT CGCTTGTTT AGT CG CT ACAACT AAC T CT ATTAT CAAAAAGGGT CTTAGAGCTT CT GTAGT CG AGTCT GT CGT CT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAG TT G AACCTTT CTTT GAT AAA GTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCG CAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTT G G ACT CTGCTT CT G AAG AGT CCAGTT CTGTTT CTTT CG AAG AT GG CTT ACAAT CC GAT CACACCT AGCAAACTT ATTGCGTTTAGT GCTT CTT AT GTT CCCGT CAGG ACT TT ACTT AATTTT CT AGTT G CTT CACAAG GT ACCGCCTT CCAG ACT CAAG CGG G AA GAG ATT CTTT CCGCG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AG AT ATT A ATT CT AGGTT CCCAAAT G CGG GTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG G CCTAT CTT CGTTT CGCTT CTT AG CTCTACG G ATACG CGTAATAG GGTCATTG A G GTTGTAG ATCCTAG CAAT CCT ACG ACT G CTG AGTCG CTTAACG CTGTAAAG CG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC T ATTT CT AAGGGTTTT GAT GTTT AT GAT AGGGCTT CATTT G AAGCCGCGTTTT CG GT AGT CTGGT CAG AGGTT ACCACCT CG AAAGCTT AGCTT CG AGGGT CTT CT GAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:29).

[0141 ] The 129 kDa protein encoded by CGMMV strain ONB has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEI IARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTK (SEQ ID NO:30).

[0142] The 186 kDa protein encoded by CGMMV strain ONB has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPI IDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEI IARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRDIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHPKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKSEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:31).

[0143] The coat protein encoded by CGMMV strain ONB has the following sequence.

MAYNPITPSKLIAFSASYVPVRTLLNFLVASQGTAFQTQAGRDSFRESLSALPSSW DINSRFPNAGFYAFLNGPVLRPIFVSLLSSTDTRNRVIEWDPSNPTTAESLNAVKRT D D AST AAR AE IDNLIESISKGFDVYDRASF E AAF SVWVS E VTT S K A (SEQ ID NO:32).

Mutant CGMMV Ontario strain ONM

[0144] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above, but using the mutagenic primers listed in Table 4, to introduce mutations (c.315G>A; c.1660C>T; and c.4144C>T) corresponding to those observed in the attenuated VIROG-43M strain (Slavokhotova, A.A. , et al, American Journal of Plant Sciences (2016), 7: 724-732). Nucleotide residues indicated in bold indicate sites of mutation. These mutations resulted in amino acid substitutions in the encoded viral proteins (G86S and S534F in the 129 kDa protein; and G86S, S534F and P1362L in the 186 kDa protein). The resulting mutant CGMMV strain was designated Ontario strain ONM.

Table 4: Primers used to produce mutant CGMMV Ontario strain ONM

[0145] The cDNA genome sequence of CGMMV strain ONM is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAAT GG CAAACATT AAT G AACAAAT CAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAGCACAGAGCACACCAGGCTTGTAACTGATGCGTATCCGGAGTTTTCGA TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT G CG G GT AGTCT G AGG C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA T AT CGG CGGTAACT AT ACG CAG CACTT GTT CAAAG GT AG AT CAT ATGTG CATT G CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATGGTCACACAACATGTACAGAGGCACAAGGGATCTGGCGGGTGCAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAGG AGGTACG AT AATT CTCCCTGTG CG GT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG G AG G CATT G CTTTT AG GTT CACCT GT AG GT AATTT AAAT AGTATT G G GG CTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATT AATTGT G ATG CGT ACTT ATTTTCCT G CTGATGATAGGTACGTGTATATTAAGGAGTTTATGGTCAAGCGTGTGGATACTTT CTT CTTT AG G TTG GT CAG AG CAG AC AC AC AT ATG CTT CAT AAAT CTGTGGGG C A CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAG CCACGTTTT CT GTGTGGTTT CCT G AG GCG AAG CGT AAG GT GTTG ATA CCCAAGTTT GAACTTT CAAG ATT CCTTT CTGGG AAT GT G AAAAT CT CTAGG ATGC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAG G CCTT AGTGT G G AAG AAT GTT CAGT CCTTTGT GG AAT CT AT ACGCT CAAG A GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT T CG CAG ACAGT CTT AAAT CCG CCGTAG AAGG ACT AGGTG ATT G CGTCTATG ATG CTCTAGTTCAAACCG G CTGGTTTG ATACCTCTAG CG ACG AACTG AAAGTTTTG C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAGG GAT GT ACG AGG CAG AT G CAAAG AT CG AG AG AG AG AGTGTCTTT G AGTT G CT CG CTT CCG GT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGT CG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GAT ACAT GT G AAG AT AT G G ATGTAACT G AAG AT AT G GAG GAT AT AGT GTTG ATG GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTTACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AG GTT AT CAAACAT G AATTT CTTT AAGGT G AG CG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT G ACTG CG AAAAGTG CAAGTGGTTACTGTTACCAG CCG AAAAG AAG CACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG G AAAG ACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGT ATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTG ACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGTG ATG CT AAG CAAAT CCCGTTT AT AAACAG AGT CAT G AATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAG CAGT CCTT GAT CAAG AGT G GGTACAAT G ACGTG AACACT GT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CGACTCCGATAGGTTTAATTGCCCGTGATTCACCACATGTACTAGTGGCCTTAA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTATAATAGCGGATGTGGAAAAGGTCGACCAGTCGATCTTGACTATGTTTGCTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT AA T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G CAG G AGTT CTAT GAT AG AT G CCTT CCT GG G AATT CCTT CGTG CT G AAT G ATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT G GTT AAG AGT GAAGCG CAG AATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCTG AT CT AG CT GG G ACT GTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AGG G CT AGTT CCATT CAAAGTTT TTCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTC AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G C AC AT GATT AAG C G G CAG C CCAAG AGT CGGTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TT GTTT AT CACCTT AAAGTG GT AAAT G CAGTTTT CGGT CCG GTTTTT AAGT ATTT G ACCACC AAGTTT CTT AG CAT G GT AG AT AGTT CT AAGTTTTT CTTTT ACACT AG G AA AAAACCAG AAG AT CT G CAGG AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT GAG ATT CTT GAG CTGG ATGTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCATT TCTCTATTG AG ATGG CAATTTG GG AAAAATTGG G G CTGG ACG ATATTTTG G CTT GGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGGA T AAAG ACG CT CATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AGG T AAT ACCTTT ATT AT CG CAG CGT GTGT AG CT AGTATGTT G CCGTT AG ACAAGT GT TTTAAAGCTAGTTTTTGTGGTGATGATTCGCTGATCTACCTTCCTAAGGGTTTGG AGT AT CCTG AT AT ACAG G CT ACT G CCAACTT G GTTT G G AATTTT GAG G CG AAACT TTT CCG AAAG AAGT ATG GTT ACTT CTGT GGG AAGT AT AT AATT CACCAT G CCAAC GGCTGTATTGTTTACCCTGACCCTTTAAAATTAATTAGTAAATTAGGTAATAAGAG TCTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGC T CAT AGTTT GTTT AAT G GT GCTT ATTT CCATTT ACT CG ACG AT G CAAT CCACG AAT T ATTT CCT AACG CTGGGGGTTG C AGTTTT GT AATT AATT GTTTG TG C AAG T ATTT GAGTGATAAGCGCCTTTTCCGTAGTCTTTATATAGATGTCTCTAAGTAAGGTGTC GGT CG AG AACT CATT GAAACCCG AG AAGTTT GTTAAAAT CT CTT GGGT CG AT AA GTTG CTCCCT AACT ATTTTT CCATT CTT AAGT ATTT AT CT AT AACT G ACTTT AG CG TAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTG TTG ATCTT AC AAAAC ACCTTT ATG T C AC ATT GTTGGGCGTTGTGGTTTCTGGTGT ATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACA CAAG G ATG CATTCTGTTG C AG AG G G AACT AT AT G C AAATTTT C AG CTCCCG CCA CCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGG AT G CCCTT CG CG AT CCTT GGT CTTT ATTT GT GAG ACT CT CT AAT GT AG GG ATT AA AGAT GGTTT CCAT CCTTT G ACCTT AG AGGT CGCTTGTTT AGT CGCT ACAACTAAC TCTATTATCAAAAAGGGTCTTAGAGCTTCTGTAGTCGAGTCTGTCGTCTCTTCCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAGTT G AACCTTT CTTT GAT AAA GTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCG CAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTT GG ACT CT GCTT CT GAAG AGT CCAGTT CTGTTT CTTT CG AAG ATGGCTTACAAT CC GAT CACACCT AGCAAACTT ATTGCGTTTAGT GCTT CTT AT GTT CCCGT CAGG ACT TT ACTT AATTTT CT AGTT G CTT CACAAG GT ACCGCCTT CCAG ACT CAAG CGG G AA GAG ATT CTTT CCGCG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AGAT ATT A ATT CT AGGTT CCCAAAT G CGG GTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG G CCTAT CTT CGTTT CGCTT CTT AGCT CT ACG GAT ACG CGT AAT AG G GTCATTG A GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC T ATTT CT AAGGGTTTT GAT GTTT AT GAT AGGGCTT CATTT G AAGCCGCGTTTT CG GTAGTCTGGTCAGAGGCTACCACCTCGAAAGCTTAGCTTCGAGGGTCTTCTGAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:39).

[0146] The 129 kDa protein encoded by CGMMV strain ONM has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEI IARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTK (SEQ ID NO:40). [0147] The 186 kDa protein encoded by CGMMV strain ONM has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRNIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHLKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKPEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:41).

[0148] The mutant clones ONB and ONM were each transformed into Agrobacterium tumefaciens strain EHA105 by electroporation and used to inoculate the cotyledon of 1-2 week old cucumber plants under laboratory greenhouse conditions using the method described in Example 1. As seen in Figure 2B, two weeks after inoculation, mutant strain ONB induced visible symptoms including mottle and mosaic symptoms, Likewise, as seen in Figure 2C, two weeks after inoculation, mutant strain ONM also induced visible symptoms, although these symptoms were milder than those induced by the mutant ONB strain. For comparison, Figure 2A shows a plant exhibiting symptoms induced by inoculation with the wild-type Ontario strain CGMMV under the same conditions, and Figure 2D shows an uninfected control plant grown under the same conditions.

