Elliot W. Kitajima
Full Text Available Preparações feitas com exsudato de plantas de fumo, girassol, picão, erva--de-Santa-Maria, cordão-de-frade, fedegoso. Chenopodium amaranticolor e Physalis floridana, sadias e infectadas pelo vínis do mosaico do picão, foram examinadas ao microscópio electrônico. Partículas com comprimento normal aproximado de 720 mm x 12-13 mm, foram encontradas nos exsudatos das oito espécies, quando afetadas, mas não nos das plantas sadias, testemunhas. Tais partículas são consideradas como sendo o vírus causador do mosaico do picão.Electron microscopical observations were made on exudates obtained from plants of Bidrus pilosa, Chenopodium amaranticolor Leonolis nepaetifolia, Helianthus annums. Nicotiana tabucum, Cassia occidentalis, Chenopodium ambrosioides, and Physalis floridona infected with a virus that induces mosaic on the first named species. The presenes of a flexible thread with a normal length 720 mm. x 12-13 mm was recorded in the exudates from the: diseased plants, but not in those from the healthy ones, and is considered to represent the causal virus.
Savithri, HS; Murthy, MRN
The amino acid sequences of the non-structural protein (molecular weight 35,000; 3a protein) from three plant viruses - cucumber mosaic, brome mosaic and alfalfa mosaic have been systematically compared using the partial genomic sequences for these three viruses already available. The 3a protein of cucumber mosaic virus has an amino acid sequence homology of 33.7% with the corresponding protein of brome mosaic virus. A similar protein from alfalfa mosaic virus has a homology of 18.2% and 14.2...
Apple mosaic virus (ApMV), a member of the ilarvirus group, naturally infects Betula, Aesculus, Humulus, and several crop genera in the family Rosaceae (Malus, Prunus, Rosa and Rubus). ApMV was first reported in Rubus in several blackberry and raspberry cultivars in the United States and subsequentl...
Cucumber mosaic virus (CMV) has been reported on red raspberry in Chile, Scotland and the Soviet Union and in Chile on blackberry. Its occurrence in Rubus is rare and seems to cause little damage. Except for one early, unconfirmed report, CMV has not been reported on Rubus in North America. This vir...
Liao, J Y; Hu, Chung-Chi; Chen, C C; Chang, C H; Deng, T C
The full-length genome of a potyvirus, previously known as sunflower chlorotic spot virus isolate SF-1 (SCSV-SF-1) which causes novel symptoms on sunflowers (Helianthus annuus), was sequenced and analyzed. The genome of SCSV-SF-1 is 9,741 nucleotides long, encoding a polyprotein of 3,071 amino acids containing the consensus motifs of potyviruses. Sequence comparison revealed that the 3'-terminus of SCSV-SF-1 shared over 96% similarities with isolates of Bidens mottle virus (BiMoV). However, SCSV-SF-1 has a very narrow host range, excluding the diagnostic host species for BiMoV, Bidens pilosa and Zinnia elegans. Therefore, SCSV-SF-1 is a distinct isolate of BiMoV. This is the first report of the full-length nucleotide sequence of BiMoV infecting sunflower in Taiwan.
Tollin, P; Bancroft, J B; Richardson, J F; Payne, N C; Beveridge, T J
X-ray and optical diffraction studies of the flexuous papaya mosaic virus are described. The virus is constructed so that there are 35 coat protein subunits in 4 turns of the helix. The virus contains about 1410 protein subunits and 6800 nucleotides and has a molecular weight of about 33 x 10(6). The structure of tubes assembled in vitro from coat protein both in the presence and absence of nucleic acid resembles that of the native virus.
McDaniel, Larry; Maratos, Marina; Farabaugh, Joan
Provides three exercises that introduce high school and college students to a common strain of the tobacco mosaic virus and the study of some basic biological processes. Activities involve inoculation of plants and observing and recording symptom development in infected plants. (DDR)
Ormrod, D.P.; Kemp, W.G.
The sensitivity of three tomato cultivars to several concentrations of ozone was evaluated after prior sequential inoculations with tobacco mosaic virus (TMV) and/or cucumber mosaic virus (CMV). Ozone injury in inoculated and uninoculated tomatoes varied from slight to severe depending on the virus, cultivar, ozone concentration and virus incubation period. The frequency of increased ozone injury was about twice as great as that of suppressed injury on infected plants. Ozone injury occurred more frequently in TMV-inoculated plants than in those inoculated with CMV. There were more increases than decreases in ozone injury after 7 or 14 days of virus infection, but mainly decreases in injury after 21 days infection. Growth was significantly reduced in plants exposed to ozone after a 21-day virus incubation period, particularly when they were inoculated with both viruses.
A 2014 nationwide survey of the distribution of Soybean mosaic virus (SMV), Soybean yellow mottle mosaic virus (SYMMV) and Soybean yellow common mosaic virus (SYCMV) major viruses in South Korean soybean fields, and changes
In 2014 symptomatic soybean samples were collected throughout Korea, and were tested for the most important soybean viruses found in Korea, namely Soybean mosaic virus (SMV), Soybean yellow common mosaic virus (SYCMV), and Soybean yellow mottle mosaic virus (SYMMV). SYMMV was most commonly detected,...
Erickson, J W; Hallett, F R; Bancroft, J B
An examination of the number of subunits in small aggregates of papaya mosaic virus (PMV) coat protein is presented based on a model system which gives results consistent with the experimental observation that the 14 S subassembly species is a double disc, composed of two rows of nine subunits each. The estimated hydration of the disc, about 0.85 g 1H20/9 protein, is unusually large and indicates a cavitated structure for the disc. Comparison with other rod-shaped viruses suggests that the flexuous nature of PMV is a consequence of sparse axial inter-subunit contacts at high radius.
Full Text Available A strain of maize dwarf mosaic virus related to sugarcane mosaic virus has been isolated from maize in Burundi. The properties (including electron microscopy and serology of the virus are described, and elements for a control strategy are reviewed.
Rajapakse, R H; Herath, H M
75 plant species from 11 families were tested in Sri Lanka for their susceptibility to transferring the papaya mosaic virus. After inoculation with this virus, six species, Cucurbita pepo, Cucumis sativus, Nicotiana tabacum, Chenopodium amaranticolor, Gomphrena globosa and Lycopersicum esculentum, developed such symptoms, and after re-isolation from the host plant the virus again infected papaya plants. Thus these species are possible alternate hosts of papaya mosaic virus in Sri Lanka.
de Assis Filho, F M; Sherwood, J L
ABSTRACT The mechanism of virus transmission through seed was studied in Arabidopsis thaliana infected with Turnip yellow mosaic virus (TYMV) and Tobacco mosaic virus (TMV). Serological and biological tests were conducted to identify the route by which the viruses reach the seed and subsequently are located in the seed. Both TYMV and TMV were detected in seed from infected plants, however only TYMV was seed-transmitted. This is the first report of transmission of TYMV in seed of A. thaliana. Estimating virus seed transmission by grow-out tests was more accurate than enzyme-linked immunosorbent assay due to the higher frequency of antigen in the seed coat than in the embryo. Virus in the seed coat did not lead to seedling infection. Thus, embryo invasion is necessary for seed transmission of TYMV in A. thaliana. Crosses between healthy and virus-infected plants indicated that TYMV from either the female or the male parent could invade the seed. Conversely, invasion from maternal tissue was the only route for TMV to invade the seed. Pollination of flowers on healthy A. thaliana with pollen from TYMV-infected plants did not result in systemic infection of healthy plants, despite TYMV being carried by pollen to the seed.
Sit, T L; Abouhaidar, M G; Holy, S
The RNA genome of papaya mosaic virus is 6656 nucleotides long [excluding the poly(A) tail] with six open reading frames (ORFs) more than 200 nucleotides long. The four nearest the 5' end each overlap with adjacent ORFs and could code for proteins with Mr 176307, 26248, 11949 and 7224 (ORFs 1 to 4). The fifth ORF produces the capsid protein of Mr 23043 and the sixth ORF, located completely within ORF1, could code for a protein with Mr 14113. The translation products of ORFs 1 to 3 show strong similarity with those of other potexviruses but the ORF 4 protein has only limited similarity with the other potexvirus ORF 4 proteins of 7K to 11K.
Geering, Andrew D W; Thomas, John E; Holton, Timothy; Hadfield, James; Varsani, Arvind
Three monocot-infecting mastreviruses from Australia, all found primarily in pasture and naturalised grasses, have been characterised at the molecular level. Here, we present the full genome sequence of a fourth, Paspalum striate mosaic virus (PSMV), isolated from Paspalum dilatatum from south-east Queensland. The genome was 2816 nt long and had an organisation typical of other monocot-infecting mastreviruses. Its nearest relative is Bromus cartharticus striate mosaic virus (BCSMV), with which it shares an overall genome identity of 75%. Phylogenetic analysis of the complete genome and each of the putative viral proteins places PSMV in a group with the other three Australian striate mosaic viruses. PSMV, BCSMV and Digitaria didactyla striate mosaic virus all contain a similar, small recombinant sequence in the small intergenic region.
程晔; 陈炯; 陈剑平
The complete sequence of a potyvirus from maize in Zhejiang Province was determined. The RNA was 9596 nucleotides long, excluding the 3′-poly (A) tail, and there was a single long open reading frame (ORF) of 9192 nts encoding a 346.1 ku polyprotein. The polyprotein had substantial amino acid sequence homology with those encoded by the RNAs of a Chinese isolate of sorghum mosaic virus (SrMV-C) and a Bulgarian isolate of maize dwarf mosaic virus, but it was most closely related to sugarcane mosaic virus (SCMV) isolates, for which only partial sequences have been published. According to the published criteria for distinguishing potyviruses, the sequence reported here is clearly a strain of SCMV, but it also showed a surprisingly high amino acid homology with SrMV-C in the HC-Pro, P3 and CI proteins.
Chen, Y.K.; Derks, A.F.L.M.; Langeveld, S.; Goldbach, R.; Prins, M.
For classification of Cucumber mosaic virus (CMV) isolates from ornamental crops of different geographical areas, these were characterized by comparing the nucleotide sequences of RNAs 4 and the encoded coat proteins. Within the ornamental-infecting CMV viruses both subgroups were represented. CMV i
Moreno Pérez, Manuel Guillermo; Pagán Muñoz, Jesús Israel; Aragón Caballero, Liliana; Cáceres, Fátima; Fraile Pérez, Aurora; García Arenal, Fernando
Virus emergence is a complex phenomenon, which generally involves spread to a new host from a wild host, followed by adaptation to the new host. Although viruses account for the largest fraction of emerging crop pathogens, knowledge about their emergence is incomplete. We address here the question of whether Pepino mosaic virus (PepMV) emergence as a major tomato pathogen worldwide could have involved spread from wild to cultivated plant species and host adaptation. For this, we surveyed natu...
Love, Andrew J; Makarov, Valentine; Yaminsky, Igor; Kalinina, Natalia O; Taliansky, Michael E
Due to the nanoscale size and the strictly controlled and consistent morphologies of viruses, there has been a recent interest in utilizing them in nanotechnology. The structure, surface chemistries and physical properties of many viruses have been well elucidated, which have allowed identification of regions of their capsids which can be modified either chemically or genetically for nanotechnological uses. In this review we focus on the use of such modifications for the functionalization and production of viruses and empty viral capsids that can be readily decorated with metals in a highly tuned manner. In particular, we discuss the use of two plant viruses (Cowpea mosaic virus and Tobacco mosaic virus) which have been extensively used for production of novel metal nanoparticles (<100nm), composites and building blocks for 2D and 3D materials, and illustrate their applications.
Full Text Available Abstract Background An effective method for obtaining resistant transgenic plants is to induce RNA silencing by expressing virus-derived dsRNA in plants and this method has been successfully implemented for the generation of different plant lines resistant to many plant viruses. Results Inverted repeats of the partial Tobacco mosaic virus (TMV movement protein (MP gene and the partial Cucumber mosaic virus (CMV replication protein (Rep gene were introduced into the plant expression vector and the recombinant plasmids were transformed into Agrobacterium tumefaciens. Agrobacterium-mediated transformation was carried out and three transgenic tobacco lines (MP16-17-3, MP16-17-29 and MP16-17-58 immune to TMV infection and three transgenic tobacco lines (Rep15-1-1, Rep15-1-7 and Rep15-1-32 immune to CMV infection were obtained. Virus inoculation assays showed that the resistance of these transgenic plants could inherit and keep stable in T4 progeny. The low temperature (15℃ did not influence the resistance of transgenic plants. There was no significant correlation between the resistance and the copy number of the transgene. CMV infection could not break the resistance to TMV in the transgenic tobacco plants expressing TMV hairpin MP RNA. Conclusions We have demonstrated that transgenic tobacco plants expressed partial TMV movement gene and partial CMV replicase gene in the form of an intermolecular intron-hairpin RNA exhibited complete resistance to TMV or CMV infection.
The complete sequence of the two RNAs of a furovirus isolate fromdurum wheat in Italy was determined. Sequence comparisons and phylogenetic analysis were done to compare the Italian virus with Soil-borne wheat mosaic virus (SBWMV) from the USA and with furovirus sequences recently published as European wheat mosaic virus (EWMV), from wheat in France, and Soil-borne rye mosaic virus (SBRMV), from rye and wheat in Germany. Over the entire genome, the Italian isolate RNA1 and RNA2 had respectively 97.5% and 98.6% nucleotide identity with EWMV, 95.5% and 85.8% with SBRMV-G and 70.6% and 64.5% with SBWMV. The Italian isolate was therefore clearly distinct from SBWMV. The European isolates all appear to belong to the same virus and the name Soil-borne cereal mosaic virus may resolve earlier ambiguities.
A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for detecting Sugarcane mosaic virus (SCMV) and Sorghum mosaic virus (SrMV) in sugarcane. Six sets of four primers corresponding to the conserved coat protein gene were designed for each virus and their succ...
Wit, de J.L.
This Thesis describes the application of conventional 13 C and 1 H high resolution Fourier Transform Nuclear Magnetic resonance (HR FT NMR) to Tobacco Mosaic Virus (TMV) and its protein oligo- and polymers and some other largebiological systems. The rod-like (TMV) consists of 2
Soilborne wheat mosaic virus (SBWMV) is a bipartite single stranded positive sense RNA virus with rigid-rod shaped virions. Taxonomically the virus is in the family Viragviridae, as are commonly used gene silencing or expression viral vectors, Tobacco rattle virus (TRV) and Barley stripe mosaic viru...
Qusus, Saba J.
In the U.S., soybean mosaic virus (SMV) is classified into seven strain groups, designated G1 to G7, based on their different responses on resistant soybean [Glycine max (L.) Merr.] cultivars. These responses are: symptomless or resistant (R), necrotic (N), and mosaic or susceptible (S). The gene-for-gene model has been proposed for SMV-soybean interactions. In the majority of cultivars, a single dominant gene, Rsv1, confers both the R and N responses. In the first part of this study, the coa...
Monika Fecury Moura
Full Text Available Weeds can act as important reservoirs for viruses. Solanum americanum (Black nightshade is a common weed in Brazil and samples showing mosaic were collected from sweet pepper crops to verify the presence of viruses. One sample showed mixed infection between Cucumber mosaic virus (CMV and Potato virus Y (PVY and one sample showed simple infection by PVY. Both virus species were transmitted by plant extract and caused mosaic in tomato (Solanum lycopersicum cv. Santa Clara, sweet pepper (Capsicum annuum cv. Magda, Nicotiana benthamiana and N. tabaccum TNN, and local lesions on Chenopodium quinoa, C. murale and C. amaranticolor. The coat protein sequences for CMV and PVY found in S. americanum are phylogenetically more related to isolates from tomato. We conclude that S. americanum can act as a reservoir for different viruses during and between sweet pepper crop seasons.
Lin, Ching-Yi; Ku, Hsin-Mei; Chiang, Yi-Hua; Ho, Hsiu-Yin; Yu, Tsong-Ann; Jan, Fuh-Jyh
Watermelon, an important fruit crop worldwide, is prone to attack by several viruses that often results in destructive yield loss. To develop a transgenic watermelon resistant to multiple virus infection, a single chimeric transgene comprising a silencer DNA from the partial N gene of Watermelon silver mottle virus (WSMoV) fused to the partial coat protein (CP) gene sequences of Cucumber mosaic virus (CMV), Cucumber green mottle mosaic virus (CGMMV) and Watermelon mosaic virus (WMV) was constructed and transformed into watermelon (cv. Feeling) via Agrobacterium-mediated transformation. Single or multiple transgene copies randomly inserted into various locations in the genome were confirmed by Southern blot analysis. Transgenic watermelon R(0) plants were individually challenged with CMV, CGMMV or WMV, or with a mixture of these three viruses for resistance evaluation. Two lines were identified to exhibit resistance to CMV, CGMMV, WMV individually, and a mixed inoculation of the three viruses. The R(1) progeny of the two resistant R(0) lines showed resistance to CMV and WMV, but not to CGMMV. Low level accumulation of transgene transcripts in resistant plants and small interfering (si) RNAs specific to CMV and WMV were readily detected in the resistant R(1) plants by northern blot analysis, indicating that the resistance was established via RNA-mediated post-transcriptional gene silencing (PTGS). Loss of the CGMMV CP-transgene fragment in R1 progeny might be the reason for the failure to resistant CGMMV infection, as shown by the absence of a hybridization signal and no detectable siRNA specific to CGMMV in Southern and northern blot analyses. In summary, this study demonstrated that fusion of different viral CP gene fragments in transgenic watermelon contributed to multiple virus resistance via PTGS. The construct and resistant watermelon lines developed in this study could be used in a watermelon breeding program for resistance to multiple viruses.
Adams, I P; Rai, S; Deka, M; Harju, V; Hodges, T; Hayward, G; Skelton, A; Fox, A; Boonham, N
The 9573-nucleotide genome of a potyvirus was sequenced from a Coriandrum sativum plant from India with viral symptoms. On analysis, this virus was shown to have greater than 85 % nucleotide sequence identity to vanilla distortion mosaic virus (VDMV). Analysis of the putative coat protein sequence confirmed that this virus was in fact VDMV, with greater than 91 % amino acid sequence identity. The genome appears to encode a 3083-amino-acid polyprotein potentially cleaved into the 10 mature proteins expected in potyviruses. Phylogenetic analysis confirmed that VDMV is a distinct but ungrouped member of the genus Potyvirus.
Haq, Q. M. I.; Ali, Arif; Malathi, V.G.
Yellow mosaic disease of cultivated legumes in South-East Asia, is caused by Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV) belonging to the genus Begomovirus of the family Geminiviridae. Efforts to engineer resistance against the genus Begomovirus are focused mainly on silencing of complementary-sense virus genes involved in virus replication. Here we have targeted a complementary-sense gene (ACI) encoding Replication initiation Protein (Rep) to develop re...
Seo, Eun–Young; Cho, Seunghee; Moon, Jae Sun; Gotoh, Takafumi; Goto, Takafumi; Kim, Hong Gi; Domier, Leslie L; Lim, Seungmo; Kim, Kil Hyun; Moon, Jung–Kyung; Hammond, John; Lim, Hyoun–Sub; Song, Ki Hak
Soybean yellow common mosaic virus (SYCMV) was recently reported from Korea, and a subsequent survey of soybean fields found that SYCMV, Soybean yellow mottle mosaic virus (SYMMV), and Soybean mosaic virus (SMV) infections were widespread. SYCMV has recently been developed into a Virus Inducing Gene Silencing (VIGS) vector for use as a reverse genetics tool for soybean, and here we report a modified SYCMV VIGS vector containing a new restriction enzyme site in the 3’ non–coding region into wh...
Gell, Gyöngyvér; Sebestyén, Endre; Balázs, Ervin
Recombination among RNA viruses is a natural phenomenon that appears to have played a significant role in the species development and the evolution of many strains. It also has particular significance for the risk assessment of plants which have been genetically modified for disease resistance by incorporating viral sequences into their genomes. However, the exact recombination events taking place in viral genomes are not investigated in detail for many virus groups. In this analysis, different single-stranded positive-sense RNA potyviruses were compared using various in silico recombination detection methods and new recombination events in the Sugarcane mosaic virus (SCMV) subgroup were detected. For an extended in silico recombination analysis, two of the analyzed Maize dwarf mosaic virus full-length genomes were sequenced additionally during this work. These results strengthen the evidence that recombination is a major driving force in virus evolution, and the emergence of new virus variants in the SCMV subgroup, paired with mutations, could generate viruses with altered biological properties. The intra- and interspecific homolog recombinations seem to be a general trait in this virus group, causing little or no changes to the amino acid of the progenies. However, we found a few breakpoints between the members of SCMV subgroup and the weed-infecting distant relatives, but only a few methods of the RDP3 package predicted these events with low significance level.
Wheat streak mosaic virus (WSMV) and Triticum mosaic virus (TriMV) are transmitted by the wheat curl mite (WCM, Aceria tosichella Keifer). Previous work has shown that different mite genotypes transmit TriMV at different rates. The objective of this research was to determine if mite genotypes differ...
Liu, Chengke; Nelson, Richard S
Successful systemic infection of a plant by Tobacco mosaic virus (TMV) requires three processes that repeat over time: initial establishment and accumulation in invaded cells, intercellular movement, and systemic transport. Accumulation and intercellular movement of TMV necessarily involves intracellular transport by complexes containing virus and host proteins and virus RNA during a dynamic process that can be visualized. Multiple membranes appear to assist TMV accumulation, while membranes, microfilaments and microtubules appear to assist TMV movement. Here we review cell biological studies that describe TMV-membrane, -cytoskeleton, and -other host protein interactions which influence virus accumulation and movement in leaves and callus tissue. The importance of understanding the developmental phase of the infection in relationship to the observed virus-membrane or -host protein interaction is emphasized. Utilizing the latest observations of TMV-membrane and -host protein interactions within our evolving understanding of the infection ontogeny, a model for TMV accumulation and intracellular spread in a cell biological context is provided.
Mackie, G A; Johnston, R; Bancroft, J B
Three classes of viral RNA were recovered from polyribosomes purified from papaya leaves infected with papaya mosaic virus (PapMV) and from barley leaves infected with foxtail mosaic virus (FoMV): full-length viral RNAs [6.8 and 6.2 kilobases (kb), respectively]; less abundant intermediate subgenomic RNAs (2.2 and 1.9 kb), and abundant, small subgenomic RNAs (1 and 0.9 kb). Small amounts of the PapMV-specified 1.0-kb subgenomic RNA were encapsidated, whereas no encapsidated subgenomic RNAs could be found in preparations of FoMV. Immunoprecipitation of the products of in vitro translation of the small subgenomic RNA of both viruses showed that it codes for the corresponding viral coat protein. FoMV genomic RNA isolated from polyribosomes also directed the efficient synthesis of a 37- to 38-kilodalton protein which was immunoprecipitated by an antiserum raised against the coat protein. We presume this product to be a readthrough protein initiated to the 5' side of and in the same reading frame as the coat protein-coding sequences in FoMV RNA. The predominant double-stranded viral-specified RNAs in tissues infected with PapMV, FoMV, and clover yellow mosaic virus were genome sized (6.8, 6.2, and 7.0 kb pairs, respectively). If double-stranded RNAs corresponding to coat protein subgenomic RNAs exist, they must be present in much lower relative abundances.
For many years, Tobacco mosaic virus (TMV) and Tomato mosaic virus (ToMV) are the two major tobamoviruses that have a serious impact on tomato productions worldwide. These seed-borne and mechanically transmitted viruses are difficult to control. The most effective disease management has been the u...
Chen, Y K; Derks, A F; Langeveld, S; Goldbach, R; Prins, M
For classification of Cucumber mosaic virus (CMV) isolates from ornamental crops of different geographical areas, these were characterized by comparing the nucleotide sequences of RNAs 4 and the encoded coat proteins. Within the ornamental-infecting CMV viruses both subgroups were represented. CMV isolates of Alstroemeria and crocus were classified as subgroup II isolates, whereas 8 other isolates, from lily, gladiolus, amaranthus, larkspur, and lisianthus, were identified as subgroup I members. In general, nucleotide sequence comparisons correlated well with geographic distribution, with one notable exception: the analyzed nucleotide sequences of 5 lily isolates showed remarkably high homology despite different origins.
Goldbach, Rob; Krijt, Jette
The protease encoded by the large (B) RNA segment of cowpea mosaic virus was tested for its ability to recognize the in vitro translation products of the small (M) RNA segment from the comoviruses squash mosaic virus, red clover mottle virus, and cowpea severe mosaic virus (CPsMV, strains Dg and Ark), and from the nepovirus tomato black ring virus. Like M RNA from cowpea mosaic virus, the M RNAs from squash mosaic virus, red clover mottle virus, CPsMV-Dg, and CPsMV-Ark were all translated int...
Overview:The aim of the studies described in this thesis was to obtain a thorough understanding of the main factors determining the spread of a potyvirus in a high plant density crop. The factors studied included the relationships between virus, host and vector, the spread of the vi
Haq, Q M I; Ali, Arif; Malathi, V G
Yellow mosaic disease of cultivated legumes in South-East Asia, is caused by Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV) belonging to the genus Begomovirus of the family Geminiviridae. Efforts to engineer resistance against the genus Begomovirus are focused mainly on silencing of complementary-sense virus genes involved in virus replication. Here we have targeted a complementary-sense gene (ACI) encoding Replication initiation Protein (Rep) to develop resistance against soybean isolate of Mungbean yellow mosaic India virus-[India:New Delhi:Soybean 2:1999], a bipartite begomovirus prevalent throughout the Indian subcontinent. We show that the legume host plants co-agroinoculated with infectious constructs of soybean isolate of Mungbean yellow mosaic India virus [India:New Delhi:Soybean 2:1999] along with this antisense Rep gene construct show resistance to the virus.
Valentin V Makarov
Full Text Available Virions of Barley stripe mosaic virus (BSMV were neglected for more than thirty years after their basic properties were determined. In this paper, the physicochemical characteristics of BSMV virions and virion-derived viral capsid protein (CP were analyzed, namely, the absorption and intrinsic fluorescence spectra, circular dichroism spectra, differential scanning calorimetry curves, and size distributions by dynamic laser light scattering. The structural properties of BSMV virions proved to be intermediate between those of Tobacco mosaic virus (TMV, a well-characterized virus with rigid rod-shaped virions, and flexuous filamentous plant viruses. The BSMV virions were found to be considerably more labile than expected from their rod-like morphology and a distant sequence relation of the BSMV and TMV CPs. The circular dichroism spectra of BSMV CP subunits incorporated into the virions, but not subunits of free CP, demonstrated a significant proportion of beta-structure elements, which were proposed to be localized mostly in the protein regions exposed on the virion outer surface. These beta-structure elements likely formed during virion assembly can comprise the N- and C-terminal protein regions unstructured in the non-virion CP and can mediate inter-subunit interactions. Based on computer-assisted structure modeling, a model for BSMV CP subunit structural fold compliant with the available experimental data was proposed.
Kumari, Reenu; Bhardwaj, Pooja; Singh, Lakhmir; Zaidi, Aijaz A; Hallan, Vipin
Cucumber mosaic virus (CMV) has a wide host range causing severe damage in many important agricultural and ornamental crops. Earlier reports showed the prevalence of CMV subgroup I isolates in India. However, some recent reports point towards increasing incidence of subgroup II isolates in the country. The complete genome of a CMV isolate causing severe mosaic in cucumber was characterized and its phylogenetic analysis with other 21 CMV isolates reported worldwide clustered it with subgroup II strains. The genome comprised of RNA 1 (3,379 nucleotides), RNA 2 (3,038 nucleotides) and RNA 3 (2,206 nucleotides). The isolate showed highest homology with subgroup II isolates: 95.1-98.7, 87.7-98.0, and 85.4-97.1 % within RNA1, RNA2, and RNA3, respectively. RNA1 and RNA2 were closely related to the Japanese isolate while RNA3 clustered with an American isolate. Host range studies revealed that isolate showed severe mosaic symptoms on Nicotiana spp. and Cucumis spp. The isolate induced leaf deformation and mild filiform type symptoms in tomato. To best of our knowledge this is the first report of complete genome of CMV subgroup II isolate from India.
Full Text Available Watermelon mosaic virus (WMV is widespread in cucurbit crops, most commonly occuring in temperate and Mediterranean regions. In Serbia WMV has been detected in single and mixed infections with Zucchini yellow mosaic virus and Cucumber mosaic virus in field-grown pumpkin and squash crops. Among pumpkin-affecting viruses WMV is the most frequent one, both by the number of localities and its incidence at each location. During the growing season of 2009, samples from 583 plants of Cucurbita pepo cvs. Olinka, Belgrade zucchini and Tosca (Zucchini group, as well as from C. maxima and C. moschata showing symptoms of virus infection were collected from 12 commercial fields at eight localities and analyzed by DAS-ELISA using polyclonal antisera specific to six most important cucurbit viruses. Interestingly, WMV was detected at fewer sites and had lower ncidence rate than in two previous years. In single infections, WMV was found in 11% of tested plants in three fields; in mixed infections with ZYMV, it was recorded in 9.9% of plants in five fields and with CMV in only 0.2% in one field. The partial coat protein gene and 3’ non-translated region from two representativeisolates of WMV originating from different localities and host plant species were amplified by RT-PCR, sequenced, and compared with the sequences available in GenBank database. The PCR-amplified fragment of predicted size of approximately 1017 bp was obtained. The sequences of isolates 137-08 (Acc. No. GQ259958 and 159-08 (GU144020 proved to be 94-99% identical at the nucleotide level with those from other parts of the world. The sequences of these two isolates differed from each other only at two nucleotide positions, without any amino acid substitution. Phylogenetic analysis of 57 isolates based on 750 bp sequences of the coat protein gene showed no correlation between isolates and their geographic origin, and italso indicated that these isolates fell into three molecular groups of
Raski, D J; Maggenti, A R; Jones, N O
Particles of fanleaf and yellow mosaic viruses are reported in the lumen of the esophagus of Xiphinerna index. Differences in cuticular morphology suggest differences in charged receptor sites which may offer an explanation for virus location and orderly arrangement.
Bernal, J J; Jiménez, I; Moreno, M; Hord, M; Rivera, C; Koenig, R; Rodríguez-Cerezo, E
ABSTRACT Chayote mosaic virus (ChMV) is a putative tymovirus isolated from chayote crops in Costa Rica. ChMV was characterized at the host range, serological, and molecular levels. ChMV was transmitted mechanically and induced disease symptoms mainly in Cucurbitaceae hosts. Asymptomatic infections were detected in other host families. Serologically, ChMV is related to the Andean potato latent virus (APLV) and the Eggplant mosaic virus (EMV), both members of the genus Tymovirus infecting solanaceous hosts in the Caribbean Basin and South America. The sequence of the genomic RNA of ChMV was determined and its genetic organization was typical of tymoviruses. Comparisons with other tymoviral sequences showed that ChMV was a new member of the genus Tymovirus. The phylogenetic analyses of the coat protein gene were consistent with serological comparisons and positioned ChMV within a cluster of tymoviruses infecting mainly cucurbit or solanaceous hosts, including APLV and EMV. Phylogenetic analyses of the replicase protein gene confirmed the close relationship of ChMV and EMV. Our results suggest that ChMV is related to two tymoviruses (APLV and EMV) of proximal geographical provenance but with different natural host ranges. ChMV is the first cucurbit-infecting tymovirus to be fully characterized at the genomic level.
