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Soybean chlorotic mottle virus (SbCMV) was first detected from soybean plants in Jiangxi Province of China by high throughput sequencing and was confirmed by PCR. The complete nucleotide sequence of NC113 was determined to be 8 210 nucleotides, and shared the highest similarity (91.7%) with sequences of SbCMV that was only reported in Japan. It encodes nine putative open reading frames (ORFs Ia, Ib and II–VIII), and contains a large intergenic region located at nucleotide 5 976–6 512 between ORFs VI and VII. Sequence analysis and phylogenetic tree indicated that NC113 is an isolate of SbCMV, and is more related to the soymoviruses Blueberry red ringspot virus (BRRSV), Peanut chlorotic streak virus (PCSV) and Cestrum yellow leaf curling virus (CmYLCV) than to other representative members in the Caulimoviridae family. Field survey of 472 legume plants from Jiangxi and Zhejiang provinces showed SbCMV was only detected from soybean in Nanchang City with a low incidence rate. This is the first report of Soybean chlorotic mottle virus identified in China.

A disease of Rudbeckia hirta (Black-eyed Susan), characterized by severe flower deformation, was observed in Minnesota during 2010–2016. A previously undescribed virus species, named Rudbeckia flower distortion virus (RuFDV, family Caulimoviridae, genus unassigned), was determined to be the causal agent of the disease. Symptoms induced by RuFDV infection resemble those characteristic of phytoplasma-induced diseases, but no phytoplasmas were detected in RuFDV-infected R. hirta. The virus, and the disease were transmitted readily by mechanical inoculation and by the aphid Myzus persicae, but only to R. hirta. Virions of RuFDV are icosahedral, 42–45 nm in diameter, and contain a circular 8222 bp dsDNA genome containing seven open reading frames (ORFs). The ORFs 2–6 have 28–52% amino acid sequence identity to the movement protein (MP), coat protein (CP), aspartic protease (AP), reverse transcriptase (RT) and RNase H, and translational transactivator (TA) domains of known caulimoviruses. The two remaining ORFs (1 and 7) have no significant amino acid sequence similarity to known viruses. Although the RuFDV ORF 6 is significantly truncated relative to those of other known caulimoviruses, neither this nor the concomitant absence of characteristic virus-encoded cytoplasmic inclusion bodies appears to adversely affect aphid transmission of this virus. Phylogenetic analysis based on the sequence of the RT region revealed no close relationship to known members of the family Caulimoviridae. Based on sequence similarity, genome organization and phylogenetic relatedness, RuFDV appears to be distinct from any currently recognized taxonomic grouping in the family Caulimoviridae.

Viral fossils in rice genomes are a best entity to understand ancient pararetrovirus activities through host plant history because of our advanced knowledge of the genomes and evolutionary history with rice and its related species. Here, we explored organization, geographic origins and genealogy of rice pararetroviruses, which were turned into endogenous rice tungro bacilliform virus-like (eRTBVL) sequences. About 300 eRTBVL sequences from three representative rice genomes were clearly classified into six families. Most of the endogenization events of the eRTBVLs were initiated before differentiation of the rice progenitor (> 160,000 years ago). We successfully followed the genealogy of old relic viruses during rice speciation, and inferred the geographical origins for these viruses. Possible virus genomic sequences were explained mostly by recombinations between different virus families. Interestingly, we discovered that only a few recombination events among the numerous occasions had determined the virus genealogy.

Microsatellites are known to exhibit ubiquitous presence across all kingdoms of life including viruses. Members of the Caulimoviridae family severely affect growth of vegetable and fruit plants and reduce economic yield in diverse cropping systems worldwide. Here, we analyzed the nature and distribution of both simple and complex microsatellites present in complete genome of 44 species of Caulimoviridae. Our results showed, in all analyzed genomes, genome size and GC content had a weak influence on number, relative abundance and relative density of microsatellites, respectively. For each genome, mono- and dinucleotide repeats were found to be highly predominant and are overrepresented in genome of majority of caulimoviruses. AT/TA and GAA/AAG/AGA was the most abundant di- and trinucleotide repeat motif, respectively. Repeats larger than trinucleotide were rarely found in these genomes. Comparative study of occurrence, abundance and density of microsatellite among available RNA and DNA viral genomes indicated that simple repeats were least abundant in genomes of caulimoviruses. Polymorphic repeats even though rare were observed in the large intergenic region of the genome, indicating strand slippage and/or unequal recombination processes do occur in caulimoviruses. To our knowledge, this is the first analysis of microsatellites occurring in any dsDNA viral genome. Characterization of such variations in repeat sequences would be important in deciphering the origin, mutational processes, and role of repeat sequences in viral genomes.