Mutant CGMMV Ontario strain ONBM

[0149] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above to introduce mutations corresponding to those of mutants ONB and ONM (c.315G>A; c.1498A>G ;

C.1660OT; C.3430OT; c.3528A>G; c.4144C>T; c.4248C>T; and c.6228C>T) .

These mutations resulted in amino acid substitutions in the encoded viral proteins (G86S, E480G , S534F and A1 124V in the 129 kDa protein; G86S, E480G , S534F, A1 124V, N 1 157D, P1362L, and P1397S in the 186 kDa protein; and A156V in the coat protein). The resulting mutant CGMMV strain was designated Ontario strain ONBM .

[0150] The cDNA genome sequence of CGMMV strain ONBM is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAATGG CAAACATTAATG AACAAATCAACAACCAACG CG ACG COG CGG CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAGCACAGAGCACACCAGGCTTGTAACTGATGCGTATCCGGAGTTTTCGA TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT G CG GGT AGTCT G AGG C T CCTT G AACT GG AAT AT AT GAT GAT G CAAGTGCCCT ACG G CT CACCTT GTTATG A TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATG GT CACACAACATGTACAG AG G CACAAGG G ATCTG G CG GGTG CAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAGG AGGTACG AT AATT CTCCCTGTG CG GT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG G AG G CATT G CTTTT AG GTT CACCTGTAG GT AATTT AAAT AGTATT G G GG CTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATTAATT GT GAT GCGT ACTT ATTTT CCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AGGTTGGT CAG AG CAG ACACACAT AT G CTT CAT AAAT CTGT GG GG CA CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAG CCACGTTTT CTGTGT G GTTT CCT GAG G CG AAGCGT AAGGT GTTG ATA CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGG GAATGTG AAAAT CT CT AG GAT GC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAG G CCTT AGTGTG G AAG AATGTTCAGTCCTTTGTG G AATCTAT ACG CTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGGAGGACTAGGTGATTGCGTCTATGATG CTCTAGTTCAAACCG G CT G GTTT GAT ACCT CT AG CG ACG AACT G AAAGTTTT G C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAG G G ATGTACG AGG CAG AT G CAAAG AT CG AG AG AG AG AGTGTCTTT G AGTT G CT CG CTT CCG GT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATTACAGT GG AGT CG AATTT GAT GT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCTATG CT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGT ACCCT CT CT CCT GAG AT G G GTG CTT CTGTTG CGTTAT CCAAT ACCT CTGTA GAT ACAT GT G AAG AT AT G GAT GT AACT G AAG AT AT GG AG G AT AT AGT GTT GAT G GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTT ACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT G AGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC T CT G AG ACCAAAGT CT AT AG CGTAATT CGT AGTTT AG AGGT ACT AAAT AAG G AAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG G AAAG ACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGTATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTG ACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGG GTTTACTTAATTTTG CGTTG AAG ATCTCAG GTTGTAAAAAG GCCTTCG T CTTT GGT GATGCTAAGCAAAT CCCGTTTATAAACAGAGTCAT GAATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CG ACT CCG AT AG GTTT AATT G CCCGT GATT CACCACAT GT ACT AGT GG CCTT AA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTAT AAT AGTG G AT GT G G AAAAGGT CG ACCAGT CG AT CTT G ACT ATGTTT GCTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT G A T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G C AG G AGTT CT AT GAT AG AT G CCTT CCT GG G AATT CCTT CGTG CT G AAT G ATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT G GTT AAG AGTG AAG CGCAGAATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCTG AT CT AG CT GG G ACT GTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AG GG CT AGTT CCATT CAAAGTTT TTCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTC AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G C AC AT GATT AAG C G G CAG C CCAAG AGT CG GTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TTGTTTATCACCTTAAAGTGGTAAATGCAGTTTTCGGTCCGGTTTTTAAGTATTTG ACCACC AAGTTT CTT AG CAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AGG AA AAAAT CAG AAG AT CTGC AG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT GAG ATT CTT GAG CTGG ATGTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCATT TCTCTATTG AG ATGG CAATTTG GG AAAAATTGG G G CTGG ACG ATATTTTG G CTT GGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGGA T AAAG ACG CT CATTT ACT AT CAACGG AAGTCT G GTG ATGTAACT ACTTT CAT AG G T AAT ACCTTT ATT AT CG CAG CGT GTGTAG CT AGT ATGTTG CCGTT AG ACAAGTGT TTT AAAG CT AGTTTTT GTGGTG AT GATT CG CTG ATCT ACCTT CCT AAGG GTTT GG AGTATCCTGATATACAGGCTACTGCCAACTTGGTTTGGAATTTTGAGGCGAAACT TTT CCG AAAG AAGT ATG GTT ACTT CTGT GGG AAGT AT AT AATT CACCAT G CCAAC GGCTGTATTGTTTACCCTGACCCTTTAAAATTAATTAGTAAATTAGGTAATAAGAG TCTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGC T CAT AGTTT GTTT AAT GGT GCTT ATTT CCATTT ACT CG ACG ATG CAAT CCACG AAT T ATTT CCT AACG CTGGGGGTTG CAG TTTT GT AATT AATT G TTT GTG CAAGT ATTT G AGTG AT AAG CG CCTTTT CCGT AGT CTTT AT AT AG ATGTCT CT AAGTAAG GTGTC GGT CG AG AACT CATT GAAACCCGAG AAGTTT GTT AAAAT CT CTT GGGT CG AT AA GTTG CTCCCT AACT ATTTTT CCATT CTT AAGTATTT AT CT AT AACT G ACTTT AGCG TAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTG TT GAT CTT ACAAAACACCTTT ATGT CACATT GTTGGGCGTTGT GGTTT CTG GTGT ATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACA CAAG G ATG CATTCTGTTG C AG AG G G AACT AT AT G C AAATTTT C AG CTCCCG CCA CCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGG AT G CCCTT CG CG AT CCTT GGT CTTT ATTT GT GAG ACT CT CT AAT GT AG GG ATT AA AG AT G GTTT CCAT CCTTT G ACCTT AG AG GT CGCTTGTTT AGT CG CT ACAACT AAC T CT ATTAT CAAAAAGGGT CTTAGAGCTT CT GTAGT CG AGTCT GT CGT CT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAGTT G AACCTTT CTTT GAT AAA GTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCG CAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTT GG ACT CT GCTT CT GAAG AGT CCAGTT CTGTTT CTTT CG AAG AT GGCTTACAAT CC GAT CACACCT AGCAAACTT ATTGCGTTTAGT GCTT CTT AT GTT CCCGT CAGG ACT TT ACTT AATTTT CT AGTT G CTT CACAAG GT ACCGCCTT CCAG ACT CAAG CGG G AA GAG ATT CTTT CCGCG AGTCCCT GT CTGCGTTACCCT CGTCT GT CGT AG AT ATT A ATT CT AGGTT CCCAAAT G CGG GTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG GCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCATTGA GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC TATTTCTAAGGGTTTTGATGTTTATGATAGGGCTTCATTTGAAGCCGCGTTTTCG GT AGT CTG GT CAG AGGTT ACCACCT CG AAAGCTT AG CTT CG AGG GT CTT CT GAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:42).

[0151 ] The cDNA genome sequence of the attenuated CGMMV strain ONBM differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO: 18) at least in that:

the nucleotide at position 315 of SEQ ID NO:42 is A;

the nucleotide at position 1498 of SEQ ID NO:42 is G;

the nucleotide at position 1660 of SEQ ID NO:42 is T;

the nucleotide at position 3430 of SEQ ID NO:42 is T;

the nucleotide at position 3528 of SEQ ID NO:42 is G;

the nucleotide at position 4144 of SEQ ID NO:42 is T;

the nucleotide at position 4248 of SEQ ID NO:42 is T; and

the nucleotide at position 6228 of SEQ ID NO:42 is T.

[0152] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVH ITLVDG VPGCG KTAEI I ARVNWKTDLVLTPG REAAAM I RRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTK (SEQ ID NO:43).

[0153] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM differs from the 129 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:63) at least in that:

position 86 of SEQ ID NO:43 is serine (S, Ser);

position 480 of SEQ ID NO:43 is glycine (G, Gly);

position 534 of SEQ ID NO:43 is phenylalanine (F, Phe); and

position 1 124 of SEQ ID NO:43 is valine (V, Val).

[0154] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRDIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHLKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKSEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:44). [0155] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that:

position 86 of SEQ ID NO:44 is serine (S, Ser);

position 480 of SEQ ID NO:44 is glycine (G, Gly);

position 534 of SEQ ID NO:44 is phenylalanine (F, Phe);

position 1 124 of SEQ ID NO:44 is valine (V, Val);

position 1 157 of SEQ ID NO:44 is aspartic acid (D, Asp);

position 1362 of SEQ ID NO:44 is leucine (L, Leu); and

position 1397 of SEQ ID NO:44 is serine (S, Ser).

[0156] The coat protein encoded by the attenuated CGMMV strain ONBM has the sequence of SEQ ID NO:32 and differs from the coat protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:65) at least in that position 156 of SEQ ID NO:32 is valine (V, Val).

Mutant CGMMV Ontario strain ONBM-2

[0157] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above to introduce mutations

corresponding to those of mutants ONB and ONM (c.315G>A; c.1498A>G;

C.1660OT; C.3430OT; c.3528A>G; c.4144C>T; C.42480T; and C.62280T) in addition to the mutation c.3334C>T introduced by randomly replacing C by T during PCR amplification. These mutations resulted in amino acid substitutions in the encoded viral proteins (G86S, E480G, S534F, A1092V and A1 124V in the 129 kDa protein; G86S, E480G, S534F, A1092V, A1 124V, N1 157D, P1362L, and P1397S in the 186 kDa protein; and A156V in the coat protein). The resulting mutant CGMMV strain was designated Ontario strain ONBM-2.