S.Obaiah; B.V. Bhaskara Reddy; N P Eswara Reddy; K. Vijay Krishna Kumar
Blackgram (Vigna mungo (L.) Hepper) is one of the major pulse crops of the tropics and sub tropics. It is the third major pulse crop cultivated in the Indian subcontinent. Pulses and grain legumes are major sources of dietary protein. These crops are subjected to yellow mosaic and golden mosaic diseases caused by white fly transmitted geminiviruses (WTG’s or begomovirus). Of these viruses, mungbean yellow mosaic virus (MYMV) is an important one, and it infects five major leguminous species...
Hollander, den P.W.
Abstract of thesis Paulus den Hollander entitled “On the involvement of host proteins in Cowpea mosaic virus intercellular spread”. Defence: 18th of November 13.30 h Abstract Intercellular spread of Cowpea mosaic virus (CPMV) occurs via movement tubules inserted into the
Yildiz, Ibrahim; Tsvetkova, Irina; Wen, Amy M.; Shukla, Sourabh; Masarapu, M. Hema; Dragnea, Bogdan; Steinmetz, Nicole F.
Viral nanoparticles (VNPs) are becoming versatile tools in platform technology development. Their well-defined structures as well as their programmability through chemical and genetic modification allow VNPs to be engineered for potential imaging and therapeutic applications. In this article, we report the application of a variety of bioconjugation chemistries to the plant VNP Brome mosaic virus (BMV). Functional BMV nanoparticles displaying multiple copies of fluorescent dyes, PEG molecules, chemotherapeutic drug moieties, targeting proteins and cell penetrating peptides were formulated. This opens the door for the application of BMV in nanomedicine. PMID:28018580
Full Text Available Cucumber mosaic virus (CMV is considered one of the most economically importantplant viruses and has a worldwide distribution and a very wide host range including plantsfrom family Cucurbitaceae. In Serbia, on cucurbits CMV was detected in single and mixedinfections with Zucchini yellow mosaic virus (ZYMV and Watermelon mosaic virus (WMV. Viruses,including CMV, are constantly present in cucurbit crops, but their frequency changesby year and locality. Surveys and sample collections were conducted in cucurbit crops inthe period from 2008 to 2009 at 15 localities in Vojvodina province, and sample testing wascarried out using the DAS-ELISA method and commercially available antisera for six economicallymost important cucurbit viruses. In 2008, a total of 51 samples were collected from13 cucurbit crops of oilseed pumpkin Olinka variety, squash, and bottle gourd and CMV wasdetected in a total of 55% of tested samples with symptoms of viral infection. The most commoninfectious type was mixed infection with ZYMV and WMV (35.3%, and then mixedinfection with ZYMV (17.7% and WMV (2%. A total of 599 symptomatic samples of oilseedpumpkin Olinka variety, zucchini squash varieties Beogradska and Tosca, squash, and wintersquash were collected in 15 cucurbits crops in 2009. CMV was present in 4.4% of totalcollected samples, in single infections in 1.3%, and in mixed with WMV or ZYMV in 1.3%, and1.8%. Five CMV isolates were obtained by mechanical inoculations of N. glutinosa and oneof them was selected for further biological characterization. Test plants which were describedto be hosts of CMV expressed symptoms characteristic for those caused by CMV afterinoculations by isolate 115-08. CMV specific primers Au1u/Au2d were used to amplify an850 bp fragment using RT-PCR method. Amplified fragment encodes the entire viral coatprotein (CP gene and partial 5’ and 3’ UTRs of two selected CMV isolates. Amplified fragmentswere sequenced and deposited in the NCBI, where
... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Coat protein of cucumber mosaic virus...-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.516 Coat protein of cucumber mosaic virus; exemption from the requirement of a tolerance. Residues of Coat Protein of Cucumber Mosaic Virus are...
Abouhaidar, M G; Bancroft, J B
The problem of the rapid multiinitiation of papaya mosaic virus or tobacco mosaic virus RNA by PMV-protein near pH 7.0 at low ionic strength has been overcome. If NaCl is added to 0.1 M, both RNAs are first encapsidated at their respective 5' ends. This shows that the initial site of helix formation depends on the protein rather than the RNA.
Farreyrol, K; Pearson, M N; Grisoni, M; Cohen, D; Beck, D
Sequence was determined for the coat protein (CP) gene and 3' non-translated region (3'NTR) of two vanilla mosaic virus (VanMV) isolates from Vanilla tahitensis, respectively from the Cook Islands (VanMV-CI) and French Polynesia (VanMV-FP). Both viruses displayed distinctive features in the N-terminal region of their CPs; for VanMV-CI, a 16-amino-acid deletion including the aphid transmission-related DAG motif, and for VanMV-FP, a stretch of GTN repeats that putatively belongs to the class of natively unfolded proteins. VanMV-FP CP also has a novel DVG motif in place of the DAG motif, and an uncommon Q//V protease cleavage site. The sequences were compared to a range of Dasheen mosaic virus (DsMV) strains and to potyviruses infecting orchids. Identity was low to DsMV strains across the entire CP coding region and across the 3'NTR, but high across the CP core and the CI-6K2-NIa region. In accordance with current ICTV criteria for species demarcation within the family Potyviridae, VanMV-CI and VanMV-FP are strains of DsMV that exclusively infect vanilla.
Mochizuki, Tomofumi; Nobuhara, Shinya; Nishimura, Miho; Ryang, Bo-Song; Naoe, Masaki; Matsumoto, Tadashi; Kosaka, Yoshitaka; Ohki, Satoshi T
We investigated the synergistic effects of co-infection by zucchini yellow mosaic virus (ZYMV) and cucumber mosaic virus (CMV) on viral distribution in the vascular tissues of cucumber. Immunohistochemical observations indicated that ZYMV was present in both the phloem and xylem tissues. ZYMV-RNA was detected in both the xylem wash and guttation fluid of ZYMV-inoculated cucumber. Steam treatment at a stem internode indicated that ZYMV enters the xylem vessels and moves through them but does not cause systemic infection in the plant. CMV distribution in singly infected cucumbers was restricted to phloem tissue. By contrast, CMV was detected in the xylem tissue of cotyledons in plants co-infected with CMV and ZYMV. Although both ZYMV-RNA and CMV-RNA were detected in the xylem wash and upper internodes of steam-treated, co-infected cucumbers grown at 24 °C, neither virus was detected in the upper leaves using an ELISA assay. Genetically modified CMV harboring the ZYMV HC-Pro gene was distributed in the xylem and phloem tissues of singly inoculated cucumber cotyledons. These results indicate that the ZYMV HC-Pro gene facilitates CMV entry into the xylem vessels of co-infected cucumbers.
Full Text Available Successful systemic infection of a plant by Tobacco mosaic virus (TMV requires three processes that repeat over time: initial establishment and accumulation in invaded cells, intercellular movement and systemic transport. Accumulation and intercellular movement of TMV necessarily involves intracellular transport by complexes containing virus and host proteins and virus RNA during a dynamic process that can be visualized. Multiple membranes appear to assist TMV accumulation, while membranes, microfilaments and microtubules appear to assist TMV movement. Here we review cell biological studies that describe TMV-membrane, -cytoskeleton and -other host protein interactions which influence virus accumulation and movement in leaves and callus tissue. The importance of understanding the developmental phase of the infection in relationship to the observed virus-membrane or -host protein interaction is emphasized. Utilizing the latest observations of TMV-membrane and -host protein interactions within our evolving understanding of the infection ontogeny, a model for TMV accumulation and intracellular spread in a cell biological context is provided.
Full Text Available Over the past decade, intensive spread of virus infections of oilseed pumpkin has resulted in significant economic losses in pumpkin crop production, which is currently expanding in our country. In 2007 and 2008, a survey for the presence and distribution of oilseed pumpkin viruses was carried out in order to identify viruses responsible for epidemics and incidences of very destructive symptoms on cucurbit leaves and fruits. Monitoring andcollecting samples of oil pumpkin, as well as other species such as winter and butternut squash and buffalo and bottle gourd with viral infection symptoms, was conducted in several localities of Vojvodina Province. The collected plant samples were tested by DAS-ELISA using polyclonal antisera specific for the detection of six most economically harmful pumpkin viruses: Cucumber mosaic virus (CMV, Zucchini yellow mosaic virus (ZYMV, Watermelon mosaic virus (WMW, Squash mosaic virus (SqMV, Papaya ringspot virus (PRSV and Tobaccoringspot virus (TRSV that are included in A1 quarantine list of harmful organisms in Serbia.Identification of viruses in the collected samples indicated the presence of three viruses, ZYMV, WMV and CMV, in individual and mixed infections. Frequency of the identified viruses varied depending on locality and year of investigations. In 2007, WMV was the most frequent virus (94.2%, while ZYMV was prevalent (98.04% in 2008. High frequency of ZYMV determined in both years of investigation indicated the need for its rapid and reliable molecular detection. During this investigation, a protocol for ZYMVdetection was developed and optimized using specific primers CPfwd/Cprev and commercial kits for total RNA extraction, as well as for RT-PCR. In RT-PCR reaction using these primers, a DNA fragment of approximately 1100 bp, which included coat protein gene, was amplified in the samples of infected pumkin leaves. Although serological methods are still useful for large-scale testing of a great number of
The recombinant clones of pTN and pNT have been constructed by exchanging the coding regions of the movement proteins (MP), coat proteins (CP) and 3′noncoding regions between the cDNAs of the tobacco mosaic virus (Chinese Isolate, TMV-Cv) and the attenuated tomato mosaic virus N14 genomes, and used as templates for in vitro runoff transcription. Their transcripts have been used for tobacco infection assays. The infection results show that the transcripts of pTN and pNT are infectious. Local lesions were observed in the leaves of Nicotiana tabacum cv. Samsun NN inoculated with pTN transcript, but were fewer than those in the same kind of plant induced by pTMV-Cv transcript. Systemic symptoms were also observed in N. tabacum cv. Huangmiaoyu induced by pTN transcript, but were slighter than those on the same kind of tobacco induced by pTMV-Cv transcript. Local lesions were shown in N. tabacum cv. Samsun NN inoculated with pNT transcript, but were more than those in the same kind of plant induced by pN14 transcript while no systemic symptom was displayed in N. tabacum cv. Huangmiaoyu. These results suggest that the recombinant viruses of TN and NT are able to propagate in the assayed tobaccos, and they keep the most same phenotypic character with pTMV-Cv and pN14 transcripts, and TMV-Cv and N14 as well. The conjunctions between the replicase and the MP, CP and 3′noncoding regions are not stringent. Apparently there is a compatible function complementation between the homologous subgenomes of TMV-Cv and N14. From those above it could be probably presumed that the mutagenized replicase gene of N14 plays a major role in contributing to the virus attenuation while its mutagenized MP gene could avianize the symptoms of the infected tobaccos.
Kobayashi, Mime; Yamashita, Ichiro; Uraoka, Yukiharu; Shiba, Kiyotaka; Tomita, Satoshi
We have succeeded in aligning gold nanoparticles (Au NPs) in three-dimensions using tobacco mosaic virus (TMV) in order to realize new optical properties. TMV is a tube-shaped plant virus about 300 nm in length with an outer- and inner-diameter of 18 nm and 4 nm. We genetically fused material-binding peptides that can promote metal crystallization, namely a gold-binding peptide (GBP) and a titanium-binding peptide (TBP), to the outer-surface of TMV. By reducing potassium chloroaurate with sodium borohydride in the presence of the engineered viruses in 5% acetic acid solution, Au NPs were deposited on the outer-surface of the viruses. Using TBP-fused TMV, NPs of 5 nm were obtained, with a standard deviation smaller than those deposited on wild-type TMV. The diameter of the NPs on GBP-fused TMV was 10 nm. These results indicate that genetically-modified TMVs are promising templates for the construction of optical metamaterials.
The camellia isolate of tomato mosaic virus (ToMV-TL) can induce local necrotic lesions on the inoculated leaves in Nicotiana tabacum, whereas the broad bean isolate of tobacco mosaic virus (TMV-B) produces the mosaic symptom on systemic leaves. To examine viral determinant for differential infection phenotype in N. tabacum, the coat protein gene and the 3′ non-coding region of TMV was replaced with that of ToMV, the chimeric virus induced similar local necrotic lesions to that induced by ToMV. The results indicate that the coat protein gene and the 3′ non-coding region of TMV and ToMV influence the virus-induced pathogenesis in N. tabacum.
Padhi, Abinash; Ramu, Karri
The sugarcane mosaic virus (SCMV) of the genus potyvirus, which primarily affects maize, sugarcane, sorghum, abaca, and grasses, occurs worldwide and causes significant economic loss. Using the full genome sequences of SCMV and several recombination detection methods, in this study we report that recombination is the major driving force in the evolution and emergence of several new variants of SCMV. We reported eight highly significant (P < 0.001) recombination break points, majority of which are located within 6K1-VPg-NIaPro-NIb region, thus indicating a region for recombination hotspot. The observation of commonalities of same recombination events among the SCMV isolates between the countries (Spain and Mexico), and within the country (within China, and within Mexico), suggests common origin of the isolates in respective regions.
Gell, Gyöngyvér; Balázs, Ervin; Petrik, Kathrin
The genetic diversity of the coat-protein (CP) region and the untranslated C-terminal region (3'UTR) of Maize dwarf mosaic virus (MDMV) was analyzed to evaluate the variability between isolates (inter-isolate sequence diversity). The results of inter-isolate sequence diversity analysis showed that the diversity of the MDMV CP gene is fairly high (p-distance: up to 0.136). During sequence analysis, a 13 amino-acid residue insertion and an 8 amino-acid residue deletion were found within the N-terminal region of the CP gene. The phylogenetic analysis showed that-unlike other potyvirus species in this subgroup-the MDMV isolates could not be distinguished on the basis of their host plants or geographic origins.
Mishra, M D; Ghosh, A; Verma, V S; Dattagupta, M
Abscisic acid (ABA) did not affect the infectivity of tobacco mosaic virus (TMV) in vitro. The same dilutions of ABA when applied on the leaves of Chenopodium amaranticolor Coste and Reyn. at different intervals before inoculation affected development of local lesions variably at different dilutions. The inhibition of local lesion formation was reduced at other intervals leading to stimulation at thirty minutes and six hours intervals. Post-inoculation treatments with 2 mg/l of ABA gave stimulation of local lesion formation, though other dilutions gave inhibition. Viral concentration was stimulated in the tomato seedlings root dipped in 0.2 mg/l of ABA for 6 hours and inoculated 24 hours after transplantation. Incorporation of different concentrations of ABA into tissue culture medium reduced the growth of the TMV infected tobacco callus tissue and stimulated the infectivity of the tissue grown over it assayed after three weeks.
Erickson, J W; Bancroft, J B
Products of specific (pH 8.0-8.5) and nonspecific (pH 6.0) assembly reactions of papaya mosaic virus (PMV) coat protein with DNA are described. The strandedness, topology, and sugar moiety of the nucleic acid are important parameters for assembly in nonspecific conditions. The linear, single-stranded form of lambda DNA, but not the double-stranded form, reacted with PMV protein to form multiply initiated particles whose helical segments apparently annealed to produce continuous tubular particles. With the circular, single-stranded DNA of phi X174, partially tubular, partially extended particles were made. Poly(dA), unlike poly(A) [Erickson JW, AbouHaidar M, Bancroft JB: Virology 90:60, 1978], was not encapsidated by PMV protein under specific assembly conditions. With all DNAs tested, extended particles were the only products formed in specific conditions at pH 8.5.
Fránová, J; Petrzik, K; Lesemann, D-E; Navrátil, M
Daphne shrubs with light green rings and mosaic on leaves contained flexuous filamentous virions (696 x 13 nm) and cylindrical inclusions typical of the subdivision III of Edwardson's classification for inclusions induced by members of the family Potyviridae. Decoration tests using antisera to 67 potyviruses revealed distant serological relations among chilli veinal mottle virus, Colombian datura virus, papaya ringspot virus, tobacco vein mottling virus and yam mosaic virus. The 3' terminal region of the virus genome was amplified by RT-PCR using primers specific for cloned and sequenced members of the family Potyviridae. The most similar sequences in the GenBank were those of isolates of wild potato mosaic virus (WPMV) and yam mild mosaic virus (YMMV), originating from Peru and Guadeloupe, respectively. The new sequence had 63.2% and 61.9% nucleotide identity to WPMV and YMMV in the coat protein gene. The results suggest that the Czech isolate from daphne should be regarded as a new member of the genus Potyvirus. The name daphne mosaic virus (DapMV) is suggested for this virus.
Full Text Available A research was undergone to study the effect of single and double infection of Cucumber Mosaic Virus (CMV and Chili Veinal Mottle Virus (ChiVMV on the growth and yield of five chilli cultivars, i.e. Prabu, Taro, Jatilaba, Laris, and Keriting Bogor. Mechanical inoculation was conducted to transmit the virus. Infection of the virus was then confirmed with DAS-ELISA. Severe symptom was observed on plant given double infection compared to those given single infection. The rate of plant growth and the amount and weight of fruits were reduced. The type of interaction between CMV and ChiVMV on most chilli cultivar can be considered as interference and additive. Synergism interaction was only observed on cultivar Laris. Based on symptom expression and reduction on yield, it can be concluded that all chilli cultivars used in this study could not hold up the virus infection.
Aliyu Taiye H
Full Text Available Abstract The study was carried out to determine the comparative pathogenic response of six cultivars of soybean; TGx 1844-18E, TGx 1448-2E, TGx 1910-8F, TGx 1019-2EN, TGx 1910-8F and TGx 1876-4E to single and mixed infections with cowpea mottle virus and cucumber mosaic virus. The experiment was conducted in the screenhouse at the crop production pavilion, Faculty of Agriculture, University of Ilorin, Ilorin, Kwara state Nigeria. The results of the experiment revealed that all soybean cultivars were susceptible to single and mixed infection of the two viruses but to seemingly different extent. The single infection with cowpea mottle virus (CMeV, however, caused the most severe symptoms on the soybean cultivars. Cucumber mosaic virus (CMV alone was not as severe as the CMeV. The mixed infection of CMeV and CMV did not cause higher severity than CMeV alone indicating that there was little or no synergistic effect between the two viruses on soybean.
The wide host range of Cucumber mosaic virus (CMV) has been expanded by the identification of Odontonema cuspidatum (firespike) and Psychotria punctata (dotted wild coffee) as CMV hosts in Florida....
Cross protection or superinfection exclusion (SE) is defined as the phenomenon whereby initial infection by one virus prevents subsequent infection by closely related viruses. The mechanisms of SE are just beginning to be understood. Wheat streak mosaic virus (WSMV; genus: Tritimovirus; family: Poty...
Kristopher J Koudelka
Full Text Available Cowpea mosaic virus (CPMV is a plant comovirus in the picornavirus superfamily, and is used for a wide variety of biomedical and material science applications. Although its replication is restricted to plants, CPMV binds to and enters mammalian cells, including endothelial cells and particularly tumor neovascular endothelium in vivo. This natural capacity has lead to the use of CPMV as a sensor for intravital imaging of vascular development. Binding of CPMV to endothelial cells occurs via interaction with a 54 kD cell-surface protein, but this protein has not previously been identified. Here we identify the CPMV binding protein as a cell-surface form of the intermediate filament vimentin. The CPMV-vimentin interaction was established using proteomic screens and confirmed by direct interaction of CPMV with purified vimentin, as well as inhibition in a vimentin-knockout cell line. Vimentin and CPMV were also co-localized in vascular endothelium of mouse and rat in vivo. Together these studies indicate that surface vimentin mediates binding and may lead to internalization of CPMV in vivo, establishing surface vimentin as an important vascular endothelial ligand for nanoparticle targeting to tumors. These results also establish vimentin as a ligand for picornaviruses in both the plant and animal kingdoms of life. Since bacterial pathogens and several other classes of viruses also bind to surface vimentin, these studies suggest a common role for surface vimentin in pathogen transmission.
Full Text Available A mosaic disease of pumpkin (Cucurbita maxima was spread widely in Sulawesi. Since the virus had not yet been identified, a study was conducted to identify the disease through mechanical inoculation, aphid vector transmission, host range, and electron microscopic test. Crude sap of infected pumpkin leaf samples was rubbed on the cotyledons of healthy pumpkin seedlings for mechanical inoculation. For insect transmission, five infective aphids were infected per seedling. Seedlings of eleven different species were inoculated mechanically for host range test. Clarified sap was examined under the electron microscope. Seeds of two pumpkin fruits from two different infected plants were planted and observed for disease transmission up to one-month old seedlings. The mosaic disease was transmitted mechanically from crude sap of different leaf samples to healthy pumpkin seedlings showing mosaic symptoms. The virus also infected eight cucurbits, i.e., cucumber (Cucumis sativus, green melon (Cucumis melo, orange/rock melon (C. melo, zucchini (Cucurbita pepo, pumpkin (Cucurbita maxima, water melon (Citrulus vulgaris, Bennicosa hispida, and blewah (Cucurbita sp.. Aphids transmitted the disease from one to other pumpkin seedlings. The virus was not transmitted by seed. The mosaic disease of pumpkin at Maros, South Sulawesi, was associated with flexious particles of approximately 750 nm length, possibly a potyvirus, such as water melon mosaic virus rather than papaya ringspot virus or zucchini yellow mosaic virus.
Naseem, Saadia; Winter, Stephan
The quantity of genomic DNA-A and DNA-B of African cassava mosaic virus (ACMV) and East African cassava mosaic virus Uganda (Uganda variant, EACMV-UG) was analysed using quantitative PCR to assess virus concentrations in plants from susceptible and tolerant cultivars. The concentrations of genome components in absolute and relative quantification experiments in single and mixed viral infections were determined. Virus concentration was much higher in symptomatic leaf tissues compared to non-symptomatic leaves and corresponded with the severity of disease symptoms. In general, higher titres were recorded for EACMV-UG Ca055 compared to ACMV DRC6. The quantitative assessment also showed that the distribution of both viruses in the moderately resistant cassava cv. TMS 30572 was not different from the highly susceptible cv. TME 117. Natural mixed infections with both viruses gave severe disease symptoms. Relative quantification of virus genomes in mixed infections showed higher concentrations of EACMV-UG DNA-A compared to ACMV DNA-A, but a marked reduction of EACMV-UG DNA-B. The higher concentrations of EACMV-UG DNA-B compared to EACMV DNA-A accumulation in single infections were consistent. Since DNA-B is implicated in virus cell-to-cell spread and systemic movement, the abundance of the EACMV-UG DNA-B may be an important factor driving cassava mosaic disease epidemic.
Yakoubi, Soumaya; Lecoq, Hervé; Desbiez, Cécile
A potyvirus was isolated from a naturally infected squash plant in Algeria in 1986. Biological and serological data have revealed that the virus, initially described as H4, is related to other cucurbit-infecting potyviruses, particularly Moroccan watermelon mosaic virus (MWMV) and Papaya ringspot virus (PRSV). To establish unequivocally the taxonomic status of H4, its full-length genome sequence was established. H4 shared identities of 70% and 65% at the amino acid level with MWMV and PRSV, respectively, indicating that H4 is a distinct species of the PRSV cluster. The name Algerian watermelon mosaic virus (AWMV) is proposed for this new potyvirus species.
YANG; Gong; (
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Full Text Available We investigated the protective effects of a viral vector based on an Apple latent spherical virus (ALSV harboring a segment of the Bean yellow mosaic virus (BYMV genome against mosaic diseases in pea, broad bean, and eustoma plants caused by BYMV infection. In pea plants pre-inoculated with the ALSV vaccine and challenge inoculated with BYMV expressing green fluorescence protein, BYMV multiplication occurred in inoculated leaves, but was markedly inhibited in the upper leaves. No mosaic symptoms due to BYMV infection were observed in the challenged plants pre-inoculated with the ALSV vaccine. Simultaneous inoculation with the ALSV vaccine and BYMV also prevented mosaic symptoms in broad bean and eustoma plants, and BYMV accumulation was strongly inhibited in the upper leaves of plants treated with the ALSV vaccine. Pea and eustoma plants were pre-inoculated with BYMV followed by inoculation with the ALSV vaccine to investigate the curative effects of the ALSV vaccine. In both plant species, recovery from mosaic symptoms was observed in upper leaves and BYMV accumulation was inhibited in leaves developing post-ALSV vaccination. These results show that ALSV vaccination not only prevents mosaic diseases in pea, broad bean, and eustoma, but that it is also effective in curing these diseases.
Satoh, Nozomi; Kon, Tatsuya; Yamagishi, Noriko; Takahashi, Tsubasa; Natsuaki, Tomohide; Yoshikawa, Nobuyuki
We investigated the protective effects of a viral vector based on an Apple latent spherical virus (ALSV) harboring a segment of the Bean yellow mosaic virus (BYMV) genome against mosaic diseases in pea, broad bean, and eustoma plants caused by BYMV infection. In pea plants pre-inoculated with the ALSV vaccine and challenge inoculated with BYMV expressing green fluorescence protein, BYMV multiplication occurred in inoculated leaves, but was markedly inhibited in the upper leaves. No mosaic symptoms due to BYMV infection were observed in the challenged plants pre-inoculated with the ALSV vaccine. Simultaneous inoculation with the ALSV vaccine and BYMV also prevented mosaic symptoms in broad bean and eustoma plants, and BYMV accumulation was strongly inhibited in the upper leaves of plants treated with the ALSV vaccine. Pea and eustoma plants were pre-inoculated with BYMV followed by inoculation with the ALSV vaccine to investigate the curative effects of the ALSV vaccine. In both plant species, recovery from mosaic symptoms was observed in upper leaves and BYMV accumulation was inhibited in leaves developing post-ALSV vaccination. These results show that ALSV vaccination not only prevents mosaic diseases in pea, broad bean, and eustoma, but that it is also effective in curing these diseases.
Blanch, Ewan W.; Robinson, David J.; Hecht, Lutz;
Potato virus X (PVX) and narcissus mosaic virus (NMV) were studied using vibrational Raman optical activity (ROA) in order to obtain new information on the structures of their coat protein subunits. The ROA spectra of the two intact virions are very similar to each other and similar to that of to......Potato virus X (PVX) and narcissus mosaic virus (NMV) were studied using vibrational Raman optical activity (ROA) in order to obtain new information on the structures of their coat protein subunits. The ROA spectra of the two intact virions are very similar to each other and similar...... that the coat protein subunit folds of PVX and NMV may be very similar to each other and similar to that of TMV. These results suggest that PVX and NMV may have coat protein subunit structures based on folds similar to the TMV helix bundle and hence that the helical architecture of the PVX and NMV particles may...... be similar to that of TMV but with different structural parameters....
Robbins, M A; Witsenboer, H; Michelmore, R W; Laliberte, J F; Fortin, M G
Presence of the dominant Tu gene in Lactuca sativa is sufficient to confer resistance to infection by turnip mosaic virus (TuMV). In order to obtain an immunological assay for the presence of TuMV in inoculated plants, the TuMV coat protein (CP) gene was cloned by amplification of a cDNA corresponding to the viral genome using degenerate primers designed from conserved potyvirus CP sequences. The TuMV CP was overexpressed in Escherichia coli, and polyclonal antibodies were produced. To locate Tu on the L. sativa genetic map, F3 families from a cross between cvs "Cobbham Green" (resistant to TuMV) and "Calmar" (susceptible) were genotyped for Tu. Families known to be recombinant in the region containing Tu were infected with TuMV and tested by the indirect enzyme-linked immunosorbent assay (ELISA) using the anti-CP serum. This assay placed Tu between two random amplified polymorphic DNA (RAPD) markers and 3.2 cM from Dm5/8 (which confers resistance to Bremia lactucae). Also, bulked segregant analysis was used to screen for additional RAPD markers tightly linked to the Tu locus. Five new markers linked to Tu were identified in this region, and their location on the genetic map was determined using informative recombinants in the region. Six markers were identified as being linked within 2.5 cM of Tu.
Trinh, Minh-Hieu; Odorico, Michael; Bellanger, Laurent; Jacquemond, Mireille; Parot, Pierre; Pellequer, Jean-Luc
The study of high-resolution topographic surfaces of isolated single molecules is one of the applications of atomic force microscopy (AFM). Since tip-induced distortions are significant in topographic images the exact AFM tip shape must be known in order to correct dilated AFM height images using mathematical morphology operators. In this work, we present a protocol to estimate the AFM tip apex radius using tobacco mosaic virus (TMV) particles. Among the many advantages of TMV, are its non-abrasivity, thermal stability, bio-compatibility with other isolated single molecules and stability when deposited on divalent ion pretreated mica. Compared to previous calibration systems, the advantage of using TMV resides in our detailed knowledge of the atomic structure of the entire rod-shaped particle. This property makes it possible to interpret AFM height images in term of the three-dimensional structure of TMV. Results obtained in this study show that when a low imaging force is used, the tip is sensing viral protein loops whereas at higher imaging force the tip is sensing the TMV particle core. The known size of the TMV particle allowed us to develop a tip-size estimation protocol which permits the successful erosion of tip-convoluted AFM height images. Our data shows that the TMV particle is a well-adapted calibrator for AFM tips for imaging single isolated biomolecules. The procedure developed in this study is easily applicable to any other spherical viral particles.
Lee, Kui Jae; Choi, Min Kyung; Lee, Wang Hyu; Rajkumar, Mani
The complete sequences of both RNAs of an isolate of barley yellow mosaic virus (BaYMV) from Haenam, Korea, were determined. RNA1 is 7639 nucleotides long [excluding the 3'-poly(A)], and codes for a 270 kDa polyprotein of 2411 amino acids which contains the capsid protein (CP) at the C terminus and seven putative non-structural proteins. RNA2 is 3582 nucleotides long and codes for a polyprotein of 890 amino acids, which contains a 28 kDa putative proteinase (P1) and a 73 kDa polypeptide (P2). The whole sequences of Korean isolate (BaYMV-K) closely resembled those of an isolate from Japan (BaYMV-J) (99.6 identical nucleotides for RNA1; 99.4 for RNA2) and china (BaYMV-C) (96.7 and 96.2%, respectively) than from Germany (BaYMV-G) (93.6 and 90.4%, respectively). The greatest differences between the BaYMV-K and BaYMV-J isolates were in the 3'-NCRs of RNA1 and 5' NCRs of RNA2 and there were also some other regions of difference in Nib Pro (RNA1) and P1 (RNA2). Further, the phylogenetic analysis of CP region showed that Asian and European isolates formed distinct clusters. However, molecular variations between isolates could not be linked to earlier results showing differences in cultivar response.
Transgenic Gladiolus plants that contain either Cucumber mosaic virus (CMV) subgroup I coat protein, CMV subgroup II coat protein, CMV replicase, a combination of the CMV subgroups I and II coat proteins, or a combination of the CMV subgroup II coat protein and replicase genes were developed. These...
Full Text Available Cucumber mosaic virus (CMV and Potato virus Y (PVY have been described among the top five important viruses infecting vegetable species worldwide. They cause severe damages in fruits and cultivated plants. There is currently no available effective pesticide to control these viral diseases. Higher plants contain a wide spectrum of secondary metabolites such as phenolics, flavonoids, quinones, tannins, essential oils, alkaloids, saponins, sterols and others. Extracts prepared from different plants have been reported to have a variety of properties including antifungal, antiviral and antibacterial properties against pathogens. Tanacetum vulgare (Tansy is native to Europe, Asia, and North Africa. It has many horticultural and pharmacological qualities. T. vulgare is principally used in traditional Asian and North African medicine as an antihelminthic, antispasmodic, stimulant to abdominal viscera, tonic, antidiabetic and diuretic, and it is antihypertensive. In our research we established antiviral effect of methanol extract from T. vulgare against CMV and PVY in tomato plants.