Endogenous pararetrovirus sequences (EPRV) belonging to the plant virus family Caulimoviridae have been discovered in the genomes of a wide range of Angiosperms. Although knowledge of EPRVs in plants is still in its infancy, it has been shown clearly in three different plant-virus pathosystems that these integrations are capable of generating functional circular viral genomes, and can thus trigger systemic infection. Here, we recapitulate information gathered over the last 15 years on how EPRVs contribute to virus replication in plants. We first present recent advances in our understanding of the molecular mechanisms involved in the transition from integrated to circular viral forms before addressing how EPRVs are controlled in planta.

To test the hypothesis that many viruses remain to be discovered in plants, a procedure was developed to sequence nucleic acids cloned randomly from virus-like particle fractions of plant homogenates. As a test of the efficiency of the procedure we targeted Ambrosia psilostachya, western ragweed, plants growing at the Tallgrass Prairie Preserve of northeastern Oklahoma. Amplifiable nucleic acid was found in the fractions from six of twelve specimens and sequences were characterized from four of them. Evidence was obtained for the presence of viruses belonging to two families (Caulimoviridae, Flexiviridae). Multiple viral species were found in two of the four specimens and their level within the isolated nucleic acid population varied from less than 1-37%. None of the sequences were derived from reported sequences of known viruses. Thus, the analysis of nucleic acid from virus-like particles is a useful tool to expand our knowledge of the universe of viruses to non-cultivated species.

Endogenous plant pararetroviruses (EPRVs) were identified as integrated counterparts of most members of the plant virus family Caulimoviridae and represent repetitive elements that are ubiquitous in the plant kingdom. They are often located in pericentromeric regions of plant chromosomes in the vicinity of retrotransposon sequences. Depending on their structure and sequence integrity, some EPRVs are able to replicate and to initiate viral infection. However, conservation of integrated sequences in plant genomes might indicate benefits for the host during evolution. Understanding EPRV activation and control by the host could have important implications for plant breeding strategies to prevent viral disease caused by EPRVs in newly generated cultivars.

Rice tungro, economically the most important viral disease of rice, is a complex disease caused by two morphologically and genomically dissimilar viruses, Rice tungro bacilliform virus (RTBV), a double stranded DNA virus replicating through RNA intermediate and Rice tungro spherical virus (RTSV), a single stranded RNA virus with 3' poly (A) tail. A novel multiplex RT-PCR technique for the simultaneous detection of RTBV and RTSV from the total RNA extracted from tungro-infected plants has been developed. It involves a one-step reaction initiating first strand cDNA synthesis by oligo (dT) primer with poly (A) tailed RTBV transcript and RTSV genomic RNA as template for the PCR amplification. The results indicate that adaptation of this technique will strengthen the screening for tungro resistance among rice varieties and hybrids.

The requirement for helper component(s) (HC) facilitating virus transmission by living organisms, termed vectors, is characteristic of plant viruses in several genera, potyviruses (genus Potyvirus, family Potyviridae) and caulimoviruses (genus Caulimovirus, family Caulimoviridae) being the most intensively studied groups of viruses. The potyviral HC-Pro fulfils several functions, including aphid transmission, long-distance movement and suppression of post-transcriptional gene silencing. The HC of Cauliflower mosaic virus plays an important role in aphid transmission, whereas no direct role in viral infectivity has so far been attributed to this protein. This review shows the current knowledge of major functions of HCs encoded by potyviruses and caulimoviruses.

We analyzed the correlation between the development of symptoms, virus titre and carbohydrate levels in turnip and Arabidopsis during infection with four isolates of Cauliflower mosaic virus (CaMV). Infection did not significantly affect sugar levels in source leaves except at the very late stages (28 d.p.i.), but induced a three-fold but short lived increase in sink leaves. Titres of severe isolates were greater than mild isolates, but we observed no obvious correlation between sugar levels and symptom appearance. In wild-type Arabidopsis, infection did not stimulate increased sugar levels, but did so in a mutant, cnr160, which shows altered growth-response to high carbon and low nitrogen. These results do not support a direct role for sugar-mediated control of symptom development during virus infection.