[0158] The cDNA genome sequence of CGMMV strain ONBM-2 is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAAT GG CAAACATT AAT G AACAAAT CAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACTG ATG CGTATCCG G AGTTTTCG A TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT G CG G GTAGTCTG AGG C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA T ATG GT C AC AC AAC AT G T AC AG AG G C AC AAG G G ATCTG G CG G GTG CAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAG G AGGTACG AT AATT CTCCCTGTGCGGT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG GAGGCATTGCTTTTAGGTTCACCTGTAGGTAATTTAAATAGTATTGGGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATTAATT GT GAT GCGT ACTT ATTTT CCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AG G TTG GT CAG AG CAG AC AC AC AT ATG CTT CAT AAAT CTGTGGGG C A CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCG AT CTT CCAA G ACAAAG CCACGTTTT CT GTGTGGTTT CCT G AG GCG AAG CGT AAG GT GTTG ATA CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGG GAATGTG AAAAT CT CT AG GAT GC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAGGCCTTAGTGTGGAAGAATGTTCAGTCCTTTGTGGAATCTATACGCTCAAGA GT AATT GT AAACG G AGTTT CG GT G AAAT CT G AAT G G AACGTACCG GTT GAT CAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGGAGGACTAGGTGATTGCGTCTATGATG CT CT AGTT CAAACCGGCT GGTTT GATACCT CT AGCG ACG AACT G AAAGTTTTGC T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAGG GAT GT ACG AGG CAG AT G CAAAG AT CG AG AG AG AG AGTGTCTTT G AGTT G CT CG CTT CCG GTG ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACG G GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GAT ACAT GT G AAG AT AT G GAT GT AACT G AAG AT AT GG AG G AT AT AGT GTT GAT G GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTT ACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG GAAAGACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGTATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTGACAACGTTAG AACTTT CG ATT CTTTTGT GAT G AATAAGAAAATCTTCAAGTTTGACGCTGTCTATGTTGACGAGGGTCTGATGGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGT GATGCT AAG CAAAT CCCGTTTATAAACAGAGTCAT G AATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CG CT ACT ACT AGT CCGGTTGT ACATT CTGT G AAG G CG ATTAAAGTGTCAG GG G C CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG GAAAG AT AAT AACGTTT AC T CAAT CT GAT AAG CAGT CCTT GAT CAAG AGT G GGTACAAT G ACGTG AACACT GT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CGACTCCGATAGGTTTAATTGTCCGTGATTCACCACATGTACTAGTGGCCTTAAC GAGG CACACTAAG G CAATG GTGTATTATACTGTTGTGTTCG ATG CAGTTACAAG T AT AAT AGT GG ATGT GG AAAAG GT CG ACCAGT CG AT CTT G ACT AT GTTT G CT AC CACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT GAT ATTTTCCTCCCTGTTAGTAAAACGGGGTTTTATACAGACATGCAGGAGTTCTATG AT AG AT GCCTT CCT G GG AATT CCTT CGTG CT G AAT G ATTT CG AT G CCGT AACCAT GCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGTAA TTTAGATCCAGTACCCGCTTTGGTTAAGAGTGAAGCGCAGAATTTTCTGATTCCC GTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATCTT GTAGCTATGATAAAGAGGAATATGAATACTCCTGATCTAGCTGGGACTGTGGAT AT AACT AAT ATGT CG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTTGTT AG AG A CG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AG GG CT AGTT CCATT CAAAGTTTT TCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTCA ATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G CACATG ATTAAGCG G CAG CC CAAG AGT CGGTTG G AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT ATT GTTT AT CACCTT AAAGT GGTAAAT GCAGTTTT CGGT CCGGTTTTT AAGT ATTT G A CCACCAAGTTT CTT AGCAT GGT AG AT AGTT CT AAGTTTTT CTTTT AC ACT AG G AAA AAAT CAG AAG AT CT GCAG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT G AG ATT CTT GAG CTGG AT GTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCATTT CTCTATTG AG ATG G CAATTTGG G AAAAATTGGG G CTGG ACG ATATTTTG G CTTG GATGTGGTCTATGGGT CACAAG AG AACT AT ACT G CAAG ATTT CCAAGCCG GG AT AAAG ACG CT CATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AG GT AATACCTTTATTATCGCAGCGTGTGTAGCTAGTATGTTGCCGTTAGACAAGTGTT TTAAAGCTAGTTTTTGTGGTGATGATTCGCTGATCTACCTTCCTAAGGGTTTGGA GTATCCT GAT AT ACAGG CT ACT G CCAACTT G GTTT GG AATTTT G AGG CG AAACTT TT CCG AAAG AAGT ATGGTT ACTT CT GTGGG AAGTATAT AATT CACCAT GCCAACG G CTGT ATTGTTT ACCCT G ACCCTTT AAAATT AATT AGT AAATT AG GT AAT AAG AGT CTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGCT CATAGTTTGTTTAATGGTGCTTATTTCCATTTACTCGACGATGCAATCCACGAATT ATTTCCTAACGCTGGGGGTTGCAGTTTTGTAATTAATTGTTTGTGCAAGTATTTG AGTGATAAGCGCCTTTTCCGTAGTCTTTATATAGATGTCTCTAAGTAAGGTGTCG GT CG AG AACT CATT G AAACCCG AG AAGTTT GTT AAAAT CT CTT G GGTCG AT AAGT TG CTCCCT AACT ATTTTT CCATT CTT AAGTATTT AT CT AT AACT G ACTTT AG CGTA GTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTGTT GAT CTT ACAAAACACCTTT AT GT CACATTGTT G G GCGTTGTG GTTTCTG GTGTAT GGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACACA AGG ATG CATTCTGTTG CAG AG GG AACT AT ATG CAAATTTTCAG CTCCCG CCACC GTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGGAT G CCCTT CG CG AT CCTT G GTCTTT ATTT GT GAG ACT CT CT AAT GT AGG GATT AAAG AT G GTTT CCAT CCTTT G ACCTT AG AG GT CG CTT GTTT AGT CG CT AC AACT AACT C T ATT AT CAAAAAG GGT CTT AG AG CTT CTGTAGT CG AGT CTGTCGTCT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAG TT G AACCTTT CTTT GAT AAAG TTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCGC AGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTTG G ACT CT G CTT CT G AAG AGT CCAGTT CT GTTT CTTT CG AAG AT G GCTT ACAAT CCG AT CACACCT AG CAAACTT ATT G CGTTT AGTG CTT CTT AT GTT CCCGTCAGG ACTT T ACTT AATTTT CT AGTT G CTT CAC AAGGTACCG CCTT CCAG ACT CAAG CGGG AA GAG ATT CTTT CCG CG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AG AT ATT A ATT CT AGGTT CCCAAAT G CGG GTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG GCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCATTGA GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC TATTTCTAAGGGTTTTGATGTTTATGATAGGGCTTCATTTGAAGCCGCGTTTTCG GT AGT CTG GT CAG AGGTT ACCACCT CG AAAG CTT AG CTT CG AGG GT CTT CT GAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:45). [0159] The cDNA genome sequence of the attenuated CGMMV strain ONBM-2 differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO: 18) at least in that:

the nucleotide at position 315 of SEQ ID NO:45 is A;

the nucleotide at position 1498 of SEQ ID NO:45 is G;

the nucleotide at position 1660 of SEQ ID NO:45 is T;

the nucleotide at position 3334 of SEQ ID NO:45 is T;

the nucleotide at position 3430 of SEQ ID NO:45 is T;

the nucleotide at position 3528 of SEQ ID NO:45 is G;

the nucleotide at position 4144 of SEQ ID NO:45 is T;

the nucleotide at position 4248 of SEQ ID NO:45 is T; and

the nucleotide at position 6228 of SEQ ID NO:45 is T.

[0160] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-2 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTK (SEQ ID NO:46).

[0161] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-2 differs from the 129 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:63) at least in that:

position 86 of SEQ ID NO:46 is serine (S, Ser);

position 480 of SEQ ID NO:46 is glycine (G, Gly);

position 534 of SEQ ID NO:46 is phenylalanine (F, Phe); position 1092 of SEQ ID NO:46 is valine (V, Val); and position 1 124 of SEQ ID NO:46 is valine (V, Val).

[0162] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-2 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRDIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHLKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKSEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:47).

[0163] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-2 differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that:

position 86 of SEQ ID NO:47 is serine (S, Ser);

position 480 of SEQ ID NO:47 is glycine (G, Gly);

position 534 of SEQ ID NO:47 is phenylalanine (F, Phe);

position 1092 of SEQ ID NO:47 is valine (V, Val);

position 1 124 of SEQ ID NO:47 is valine (V, Val);

position 1 157 of SEQ ID NO:47 is aspartic acid (D, Asp);

position 1362 of SEQ ID NO:47 is leucine (L, Leu); and

position 1397 of SEQ ID NO:47 is serine (S, Ser). [0164] The coat protein encoded by the attenuated CGMMV ONBM-2 strain has the sequence of SEQ ID NO:32 and differs from the coat protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:65) at least in that position 156 of SEQ ID NO:32 is valine (V, Val).

Mutant CGMMV Ontario strain ONBM-3

[0165] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above to introduce mutations

corresponding to those induced in the cDNA genome of the cloned CGMMV Ontario strain mutants ONB and ONM (c.315G>A; c.1498A>G; c.1660C>T; c.3430C>T; c.3528A>G; c.4144C>T; c.4248C>T; and c.6228C>T) in addition to the mutation c.4969G>A introduced by randomly replacing G by A during PCR amplification.

These mutations resulted in amino acid substitutions in the encoded viral proteins (G86S, E480G, S534F and A1 124V in the 129 kDa protein; G86S, E480G, S534F, A1 124V, N1 157D, P1362L, P1397S and R1637H in the 186 kDa protein; and A156V in the coat protein). The resulting mutant CGMMV strain was designated Ontario strain ONBM-3.

