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The article offers information on the emergence of coffee ringspot virus (CoRSV). Topics discussed include information on the genetic features of coffee ringspot virus same as plant viruses; role of climate change in CoRSV emergence in terms of temperature-dependent; and views on the CoRSV research regarding the replication of virus.

A generic RT-PCR assay was developed for the universal detection of viruses of the genus Tobamovirus using a novel pair of degenerate primers designed based on conserved regions on replicase genes of 32 tobamoviruses. The assay detected nine tobamoviruses, including six Solanaceae-infecting subgroup tobamoviruses of Tobacco mosaic virus (TMV), Tomato mosaic virus (ToMV), Tomato mottle mosaic virus (ToMMV), Tobacco mottle green mosaic virus (TMGMV), Pepper mild mottle virus (PMMoV), Paprika mild mottle virus (PaMMV), one Orchidaceae-infecting tobamovirus of Odontoglossum ringspot virus (ORSV) and two Cucurbitaceae-infecting subgroup tobamoviruses of Cucumber green mottle mosaic virus (CGMMV) and Zucchini green mottle mosaic virus (ZGMMV), with high amplification efficiency, specificity and sensitivity. The assay was applied to detect tobamoviruses in pepper and tomato fields. Five tobamoviruses, PMMoV, TMV, ToMV, ToMMV and TMGMV, were detected from the pepper fields in single and mixed infections. Single infections of PMMoV, ToMV and ToMMV and mix-infection of ToMV + PMMoV were detected from the tomato fields. Among these viruses, PMMoV was first detected from tomato worldwide, while ToMMV was first detected from tomato plants in China. This generic assay is simple, cost-effective and has great potential to detect more tobamoviruses in the field. [ABSTRACT FROM AUTHOR]

Soybean blotchy mosaic virus (SbBMV), a plant virus of the genus Cytorhabdovirus is an economically important virus of soybean reported only from the warmer, lower-lying soybean production areas in South Africa. The virus consistently appears in soybean crops annually in spite of the absence of soybean plants in winter. One possible reason for this may be that the virus replicates and hence persists in the SbBMV vector, a leafhopper, Peragallia caboverdensis . RNA viruses with antisense genomes as inferred for SbBMV produce positive sense RNAs as intermediate replicative forms during replication in their hosts, and detection of the positive strand in the plant host or vector is evidence of virus replication. In this study, a positive-strand specific RT-PCR (pss-RT-PCR) was developed to detect the positive RNA strand of SbBMV and validated on nine SbBMV isolates from soybean. The effect of tagged reverse transcription (RT) primers for cDNA synthesis, coupled with PCR using a tag-specific primer, as well as removal of unincorporated RT primers following cDNA synthesis was assessed. The positive RNA strand of SbBMV in infected plants was successfully detected following this protocol. Reverse transcription with forward and unmodified reverse primers confirmed that the assay was not able to detect the genomic sense RNA or self-primed cDNAs, lacking the non-viral tag, respectively. However, Exonuclease I (ExoI) treatment of cDNA was required to eliminate false-positive results during PCR amplification. [ABSTRACT FROM AUTHOR]

A Tospovirus species was isolated from peanut plants showing chlorotic ring spots and chlorosis, and identified as Tomato yellow fruit ring virus (TYFRV) on the basis of its biological, serological, and molecular properties. In host range studies, a broad range of indicator plants was infected by the five isolates studied; all the isolates systemically infected Nicotiana tabacum cultivars and, thus, they were classified into the N-host-infecting type isolates of the virus. These isolates strongly reacted with TYFRV antibodies but not with the specific antibodies of other tospoviruses tested. Recombination analyses showed that the nucleoprotein gene of the peanut isolates and other isolates studied were nonrecombinant. In phylogenetic trees, the virus isolates were clustered in three genogroups: IRN-1, IRN-2, and a new group, POL; the peanut isolates fell into IRN-2 group. Multiple sequence alignments showed some genogroup-specific amino acid substitutions among the virus isolates studied. The results revealed the presence of negative selection in TYFRV populations. Also, the Iranian populations had higher nucleotide diversity compared with the Polish population. Genetic differentiation and gene flow analyses indicated that the populations from Iran and Poland and those belonging to different genogroups were partially differentiated populations. Our findings seem to suggest that there has been frequent gene flow between some populations of the virus in the mid-Eurasian region of Iran. [ABSTRACT FROM AUTHOR]

A bipartite begomovirus was identified from a Boehmeria leiophylla plant sample exhibiting yellow mosaic symptoms collected in Nabanhe National Nature Reserve, Xishuangbanna, Yunnan, China. Five complete DNA-A and four DNA-B genome sequences were obtained by rolling-circle amplification (RCA), cloned, and sequenced. All DNA-A sequences were determined to be 2759 nucleotides in size, sharing 99.9%-100% nucleotide sequence identity with each other. The DNA-B sequences were comprised of 2673 nucleotides, sharing 98.6-100% nucleotide sequence identity with each other. Genomic organization of the begomovirus was typical of Old World bipartite begomoviruses. Sequence analysis revealed 81.84% nucleotide sequence identity to tomato leaf curl Hsinchu virus (ToLCHsV) from China for the DNA A component and 67.23% identity to the squash leaf curl China virus (SLCCNV) from India for the DNA B component. The sequence comparisons suggest that this bipartite begomovirus represents a novel species for which we propose the name “Ramie yellow mosaic virus”. [ABSTRACT FROM AUTHOR]

A new virus causing a serious stunt disease of chrysanthemum was identified in China by high-throughput sequencing (HTS) and named chrysanthemum virus R (CVR). The complete sequence of CVR was determined by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The genomic RNA of CVR consists of 8,874 nucleotides (nt), excluding the poly(A) tail, contains six putative open reading frames (ORFs), and has a genomic organization typical of members of the genus Carlavirus. BLAST analysis of the full genome sequence showed low similarity (38%-56% sequence identity) to other members of the genus Carlavirus. BLAST analysis and phylogenetic analysis based on the amino acid (aa) sequences of the CVR replicase and coat protein (CP) confirmed that CVR is a distinct member of the genus Carlavirus. [ABSTRACT FROM AUTHOR]

