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Over the past decades, several studies have examined the subcellular localization of the cauliflower mosaic virus (CaMV) P6 protein by tagging it with GFP (P6-GFP). These investigations have been essential in the development of models for inclusion body formation, nuclear transport, and microfilament-associated intracellular movement of P6 inclusion bodies for delivery of virions to plasmodesmata. Although it was shown early on that the translational transactivation function of P6-GFP was comparable to wild type P6, it has not been possible to incorporate a P6-GFP gene into an infectious clone of CaMV. Consequently, it has not been possible to formally prove that a P6-GFP fusion is comparable in function to the unmodified P6 protein. Here we show that transient expression of P6-GFP can complement a defective CaMV replicon through gene expression, replication and encapsidation, which validates the relevance of P6-GFP subcellular localization studies for understanding the development of CaMV infections. • CaMV P6-GFP fusions have been integral for elucidating the subcellular location of the multifunctional P6 protein. • Separate delivery of either P6 or P6-GFP can complement a defective CaMV isolate through replication and virion formation. • This work validates the previously published subcellular localization studies conducted with P6-GFP. [ABSTRACT FROM AUTHOR]

Cauliflower mosaic virus gene VI product (P6) is an essential protein that forms cytoplasmic, inclusion bodies (IBs). P6 contains four regions involved in self-association, termed D1–D4. D3 binds to D1, along with D4 and contains a spacer region (termed D3b) between two RNA-binding domains. Here we show D3b binds full-length P6 along with D1 and D4. Full-length P6s harboring single amino acid substitutions within D3b showed reduced binding to both D1 and D4. Full-length P6s containing D3b mutations and fused with green fluorescent protein formed inclusion-like bodies (IL-Bs) when expressed in Nicotiana benthamiana leaves. However, mutant P6s with reduced binding to D1 and D4, showed smaller IL-Bs, than wild type. Likewise, viruses containing these mutations showed a decrease in inoculated leaf viral DNA levels and reduced efficiency of systemic infection. These data suggest that mutations influencing P6 self-association alter IB formation and reduce virus infection. [ABSTRACT FROM AUTHOR]

The Panel on Plant Health performed a pest categorisation of Strawberry vein banding virus (SVBV) for the European Union (EU) territory. SVBV is a well-defined virus species of the genus Caulimovirus for which the entire genome sequence is known and molecular detection assays are available. SVBV is transmitted by vegetative multiplication of infected hosts and through the activity of aphid vectors, the most efficient being Chaetosiphon spp. The virus is reported from all continents and is present in three EU Member States: the Czech Republic, Italy and Slovakia. The host range of SVBV is restricted to cultivated and wild strawberries. It is listed in Annex IAI of Directive 2000/29/EC. SVBV is not expected to be affected by ecoclimatic conditions wherever its hosts are present and has the potential to establish in large parts of the EU territory, and to subsequently spread through the action of its Chaetosiphon fragaefolii vector, which is present in many Member States. SVBV does not cause severe symptoms, and modern cultivars are mostly symptomless if infected with SVBV alone. SVBV can, however, contribute to more severe symptoms when it occurs in mixed infections with other strawberry viruses. Despite this, SVBV is considered a minor problem in strawberry production as a consequence of modern practices including the systematic use of certified planting materials and the use of short crop cycles, which have greatly reduced the impact of strawberry viruses. Overall, SVBV does not have the potential to be a quarantine pest as, given current agricultural practices, it does not fulfil the pest categorisation criteria defined in the International Standards for Phytosanitary Measures No 11 of having a severe impact. However, SVBV has the potential to be a regulated non-quarantine pest because it fulfils all pest categorisation criteria defined in the International Standards for Phytosanitary Measures No 21. [ABSTRACT FROM AUTHOR]

In this study a new real-time PCR assay for the detection of figwort mosaic virus (FMV) DNA is described. This assay targets a 113-bp-long sequence of the FMV open reading frame Vll, a non-conserved coding region among the caulimoviruses. Detection of FMV DNA is useful to complement screening for the FMV 34S promoter (P-FMV), a genetic element present in several genetically modified (GM) plants. The specificity of the assay was assessed against closely related plant viruses, plant species of agronomic importance or susceptible to infection by FMV, and various GM plants containing the P-FMV sequence. No cross-reactivity of the assay against the tested nontarget organisms was observed. The limit of detection of the FMV real-time PCR assay was determined at approximately 5 copies per reaction. The robustness of the method was tested using different real-time PCR instruments and PCR master mixes with no negative effects on the PCR efficiency and linearity being observed. When amplifying the FMV target DNA at a concentration level close to the detection limit, no negative influence on the PCR performance was observed in the presence of background genomic DNA from various plant species. The precision data of the in-house validation were in line with generally accepted performance requirements. In summary, the method appears fit for the purpose of a specific identification test for the presence of genomic FMV DNA, while preventing false-positive results during P-FMV screening. [ABSTRACT FROM AUTHOR]