[0166] The cDNA genome sequence of CGMMV strain ONBM-3 is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAATGG CAAACATTAATG AACAAATCAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TG GT CAG CACAG AG CACACCAG G CTTGTAACT G ATG CG T AT CCG G AGTTTT CG A TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT G CG GGT AGTCT G AGG C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATGGTCACACAACATGTACAGAGGCACAAGGGATCTGGCGGGTGCAGACCTC TTCCAACTTTTCAGATAGATGCATTCAGGAGGTACGATAATTCTCCCTGTGCGGT CACCTGTT CAG ACGTTTT CCAAG AGT GTTCCT AT G ATTTTGG G AG CG GT AGG G A T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG G AG G CATT G CTTTT AG GTT CACCTGTAG GT AATTT AAAT AGTATT G GGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATT AATTGT GAT G CGT ACTT ATTTT CCTG CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AGGTTGGT CAG AG CAGACACACAT AT GCTT CAT AAAT CTGTGGGG CA CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA GACAAAGCCACGTTTTCTGTGTGGTTTCCTGAGGCGAAGCGTAAGGTGTTGATA CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGGG AAT GT G AAAAT CT CTAGG ATGC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAG G CCTT AGTGT G G AAG AAT GTT CAGTCCTTT GTG G AAT CT AT ACG CTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GT ACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGGAGGACTAGGTGATTGCGTCTATGATG CTCTAGTTCAAACCGG CTG GTTTG ATACCTCTAG CG ACG AACTG AAAGTTTTG C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAG GG AT GT ACG AG GCAG AT G CAAAG AT CG AG AG AG AG AGTGTCTTT G AGTT G CT CG CTT CCG GT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CAT G TT ATCG AAG CT ATTTTTT CG C AG AAAG CTG G G G T G AC AG T AACG G GT CTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GATACATGTGAAGATATGGATGTAACTGAAGATATGGAGGATATAGTGTTGATG GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTTACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATT GATT ACTT ATCG GCCTCT CTTGGTT CTACG CTG GGT AAT CT G GAG AG AATT G TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC T CT G AG ACCAAAGT CT AT AG CGTAATT CGT AGTTT AG AGGT ACT AAAT AAG G AAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG GAAAGACCGCCGAAATTATAGCGAGGGTCAATTGGAAAACCGATCTAGTATTGA CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGTCACCTGTGGCAACCAGTGACAACGTTAGAACTTTCGATTCTTTTGTGATG AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CAT ACGGGTTT ACTT AATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGTG ATG CT AAG CAAAT CCCGTTT AT AAACAG AGT CAT G AATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT GAGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CG ACT CCG AT AG GTTT AATT GCCCGT GATT CACCACAT GT ACT AGT GG CCTT AA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTAT AAT AGTG G AT GT G G AAAAGGT CG ACCAGT CG AT CTT G ACT ATGTTT GCTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT G A T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G CAG G AGTT CTAT GAT AG AT G CCTT CCT GG G AATT CCTT CGTG CT G AAT G ATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT G GTT AAG AGTG AAG CGCAG AATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCT GAT CT AG CT G GG ACTGTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AG GG CT AGTT CCATT CAAAGTTT TTCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTC AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G C AC AT GATT AAG C G G CAG C CCAAG AGT CGGTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TTGTTTATCACCTTAAAGTGGTAAATGCAGTTTTCGGTCCGGTTTTTAAGTATTTG ACCACCAAGTTT CTT AG CAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AGG AA AAAAT CAG AAG AT CTGC AG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT GAG ATT CTT GAG CTGG ATGTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCATT TCTCTATTG AG ATGG CAATTTG GG AAAAATTGG G G CTGG ACG ATATTTTG G CTT GGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGGA T AAAG ACG CT CATTT ACT AT CAACGG AAGTCT G GTG ATGTAACT ACTTT CAT AG G T AAT ACCTTT ATT AT CG CAG CGT GTGTAG CT AGT ATGTTG CCGTT AG ACAAGTGT TTT AAAG CT AGTTTTT GTGGTG AT GATT CG CTG ATCT ACCTT CCT AAGG GTTT GG AGTATCCTG ATATACAG G CT ACT G CCAACTT G GTTT G G AATTTT GAG G CG AAACT TTT CCG AAAG AAGT ATG GTT ACTT CTGT GG G AAGT AT AT AATT CACCAT GCCAAC GGCTGTATTGTTTACCCTGACCCTTTAAAATTAATTAGTAAATTAGGTAATAAGAG TCTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGC T CAT AG TTT GTTT AAT G GT GCTT ATTT CCATTT ACT CG ACG ATG CAAT CCACG AAT T ATTT CCT AACG CTGGGGGTTG C AGTTTT G T AATT AATT G TTT GTG C AAGT ATTT G AGTG AT AAG CACCTTTT CCGT AGTCTTT AT AT AG AT GTCTCT AAGTAAG GTGT C G GT CG AG AACT CATT G AAACCCG AG AAGTTT GTT AAAAT CT CTT G GGTCG AT AA GTTG CTCCCT AACT ATTTTT CCATT CTT AAGT ATTT AT CT AT AACT G ACTTT AG CG TAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTG TTGATCTTACAAAACACCTTTATGTCACATTGTTGGGCGTTGTGGTTTCTGGTGT ATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACA CAAG G ATG CATTCTGTTG CAG AG GG AACTATATGC AAATTTTCAG CTCCCG CCA CCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGG AT G CCCTT CG CG AT CCTT GGT CTTT ATTT GT GAG ACT CT CT AAT GT AG G G ATT AA AG AT GGTTT CCAT CCTTT G ACCTT AG AGGT CG CTT GTTT AGT CGCT ACAACT AAC T CT ATTAT CAAAAAGGGT CTTAGAGCTT CT GTAGT CG AGTCT GT CGT CT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAG TT G AACCTTT CTTT GAT AAA GTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCG CAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTT GG ACT CT GCTT CT GAAG AGT CCAGTT CTGTTT CTTT CG AAG AT GGCTTACAAT CC GAT CACACCT AGCAAACTT ATTGCGTTTAGT GCTT CTT AT GTT CCCGT CAGG ACT TT ACTT AATTTT CT AGTT G CTT CACAAG GT ACCGCCTT CCAG ACT CAAG CGG G AA GAG ATT CTTT CCGCG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AG AT ATT A ATT CT AGGTT CCCAAATGCGGGTTTTT ACGCTTT CCT CAACGGTCCTGTGTT GAG GCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCATTGA GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC T ATTT CT AAGGGTTTT GAT GTTT AT GAT AGGGCTT CATTT G AAGCCGCGTTTT CG GTAGTCTGGTCAGAGGTTACCACCTCGAAAGCTTAGCTTCGAGGGTCTTCTGAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:48).

[0167] The cDNA genome sequence of the attenuated CGMMV strain ONBM-3 differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO: 18) at least in that:

the nucleotide at position 315 of SEQ ID NO:48 is A;

the nucleotide at position 1498 of SEQ ID NO:48 is G;

the nucleotide at position 1660 of SEQ ID NO:48 is T;

the nucleotide at position 3430 of SEQ ID NO:48 is T;

the nucleotide at position 3528 of SEQ ID NO:48 is G;

the nucleotide at position 4144 of SEQ ID NO:48 is T;

the nucleotide at position 4248 of SEQ ID NO:48 is T;

the nucleotide at position 4969 of SEQ ID NO:48 is A; and the nucleotide at position 6228 of SEQ ID NO:48 is T.

[0168] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-3 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTK (SEQ ID NO:49).

[0169] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-3 differs from the 129 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:63) at least in that:

position 86 of SEQ ID NO:49 is serine (S, Ser);

position 480 of SEQ ID NO:49 is glycine (G, Gly);

position 534 of SEQ ID NO:49 is phenylalanine (F, Phe); and

position 1 124 of SEQ ID NO:49 is valine (V, Val).

[0170] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-3 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRDIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHLKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKSEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKHLFRSLYIDVSK (SEQ ID

NO:50).

[0171] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-3 differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that:

position 86 of SEQ ID NO:50 is serine (S, Ser);

position 480 of SEQ ID NO:50 is glycine (G, Gly);

position 534 of SEQ ID NO:50 is phenylalanine (F, Phe);

position 1 124 of SEQ ID NO:50 is valine (V, Val);

position 1 157 of SEQ ID NO:50 is aspartic acid (D, Asp);

position 1362 of SEQ ID NO:50 is leucine (L, Leu);

position 1397 of SEQ ID NO:50 is serine (S, Ser); and

position 1637 of SEQ ID NO:50 is histidine (H, His).

[0172] The coat protein encoded by the attenuated CGMMV strain ONBM-3 has the sequence of SEQ ID NO:32 and differs from the coat protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:65) at least in that position 156 of SEQ ID NO:32 is valine (V, Val).

Mutant CGMMV Ontario strain ONAL-1

[0173] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above, using the mutagenic primers listed in Table 5 to introduce the mutation c.3334C>T. Nucleotide residues indicated in bold indicate sites of mutation. This mutation resulted in an A1092V amino acid substitution in the encoded viral 129 kDa and 186 kDa proteins. The resulting mutant CGMMV strain was designated Ontario strain ONAL-1. Table 5: Primers used to produce mutant CGMMV Ontario strain ONAL-1