Full Text Available Abstract Viral diseases constitute obstacles to pepper production in the world. In Nigeria, pepper plants are primarily affected by pepper veinal mottle virus (PVMV, Cucumber mosaic virus (CMV, Pepper leaf curl Virus (TLCV, Tobacco mosaic virus (TMV, Pepper mottle virus (PMV and a host of other viruses. The experiment was carried out with a diagnostic survey on the experimental field of the National Horticultural Research Institute, Ibadan, Nigeria and on pepper farms in six local government areas within Ibadan Oyo State, Nigeria, forty samples were collected from each of the farms. Diseased samples were obtained from the field and taken to the laboratory for indexing. In ELISA test some of the samples from the pepper farms showed positive reaction to single infection with PVMV (36.79%, CMV (22.14% while some others showed positive reaction to mixed infection of the two viruses (10% but some also negative reaction to PVMV and CMV antisera (31.07.
Cowpea aphid-borne mosaic potyvirus causes severe yield losses in cowpea, an important legume crop in semi-arid regions of Africa. We have elucidated the genomic sequence of the virus and subsequently focused our attention on the so-called helper component-proteinase (HC-Pro), a virus-encoded multif
Tropical soda apple mosaic virus (TSAMV) was first identified in tropical soda apple (Solanum viarum), a noxious weed, in Florida in 2002. This report provides the first full genome sequence of TSAMV. The full genome sequence of this virus will enable research scientists to develop additional spec...
Naderpour, Masoud; Johansen, Ida Elisabeth
Reporter tagged virus clones can provide detailed information on virus–host interactions. In Phaseolus vulgaris (bean), four recessive and one dominant gene are known to control infection by strains of the potyvirus species Bean common mosaic virus (BCMV). To study the interactions between BCMV...
Gopinath, K.; Bertens, P.; Pouwels, J.; Marks, H.; Lent, van J.W.M.; Wellink, J.E.; Kammen, van A.
Cowpea mosaic virus (CPMV) derivatives expressing movement protein (MP) green fluorescent protein (GFP) fusions (MP:GFP) were used to study the intracellular targeting and localization of the MP in cowpea protoplasts and plants. In protoplasts, a virus coding for a wild type MP:GFP (MPfGFP) induced
Cowpea mosaic virus (CPMV) is the type member of the comoviridae , a group of 14 different plant viruses that have a divided genome consisting of two plus-strand RNAs. These RNAs, designated B-RNA and M-RNA, have a small protein, VPg, attached to the 5'-end and a poly(A) tail at the 3'-end and are s
In this thesis, the results of various 31P NMR experiments observed for intact virus particles of bacteriophage M13 and Tobacco Mosaic Virus (TMV), are presented. To explain the results in a consistent way, models are developed and tested. 31
Padmanabhan, Chellappan; Zheng, Yi; Li, Rugang; Martin, Gregory B.; Fei, Zhangjun; Ling, Kai-Shu
The complete genome sequence of an isolate of tomato mottle mosaic virus (ToMMV) infecting tomatoes in New York was obtained using small RNA (sRNA) deep sequencing. ToMMV_NY-13 shared 99% sequence identity with isolates from Mexico and Florida. Broader distribution of this emerging virus is a cause for concern to the tomato industry.
Reação de genótipos de feijão-caupi revela resistência às coinfecções pelo Cucumber mosaic virus, Cowpea aphid-borne mosaic virus e Cowpea severe mosaic virus Reaction of cowpea genotypes reveals resistance to co-infection by Cucumber mosaic virus, Cowpea aphid-borne mosaic virus and Cowpea severe mosaic virus
Cláudia Roberta Ribeiro de Oliveira
Full Text Available O rendimento do feijão-caupi pode ser afetado por diversos fatores, em especial as viroses. As principais espécies de vírus que infectam o feijão-caupi, no Brasil, são: Cucumber mosaic virus (CMV, Cowpea aphid-borne mosaic virus (CABMV, Cowpea severe mosaic virus (CPSMV e o Bean golden mosaic virus (BGMV. Este trabalho foi realizado em duas etapas e teve como objetivo avaliar a reação de genótipos de feijão-caupi quanto à resistência à infecção simples pelo CMV e mista nas combinações CMV+CABMV, CMV+CPSMV-I e CMV+CABMV+CPSMV-I. Inicialmente, foram incluídos 57 genótipos, sendo três avaliações em gaiolas com tela antiafídeos sob infecção controlada, e uma em condição de campo sob infecção natural. Em seguida, foram selecionados 18 genótipos para serem desenvolvidos em nove ensaios, oito em gaiolas com tela antiafídeos sob infecção controlada, e um em campo sob infecção natural. Nesses ensaios, avaliaram-se os efeitos qualitativos e quantitativos resultantes das infecções. No ensaio de campo, foram avaliados o número de plantas assintomáticas, comprimento de vagem, número de grãos por vagem, massa de cem grãos e produtividade. As coinfecções reduziram a altura da planta e a massa seca. Além disso, nas infecções envolvendo os três vírus ocorreu a morte prematura de alguns genótipos. Os genótipos BR17-Gurguéia, Epace V-96, TE97-309G-9, TE97-309G-22, TE97-309G-24 e Patativa, além de bom comportamento diante das coinfecções virais, têm sementes com padrão comercial, podendo ser empregadas diretamente em programas de melhoramento.Many factors can affect the yield of cowpea, especially viruses. The main species of viruses infecting cowpea in Brazil are Cucumber mosaic virus (CMV, Cowpea aphid-borne mosaic virus (CABMV, Cowpea severe mosaic virus (CPSMV and Cowpea golden mosaic virus (CPGMV. This study aimed to evaluate the reaction of cowpea genotypes for resistance to CMV in single or in co
Huo, P; Shen, W T; Yan, P; Tuo, D C; Li, X Y; Zhou, P
Both the single infection of papaya ringspot virus (PRSV), papaya leaf distortion mosaic virus (PLDMV) or papaya mosaic virus (PapMV) and double infection of PRSV and PLDMV or PapMV which cause indistinguishable symptoms, threaten the papaya industry in Hainan Island, China. In this study, a multiplex real-time reverse transcription PCR (RT-PCR) was developed to detect simultaneously the three viruses based on their distinctive melting temperatures (Tms): 81.0±0.8°C for PRSV, 84.7±0.6°C for PLDMV, and 88.7±0.4°C for PapMV. The multiplex real-time RT-PCR method was specific and sensitive in detecting the three viruses, with a detection limit of 1.0×10(1), 1.0×10(2), and 1.0×10(2) copies for PRSV, PLDMV, and PapMV, respectively. Indeed, the reaction was 100 times more sensitive than the multiplex RT-PCR for PRSV, and 10 times more sensitive than multiplex RT-PCR for PLDMV. Field application of the multiplex real-time RT-PCR demonstrated that some non-symptomatic samples were positive for PLDMV by multiplex real-time RT-PCR but negative by multiplex RT-PCR, whereas some samples were positive for both PRSV and PLDMV by multiplex real-time RT-PCR assay but only positive for PLDMV by multiplex RT-PCR. Therefore, this multiplex real-time RT-PCR assay provides a more rapid, sensitive and reliable method for simultaneous detection of PRSV, PLDMV, PapMV and their mixed infections in papaya.
Fuji, S; Terami, F; Furuya, H; Naito, H; Fukumoto, F
The nucleotide sequences of the 3' terminal region of the genomes of Alstroemeria mosaic virus (AlsMV) and the Amazon lily mosaic virus (ALiMV) have been determined. These sequences contain the complete coding region of the viral coat protein (CP) gene followed by a 3'-untranslated region (3'-UTR). AlsMV and ALiMV share 74.9% identity in the amino acid sequence of the CP, and 55.6% identity in the nucleotide sequence of the 3'-UTR. Phylogenetic analysis of these CP genes and 3'-UTRs in relation to those of 79 potyvirus species revealed that AlsMV and ALiMV should be assigned to the Potato virus Y (PVY) subgroup. AlsMV and ALiMV were concluded to have arisen independently within the PVY subgroup.
Oteng-Frimpong, R; Levy, Y; Torkpo, S K; Danquah, E Y; Offei, S K; Gafni, Y
Cassava mosaic disease (CMV), caused by one or a combination of cassava mosaic geminiviruses, is ranked among the most important constraints to profitable and efficient production of cassava. Effective control measures require in-depth knowledge of the viral causative agent. Using rolling-circle amplification and unique enzymes, the full genome of two species of cassava mosaic geminivirus isolated from infected cassava plants in Ghana were cloned into pCambia 1300 and pET-28b. The sequences of the genome were determined on an ABI sequencer and a pairwise comparison was performed with other cassava-infecting geminiviruses from different countries. It was revealed that cassava grown in Ghana is attacked by two species of geminivirus in either single or mixed infections. These are the African cassava mosaic virus (ACMV) and the East African cassava mosaic virus (EACMV)-like, with high sequence similarity of 94% and 80%, respectively, between the DNA-A and DNA-B components of each virus, and 66% and 41% similarity of the common region (CR) (for A and B accordingly). The DNA-A of ACMV and EACMV-like contained 2781 and 2800 nucleotides, respectively, while their DNA-B components had 2725 and 2734 nucleotides, respectively. ACMV DNA-A was over 97% similar to those of other ACMVs from the continent. In contrast, EACMV-like DNA-A was over 98% similar to the isolates from Cameroon and other West African countries, and less than 88% similar to other EACMV species. Thus ACMV and EACMV-like were named African cassava mosaic virus-Ghana and East African cassava mosaic Cameroon virus-Ghana. Computer analysis revealed that their genome arrangement follows the typical old world bipartite begomovirus genome. The association of these two species and their interaction might account for the severe symptoms observed on infected plants in the field and in the greenhouse.
Gómez, Pedro; Sempere, Raqueln; Aranda, Miguel A
The molecular biology, epidemiology, and evolutionary dynamics of Pepino mosaic virus (PepMV) are much better understood than those of Tomato torrado virus (ToTV). The earliest descriptions of PepMV suggest a recent jump from nontomato species (e.g., pepino; Solanum muricatum) to tomato (Solanum lycopersicum). Its stability in contaminated plant tissues, its transmission through seeds, and the global trade of tomato seeds and fruits may have facilitated the global spread of PepMV. Stability and seed transmission also probably account for the devastating epidemics caused by already-established PepMV strains, although additional contributing factors may include the efficient transmission of PepMV by contact and the often-inconspicuous symptoms in vegetative tomato tissues. The genetic variability of PepMV is likely to have promoted the first phase of emergence (i.e., the species jump) and it continues to play an important role as the virus becomes more pervasive, progressing from regional outbreaks to pandemics. In contrast, the long-term progression of ToTV outbreaks is not yet clear and this may reflect factors such as the limited accumulation of the virus in infected plants, which has been shown to be approximately two orders of magnitude less than PepMV. The efficient dispersion of ToTV may therefore depend on dense populations of its principal vectors, Bemisia tabaci and Trialeurodes vaporariorum, as has been proposed for the necrogenic satellite RNA of Cucumber mosaic virus.
Haq, Q M I; Rouhibakhsh, A; Ali, Arif; Malathi, V G
Yellow mosaic disease in grain legumes in Indian subcontinent is caused by two important virus species viz. Mungbean yellow mosaic virus (MYMV) and Mungbean yellow mosaic India virus (MYMIV), belonging to the genus Begomovirus of the family Geminiviridae. The genomic components of a begomovirus causing yellow mosaic disease in blackgram in southern India were cloned and sequenced. Nucleotide sequence comparison of DNA A component shows the virus isolate to be a variant of Mungbean yellow mosaic virus:-(MYMV-[IN:Vam:05]). However, DNA B component of the present virus isolate has greater similarity (92%) to Mungbean yellow mosaic India virus. Agroinoculations of the viral clones produced typical yellow mosaic symptoms in blackgram and mungbean, severe leaf curl and stunting in French bean, similar to blackgram isolate of MYMIV. Blackgram isolates of both the virus species were only mildly infectious on cowpea, produced atypical leaf curl symptoms and not yellow or golden mosaic. In agroinoculations done by exchanging genomic components, symptom expression was seen only in French bean. In cowpea, blackgram and mungbean there was no visible symptoms though viral DNA could be detected by PCR.
Hille, Jacques; Goldbach, Rob
A full-length DNA copy of the M-RNA of cowpea mosaic virus (CPMV), supplied with either the 35S promoter from cauliflower mosaic virus (CaMV) or the nopaline synthase promoter from Agrobacterium tumefaciens, was introduced into the T-DNA region of a Ti-plasmid-derived gene vector and transferred to
Siljo, A; Bhat, A I; Biju, C N
Cardamom being perennial, propagated vegetatively, detecting viruses in planting material is important to check the spread of viruses through infected material. Thus development of effective and sensitive assay for detection of viruses is need of the time. In this view, assay for the detection of Cardamom mosaic virus (CdMV) and Banana bract mosaic virus (BBrMV), infecting cardamom was developed using SYBR Green one step reverse transcription-quantitative PCR (RT-qPCR). The RT-qPCR assay amplified all isolates of CdMV and BBrMV tested but no amplification was obtained with RNA of healthy plants. Recombinant plasmids carrying target virus regions corresponding to both viruses were quantified, serially diluted and used as standards in qPCR to develop standard curve to enable quantification. When tenfold serial dilutions of the total RNAs from infected plants were tested through RT-qPCR, the detection limit of the assay was estimated to be 16 copies for CdMV and 10 copies for BBrMV, which was approximately 1,000-fold higher than the conventional RT-PCR. The RT-qPCR assay was validated by testing field samples collected from different cardamom growing regions of India. This is the first report of RT-qPCR assay for the detection of CdMV and BBrMV in cardamom.
Korber, Bette T [Los Alamos, NM; Perkins, Simon [Los Alamos, NM; Bhattacharya, Tanmoy [Los Alamos, NM; Fischer, William M [Los Alamos, NM; Theiler, James [Los Alamos, NM; Letvin, Norman [Boston, MA; Haynes, Barton F [Durham, NC; Hahn, Beatrice H [Birmingham, AL; Yusim, Karina [Los Alamos, NM; Kuiken, Carla [Los Alamos, NM
The present invention relates to mosaic clade M HIV-1 Nef polypeptides and to compositions comprising same. The polypeptides of the invention are suitable for use in inducing an immune response to HIV-1 in a human.
Karthikeyan, Chockalingam; Patil, Basavaprabhu L.; Borah, Basanta K.; Resmi, Thulasi R.; Turco, Silvia; Pooggin, Mikhail M.; Hohn, Thomas; Veluthambi, Karuppannan
The major threat for cassava cultivation on the Indian subcontinent is cassava mosaic disease (CMD) caused by cassava mosaic geminiviruses which are bipartite begomoviruses with DNA A and DNA B components. Indian cassava mosaic virus (ICMV) and Sri Lankan cassava mosaic virus (SLCMV) cause CMD in India. Two isolates of SLCMV infected the cassava cultivar Sengutchi in the fields near Malappuram and Thiruvananthapuram cities of Kerala State, India. The Malappuram isolate was persistent when maintained in the Madurai Kamaraj University (MKU, Madurai, Tamil Nadu, India) greenhouse, whereas the Thiruvananthapuram isolate did not persist. The recovered cassava plants with the non-persistent SLCMV, which were maintained vegetative in quarantine in the University of Basel (Basel, Switzerland) greenhouse, displayed re-emergence of CMD after a six-month period. Interestingly, these plants did not carry SLCMV but carried ICMV. It is interpreted that the field-collected, SLCMV-infected cassava plants were co-infected with low levels of ICMV. The loss of SLCMV in recovered cassava plants, under greenhouse conditions, then facilitated the re-emergence of ICMV. The partial dimer clones of the persistent and non-persistent isolates of SLCMV and the re-emerged isolate of ICMV were infective in Nicotiana benthamiana upon agroinoculation. Studies on pseudo-recombination between SLCMV and ICMV in N. benthamiana provided evidence for trans-replication of ICMV DNA B by SLCMV DNA A. PMID:27690084
Yu, Tsong-Ann; Chiang, Chu-Hui; Wu, Hui-Wen; Li, Chin-Mei; Yang, Ching-Fu; Chen, Jun-Han; Chen, Yu-Wen; Yeh, Shyi-Dong
Zucchini yellow mosaic virus (ZYMV) and Papaya ringspot virus type W (PRSV W) are major limiting factors for production of watermelon worldwide. For the effective control of these two viruses by transgenic resistance, an untranslatable chimeric construct containing truncated ZYMV coat protein (CP) and PRSV W CP genes was transferred to commercial watermelon cultivars by Agrobacterium-mediated transformation. Using our protocol, a total of 27 putative transgenic lines were obtained from three cultivars of 'Feeling' (23 lines), 'China baby' (3 lines), and 'Quality' (1 line). PCR and Southern blot analyses confirmed that the chimeric construct was incorporated into the genomic DNA of the transformants. Greenhouse evaluation of the selected ten transgenic lines of 'Feeling' cultivar revealed that two immune lines conferred complete resistance to ZYMV and PRSV W, from which virus accumulation were not detected by Western blotting 4 weeks after inoculation. The transgenic transcript was not detected, but small interfering RNA (siRNA) was readily detected from the two immune lines and T(1) progeny of line ZW 10 before inoculation, indicating that RNA-mediated post-transcriptional gene silencing (PTGS) is the underlying mechanism for the double-virus resistance. The segregation ratio of T(1) progeny of the immune line ZW10 indicated that the single inserted transgene is nuclearly inherited and associated with the phenotype of double-virus resistance as a dominant trait. The transgenic lines derived from the commercial watermelon cultivars have great potential for control of the two important viruses and can be implemented directly without further breeding.
Lockney, Dustin M.
Red clover necrotic mosaic virus (RCNMV) is a highly robust (Tm=60 °C), 36 nm icosahedral plant virus. The capsid of RCNMV is assembled from 180 chemically equivalent coat proteins (CPs). The CPs arrange in a T=3 symmetry, in 1 of 3 conformations forming the asymmetric subunit (ASU). There are two Ca(II) binding sites per CP; the removal of divalent cations causes the CP subunits of the ASU to rotate away from each other forming a ˜13 A channel. These channels lead to the highly organized bipartite genome of RCNMV and can be closed by adding back Ca(II). Titrimetric analysis and tryptophan fluorescence was used to determine the affinity of RCNMV for Ca(II) to be ˜Kd buffer was changed from 50 mM Tris-HCl/50 mM NaOAc/50 mM EDTA or 200 mM EDTA at pH 8.0 to 5 mM HEPES/5 mM Na4EDTA/10 mM NaCl pH 7.8. The Dox:RCNMV infusion mole ratio was also lowered from 5000:1 to 500:1 and the incubation temperature was changed from 4 °C to 22 °C for <12 hours, opposed to 24 hours. To impart targeting functionality to RCNMV, biomimetic peptides were conjugated to either the surface capsid lysines or cysteines using standard bioconjugation methods. For all of the biomimetic peptides screened, sulfosuccinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (sulfo-SMCC) was used to orthogonally attach the cysteineterminated peptides to the surface lysines of RCNMV. The efficacy of a plant virus nanoparticle (PVN), loaded with a cargo (Dox or propidium iodide) and armed with a targeting peptide, was tested in vitro against several cell line using cell viability assays. ADH304, an N-cadherin targeting peptide, was synthesized to contain a lys[maleimide]. Dox infused RCNMV was armed with ADH304maleimide by conjugating the peptide to the endogenous C267 located in the P-domain. The use of a cysteine reactive peptide increased the PVN yield from ˜30% to ˜70% by eliminating several synthetic steps. The efficacy of this PVN formulation was tested on a human melanoma tumor in a
YU; Cui; HU; Dongwei; DONG; Jiahong; CUI; Xiaofeng; WU; Jun
Tobacco mosaic virus (TMV) and Tomato mosaic virus (ToMV) are two closely related viruses in the genus Tobamovirus, but they induce obviously different sizes of necrotic lesions in tobacco plants containing the N gene. Comparison of the symptoms produced by TMV, ToMV and a chimaeric virus (T/OMP), in which the TMV movement protein (MP) gene was replaced by the ToMV MP gene, showed T/OMP caused necrotic lesions that were similar in size to those of ToMV in tobacco plants containing the N gene. The coat protein and MP of the three viruses accumulated in planta with similar levels, and the replication level of TMV and T/OMP in protoplasts also had no difference. Comparison of the activities of defense-related enzymes (PAL, POD and PPO) induced by the three viruses also showed that the variability of enzyme activity induced by T/OMP was similar to that induced by TMV, but different from that induced by ToMV. The results indicate that the size difference of necrotic lesions induced by TMV and ToMV in tobacco plants containing the N gene results from the functional difference of their MP genes.
Siegel, A.; Hari, V.; Kolacz, K.
The nature and rate of virus-specific protein synthesis were determined in tobacco mosaic virus-infected protoplasts as a function of time after inoculation. Samples of infected and mock-infected protoplasts were exposed to radioactive amino acid for relatively short sequential time periods and the consequent labeled proteins were assessed following SDS-polyacrylamide gel electrophoresis and fluorography. The synthesis of three virus-specific proteins of molecular weights 160,000, 135,000, and 17,500 was confirmed. Synthesis of all three proteins was first detected during the 5- to 7-hr postinoculation period at which time the synthetic rate of the 135,000-dalton protein was greatest.This was soon overtaken by the 17,500-dalton capsid protein, the synthetic rate of which kept increasing until it accounted for a major portion of total protoplast protein synthesis. At 1 day postinoculation, it accounted for 50% and, at not quite 2 days, 70% of the total protein synthesis. Evidence is presented to suggest that virus-specific protein synthesis occurs in addition to, rather than at the expense of, normal cellular protein synthesis.
Full Text Available A viral disease causing severe mosaic, necrotic, and yellow symptoms on Vigna angularis var. nipponensis was prevalent around Suwon area in Korea. The causal virus was characterized as Cucumber mosaic virus (CMV on the basis of biological and nucleotide sequence properties of RNAs 1, 2 and 3 and named as CMV-wVa. CMV-wVa isolate caused mosaic symptoms on indicator plants, Nicotiana tabacum cv. Xanthi-nc, Petunia hybrida, and Cucumis sativus. Strikingly, CMV-wVa induced severe mosaic and malformation on Cucurbita pepo, and Solanum lycopersicum. Moreover, it caused necrotic or mosaic symptoms on V. angularis and V. radiate of Fabaceae. Symptoms of necrotic local or pin point were observed on inoculated leaves of V. unguiculata, Vicia fava, Pisum sativum and Phaseolus vulgaris. However, CMV-wVa isolate failed to infect in Glycine max cvs. ‘Sorok’, ‘Sodam’ and ‘Somyeong’. To assess genetic variation between CMV-wVa and the other known CMV isolates, phylogenetic analysis using 16 complete nucleotide sequences of CMV RNA1, RNA2, and RNA3 including CMV-wVa was performed. CMV-wVa was more closely related to CMV isolates belonging to CMV subgroup I showing about 85.1–100% nucleotide sequences identity to those of subgroup I isolates. This is the first report of CMV as the causal virus infecting wild Vigna angularis var. nipponensis in Korea.
Kim, Mi-Kyeong; Jeong, Rae-Dong; Kwak, Hae-Ryun; Lee, Su-Heon; Kim, Jeong-Soo; Kim, Kook-Hyung; Cha, Byeongjin; Choi, Hong-Soo
A viral disease causing severe mosaic, necrotic, and yellow symptoms on Vigna angularis var. nipponensis was prevalent around Suwon area in Korea. The causal virus was characterized as Cucumber mosaic virus (CMV) on the basis of biological and nucleotide sequence properties of RNAs 1, 2 and 3 and named as CMV-wVa. CMV-wVa isolate caused mosaic symptoms on indicator plants, Nicotiana tabacum cv. Xanthi-nc, Petunia hybrida, and Cucumis sativus. Strikingly, CMV-wVa induced severe mosaic and malformation on Cucurbita pepo, and Solanum lycopersicum. Moreover, it caused necrotic or mosaic symptoms on V. angularis and V. radiate of Fabaceae. Symptoms of necrotic local or pin point were observed on inoculated leaves of V. unguiculata, Vicia fava, Pisum sativum and Phaseolus vulgaris. However, CMV-wVa isolate failed to infect in Glycine max cvs. 'Sorok', 'Sodam' and 'Somyeong'. To assess genetic variation between CMV-wVa and the other known CMV isolates, phylogenetic analysis using 16 complete nucleotide sequences of CMV RNA1, RNA2, and RNA3 including CMV-wVa was performed. CMV-wVa was more closely related to CMV isolates belonging to CMV subgroup I showing about 85.1-100% nucleotide sequences identity to those of subgroup I isolates. This is the first report of CMV as the causal virus infecting wild Vigna angularis var. nipponensis in Korea.
Côté, Fabien; Paré, Christine; Majeau, Nathalie; Bolduc, Marilène; Leblanc, Eric; Bergeron, Michel G; Bernardy, Michael G; Leclerc, Denis
A filamentous virus isolated from Malva neglecta Wallr. (common mallow) and propagated in Chenopodium quinoa was grown, cloned and the complete nucleotide sequence was determined (GenBank accession # DQ660333). The genomic RNA is 6858 nt in length and contains five major open reading frames (ORFs). The genomic organization is similar to members and the viral encoded proteins shared homology with the group of the Potexvirus genus in the Flexiviridae family. Phylogenetic analysis revealed a close relationship with narcissus mosaic virus (NMV), scallion virus X (ScaVX) and, to a lesser extent, to Alstroemeria virus X (AlsVX) and pepino mosaic virus (PepMV). A novel putative pseudoknot structure is predicted in the 3'-UTR of a subgroup of potexviruses, including this newly described virus. The consensus GAAAA sequence is detected at the 5'-end of the genomic RNA and experimental data strongly suggest that this motif could be a distinctive hallmark of this genus. The name Malva mosaic virus is proposed.
The aim of the studies described in this thesis was to gain more insight in the complex molecular mechanisms underlying the RNA replication of the cowpea mosaic virus genome. Previously the replication of CPMV RNA has been examined extensively with crude membrane fractions prepared from CPMV inf
The complete genome sequence of Triticum mosaic virus (TriMV), a member in the family Potyviridae, has been determined to be 10,266 nucleotides excluding the 3’-polyadenylated tail. The genome encodes a large polyprotein of 3,112 amino acids with the ‘hall-mark proteins’ of potyviruses including a s...
Chen, Yuh-Kun; Goldbach, Rob; Prins, Marcel
In four distinct alstroemeria-infecting cucumber mosaic virus (CMV) isolates, additional sequences of various lengths were present in the 3' nontranslated regions of their RNAs 2 and 3, apparently the result of intra- and intermolecular recombination events. Competition experiments revealed that these recombined RNA 2 and 3 segments increased the biological fitness of CMV in alstroemeria.
Since its first discovery in 2013 in Mexico, Tomato mottle mosaic virus (ToMMV), a new tomato-infecting tobamovirus is now present in a number of countries (i.e., Brazil, China, and Israel) and several states in the U.S. There is little information available on the molecular and biological properti...
The complete genome sequence (6,423 nt) of an emerging Cucumber green mottle mosaic virus (CGMMV) isolate on cucumber in North America was determined through deep sequencing of sRNA and rapid amplification of cDNA ends. It shares 99% nucleotide sequence identity to the Asian genotype, but only 90% t...
Kodetham Gopinath,; Wellink, J.; Porta, C.; Taylor, K.M.; Lomonossoff, G.P.; Kammen, van A.
series of new cowpea mosaic virus (CPMV) RNA-2-based expression vectors were designed. The jellyfish green fluorescent protein (GFP) was introduced between the movement protein (MP) and the large (L) coat protein or downstream of the small (S) coat protein. Release of the GFP inserted between the MP
Lim, Seungmo; Zhao, Fumei; Yoo, Ran Hee; Igori, Davaajargal; Jeong, Jae Cheol; Lee, Haeng-Soon; Kwak, Sang-Soo; Moon, Jae Sun
The complete genome sequence of a South Korean isolate of Rehmannia mosaic virus (ReMV) infecting Rehmannia glutinosa was determined through next-generation sequencing and Sanger sequencing. To our knowledge, this is the first report of a natural infection of R. glutinosa by ReMV in South Korea.
Mlotshwa, S.; Verver, J.; Sithole-Niang, I.; Gopinath, K.; Carette, J.; Kammen, van A.; Wellink, J.
The helper component-proteinase (HC-Pro) of Cowpea aphid-borne mosaic virus (CABMV) was expressed in Escherichia coli and used to obtain HC-Pro antiserum that was used as an analytical tool for HC-Pro studies. The antiserum was used in immunofluorescence assays to study the subcellular location of H
Transgenic plants of Gladiolus ‘Peter Pears’ or ‘Jenny Lee’ were developed that contain single-chain variable fragments (scFv) to Cucumber mosaic virus (CMV) subgroup I or II. The CMV subgroup I heavy and light chain scFv fragments were placed under control of either the duplicated CaMV 35S or suga...
Chen, Y.K.; Goldbach, R.W.; Prins, M.W.
In four distinct alstroemeria-infecting cucumber mosaic virus (CMV) isolates, additional sequences of various lengths were present in the 3' nontranslated regions of their RNAs 2 and 3, apparently the result of intra- and intermolecular recombination events. Competition experiments revealed that the
This thesis described the characterization of a range of ornamental-infecting Cucumber mosaic virus strains and the development of novel transgene constructs to improve the efficiency of obtaining resistant transformants which is essential for most ornamental plants that are diffi
Chen, Y.K.; Prins, M.W.; Derks, A.F.L.M.; Langeveld, S.A.; Goldbach, R.W.
The coat protein (CP) genes and flanking regions of three alstroemeria-infecting cucumber mosaic virus isolates (CMV-ALS), denoted ALS-LBO, ALS-IPO, and ALS-NAK, were cloned and their nucleotide sequence determined and compared at both nucleic acid and deduced protein level with the published sequen
Full Text Available Blackgram (Vigna mungo (L. Hepper is one of the major pulse crops of the tropics and sub tropics. It is the third major pulse crop cultivated in the Indian subcontinent. Pulses and grain legumes are major sources of dietary protein. These crops are subjected to yellow mosaic and golden mosaic diseases caused by white fly transmitted geminiviruses (WTG’s or begomovirus. Of these viruses, mungbean yellow mosaic virus (MYMV is an important one, and it infects five major leguminous species, such as blackgram, greengram, Frenchbean, pigeonpea and soybean causing an annual yield loss of about US $ 300 million (Varma et al., 1992. The MYMV causes 85-100 per cent yield loss in the plants that are infected at the seedling stage (Nene, 1973.MYMV was first observed in Delhi in the late fifties (Nariani, 1960. Virus particles were first observed by Thongmeearkom et al. (1981 and purified by Honda et al. (1983. Hence the characterisation of Yellow Mosaic Virus is essential to study the variability and to identify any new strains/ variants of YMV prevalent in India and Abroad at molecular level for developing the new resistant genotypes.
Cho, Sang-Yun; Lim, Seungmo; Kim, Hongsup; Yi, Seung-In; Moon, Jae Sun
We report here the first complete genome sequence of Ornithogalum mosaic virus (OrMV) isolated from Taean, South Korea, in 2011, which was obtained by next-generation sequencing and Sanger sequencing. The sequence information provided here may serve as a potential reference for other OrMV isolates.
Igori, Davaajargal; Lim, Seungmo; Zhao, Fumei; Baek, Dasom; Moon, Jae Sun
Habenaria mosaic virus (HaMV), a member of the genus Potyvirus in the family Potyviridae, was first discovered from Habenaria radiata in Japan. The complete genomic sequence of a South Korean isolate (PA1) of HaMV infecting Plantago asiatica L. was determined with high-throughput RNA sequencing.