Because so few viruses in the family Barnaviridae have been reported, we searched for more of them in public sequence databases. Here, we report the complete coding sequence of Colobanthus quitensis associated barnavirus 1, mined from a transcriptome of the Antarctic pearlwort Colobanthus quitensis. The 4.2-kb plus-strand sequence of this virus encompasses four main open reading frames (ORFs), as expected for barnaviruses, including ORFs for a protease-containing polyprotein, an RNA-dependent RNA polymerase whose translation appears to rely on − 1 ribosomal frameshifting, and a capsid protein that is likely to be translated from a subgenomic RNA. The possible derivation of this virus from a fungus associated with C. quitensis is discussed. [ABSTRACT FROM AUTHOR]

Strain differentiating marker profiles of citrus tristeza virus (CTV) isolates from California have shown the presence of multiple genotypes. To better define the genetic diversity involved, full-length genome sequences from four California CTV isolates were determined by small-interfering RNA sequencing. Phylogenetic analysis and nucleotide sequence comparisons differentiated these isolates into the genotypes VT (CA-VT-AT39), T30 (CA-T30-AT4), and a new strain called S1 (CA-S1-L and CA-S1-L65). S1 isolates had three common recombination events within portions of genes from VT, T36 and RB strains and were transmissible by Aphis gossypii. Virus indexing showed that CA-VT-AT39 could be classified as a severe strain, whereas CA-T30-AT4, CA-S1-L and CA-S1-L65 were mild. CA-VT-AT39, CA-S1-L, and CA-S1-L65 reacted with monoclonal antibody MCA13, whereas CA-T30-AT4 did not. RT-PCR and RT-qPCR detection assays for the S1 strain were developed and used to screen MCA13-reactive isolates in a CTV collection from central California collected from 1968 to 2011. Forty-two isolates were found to contain the S1 strain, alone or in combinations with other genotypes. BLAST and phylogenetic analysis of the S1 p25 gene region with other extant CTV sequences from the NCBI database suggested that putative S1-like isolates might occur elsewhere (e.g., China, South Korea, Turkey, Bosnia and Croatia). This information is important for CTV evolution, detection of specific strains, and cross-protection. [ABSTRACT FROM AUTHOR]

We investigated the molecular characteristics of an Argentinean isolate of alfalfa leaf curl virus (ALCV-Arg), a virus of the genus Capulavirus in the family Geminiviridae that was isolated from alfalfa plants showing dwarfism. The genome was found to be 2,750 nucleotides in length. In pairwise comparisons, this ALCV isolate shared 83.2% to 92.6% sequence identity with European ALCV isolates. Sequence comparisons and phylogenetic analysis showed that this isolate combines features of strains A and B of ALCV. Recombination analysis showed that ALCV-Arg is a recombinant isolate that was generated by intraspecific recombination between ALCV strains A and B. The results of this study not only show that ALCV-Arg is unique because it combines features of strains A and B but also show that ALCV naturally infects this forage crop on the American continent. [ABSTRACT FROM AUTHOR]

Maize Iranian mosaic virus (MIMV) is a negative-sense single-stranded RNA virus that is classified in the genus Nucleorhabdovirus, family Rhabdoviridae. The MIMV genome contains six open reading frames (ORFs) that encode in 3΄ to 5΄ order the nucleocapsid protein (N), phosphoprotein (P), putative movement protein (P3), matrix protein (M), glycoprotein (G) and RNA-dependent RNA polymerase (L). In this study, we determined the first complete genome sequence of MIMV using Illumina RNA-Seq and 3′/5′ RACE. MIMV genome (‘Fars’ isolate) is 12,426 nucleotides in length. Unexpectedly, the predicted N gene ORF of this isolate and of four other Iranian isolates is 143 nucleotides shorter than that of the MIMV coding-complete reference isolate ‘Shiraz 1’ (Genbank NC_011542), possibly due to a minor error in the previous sequence. Genetic variability among the N, P, P3 and G ORFs of Iranian MIMV isolates was limited, but highest in the G gene ORF. Phylogenetic analysis of complete nucleorhabdovirus genomes demonstrated a close evolutionary relationship between MIMV, maize mosaic virus and taro vein chlorosis virus. [ABSTRACT FROM AUTHOR]

Nanoviruses are multi-component plant-infecting single-stranded DNA viruses. Using a viral metagenomics-informed approach, a new nanovirus and two associated alphasatellite molecules have been identified in an uncultivated asymptomatic Vicia cracca plant in the Rhône region of France. This novel nanovirus genome includes eight genomic components (named DNA-R, DNA-S, DNA-M, DNA-C, DNA-N, DNA-U1, DNA-U2 and DNA-U4) and, across all components, shares < 66% pairwise sequence identity with other nanovirus genomes. The two associated alphasatellites share 62% identity with each other and < 81% identity will all other nanovirus-associated alphasatellites. [ABSTRACT FROM AUTHOR]

Narcissus plants (Narcissus tazetta var. chinensis) showing mosaic or striping leaves were collected from around Japan, and tested for virus infections using potyvirus-specific primers. Many were found to be infected with a macluravirus and mixtures of different potyviruses, one third of them narcissus yellow stripe virus (NYSV)-like viruses. Genomes of nine of the NYSV-like viruses were sequenced and, together with four already published, provided data for phylogenetic and pairwise identity analyses of their place in the turnip mosaic virus (TuMV) phylogenetic group. Using existing ICTV criteria for defining potyvirus species, the narcissus viruses in TuMV group were found to be from five species; the previously described NLSYV, and four new species we call narcissus virus 1 (NV-1) and narcissus yellow stripe-1 to -3 (NYSV-1, NYSV-2 and NYSV-3). However, as all are from a single host species, and natural recombinants with NV-1 and NYSV-3 'parents have been found in China and India, we also conclude that they could be considered to be members of a single mega-species, narcissus virus; the criteria for defining such a potyvirus species would then be that their polyprotein sequences have greater than 69% identical nucleotides and greater than 75% identical amino acids. [ABSTRACT FROM AUTHOR]