[0174] The cDNA genome sequence of CGMMV strain ONAL-1 is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAAT GG CAAACATT AAT G AACAAAT CAACAACCAACGCG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACT G ATGCGT AT CCGG AGTTTTCGA TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT GCGGGTGGTCT GAG G C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATG GT CACACAACATGTACAG AG G CACAAGG G ATCTG G CG G GTG CAGACCTC TT CCAACTTTT CAG AT AG AT G CATT CAGG AGGTACG AT AATT CTCCCTGTG CG GT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG GAGGCATTGCTTTTAGGTTCACCTGTAGGTAATTTAAATAGTATTGGGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATTAATT GT GAT GCGT ACTT ATTTT CCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTTCTTTAGGTTGGTCAG AG CAGACACACATATGCTTCAT AAAT CTGTGGGGCA CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAG CCACGTTTT CT GTGTGGTTT CCT G AG GCG AAG CGT AAG GT GTTG ATA CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGG GAATGTG AAAAT CT CT AG GAT GC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAGGCCTTAGTGTGGAAGAATGTTCAGTCCTTTGTGGAATCTATACGCTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGAAGGACTAGGTGATTGCGTCTATGATG CT CT AGTT CAAACCGG CT GGTTT GAT ACCT CT AG CG ACG AACT G AAAGTTTT G C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAGG GAT GT ACG AGG CAG ATGCAAAG AT CG AG AG AG AG AGTGTCT CT G AGTTGCT CGCTT CCGGT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AGAG AAATT CCAGG AATTTT GCAAGG AACT G AAT GTTAAT CCT ATGCT AATT GGC CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GAT ACAT GT G AAG AT AT G GAT GT AACT G AAG AT AT GG AG G AT AT AGT GTT GAT G GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTT ACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAATG AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTG GTT CT CCCAT AATT GAT AAG AG AAACT GG AAG CG ATTT G CTGTTT G C TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG G AAAG ACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGTATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGT GACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGT GATGCTAAGCAAAT CCCGTTTATAAACAGAGTCAT GAATTTT GATT AT CCTAAGGAGTTAAGAACTTTAATAGTCGATAATGTAGAGCGTAGGTATGTTACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CGACTCCGATAGGTTTAATTGTCCGTGATTCACCACATGTACTAGTGGCCTTAAC G AG G CACACTAAG G CAATG GTGTATTATACTGTTGTGTTCG ATG CAGTTACAAG TATAATAGCGGATGTGGAAAAGGTCGACCAGTCGATCTTGACTATGTTTGCTAC CACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT AAT ATTTTCCTCCCTGTTAGTAAAACGGGGTTTTATACAGACATGCAGGAGTTCTATG ATAGATGCCTTCCTGGGAATTCCTTCGTGCTGAATGATTTCGATGCCGTAACCAT GCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGTAA TTTAGATCCAGTACCCGCTTTGGTTAAGAGTGAAGCGCAGAATTTTCTGATTCCC GTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATCTT GT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCT GAT CT AG CT GG G ACT GTGG AT AT AACT AAT ATGT CG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG A CG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AG GG CT AGTT CCATT CAAAGTTTT TCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTCA ATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G CACATG ATTAAG CGG CAG CC CAAG AGT CGGTTG G AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT ATT GTTTATCACCCTAAAGTGGTAAATGCAGTTTTCGGTCCGGTTTTTAAGTATTTGA CCACCAAGTTT CTT AGCAT GGT AG AT AGTT CT AAGTTTTT CTTTT AC ACT AG G AAA AAACCAG AAG AT CT GCAG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT G AG ATT CTT GAG CTGG AT GTTT CT AAAT AT G ACAAGT CACAAT CCG ATTTCCATTT CTCTATTG AG ATG G CAATTTGG G AAAAATTGGG G CTGG ACG ATATTTTG G CTTG GATGTGGTCTATGGGT CACAAG AG AACT AT ACT G CAAG ATTT CCAAGCCG GG AT AAAG ACG CT CATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AG GT AATACCTTTATTATCGCAGCGTGTGTAGCTAGTATGTTGCCGTTAGACAAGTGTT TT AAAG CT AGTTTTT GTG GTG AT GATT CG CTG ATCT ACCTT CCT AAG GGTTT G G A GTATCCT GAT AT ACAGG CT ACT G CCAACTT G GTTT GG AATTTT G AGGCG AAACTT TT CCG AAAG AAGT ATGGTT ACTT CT GTGGG AAGTATAT AATT CACCAT GCCAACG G CTGT ATTGTTT ACCCT G ACCCTTT AAAATT AATT AGT AAATT AG GT AAT AAG AGT CTT GT AG G GTAT GAG CATGTT G AG G AGTTT CGTATATCTCTCCTCG ACGTCGCT CATAGTTTGTTTAATGGTGCTTATTTCCATTTACTCGACGATGCAATCCACGAATT ATTT CCT AACG CTGG GG GTT G CAGTTTTGTAATT AATT GTTT GT GC AAGT ATTT G AGTGATAAGCGCCTTTTCCGTAGTCTTTATATAGATGTCTCTAAGTAAGGTGTCG GT CG AG AACT CATT G AAACCCG AG AAGTTT GTT AAAAT CT CTT G G GT CG AT AAGT TG CTCCCT AACT ATTTTT CCATT CTT AAGT ATTT AT CT AT AACT G ACTTT AG CGTA GTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTGTT GAT CTT ACAAAACACCTTT AT GT CACATTGTT G G GCGTTGTG GTTTCTG GTGTAT GGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACACA AGG ATG CATTCTGTTG CAG AG GG AACT AT ATG CAAATTTTCAG CTCCCG CCACC GTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGGAT G CCCTT CG CG AT CCTT G GTCTTT ATTT GT GAG ACT CT CT AATGTAG GG ATT AAAG AT G GTTT CCAT CCTTT G ACCTT AG AG GT CG CTT GTTT AGT CG CT AC AACT AACT C T ATT AT CAAAAAG G GT CTT AG AG CTT CTGTAGT CG AGT CTGTCGTCT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAGTT G AACCTTT CTTT GAT AAAG TTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCGC AGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTTG G ACT CT G CTT CT G AAG AGT CCAGTT CT GTTT CTTT CG AAG AT G GCTT ACAAT CCG AT CACACCT AG CAAACTT ATT G CGTTT AGTG CTT CTT AT GTT CCCGTCAGG ACTT T ACTT AATTTT CT AGTT G CTTCAC AAG GTACCG CCTTCCAG ACTCAAG CG GG AA GAG ATT CTTT CCG CG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AG AT ATT A ATT CT AGGTT CCCAAAT G CGG GTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG GCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCATTGA GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG T ACT GAT G ACG CAT CTACG G CCG CT AGG G CT G AAAT AG AT AATTT AAT AG AGTC T ATTT CT AAGGGTTTT GAT GTTT AT GAT AGGGCTT CATTT G AAGCCGCGTTTT CG GT AGT CTG GT CAG AGG CT ACCACCT CG AAAGCTT AG CTT CG AG G GT CTT CT GAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:53).

[0175] The cDNA genome sequence of the attenuated CGMMV strain ONAL-1 differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO: 18) at least in that the nucleotide at position 3334 of SEQ ID NO:53 is T.

[0176] The 129 kDa protein encoded by the attenuated CGMMV strain ONAL-1 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEI IARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTK (SEQ ID NO:54).

[0177] The 129 kDa protein encoded by the attenuated CGMMV strain ONAL-1 differs from the 129 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:63) at least in that position 1092 of SEQ ID NO:54 is valine (V, Val).

[0178] The 186 kDa protein encoded by the attenuated CGMMV strain ONAL-1 has the following sequence. MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRNIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHPKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKPEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKRLFRSLYIDVSK (SEQ ID

NO:55).

[0179] The 186 kDa protein encoded by the attenuated CGMMV strain ONAL-1 differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that position 1092 of SEQ ID NO:55 is valine (V, Val).

Mutant CGMMV Ontario strain ONAL-2

[0180] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above, using the mutagenic primers listed in Table 6 to introduce the mutation c.4969G>A. Nucleotide residues indicated in bold indicate sites of mutation. This mutation resulted in an R1637H amino acid substitution in the encoded viral 186 kDa protein. The resulting mutant CGMMV strain was designated Ontario strain ONAL-2.

Table 6: Primers used to produce mutant CGMMV Ontario strain ONAL-2

[0181] The cDNA genome sequence of CGMMV strain ONAL-2 is shown below. GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAATGG CAAACATTAATG AACAAATCAACAACCAACG CG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACTG ATG CGTATCCG G AGTTTTCG A TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT GCGGGTGGTCT G AG GC TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATG GT CACACAACATGTACAG AG G CACAAGG G ATCTG G CG G GTG CAGACCTC TT CCAACTTTT CAG ATAG ATGCATT CAGG AGGTACG AT AATT CT CCCTGT GCGGT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG GAGGCATTGCTTTTAGGTTCACCTGTAGGTAATTTAAATAGTATTGGGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATTAATT GT GAT GCGT ACTT ATTTT CCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AG G TTG GT CAG AG CAG AC AC AC AT ATG CTT CAT AAAT CTGTGGGG C A CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAGCCACGTTTT CT GTGTGGTTT CCT GAGGCG AAGCGT AAGGT GTT GAT A CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGG GAATGTG AAAAT CT CT AG GAT GC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAGGCCTTAGTGTGGAAGAATGTTCAGTCCTTTGTGGAATCTATACGCTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT T CG CAG ACAGT CTT AAAT CCG CCGTAG AAGG ACT AGGTG ATT G CGTCTATG ATG CT CT AGTT CAAACCGG CT GGTTT GAT ACCT CT AG CG ACG AACT G AAAGTTTT G C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAG GG AT GT ACG AG GCAG ATGCAAAG AT CG AG AG AG AG AGTGTCT CT G AGTTGCT CGCTT CCGGT G ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GAT ACAT GT G AAG AT AT G GAT GT AACT G AAG AT AT GG AG G AT AT AGT GTT GAT G GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTT ACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG GAAAG ACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGTATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTGACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGT GATGCT AAG CAAAT CCCGTTTATAAACAGAGTCAT G AATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CGACTCCGATAGGTTTAATTGCCCGTGATTCACCACATGTACTAGTGGCCTTAA CGAGGCACACTAAGGCAATGGTGTATTATACTGTTGTGTTCGATGCAGTTACAA GTATAATAGCGGATGTGGAAAAGGTCGACCAGTCGATCTTGACTATGTTTGCTA CCACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT AA T ATTTT CCTCCCTGTT AGTAAAACG GG GTTTT AT ACAG ACAT G CAG G AGTT CTAT GAT AG AT G CCTT CCT GG G AATT CCTT CGTG CT G AAT G ATTT CG AT GCCGT AACC ATGCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGT AATTT AG AT CCAGT ACCCG CTTT G GTT AAG AGTG AAG CGCAG AATTTT CT GATT C CCGTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATC TTGT AG CT AT GAT AAAG AGG AAT AT G AAT ACT CCT GAT CTAG CTGG G ACT GTGG AT AT AACT AAT AT GTCG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG ACG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AGG G CT AGTT CCATT CAAAGTTT TTCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTC AATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G C AC AT GATT AAG C G G CAG C CCAAG AGT CGGTTGG AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT A TTGTTTATCACCCTAAAGTGGTAAATGCAGTTTTCGGTCCGGTTTTTAAGTATTT G ACCACCAAGTTT CTTAGCAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AGG AAAAAACCAG AAG AT CT G CAG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT GAG ATT CTT GAG CT G G ATGTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCA TTTCTCTATTGAGATGGCAATTTGGGAAAAATTGGGGCTGGACGATATTTTGGCT TGGATGTGGTCTATGGGTCACAAGAGAACTATACTGCAAGATTTCCAAGCCGGG AT AAAG ACG CT CATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AG GT AAT ACCTTT ATT AT CG CAG CGTGT GT AGCT AGT ATGTTG CCGTT AG ACAAGT G TTTT AAAG CT AGTTTTTGTG GT GAT GATT CG CT GAT CT ACCTT CCT AAGG GTTT G G AGTATCCTG AT AT ACAG GCTACTG CCAACTTG GTTTG G AATTTTG AG G CG AAA CTTTT CCG AAAG AAGTAT G GTT ACTT CT GTGG G AAGT AT AT AATT CACCAT G CCA ACGG CTGTATT GTTT ACCCT G ACCCTTT AAAATT AATT AGT AAATT AG GT AAT AAG AGTCTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTC G CT CAT AGTTT GTTT AAT GGT GCTT ATTT CCATTT ACT CG ACG AT G CAAT CCACG AATT ATTT CCT AACG CT G GG G GTT GCAGTTTT GT AATT AATT GTTT GTG CAAG T A TTT G AGTG AT AAG CACCTTTT CCGT AGT CTTT AT AT AG AT GTCTCT AAGT AAGGT GTCGGTCGAGAACTCATTGAAACCCGAGAAGTTTGTTAAAATCTCTTGGGTCGA T AAGTT G CTCCCT AACT ATTTTT CCATT CTT AAGT ATTT AT CT AT AACT G ACTTT AG CGTAGTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGG TGTTGATCTTACAAAACACCTTTATGTCACATTGTTGGGCGTTGTGGTTTCTGGT GTATGGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGA CACAAG G AT GC ATT CTGTTG CAG AGG G AACT AT AT G CAAATTTT CAGCT CCCG C CACCGTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGC GGATGCCCTTCGCGATCCTTGGTCTTTATTTGTGAGACTCTCTAATGTAGGGATT AAAG ATGGTTT CCAT CCTTT GACCTT AGAGGT CGCTTGTTT AGT CGCTACAACTA ACTCTATTATCAAAAAGGGTCTTAGAGCTTCTGTAGTCGAGTCTGTCGTCTCTTC CG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAG TT G AACCTTT CTTT GAT AAAGTTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGG TCGCAGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAG GTT G G ACT CT G CTT CT G AAG AGT CCAGTT CTGTTT CTTT CG AAG AT GG CTT ACAA TCCGATCACACCTAGCAAACTTATTGCGTTTAGTGCTTCTTATGTTCCCGTCAGG ACTTT ACTT AATTTT CT AGTT GCTT CACAAG GT ACCG CCTT CCAG ACT CAAG CG G G AAG AG ATT CTTT CCG CG AGT CCCTGTCTGCGTTACCCTCGTCTGTCGT AG AT A TT AATT CTAGGTT CCCAAATGCGGGTTTTTACGCTTT CCT CAACGGTCCT GT GTT GAG G CCTATCTTCGTTTCG CTTCTTAG CTCTACG G ATACG CGTAATAG GGTCAT

TGAGGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAA

GCGTACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGA

GTCTATTTCTAAGGGTTTTGATGTTTATGATAGGGCTTCATTTGAAGCCGCGTTT

TCGGTAGTCTGGTCAGAGGCTACCACCTCGAAAGCTTAGCTTCGAGGGTCTTCT

GAT GGTGGT GCACACCAAAGTGCATAGT GCTTT CCCGTT CACTT AAAT CGAACG

GTTTGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGG

GCAGTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA

(SEQ ID NO: 58).

[0182] The cDNA genome sequence of the attenuated CGMMV strain ONAL-2 differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO:18) at least in that the nucleotide at position 4969 of SEQ ID NO:58 is A.

[0183] The 186 kDa protein encoded by the attenuated CGMMV strain ONAL-2 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGGLRLLELEYMMMQVPYGSPCYDIGGN

YTQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQ

IDAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVEGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVSELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIARDSPHVLVALTRHTKAMVYYTWFDAVTSIIADVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRNIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHPKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKPEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKHLFRSLYIDVSK (SEQ ID

NO:59).

[0184] The 186 kDa protein encoded by the attenuated CGMMV strain ONAL-2 differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that position 1637 of SEQ ID NO:59 is histidine (H, His). Mutant CGMMV Ontario strain ONBM-32

[0185] Directed mutation of the cDNA genome of the cloned CGMMV Ontario strain (Example 1) was carried out as described above to introduce mutations

corresponding to those induced in the cDNA genome of the cloned CGMMV Ontario strain mutants ONBM, ONAL-1 and ONAL-2 (c.315G>A; c.1498A>G; C.1660OT; C.33340T C.3430OT; c.3528A>G; c.4144C>T; C.42480T; c.4969G>A; and c.6228C>T). These mutations resulted in amino acid substitutions in the encoded viral proteins (G86S, E480G, S534F, A1092V and A1 124V in the 129 kDa protein; G86S, E480G, S534F, A1092V, A1 124V, N1 157D, P1362L, P1397S and R1637H in the 186 kDa protein; and A156V in the coat protein). The resulting mutant CGMMV strain was designated Ontario strain ONBM-32.

[0186] The cDNA genome sequence of CGMMV strain ONBM-32 is shown below.