Komatsu, Ken; Yamashita, Kazuo; Sugawara, Kota; Verbeek, Martin; Fujita, Naoko; Hanada, Kaoru; Uehara-Ichiki, Tamaki; Fuji, Shin Ichi
Plantago asiatica mosaic virus (PlAMV) is a member of the genus Potexvirus and has an exceptionally wide host range. It causes severe damage to lilies. Here we report on the complete nucleotide sequences of two new Japanese PlAMV isolates, one from the eudicot weed Viola grypoceras (PlAMV-Vi), and t
The aim of the research described in this thesis was the purification and identification of the RNA-dependent RNA polymerase engaged in replicating viral RNA in cowpea mosaic virus (CPMV)- infected cowpea leaves.Previously, an RNA-dependent RNA polymerase produced upon infection of Vigna unguiculata
Londoño, M. A.; Cohen, A. L.; Padilla-Rodriguez, M.; Rosario, K.; Breitbart, M.
ABSTRACT Euphorbia mosaic virus (EuMV) was found in a symptomatic passionfruit (Passiflora edulis) plant from Homestead, Florida, USA, as well as in the symptomatic weed Euphorbia heterophylla. This is the first identification of EuMV in Florida and the United States and the first report of a natural infection of passionfruit by EuMV. PMID:28254981
Chen, Ying-Juan; Zhang, Jing; Liu, Jian; Deng, Xing-Guang; Zhang, Ping; Zhu, Tong; Chen, Li-Juan; Bao, Wei-Kai; Xi, De-Hui; Lin, Hong-Hui
Infection of plants by multiple viruses is common in nature. Cucumber mosaic virus (CMV) and Turnip crinkle virus (TCV) belong to different families, but Arabidopsis thaliana and Nicotiana benthamiana are commonly shared hosts for both viruses. In this study, we found that TCV provides effective resistance to infection by CMV in Arabidopsis plants co-infected by both viruses, and this antagonistic effect is much weaker when the two viruses are inoculated into different leaves of the same plant. However, similar antagonism is not observed in N. benthamiana plants. We further demonstrate that disrupting the RNA silencing-mediated defense of the Arabidopsis host does not affect this antagonism, but capsid protein (CP or p38)-defective mutant TCV loses the ability to repress CMV, suggesting that TCV CP plays an important role in the antagonistic effect of TCV toward CMV in Arabidopsis plants co-infected with both viruses.
Wang, Zhao; Hryc, Corey F.; Bammes, Benjamin; Afonine, Pavel V.; Jakana, Joanita; Chen, Dong-Hua; Liu, Xiangan; Baker, Matthew L.; Kao, Cheng; Ludtke, Steven J; Schmid, Michael F.; Adams, Paul D.; Chiu, Wah
Advances in electron cryo-microscopy have enabled structure determination of macromolecules at near-atomic resolution. However, structure determination, even using de novo methods, remains susceptible to model bias and overfitting. Here we describe a complete workflow for data acquisition, image processing, all-atom modelling and validation of brome mosaic virus, an RNA virus. Data were collected with a direct electron detector in integrating mode and an exposure beyond the traditional radiat...
Glasa, M; Kollerova, E
The seed transmission of the Zucchini yellow mosaic virus (ZYMV) was studied in cucumber using two isolates unrelated in their biological characteristics. Although the virus could be readily detected in mature seeds harvested from infected cucumbers, the seedlings obtained from infected germinated seeds tested negative for ZYMV using both ELISA and RT-PCR assays. No evidence was obtained for transmission of two ZYMV isolates through seeds.
Aqueous suspensions of mixtures of the rodlike virus tobacco mosaic virus (TMV) with globular macromolecules such as polyethylene oxide (PEO) or bovine serum albumin (BSA) phase separate and exhibit rich and strikingly similar phase behavior. Isotropic, nematic, lamellar, and crystalline phases are observed as a function of the concentration of the constituents and ionic strength. The observed phase behavior is considered to arise from attractions between the two particles induced by the pres...
Suzuki, Masashi; Yoshida, Megumi; Yoshinuma, Toshio; Hibi, Tadaaki
Cucumber mosaic virus (CMV) and Peanut stunt virus (PSV) each have genomes consisting of three single-stranded RNA molecules: RNA 1, 2 and 3. RNAs 1 and 2 encode the 1a and 2a proteins, respectively, which are necessary for replication of the viral genome. Although RNA 3 is exchangeable between CMV and PSV, exchange of RNA 1 and 2 between the two viruses has been unsuccessful. In this study, reassortants containing PSV RNA 1 and CMV RNA 2 together with RNA 3 of CMV or PSV were shown to be able to replicate their genomic RNA, but not to transcribe subgenomic RNA 4 in tobacco protoplasts. Conversely, the reassortant consisting of CMV RNA 1 and PSV RNA 2 together with RNA 3 of CMV or PSV could not replicate. Subsequently, a yeast two-hybrid system was used to analyse the in vivo interaction between the 1a and 2a proteins. The C-terminal half of PSV-1a protein interacted with the N-terminal region of 2a protein of both PSV and CMV, but the C-terminal half of CMV-1a and the N-terminal region of PSV-2a did not interact. These results suggest that RNA replication in the interspecific reassortant between CMV and PSV requires compatibility between the C-terminal half of the 1a protein and the N-terminal region of the 2a protein, and this compatibility is insufficient for transcription of subgenomic RNA 4.
Di Carli, Mariasole; Villani, Maria Elena; Bianco, Linda; Lombardi, Raffaele; Perrotta, Gaetano; Benvenuto, Eugenio; Donini, Marcello
Cucumber mosaic virus (CMV), a member of the Cucumovirus genus, is the causal agent of several plant diseases in a wide range of host species, causing important economic losses in agriculture. Because of the lack of natural resistance genes in most crops, different genetic engineering strategies have been adopted to obtain virus-resistant plants. In a previous study, we described the engineering of transgenic tomato plants expressing a single-chain variable fragment antibody (scFv G4) that are specifically protected from CMV infection. In this work, we characterized the leaf proteome expressed during compatible plant-virus interaction in wild type and transgenic tomato. Protein changes in both inoculated and apical leaves were revealed using two-dimensional gel electrophoresis (2-DE) coupled to differential in gel electrophoresis (DIGE) technology. A total of 2084 spots were detected, and 50 differentially expressed proteins were identified by nanoscale liquid chromatographic-electrospray ionization-ion trap-tandem mass spectrometry (nLC-ESI-IT-MS/MS). The majority of these proteins were related to photosynthesis (38%), primary metabolism (18%), and defense activity (14%) and demonstrated to be actively down regulated by CMV in infected leaves. Moreover, our analysis revealed that asymptomatic apical leaves of transgenic inoculated plants had no protein profile alteration as compared to control wild type uninfected plants demonstrating that virus infection is confined to the inoculated leaves and systemic spread is hindered by the CMV coat protein (CP)-specific scFv G4 molecules. Our work is the first comparative study on compatible plant-virus interactions between engineered immunoprotected and susceptible wild type tomato plants, contributing to the understanding of antibody-mediated disease resistance mechanisms.
In Ecuador, tamarillo (Solanum betaceum) represents an important cash crop for hundreds of small farmers. In 2013, leaves from tamarillo plants showing severe virus-like symptoms (mosaic, mottling and leaf deformation) were collected from old orchards in Pichincha and Tungurahua. Double-stranded RN...
Full Text Available Understanding the molecular mechanisms involved in plant virus–vector interactions is essential for the development of effective control measures for aphid-vectored epidemic plant diseases. The coat proteins (CP are the main component of the viral capsids, and they are implicated in practically every stage of the viral infection cycle. Pea enation mosaic virus 1 (PEMV1, Enamovirus, Luteoviridae and Pea enation mosaic virus 2 (PEMV2, Umbravirus, Tombusviridae are two RNA viruses in an obligate symbiosis causing the pea enation mosaic disease. Sixteen mutant viruses were generated with mutations in different domains of the CP to evaluate the role of specific amino acids in viral replication, virion assembly, long-distance movement in Pisum sativum, and aphid transmission. Twelve mutant viruses were unable to assemble but were able to replicate in inoculated leaves, move long-distance, and express the CP in newly infected leaves. Four mutant viruses produced virions, but three were not transmissible by the pea aphid, Acyrthosiphon pisum. Three-dimensional modeling of the PEMV CP, combined with biological assays for virion assembly and aphid transmission, allowed for a model of the assembly of PEMV coat protein subunits.
Yang, Shaoqing; Wang, Tao; Bohon, Jen; Gagné, Marie-Ève Laliberté; Bolduc, Marilène; Leclerc, Denis; Li, Huilin
Papaya mosaic virus (PapMV) is a filamentous plant virus that belongs to the Alphaflexiviridae family. Flexible filamentous viruses have defied more than two decades of effort in fiber diffraction, and no high-resolution structure is available for any member of the Alphaflexiviridae family. Here, we report our structural characterization of PapMV by X-ray crystallography and cryo-electron microscopy three-dimensional reconstruction. We found that PapMV is 135Å in diameter with a helical symmetry of ~10 subunits per turn. Crystal structure of the C-terminal truncated PapMV coat protein (CP) reveals a novel all-helix fold with seven α-helices. Thus, the PapMVCP structure is different from the four-helix-bundle fold of tobacco mosaic virus in which helix bundling dominates the subunit interface in tobacco mosaic virus and conveys rigidity to the rod virus. PapMV CP was crystallized as an asymmetrical dimer in which one protein lassoes the other by the N-terminal peptide. Mutation of residues critical to the inter-subunit lasso interaction abolishes CP polymerization. The crystal structure suggests that PapMV may polymerize via the consecutive N-terminal loop lassoing mechanism. The structure of PapMV will be useful for rational design and engineering of the PapMV nanoparticles into innovative vaccines.
Yusim, Karina; Korber, Bette T. M.; Kuiken, Carla L.; Fischer, William M.
The invention relates to immunogenic compositions useful as HCV vaccines. Provided are HCV mosaic polypeptide and nucleic acid compositions which provide higher levels of T-cell epitope coverage while minimizing the occurrence of unnatural and rare epitopes compared to natural HCV polypeptides and consensus HCV sequences.
A bench-scale, cost effective and simple method to elicit Lycopersicon esculentum cv. PKM1 (tomato) plants against Cucumber mosaic virus attack using ozone-mediated inactivated Cucumber mosaic virus inoculum.
Sudhakar, N; Nagendra-Prasad, D; Mohan, N; Murugesan, K
Studies were undertaken to evaluate ozone for inactivation of Cucumber mosaic virus present in the inoculum and to stimulate Lycopersicon esculentum cv. PKM1 (tomato) plants against Cucumber mosaic virus infection by using the inactivated Cucumber mosaic virus inoculum. Application of a T(4) (0.4mg/l) concentration of ozone to the inoculum containing Cucumber mosaic virus resulted in complete inactivation of the virus. The inactivated viral inoculum was mixed with a penetrator (delivery agent), referred to as T(4) preparation, and it was evaluated for the development of systemic acquired resistance in the tomato plants. Application of a T(4) preparation 5 days before inoculation with the Cucumber mosaic virus protected tomato plants from the effects of Cucumber mosaic virus. Among the components of the inactivated virus tested, coat protein subunits and aggregates were responsible for the acquired resistance in tomato plants. In field trials, the results of enzyme-linked immunosorbent assay revealed that, Cucumber mosaic virus accumulation was significantly less for all the test plants (16%) sprayed with the T(4) preparation than untreated control plants (89.5%) at 28 days postinoculation (dpi). A remarkable increase in the activities of the total soluble phenolics (10-fold) and salicylic acid (16-fold) was detected 5 days after the treatment in foliar extracts of test plants relative to untreated control plants. The results showed that treatment of tomato plants with inactivated viral inoculum led to a significant enhancement of protection against Cucumber mosaic virus attack in a manner that mimics a real pathogen and induces systemic acquired resistance.
Zeng, Rong; Liao, Qiansheng; Feng, Junli; Li, Dingjun; Chen, Jishuang
Cucumber mosaic virus (CMV) and zucchini yellow mosaic virus (ZYMV) are two principal viruses infecting cucurbitaceous crops, and their synergy has been repeatedly observed. In our present work, a real-time reverse transcription-polymerase chain reaction procedure was established to study the accumulation kinetics of these two viruses in single and combined infections at the molecular level. The accumulations of open reading frames (ORFs) for 1a, 2a, 3a and coat protein (CP) of CMV and CP of ZYMV were tested. In the single infection, CMV-Fny ORFs accumulated to their maxima in cucumber or bottle gourd at 14 d post-inoculation (dpi), and gradually declined thereafter. ZYMV-SD CP ORF reached maximal accumulation at 14 and 28 dpi on cucumber and bottle gourd, respectively. However, when co-infected with CMV-Fny and ZYMV-SD, the maximal accumulation levels of all viral ORFs were delayed. CMV-Fny ORFs reached their maxima at 21 dpi on both hosts, and ZYMV-SDCP ORF reached maximal accumulation at 21 and 28 dpi on cucumber and bottle gourd, respectively. Generally, the accumulation levels of CMV-Fny ORFs in the co-infection were higher than those in the single infection, whereas the accumulation of ZYMV-SD CP ORF showed a reverse result.
Zhang, H; Todderud, E; Stubbs, G
Papaya mosaic virus coat protein has been treated with trypsin and a large fragment of the intact protein has been crystallized in space group P3(1)21 or P3(2)21 (unit cell dimensions: a = b = 110 A, c = 237 A). The crystals diffract to 3.5 A resolution. Crystals of the untreated protein have also been grown. The untreated protein crystals diffract to 4 A resolution, but have a large mosaic spread. They have the same space group as the trypsin-treated protein crystals, but a much smaller unit cell (a = b = 72 A, c = 240 A).
Lee, Hoonsoo; Lim, Hyoun-Sub; Cho, Byoung-Kwan
The Cucumber Green Mottle Mosaic Virus (CGMMV) is a globally distributed plant virus. CGMMV-infected plants exhibit severe mosaic symptoms, discoloration, and deformation. Therefore, rapid and early detection of CGMMV infected seeds is very important for preventing disease damage and yield losses. Raman spectroscopy was investigated in this study as a potential tool for rapid, accurate, and nondestructive detection of infected seeds. Raman spectra of healthy and infected seeds were acquired in the 400 cm-1 to 1800 cm-1 wavenumber range and an algorithm based on partial least-squares discriminant analysis was developed to classify infected and healthy seeds. The classification model's accuracies for calibration and prediction data sets were 100% and 86%, respectively. Results showed that the Raman spectroscopic technique has good potential for nondestructive detection of virus-infected seeds.
Aljabali, Alaa A. A.; Barclay, J. Elaine; Steinmetz, Nicole F.; Lomonossoff, George P.; Evans, David J.
Immobilisation of horseradish peroxidase (HRP) and glucose oxidase (GOX) via covalent attachment of modified enzyme carbohydrate to the exterior of the cowpea mosaic virus (CPMV) capsid gave high retention of enzymatic activity. The number of enzymes bound per virus was determined to be about eleven for HRP and 2-3 for GOX. This illustrates that relatively large biomacromolecules can be readily coupled to the virus surface using simple conjugation strategies. Virus-biomacromolecule hybrids have great potential for uses in catalysis, diagnostic assays or biosensors.Immobilisation of horseradish peroxidase (HRP) and glucose oxidase (GOX) via covalent attachment of modified enzyme carbohydrate to the exterior of the cowpea mosaic virus (CPMV) capsid gave high retention of enzymatic activity. The number of enzymes bound per virus was determined to be about eleven for HRP and 2-3 for GOX. This illustrates that relatively large biomacromolecules can be readily coupled to the virus surface using simple conjugation strategies. Virus-biomacromolecule hybrids have great potential for uses in catalysis, diagnostic assays or biosensors. Electronic supplementary information (ESI) available: Alternative conjugation strategies, agarose gel electrophoresis of CPMV and CPMV-HRP conjugates, UV-vis spectrum of HRP-ADHCPMV, agarose gel electrophoresis of GOX-ADHCPMV particles and corresponding TEM image, calibration curves for HRP-ADHCPMV and GOX-ADHCPMV, DLS data for GOX-ADHCPMV are made available. See DOI: 10.1039/c2nr31485a
Huynh, Nhung T; Hesketh, Emma L; Saxena, Pooja; Meshcheriakova, Yulia; Ku, You-Chan; Hoang, Linh T; Johnson, John E; Ranson, Neil A; Lomonossoff, George P; Reddy, Vijay S
Empty virus-like particles (eVLPs) of Cowpea mosaic virus (CPMV) are currently being utilized as reagents in various biomedical and nanotechnology applications. Here, we report the crystal structure of CPMV eVLPs determined using X-ray crystallography at 2.3 Å resolution and compare it with previously reported cryo-electron microscopy (cryo-EM) of eVLPs and virion crystal structures. Although the X-ray and cryo-EM structures of eVLPs are mostly similar, there exist significant differences at the C terminus of the small (S) subunit. The intact C terminus of the S subunit plays a critical role in enabling the efficient assembly of CPMV virions and eVLPs, but undergoes proteolysis after particle formation. In addition, we report the results of mass spectrometry-based proteomics analysis of coat protein subunits from CPMV eVLPs and virions that identify the C termini of S subunits undergo proteolytic cleavages at multiple sites instead of a single cleavage site as previously observed.
Pepino mosaic, once an emerging disease a decade ago, has become endemic on greenhouse tomatoes worldwide in recent years. Three distinct genotypes of Pepino mosaic virus (PepMV), including EU, US1 and CH2 have been recognized. Our earlier study in 2006-2007 demonstrated a predominant EU genotype ...
Grdzelishvili, Valery Z.; Garcia-Ruiz, Hernan; Watanabe, Tokiko; Ahlquist, Paul
Replication by many positive-strand RNA viruses includes genomic RNA amplification and subgenomic mRNA (sgRNA) transcription. For brome mosaic virus (BMV), both processes occur in virus-induced, membrane-associated compartments, require BMV replication factors 1a and 2a, and use negative-strand RNA3 as a template for genomic RNA3 and sgRNA syntheses. To begin elucidating their relations, we examined the interaction of RNA3 replication and sgRNA transcription in Saccharomyces cerevisiae expres...
Jeske, Holger; Gotthardt, Diether; Kober, Sigrid
The circular single-stranded DNAs of geminiviruses are multiplied efficiently and preferentially by rolling circle amplification (RCA), and can be diagnosed readily by restriction fragment length polymorphism (RFLP) and direct sequencing of the RCA product. Two strategies are described for cloning geminiviruses from plants harboring mixed infections by using RCA and RFLP with plant-derived nucleic acids without the need for bacterial amplification. By combining both these approaches, the true Sida micrantha mosaic virus was identified. The advantages of maintaining the quasispecies nature of a virus during in planta cloning is discussed with respect to reliable virus identification and resistance breeding.
Wyant, Patrícia Soares; Gotthardt, Diether; Schäfer, Benjamin; Krenz, Björn; Jeske, Holger
Begomovirus is the largest genus within the family Geminiviridae and includes economically important plant DNA viruses infecting a broad range of plant species and causing devastating crop diseases, mainly in subtropical and tropical countries. Besides cultivated plants, many weeds act as virus reservoirs. Eight begomovirus isolates from Bolivian weeds were examined using rolling-circle amplification (RCA) and restriction fragment length polymorphism (RFLP). An efficient, novel cloning strategy using limited Sau3A digestion to obtain tandem-repeat inserts allowed the sequencing of the complete genomes. The viruses were classified by phylogenetic analysis as typical bipartite New World begomoviruses. Four of them represented distinct new virus species, for which the names Solanum mosaic Bolivia virus, Sida mosaic Bolivia virus 1, Sida mosaic Bolivia virus 2, and Abutilon mosaic Bolivia virus are proposed. Three were variants of a new strain of Sida micrantha mosaic virus (SimMV), SimMV-rho[BoVi07], SimMV-rho[Bo:CF1:07] and SimMV-rho[Bo:CF2:07], and one was a new variant of a previously described SimMV, SimMV-MGS2:07-Bo.
Xu, D-L; Park, J-W; Mirkov, T E; Zhou, G-H
A survey of cultivated hybrid sugarcane (Saccharum inter-specific hybrid) and noble sugarcane (Saccharum officinarum) in southern China for the presence of Sugarcane mosaic virus (SCMV), Sorghum mosaic virus (SrMV) and Sugarcane streak mosaic virus (SCSMV) was conducted by RT-PCR from the years 2003 to 2006. SCMV and SrMV, but not SCSMV, were found. A high incidence of SCMV and SrMV coinfection was revealed in both hybrid and noble sugarcanes. All coinfected plants showed mosaic symptom, whereas plants infected with a single virus were symptomatic or asymptomatic. It appears that virus mixtures are more virulent than single infections. The nucleotide sequences of the coat protein (CP) gene of 33 SCMV and 10 SrMV isolates from this study were compared to those of CP genes of SCMV and SrMV reported in GenBank. One hundred and seventy-three SCMV isolates, with the exception of MDB and Abaca strains, can be grouped into five groups, which include three previously known groups, the sugarcane (SCE), maize (MZ), and Thailand groups, and two newly identified groups, the noble sugarcane (NSCE) and Brazil groups. Twenty-two SrMV isolates were divided into two groups, HS (hybrid sugarcane) and NS (noble sugarcane) groups. Five out of eight SrMV hybrid isolates belonged to the HS group, and two SrMV noble isolates and three hybrid isolates were within the NS group. Interestingly, the three hybrid isolates within the NS group were isolated from hybrid sugarcane co-infected with SCMV. This indicates that SCMV helps the NS group SrMV to infect hybrid sugarcane.
Choi, Hoseong; Jo, Yeonhwa; Lian, Sen; Jo, Kyoung-Min; Chu, Hyosub; Yoon, Ju-Yeon; Choi, Seung-Kook; Kim, Kook-Hyung; Cho, Won Kyong
The chrysanthemum is one of popular flowers in the world and a host for several viruses. So far, molecular interaction studies between the chrysanthemum and viruses are limited. In this study, we carried out a transcriptome analysis of chrysanthemum in response to three different viruses including Cucumber mosaic virus (CMV), Tomato spotted wilt virus (TSWV) and Potato virus X (PVX). A chrysanthemum 135K microarray derived from expressed sequence tags was successfully applied for the expression profiles of the chrysanthemum at early stage of virus infection. Finally, we identified a total of 125, 70 and 124 differentially expressed genes (DEGs) for CMV, TSWV and PVX, respectively. Many DEGs were virus specific; however, 33 DEGs were commonly regulated by three viruses. Gene ontology (GO) enrichment analysis identified a total of 132 GO terms, and of them, six GO terms related stress response and MCM complex were commonly identified for three viruses. Several genes functioning in stress response such as chitin response and ethylene mediated signaling pathway were up-regulated indicating their involvement in establishment of host immune system. In particular, TSWV infection significantly down-regulated genes related to DNA metabolic process including DNA replication, chromatin organization, histone modification and cytokinesis, and they are mostly targeted to nucleosome and MCM complex. Taken together, our comparative transcriptome analysis revealed several genes related to hormone mediated viral stress response and DNA modification. The identified chrysanthemums genes could be good candidates for further functional study associated with resistant to various plant viruses.
Full Text Available Abstract Background Plant viruses can be employed as versatile vectors for the production of vaccines by expressing immunogenic epitopes on the surface of chimeric viral particles. Although several viruses, including tobacco mosaic virus, potato virus X and cowpea mosaic virus, have been developed as vectors, we aimed to develop a new viral vaccine delivery system, a bamboo mosaic virus (BaMV, that would carry larger transgene loads, and generate better immunity in the target animals with fewer adverse environmental effects. Methods We engineered the BaMV as a vaccine vector expressing the antigenic epitope(s of the capsid protein VP1 of foot-and-mouth disease virus (FMDV. The recombinant BaMV plasmid (pBVP1 was constructed by replacing DNA encoding the 35 N-terminal amino acid residues of the BaMV coat protein with that encoding 37 amino acid residues (T128-N164 of FMDV VP1. Results The pBVP1 was able to infect host plants and to generate a chimeric virion BVP1 expressing VP1 epitopes in its coat protein. Inoculation of swine with BVP1 virions resulted in the production of anti-FMDV neutralizing antibodies. Real-time PCR analysis of peripheral blood mononuclear cells from the BVP1-immunized swine revealed that they produced VP1-specific IFN-γ. Furthermore, all BVP1-immunized swine were protected against FMDV challenge. Conclusion Chimeric BaMV virions that express partial sequence of FMDV VP1 can effectively induce not only humoral and cell-mediated immune responses but also full protection against FMDV in target animals. This BaMV-based vector technology may be applied to other vaccines that require correct expression of antigens on chimeric viral particles.
Kim, Sang-Min; Nam, Sang-Hyun; Lee, Jung-Myung; Yim, Kyu-Ock; Kim, Kook-Hyung
Heat treatment is commonly used to control viral contamination of seeds. To study virus inactivation, virus was purified from seeds contaminated with Cucumber green mottle mosaic virus (CGMMV) after various heat treatments. CGMMV particles were observed to be physically disrupted by high temperature. Analysis of viral RNA revealed that the 5' and 3' termini of the genome were protected whereas regions between 2-2.5, 3.2-3.7 and 4-4.8 kb from the 5' terminus were not. Heat inactivation of virus on seeds was confirmed by RT-PCR using primers for a heat-sensitive region. The RT-PCR approach developed here may prove preferable to time- and labor-intensive bioassays for assessing virus heat inactivation.
Kothandaraman, Satya Vijayalakshmi; Devadason, Alice; Ganesan, Malathi Varagur
The yellow mosaic viruses (YMV) infecting legumes are considered to be the most devastating begomoviruses as they incite considerable yield loss. The yellow discoloration of pods and seeds of infected plants and symptom emergence in the very first trifoliate leaf of the plants in the field were suggestive that the virus may be seed borne, which was investigated in the present study. The distribution of the virus in various parts of the seeds of black gram (Vigna mungo L. Hepper) plants naturally infected in the field was determined by polymerase chain reaction (PCR), Southern blot analysis, and sequencing. Nucleotide sequencing of the PCR amplicons from the seed parts from groups of ten seeds revealed the presence of mung bean yellow mosaic virus (MYMV) in the seed coat, cotyledon, and embryonic axes. The presence of virion particles was confirmed through double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) and immunosorbent electron microscopy (ISEM) even in a single whole seed. In confocal microscopy, positive fluorescent signals were obtained using coat protein gene-specific primers in the embryonic axes. However, in the growth tests performed with the same batch of seeds, there was no symptom development in the seedlings though the virus (both DNA A and B components) was detected in 32 % of tested seedlings. In this study, the MYMV was detected in seed coat, cotyledon, and embryo. This study revealed that the MYMV is a seed-borne virus.
Jebasingh, T; Pandaranayaka, Eswari P J; Mahalakshmi, A; Kasin Yadunandam, A; Krishnaswamy, S; Usha, R
The NIa protease of Potyviridae is the major viral protease that processes potyviral polyproteins. The NIa protease coding region of Cardamom mosaic virus (CdMV) is amplified from the viral cDNA, cloned and expressed in Escherichia coli. NIa protease forms inclusion bodies in E.coli. The inclusion bodies are solubilized with 8 M urea, refolded and purified by Nickel-Nitrilotriacetic acid affinity chromatography. Three-dimensional modeling of the CdMV NIa protease is achieved by threading approach using the homologous X-ray crystallographic structure of Tobacco etch mosaic virus NIa protease. The model gave an insight in to the substrate specificities of the NIa proteases and predicted the complementation of nearby residues in the catalytic triad (H42, D74 and C141) mutants in the cis protease activity of CdMV NIa protease.
于嘉林; 晏立英; 苏宁; 侯占军; 李大伟; 韩成贵; 杨莉莉; 蔡祝南; 刘仪
Wheat yellow mosaic virus (WYMV) isolate HC was used for viral cDNA synthesis and sequencing. The results show that the viral RNA1 is 7629 nueleotides encoding a polyprotein with 2407 amino acids, from which seven putative proteins may be produced by an autolytie cleavage processing besides the viral coat protein. The RNA2 is 3639 nueleotides and codes for a polypretein of 903 amino acids, which may contain two putative non-structural proteins. Although WYMV shares a similarity in genetic organization to wheat spindle streak mosaic virus (WSSMV), the identities in their nucleotide sequences or deduced amino acid sequences are as low as 70% and 75 % respectively. Based on this result, it is confirmed that WYMV and WSSMV are different species within Bymovirus.
AKM Mahbubul Alam; Prakit Somta; Choosak Jompuk; Prasert Chatwachirawong; Peerasak Srinives
This work was conducted to identify mungbean genotypes showing yield stability and resistance to mungbean yellow mosaic virus (MYMV) disease. Sixteen genotypes were evaluated in a randomized complete block design with two replications for two years (2011 and 2012) at three locations (Gazipur, Ishurdi and Madaripur) of the Bangladesh Agricultural Research Institute. An analysis of variance exhibited significant effects of genotype (G), environment (E), and genotype × environment (G×E) on grain...
Singh, D P
The inheritance of resistance to mungbean yellow mosaic virus (MYMV) was studied in blackgram (Vigna mungo (L.) Hepper). The highly resistant donors Pant U-84 and UPU-2 and a highly susceptible line, UL-2, their F1's, F2's and backcrosses were grown with spreader located every 5 to 6 rows. The resistance was found to be digenic and recessive in all the crosses and free from cytoplasmic effect.
Jianzhong Liu; Yuan Fang; Hongxi Pang
Soybean mosaic virus (SMV) is one of the most devastating pathogens that cost huge economic losses in soybean production worldwide. Due to the duplicated genome, clustered and highly homologous nature of R genes, as well as recalcitrant to transformation, soybean disease resistance studies is largely lagging compared with other diploid crops. In this review, we focus on the major advances that have been made in identifying both the virulence/avirulence factors of SMV and mapping of SMV resist...
Takamatsu, N; Watanabe, Y.; Iwasaki, T.; Shiba, T.; Meshi, T; Okada, Y.
To determine the sequences essential for viral multiplication in the 5' untranslated leader sequence of tobacco mosaic virus RNA, mutant TMV-L (a tomato strain) RNAs which carry several deletions in this 71-nucleotide sequence were constructed by an in vitro transcription system and their multiplication was analyzed by introducing mutant RNA into tobacco protoplasts by electroporation. Large deletions of the sequence from nucleotides 9 to 47 or 25 to 71 abolished viral multiplication; when ab...
Wang, Y; Lee, K C; Gaba, V; Wong, S M; Palukaitis, P; Gal-On, A
Resistance to the cucumovirus Cucumber mosaic virus (CMV) in cucumber cv. Delila was manifested as a very low level of accumulation of viral RNA and capsid protein, and an absence of CMV-induced symptoms. In addition, resistance was observed at the single cell level, with a reduction in accumulation of CMV RNAs, compared to accumulation in cells of the susceptible cucumber cv. Bet Alpha. Resistance to CMV in cv. Delila was broken by co-infection with the potyvirus Zucchini yellow mosaic virus (ZYMV). Resistance breakage in cv. Delila plants was manifested by an increase in the accumulation of (+) and (-) CMV RNA as well as CMV capsid protein, with no increase in the level of accumulation of ZYMV. Resistance breakage in the resistant cultivar by ZYMV also occurred at the single cell level. Thus, synergistic interactions known to occur between a potyvirus and a cucumovirus led to resistance breakage during a double infection. However, resistance breakage was not accompanied by an increase in disease symptoms beyond those induced by ZYMV itself. On co-inoculation with an asymptomatic variant of ZYMV-AG an enhancement of CMV infection occurred without disease manifestation. Consequently, intensification of viral RNA and capsid protein accumulation can occur without a corresponding increase in disease development, suggesting that different host genes regulate viral accumulation and disease development in the CMV-resistant cucumber plants.