A full-length transcript (FLt) promoter was isolated from a genomic clone of Strawberry vein banding virus(SVBV), a double-stranded DNA plant pararetrovirus belonging to the Caulimoviridae family, and its activity was analyzed both in protoplasts and transgenic plants. The 5 � –3 � -boundaries required for maximal promoter activity were determined by 5 � - and 3 � -end deletion analysis of the SVBV promoter fused to a -glucuronidase (GUS) reporter gene. A 371-bp promoter fragment (−352 to +19 from the transcription start site; TSS) was found sufficient for maximal promoter activity in a transient protoplast expression assay, and this was chosen for further analysis. Finer deletion analysis of a 90-bp sequence (coordinates −392 to −302 from TSS) of the SVBV FLt promoter revealed the presence of a negative and a positive regulatory element in this region. In gain-of-function experiment, the fusion of the putative positive regulatory elements with minimal promoter showed very little increment in promoter activity, suggesting that a combinatorial action of various cis-sequences is involved in promoter function. The transcription start site of the full-length transcript promoter was mapped to an A-residue that is located 25-bp downstream of the TATA-box. In protoplast transient expression analysis, the SVBV FLt promoter showed about six-fold higher activity in tobacco compared to maize. A quantitative GUS activity assay showed that in transgenic tobacco plants the average promoter activity was about three-fold higher in roots than in leaves, and this higher activity was due to the accumulation of more GUS specific mRNA in roots. Real-time qRT-PCR analysis and quantitative GUS activity assay showed that the relative strength of the SVBV FLt promoter was greater than the CaMV35S promoter in transgenic tobacco plants. The SVBV FLt promoter is a strong, constitutive promoter and has great application potential in expression of foreign genes in plants. © 2004 Elsevier Ireland Ltd. All rights reserved.

The principal aphid-borne viruses infecting Strawberry (Fragaria spp.) Strawberry crinkle virus (SCV), Strawberry mild yellow edge virus (SMYEV), Strawberry mottle virus (SMoV) and Strawberry vein banding virus (SVBV) can cause serious crop losses. In this paper, a multiplex reverse transcriptase polymerase chain reaction (RT-PCR) method is described for the simultaneous detection of all four viruses in combination with a plant mRNA specific internal control which can be used as an indicator of the effectiveness of the extraction and RT-PCR. In total, 18 strawberry isolates infected naturally were analysed by this method. Every combination of RNA virus was able to be detected and a full complement of all four viruses were found together in three isolates, all taken from wild strawberry (Fragaria chiloensis (L.) Duch.) in Chile. The upper detection limit for the four viruses was at an extract dilution of 1/200. The broad applicability of the RNA specific internal control primers—which produced a PCR fragment of the expected size in 25 of 27 plant species tested—combined with improvements, made in extraction methods described provides potentially a standard method for comparable RT-PCR analyses in a wide variety of plant species.

The genus Caulimovirus consists of several distinct virus species with a double-stranded DNA genome that infect diverse plant species. A comparative analysis of the sequences of known Caulimovirus species revealed two regions that are conserved in all Caulimovirus species with the exception of Strawberry vein banding virus. Degenerate primers based on these two regions were designed and tested in a polymerase chain reaction-based assay for broad spectrum detection of members of this genus. Cauliflower mosaic virus, Figwort mosaic virus and three distinct caulimoviruses associated with dahlia (Dahlia variabilis) were used to show the utility of this test in detecting diverse caulimoviruses. The primer pair gave an amplicon of expected size (840 bp). Amplicons from each virus were cloned and sequenced to verify their identity. The primer pair and the PCR assay provide approach for the broad spectrum detection of several members of the genus Caulimovirus.

Breeding and tissue culture of certain cultivars of bananas (Musa) have led to high levels of banana streak badnavirus (BSV) infection in progeny from symptomless parents. BSV DNA hybridized to genomic DNA of one such parent, Obino l'Ewai, suggesting integration of viral sequences. Sequencing of clones of Obino l'Ewai genomic DNA revealed an interface between BSV andMusasequences and a complex BSV integrant.In situhybridization revealed two different BSV sequence locations in Obino l'Ewai chromosomes and a complex arrangement of BSV andMusasequences was shown by probing stretched DNA fibers. This is the first report of integrated sequences that possibly lead to a plant pararetrovirus episomal infection by a mechanism differing markedly from animal retroviral systems.

Petunia vein-clearing virus (PVCV) is a plant pararetrovirus that has some features of retrotransposons. It encapsidates dsDNA and has isometric particles and inclusion bodies similar to those of caulimoviruses. The PVCV genome of 7205 bp has two large ORFs in the transcribed strand and a methionine tRNA primer-binding site in its 663-bp intergenic region. The N-terminal position of the large protein (126 kDa) encoded by ORF I has similarity to the movement protein of caulimoviruses. Toward the C-terminus of this same polyprotein are the two distinctive sequence elements [HHCC and DD(35)E] of the integrase function of retroviruses and retrotransposons. ORF II of PVCV encodes a protein of 125 kDa with domains for an RNA-binding element, common to thegaggene of retroelements, followed by consensus sequences for an acid protease, reverse transcriptase, and ribonuclease H. Hence, thegagequivalent (capsid protein) andpolgene of PVCV are part of the same polyprotein. Phylogenetic comparison of the reverse transcriptase of PVCV with that of various other retroelements grouped PVCV between caulimoviruses and the Ty3/gypsyretrotransposons, suggesting that PVCV is a divergent member of the caulimoviruses.