For the exploration of genetic diversity, 151 exotic cotton germplasm were evaluated for CLCuV, yield and fiber traits at the environmental conditions of central zone of Sakrand, Sindh. Basic statistical parameters, correlation and principal components were employed to explore the potential source for the utilization in breeding program to maintain the desired traits and develop CLCuV resistant/tolerant line. The significant positive association was observed among traits. Seed cotton yield was related positively significantly with traits sympodial branches, bolls plant-1, boll weight, ginning outturn, staple length and uniformity index. Whereas the negative correlation was observed with number of monopodial branches, plant height, seed index, micronaire value and seed index. Principal components were employed to assess the genetic variability. Out of 13 PCs 5 components exhibited >1 Eigen value, which shows 67.5% variability, whereas the remaining 32.5% variability contributed by further 8 components. The PC1 contributed highest genetic variability (22.2%) followed by PC2 (14.8%), PC3 (12.5%), PC4 (9.2%) and PC5 (8.8%), respectively. The principal component analysis (PCA) authenticated the amount of variability among traits and the material which may be tested exploited in breeding program to maintain the desirable traits, CLCuV resistant/tolerant, yield, its components, GOT and fiber quality traits. For yield, its components, GOT, fiber traits and cotton leaf curl virus USG2554, USG2570, USG2807, USG2810, USG2813, USG2833 and USG2848 can be utilized in breeding program to maintain the desirable traits. [ABSTRACT FROM AUTHOR]

Most of the positive-strand RNA plant viruses lack the 5'-cap and/or the poly(A)-tail that act synergistically to stimulate canonical translation of cellular mRNAs. However, they have RNA elements in the 5' - or 3'-untranslated regions of their RNAs that are required for their cap-independent translation. Cap-independent translation enhancers (CITEs) have been identified in the genomic 3'-end of viruses belonging to the family Tombusviridae and the genus Luteovirus. Seven classes of 3'-CITEs have been described to date based on their different RNA structures. They generally control the efficient formation of the translation initiation complex by varying mechanisms. Some 3'-CITEs bind eukaryotic translation initiation factors, others ribosomal subunits, bridging these to the 5'-end by different mechanisms, often long-distance RNA--RNA interactions. As previously proposed and recently found in one case in nature, 3'- CITEs are functionally independent elements that are transferable through recombination between viral genomes, leading to potential advantages for virus multiplication. In this review, the knowledge on 3'-CITEs and their functioning is updated. We also suggest that there is local structural conservation in the regions interacting with eIF4E of 3'-CITEs belonging to different classes. [ABSTRACT FROM AUTHOR]

A novel Rhizobium radiobacter (synonym Agrobacterium tumefaciens )-mediated approach was developed to generate stable infectious clones of plant viruses. This method uses R. radiobacter for both cloning and inoculation of infectious clones, bypassing the requirement of cloning in E. coli to avoid the instability. Only three steps are included in this method: (i) construct viral genome-encoding plasmids in vitro by one-step Gibson assembly; (ii) transform the assembled DNA products into R. radiobacter ; (iii) inoculate plants with the R. radiobacter clones containing the viral genome. Stable infectious clones were obtained from two potyviruses papaya ringspot virus (PRSV) and papaya leaf distortion mosaic virus (PLDMV) using this method, whereas attempts utilizing "classical" E. coli cloning system failed repeatedly. This method is simple and efficient, and is promising for a wide application in generation of infectious clones of plant virus, especially for those which are instable in E. coli . [ABSTRACT FROM AUTHOR]

Key message: Identification of three genomic regions and underlying candidate genes controlling the high level of resistance to ToLCNDV derived from a wild melon. SNP markers appropriated for MAS management of resistance. Abstract: Tomato leaf curl New Delhi virus (ToLCNDV) is a bipartite begomovirus that severely affects melon crop ( Cucumis melo) in the main production areas of Spain since 2012. In this work, we evaluated the degree of resistance of four accessions (two belonging to the subsp. agrestis var. momordica and two to the wild agrestis group) and their corresponding hybrids with a susceptible commercial melon belonging to the subsp. melo (Piel de Sapo, PS). The analysis using quantitative PCR (qPCR) allowed us to select one wild agrestis genotype (WM-7) with a high level of resistance and use it to construct segregating populations ( F and backcrosses). These populations were phenotyped for symptom severity and virus content using qPCR, and genotyped with different sets of SNP markers. Phenotyping and genotyping results in the F and BC1s populations derived from the WM-7 × PS cross were used for QTL analysis. Three genomic regions controlling resistance to ToLCNDV were found, one major locus in chromosome 11 and two additional regions in chromosomes 12 and 2. The highest level of resistance (no or mild symptoms and very low viral titer) was obtained with the homozygous WM-7WM-7 genotype at the major QTL in chromosome 11, even with PSPS genotypes at the other two loci. The resistance derived from WM-7 is useful to develop new melon cultivars and the linked SNPs selected in this paper will be highly useful in marker-assisted breeding for ToLCNDV resistance in melon. [ABSTRACT FROM AUTHOR]

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The coat protein (CP) of Melon necrotic spot virus (MNSV) is structurally composed of three major domains. The middle S-domain builds a robust protein shell around the viral genome, whereas the C-terminal protruding domain, or P-domain, is involved in the attachment of virions to the transmission vector. Here, we have shown that the N-terminal domain, or R-domain, and the arm region, which connects the R-domain and S-domain, are involved in different key steps of the viral cycle, such as cell-to-cell movement and the suppression of RNA silencing and pathogenesis through their RNA-binding capabilities. Deletion mutants revealed that the CP RNA-binding ability was abolished only after complete, but not partial, deletion of the R-domain and the arm region. However, a comparison of the apparent dissociation constants for the CP RNA-binding reaction of several partial deletion mutants showed that the arm region played a more relevant role than the R-domain in in vitro RNA binding. Similar results were obtained in in vivo assays, although, in this case, full-length CPs were required to encapsidate full-length genomes. We also found that the R-domain carboxyl portion and the arm region were essential for efficient cell-to-cell movement, for enhancement of Potato virus X pathogenicity, for suppression of systemic RNA silencing and for binding of small RNAs. Therefore, unlike other carmovirus CPs, the R-domain and the arm region of MNSV CP have acquired, in addition to other essential functions such as genome binding and encapsidation functions, the ability to suppress RNA silencing by preventing systemic small RNA transport. [ABSTRACT FROM AUTHOR]