Rodriguez-Alvarado, G.; Kurath, G.; Dodds, J.A.
Twenty-four cucumber mosaic cucumovirus (CMV) field isolates from pepper crops in Cali-fornia were characterized and compared by nucleic acid hybridization subgrouping, virion electrophoresis, and biological effects in several hosts. Isolates, belonging to subgroup I or subgroup II, were found that induced severe symptoms in mechanically inoculated bell pep-pers. Only two isolates, both from subgroup II, were mild. A group of 19 isolates collected from a single field were all in subgroup II and appeared identical by virion electrophoresis, but they exhibited varying degrees of symptom severity in peppers. As a more detailed indicator of heterogeneity, these 19 isolates were examined by RNase protection assays to delect sequence variation in the coat protein gene region of their genomes. The patterns of bands observed were complex and a high degree of genomic heterogeneity was detected between isolates, with no apparent correlation to symptomatology in bell pepper.
Choudhury, Nirupam Roy; Malik, Punjab Singh; Singh, Dharmendra Kumar; Islam, Mohammad Nurul; Kaliappan, Kosalai; Mukherjee, Sunil Kumar
Geminiviruses replicate by rolling circle mode of replication (RCR) and the viral Rep protein initiates RCR by the site-specific nicking at a conserved nonamer (TAATATT downward arrow AC) sequence. The mechanism of subsequent steps of the replication process, e.g. helicase activity to drive fork-elongation, etc. has largely remained obscure. Here we show that Rep of a geminivirus, namely, Mungbean yellow mosaic India virus (MYMIV), acts as a replicative helicase. The Rep-helicase, requiring > or =6 nt space for its efficient activity, translocates in the 3'-->5' direction, and the presence of forked junction in the substrate does not influence the activity to any great extent. Rep forms a large oligomeric complex and the helicase activity is dependent on the oligomeric conformation ( approximately 24mer). The role of Rep as a replicative helicase has been demonstrated through ex vivo studies in Saccharomyces cerevisiae and in planta analyses in Nicotiana tabacum. We also establish that such helicase activity is not confined to the MYMIV system alone, but is also true with at least two other begomoviruses, viz., Mungbean yellow mosaic virus (MYMV) and Indian cassava mosaic virus (ICMV).
Full Text Available Virus infection of plants may induce a variety of disease symptoms. However, little is known about the molecular mechanism of systemic symptom development in infected plants. Here we performed the first next-generation sequencing study to identify gene expression changes associated with disease development in tobacco plants (Nicotiana tabacum cv. Xanthi nc induced by infection with the M strain of Cucumber mosaic virus (M-CMV. Analysis of the tobacco transcriptome by RNA-Seq identified 95,916 unigenes, 34,408 of which were new transcripts by database searches. Deep sequencing was subsequently used to compare the digital gene expression (DGE profiles of the healthy plants with the infected plants at six sequential disease development stages, including vein clearing, mosaic, severe chlorosis, partial and complete recovery, and secondary mosaic. Thousands of differentially expressed genes were identified, and KEGG pathway analysis of these genes suggested that many biological processes, such as photosynthesis, pigment metabolism and plant-pathogen interaction, were involved in systemic symptom development. Our systematic analysis provides comprehensive transcriptomic information regarding systemic symptom development in virus-infected plants. This information will help further our understanding of the detailed mechanisms of plant responses to viral infection.
Babu, Binoy; Hegde, Vinayaka; Makeshkumar, T; Jeeva, M L
Reverse transcription polymerase chain reaction of the infected leaf samples of Colocasia esculenta plants showing severe whitish feathery symptoms were carried out using Potyvirus group specific primers, resulting in an amplicon of 327 bp, encoding the core region of the coat protein gene. Sequencing and BLAST analysis showed that the virus is distinct, closely related to Dasheen mosaic virus (DsMV). Sequence analysis revealed 86 and 96% identity at the nucleotide and amino acid level respectively with the DsMV isolate SY1(accession Number AJ628756). This is the first molecular level characterisation of the DsMV infecting C. esculenta in India.
D. R. Turner; Butler, P J
The assembly origin of tobacco mosaic virus RNA contains three stable hairpin loops. Coat protein disks bind first to loop 1 (the 3' most) during virus assembly, but the whole region is coated in a concerted fashion even in conditions of limiting protein. It is shown by in vitro packaging assays using mutant assembly origin transcripts that rapid and specific assembly initiation occurs in the absence of loops 2 and 3, but is abolished on removal of loop 1. Deletion or alteration of the unpair...
Bruun-Rasmussen, Marianne; Madsen, Christian Toft; Jessing, Stine;
Virus-induced gene silencing (VIGS) can be used as a powerful tool for functional genomics studies in plants. With this approach, it is possible to target most genes and downregulate the messenger (m)RNA in a sequence-specific manner. Barley stripe mosaic virus (BSMV) is an established VIGS vecto...... inoculation, although large parts of the insert had been lost from the virus vector. The instability of the insert, observed consistently throughout our experiments, offers an explanation for the transient nature of silencing when using BSMV as a VIGS vector.......Virus-induced gene silencing (VIGS) can be used as a powerful tool for functional genomics studies in plants. With this approach, it is possible to target most genes and downregulate the messenger (m)RNA in a sequence-specific manner. Barley stripe mosaic virus (BSMV) is an established VIGS vector...... for barley and wheat; however, silencing using this vector is generally transient, with efficient silencing often being confined to the first two or three systemically infected leaves. To investigate this further, part of the barley Phytoene desaturase (PDS) gene was inserted into BSMV and the resulting...
Bau, H-J; Kung, Y-J; Raja, J A J; Chan, S-J; Chen, K-C; Chen, Y-K; Wu, H-W; Yeh, S-D
A virus identified as a new pathotype of Papaya leaf distortion mosaic virus (PLDMV, P-TW-WF) was isolated from diseased papaya in an isolated test-field in central Taiwan, where transgenic papaya lines resistant to Papaya ringspot virus (PRSV) were evaluated. The infected plants displayed severe mosaic, distortion and shoe-stringing on leaves; stunting in apex; and water-soaking on petioles and stems. This virus, which did not react in enzyme-linked immunosorbent assay with the antiserum to the PRSV coat protein, infected only papaya, but not the other 18 plant species tested. Virions studied under electron microscope exhibited morphology and dimensions of potyvirus particles. Reverse transcription-polymerase chain reaction conducted using potyvirus-specific primers generated a 1,927-nucleotide product corresponding to the 3' region of a potyvirus, showing high sequence identity to the CP gene and 3' noncoding region of PLDMV. Search for similar isolates with the antiserum against CP of P-TW-WF revealed scattered occurrence of PLDMV in Taiwan. Phylogenetic analysis of PLDMV isolates of Taiwan and Japan indicated that the Taiwan isolates belong to a separate genetic cluster. Since all the Taiwan isolates infected only papaya, unlike the cucurbit-infecting Japanese P type isolates, the Taiwan isolates are considered a new pathotype of PLDMV. Susceptibility of all our PRSV-resistant transgenic papaya lines to PLDMV indicates that the virus is an emerging threat for the application of PRSV-resistant transgenic papaya in Taiwan and elsewhere.
GRAZIELA DA SILVA BARBOSA
Full Text Available Mixed infections in cucurbits are frequently observed in natural conditions between viruses from the Potyvirus genus and Cucumber mosaic virus (CMV, which significantly decreases productivity. The objectives of the present study was to compare the host range of PRSV - W, WMV, and ZYMV isolates and evaluate the effects of mixed infections with CMV in zucchini plants ( Cucurbita pepo L.. Host range studies comprising 23 plant species confirmed some similarities and biological differences among the isolates of PRSV - W, ZYMV, and WMV. RT - PCR confirmed the amplification of DNA fragments of the PRSV - W, WMV, and ZYMV coat protein gene ( cp and cytoplasm inclusion gene ( ci . The virus interaction studies in zucchini Caserta plants indicated synergistic interactions, particularly among species from the Potyvirus genus, and some CMV interference with some virus combinations.
Maoka, Tetsuo; Hataya, Tatsuji
ABSTRACT The complete nucleotide sequence of the genome of Papaya leaf distortion mosaic virus (PLDMV) was determined. The viral RNA genome of strain LDM (leaf distortion mosaic) comprised 10,153 nucleotides, excluding the poly(A) tail, and contained one long open reading frame encoding a polyprotein of 3,269 amino acids (molecular weight 373,347). The polyprotein contained nine putative proteolytic cleavage sites and some motifs conserved in other potyviral polyproteins with 44 to 50% identities, indicating that PLDMV is a distinct species in the genus Potyvirus. Like the W biotype of Papaya ringspot virus (PRSV), the non-papaya-infecting biotype of PLDMV (PLDMV-C) was found in plants of the family Cucurbitaceae. The coat protein (CP) sequence of PLDMV-C in naturally infected-Trichosanthes bracteata was compared with those of three strains of the P biotype (PLDMV-P), LDM and two additional strains M (mosaic) and YM (yellow mosaic), which are biologically different from each other. The CP sequences of three strains of PLDMV-P share high identities of 95 to 97%, while they share lower identities of 88 to 89% with that of PLDMV-C. Significant changes in hydrophobicity and a deletion of two amino acids at the N-terminal region of the CP of PLDMV-C were observed. The finding of two biotypes of PLDMV implies the possibility that the papaya-infecting biotype evolved from the cucurbitaceae-infecting potyvirus, as has been previously suggested for PRSV. In addition, a similar evolutionary event acquiring infectivity to papaya may arise frequently in viruses in the family Cucurbitaceae.
Praveen, Shelly; Mangrauthia, Satendra K; Singh, Priyanka; Mishra, Anil K
The 2b protein encoded by Cucumber mosaic virus (CMV) has been shown as a virus counter defense factor that interferes with the RNAi pathway. The 2b gene from CMV-banana, New Delhi isolate (CMV-NDLS) was amplified from CMV infected cucumber plants to generate the sense and antisense binary vector constructs for 2b expression and repression in planta. Constitutive expression of 2b gene in healthy Nicotiana tabacum caused phenotypic aberrations during somatic embryogenesis, which were not observed when expressed in CMV infected N. tabacum. Further, the established virus population in CMV infected N. tabacum was not affected by constitutive expression and repression of 2b gene. Thus, indicating its role in initiation of gene silencing, at the early stage of viral infection. This is the first demonstration of differential behavior of 2b suppressor protein in host development in the absence and presence of virus.
杨恭; 邱并生; 魏军亚; 刘广超
Our previous reports showed that one opal mutation (UGA) and one ochre mutation (UAA) respectively located in the replicase and movement protein (MP) genes of the attenuated tomato mosaic virus K(ToMV-K) contribute to the viral attenuation. To explore a wider application of this attenuation pattern to other plant viruses, we have constructed three mutants which respectively contain one opal mutation of the replicase gene and/or one ochre mutation of the MP using PCR-mediated site-directed mutagenesis from a virulent tobacco mosaic virus isolated from China (TMV-Cv). Plant infection performed by in vitro transcripts revealed that the MP truncated mutant TMV-Cvmp and the replicase-MP truncated mutant TMV-Cvrase-mp were infectious on both local lesion (Nicotiana tabacum cv. Xanthi NC) and systemic (N. tabacum cv. K326) host plants, while the replicase truncated mutant TMV-Cvrase was non-infectious. The K326 plant infected by TMV- Cvrease-mp displayed only a little mild mosaic. By electronic microscopy (EM), plant re-inoculation, RNA Dot-blot, RT-PCR and sequencing we demonstrated that the progeny viruses of TMV-Cvmp and TMV-Cvrease-mp shared similar morphological character with TMV-Cv, owned the abilities to infect, replicate and propagate in the assayed plants, and maintained the mutated sites during infection. These data showed that both the opal and the ochre mutations are able to cooperatively induce the attenuated phenotypes of TMV-Cvrase-mp on plants, indicating that the mutation pattern of ToMV-K could be used to attenuate other virulent plant viruses.
Replication of many positive-stranded RNA viruses takes place in association with intracellular membranes. Often these membranes are induced upon infection by vesiculation or rearrangement of membranes from different organelles including the early and late endomembrane system. Upon infection of cowp
Wel, van der N.N.
For a full infection of a host, plant viruses should be able to multiply in the initially infected cell and to spread to neighbouring cells as to eventually invade the entire plant. The viral transport pathway can in principle be divided into two steps, i.e. cell-to-cell movement within tissues, and
Bhat, Sumana; Folimonova, Svetlana Y; Cole, Anthony B; Ballard, Kimberly D; Lei, Zhentian; Watson, Bonnie S; Sumner, Lloyd W; Nelson, Richard S
Tobacco mosaic virus (TMV) forms dense cytoplasmic bodies containing replication-associated proteins (virus replication complexes [VRCs]) upon infection. To identify host proteins that interact with individual viral components of VRCs or VRCs in toto, we isolated viral replicase- and VRC-enriched fractions from TMV-infected Nicotiana tabacum plants. Two host proteins in enriched fractions, ATP-synthase γ-subunit (AtpC) and Rubisco activase (RCA) were identified by matrix-assisted laser-desorption ionization time-of-flight mass spectrometry or liquid chromatography-tandem mass spectrometry. Through pull-down analysis, RCA bound predominantly to the region between the methyltransferase and helicase domains of the TMV replicase. Tobamovirus, but not Cucumber mosaic virus or Potato virus X, infection of N. tabacum plants resulted in 50% reductions in Rca and AtpC messenger RNA levels. To investigate the role of these host proteins in TMV accumulation and plant defense, we used a Tobacco rattle virus vector to silence these genes in Nicotiana benthamiana plants prior to challenge with TMV expressing green fluorescent protein. TMV-induced fluorescent lesions on Rca- or AtpC-silenced leaves were, respectively, similar or twice the size of those on leaves expressing these genes. Silencing Rca and AtpC did not influence the spread of Tomato bushy stunt virus and Potato virus X. In AtpC- and Rca-silenced leaves TMV accumulation and pathogenicity were greatly enhanced, suggesting a role of both host-encoded proteins in a defense response against TMV. In addition, silencing these host genes altered the phenotype of the TMV infection foci and VRCs, yielding foci with concentric fluorescent rings and dramatically more but smaller VRCs. The concentric rings occurred through renewed virus accumulation internal to the infection front.
Heijden, van der M.W.; Carette, J.E.; Reinhoud, P.J.; Haegi, A.; Bol, J.F.
Replication of Alfalfa mosaic virus (AMV) RNAs depends on the virus-encoded proteins P1 and P2. P1 contains methyltransferase- and helicase-like domains, and P2 contains a polymerase-like domain. Coimmunoprecipitation experiments revealed an interaction between in vitro translated-P1 and P2 and show
Full Text Available As one of the critical diseases of sugarcane, sugarcane mosaic disease can lead to serious decline in stalk yield and sucrose content. It is mainly caused by Potyvirus sugarcane mosaic virus (SCMV and/or Sorghum mosaic virus (SrMV, with additional differences in viral strains. RNA interference (RNAi is a novel strategy for producing viral resistant plants. In this study, based on multiple sequence alignment conducted on genomic sequences of different strains and isolates of SrMV, the conserved region of coat protein (CP genes was selected as the target gene and the interference sequence with size of 423 bp in length was obtained through PCR amplification. The RNAi vector pGII00-HACP with an expression cassette containing both hairpin interference sequence and cp4-epsps herbicide-tolerant gene was transferred to sugarcane cultivar ROC22 via Agrobacterium-mediated transformation. After herbicide screening, PCR molecular identification, and artificial inoculation challenge, anti-SrMV positive transgenic lines were successfully obtained. SrMV resistance rate of the transgenic lines with the interference sequence was 87.5% based on SrMV challenge by artificial inoculation. The genetically modified SrMV-resistant lines of cultivar ROC22 provide resistant germplasm for breeding lines and can also serve as resistant lines having the same genetic background for study of resistance mechanisms.
Borah, Basanta K; Johnson, A M Anthony; Sai Gopal, D V R; Dasgupta, Indranil
Citrus yellow mosaic badna virus (CMBV), a member of the Family Caulimoviridae, Genus Badnavirus, is the causative agent of Citrus mosaic disease in India. Although the virus has been detected in several citrus species, only two full-length genomes, one each from Sweet orange and Rangpur lime, are available in publicly accessible databases. In order to obtain a better understanding of the genetic variability of the virus in other citrus mosaic-affected citrus species, we performed the cloning and sequence analysis of complete genomes of CMBV from two additional citrus species, Acid lime and Pummelo. We show that CMBV genomes from the two hosts share high homology with previously reported CMBV sequences and hence conclude that the new isolates represent variants of the virus present in these species. Based on in silico sequence analysis, we predict the possible function of the protein encoded by one of the five ORFs.
Madhubala, R; Bhadramurthy, V; Bhat, A I; Hareesh, P S; Retheesh, S T; Bhai, R S
Cucumber mosaic virus (CMV) causing mosaic, leaf distortion and stunting of vanilla (Vanilla planifolia Andrews) in India was characterized on the basis of biological and coat protein (CP) nucleotide sequence properties. In mechanical inoculation tests, the virus was found to infect members of Chenopodiaceae, Cucurbitaceae, Fabaceae and Solanaceae. Nicotiana benthamiana was found to be a suitable host for the propagation of CMV. The virus was purified from inoculated N. benthamiana plants and negatively stained purified preparations contained isometric particles of about 28 nm in diameter. The molecular weight of the viral coat protein subunits was found to be 25.0 kDa. Polyclonal antiserum was produced in New Zealand white rabbit, immunoglobulin G (IgG) was purified and conjugated with alkaline phosphatase enzyme. Double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) method was standardized for the detection of CMV infection in vanilla plants. CP gene of the virus was amplified using reverse transcriptase-polymerase chain reaction (RT-PCR), cloned and sequenced. Sequenced region contained a single open reading frame of 657 nucleotides potentially coding for 218 amino acids. Sequence analyses with other CMV isolates revealed the greatest identity with black pepper isolate of CMV (99%) and the phylogram clearly showed that CMV infecting vanilla belongs to subgroup IB. This is the first report of occurrence of CMV on V. planifolia from India.
R Madhubala; V Bhadramurthy; A I Bhat; P S Hareesh; S T Retheesh; R S Bhai
Cucumber mosaic virus (CMV) causing mosaic, leaf distortion and stunting of vanilla (Vanilla planifolia Andrews) in India was characterized on the basis of biological and coat protein (CP) nucleotide sequence properties. In mechanical inoculation tests, the virus was found to infect members of Chenopodiaceae, Cucurbitaceae, Fabaceae and Solanaceae. Nicotiana benthamiana was found to be a suitable host for the propagation of CMV. The virus was purified from inoculated N. benthamiana plants and negatively stained purified preparations contained isometric particles of about 28 nm in diameter. The molecular weight of the viral coat protein subunits was found to be 25.0 kDa. Polyclonal antiserum was produced in New Zealand white rabbit, immunoglobulin G (IgG) was purified and conjugated with alkaline phosphatase enzyme. Double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) method was standardized for the detection of CMV infection in vanilla plants. CP gene of the virus was amplified using reverse transcriptase-polymerase chain reaction (RT-PCR), cloned and sequenced. Sequenced region contained a single open reading frame of 657 nucleotides potentially coding for 218 amino acids. Sequence analyses with other CMV isolates revealed the greatest identity with black pepper isolate of CMV (99%) and the phylogram clearly showed that CMV infecting vanilla belongs to subgroup IB. This is the first report of occurrence of CMV on V. planifolia from India.
Ming-Min ZHAO; De-Rong AN; Jian ZHAO; Guang-Hua HUANG; Zu-Hua HE; Jiang-Ye CHEN
RNA interference (RNAi) silences gene expression by guiding mRNA degradation in asequence-specific fashion. Small interfering RNA (siRNA), an intermediate of the RNAi pathway, has been shown to be very effective in inhibiting virus infection in mammalian cells and cultured plant cells. Here, we report that Agrobacterium tumefaciens-mediated transient expression of short hairpin RNA (shRNA) could inhibit tobacco mosaic virus (TMV) RNA accumulation by targeting the gene encoding the replication-associated 126 kDa protein in intact plant tissue. Our results indicate that transiently expressed shRNA efficiently interfered with TMV infection. The interference observed is sequence-specific, and time- and site-dependent.Transiently expressed shRNA corresponding to the TMV 126 kDa protein gene did not inhibit cucumber mosaic virus (CMV), an unrelated tobamovirus. In order to interfere with TMV accumulation in tobacco leaves, it is essential for the shRNA constructs to be infiltrated into the same leaves as TMV inoculation. Our results support the view that RNAi opens the door for novel therapeutic procedures against virus diseases.We propose that a combination of the RNAi technique and Agrobacterium-mediated transient expression could be employed as a potent antiviral treatment in plants.
Rouhibakhsh, A; Choudhury, N R; Mukherjee, S K; Malathi, V G
Yellow mosaic disease causes severe yield loss in grain legumes in Indian subcontinent and south east Asia. The disease is caused by two virus species, Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV). They have genome organization typical of Old World begomoviruses, the unique feature being the presence of an open reading frame (ORF) AV2 upstream of coat protein gene. In order to elucidate its function, ORF AV2 of blackgram isolate, Mungbean yellow mosaic India virus-[India:New Delhi:Blackgram 3:1991] MYMIV-[IN:ND:Bg3:91] and cowpea isolate, Mungbean yellow mosaic India virus-[India:New Delhi:Cowpea7:1998] MYMIV-[IN:ND:Cp7:98], respectively, were over expressed in Escherichia coli in fusion with maltose binding protein (MBP). The recombinant protein did not show efficient binding to DNA. However, both MBP-BgAV2 and MBP-CpAV2 proteins modulated nicking and ATPase activity of replication initiation protein (Rep). Even low concentration, 20 ng of MBP-BgAV2 and MBP-CpAV2 could bring 20 folds increase in nicking activity of Rep. Similarly in the presence of AV2 protein, two to three fold increase in ATPase activity was observed. It is hypothesized that AV2 protein may play a role of accessory protein modulating Rep activities.
Balint, R.; Cohen, S.S.
Turnip yellow mosaic virus contains large amounts of nonexchangeable spermidine and induces an accumulation of spermidine in infected Chinese cabbage. By 7 days after inoculation, a majority of protoplasts isolated from newly emerging leaves stain with fluorescent antibody to the virus. (/sup 14/C)Spermidine (10 microM) was taken up by these cells in amounts comparable to the original endogenous pool within 24 hr. However, after an initial rise, the spermidine content of the cell returned to its original level, implying considerable regulation of the endogenous pool(s). Putrescine and spermine were major products of the metabolism of exogenous spermidine. Radioactivity from exogenous (/sup 14/C)spermidine was also readily incorporated into the ribonucleoprotein component(s) of the virus, where it appeared as both spermidine and spermine. The specific radioactivities of the viral polyamines were approximately twice those of spermidine and spermine extracted from the whole cell. Radioactivity from (2-/sup 14/C)methionine was readily incorporated into the protein, spermidine, and spermine of the virus. Again, the specific activities of these amines were substantially higher in the virus than in the whole cell. Thus, newly formed virus contained predominantly newly synthesized spermidine and spermine. However, inhibition of spermidine synthesis by dicyclohexylamine led to incorporation of preexisting spermidine and increased amounts of spermine into newly formed virus.
Full Text Available A virus was isolated, using mechanical inoculation, from hill mustard (Bunias orientalis L. plants exhibiting yellow mottling and blistering on leaves, which were frequently accompanied by asymmetric leaf narrowing. It systemically infected certain plants from the family Brassicaceae (Brassica rapa, Bunias orientalis, Hesperis matronalis, Sinapis alba as well as Cleome spinosa and Nicotiana clevelandii, and locally Atriplex hortensis, Chenopodium quinoa, Ch. amaranticolor, N. tabacum. In the sap, it maintained infectivity for 3-4 days and lost it after heating for 10 min. at a temperature of 55 - 60oC or when diluted with water at 10-3. Virus particles were thread- like with a length of 675 - 710 nm. Based on an analysis of biological properties of the pathogen, serological response, particle morphology and data from field observations, it was identified as an isolate of Turnip mosaic virus (TuMV, and hill mustard was recognised as a natural overwintering host for this pathogen.
Kondo, Hideki; Takemoto, Shogo; Maruyama, Kazuyuki; Chiba, Sotaro; Andika, Ida Bagus; Suzuki, Nobuhiro
Cymbidium chlorotic mosaic virus (CyCMV), isolated from a spring orchid (Cymbidium goeringii), was characterized molecularly. CyCMV isometric virions comprise a single, positive-strand RNA genome of 4,083 nucleotides and 30-kDa coat protein. The virus genome contains five overlapping open reading frames with a genomic organization similar to that of sobemoviruses. BLAST searches and phylogenetic analysis revealed that CyCMV is most closely related to papaya lethal yellowing virus, a proposed dicot-infecting sobemovirus (58.8 % nucleotide sequence identity), but has a relatively distant relationship to monocot-infecting sobemoviruses, with only modest sequence identities. This suggests that CyCMV is a new monocot-infecting member of the floating genus Sobemovirus.
Su, Xiu; Fu, Shuai; Qian, Yajuan; Zhang, Liqin; Xu, Yi; Zhou, Xueping
A novel potyvirus was discovered in pecan (Carya illinoensis) showing leaf mosaic symptom through the use of deep sequencing of small RNAs. The complete genome of this virus was determined to comprise of 9,310 nucleotides (nt), and shared 24.0% to 58.9% nucleotide similarities with that of other Potyviridae viruses. The genome was deduced to encode a single open reading frame (polyprotein) on the plus strand. Phylogenetic analysis based on the whole genome sequence and coat protein amino acid sequence showed that this virus is most closely related to Lettuce mosaic virus. Using electron microscopy, the typical Potyvirus filamentous particles were identified in infected pecan leaves with mosaic symptoms. Our results clearly show that this virus is a new member of the genus Potyvirus in the family Potyviridae. The virus is tentatively named Pecan mosaic-associated virus (PMaV). Additionally, profiling of the PMaV-derived small RNA (PMaV-sRNA) showed that the most abundant PMaV-sRNAs were 21-nt in length. There are several hotspots for small RNA production along the PMaV genome; two 21-nt PMaV-sRNAs starting at 811 nt and 610 nt of the minus-strand genome were highly repeated.
Li, Yanan; Zhang, Jing; Zhao, Feifei; Ren, Han; Zhu, Lin; Xi, Dehui; Lin, Honghui
Cross protection is a common phenomenon among closely related strain viruses in co-infected plants. However, unrelated viruses, Turnip crinkle virus (TCV) and Cucumber mosaic virus (CMV), also show an antagonistic effect in co-infected Arabidopsis plants. In many cases, viral suppressors of RNA silencing (VSRs) have important roles in the interactions between viruses in mixed infections. CMV 2b and TCV p38 are multifunctional proteins and both of them are well characterized VSRs and have important roles in operation synergistic interactions with other viruses. Here, we demonstrated antagonistic effects of TCV toward CMV and showed that RNA silencing-mediated resistance protein, RCY1 and TCV-interacting protein (TIP) of Arabidopsis plants did not affect this antagonism effect. We further showed that TCV p38 and CMV 2b could interact with each other in vivo but not in vitro. Further mutational analysis showed that C-terminal of 2b and middle domains of p38 had more important roles in the interaction between the two viruses.
Desbiez, C; Justafre, I; Lecoq, H
Zucchini yellow fleck virus (ZYFV, genus Potyvirus) infects cultivated or wild cucurbits in the Mediterranean basin and occasionally causes severe damage in crops. Biological and serological data tend to indicate that ZYFV is related to other cucurbit-infecting potyviruses, mainly papaya ringspot virus (PRSV) and Moroccan watermelon mosaic virus (MWMV). In order to establish unambiguously the taxonomic status of ZYFV, the sequence of the 3' part of the genome - encompassing the CP coding region - of two ZYFV strains originating from Italy and France was obtained and compared with other potyviruses. The results obtained indicate that ZYFV belongs to a distinct potyvirus species, related to but different from PRSV and MWMV.
Dynamics of small RNA profiles of virus and host origin in wheat cultivars synergistically infected by Wheat streak mosaic virus and Triticum mosaic virus: virus infection caused a drastic shift in the endogenous small RNA profile.
Tatineni, Satyanarayana; Riethoven, Jean-Jack M; Graybosch, Robert A; French, Roy; Mitra, Amitava
Co-infection of wheat (Triticum aestivum L.) by Wheat streak mosaic virus (WSMV, a Tritimovirus) and Triticum mosaic virus (TriMV, a Poacevirus) of the family Potyviridae causes synergistic interaction. In this study, the effects of the synergistic interaction between WSMV and TriMV on endogenous and virus-derived small interfering RNAs (vsiRNAs) were examined in susceptible ('Arapahoe') and temperature-sensitive resistant ('Mace') wheat cultivars at 18°C and 27°C. Single and double infections in wheat caused a shift in the profile of endogenous small RNAs from 24 nt being the most predominant in healthy plants to 21 nt in infected wheat. Massive amounts of 21 and 22 nt vsiRNAs accumulated in singly and doubly infected Arapahoe at both temperatures and in Mace at 27°C but not 18°C. The plus- and minus-sense vsiRNAs were distributed throughout the genomic RNAs in Arapahoe at both temperature regimens and in Mace at 27°C, although some regions served as hot-spots, spawning an excessive number of vsiRNAs. The vsiRNA peaks were conserved among cultivars, suggesting that the Dicer-like enzymes in susceptible and resistant cultivars similarly accessed the genomic RNAs of WSMV or TriMV. Accumulation of large amounts of vsiRNAs in doubly infected plants suggests that the silencing suppressor proteins encoded by TriMV and WSMV do not prevent the formation of vsiRNAs; thus, the synergistic effect observed is independent from RNA-silencing mediated vsiRNA biogenesis. The high-resolution map of endogenous and vsiRNAs from WSMV- and/or TriMV-infected wheat cultivars may form a foundation for understanding the virus-host interactions, the effect of synergistic interactions on host defense, and virus resistance mechanisms in wheat.
Mi Sa Vo Phan
Full Text Available Recently, a Cucumber mosaic virus (CMV strain, named as CMV-209, was isolated from Glycine soja. In this study, symptom expression of CMV-209 was analyzed in detail in Nicotiana benthamiana by comparing with that of CMV-Fny, which is a representative strain of CMV. Using infectious cDNA clones of CMV strains 209 and Fny, symptom expression of various pseudorecombinants between these two strains were examined in the early and late infection stages. In the early infection stage, the pseudorecombinants containing Fny-RNA2 induced stunting and leaf distortion on the newly emerged leaves whereas the pseudorecombinants containing 209-RNA2 caused no obvious symptoms. In the late infection stage, the pseudorecombinants containing 209-RNA1 and Fny-RNA2 induced severe leaf distortion and stunting, while CMV-209 induced mild symptom and CMV-Fny caused typical mosaic, general stunting, and leaf distortion symptoms, indicating that RNA 2 encodes a symptom determinant(s of CMV, which is capable of enhancing symptoms. Furthermore, our results support the possibility that natural recombination between compatible viruses can result in emergence of novel viruses causing severe damages in crop fields.