Selected wild Dahlia species in their natural habitats from west-central Mexico were tested for the presence of three caulimoviruses known to be associated with cultivated dahlia ( Dahlia variabilis), viz. Dahlia mosaic virus (DMV), DMV-D10 and Dahlia common mosaic virus. Virus species-specific primers and PCR were used followed by cloning and sequencing of the amplicons. Results showed that the wild dahlia species in their natural habitat contained DMV-D10, which is an endogenous plant pararetrovirus. Viral sequences were found in 91% of the samples ( n = 56) representing four different wild species. The gene coding for the movement protein of DMV-D10 from Dahlia coccinea and all other species was cloned and sequenced. Sequence comparisons showed divergence of this gene when compared to that of DMV-D10 from cultivated dahlias. The discovery of plant pararetroviruses in wild dahlia species in their natural habitats suggests a possible emergence, co-existence and co-evolution of pararetroviruses and their host plants. [ABSTRACT FROM AUTHOR]

Dahlia mosaic, caused by Dahlia mosaic virus (DMV), is one of the most important viral diseases of dahlia. Molecular characterization of DMV showed the association of two distinct caulimoviruses (DMV-D10, DMV-Portland) and a D10-like sequence variant (DMV-Holland) with the disease. Using primers specific to these two viruses and the sequence variant, a polymerase chain reaction-based assay was used to determine their relative incidence in several dahlia samples from the United States and the Netherlands. Testing was done on samples collected in 2005 and 2006 in the United States and in 2006 in the Netherlands. Results indicated the predominance of DMV-D10 over DMV-Portland and DMV-Holland in both the United States and the Netherlands. Using conserved regions of the viral genome, primers were designed and used to detect all three sequences. Results suggested that DMV-D10 is predominantly associated with dahlia mosaic, but diagnostics should also include testing for DMV-Portland and DMV-Holland. [ABSTRACT FROM AUTHOR]

Dahlia mosaic is a serious disease affecting dahlias. In addition to the Dahlia mosaic virus (DMV) reported previously, we characterized two putative new caulimoviruses, tentatively designated as DMV-D10 and Dahlia common mosaic virus (DCMV), from dahlia. To better understand their relative incidence in dahlia, a total of 213 samples were collected during 2007 and 2008 from several varieties of cultivated dahlia (D. variabilis) in the United States. Samples were tested for the three caulimoviruses using virus-specific primers in a polymerase chain reaction. Amplicons were cloned and sequenced to confirm the infection of dahlia with these viruses. Results showed that DMV-D10 was the most prevalent (94%) followed by DCMV (48.5%) and DMV (23%). Mixed infections were common and viruses were detected irrespective of symptom expression at the time of sampling. Two percent of the samples were not infected by any of the three tested caulimoviruses. Results suggest that caulimovirus infectious are widespread in dahlia and highlight the need for testing and production of virus-free material to reduce their spread. [ABSTRACT FROM AUTHOR]

The strawberry vein banding virus (SVBV) open reading frame (ORF) VI encodes a P6 protein known as the RNA silencing suppressor. This protein is known to form inclusion like granules of various sizes and accumulate in both the nuclei and the cytoplasm of SVBV-infected plant cells. In this study, we have determined that the P6 protein is the only trans-activator (TAV) encoded by SVBV, and can efficiently trans-activate the translation of downstream gfp mRNA in a bicistron derived from the SVBV. Furthermore, the P6 protein can trans-activate the expression of different bicistrons expressed by different caulimovirus promoters. The P6 protein encoded by SVBV from an infectious clone can also trans-activate the expression of bicistron. Through protein-protein interaction assays, we determined that the P6 protein could interact with the cell translation initiation factor FveIF3g of Fragaria vesca and co-localize with it in the nuclei of Nicotiana benthamiana cells. This interaction reduced the formation of P6 granules in cells and its trans-activation activity on translation. [ABSTRACT FROM AUTHOR]