Plant-made biopharmaceuticals have long been considered a promising technology for providing inexpensive and efficacious medicines for developing countries, as well as for combating pandemic infectious diseases and for use in personalized medicine. Plant virus expression vectors produce high levels of pharmaceutical proteins within a very short time period. Recently, plant viruses have been employed as nanoparticles for novel forms of cancer treatment. This review provides a glimpse into the development of plant virus expression systems both for pharmaceutical production as well as for immunotherapy. [ABSTRACT FROM AUTHOR]

Recessive resistances to plant viruses in the Potyvirus genus have been found to be based on mutations in the plant eukaryotic translation initiation factors, eIF4E and eIF4G or their isoforms. Here we report that natural, monogenic recessive resistance to the Potyvirus Turnip mosaic virus (Tu MV) has been found in a number of mustard ( Brassica juncea) accessions. Bulked segregant analysis and sequencing of resistant and susceptible plant lines indicated the resistance is controlled by a single recessive gene, re cessive T u MV r esistance 03 ( retr03), an allele of the eukaryotic translation initiation factor 2B-beta ( eIF2Bβ). Silencing of eIF2Bβ in a Tu MV-susceptible mustard plant line and expression of eIF2Bβ from a Tu MV-susceptible line in a Tu MV-resistant mustard plant line confirmed the new resistance mechanism. A functional copy of a specific allele of eIF2Bβ is required for efficient Tu MV infection. eIF2Bβ represents a new class of virus resistance gene conferring resistance to any pathogen. eIF2B acts as a guanine nucleotide exchange factor ( GEF) for its GTP-binding protein partner eIF2 via interaction with eIF2· GTP at an early step in translation initiation. Further genotyping indicated that a single non-synonymous substitution (A120G) in the N-terminal region of eIF2Bβ was responsible for the Tu MV resistance. A reproducible marker has been developed, facilitating marker-assisted selection for Tu MV resistance in B. juncea. Our findings provide a new target for seeking natural resistance to potyviruses and new opportunities for the control of potyviruses using genome editing techniques targeted on eIF2Bβ. [ABSTRACT FROM AUTHOR]

Various viral pathogens infect Cucurbitaceae and cause economic losses. The aim of the present study was to detect plant viral pathogens including Cucumber mosaic virus (CMV), Cucumber green mottle mosaic virus (CGMMV), Zucchini yellow mosaic virus (ZYMV), Cucurbit yellow stunting disorder virus (CYSDV) and Cucurbit chlorotic yellows virus (CCYV) in Lorestan province, in western Iran, and also to determine CMV genetic diversity in Iranian populations. A total of 569 symptomatic leaf samples were collected in 2013 and 2014 from cucurbits growing regions in Lorestan province. The collected samples were assessed for viral diseases by ELISA. The results showed virus incidences in most regions. Then, the infection of 40 samples to CMV was confirmed by RT-PCR. Moreover, to distinguish between the two groups (I and II) of CMV, PCR products were digested by two restriction enzymes XhoI and EcoRI. Results of the digestion showed that the isolates of Lorestan belonged to group I. The CMV-coat protein gene of eight isolates from different regions and hosts was sequenced and phylogenetic analysis was performed. Subsequent analyses showed even more genetic variation among Lorestan isolates. The phylogenetic tree revealed that Lorestan province isolates belonged to two IA and IB subgroups and could be classified together with East Azerbaijan province isolates. The results of the present study indicate a wide distribution of CMV, ZYMV, CGMMV, CYSDV and CCYV viruses in cucurbits fields of Lorestan province and for the first time subgroup IB of CMV was reported on melon from Iran. [ABSTRACT FROM AUTHOR]

Chickpea ( Cicer arietinum L.) is an important legume crop and widely cultivated in northwestern provinces of Iran. During a survey in the 2015 growing season a total of 170 selected chickpea plants with general yellowing symptoms including stunting and leaf bronzing were collected. Serological Elisa and tissue blot immunoassay (TIBA) tests revealed the presence of Bean leaf roll virus (BLRV) and Chickpea chlorotic stunt virus (CpCSV) as the predominant viruses in the region. Some serologically positive samples of BLRV and CpCSV were selected and rechecked by RT-PCR. The results of amplified PCR products using a specific pair of primers towards the Cp gene region of the viruses were approximately 413 bp for CpCSV and 391 bp for BLRV. Results obtained from sequence comparison of BLRV (IR-F-Lor-5) isolate form two subgroups with eight other BLRV isolates from GeneBank indicating a high homology of 96% with isolates from Argentina, Germany, Tunisia, USA, Spain, and Colombia. An isolate from Norabad (Iran) (IR-Nor) had 98% homology with HQ840727 Libyan isolate. CpCSV sequence comparison with six other GeneBank isolates indicated 98% homology with isolates from Tunisia and Azerbaijan. The overall results of this research revealed the CpCSV and BLRV (luteoviruses) associated with the yellowing disease syndrome of chickpea crops in the surveyed region. [ABSTRACT FROM AUTHOR]

Small-scale sequencing has improved substantially in recent decades, culminating in the development of next-generation sequencing (NGS) technologies. Modern NGS methods have helped the discovery of many new plant viruses. Nevertheless, there is still a need to establish solid assembly pipelines targeting small genomes characterised by low identities to known viral sequences. Here, we describe and discuss the fundamental steps required for discovering and sequencing new plant viral genomes by NGS. A practical pipeline and standard alternative tools used in NGS analysis are presented. [ABSTRACT FROM AUTHOR]

The article provides information on a study based on genome sizes and functions found in plant viruses and the proteins released by them. Particular focus is given to the initial stages of transforming viral proteins into a complete virus life cycle, a resistance of recessive genes against potyvirus to enhance breeding programs, and details related to susceptibility genomes.