Phan, Mi Sa Vo; Seo, Jang-Kyun; Choi, Hong-Soo; Lee, Su-Heon; Kim, Kook-Hyung
Recently, a Cucumber mosaic virus (CMV) strain, named as CMV-209, was isolated from Glycine soja. In this study, symptom expression of CMV-209 was analyzed in detail in Nicotiana benthamiana by comparing with that of CMV-Fny, which is a representative strain of CMV. Using infectious cDNA clones of CMV strains 209 and Fny, symptom expression of various pseudorecombinants between these two strains were examined in the early and late infection stages. In the early infection stage, the pseudorecombinants containing Fny-RNA2 induced stunting and leaf distortion on the newly emerged leaves whereas the pseudorecombinants containing 209-RNA2 caused no obvious symptoms. In the late infection stage, the pseudorecombinants containing 209-RNA1 and Fny-RNA2 induced severe leaf distortion and stunting, while CMV-209 induced mild symptom and CMV-Fny caused typical mosaic, general stunting, and leaf distortion symptoms, indicating that RNA 2 encodes a symptom determinant(s) of CMV, which is capable of enhancing symptoms. Furthermore, our results support the possibility that natural recombination between compatible viruses can result in emergence of novel viruses causing severe damages in crop fields.
Marcos Cesar Gonçalves
Full Text Available O objetivo deste trabalho foi caracterizar biológica e molecularmente três isolados de Sugarcane mosaic virus (SCMV de lavouras de milho, analisá-los filogeneticamente e discriminar polimorfismos do genoma. Plantas com sintomas de mosaico e nanismo foram coletadas em lavouras de milho, no Estado de São Paulo e no Município de Rio Verde, GO, e seus extratos foliares foram inoculados em plantas indicadoras e submetidos à análise sorológica com antissoros contra o SCMV, contra o Maize dwarf mosaic virus (MDMV e contra o Johnsongrass mosaic virus (JGMV. Mudas de sorgo 'Rio' e 'TX 2786' apresentaram sintomas de mosaico após a inoculação dos três isolados, e o DAS-ELISA confirmou a infecção pelo SCMV. O RNA total foi extraído e usado para amplificação por transcriptase reversa seguida de reação em cadeia de polimerase (RT-PCR. Fragmentos específicos foram amplificados, submetidos à análise por polimorfismo de comprimento de fragmento de restrição (RFLP e sequenciados. Foi possível discriminar os genótipos de SCMV isolados de milho de outros isolados brasileiros do vírus. Alinhamentos múltiplos e análises dos perfis filogenéticos corroboram esses dados e mostram diversidade nas sequências de nucleotídeos que codificam para a proteína capsidial, o que explica o agrupamento separado desses isolados e sugere sua classificação como estirpes distintas, em lugar de simples isolados geográficos.
Wang, Lin; Liu, Zhanmin; Xia, Xueying; Yang, Cuiyun; Huang, Junyi; Wan, Sibao
Cucumber green mottle mosaic virus (CGMMV)causes a severe mosaic symptom of watermelon and cucumber, and can be transmitted via infected cucumber seeds, leaves and soil. It remains a challenge to detect this virus to prevent its introduction and infection and spread in fields. For this purpose, a simple and sensitive label-free colorimetric detection method for CGMMV has been developed with unmodified gold nanoparticles (AuNPs) as colorimetric probes. The method is based on the finding that the presence of RT-PCR target products of CGMMV and species-specific probes results in color change of AuNPs from red to blue after NaCl induction. Normally, species-specific probes attach to the surface of AuNPs and thereby increasing their resistance to NaCl-induced aggregation. The concentration of sodium, probes in the reaction system and evaluation of specificity and sensitivity of a novel assay, visual detection of Cucumber green mottle mosaic virus using unmodified AuNPs has been carried out with simple preparation of samples in our study. Through this assay, as low as 30pg/μL of CGMMV RNA was thus detected visually, by the naked eye, without the need for any sophisticated, expensive instrumentation and biochemical reagents. The specificity was 100% and exhibited good reproducibility in our assays. The results note that this assay is highly species-specific, simple, low-cost, and visual for easy detection of CGMMV in plant tissues. Therefore, visual assay is a potentially useful tool for middle or small-scales corporations and entry-exit inspection and quarantine bureau to detect CGMMV in cucumber seeds or plant tissues.
Maredza, A T; Allie, F; Plata, G; Rey, M E C
Cassava is an important food security crop in Sub-Saharan Africa. Two episomal begomovirus-associated sequences, named Sequences Enhancing Geminivirus Symptoms (SEGS1 and SEGS2), were identified in field cassava affected by the devastating cassava mosaic disease (CMD). The sequences reportedly exacerbated CMD symptoms in the tolerant cassava landrace TME3, and the model plants Arabidopsis thaliana and Nicotiana benthamiana, when biolistically co-inoculated with African cassava mosaic virus-Cameroon (ACMV-CM) or East African cassava mosaic virus-UG2 (EACMV-UG2). Following the identification of small SEGS fragments in the cassava EST database, the intention of this study was to confirm their presence in the genome, and investigate a possible role for these sequences in CMD. We report that multiple copies of varying lengths of both SEGS1 and SEGS2 are widely distributed in the sequenced cassava genome and are present in several other cassava accessions screened by PCR. The endogenous SEGS1 and SEGS2 are in close proximity or overlapping with cassava genes, suggesting a possible role in regulation of specific biological processes. We confirm the expression of SEGS in planta using EST data and RT-PCR. The sequence features of endogenous SEGS (iSEGS) are unique but resemble non-autonomous transposable elements (TEs) such as MITEs and helitrons. Furthermore, many SEGS-associated genes, some involved in virus-host interactions, are differentially expressed in susceptible (T200) and tolerant TME3) cassava landraces infected by South African cassava mosaic virus (SACMV) of susceptible (T200) and tolerant (TME3) cassava landraces. Abundant SEGS-derived small RNAs were also present in mock-inoculated and SACMV-infected T200 and TME3 leaves. Given the known role of TEs and associated genes in gene regulation and plant immune responses, our observations are consistent with a role of these DNA elements in the host's regulatory response to geminiviruses.
Kuruba, B L; Buvani, A P; Veluthambi, K
Mungbean yellow mosaic virus-[India:Vigna] (MYMV-[IN:Vig]), a blackgram isolate of MYMV, causes yellow mosaic disease in blackgram and mungbean. Two variable DNA-B components, KA22 and KA27, cause distinct symptoms in blackgram [V. mungo (L.) Hepper] with the same DNA-A component. KA22 + DNA-A-agroinoculated blackgram plants displayed yellow mosaic symptom and accumulated high levels of viral single-stranded (ss) DNA. KA27 + DNA-A-agroinoculated blackgram plants displayed severe stunting symptom and accumulated very low levels of viral ssDNA. However, in mungbean [V. radiata (L.) Wilczek], KA27 + DNA-A caused yellow mosaic symptom and a high level of viral ssDNA accumulated. Swapping of KA27 DNA-B with the nuclear shuttle protein gene (NSP) of KA22 DNA-B (KA27xKA22 NSP) caused yellow mosaic symptom in blackgram, suggesting that KA22 NSP is the determinant of yellow mosaic symptom. Interestingly, KA27xKA22 NSP-infected blackgram plants accumulated high levels of viral ssDNA, comparable to that of KA22 DNA-B infection, suggesting that the KA22 NSP is responsible for accumulation of high levels of viral ssDNA. MYMV distribution was studied in blackgram and mungbean plants by leaf tissue hybridization, which showed mesophyll spread of the virus in KA22-infected blackgram leaflets and in KA27-infected mungbean leaflets, both of which displayed yellow mosaic symptom. However, the virus did not accumulate in the mesophyll in the case of KA27-infected blackgram leaflets. Interestingly, the swapped KA27xKA22 NSP-infected blackgram leaflets showed mesophyll accumulation of the virus, suggesting that KA22 NSP determines its mesophyll spread.
Morinaga, T; Ikegami, M; Miura, K
Viral single-stranded DNA of mung bean yellow mosaic virus (MYMV) was converted to the double-stranded state in vitro, and physical mapping was carried out. The genome of MYMV was found to consist of two major components (designated as DNA 1 and DNA 2). In addition, some minor components were detected. Molecular cloning of the major components was carried out, using in vitro double-stranded DNA and replicative intermediate DNAs. DNA 1 is about 2.72 and DNA 2 about 2.67 kilobase pairs. No similarities were observed when the two restriction maps of DNA 1 and 2 were compared.
Full Text Available We developed a loop-mediated isothermal amplification (LAMP method to rapidly diagnose Wheat streak mosaic virus (WSMV during quarantine inspections of imported wheat, corn, oats, and millet. The LAMP method was developed as a plant quarantine inspection method for the first time, and its simplicity, quickness, specificity and sensitivity were verified compared to current reverse transcription-polymerase chain reaction (RT-PCR and nested PCR quarantine methods. We were able to quickly screen for WSMV at quarantine sites with many test samples; thus, this method is expected to contribute to plant quarantine inspections.
He, Zhen; Jiang, Dongmei; Liu, Aiqin; Sang, Liwei; Li, Wenfeng; Li, Shifang
The complete nucleotide sequence of Cymbidium mosaic virus (CymMV) isolated from vanilla in Hainan province, China was determined for the first time. It comprised 6,224 nucleotides; sequence analysis suggested that the isolate we obtained was a member of the genus Potexvirus, and its sequence shared 86.67-96.61% identities with previously reported sequences. Phylogenetic analysis suggested that CymMV from vanilla fragrans was clustered into subgroup A and the isolates in this subgroup displayed little regional difference.
Andrew John Love
Full Text Available 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 (WT 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.
Verde, C; Malorni, A; Parente, A
The presence of an acetyl blocking group at the N-terminus of the coat protein of papaya mosaic virus has been identified by FAB mass spectrometry. Furthermore, we have found that the N-terminal sequence of the protein is four amino-acid residues (AC-Ser-Lys-Ser-Ser-) longer than that previously reported, while Glu instead of Gln is the C-terminal residue. The present paper shows that PMV-protein is made up of 215 amino acid residues, with a molecular mass of 22,960 Da.
Zhou, Min; Zhou, Kun; Xiang, Neng-Jun; Yang, Liu; Zhang, Cheng-Ming; Wang, Yue-De; Dong, Wei; Lou, Jie; Ji, Bing-Kun; Gao, Xue-Mei; Miao, Ming-Ming; Hu, Qiu-Fen
Two new flavones, siameflavones A and B (1 and 2), together with five known flavones (3-7) were isolated from the stem of Cassia siamea. Their structures were elucidated by spectroscopic methods including extensive 1D and 2D NMR techniques. Compounds 1-5 were evaluated for their anti-tobacco mosaic virus (Anti-TMV) activity. The results showed that compounds 1-5 showed weak anti-TMV activity with inhibition rates in the range of 11.6-18.5%.
Vargas-Asencio, José; Wojciechowska, Klaudia; Baskerville, Maia; Gomez, Annika L; Perry, Keith L; Thompson, Jeremy R
In analyzing grapevine clones infected with grapevine red blotch associated virus, we identified a small number of isometric particles of approximately 30nm in diameter from an enriched fraction of leaf extract. A dominant protein of 25kDa was isolated from this fraction using SDS-PAGE and was identified by mass spectrometry as belonging to grapevine asteroid mosaic associated virus (GAMaV). Using a combination of three methods RNA-Seq, sRNA-Seq, and Sanger sequencing of RT- and RACE-PCR products, we obtained a full-length genome sequence consisting of 6719 nucleotides without the poly(A) tail. The virus possesses all of the typical conserved functional domains concordant with the genus Marafivirus and lies evolutionarily between citrus sudden death associated virus and oat blue dwarf virus. A large shift in RNA-Seq coverage coincided with the predicted location of the subgenomic RNA involved in coat protein (CP) expression. Genus wide sequence alignments confirmed the cleavage motif LxG(G/A) to be dominant between the helicase and RNA dependent RNA polymerase (RdRp), and the RdRp and CP domains. A putative overlapping protein (OP) ORF lacking a canonical translational start codon was identified with a reading frame context more consistent with the putative OPs of tymoviruses and fig fleck associated virus than with those of marafiviruses. BLAST analysis of the predicted GAMaV OP showed a unique relatedness to the OPs of members of the genus Tymovirus.
Geering, A D; Thomas, J E
We have isolated a previously undescribed potexvirus from Alternanthera pungens (Amaranthaceae) in southern Queensland, Australia. This virus was shown to have a moderately wide experimental host range, infecting plants in nine of the twelve families tested. Using specific antibodies, a plate trapped antigen ELISA was developed, allowing detection of virions down to 0.8 microgram/ml of leaf extract. Virions averaged 554 nm long and had a capsid protein with a M(r) of 23.1 x 10(3). A portion of the genome containing the capsid protein ORF and 3' untranslated region was cloned and sequenced. From both serological and amino acid sequence comparisons, the virus was shown to be closely related to papaya mosaic potexvirus (PMV). To determine the taxonomic status of the virus, we assessed variation in the amino acid sequence of capsid proteins of distinct species within the potexvirus genus, as well as variation between strains of the same virus. When the core region of the capsid proteins were compared, distinct species had a maximum of 62.2% sequence identity, whereas strains had a minimum of 88.8% identity. By comparison, the core region of the capsid proteins of the Alternanthera virus and PMV had 79.8% identity. We have concluded that the Alternanthera virus is a different species from PMV, and its relationship with PMV resembles that of potyvirus subgroup members.
Meng, Menghsiao; Lee, Cheng-Cheng
The genus Potexvirus is one of the eight genera belonging to the family Alphaflexiviridae according to the Virus Taxonomy 2015 released by International Committee on Taxonomy of Viruses (www.ictvonline.org/index.asp). Currently, the genus contains 35 known species including many agricultural important viruses, e.g., Potato virus X (PVX). Members of this genus are characterized by flexuous, filamentous virions of 13 nm in diameter and 470–580 nm in length. A potexvirus has a monopartite positive-strand RNA genome, encoding five open-reading frames (ORFs), with a cap structure at the 5′ end and a poly(A) tail at the 3′ end. Besides PVX, Bamboo mosaic virus (BaMV) is another potexvirus that has received intensive attention due to the wealth of knowledge on the molecular biology of the virus. In this review, we discuss the enzymatic activities associated with each of the functional domains of the BaMV replication protein, a 155-kDa polypeptide encoded by ORF1. The unique cap formation mechanism, which may be conserved across the alphavirus superfamily, is particularly addressed. The recently identified interactions between the replication protein and the plant host factors are also described.
Liu, Jiao; Yang, Jun; Bi, Huiping; Zhang, Peng
Cassava mosaic disease, caused by cassava begomoviruses, is the most serious disease for cassava in Africa. However, the pathogenesis of this disease is poorly understood. We employed high throughput digital gene expression profiling based on the Illumina Solexa sequencing technology to investigate the global transcriptional response of cassava to African cassava mosaic virus infection. We found that 3,210 genes were differentially expressed in virus-infected cassava leaves. Gene ontology term and Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that genes implicated in photosynthesis were most affected, consistent with the chlorotic symptoms observed in infected leaves. The upregulation of chlorophyll degradation genes, including the genes encoding chlorophyllase, pheophytinase, and pheophorbide a oxygenase, and downregulation of genes encoding the major apoproteins in light-harvesting complex II were confirmed by qRT-PCR. These findings, together with the reduction of chlorophyll b content and fewer grana stacks in the infected leaf cells, reveal that the degradation of chlorophyll plays an important role in African cassava mosaic virus symptom development. This study will provide a road map for future investigations into viral pathogenesis.
Danesh-Amuz, S; Rakhshandehroo, F; Rezaee, S
Commercial and outdoor fig orchards in four Iranian provinces were surveyed for the incidence of fig mosaic virus (FMV), fig leaf mottle associated virus 2 (FLMaV-2) and fig mild mottle associated virus (FMMaV) from March 2011 to October 2012. A total of 350 asymptomatic and symptomatic fig samples were collected and tested by dot-immunobinding assay (DIBA) for the fig mosaic disease (FMD) using a polyclonal antiserum. According to DIBA results, FMD was present in 73% of the collected symptomatic samples from all visited regions. Samples with positive reactions in DIBA were then analyzed by RT-PCR using with specific primers. PCR results showed that about 14.8% of the FMD-positive samples from three inspected provinces are infected with at least one virus. FMV was the most widely spread virus (14%) followed by FLMaV-2 (1.5%), whereas FMMaV was not found. Phylogenetic analysis of the glycoprotein nucleotide and amino acid sequences of known FMV isolates showed two independent groups with high bootstrap values, with all Iranian isolates distinctly clustered in group I, subgroup IA beside those reported in Turkey. Nucleotide diversity was high within but low between different selected geographic regions and except for Europe, nucleotide distance within geographic regions was low. Statistical analyses indicated a correlation between the genetic structure of the FMV isolates and the geographical origin of isolation. Our analyses suggested that the FMV population is in a state of increase following a bottleneck or founder event in Iran.
Mohammed A. AL-Saleh
Full Text Available In 2011–2012, sixty nine samples were collected from alfalfa plants showing viral infection symptoms in Riyadh region. Mechanical inoculation with sap prepared from two collected samples out of twenty five possitive for Alfalfa mosaic virus (AMV by ELISA were produced systemic mosaic on Vigna unguiculata and Nicotiana tabacum, local lesion on Chenopodium amaranticolor and C. quinoa. Vicia faba indicator plants that induce mosaic and mottle with AMV-Sagir isolate and no infection with AMV-Wadi aldawasser isolate. Approximately 700-bp was formed by RT-PCR using AMV coat protein specific primer. Samples from infected alfalfa gave positive results, while healthy plant gave negative result using dot blot hybridization assay. The nucleotide sequences of the Saudi isolates were compared with corresponding viral nucleotide sequences reported in GenBank. The obtained results showed that the AMV from Australia, Brazil, Puglia and China had the highest similarity with AMV-Sajer isolate. While, the AMV from Spain and New Zealaland had the lowest similarity with AMV-Sajer and Wadi aldawasser isolates. The data obtained in this study has been deposited in the GenBank under the accession numbers KC434083 and KC434084 for AMV-Sajer and AMV- Wadialdawasser respectively. This is the first report regarding the gnetic make up of AMV in Saudi Arabia.
Sivalingam, P N; Satheesh, V; John, P; Chandramohan, S; Malathi, V G
Clerodendron inerme, a common hedge plant grown in India, is affected by a yellow mosaic disease caused by a begomovirus. In the present study, the complete genome (DNA A) of this virus was cloned and sequenced. The total size of DNA A is 2760 nucleotides. The genome of this virus contains six open reading frames and a non-coding intergenic region of 293 nucleotides. Nucleotide sequence comparison analysis revealed maximum sequence identity with Papaya leaf curl virus-Pakistan [Pakistan:Cotton:2002] (73.9%). As this virus had less than 89% identity with other begomoviruses, it was identified as a new begomovirus species and tentatively, named as Clerodendron yellow mosaic virus-[India:New Delhi:2007] (ClYMV-[IN:ND:07]).
Rasoulpour, Rasoul; Afsharifar, Alireza; Izadpanah, Keramat
Melon seedlings showing systemic chlorotic spots and mosaic symptoms were collected in central part of Iran, and a virus was isolated from diseased plants by mechanical inoculation. The virus systemically infected the most inoculated test plants by inducing mosaic symptoms, while, in the members of Fabaceae family and Chenopodium quinoa induced local lesions. Agar gel diffusion test using a polyclonal antiserum against a squash Cucumber mosaic virus (CMV) isolate showed the presence of CMV in the mechanically inoculated plants (designated CMV-Me). The virus was purified by polyethylene glycol precipitation and differential centrifugation. A polyclonal antiserum was produced against the virus that reacted specifically with virus antigen in ELISA and agar gel diffusion tests. The virus was molecularly characterized by PCR amplification of the full length of the coat protein gene using cucumovirus genus specific primer pair CPTALL-3/CPTALL-5 and sequence analysis of the resulting product. No RNA satellite was detected using the primer pair CMVsat3H/sat5T7P. Phylogenetic analysis based on the coat protein amino acid sequences showed that CMV-Me belongs to Subgroup IB. These results may be helpful in melon breeding programs, focusing on plant resistance to plant viruses including CMV.
Sivanandam, Venkatesh; Varady, Erika; Rao, A L N
To investigate the extent of the 3' end repair in a satellite RNA of Cucumber mosaic virus (CMV) strain Q (Q(sat)) by a heterologous Tomato aspermy virus (TAV), a set of biologically active agrotransformants corresponding to the three genomic RNAs of TAV was developed. Analysis of Nicotiana benthamiana plants agroinfiltrated with TAV and either wild type or each of the six 3' deletion mutants of Q(sat) revealed that (i) heterologous replicase failed to generate Q(sat) multimers, a hallmark feature of homologous replicase dependent replication of Qsat; (ii) manifestation of severe symptom phenotypes and progeny analysis suggested that heterologous replicase was competent to repair Q(sat) deletion mutants lacking up to 3'13 nucleotides (nt) but not beyond and (iii) comparative in silico analysis indicated that the 3' secondary structural features of the repaired Q(sat) progeny from heterologous vs homologous driven replicases are remarkably very similar. The significance of these observations is discussed.
Kim, Sang-Min; Lee, Jung-Myung; Yim, Kyu-Ock; Oh, Man-Ho; Park, Jin-Woo; Kim, Kook-Hyung
The nucleotide sequences of the genomic RNAs of Cucumber green mottle mosaic virus Korean watermelon isolate (CGMMV-KW) and Korean oriental melon isolate (CGMMV-KOM) were determined and compared to the sequences of other tobamoviruses including CGMMV strains W and SH. Each CGMMV isolate had a genome of 6,424 nucleotides. Each also had 60 and 176 nucleotides of 5' and 3' untranslated regions (UTRs), respectively, and four open reading frames (ORF1-4). ORFs 1 to 4 encode proteins of 129, 186, 29, and 17.4 kDa, respectively. The nucleotide and deduced amino acid sequences of CGMMV-KOM and CGMMV-KW were more than 98.3% identical. When compared to other CGMMV strains in a phylogenetic analysis they were found to form a distinct virus clade, and were more distantly related to other tobamoviruses (23.5-56.7% identity).
Li, Yongqiang; Deng, Congliang; Shang, Qiaoxia; Zhao, Xiaoli; Liu, Xingliang; Zhou, Qi
Virus-derived small interfering RNAs (vsiRNAs) of cucumber green mottle mosaic virus (CGMMV), a member of the genus Tobamovirus, were characterised in cucumber plants by deep sequencing. CGMMV vsiRNAs of 21-22 nt in length predominated, suggesting that there might be a conserved mechanism of DCL2 and DCL4 involvement in the biogenesis of vsiRNAs, as well as a common RNA silencing pathway in CGMMV-infected cucumber plants. The 5'-terminal base of vsiRNAs was biased towards C/A/U, suggesting that CGMMV vsiRNAs might be loaded into diverse AGO-containing RISCs to disturb the gene expression of host plants. Possible targets for some of the vsiRNAs were also predicted.
Kil Hyun Kim; Seungmo Lim; Yang Jae Kang; Min Young Yoon; Moon Nam; Tae Hwan Jun; Min-Jung Seo; Seong-Bum Baek; Jeom-Ho Lee; Jung-Kyung Moon; Suk-Ha Lee; Su-Heon Lee; Hyoun-Sub Lim; Jae Sun Moon; Chang-Hwan Park
Virus-induced gene silencing (VIGS) is an effective tool for the study of soybean gene function. Successful VIGS depends on the interaction between virus spread and plant growth, which can be influenced by environmental conditions. Recently, we developed a new VIGS system derived from the Soybean yellow common mosaic virus (SYCMV). Here, we investigated several environmental and developmental factors to improve the efficiency of a SYCMV-based VIGS system to optimize the functional analysis of...
GUO Xing-qi; ZHU Xiao-ping; ZHANG Jie-dao; GUO Yan-kui
Ultrastructural alterations in foliar cells were studied in leaves of resistant maize varietyLuyu16 and susceptible maize inbred line Luyuan92 infected by maize dwarf mosaic virus Shandong isolate(MDMV-SD), respectively. The results showed that marked cytopathological alterations were observed both inresistant plants and in susceptible plants, compared with that in healthy plants. However, some ultrastructur-al alterations, which observed in resistant plants, were different from those in susceptible plants. In resistantplants, which infected with the virus, the main organelles, including chloroplasts and mitochondria, wereslightly destroyed, the amount of mitochondria and peroxisome were increased. A few or no plasmodesmatawere observed. There were three kinds of inclusions including pinwheel, bundle and laminated aggregate, andthe virus particles in the cytoplasm. In susceptible plants, which infected with the virus, the chloroplasts wereheavily disrupted, including thylakoid swelling and envelope broking. The virus particles were more than thosein the resistant variety. Four kinds of inclusions including pinwheel, bundle, laminated aggregate and highelecton-dense body appeared in cytoplasm. Plasmodesmata and plasma membrane were abundant, and therewere frequent invaginations of the plasma membrane that led to the formation of vesicles and myelin-likestructures.
Ang Rinzing Sherpa; Vipin Hallan; Promila Pathak; Aijaz Asghar Zaidi
The complete nucleotide sequence of an Indian strain of Cymbidium mosaic virus (CymMV) was determined and compared with other potexviruses. Phylogenetic analyses on the basis of RNA-dependent RNA polymerase (RdRp), triple gene block protein and coat protein (CP) amino acid sequences revealed that CymMV is closely related to the Narcissus mosaic virus (NMV), Scallion virus X (SVX), Pepino mosaic virus (PepMV) and Potato aucuba mosaic virus (PAMV). Different sets of primers were used for the amplification of different regions of the genome through RT-PCR and the amplified genes were cloned in a suitable vector. The full genome of the Indian isolate of CymMV from Phaius tankervilliae shares 96–97% similarity with isolates reported from other countries. It was found that the CP gene of CymMV shares a high similarity with each other and other potexviruses. One of the Indian isolates seems to be a recombinant formed by the intermolecular recombination of two other CymMV isolates. The phylogenetic analyses, Recombination Detection Program (RDP2) analyses and sequence alignment survey provided evidence for the occurrence of a recombination between an Indian isolate (AM055720) as the major parent, and a Korean type-2 isolate (AF016914) as the minor parent. Recombination was also observed between a Singapore isolate (U62963) as the major parent, and a Taiwan CymMV (AY571289) as the minor parent.
Robinson, Megan D; Murray, Timothy D
Wheat streak mosaic virus (WSMV), the cause of wheat streak mosaic, is a widespread and damaging pathogen of wheat. WSMV is not a chronic problem of annual wheat in the United States Pacific Northwest but could negatively affect the establishment of perennial wheat, which is being developed as an alternative to annual wheat to prevent soil erosion. Fifty local isolates of WSMV were collected from 2008 to 2010 near Lewiston, ID, Pullman, WA, and the United States Department of Agriculture Central Ferry Research Station, near Pomeroy, WA to determine the amount of genetic variation present in the region. The coat protein gene from each isolate was sequenced and the data subjected to four different methods of phylogenetic analyses. Two well-supported clades of WSMV were identified. Isolates in clade I share sequence similarity with isolates from Central Europe; this is the first report of isolates from Central Europe being reported in the United States. Isolates in clade II are similar to isolates originating from Australia, Argentina, and the American Pacific Northwest. Nine isolates showed evidence of recombination and the same two well-supported clades were observed when recombinant isolates were omitted from the analysis. More polymorphic sites, parsimony informative sites, and increased diversity were observed in clade II than clade I, suggesting more recent establishment of the virus in the latter. The observed diversity within both clades could make breeding for durable disease resistance in perennial wheat difficult if there is a differential response of WSMV resistance genes to isolates from different clades.
Kumar, S; Gautam, K K; Raj, S K
Cucumber mosaic virus (CMV) is an important virus since it causes severe losses to many economically important crops worldwide. Five new isolates of CMV were isolated from naturally infected Hippeastrum hybridum, Dahlia pinnata, Hemerocallis fulva, Acorus calamus and Typhonium trilobatum plants, all exhibiting severe leaf mosaic symptoms. For molecular identification and sequence analyses, the complete coat protein (CP) gene of these isolates was amplified by RT-PCR. The resulting amplicons were cloned and sequenced and isolates were designated as HH (KP698590), DP (JF682239), HF (KP698589), AC (KP698588) and TT (JX570732). For study of genetic diversity among these isolates, the sequence data were analysed by BLASTn, multiple alignment and generating phylogenetic trees along with the respective sequences of other CMV isolates available in GenBank Database were done. The isolates under study showed 82-99% sequence diversity among them at nucleotide and amino acid levels; however they showed close relationships with CMV isolates of subgroup IB. In alignment analysis of amino acid sequences of HH and AC isolates, we have found fifteen and twelve unique substitutions, compared to HF, DP and TT isolates, suggesting the cause of high genetic diversity.
Qiansheng LIAO; Liping ZHU; Zhiyou DU; Rong ZENG; Junli FENG; Jishuang CHEN
Satellite RNAs (satRNAs) are molecular parasites that interfere with the pathogenesis of the helper viruses.In this study,the relative accumulation of cucumber mosaic virus (CMV)-Fny genomic RNAs with or without satRNAs were quantitatively analyzed by real-time RT-PCR.The results showed that satRs apparently attenuated the symptoms of CMV-Fny on Nicotiana tabacum by depressing the accumulation of CMV-Fny genomic RNAs,tested as open reading frames.The accumulation of CMV-Fny la,2a,2b,3a,and CP genes was much higher than that of CMV-Fny with satRs added(CMV-Fsat),at different inoculation times.CMV-Fny△2b,in which the complete 2b gene and 41 amino acids at the C-terminal of the 2a gene were deleted,caused only a slight mosaic effect on N.tabacum seedlings,similar to that of CMVFsat,but the addition of satRs to CMV-Fny△2b showed further decrease in the accumulation of CMVFny△2b genomic RNAs.Our results indicated that the attenuation of CMV,by adding satRs or deleting the 2b gene,was due to the low accumulation of CMV genomic RNAs,and that satRNA-mediated reduction of CMV genomic RNAs accumulation in N.tabacum was possibly related to the 2b gene.
Ibrahim, Amr [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States); Present address: Genomics Facility, Agricultural Genetic Engineering Research Institute, Agricultural Research Center, Giza 12619 (Egypt); Hutchens, Heather M. [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States); Howard Berg, R. [Integrated Microscopy Facility, Donald Danforth Plant Science Center, Saint Louis, MO 63132 (United States); Sue Loesch-Fries, L., E-mail: email@example.com [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States)
To identify the virus components important for assembly of the Alfalfa mosaic virus replicase complex, we used live cell imaging of Arabidopsis thaliana protoplasts that expressed various virus cDNAs encoding native and GFP-fusion proteins of P1 and P2 replicase proteins and full-length virus RNAs. Expression of P1-GFP alone resulted in fluorescent vesicle-like bodies in the cytoplasm that colocalized with FM4-64, an endocytic marker, and RFP-AtVSR2, RabF2a/Rha1-mCherry, and RabF2b/Ara7-mCherry, all of which localize to multivesicular bodies (MVBs), which are also called prevacuolar compartments, that mediate traffic to the lytic vacuole. GFP-P2 was driven from the cytosol to MVBs when expressed with P1 indicating that P1 recruited GFP-P2. P1-GFP localized on the tonoplast, which surrounds the vacuole, in the presence of infectious virus RNA, replication competent RNA2, or P2 and replication competent RNA1 or RNA3. This suggests that a functional replication complex containing P1, P2, and a full-length AMV RNA assembles on MVBs to traffic to the tonoplast.
Romay, G; Lecoq, H; Desbiez, C
In recent years, three new potyviruses have been described in the papaya ringspot virus (PRSV) cluster. In addition, two types of PRSV are recognized, type W, infecting cucurbit plants, and type P, infecting papaya and also cucurbits. A third type, PRSV-T, was also partially described in Guadeloupe. Complete genome sequencing of four PRSV-T isolates showed that this virus is a related virus that is distinct from PRSV, and the name zucchini tigré mosaic virus (ZTMV) is proposed, in reference to the typical symptoms observed in zucchini squash. Eleven other viral isolates from different geographic origins were confirmed as ZTMV isolates using the complete sequence of the cylindrical inclusion (CI) coding region, whereas pairwise sequence similarities in the coat protein (CP) coding region did not unambiguously distinguish ZTMV isolates from PRSV isolates. The use of the CI coding region for species demarcation appears more suitable than the CP coding region for closely related viruses. Principal coordinates analysis based on the biological behavior of the viral isolates studied clustered PRSV-P, PRSV-W and ZTMV isolates into three different groups. Therefore, ZTMV is different from PRSV in its molecular and biological properties.