GTTTT AATTTTT AAAATT AAACAAACAACAACAACAACAACAAACAATTT AAAACA ACAATGG CAAACATTAATG AACAAATCAACAACCAACG CG ACG CCG CG G CCAG CGGGAGAAACAATCTCGTTAGCCAATTGGCGTCAAAAAGGGTGTATGACGAGG CTGTTCG CTCGTTGG AT CAT CAAG ACAG ACGCCCAAAAAT G AACTTTT CTCGTG TGGTCAG CACAG AG CACACCAG G CTTGTAACTG ATG CGTATCCG G AGTTTTCG A TTAG CTTT ACCG CCACCAAG AACT CT GTACACT CCCTT G CG GGT AGTCT GAG G C TCCTTGAACTGGAATATATGATGATGCAAGTGCCCTACGGCTCACCTTGTTATGA TATCGGCGGTAACTATACGCAGCACTTGTTCAAAGGTAGATCATATGTGCATTG CTGCAATCCGTGCCTGGATCTTAAAGATGTTGCGAGGAACGTGATGTATAACGA TATG GT CACACAACATGTACAG AG G CACAAGG G ATCTG G CG G GTG CAGACCTC TT CCAACTTTT CAG ATAG ATGCATT CAGG AGGTACG AT AATT CT CCCTGT GCGGT CACCTGTTCAGACGTTTTCCAAGAGTGTTCCTATGATTTTGGGAGCGGTAGGGA T AAT CAT G CAGT CT CG CT G CATT CAAT CT ACG AT AT CCCTT ATT CTT CG ATCGG A CCTG CT CTT CAT AG G AAG AACGT G CG AGTTTGTT AT G CAG CCTTT CACTT CTCG GAGGCATTGCTTTTAGGTTCACCTGTAGGTAATTTAAATAGTATTGGGGCTCAGT TTAGGGTCGATGGTGATGATGTGCATTTTCTTTTTAGTGAAGAGTCTACTTTGCA TT AT ACT CAT AGTTT AG AAAAT AT CAAATTAATT GT GAT GCGT ACTT ATTTT CCT G CT G ATG AT AGGT ACGT GTAT ATT AAGG AGTTT ATG GT CAAG CGT GTGG AT ACTTT CTT CTTT AG G TTG GT CAG AG CAG AC AC AC AT ATG CTT CAT AAAT CTGTGGGG C A CT ATT CAAAAT CGAAAT CT G AGTACTTTGCGCT G AATACCCCT CCGAT CTT CCAA G ACAAAGCCACGTTTT CT GTGTGGTTT CCT GAGGCG AAGCGT AAGGT GTT GAT A CCCAAGTTT G AACTTT CAAG ATT CCTTT CTGG GAATGTG AAAAT CT CT AG GAT GC TTGTCG ATG CT G ATTT CGT CCAT ACCATT ATT AAT CACATT AG CACGT AT GAT AAT AAGGCCTTAGTGTGGAAGAATGTTCAGTCCTTTGTGGAATCTATACGCTCAAGA GTAATTGTAAACGGAGTTTCGGTGAAATCTGAATGGAACGTACCGGTTGATCAG CT CACT GAT AT CTCGTTCTCG AT ATT CCTT CT CGTG AAG GTT AGG AAG GTACAG A TCGAGTTAATGTCTGATAAAGTTGTAATCGAGGCGAGGGGCTTGCTCCGGAGGT TCGCAGACAGTCTTAAATCCGCCGTAGGAGGACTAGGTGATTGCGTCTATGATG CT CT AGTT CAAACCGG CT GGTTT GAT ACCT CT AG CG ACG AACT G AAAGTTTT G C T ACCT G AACCGTTT AT G ACCTTTT CG G ATT AT CTT G AAG GG AT GT ACG AG GCAG AT G CAAAG AT CG AG AG AG AG AGTGTCTTT G AGTT G CT CG CTT CCG GTG ACG ATT TGTT CAAG AAAAT CG AT GAG AT AAG AAACAATT ACAGT G G AGTCG AATTT G ATGT AG AG AAATT CCAG G AATTTT G CAAGG AACT G AAT GTT AAT CCT AT GCT AATT GG C CATGTTATCG AAG CTATTTTTTCG CAG AAAGCTG G GGTG ACAGTAACGG GTCTG GGTACCCTCTCTCCTGAGATGGGTGCTTCTGTTGCGTTATCCAATACCTCTGTA GAT ACAT GT G AAG AT AT G GAT GT AACT G AAG AT AT GG AG G AT AT AGT GTT GAT G GCGG ACAAG AGT CATT CTT ACAT GT CCCCAGAAAT GGCG AG AT GGGCT G ATGTA AAATACGACAACAATAAAGGGGGCCTGGTCGAATACAAAGTCGGAACCTCGATG ACTTTACCTGCCACCTGGGCAGAGAAGGGTAAGGCTGTCTTACCGTTGTCGGG GAT CT GT GT G AGG AAACCCCAATTTT CG AAGCCGCTT GAT GAGG AAG ACG ACTT G AGGTTAT CAAACAT GAATTT CTTT AAGGT G AGCG AT CT G AAGTT G AAG AAAACT AT CACT CCAGTT GTTT ACACT G G G ACCATT CG AG AG AGG CAAAT G AAG AATT AT ATTGATTACTTATCGGCCTCTCTTGGTTCTACGCTGGGTAATCTGGAGAGAATTG TGCGGAGTGATTGGAACGGTACCGAGGAGAGTATGCAAACGTTCGGGTTGTAT GACTGCGAAAAGTGCAAGTGGTTACTGTTACCAGCCGAAAAGAAGCACGCATG GGCTGTGGTTCTGGCAAGTGATGATACCACTCGCATAATCTTCCTCTCATATGA CG AAT CTGGTT CT CCCAT AATT GAT AAG AG AAACTGGAAGCG ATTT GCT GTTTGC TCTGAGACCAAAGTCTATAGCGTAATTCGTAGTTTAGAGGTACTAAATAAGGAAG CAATAGTCGACCCCGGGGTTCATATAACATTAGTTGACGGAGTGCCGGGTTGTG G AAAG ACCG CCG AAATT AT AG CG AG GGTCAATT G G AAAACCG AT CT AGTATT G A CTCCCGGGAGGGAGGCGGCTGCTATGATTAGGCGGAGGGCCTGCGCCCTGCA CAAGT CACCT GT GGCAACCAGTG ACAACGTTAG AACTTT CG ATT CTTTT GT GAT G AAT AAG AAAAT CTT CAAGTTT G ACGCT GT CT AT GTT G ACG AGGGT CT GAT GGTC CATACGGGTTTACTTAATTTTGCGTTGAAGATCTCAGGTTGTAAAAAGGCCTTCG T CTTT GGT GATGCT AAG CAAAT CCCGTTTATAAACAGAGTCAT GAATTTT GATT AT CCT AAGG AGTT AAG AACTTT AAT AGT CG AT AAT GT AG AG CGT AGGTAT GTT ACCC ATAGGTGTCCTAGAGATGTCACTAGTTTTCTTAATACTATTTACAAAGCCGCTGT CGCTACTACTAGTCCGGTTGTACATTCTGTGAAGGCGATTAAAGTGTCAGGGGC CG GT ATT CT G AGG CCCG AGTT G ACG AAG AT CAAAG G AAAG AT AAT AACGTTT AC T CAAT CT GAT AAGCAGT CCTT GAT CAAG AGTGGGTACAAT G ACGT G AACACTGT GCATGAAATTCAGGGAGAAACCTTTGAAGAGACGGCGGTTGTGCGTGCCACCC CG ACT CCG AT AG GTTT AATT GTCCGT GATT CACCACAT GT ACT AGTG GCCTT AAC GAG G CACACT AAG G CAAT G GTGT ATT AT ACTGTT GTGTT CG AT GC AGTT ACAAG T AT AAT AGT GG ATGT GG AAAAG GT CG ACCAGT CG AT CTT G ACT AT GTTT G CT AC CACTGTG CCT ACCAAAT AG CAATT AAT G CAG AACT CACT GTATGT CCAT CGT GAT ATTTTCCTCCCTGTTAGTAAAACGGGGTTTTATACAGACATGCAGGAGTTCTATG ATAGATGCCTTCCTGGGAATTCCTTCGTGCTGAATGATTTCGATGCCGTAACCAT GCGGTTGAGGGACAACGAATTTAACCTACAACCTTGTAGGCTAACCTTAAGTAA TTTAGATCCAGTACCCGCTTTGGTTAAGAGTGAAGCGCAGAATTTTCTGATTCCC GTTTTGCGTACGGCCTGTGAAAGGCCGCGCATTCCAGGTCTCCTTGAAAATCTT GTAGCTATGATAAAGAGGAATATGAATACTCCTGATCTAGCTGGGACTGTGGAT AT AACT AAT ATGT CG ATTT CT AT AGT AG AT AACTT CTTTT CTT CTTTT GTT AG AG A CG AG GTTTT G CTT GAT CATTT AG ATT GTGTT AG GG CT AGTT CCATT CAAAGTTTT TCTGATTGGTTTTCGTGTCAGCCAACCTCGGCGGTTGGTCAATTAGCTAATTTCA ATTT CAT AG ATTT G CCT G CCTTT GAT ACTT AT AT G CACATG ATTAAGCG G CAG CC CAAG AGT CGGTTG G AT ACTT CG ATT CAGT CT G AAT AT CCG G CCTT G CAAACT ATT GTTTATCACCTTAAAGTG GTAAATG CAGTTTT CG GT CCG GTTTTT AAGT ATTT G A CCACCAAGTTT CTT AGCAT GGT AG AT AGTT CT AAGTTTTT CTTTT ACACT AG G AAA AAAT CAG AAG AT CT GCAG G AATTTTT CT CAG AT CT CT CTT CCCATT CT GATT AT G AG ATT CTT GAG CTGG AT GTTT CT AAAT AT G ACAAGT CACAAT CCG ATTT CCATTT CTCTATTG AG ATG G CAATTTGG G AAAAATTGGG G CTGG ACG ATATTTTG G CTTG GAT GT GGT CT ATGGGT CACAAG AGAACT AT ACTGCAAG ATTT CCAAGCCGGG AT AAAG ACG CT CATTT ACT AT CAACG G AAGT CTGGTG ATGT AACT ACTTT CAT AG GT AATACCTTTATTATCGCAGCGTGTGTAGCTAGTATGTTGCCGTTAGACAAGTGTT TT AAAGCT AGTTTTT GT GGTG AT GATT CGCT GAT CT ACCTT CCT AAGGGTTTGGA GTATCCT GAT AT ACAGG CT ACT G CCAACTT G GTTT GG AATTTT G AGG CG AAACTT TT CCG AAAG AAGT ATGGTT ACTT CT GTGGG AAGTATAT AATT CACCAT GCCAACG G CTGT ATTGTTT ACCCT G ACCCTTT AAAATT AATT AGT AAATT AG GT AAT AAG AGT CTTGTAGGGTATGAGCATGTTGAGGAGTTTCGTATATCTCTCCTCGACGTCGCT CATAGTTTGTTTAATGGTGCTTATTTCCATTTACTCGACGATGCAATCCACGAATT ATTTCCTAACG CTGG GG GTTG CAGTTTT GT AATT AATT GTTTGTG CAAGT ATTT G AGTGATAAGCACCTTTTCCGTAGTCTTTATATAGATGTCTCTAAGTAAGGTGTCG GT CG AG AACT CATT G AAACCCG AG AAGTTT GTT AAAAT CT CTT G G GT CG AT AAGT TG CTCCCT AACT ATTTTT CCATT CTT AAGTATTT AT CT AT AACT G ACTTT AG CGTA GTTAAAGCTCAGAGCTATGAATCCCTCGTGCCTGTCAAGTTGTTGCGTGGTGTT GAT CTT ACAAAAC ACCTTT ATGTC ACATTGTT GG G CGTT GT G GTTT CTG GTGTAT GGAACGTACCGGAATCCTGTAGGGGTGGTGCTACTGTTGCTCTGGTTGACACA AGG AT G CATT CTGTTG CAG AG GG AACTATATG CAAATTTTCAG CTCCCG CCACC GTCCGCGAATTCTCTGTTAGGTTCATACCTAACTATTCTGTCGTGGCTGCGGAT G CCCTT CG CG AT CCTT G GTCTTT ATTT GT GAG ACT CT CT AATGTAG GG ATT AAAG AT G GTTT CCAT CCTTT G ACCTT AG AG GT CG CTT GTTT AGT CG CT AC AACT AACT C T ATT AT CAAAAAG G GT CTT AG AG CTT CTGTAGT CG AGT CTGTCGTCT CTT CCG AT CAGT CCATT GTCCT AG ATT CTTT AT CCG AG AAAGTT G AACCTTT CTTT GAT AAAG TTCCTATTTCGGCGGCTGTGATGGCAAGAGACCCCAGTTATAGGTCTAGGTCGC AGTCTGTCGGTGGTCGTGGTAAGCGGCATTCTAAACCTCCAAATCGGAGGTTG G ACT CT G CTT CT G AAG AGT CCAGTT CT GTTT CTTT CG AAG AT G GCTT ACAAT CCG AT CACACCT AG CAAACTT ATT G CGTTT AGTG CTT CTT AT GTT CCCGTCAGG ACTT T ACTT AATTTT CT AGTT G CTT CAC AAGGTACCG CCTT CCAG ACT CAAG CGGG AA GAG ATT CTTT CCG CG AGTCCCT GTCTGCGTTACCCTCGTCTGTCGT AG AT ATT A ATT CT AGGTT CCCAAAT G CG GGTTTTT ACG CTTT CCT CAACG GT CCTGTGTT GAG GCCTATCTTCGTTTCGCTTCTTAGCTCTACGGATACGCGTAATAGGGTCATTGA GGTTGTAGATCCTAGCAATCCTACGACTGCTGAGTCGCTTAACGCTGTAAAGCG TACTGATGACGCATCTACGGCCGCTAGGGCTGAAATAGATAATTTAATAGAGTC TATTTCTAAGGGTTTTGATGTTTATGATAGGGCTTCATTTGAAGCCGCGTTTTCG GT AGT CTGGT CAG AGGTT ACCACCT CG AAAGCTTAGCTT CG AGGGTCTT CT GAT GGTGGTGCACACCAAAGTGCATAGTGCTTTCCCGTTCACTTAAATCGAACGGTT TGCTCATTGGTTTGCGGAAACCTCTCACGTGTGGCGTTGAAGTTTCTATGGGCA GTAATTCTGCAAGGGGTTCGAATCCCCCCTTTCCCCGGGTAGGGGCCCA (SEQ ID NO:60).

[0187] The cDNA genome sequence of the attenuated CGMMV strain ONBM-32 differs from the cDNA genome sequence of the wild type CGMMV Ontario strain (SEQ ID NO: 18) at least in that:

the nucleotide at position 315 of SEQ ID NO:60 is A;

the nucleotide at position 1498 of SEQ ID NO:60 is G;

the nucleotide at position 1660 of SEQ ID NO:60 is T;

the nucleotide at position 3334 of SEQ ID NO:60 is T;

the nucleotide at position 3430 of SEQ ID NO:60 is T;

the nucleotide at position 3528 of SEQ ID NO:60 is G;

the nucleotide at position 4144 of SEQ ID NO:60 is T;

the nucleotide at position 4248 of SEQ ID NO:60 is T;

the nucleotide at position 4969 of SEQ ID NO:60 is A; and

the nucleotide at position 6228 of SEQ ID NO:60 is T.

[0188] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-32 has the following sequence. MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK

QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTK (SEQ ID N0:61)

[0189] The 129 kDa protein encoded by the attenuated CGMMV strain ONBM-32 differs from the 129 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:63) at least in that:

position 86 of SEQ ID NO:61 is serine (S, Ser);

position 480 of SEQ ID NO:61 is glycine (G, Gly);

position 534 of SEQ ID NO:61 is phenylalanine (F, Phe);

position 1092 of SEQ ID NO:61 is valine (V, Val); and

position 1 124 of SEQ ID NO:61 is valine (V, Val).

[0190] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-32 has the following sequence.