Plum pox virus (PPV) is the causal agent of sharka disease of stone fruit trees. Since the late 1960s, PPV infection has been reported in different regions of Turkey. In this study, we aim to discover PPV strains in infected regions and determine its genetic variability in Turkey. For this reason, 612 samples were collected from distant locations, where PPV was previously detected in most cases. First, PPV presence in the samples was tested with serological and molecular methods to confirm the disease. Then 314 positive samples were sequence analyzed at a 664-nucleotide length, including the P3-6K1 gene region, one of the most variable regions among Potyvirus species and quite conserved among different strains of a viral species. PPV-D and PPV-T strains were identified mostly in residential gardens, whereas PPV-M was mostly detected in the orchards, except one isolate that was identified as PPV-Rec in Bursa. PPV-T was found to be dominant in the Turkish PPV pool. Estimates of average evolutionary divergence over sequence pairs of P3-6K1 gene regions revealed that the mean intragroup diversity was 0.049 for W; 0.017 for Rec, T, and M; 0.14 for C; 0.012 for Turkish D; 0.009 for global D; and 0.007 for CR. [ABSTRACT FROM AUTHOR]

Next-generation sequencing technologies enable the rapid identification of viral infection of diseased organisms. However, despite a consistent decrease in sequencing costs, it is difficult to justify their use in large-scale surveys without a virus sequence enrichment technique. As the majority of plant viruses have an RNA genome, a common approach is to extract the double-stranded RNA (ds RNA) replicative form, to enrich the replicating virus genetic material over the host background. The traditional ds RNA extraction is time-consuming and labour-intensive. We present an alternative method to enrich ds RNA from plant extracts using anti-ds RNA monoclonal antibodies in a pull-down assay. The extracted ds RNA can be amplified by reverse transcriptase-polymerase chain reaction and sequenced by next-generation sequencing. In our study, we have selected three distinct plant hosts: Māori potato ( Solanum tuberosum), rengarenga ( Arthropodium cirratum) and broadleaved dock ( Rumex obtusifolius) representing a cultivated crop, a New Zealand-native ornamental plant and a weed, respectively. Of the sequence data obtained, 31-74% of the reads were of viral origin, and we identified five viruses including Potato virus Y and Potato virus S in potato; Turnip mosaic virus in rengarenga (a new host record); and in the dock sample Cherry leaf roll virus and a novel virus belonging to the genus Macluravirus. We believe that this new assay represents a significant opportunity to upscale virus ecology studies from environmental, primary industry and/or medical samples. [ABSTRACT FROM AUTHOR]

The complete genome sequences of four new isolates of strawberry mild yellow edge virus ( SMYEV) were determined and analysed. The isolates, designated as AB41-01, AB41-02, NB1165 and NS26, were found from strawberry fields showing strawberry decline symptoms in eastern Canada. AB41-01 and AB41-02 were from a single-plant sample originating from Prince Edward Island, while NB1165 came from New Brunswick and NS26 from Nova Scotia. Nucleotide sequence identities are 95.8% between AB41-01 and NB1165, 99.6% between AB41-02 and NS26, and 84% between AB41-01/ NB1165 and AB41-02/ NS26. The four isolates share nucleotide sequence identities of 83.5-89.9% to two previously identified SMYEV isolates, namely MY18 and D7. Phylogenetic analysis indicates that the four Canadian isolates represented two new SMYEV strain types and the strain divergences were not likely from recombination events among all presently known SMYEV isolates. [ABSTRACT FROM AUTHOR]

The full-length nucleotide sequence and genomic organization of a melon necrotic spot virus isolate from Haimen, China ( MNSV- HM), were determined. The MNSV- HM genome consists of a positive sense single-stranded RNA, 4267 nt in length, with at least five open reading frames ( ORFs) encoding p29, p89, p42, and two small 7- kDa proteins (p7A and p7B). p89 shares a common start codon with p29 and continues through the amber stop codon of p29 to produce an 89- kDa protein. The p7A ORF terminates in an amber codon whose read-through could generate a 14- kDa protein. Phylogenetic analyses based on the p42 amino acid sequence and complete genomic sequence placed MNSV- HM and Spanish isolates of the virus in a major cluster, indicating a close genetic relationship. In conclusion, we report the full-length sequence of MNSV- HM and its translation strategy. The obtained genomic organization and phylogenetic trees indicate that MNSV- HM belongs to the MNSV genus Carmovirus. To the best of our knowledge, this is the first demonstration of the complete nucleotide sequence of an MNSV isolate from China. [ABSTRACT FROM AUTHOR]

Background: Viruses are among the most destructive and difficult to control plant pathogens. Melon (Cucumis melo L.) has become the model species for the agriculturally important Cucurbitaceae family. Approaches that take advantage of recently developed genomic tools in melon have been extremely useful for understanding viral pathogenesis and can contribute to the identification of target genes for breeding new resistant cultivars. In this work, we have used a recently described melon microarray for transcriptome profiling of two melon cultivars infected with two strains of Melon necrotic spot virus (MNSV) that only differ on their 3'-untranslated regions. Results: Melon plant tissues from the cultivars Tendral or Planters Jumbo were locally infected with either MNSV-Mα5 or MNSV-Mα5/3'264 and analysed in a time-course experiment. Principal component and hierarchical clustering analyses identified treatment (healthy vs. infected) and sampling date (3 vs. 5 dpi) as the primary and secondary variables, respectively. Out of 7566 and 7074 genes deregulated by MNSV-Mα5 and MNSV-Mα5/3'264, 1851 and 1356, respectively, were strain-specific. Likewise, MNSV-Mα5/3'264 specifically deregulated 2925 and 1618 genes in Tendral and Planters Jumbo, respectively. The GO categories that were significantly affected were clearly different for the different virus/host combinations. Grouping genes according to their patterns of expression allowed for the identification of two groups that were specifically deregulated by MNSV-Mα5/3'264 with respect to MNSV-Mα5 in Tendral, and one group that was antagonistically regulated in Planters Jumbo vs. Tendral after MNSV-Mα5/3'264 infection. Genes in these three groups belonged to diverse functional classes, and no obvious regulatory commonalities were identified. When data on MNSV-Mα5/Tendral infections were compared to equivalent data on cucumber mosaic virus or watermelon mosaic virus infections, cytokinin-O-glucosyltransferase2 was identified as the only gene that was deregulated by all three viruses, with infection dynamics correlating with the amplitude of transcriptome remodeling. Conclusions: Strain-specific changes, as well as cultivar-specific changes, were identified by profiling the transcriptomes of plants from two melon cultivars infected with two MNSV strains. No obvious regulatory features shared among deregulated genes have been identified, pointing toward regulation through differential functional pathways. [ABSTRACT FROM AUTHOR]