Yao, Wei; Ruan, Miaohong; Qin, Lifang; Yang, Chuanyu; Chen, Rukai; Chen, Baoshan; Zhang, Muqing
Sugarcane mosaic disease is mainly caused by the sugarcane mosaic virus (SCMV), which can significantly reduce stalk yield and sucrose content of sugarcane in the field. Coat protein mediated protection (CPMP) is an effective strategy to improve virus resistance. A 2-year field study was conducted to compare five independent transgenic sugarcane lines carrying the SCMV-CP gene (i.e., B2, B36, B38, B48, and B51) with the wild-type parental clone Badila (WT). Agronomic performance, resistance to SCMV infection, and transgene stability were evaluated and compared with the wild-type parental clone Badila (WT) at four experimental locations in China across two successive seasons, i.e., plant cane (PC) and 1st ratoon cane (1R). All transgenic lines derived from Badila had significantly greater tons of cane per hectare (TCH) and tons of sucrose per hectare (TSH) as well as lower SCMV disease incidence than those from Badila in the PC and 1R crops. The transgenic line B48 was highly resistant to SCMV with less than 3% incidence of infection. The recovery phenotype of transgenic line B36 was infected soon after virus inoculation, but the subsequent leaves showed no symptoms of infection. Most control plants developed symptoms that persisted and spread throughout the plant with more than 50% incidence. B48 recorded an average of 102.72 t/ha, which was 67.2% more than that for Badila. The expression of the transgene was stable over many generations with vegetative propagation. These results show that SCMV-resistant transgenic lines derived from Badila can provide resistant germplasm for sugarcane breeding and can also be used to study virus resistance mechanisms. This is the first report on the development and field performance of transgenic sugarcane plants that are resistant to SCMV infection in China. PMID:28228765
Gislanne Brito Barros
Full Text Available Dentre os vírus que infectam o feijão-caupi (Vigna unguiculata L. Walp. destacam-se, respectivamente, pela severidade e ampla ocorrência o Cowpea severe mosaic virus (CPSMV e o Cowpea aphid-borne mosaic virus (CABMV. Portanto, objetivaram-se, no presente trabalho, obter e avaliar plantas de feijão-caupi com resistência ao CPSMV e ao CABMV, visando ao desenvolvimento de cultivares essencialmente derivadas e novas cultivares. Realizaram-se oito cruzamentos seguidos de retrocruzamentos, utilizando a linhagem TE 97-309G-9 e a cultivar Patativa como genitores resistentes, e as cultivares BR3-Tracuateua, BRS-Urubuquara, BRS-Novaera, BRS-Guariba e Pretinho como genitores suscetíveis. As gerações F2 e F2RC1 foram desafiadas quanto à resistência por meio de inoculação mecânica com isolados do CPSMV e do CABMV. Nas gerações F2RC1, além da resistência foram avaliados os caracteres: número de dias para o início da floração, comprimento das vagens, número de grãos. vagem-1, peso de cem grãos e produção de grãos.planta-1. Todos os indivíduos F2 e F2RC1 foram analisados pelo teste χ² e se ajustaram à frequência esperada de 15 plantas suscetíveis 1 planta resistente a ambos os vírus. As médias das plantas F2RC1 resistentes, de cada retrocruzamento, foram comparadas com a média do seu respectivo genitor recorrente pelo teste 't' e as médias dos retrocruzamentos foram comparadas pelo teste de Scott-Knott. Foi detectada variabilidade genética entre os retrocruzamentos para todos os caracteres. Todos os retrocruzamentos foram considerados promissores para produção de cultivares essencialmente derivadas resistentes ao CPSMV e ao CABMV e as plantas selecionadas possuem características que possibilitam a seleção de linhagens com grãos de bom padrão comercial e altamente produtivas.
Davies Kevin M
Full Text Available Abstract Background Daffodils (Narcissus pseudonarcissus are one of the world's most popular ornamentals. They also provide a scientific model for studying the carotenoid pigments responsible for their yellow and orange flower colours. In reverse bicolour daffodils, the yellow flower trumpet fades to white with age. The flowers of this type of daffodil are particularly prone to colour break whereby, upon opening, the yellow colour of the perianth is observed to be 'broken' into patches of white. This colour break symptom is characteristic of potyviral infections in other ornamentals such as tulips whose colour break is due to alterations in the presence of anthocyanins. However, reverse bicolour flowers displaying colour break show no other virus-like symptoms such as leaf mottling or plant stunting, leading some to argue that the carotenoid-based colour breaking in reverse bicolour flowers may not be caused by virus infection. Results Although potyviruses have been reported to cause colour break in other flower species, enzyme-linked-immunoassays with an antibody specific to the potyviral family showed that potyviruses were not responsible for the occurrence of colour break in reverse bicolour daffodils. Colour break in this type of daffodil was clearly associated with the presence of large quantities of rod-shaped viral particles of lengths 502-580 nm in tepals. Sap from flowers displaying colour break caused red necrotic lesions on Gomphrena globosa, suggesting the presence of potexvirus. Red necrotic lesions were not observed in this indicator plant when sap from reverse bicolour flowers not showing colour break was used. The reverse transcriptase polymerase reactions using degenerate primers to carla-, potex- and poty-viruses linked viral RNA with colour break and sequencing of the amplified products indicated that the potexvirus Narcissisus mosaic virus was the predominant virus associated with the occurrence of the colour break
Multifunctional viral coat proteins (CPs) play important roles in the virus life-cycle. The CP determinants and mechanisms involved in extension of host range of monocot-infecting viruses are poorly understood. The role of the C-terminal region of Wheat streak mosaic virus (WSMV) CP in virus transpo...
［1］Palukaitis, P., Roossinck, M. J., Dietzgen, R. G. et al., Cucumber mosaic virus, Adv. Virus Res., 1992, 41: 281-348.［2］Hayes, R. J., Buck, K. W., Complete replication of an eukaryotic virus RNA in vitro by a purified RNA-dependent RNA polymerase, Cell, 1990, 63: 363-369.［3］Nitta, N., Takanami, Y., Kuwata, S. et al., Inoculation with RNAs 1 and 2 of cucumber mosaic virus induces viral RNA replicase activity in tobacco mesophyll protoplasts, J. Gen. Virol., 1988, 69: 2695-2700.［4］Suzuki, M., Kuwata, S., Kataoda, J. et al., Functional analysis of deletion mutants of cucumber mosaic virus RNA3 using an in vitro transcription system, Virology, 1991, 183: 106-113.［5］Canto, T., Prior, D. A. M., Hellwald, K. H. et al., Characterization of cucumber mosaic virus (IV)--Movement protein and coat protein are both essential for cell-to-cell movement of cucumber mosaic virus, Virology, 1997, 237:237-248.［6］Takanami, Y., A striking change in symptoms on cucumber mosaic virus-infected tobacco plants induced by a satellite RNA, Virology, 1981, 109: 120-126.［7］DeBorde, D. C., Naeve, C. W., Herlocher, M. L. et al., Resolution of a common RNA sequencing ambiguity by terminal deoxynucleotidyl transferase, Anal. Biochem., 1986, 157: 275-282.［8］Haseloff, J., Siemering, K. R., Prasher, D. C. et al., Removal of a cryptic intron and subcellular localization of green fluo-rescent protein are required to mark transgenic Arabidopsis plants brightly, Proc. Natl. Acad. Sci. USA, 1997, 94: 2122-2127.［9］Shi, B. J., Ding, S. W., Symons, R. H., Plasmid vector for cloning infectious cDNAs from plant RNA viruses: high infec-tivity of cDNA clones of tomato aspermy cucumovirus, J. Gen. Virol., 1997, 78: 1181-1185.［10］Rizzo, T. M., Palukaitis, P., Construction of full-length cDNA clones of cucumber mosaic virus RNAs 1, 2 and 3: Genera-tion of infectious RNA transcripts, Mol. Gen. Genet., 1990, 222: 249-256.［11］Hall, R. D., The initiation and maintenance of
A Srivastava; S K Raj
Coat protein (CP)-mediated resistance against an Indian isolate of the Cucumber mosaic virus (CMV) subgroup IB was demonstrated in transgenic lines of Nicotiana benthamiana through Agrobacterium tumefaciens-mediated transformation. Out of the fourteen independently transformed lines developed, two lines were tested for resistance against CMV by challenge inoculations. The transgenic lines exhibiting complete resistance remained symptomless throughout life and showed reduced or no virus accumulation in their systemic leaves after virus challenge. These lines also showed virus resistance against two closely related strains of CMV. This is the first report of CP-mediated transgenic resistance against a CMV subgroup IB member isolated from India.
Pinto, Zayame Vegette [UNESP; Rezende,Jorge Alberto Marques; Yuki, Valdir Atsushi; Piedade,Sônia Maria de Stefano
The main objective of this work was to investigate the ability of Aphis gossypii and Myzus persicae to transmit Cucumber mosaic virus (CMV) singly and mixed with two potyviruses (Papaya ringspot virus - type W, PRSV-W and Zucchini yellow mosaic virus, ZYMV), to zucchini squash plants (Cucurbita pepo). The results showed that the potyviruses in general were more efficiently transmitted by both species of aphids as compared to CMV. The transmission of PRSV-W, ZYMV and CMV separately was more ef...
Ming-Min ZHAO; De-Rong AN; Guang-Hua HUANG; Zu-Hua HE; Jiang-Ye CHEN
Plant viruses encode suppressors of post-transcriptional gene silencing (PTGS), an adaptive defense response that limits virus replication and its spread in plants. The helper component proteinase (HCPro) of the potato virus A (PVA, genus Potyvirus) suppresses PTGS of silenced transgenes. Here, the effect of HC-Pro on siRNA-directed interference in the tobacco mosaic virus (TMV) was examined by using a transient Agrobacterium tumefaciens-based delivery system in intact tissues. It was shown that the interference effect was completely blocked by co-infiltration with HC-Pro plus siRNA constructs in both systemic and hypersensitive hosts. In the system host, all plants agro-infiltrated with HC-Pro plus siRNA constructs displayed the same symptoms as the negative control. Meanwhile, TMV RNA accumulation was found to be abundant in the upper leaves using reverse transcriptase-PCR (RT-PCR) and Northern blot assays. On the contrary, plants agro-infiltrated with the siRNA construct alone were free of symptoms. Therefore, our study suggests that the transient expression of HC-Pro inhibited the siRNA-directed host defenses against TMV infection.
Full Text Available Fig mosaic virus (FMV and Fig badnavirus-1 (FBV-1 are two of the most important fig infecting viruses. The incidence and distribution of FBV-1 and FMV were determined by testing in PCR 138 asymptomatic and symptomatic samples. These samples were collected from 60 fig gardens and agricultural fields in three provinces of Iran. The fig infecting viruses FBV-1 and FMV, respectively, were detected in 92 (66.6% and 34 (24.6% samples collected from all the surveyed fields. Overall, 24 out of 138 (17.3% samples showed mixed infections. The sequence analysis of a genomic fragment of 922 nt, comprising the entire ORF-2 and part of the 5’ termini of the ORF-3 of 10 selected FBV-1 Iranian isolates from different provinces, and of the type member from GenBank (Acc. No: JF411989, showed a variation ranging from 1 to 3% at nucleotide level and 1% at the amino acid level. The phylogenetic analysis grouped the FBV-1 isolates into two groups, with the Iranian isolates clustered in two distinct subgroups of group I, according to their geographical origin. In our research, the prevalence and sequence analysis of FBV-1 as the only identified DNA virus infecting fig trees, was studied for the first time in Iran.
Creager, Angela N H; Morgan, Gregory J
Rosalind Franklin is best known for her informative X-ray diffraction patterns of DNA that provided vital clues for James Watson and Francis Crick's double-stranded helical model. Her scientific career did not end when she left the DNA work at King's College, however. In 1953 Franklin moved to J. D. Bernal's crystallography laboratory at Birkbeck College, where she shifted her focus to the three-dimensional structure of viruses, obtaining diffraction patterns of Tobacco mosaic virus (TMV) of unprecedented detail and clarity. During the next five years, while making significant headway on the structural determination of TMV, Franklin maintained an active correspondence with both Watson and Crick, who were also studying aspects of virus structure. Developments in TMV research during the 1950s illustrate the connections in the emerging field of molecular biology between structural studies of nucleic acids and of proteins and viruses. They also reveal how the protagonists of the "race for the double helix" continued to interact personally and professionally during the years when Watson and Crick's model for the double-helical structure of DNA was debated and confirmed.
Yakoubi, S; Desbiez, C; Fakhfakh, H; Wipf-Scheibel, C; Marrakchi, M; Lecoq, H
During a survey conducted in October 2005, cucurbit leaf samples showing virus-like symptoms were collected from the major cucurbit-growing areas in Tunisia. DAS-ELISA showed the presence of Moroccan watermelon mosaic virus (MWMV, Potyvirus), detected for the first time in Tunisia, in samples from the region of Cap Bon (Northern Tunisia). MWMV isolate TN05-76 (MWMV-Tn) was characterized biologically and its full-length genome sequence was established. MWMV-Tn was found to have biological properties similar to those reported for the MWMV type strain from Morocco. Phylogenetic analysis including the comparison of complete amino-acid sequences of 42 potyviruses confirmed that MWMV-Tn is related (65% amino-acid sequence identity) to Papaya ringspot virus (PRSV) isolates but is a member of a distinct virus species. Sequence analysis on parts of the CP gene of MWMV isolates from different geographical origins revealed some geographic structure of MWMV variability, with three different clusters: one cluster including isolates from the Mediterranean region, a second including isolates from western and central Africa, and a third one including isolates from the southern part of Africa. A significant correlation was observed between geographic and genetic distances between isolates. Isolates from countries in the Mediterranean region where MWMV has recently emerged (France, Spain, Portugal) have highly conserved sequences, suggesting that they may have a common and recent origin. MWMV from Sudan, a highly divergent variant, may be considered an evolutionary intermediate between MWMV and PRSV.
The experiments described in this thesis were designed to unravel various aspects of the mechanism of gene expression of cowpea mosaic virus (CPMV). For this purpose full-length DNA copies of both genomic RNAs of CPMV were constructed. Using powerful invitro transcription systems RNA t
Next generation sequencing (NGS) technology was used to analyze the occurrence of viruses in Sorghum almum plants in Florida exhibiting mosaic symptoms. Total RNA was extracted from symptomatic leaves and used as a template for cDNA library preparation. The resulting library was sequenced on an Illu...
Pyramiding of alien-derived Wheat streak mosaic virus (WSMV) resistance and resistance enhancing genes in wheat is a costeffective and environmentally safe strategy for disease control. PCR-based markers and cytogenetic analysis with genomic in situ hybridisation were applied to identify alien chrom...
Wheat streak mosaic virus (WSMV) can cause significant yield loss in wheat (Triticum aestivum L.) in the Great Plains of North America. A recently identified WSMV resistance gene, Wsm2, was mapped to chromosome 3BS in germplasm line ‘CO960293–2’. Effective genetic markers tightly linked to the gene ...
Hanssen, I.M.; Gutiérrez-Aguirre, I.; Paeleman, A.; Goen, K.; Wittemans, L.; Lievens, B.; Vanachter, A.C.R.C.; Ravnikar, M.; Thomma, B.P.H.J.
The potential of three mild Pepino mosaic virus (PepMV) isolates, belonging to the CH2, EU and LP genotypes, to protect a tomato (Solanum lycopersicum) crop against an aggressive challenge isolate of the CH2 genotype was assessed in greenhouse trials and PepMV symptoms were rated at regular time poi
Lee, Joong-Hwan; Son, Chang-Gi; Kwon, Joong-Bae; Nam, Hyo-Hun; Kim, Yeongtae; Lee, Su-Heon; Zhao, Fumei; Moon, Jae Sun
The complete genome sequence of Chinese yam necrotic mosaic virus (ChYNMV) consisting of 8,213 nucleotides containing one open reading frame was determined by the transcriptome data generated from Discorea opposita This is the first report of the complete nucleotide sequence of ChYNMV from Dioscorea opposita in the Republic of Korea.
Pinheiro, Patrícia V; Quintela, Eliane D; Ana Maria R. Junqueira; Aragão, Francisco JL; Faria, Josias C
Genetically modified (GM) crops is considered the fastest adopted crop technology in the history of modern agriculture. However, possible undesirable and unintended effects must be considered during the research steps toward development of a commercial product. In this report we evaluated effects of a common bean virus resistant line on arthropod populations, considered as non-target organisms. This GM bean line (named M1/4) was modified for resistance against Bean golden mosaic virus (BGMV) ...
Full Text Available From Anemone coronaria cucumber mosaic virus (Cucumis virus 1 Smith was isolated. It caused a general chlorosis, reduction of leaves blades and of the whole plants. 66 species of test plants were inoculated with the sap from infected cucumber plants. 33 of these were infected systemically and 11 only locally. Among 22 noninfected plants was Anemone coronaria which indicated as it was experimentally proved, that this species is infected only by the aphids – Myzus persicae Sulz.
de Wispelaere, Melissanne; Rao, A L N
Cucumber Mosaic Virus (CMV) is a plant infecting tripartite positive-strand RNA virus. In addition to three genomic and two known subgenomic RNAs, CMV strains of subgroup II (e.g. Q-CMV), but not subgroup I (e.g. Fny-CMV), produce and package a redundant RNA5 encompassing the 3' 304-307 nucleotides of RNAs 2 and 3. The mechanism regulating RNA5 production and its role in CMV life cycle is unknown. In this study, transient expression of Q2 or Q3 by agroinfiltration into Nicotiana benthamiana plants resulted in efficient accumulation of RNA5 suggesting that its production is independent of CMV replication. Deletion and point mutations engineered into a highly conserved region (Box1) adjacent to the 5' end of RNA5 identified sequences required for its efficient production. An experimental system, involving a chimera of Q3 (Q3B3) characterized by having a 3' tRNA-like structure (3'TLS) from Brome mosaic virus (BMV) and RNA5 defective variants of Q1 (Q1Delta), Q2 (Q2Delta) and Q3B3 (Q3DeltaB3), was used to evaluate in vivo the contribution of RNA5 in promoting RNA recombination. Generation of precise homologous recombinants was strictly dependent on sequence identity. When both parental RNAs carried the Box1, recombination occurred preferentially within the Box1. In contrast, generation of non-homologous recombinants occurred only when Q1 and Q2 were competent to produce RNA5. A mechanistic model explaining the functional role played by the RNA5 in generating CMV recombinants was presented.
Kumari, Reenu; Kumar, Surender; Singh, Lakhmir; Hallan, Vipin
Plant viral movement proteins facilitate virion movement mainly through interaction with a number of factors from the host. We report the association of a cell wall localized ascorbate oxidase (CsAO4) from Cucumis sativus with the movement protein (MP) of Cucumber mosaic virus (CMV). This was identified first in a yeast two-hybrid screen and validated by in vivo pull down and bimolecular fluorescence complementation (BiFC) assays. The BiFC assay showed localization of the bimolecular complexes of these proteins around the cell wall periphery as punctate spots. The expression of CsAO4 was induced during the initial infection period (up to 72 h) in CMV infected Nicotiana benthamiana plants. To functionally validate its role in viral spread, we analyzed the virus accumulation in CsAO4 overexpressing Arabidopsis thaliana and transiently silenced N. benthamiana plants (through a Tobacco rattle virus vector). Overexpression had no evident effect on virus accumulation in upper non-inoculated leaves of transgenic lines in comparison to WT plants at 7 days post inoculation (dpi). However, knockdown resulted in reduced CMV accumulation in systemic (non-inoculated) leaves of NbΔAO-pTRV2 silenced plants as compared to TRV inoculated control plants at 5 dpi (up to 1.3 fold difference). In addition, functional validation supported the importance of AO in plant development. These findings suggest that AO and viral MP interaction helps in early viral movement; however, it had no major effect on viral accumulation after 7 dpi. This study suggests that initial induction of expression of AO on virus infection and its association with viral MP helps both towards targeting of the MP to the apoplast and disrupting formation of functional AO dimers for spread of virus to nearby cells, reducing the redox defense of the plant during initial stages of infection.
Full Text Available Homologous recombination is a dominant force in evolution and results in genetic mosaics. To detect evidence of recombination events and assess the biological significance of genetic mosaics, genome sequences for various viral populations of reasonably large size are now available in the GenBank. We studied a multi-functional viral gene, the adeno-associated virus (AAV cap gene, which codes for three capsid proteins, VP1, VP2 and VP3. VP1-3 share a common C-terminal domain corresponding to VP3, which forms the viral core structure, while the VP1 unique N-terminal part contains an enzymatic domain with phospholipase A2 activity. Our recombinant detection program (RecI revealed five novel recombination events, four of which have their cross-over points in the N-terminal, VP1 and VP2 unique region. Comparison of phylogenetic trees for different cap gene regions confirmed discordant phylogenies for the recombinant sequences. Furthermore, differences in the phylogenetic tree structures for the VP1 unique (VP1u region and the rest of cap highlighted the mosaic nature of cap gene in the AAV population: two dominant forms of VP1u sequences were identified and these forms are linked to diverse sequences in the rest of cap gene. This observation together with the finding of frequent recombination in the VP1 and 2 unique regions suggests that this region is a recombination hot spot. Recombination events in this region preserve protein blocks of distinctive functions and contribute to convergence in VP1u and divergence of the rest of cap. Additionally the possible biological significance of two dominant VP1u forms is inferred.
José Evando Aguiar Beserra Júnior
Full Text Available A análise SDS-PAGE do Cowpea severe mosaic virus (CPSMV purificado revelou a migração de três frações protéicas estimadas em 43, 23 e 21 kDa, correspondentes às proteínas do capsídeo: denominadas proteína maior (43 kDa e menor (23 kDa; intacta e 21 kDa; clivada. As proteínas do capsídeo, na sua forma nativa, foram utilizadas na imunização de camundongos pelas vias oral e nasal, durante 10 dias consecutivos. As frações protéicas de 43 e 23 kDa, em sua forma desnaturada, foram utilizadas para imunização subcutânea. A resposta imunológica da mucosa foi avaliada pela proliferação celular das placas de Peyer de camundongos imunizados pela via oral com o CPSMV purificado. Ficou demonstrado que o CPSMV induz resposta imunológica, evidenciada pela síntese de anticorpos séricos, quando administrado na sua forma nativa pelas vias oral e nasal ou através de suas proteínas do capsídeo desnaturadas, pela via subcutânea. Não foi necessário o uso de adjuvantes, quer por via oral quer por via nasal. As frações protéicas de 43 e 23 kDa mostraram-se responsáveis pela imunogenicidade do vírus, como foi evidenciado pela síntese de anticorpos específicos detectados por ELISA. A análise da proliferação celular da placas de Peyer revelou um aumento (r=0,88 do número de leucócitos ao longo de 42 dias após a imunização. Esses resultados reforçam a possibilidade do uso do CPSMV como vetor seguro de antígenos de doenças humanas/animais pouco imunogênicos para produção de vacinas.SDS-PAGE analysis of purified Cowpea severe mosaic virus (CPSMV revealed the migration of three protein fractions of 43, 23 and 21 kDa, corresponding to the capsid protein called large protein (43 kDa and small protein (23 kDa; intact and 21 kDa; cleaved. The capsid proteins, in their native form, were used to immunize mice through oral and nasal routes for ten consecutive days. The denatured form of the 43 and 23 kDa protein fractions were
Zhou, Wen-Wen; Zhang, Li-Xiang; Zhang, Bin; Wang, Fei; Liang, Zhi-Hong; Niu, Tian-Gui
A large number of bacteria were isolated from plant samples and screened for antiviral activity against the Tobacco mosaic virus (TMV). The bacterium ZH14, which was isolated from Chinese Anxi oolong tea, secreted the antiviral substances, having 94.2% virus inhibition when the bacterial culture filtrate and TMV extract were mixed at a ratio of 1:1. The ZH14 strain is a gram-positive, spore-forming rod and has the ability to degrade ribonucleic acid. Based on its effectiveness on virus inhibition, ZH14 was selected for characterization and was identified as a strain of the Bacillus cereus group based on phenotypic tests and comparative analysis of its 16S rDNA sequence. At the same time, we determined the antiviral product of ZH14 as an extracellular protein with high molecular mass, having an optimum temperature of 15-60 degrees C and an optimum pH of 6-10. Hence, the ZH14 strain and its culture filtrate have potential application in controlling plant diseases caused by TMV.
Rioux, Gervais; Majeau, Nathalie; Leclerc, Denis
In general, the structure of the papaya mosaic virus (PapMV) and other members of the potexviruses is poorly understood. Production of PapMV coat proteins in a bacterial expression system and their self-assembly in vitro into nanoparticles is a very useful tool to study the structure of this virus. Using recombinant PapMV nanoparticles that are similar in shape and appearance to the plant virus, we evaluated surface-exposed regions by two different methods, immunoblot assay and chemical modification with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide or diethyl-pyrocarbonate followed by mass spectrometry. Three regions were targeted by the two techniques. The N- and C-termini were shown to be surfaced exposed as expected. However, the region 125-136 was revealed for the first time as the major surface-exposed region of the nanoparticles. The presence of linear peptides at the surface was finally confirmed using antibodies directed to those peptides. It is likely that region 125-136 plays a key role in the lifecycle of PapMV and other members of the potexvirus group.
Shen, Wentao; Tuo, Decai; Yan, Pu; Li, Xiaoying; Zhou, Peng
Papaya leaf distortion mosaic virus (PLDMV) can infect transgenic papaya resistant to a related pathogen, Papaya ringspot virus (PRSV), posing a substantial threat to papaya production in China. Current detection methods, however, are unable to be used for rapid detection in the field. Here, a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of PLDMV, using a set of four RT-LAMP primers designed based on the conserved sequence of PLDMV CP. The RT-LAMP method detected specifically PLDMV and was highly sensitive, with a detection limit of 1.32×10(-6) μg of total RNA per reaction. Indeed, the reaction was 10 times more sensitive than one-step RT-PCR, while also requiring significantly less time and equipment. The effectiveness of RT-LAMP and one-step RT-PCR in detecting the virus were compared using 90 field samples of non-transgenic papaya and 90 field samples of commercialized PRSV-resistant transgenic papaya from Hainan Island. None of the non-transgenic papaya tested positive for PLDMV using either method. In contrast, 19 of the commercialized PRSV-resistant transgenic papaya samples tested positive by RT-LAMP assay, and 6 of those tested negative by RT-PCR. Therefore, the PLDMV-specific RT-LAMP is a simple, rapid, sensitive, and cost-effective tool in the field diagnosis and control of PLDMV.
Bamunusinghe, Devinka; Seo, Jang-Kyun; Rao, A L N
Genome packaging in the plant-infecting Brome mosaic virus (BMV), a member of the alphavirus-like superfamily, as well as in other positive-strand RNA viruses pathogenic to humans (e.g., poliovirus) and animals (e.g., Flock House virus), is functionally coupled to replication. Although the subcellular localization site of BMV replication has been identified, that of the capsid protein (CP) has remained elusive. In this study, the application of immunofluorescence confocal microscopy to Nicotiana benthamiana leaves expressing replication-derived BMV CP as a green fluorescent protein (GFP) fusion, in conjunction with antibodies to the CP and double-stranded RNA, a presumed marker of RNA replication, revealed that the subcellular localization sites of replication and CP overlap. Our temporal analysis by transmission electron microscopy of ultrastructural modifications induced in BMV-infected N. benthamiana leaves revealed a reticulovesicular network of modified endoplasmic reticulum (ER) incorporating large assemblies of vesicles derived from ER accumulated in the cytoplasm during BMV infection. Additionally, for the first time, we have found by ectopic expression experiments that BMV CP itself has the intrinsic property of modifying ER to induce vesicles similar to those present in BMV infections. The significance of CP-induced vesicles in relation to CP-organized viral functions that are linked to replication-coupled packaging is discussed.
Full Text Available Potential porcine circovirus type 2 (PCV2 capsid protein epitopes, suitable for expression on the surface of cucumber mosaic virus (CMV particles were determined by a thorough analysis of the predicted PCV capsid protein structure. The ab initio protein structure prediction was carried out with fold recognition and threading methods. The putative PCV epitopes were selected on the basis of PCV virion models and integrated into the plant virus coat protein, after amino acid position 131. The recombinants were tested for infectivity and stability on different Nicotiana species and stable recombinant virus particles were purified. The particles were tested for their ability to bind to PCV induced porcine antibodies and used for specific antibody induction in mice and pigs. The results showed that PCV epitopes expressed on the CMV surface were recognized by the porcine antibodies and they were also able to induce PCV specific antibody response. Challenge experiment with PCV2 carried out in immunized pigs showed partial protection against the infection. Based on these results it was concluded that specific antiviral vaccine production for the given pathogen was feasible, offering an inexpensive way for the mass production of such vaccines.
Holz, S; Kube, M; Bartoszewski, G; Büttner, C
The role of soluble silicon (Si) in alleviating viral plant infections is largely unknown. In order to analyse this gap in knowledge, this study provides insights into the relative gene expression data obtained from 1) control, 2) Cucumber mosaic virus (CMV)-infected and 3) sodium silica-treated, CMV-infected Cucumis sativus line B10 tissue cultures regenerated plants. The absence or presence of CMV was determined through RT-PCR, six days' post-inoculation. qRT-PCR was performed on five selected host genes related to CMV-defence (argonaute protein, WRKY transcription factor) and replication (chaperone, heat shock cognate protein, aquaporin). Relative gene expressions from Si-treated, CMV-infected clones were not significantly different from CMV-infected clones, but they were significantly different from the control plants. The upregulation of the chaperone, and heat shock cognate genes in Si-treated clones, is associated with enhanced virus replication, while the gene expression of the transcription factor increases and is related to defence, in contrast to decreased expression in CMV-infected clones. Aquaporin gene expression was downregulated and the argonaute expression was unaffected in both Si-treated, CMV-infected as well as CMV-infected clones. Since both alleviating and supportive gene shifts are observed in Si-treated plantlets for key genes related to the virus infection examined herein, sodium silica is suggested to have a neutral and limited impact on CMV infection in cucumber cultures.
Wei Chen; Wenting Liu; Honghong Jiao; Huawei Zhang; Julong Cheng; Yunfeng Wu
Tobacco mosaic virus (TMV) causes significant yield loss in susceptible crops irrigated with contaminated water. However, detection of TMV in water is difficult owing to extremely low concentrations of the virus. Here, we developed a simple method for the detection and quantiifcation of TMV in irrigation water. TMV was reliably detected at concentrations as low as 10 viral copies/µL with real-time PCR. The sensitivity of detection was further improved using polyethylene glycol 6000 (PEG6000, MW 6000) to concentrate TMV from water samples. Among the 28 samples from Shaanxi Province examined with our method, 17 were tested positive after virus concentration. Infectivity of TMV in the original water sample as well as after concentration was conifrmed using PCR. The limiting concentration of TMV in water to re-infect plants was determined as 102 viral copies/mL. The method developed in this study offers a novel approach to detect TMV in irrigation water, and may provide an effective tool to control crop infection.