MANINEQINNQRDAAASGRNNLVSQLASKRVYDEAVRSLDHQDRRPKMNFSRWS

TEHTRLVTDAYPEFSISFTATKNSVHSLAGSLRLLELEYMMMQVPYGSPCYDIGGNY

TQHLFKGRSYVHCCNPCLDLKDVARNVMYNDMVTQHVQRHKGSGGCRPLPTFQI

DAFRRYDNSPCAVTCSDVFQECSYDFGSGRDNHAVSLHSIYDIPYSSIGPALHRKN

VRVCYAAFHFSEALLLGSPVGNLNSIGAQFRVDGDDVHFLFSEESTLHYTHSLENIK

LIVMRTYFPADDRYVYIKEFMVKRVDTFFFRLVRADTHMLHKSVGHYSKSKSEYFAL

NTPPIFQDKATFSVWFPEAKRKVLIPKFELSRFLSGNVKISRMLVDADFVHTIINHI ST

YDNKALVWKNVQSFVESIRSRVIVNGVSVKSEWNVPVDQLTDISFSIFLLVKVRKVQ I

ELMSDKWIEARGLLRRFADSLKSAVGGLGDCVYDALVQTGWFDTSSDELKVLLPE

PFMTFSDYLEGMYEADAKIERESVFELLASGDDLFKKIDEIRNNYSGVEFDVEKFQE

FCKELNVNPMLIGHVIEAIFSQKAGVTVTGLGTLSPEMGASVALSNTSVDTCEDMDV

TEDMEDIVLMADKSHSYMSPEMARWADVKYDNNKGGLVEYKVGTSMTLPATWAE

KGKAVLPLSGICVRKPQFSKPLDEEDDLRLSNMNFFKVSDLKLKKTITPWYTGTIRE

RQMKNYIDYLSASLGSTLGNLERIVRSDWNGTEESMQTFGLYDCEKCKWLLLPAEK

KHAWAWLASDDTTRIIFLSYDESGSPIIDKRNWKRFAVCSETKVYSVIRSLEVLNKE

AIVDPGVHITLVDGVPGCGKTAEIIARVNWKTDLVLTPGREAAAMIRRRACALHKSP

VATSDNVRTFDSFVMNKKIFKFDAVYVDEGLMVHTGLLNFALKISGCKKAFVFGDAK QIPFINRVMNFDYPKELRTLIVDNVERRYVTHRCPRDVTSFLNTIYKAAVATTSPWH

SVKAIKVSGAGILRPELTKIKGKIITFTQSDKQSLIKSGYNDVNTVHEIQGETFEET AW

RATPTPIGLIVRDSPHVLVALTRHTKAMVYYTWFDAVTSIIVDVEKVDQSILTMFAT T

VPTKXQLMQNSLYVHRDIFLPVSKTGFYTDMQEFYDRCLPGNSFVLNDFDAVTMRL

RDNEFNLQPCRLTLSNLDPVPALVKSEAQNFLIPVLRTACERPRIPGLLENLVAMIK R

NMNTPDLAGTVDITNMSISIVDNFFSSFVRDEVLLDHLDCVRASSIQSFSDWFSCQP

TSAVGQLANFNFIDLPAFDTYMHMIKRQPKSRLDTSIQSEYPALQTIVYHLKWNAVF

GPVFKYLTTKFLSMVDSSKFFFYTRKKSEDLQEFFSDLSSHSDYEILELDVSKYDKS

QSDFHFSIEMAIWEKLGLDDILAWMWSMGHKRTILQDFQAGIKTLIYYQRKSGDVTT

FIGNTFIIAACVASMLPLDKCFKASFCGDDSLIYLPKGLEYPDIQATANLVWNFEAK LF

RKKYGYFCGKYIIHHANGCIVYPDPLKLISKLGNKSLVGYEHVEEFRISLLDVAHSL FN

GAYFHLLDDAIHELFPNAGGCSFVINCLCKYLSDKHLFRSLYIDVSK (SEQ ID

NO:62).

[0191] The 186 kDa protein encoded by the attenuated CGMMV strain ONBM-32 differs from the 186 kDa protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:64) at least in that:

position 86 of SEQ ID NO:62 is serine (S, Ser);

position 480 of SEQ ID NO:62 is glycine (G, Gly);

position 534 of SEQ ID NO:62 is phenylalanine (F, Phe);

position 1092 of SEQ ID NO:62 is valine (V, Val);

position 1 124 of SEQ ID NO:62 is valine (V, Val);

position 1 157 of SEQ ID NO:62 is aspartic acid (D, Asp);

position 1362 of SEQ ID NO:62 is leucine (L, Leu);

position 1397 of SEQ ID NO:62 is serine (S, Ser); and

position 1637 of SEQ ID NO:62 is histidine (H, His).

[0192] The coat protein encoded by the attenuated CGMMV strain ONBM-32 has the sequence of SEQ ID NO:32 and differs from the coat protein encoded by the wild type CGMMV Ontario strain (SEQ ID NO:65) at least in that position 156 of SEQ ID NO:32 is valine (V, Val).

Example 3: Inoculation of cucumber plants with the attenuated CGMMV ONBM, ONBM-2 and ONBM-3 strains

[0193] The attenuated CGMMV Ontario strains ONBM, ONBM-2 and ONBM-3 were transformed into Agrobacterium tumefaciens strain EHA105 by electroporation and used to inoculate the cotyledon of 1-2 week old cucumber plants as described in Example 1 . Two weeks after inoculation, plants were inspected for visible symptoms of CGMMV infection and leaf tissue was collected for detection of virus infection. The attenuated CGMMV Ontario strains ONBM, ONBM-2 and ONBM-3 were detected in leaf tissue by real-time TaqMan reverse-transcription PCR (Chen et al., Journal of Virological Methods (2008), 149: 326-329). As seen from the results shown in Figures 3A to 3E, no symptoms were induced by inoculation with ONBM (Figure 3C), ONBM-2 (Figure 3D) or ONBM-3 (Figure 3E) while the typical green mottle and mosaic symptoms were induced by inoculation with wild-type CGMMV under laboratory greenhouse conditions (Figure 3A). Figure 3B shows leaves of a healthy cucumber plant grown as a control.

[0194] The attenuated CGMMV Ontario strains ONBM, ONBM-2 and ONBM-3 were tested for the protection of cucumber plants from infection by wild-type CGMMV. Seven day old cucumber seedlings were inoculated with CGMMV Ontario strains ONBM, ONBM-2 or ONBM-3 or were not inoculated, under laboratory greenhouse conditions as described in Example 1. Two weeks after inoculation, the plants were challenged with wild-type CGMMV Ontario strain. As seen from the results shown in Figure 4A to 4E, no symptoms were observed 4 weeks after challenge with the wild- type CGMMV on plants inoculated with ONBM-2 (Figure 4B) or ONBM-3 (Figure 4C), and very mild or undetectable symptoms were observed 4 weeks after challenge with the wild-type CGMMV on plants inoculated with ONBM (Figure 4A). In contrast, the typical green mottle and mosaic symptoms were observed in uninoculated plants challenged with wild-type CGMMV (Figure 4D). Figure 4E shows leaves of an uninoculated and unchallenged healthy cucumber plant (control).

[0195] The attenuated CGMMV Ontario strains ONBM-2 and ONBM-3 were tested for the protection of cucumber plants from natural infection by wild-type CGMMV under commercial greenhouse conditions. Cotyledons of 1-2 week old cucumber plants were inoculated with the strains ONBM-2 and ONBM-3 as described in Example 1. Uninoculated cucumber plants were used as a control. As seen from the results presented in Figures 5A to 5E, no visible symptoms were observed on the cucumber plants after inoculation with ONBM-2 or ONBM-3 for a period of 100 days (until the end of production) in a commercial greenhouse. In contrast, uninoculated plants showed symptoms of CGMMV infection of leaves, showing green mottle and mosaic symptoms (Figure 5A), and CGMMV infection of fruits (Figure 5B and Figure 5C). The cucumber fruits produced from the cucumber plants inoculated with the attenuated strains ONBM-2 (Figure 5D) or ONBM-3 (Figure 5E) were healthy, while the uninoculated cucumber plants produced diseased and unmarketable cucumber fruits showing mosaic and soft (Figure 5B) or curling (Figure 5C) symptoms. In addition, over a harvest period of two months, the cucumber fruit yield was increased by 12.7% during the second month of harvest for plants treated with the attenuated CGMMV ONBM-3 strain compared with untreated cucumber plants exposed to natural CGMMV infection in commercial greenhouse conditions. Example 4: Inoculation of cucumber plants with the attenuated CGMMV ONAL- 1 , ONAL-2 and ONBM-32 strains

[0196] The attenuated CGMMV Ontario strains ONAL-1 , ONAL-2, and ONBM-32 were transformed into Agrobacterium tumefaciens strain EHA105 by electroporation. Agrobacterium tumefaciens containing mutant CGMMV strains ONAL-1 , ONAL-2, ONBM-32 and ONB (Example 2) were used to inoculate the cotyledon of 1 -2 week old cucumber plants using the method described in Example 1. Two weeks after inoculation, plants were inspected for visible symptoms of CGMMV infection and leaf tissue was collected for detection of virus infection. The attenuated CGMMV Ontario strains ONAL-1 , ONAL-2, ONBM-32 and ONB were detected in leaf tissue by realtime TaqMan reverse-transcription PCR (Chen et al., Journal of Virological Methods (2008), 149: 326-329).

[0197] As seen from the results shown in Figures 6A to 6F, plants inoculated with either mutant strains ONAL-1 (Figure 6A) or ONAL-2 (Figure 6B) under laboratory greenhouse conditions showed mild symptoms compared with plants inoculated with the wild-type CGMMV Ontario strain, which showed typical green mottle and mosaic symptoms (Figure 6F). No symptoms were observed after inoculation with the mutant strain ONBM-32 (Figure 6C). The symptoms observed after inoculation with mutant strain ONAL-2 (Figure 6B) were milder than those observed after inoculation with mutant strain ONAL-1 (Figure 6A), and milder than those observed after inoculation with mutant strain ONB (Figure 6D). Figure 1 E shows leaves of a healthy cucumber plant grown as a control.

[0198] The embodiments described herein are intended to be illustrative of the present compositions and methods and are not intended to limit the scope of the present invention. Various modifications and changes consistent with the description as a whole and which are readily apparent to the person of skill in the art are intended to be included. The appended claims should not be limited by the specific embodiments set forth in the examples, but should be given the broadest interpretation consistent with the description as a whole.