To counter the host RNA silencing defense mechanism, many plant viruses encode RNA silencing suppressor proteins. These groups of proteins share very low sequence and structural similarities among them, which consequently hamper their annotation using sequence similarity-based search methods. Alternatively the machine learning-based methods can become a suitable choice, but the optimal performance through machine learning-based methods is being affected by various factors such as class imbalance, incomplete learning, selection of inappropriate features, etc. In this paper, we have proposed a novel approach to deal with the class imbalance problem by finding the optimal class distribution for enhancing the prediction accuracy for the RNA silencing suppressors. The optimal class distribution was obtained using different resampling techniques with varying degrees of class distribution starting from natural distribution to ideal distribution, i.e., equal distribution. The experimental results support the fact that optimal class distribution plays an important role to achieve near perfect learning. The best prediction results are obtained with Sequential Minimal Optimization (SMO) learning algorithm. We could achieve a sensitivity of 98.5%, specificity of 92.6% with an overall accuracy of 95.3% on a tenfold cross validation and is further validated using leave one out cross validation test. It was also observed that the machine learning models trained on oversampled training sets using synthetic minority oversampling technique (SMOTE) have relatively performed better than on both randomly undersampled and imbalanced training data sets. Further, we have characterized the important discriminatory sequence features of RNA-silencing suppressors which distinguish these groups of proteins from other protein families. [ABSTRACT FROM AUTHOR]

This paper describes the nucleotide sequence and genome organization of a novel RNA virus detected in grapevine (Vitis vinifera) cultivar ‘Kizil Sapak’ by high-throughput sequencing (HTS) and tentatively named “grapevine virus J” (GVJ). The full genome of GVJ is 7,390 nucleotides in length, which comprises five open reading frames (ORFs), including a 20K ORF (ORF 2) between the replicase (ORF 1) and the movement protein (ORF 3) genes. According to the level of sequence homology and phylogenetics, GVJ is proposed as a new member of the genus Vitivirus (subfamily Trivirinae; family Betaflexiviridae), with the closest characterized virus being grapevine virus D (GVD). [ABSTRACT FROM AUTHOR]

Background: Plant viruses in the genus Begomovirus, family Geminiviridae often cause substantial crop losses. These viruses have been emerging in many locations throughout the tropics and subtropics. Like many plant viruses, they are often not recognized by plant diagnostic clinics due in large part to the lack of rapid and cost effective assays. An isothermal amplification assay, Recombinase polymerase amplification (RPA), was evaluated for its ability to detect three begomoviruses and for its suitability for use in plant diagnostic clinics. Methods for DNA extraction and separation of amplicons from proteins used in the assay were modified and compared to RPA manufacturer's protocols. The modified RPA assays were compared to PCR assays for sensitivity, use in downstream applications, cost, and speed. Results: Recombinase polymerase amplification (RPA) assays for the detection of Bean golden yellow mosaic virus, Tomato mottle virus and Tomato yellow leaf curl virus (TYLCV) were specific, only amplifying the target viruses in three different host species. RPA was able to detect the target virus when the template was in a crude extract generated using a simple inexpensive extraction method, while PCR was not. Separation of RPA-generated amplicons from DNA-binding proteins could be accomplished by several methods, all of which were faster and less expensive than that recommended by the manufacturer. Use of these modifications resulted in an RPA assay that was faster than PCR but with a similar reagent cost. This modified RPA was the more cost effective assay when labor is added to the cost since RPA can be performed much faster than PCR. RPA had a sensitivity approximate to that of ELISA when crude extract was used as template. RPA-generated amplicons could be used in downstream applications (TA cloning, digestion with a restriction endonuclease, direct sequencing) similar to PCR but unlike some other isothermal reactions. Conclusions: RPA could prove useful for the cost effective detection of plant viruses by plant diagnostic clinics. It can be performed in one hour or less with a reagent cost similar to that of PCR but with a lower labor cost, and with an acceptable level of sensitivity and specificity. [ABSTRACT FROM AUTHOR]

Sugarcane streak mosaic virus (SCSMV) causes mosaic and streak symptoms on sugarcane and sorghum crops, and has a broad host range. SCSMV is a member of the genus Poacevirus in the family Potyviridae. Ten SCSMV isolates were collected from sugarcane plants showing mosaic and streaking in Southern China from 2009–2011. Sequence-based phylogenetic and population genetic analyses were conducted using four partial genomic sequences covering the full genomes. These analyses were used to estimate the subpopulation differentiation and divergence within the Chinese virus population, and were compared with isolates from India. SCSMV-infected sugarcane plants in the field commonly harbor virus quasispecies (mutant cloud), and often have mixed infections with the same virus isolates. Inter- and intra-lineage recombination sites were identified in the protein 1, helper-component proteinase, coat protein and 3′ non-coding regions of the Chinese isolates. All the Chinese non-recombinant isolates fell into at least nine lineages, and many clustered with Indian isolates. However, estimates of genetic differentiation and gene flow indicated that the SCSMV populations in China and India are genetically independent. Our genetic study of a poacevirus population in South Asia regions indicates the importance of the evolutionary-based design to control viruses. [ABSTRACT FROM AUTHOR]

The main objective was to develop a method for the specific diagnosis of PepMoV based on RTPCR. Primers were designed using as reference the genomic sequence of the Florida isolate to amplify a fragment of the 5'untranslatable end and amino-terminus of P1 protein of the genome. The Oligo program (version 7.4) was used to carry out the analysis in silico and establish the initial parameters of the reaction. The optimal temperature of primer hybridization, primer concentration, intra- and inter-assay repeatability, detection limit, and the analytic specificity were evaluated. The optimal temperature of primer hybridization was 59°C, and 0,32 μM was selected as the optimal concentration of the primers. The aspects of the assay evaluated allowed establishing the optimal conditions of the technique. The RT-PCR method behaved with a high specificity for the diagnosis of PepMoV. It showed a detection limit of 94 pg.μl-1. The molecular tool obtained will allow supporting the breeding programs and the integrated pest management of this crop in the country. [ABSTRACT FROM AUTHOR]

Maize (Zea mays L.) is a host to numerous pathogenic species that impose serious diseases to its ear and foliage, negatively affecting the yield and the quality of the maize crop. A considerable amount of research has been carried out to elucidate mechanisms of maize-pathogen interactions with a major goal to identify defense-associated proteins. In this review, we summarize interactions of maize with its agriculturally important pathogens that were assessed at the proteome level. Employing differential analyses, such as the comparison of pathogen-resistant and susceptible maize varieties, as well as changes in maize proteomes after pathogen challenge, numerous proteins were identified as possible candidates in maize resistance. We describe findings of various research groups that used mainly mass spectrometry-based, high through-put proteomic tools to investigate maize interactions with fungal pathogens Aspergillus flavus, Fusarium spp., and Curvularia lunata, and viral agents Rice Black-streaked Dwarf Virus and Sugarcane Mosaic Virus. [ABSTRACT FROM AUTHOR]

We genetically modified tobacco mosaic virus (TMV) to surface display a characterized peptide with potent metal ion binding and reducing capacity (MBP TMV), and demonstrate that unlike wild type TMV, this construct can lead to the formation of discrete 10-40 nm gold nanoparticles when mixed with 3 mM potassium tetrachloroaurate. Using a variety of analytical physicochemical approaches it was found that these nanoparticles were crystalline in nature and stable. Given that the MBP TMV can produce metal nanomaterials in the absence of chemical reductants, it may have utility in the green production of metal nanomaterials. [ABSTRACT FROM AUTHOR]

Plant virus species of the family Nanoviridae have segmented genomes with the highest known number of segments encapsidated individually. They thus likely represent the most extreme case of the so-called multipartite, or multicomponent, viruses. All species of the family are believed to be transmitted in a circulative nonpropagative manner by aphid vectors, meaning that the virus simply crosses cellular barriers within the aphid body, from the gut to the salivary glands, without replicating or even expressing any of its genes. However, this assumption is largely based on analogy with the transmission of other plant viruses, such as geminiviruses or luteoviruses, and the details of the molecular and cellular interactions between aphids and nanoviruses are poorly investigated. When comparing the relative frequencies of the eight genome segments in populations of the species Faba bean necrotic stunt virus (FBNSV) (genus Nanovirus) within host plants and within aphid vectors fed on these plants, we unexpectedly found evidence of reproducible changes in the frequencies of some specific segments. We further show that these changes occur within the gut during early stages of the virus cycle in the aphid and not later, when the virus is translocated into the salivary glands. This peculiar observation, which was similarly confirmed in three aphid vector species, Acyrthosiphon pisum, Aphis craccivora, and Myzus persicae, calls for revisiting of the mechanisms of nanovirus transmission. It reveals an unexpected intimate interaction that may not fit the canonical circulative nonpropagative transmission. [ABSTRACT FROM AUTHOR]

Genome uncoating is a prerequisite for the successful infection of plant viruses in host plants. Thus far, little is known about the genome uncoating of the Carnation mottle virus (CarMV). Here, we obtained two reconstructions of CarMV at pH7 in the presence (Ca-pH7) and absence (EDTA-pH7) of calcium ions by Cryo-EM single particle analysis, which achieved 6.4 Å and 8 Å resolutions respectively. Our results showed that chelation of the calcium ions under EDTA-pH7 resulted in reduced interaction between the subunits near the center of the asymmetric unit but not overall size change of the viral particles, which indicated that the role of the calcium ions in CarMV was not predominantly for the structural preservation. Part of the genomic RNA closest to the capsid was found to be located near the center of the asymmetric unit, which might result from the interaction between genomic RNA and Lys194 residues. Together with the electrostatic potential analysis on the inner surface of the asymmetric unit, the reduced interaction near the center of the asymmetric unit under EDTA-pH7 suggested that the genome release of CarMV might be realized through the center of the asymmetric unit. [ABSTRACT FROM AUTHOR]

A recent proposal that the genus Rymovirus be assimilated into the genus Potyvirus is examined, discussed, and rejected. It illustrates the danger of using ‘sequence identity’ as a proxy for phylogenetic relatedness to distinguish closely related but distinct groups of viruses. [ABSTRACT FROM AUTHOR]

Contigs with sequence homologies to cherry-associated luteovirus were identified by high-throughput sequencing analysis in two peach accessions. Complete genomic sequences of the two isolates of this virus were determined to be 5,819 and 5,814 nucleotides long, respectively. The genome of the new virus is typical of luteoviruses, containing eight open reading frames in a very similar arrangement. Its genomic sequence is 58-74% identical to those of other members of the genus Luteovirus. These sequences thus belong to a new virus, which we have named 'peach-associated luteovirus'. [ABSTRACT FROM AUTHOR]

Begomovirus is a genus of highly destructive plant viruses that belongs to the family Geminiviriade and spreads through a single vector, the whitefly Bemisia tabaci . Although an old family of organisms, begomovirus has recently emerged as a potent threat to large-scale cultivation of crops. It is a threat with a very wide domain of influence and it can infect a very wide range of green plants cultivated for both sustenance and as cash crops. We introduce a mathematical model that simulates the complex dynamic interaction between begomovirus genetics, their adaptability to certain plants, and the availability of those plants to the virus under different cropping patterns. The model captures many empirically observed patterns of begomovirus epidemic outbursts, even if the equations do not directly depend on the genetics of begomovirus strains, and simply assume the vector to be ubiquitous. The model is formulated as a population dynamic system of differential equations, with nonlinear interactions between plant availability and begomovirus dynamic adaptability to different crop species. It also includes spatial diffusion allowing simulations over multiple neighboring regions. The ability of such a model to reproduce qualitatively correct epidemic structures indicates that the main reason for epidemic outbursts and the global spreading of the disease could be found in the patterns of inter-species interactions, many of which are human-induced. If this is the case, then the key to mitigating begomovirus epidemics would be the modification of agricultural practices. In particular, the use of intensive cropping patterns and resistant cultivars triggers aggressive virus adaptability through mutation speed-up. It seems that the only simple recourse would be to develop more diverse and less concentrated cropping patterns, both in cropland extent and in time. [ABSTRACT FROM AUTHOR]

The diversity of begomovirus and associated betasatellite complexes was analyzed from infected leaf samples of radish, tomato, chili, torai (ridge gourd), cotton, spinach, citrus and guar bean collected from different geographical regions of northern India. Leaves showing the characteristic begomovirus symptoms were used for cloning and sequencing for further characterization of the begomovirus complexes. In the present study, coat protein (CP) was amplified from eight different infected crop samples and betasatellites from only six. Our results showed significant diversity of CP and betasatellites of monopartite begomovirus in different crops in northern India. Phylogenetic analysis of CP and betasatellites test sequences exhibit a close relationship to diverse crops infecting begomovirus complexes. This strengthens the increase of host range of begomovirus in India. [ABSTRACT FROM AUTHOR]

Satellite RNAs (satRNAs) are small noncoding subviral RNA pathogens in plants that depend on helper viruses for replication and spread. Despite many decades of research, the origin of satRNAs remains unknown. In this study we show that a β-glucuronidase (GUS) transgene fused with a Cucumber mosaic virus (CMV) Y satellite RNA (Y-Sat) sequence (35S-GUS:Sat) was transcriptionally repressed in N. tabacum in comparison to a 35S-GUS transgene that did not contain the Y-Sat sequence. This repression was not due to DNA methylation at the 35S promoter, but was associated with specific DNA methylation at the Y-Sat sequence. Both northern blot hybridization and small RNA deep sequencing detected 24-nt siRNAs in wild-type Nicotiana plants with sequence homology to Y-Sat, suggesting that the N. tabacum genome contains Y-Sat-like sequences that give rise to 24-nt sRNAs capable of guiding RNA-directed DNA methylation (RdDM) to the Y-Sat sequence in the 35S-GUS:Sat transgene. Consistent with this, Southern blot hybridization detected multiple DNA bands in Nicotiana plants that had sequence homology to Y-Sat, suggesting that Y-Sat-like sequences exist in the Nicotiana genome as repetitive DNA, a DNA feature associated with 24-nt sRNAs. Our results point to a host genome origin for CMV satRNAs, and suggest novel approach of using small RNA sequences for finding the origin of other satRNAs. [ABSTRACT FROM AUTHOR]

Stromules are stroma-filled tubules, extruding from the plastid and surrounded by both envelope membranes, but so far, stromules remain enigmatic structures and their function unknown. Stromules can interconnect plastids and have been found to associate with the nucleus, endoplasmic reticulum, Golgi complex, plasma membrane, mitochondria and peroxisomes. This minireview briefly summarizes markers to visualize stromules, inducers of stromules and provides new data about plant virus induced stromules. [ABSTRACT FROM AUTHOR]

Several species of ampeloviruses and badnaviruses infect pineapple plants around the world. Pineapple mealybug wilt-associated ampeloviruses have been associated with mealybug wilt of pineapple (MWP), the major viral disease threatening this crop. Conversely, infection by the badnaviruses Pineapple bacilliform comosus virus (PBCOV) and Pineapple bacilliform erectifolius virus (PBERV) is asymptomatic. To investigate the status of infection of the pineapple crop in Cuba, a diagnostic survey was developed in commercial areas during the period 2009–2012. Incidence of MWP disease was found in up to 100% of the plants in some fields of Central and Eastern regions of the island. Molecular assays revealed the presence of PMWaV-1 for the first time in the Caribbean basin and PMWaV-2, PMWaV-3, either as mixed infections or in combination with PBCOV throughout the country. Furthermore, they revealed for the first time the presence of PMWaV-2 in Bromelia pinguin L., a plant commonly used in Cuba as hedgerow. Sequence analysis of partial heat shock protein 70h and complete coat protein gene of Cuban isolates of PMWaV-1, -2 and -3 showed nucleotide identities above 97% with cognate sequences of viruses isolated from other countries. This work discloses the presence of a complex of viruses associated with the pineapple crop in Cuba, highlights the potential role of B. pinguin in the PMWaV-mealybug-pineapple pathosystem and makes available diagnostic tools for the detection of viruses affecting pineapple for a seed certified production system in Cuba. [ABSTRACT FROM AUTHOR]

Analysis of four genomic regions from 37 geographically diverse isolates of broad bean wilt virus 1 (BBWV-1) showed high genetic diversity in comparison to most plant viruses. Comparison of synonymous and nonsynonymous substitutions of the small coat protein gene (SCP) revealed negative selection for most amino acid positions. Phylogenetic analysis of SCP showed that some BBWV-1 isolates from distant geographical areas were genetically close, suggesting long-distance migration. Analysis of genetic differentiation revealed high gene flow between Spanish and Near Eastern subpopulations, which were separated from North-Central and South-Eastern European subpopulations. Finally, putative recombinant and reassortant genomes were also identified. [ABSTRACT FROM AUTHOR]

A virus species provisionally named Grapevine virus F (GVF) was found in grapevine by deep sequencing. GVF is a vitivirus distinctly different from the other four members of this genus, as shown by nucleotide sequence and phylogenetic analysis. An RT-PCR test was developed for the detection of this virus. In a collection of 454 grapevine accessions from worldwide sources, an infection rate of 7% was found. [ABSTRACT FROM AUTHOR]

Iris yellow spot virus (IYSV) is an emerging pathogen of onion and other Allium crops worldwide. This study focused on the incidence, epidemiology and yield effects of IYSV in bulb onion crops in Spain. Surveys were conducted from 2005 to 2009. Samplings were performed in 101 onion fields with 2,677 onion plants tested in total. Onion transplants, winter onion crops and potential alternative hosts were also sampled. IYSV infection showed a temporal pattern of spread, and the proportion of sites with IYSV-infected plants began to increase rapidly after August. Two early infected fields were detected, and were the only ones with severe economic losses (50-60% crop reduction) due to IYSV. In both cases, onions were grown from IYSV-infected transplants imported from another region. There was no evidence that weeds and volunteer onion can act as alternative host. The use of virus (and vector)-free transplants must play an important role in IYSV management strategies by delaying early-season infection of onion in Spain. [ABSTRACT FROM AUTHOR]