Estela Bonilha; Ricardo Gioria; Rômulo Fujito Kobori; Paulo Tarcísio Della Vecchia; Sônia Maria de Stefano Piedade; Jorge Alberto Marques Rezende
Papaya ringspot virus - type W (PRSV-W) and Zucchini yellow mosaic virus (ZYMV) are the most prevalent viruses in cucurbit crops in Brazil and responsible for frequent yield losses. Diseases caused by these viruses are difficult to control. The objective of this work was to evaluate the effects of mild strains PRSV-W-1 and ZYMV-M on the yield of Cucurbita pepo L. cvs. Samira, Novita Plus, AF 2847, and Yasmin, under plastic greenhouse and field conditions. Plants infected with ZYMV-M and grown...
Karpova, O V; Lednev, V N; Mironova, T V; Oshurko, V B; Pershin, S M; Petrova, E K; Tcherniega, N V; Zemskov, K I
Laser pulses interaction with tobacco mosaic virus (TMV) in Tris-HCl pH7.5 buffer and in water has been investigated. 20 ns ruby laser pulses have been used for excitation. Spectrum of the light passing through the sample was registered with the help of Fabri-Perot interferometer. In the case of TMV in water we observed in the spectrum only one line of the exciting laser light, for TMV in Tris-HCl pH7.5 buffer second line appeared, corresponding to the stimulated low-frequency Raman scattering (SLFRS) on the breathing radial mode of TMV. SLFRS frequency shift by 2 cm-1, (60 GHz), conversion efficiency and threshold are measured for the first time to the best of our knowledge.
López, C; Aramburu, J; Galipienso, L; Nuez, F
Preparations of double-stranded RNAs (dsRNAs) extracted from Nicotiana tabacum cv Xanthi plants infected with a subgroup IB isolate of Cucumber mosaic virus (CMV) were found to contain a heterogeneous population of defective RNAs (D-RNAs) derived from RNA 3. Characterised D-RNAs ranged in size from 1.5 to 1.9 kb and were derived either by a single in-frame deletion within the 3a or 3b genes or by means of double in-frame deletions within both genes. Also, northern blot hybridisation showed two other types of RNA derived from RNA 3: (a) RNA species of ca. 0.7 kb containing the 3'-terminus but lacking the 5'-terminus, which could be 3'-coterminal subgenomic of D-RNAs derived from the 3b gene and (b) RNA species of unknown origin of ca. 0.8 kb containing the 5'-terminus but lacking the 3'-terminus.
Karpova, O. V.; Kudryavtseva, A. D.; Lednev, V. N.; Mironova, T. V.; Oshurko, V. B.; Pershin, S. M.; Petrova, E. K.; Tcherniega, N. V.; Zemskov, K. I.
The interaction of laser pulses with tobacco mosaic virus (TMV) in Tris-HCl pH7.5 buffer and in water has been investigated. Ruby laser pulses of 20 ns duration have been used for excitation. The spectrum of the light passing through the sample was registered with the help of a Fabry-Perot interferometer. In the case of TMV in water we observed in the spectrum only one line of the exciting laser light, but for TMV in Tris-HCl pH7.5 buffer a second line appeared, corresponding to stimulated low-frequency Raman scattering (SLFRS) on the breathing radial mode of TMV. The frequency shift of the SLFRS by 2 cm-1 (60 GHz), the conversion efficiency and the threshold are measured for the first time to the best of our knowledge.
Jie-hong ZHAO; Ji-shun ZHANG; Yi WANG; Ren-gang WANG; Chun WU; Long-jiang FAN; Xue-liang REN
DNA methylation plays an important role in the epigenetic regulation of gene expression during plant growth,development,and polyploidization.However,there is still no distinct evidence in tobacco regarding the distribution of the methylation pattern and whether it contributes to qualitative characteristics.We studied the levels and patterns of methylation polymorphism at CCGG sites in 48 accessions of allotetraploid flue-cured tobacco,Nicotiana tabacum,using a methylation-sensitive amplified polymorphism (MSAP) technique.The results showed that methylation existed at a high level among tobacco accessions,among which 49.3％ sites were methylated and 69.9％ allelic sites were polymorphic.A cluster analysis revealed distinct patterns of geography-specific groups.In addition,three polymorphic sites significantly related to tobacco mosaic virus (TMV) resistance were explored.This suggests that tobacco breeders should pay more attention to epigenetic traits.
Moore, Lauren; Hamorsky, Krystal; Matoba, Nobuyuki
Here, we describe a method to produce a recombinant cholera toxin B subunit in Nicotiana benthamiana plants (CTBp) using the GENEWARE(®) tobacco mosaic virus vector system. Infectious transcripts of the vector RNA are generated in vitro and inoculated on N. benthamiana seedlings. After 11 days, CTBp is extracted in a simple tris buffer at room temperature. No protease inhibitor is required. The leaf homogenate is treated with mild heat and a pH shift to selectively precipitate host-derived proteins. CTBp is purified to >95 % homogeneity by two-step chromatography using immobilized metal affinity and ceramic hydroxyapatite resins. This procedure yields on average 400 mg of low-endotoxin CTBp from 1 kg of fresh leaf material.
Eber, Fabian J.; Eiben, Sabine; Jeske, Holger; Wege, Christina
The in vitro assembly of artificial nanotubular nucleoprotein shapes based on tobacco mosaic virus-(TMV-)-derived building blocks yielded different spatial organizations of viral coat protein subunits on genetically engineered RNA molecules, containing two or multiple TMV origins of assembly (OAs). The growth of kinked nanoboomerangs as well as of branched multipods was determined by the encapsidated RNAs. A largely simultaneous initiation at two origins and subsequent bidirectional tube elongation could be visualized by transmission electron microscopy of intermediates and final products. Collision of the nascent tubes' ends produced angular particles with well-defined arm lengths. RNAs with three to five OAs generated branched multipods with a maximum of four arms. The potential of such an RNA-directed self-assembly of uncommon nanotubular architectures for the fabrication of complex multivalent nanotemplates used in functional hybrid materials is discussed.
Slavokhotova, A A; Andreeva, E N; Shiian, A N; Odintsova, T I; Pukhal'skiĭ, V A
The primary structure of the coat protein (CP) gene was examined for pathogenic strain MS-1 and vaccine strain VIROG-43M of the cucumber green mottle mosaic virus (CGMMV). In CP amino acid composition, strains MS-1 and VIROG-43M are typical representatives of CGMMV: their CPs have 98-100% homology to CPs of other tobamoviruses of the group. The CP gene has the same nucleotide composition in pathogenic MS-1 and vaccine VIROG-43M, indicating that strain attenuation is not determined by this gene. The CP amino acid sequences of the two Russian strains are fully identical to the CP sequences of two Greek strains, GR-3 and GR-5. However, the nucleotide sequences of their genes differ in 13 bp, testifying to the difference between the Russian and Greek strains.
WIWIEK SRI WAHYUNI
Full Text Available In semiaerobic conditions, different composting processes of vegetable wastes have different characteristics. When compost extracts amended with the effective microorganism-4 (EM4, +E and Pseudomonas aeruginosa Ch1 (+B stored for 40 days, the bacteria population and P-content increased. Tobacco plants treated with compost extracts amended with +E+B and [+E+B] directly to organic materials and inoculated with Cucumber mosaic virus (CMV both sprayed or watered applications reduced the disease severity. This is due to the higher bacteria population in the root and rhizosphere, particularly the activities of P. aeruginosa Ch1 as plant growth promoting rhizobacteria (PGPR rather than the activities of bacteria from EM4. The role of P. aeruginosa Ch1 to induce resistance of the plants to CMV was suggested by producing siderophores under the limited Fe conditions,17-20 ppm.
赛吉庆; 康良仪; 黄忠; 史春霖; 田波; 谢友菊
The 3’-terminal 1 279 nucleotide sequence of maize dwarf mosaic virus (MDMV) genome has been determined. This sequence contains an open reading frame of 1023 nudeotides and a 3’ -non-coding region of 256 nucleotides. The open reading frame includes all of the coding regions for the viral capsid protein (CP) and part of the viral nuclear inclusion protein (Nib). The predicted viral CP consists of 313 amino acid residues with a calculated molecular weight of 35400. The amino acid sequence of the viral CP derived from MDMV cDNA shows about 47%-54% homology to that of 4 other potyviruses. The viral CP gene was constructed in frame with the lacZ gene in pUC19 plasmid and expressed in E. coli cells. The fusion polypeptide positively reacted in Western blot with an antiserum prepared against the native viral CP.
Ma, Yuan; Mao, Xin-Ying; Huang, Lie-Jun; Fan, Yi-Min; Gu, Wei; Yan, Chen; Huang, Tao; Zhang, Jian-Xin; Yuan, Chun-Mao; Hao, Xiao-Jiang
Five new naturally occurring natural products, including two atisine-type diterpene alkaloids (1 and 2), two atisane-type diterpenes (3 and 4), and a new natural product spiramine C2 (5), along with nine known ones (6-14), were isolated from the ethanolic extracts of the whole plant of Spiraea japonica var. acuminata Franch. Their structures were elucidated by extensive spectroscopic analysis. The anti-tobacco mosaic virus (TMV) activities of all the compounds were evaluated by the conventional half-leaf method. Six compounds (2, 3, 6, 7, 11, and 12) exhibited moderate activities at 100 μg/mL with inhibition rates in the range of 69.4-92.9%, which were higher than that of the positive control, ningnanmycin. Their preliminary structure-activity relationships were also discussed.
Li, Na; Deng, Zhaoqi; Lin, Yuan; Zhang, Xiaojie; Geng, Yanhou; Ma, Dongge; Su, Zhaohui
Layer-by-layer assembly technique was used to construct ultrathin film of cowpea mosaic virus (CPMV) by electrostatic interactions, and the film was employed as a precursor on which an OF8T2 film was deposited by spin coating. Amplified spontaneous emission (ASE) was observed and improved for the OF8T2 film. Compared with OF8T2 film on quartz, the introduction of CPMV nanoparticles reduced the threshold and loss, and remarkably increased the net gain. The threshold, loss, and gain reached 0.05 mJ/pulse, 6.9 cm-1, and 82 cm-1, respectively. CPMV nanoparticles may enormously scatter light, resulting in a positive feedback, thus the ASE is easily obtained and improved.
Hasiów-Jaroszewska, Beata; Chrzanowski, Mateusz; Budzyńska, Daria; Rymelska, Natalia; Borodynko-Filas, Natasza
In recent years, the occurrence of cucumber mosaic virus (CMV) has been noted in zucchini crops in Poland. Beside characteristic isolates, which displayed mosaics and chlorosis on infected plants, new necrotic isolates have also been identified. Here, we analysed the molecular variability of 27 isolates of CMV collected from zucchini in various regions of the country. Sequence and phylogenetic analysis based on the genes encoding the coat (CP) and movement (MP) proteins revealed that the Polish isolates belong to two subgroups: IA and II, with the prevalence of subgroup II. New recombinant variants with an IA-MP/II-CP pattern for RNA3 were also detected.
Full Text Available Previous studies with Paenibacillus lentimorbus B-30488" (hereafter referred as B-30488, a plant growth promoting rhizobacteria (PGPR isolated from cow's milk, revealed its capabilities to improve plant quality under normal and stress conditions. Present study investigates its potential as a biocontrol agent against an economically important virus, Cucumber mosaic virus (CMV, in Nicotiana tabacum cv. White Burley plants and delineates the physical, biophysical, biochemical and molecular perturbations due to the trilateral interactions of PGPR-host-CMV. Soil inoculation of B-30488 enhanced the plant vigor while significantly decreased the virulence and virus RNA accumulation by ~12 fold (91% in systemic leaves of CMV infected tobacco plants as compared to the control ones. Histology of these leaves revealed the improved tissue's health and least aging signs in B-30488 inoculated tobacco plants, with or without CMV infection, and showed lesser intercellular spaces between collenchyma cells, reduced amount of xyloglucans and pectins in connecting primary cells, and higher polyphenol accumulation in hypodermis layer extending to collenchyma cells. B-30488 inoculation has favorably maneuvered the essential biophysical (ion leakage and photosynthetic efficiency and biochemical (sugar, proline, chlorophyll, malondialdehyde, acid phosphatase and alkaline phosphatase attributes of tobacco plants to positively regulate and release the virus stress. Moreover, activities of defense related enzymes (ascorbate peroxidase, guaiacol peroxidase, superoxide dismutase and catalase induced due to CMV-infection were ameliorated with inoculation of B-30488, suggesting systemic induced resistance mediated protection against CMV in tobacco. The quantitative RT-PCR analyses of the genes related to normal plant development, stress and pathogenesis also corroborate well with the biochemical data and revealed the regulation (either up or down of these genes in favor of
Palama, Tony Lionel; Grisoni, Michel; Fock-Bastide, Isabelle; Jade, Katia; Bartet, Laetitia; Choi, Young Hae; Verpoorte, Robert; Kodja, Hippolyte
The genus Vanilla which belongs to the Orchidaceae family comprises more than 110 species of which two are commercially cultivated (Vanilla planifolia and Vanilla xtahitensis). The cured pods of these species are the source of natural vanilla flavor. In intensive cultivation systems the vines are threatened by viruses such as Cymbidium mosaic virus (CymMV). In order to investigate the effect of CymMV on the growth and metabolome of vanilla plants, four accessions grown in intensive cultivation systems under shadehouse, CR01 (V. planifolia), CR17 (V. xtahitensis), CR03 (V. planifolia × V. xtahitensis) and CR18 (Vanilla pompona), were challenged with an isolate of CymMV. CymMV infected plants of CR01, CR03 and CR17 had a reduced growth compared to healthy plants, while there was no significant difference in the growth of CR18 vines. Interestingly, CR18 had qualitatively more phenolic compounds in leaves and a virus titre that diminished over time. No differences in the metabolomic profiles of the shadehouse samples obtained by nuclear magnetic resonance (NMR) were observed between the virus infected vs. healthy plants. However, using in- vitro V. planifolia plants, the metabolomic profiles were affected by virus infection. Under these controlled conditions the levels of amino acids and sugars present in the leaves were increased in CymMV infected plants, compared to uninfected ones, whereas the levels of phenolic compounds and malic acid were decreased. The metabolism, growth and viral status of V. pompona accession CR18 contrasted from that of the other species suggesting the existence of partial resistance to CymMV in the vanilla germplasm.
Wang, Jinbo; Turina, Massimo; Medina, Vicente; Falk, Bryce W
Lettuce infectious yellows virus (LIYV), the type member of the genus Crinivirus in the family Closteroviridae, is specifically transmitted by the sweet potato whitefly (Bemisia tabaci) in a semipersistent manner. LIYV infections result in a low virus titer in plants and protoplasts, impeding reverse genetic efforts to analyze LIYV gene/protein functions. We found that synergistic interactions occurred in mixed infections of LIYV and Turnip mosaic virus (TuMV) in Nicotiana benthamiana plants, and these resulted in enhanced accumulation of LIYV. Furthermore, we examined the ability of transgenic plants and protoplasts expressing only the TuMV P1/HC-Pro sequence to enhance the accumulation of LIYV. LIYV RNA and protein titers increased by as much as 8-fold in these plants and protoplasts relative to control plants. LIYV infections remained phloem-limited in P1/HC-Pro transgenic plants, suggesting that enhanced accumulation of LIYV in these plants was due primarily to increased replication efficiency, not to greater spread.
Junli Feng; Xin Liu; Leiyu Lai; Jishuang Chen
MicroRNAs (miRNAs) play vital roles in regulating plant growth and development. Recent work has shown that miRNA-mediated regulation of cellular mRNA expression is involved in pathogen-host interactions. However, knowledge about the timing and spatial regulation of plant miRNA expression is still limited. Here, we use stem-loop real-time reverse transcription-polymerase chain reaction to quantify the expression changes of seven miRNAs and their target mRNAs in different tomato tissues during the pathogenic processes. Compared with mock-inoculated plants, the expression levels of investigated miRNAs and mRNAs were enhanced by different degrees upon Cucumber mosaic virus (CMV)-Fny and Tomato aspermy virus-Bj infections, but were almost unchanged in CMV-FnyA2b (a CMV-Fny 2b-deletion mutant)-infected tomato seedlings. In addition, the obvious up-regulation of several miRNAs and target mRNAs in some tomato tissues suggested their special roles in these tissues' organogenesis and development. Temporal analyses also revealed that the expressions of these miRNAs and mRNAs were highly regulated by different viral infections. Taken together, the observed spatially and temporally changes in miRNAs and target mRNAs expression levels indicate the abilities of different viruses to interfere with miRNA pathway, and are correlated with their respective functions in phenotype determination in different tomato tissues.
Kung, Yi-Jung; Bau, Huey-Jiunn; Wu, Yi-Ling; Huang, Chiung-Huei; Chen, Tsui-Miao; Yeh, Shyi-Dong
During the field tests of coat protein (CP)-transgenic papaya lines resistant to Papaya ringspot virus (PRSV), another Potyvirus sp., Papaya leaf-distortion mosaic virus (PLDMV), appeared as an emerging threat to the transgenic papaya. In this investigation, an untranslatable chimeric construct containing the truncated CP coding region of the PLDMV P-TW-WF isolate and the truncated CP coding region with the complete 3' untranslated region of PRSV YK isolate was transferred into papaya (Carica papaya cv. Thailand) via Agrobacterium-mediated transformation to generate transgenic plants with resistance to PLDMV and PRSV. Seventy-five transgenic lines were obtained and challenged with PRSV YK or PLDMV P-TW-WF by mechanical inoculation under greenhouse conditions. Thirty-eight transgenic lines showing no symptoms 1 month after inoculation were regarded as highly resistant lines. Southern and Northern analyses revealed that four weakly resistant lines have one or two inserts of the construct and accumulate detectable amounts of transgene transcript, whereas nine resistant lines contain two or three inserts without significant accumulation of transgene transcript. The results indicated that double virus resistance in transgenic lines resulted from double or more copies of the insert through the mechanism of RNA-mediated posttranscriptional gene silencing. Furthermore, three of nine resistant lines showed high levels of resistance to heterologous PRSV strains originating from Hawaii, Thailand, and Mexico. Our transgenic lines have great potential for controlling a number of PRSV strains and PLDMV in Taiwan and elsewhere.
Panyukov, Yuliy; Yudin, Igor; Drachev, Vladimir; Dobrov, Evgeny; Kurganov, Boris
The kinetics of heat-induced and cetyltrimethylammonium bromide induced amorphous aggregation of tobacco mosaic virus coat protein in Na(+)/Na(+) phosphate buffer, pH 8.0, have been studied using dynamic light scattering. In the case of thermal aggregation (52 degrees C) the character of the dependence of the hydrodynamic radius (R(h)) on time indicates that at certain instant the population of aggregates is split into two components. The size of the aggregates of one kind remains practically constant in time, whereas the size of aggregates of other kind increases monotonously in time reaching the values characteristic of aggregates prone to precipitation (R(h)=900-1500 nm). The construction of the light scattering intensity versus R(h) plots shows that the large aggregates (the start aggregates) exist in the system at the instant the initial increase in the light scattering intensity is observed. For thermal aggregation the R(h) value for the start aggregates is independent of the protein concentration and equal to 21.6 nm. In the case of the surfactant-induced aggregation (at 25 degrees C) no splitting of the aggregates into two components is observed and the size of the start aggregates turns out to be much larger (107 nm) than on the thermal aggregation. The dependence of R(h) on time for both heat-induced aggregation and surfactant-induced aggregation after a lapse of time follows the power law indicating that the aggregation process proceeds in the kinetic regime of diffusion-limited cluster-cluster aggregation. Fractal dimension is close to 1.8. The molecular chaperone alpha-crystallin does not affect the kinetics of tobacco mosaic virus coat protein thermal aggregation.
Pepino mosaic, once an emerging disease a decade ago, has become endemic on greenhouse tomatoes worldwide in recent years. Three distinct genotypes of Pepino mosaic virus (PepMV), including EU, US1 and CH2 have been recognized. Our earlier study conducted in 2006-2007 demonstrated a predominant EU...
Balint, R.; Cohen, S.S.
The authors have reported that protoplasts from plants infected with turnip yellow mosaic virus (TYMV) continue to produce virus in culture and that newly formed virus particles contained predominantly newly synthesized spermidine and spermine. They now report similar results with healthy protoplasts infected in vitro, in which essentially all of the virus is newly formed. Again, newly synthesized spermidine and spermine were preferentially incorporated into virus. DCHA inhibited spermidine synthesis by 85%, leading in 20 hr to a 60% depletion of the cellular spermidine and a 30% reduction in the amount of spermidine per virion. Spermine synthesis increased, however, producing a 40% increase in cellular spermine and 50-100% increase in the amount of spermine per virion. Thus, in spite of spermidine depletion, the total positive charge contributed by polyamines to the virus was essentially conserved.
Dubey, Vimal Kumar; Aminuddin; Singh, Vijai Pal
The majority of Gladiolus plants growing in the botanical garden at NBRI, Lucknow, India and adjoining areas exhibited symptoms of mosaic, color breaking, stunting of spikes and reduction in flower size. The occurrence of Cucumber mosaic virus (CMV) was suspected in symptomatic Gladiolus plants. Cucumber mosaic virus, the type species of the genus Cucumovirus of the family Bromoviridae, is an important plant virus worldwide, which infects many plants and causes quantity and quality losses. For virus characterization, total RNA was isolated from leaves of infected plants and used in reverse transcriptase polymerase chain reaction with a primer set designed in the Cucumber mosaic virus coat protein region. Viral amplicons of the expected 657 bp size were obtained from infected plants. No viral amplicon was obtained from healthy control plants. Viral amplicons were cloned and sequenced (DQ295914). Molecular characterization was performed and phylogenetic relationship determined by the comparison of coat protein gene nucleotide and amino acid sequences with other Cucumber mosaic virus isolates reported from India and worldwide. The nucleotide and amino acid percentage comparison and phylogenetic tree results revealed that Cucumber mosaic virus infecting Gladiolus show resemblance with the Fny strain, which is not common in the Asian continent.
Full Text Available Cucumber mosaic virus (CMV has a worldwide distribution and the widest host range of any known plant virus. From 2000 to 2012, epidemics of CMV severely affected the production of snap bean (Phaseulos vulgaris L. in the Midwest and Northeastern United States. Virus diversity leading to emergence of new strains is often considered a significant factor in virus epidemics. In addition to epidemics, new disease phenotypes arising from genetic exchanges or mutation can compromise effectiveness of plant disease management strategies. Here, we captured a snapshot of genetic variation of 32 CMV isolates collected from different regions of the U.S including new field as well as historic isolates. Nucleotide diversity (π was low for U.S. CMV isolates. Sequence and phylogenetic analyses revealed that CMV subgroup I is predominant in the US and further showed that the CMV population is a mixture of subgroups IA and IB. Furthermore, phylogenetic analysis suggests likely reassortment between subgroups IA and IB within five CMV isolates. Based on phylogenetic and computational analysis, recombination between subgroups I and II as well as IA and IB in RNA 3 was detected. This is the first report of recombination between CMV subgroups I and II. Neutrality tests illustrated that negative selection was the major force operating upon the CMV genome, although some positively selected sites were detected for all encoded proteins. Together, these data suggest that different regions of the CMV genome are under different evolutionary constraints. These results also delineate composition of the CMV population in the US, and further suggest that recombination and reassortment among strain subgroups does occur but at a low frequency, and point towards CMV genomic regions that differ in types of selection pressure.
研究菌克毒克对烟草花叶病的防治效果，结果表明：菌克毒克(8%宁南霉素水剂内销品)对烟草花叶病有较好的防治效果，平均防治效果达66.7%，可促进烟株的生长，经济效益显著。%Effect of Junkeduke on tobacco mosaic virus(TMV)and cucumber mosaic virus(CMV)were studied . The results showed that Junkeduke(8%Ningnanmycin domestic product)had better control efficincy,the average control efficiency reached 66.7%.It could promote the growth of tobacco plants,economic benefits was significant.
Full Text Available The rod-shaped nanoparticles of the widespread plant pathogen tobacco mosaic virus (TMV have been a matter of intense debates and cutting-edge research for more than a hundred years. During the late 19th century, their behavior in filtration tests applied to the agent causing the 'plant mosaic disease' eventually led to the discrimination of viruses from bacteria. Thereafter, they promoted the development of biophysical cornerstone techniques such as electron microscopy and ultracentrifugation. Since the 1950s, the robust, helically arranged nucleoprotein complexes consisting of a single RNA and more than 2100 identical coat protein subunits have enabled molecular studies which have pioneered the understanding of viral replication and self-assembly, and elucidated major aspects of virus–host interplay, which can lead to agronomically relevant diseases. However, during the last decades, TMV has acquired a new reputation as a well-defined high-yield nanotemplate with multivalent protein surfaces, allowing for an ordered high-density presentation of multiple active molecules or synthetic compounds. Amino acid side chains exposed on the viral coat may be tailored genetically or biochemically to meet the demands for selective conjugation reactions, or to directly engineer novel functionality on TMV-derived nanosticks. The natural TMV size (length: 300 nm in combination with functional ligands such as peptides, enzymes, dyes, drugs or inorganic materials is advantageous for applications ranging from biomedical imaging and therapy approaches over surface enlargement of battery electrodes to the immobilization of enzymes. TMV building blocks are also amenable to external control of in vitro assembly and re-organization into technically expedient new shapes or arrays, which bears a unique potential for the development of 'smart' functional 3D structures. Among those, materials designed for enzyme-based biodetection layouts, which are routinely applied
Peng, Jiyu; Song, Kunlin; Zhu, Hongyan; Kong, Wenwen; Liu, Fei; Shen, Tingting; He, Yong
Tobacco mosaic virus (TMV) is one of the most devastating viruses to crops, which can cause severe production loss and affect the quality of products. In this study, we have proposed a novel approach to discriminate TMV-infected tobacco based on laser-induced breakdown spectroscopy (LIBS). Two different kinds of tobacco samples (fresh leaves and dried leaf pellets) were collected for spectral acquisition, and partial least squared discrimination analysis (PLS-DA) was used to establish classification models based on full spectrum and observed emission lines. The influences of moisture content on spectral profile, signal stability and plasma parameters (temperature and electron density) were also analysed. The results revealed that moisture content in fresh tobacco leaves would worsen the stability of analysis, and have a detrimental effect on the classification results. Good classification results were achieved based on the data from both full spectrum and observed emission lines of dried leaves, approaching 97.2% and 88.9% in the prediction set, respectively. In addition, support vector machine (SVM) could improve the classification results and eliminate influences of moisture content. The preliminary results indicate that LIBS coupled with chemometrics could provide a fast, efficient and low-cost approach for TMV-infected disease detection in tobacco leaves.
Babu, B; Hegde, V
Dasheen mosaic virus (DsMV) infecting three major edible aroids namely Amorphophallus paeoniifolius, Colocasia esculenta, and Xanthosoma sagittifolium cultivated in India was characterized. Infected plants showing typical DsMV symptoms were subjected to reverse transcription-polymerase chain reaction, and an amplification of a 963 bp fragment which encoded the coat protein (CP) gene was obtained. BLAST analysis of the cloned DNA amplicon revealed the identity of the virus to be that of DsMV. Sequence identity matrix of the nucleotide sequences among the three isolates showed that the DsMV isolate infecting A. paeoniifolius and C. esculenta shared an identity as high as 93%, while the DsMV isolate from X. sagittifolium shared an identity of only 73% and 76% with the DsMV isolates from A. paeoniifolius and C. esculenta, respectively. Comparative analysis of the coat protein of the three DsMV isolates showed the presence of DVG motif (A. paeoniifolius and C. esculenta) and DTG motif in X. sagittifolium and several varying potential threonine and asparagine rich N-glycosylation motifs. Single amino acid substitution of the several conserved motifs occurs in all the three DsMV isolates. This is the first characterization of DsMV isolates infecting A. paeoniifolius, C. esculenta, and X. sagittifolium plants in India.
Wang, Zhao; Hryc, Corey F; Bammes, Benjamin; Afonine, Pavel V; Jakana, Joanita; Chen, Dong-Hua; Liu, Xiangan; Baker, Matthew L; Kao, Cheng; Ludtke, Steven J; Schmid, Michael F; Adams, Paul D; Chiu, Wah
Advances in electron cryo-microscopy have enabled structure determination of macromolecules at near-atomic resolution. However, structure determination, even using de novo methods, remains susceptible to model bias and overfitting. Here we describe a complete workflow for data acquisition, image processing, all-atom modelling and validation of brome mosaic virus, an RNA virus. Data were collected with a direct electron detector in integrating mode and an exposure beyond the traditional radiation damage limit. The final density map has a resolution of 3.8 Å as assessed by two independent data sets and maps. We used the map to derive an all-atom model with a newly implemented real-space optimization protocol. The validity of the model was verified by its match with the density map and a previous model from X-ray crystallography, as well as the internal consistency of models from independent maps. This study demonstrates a practical approach to obtain a rigorously validated atomic resolution electron cryo-microscopy structure.
Grangeon, Romain; Agbeci, Maxime; Chen, Jun; Grondin, Gilles; Zheng, Huanquan; Laliberté, Jean-François
The impact of turnip mosaic virus (TuMV) infection on the endomembranes of the host early secretory pathway was investigated using an infectious clone that has been engineered for tagging viral membrane structures with a fluorescent protein fused to the viral protein 6K(2). TuMV infection led to the amalgamation of the endoplasmic reticulum (ER), Golgi apparatus, COPII coatamers, and chloroplasts into a perinuclear globular structure that also contained viral proteins. One consequence of TuMV infection was that protein secretion was blocked at the ER-Golgi interface. Fluorescence recovery after photobleaching (FRAP) experiments indicated that the perinuclear structure cannot be restocked in viral components but was dynamically connected to the bulk of the Golgi apparatus and the ER. Experiments with 6K(2) fused to photoactivable green fluorescent protein (GFP) showed that production of motile peripheral 6K(2) vesicles was functionally linked to the perinuclear structure. Disruption of the early secretory pathway did not prevent the formation of the perinuclear globular structure, enhanced the clustering of peripheral 6K(2) vesicles with COPII coatamers, and led to inhibition of cell-to-cell virus movement. This suggests that a functional secretory pathway is not required for the formation of the TuMV perinuclear globular structure and peripheral vesicles but is needed for successful viral intercellular propagation.
Peart, Jack R; Cook, Graeme; Feys, Bart J; Parker, Jane E; Baulcombe, David C
In Arabidopsis, EDS1 is essential for disease resistance conferred by a structural subset of resistance (R) proteins containing a nucleotide-binding site, leucine-rich-repeats and amino-terminal similarity to animal Toll and Interleukin-1 (so-called TIR-NBS-LRR proteins). EDS1 is not required by NBS-LRR proteins that possess an amino-terminal coiled-coil motif (CC-NBS-LRR proteins). Using virus-induced gene silencing (VIGS) of a Nicotiana benthaminana EDS1 orthologue, we investigated the role of EDS1 in resistance specified by structurally distinct R genes in transgenic N. benthamiana. Resistance against tobacco mosaic virus mediated by tobacco N, a TIR-NBS-LRR protein, was EDS1-dependent. Two other R proteins, Pto (a protein kinase), and Rx (a CC-NBS-LRR protein) recognizing, respectively, a bacterial and viral pathogen did not require EDS1. These data, together with the finding that expression of N. benthamiana and Arabidopsis EDS1 mRNAs are similarly regulated, lead us to conclude that recruitment of EDS1 by TIR-NBS-LRR proteins is evolutionarily conserved between dicotyledenous plant species in resistance against bacterial, oomycete and viral pathogens. We further demonstrate that VIGS is a useful approach to dissect resistance signaling pathways in a genetically intractable plant species.
Full Text Available Soybean mosaic virus (SMV is one of the most devastating pathogens that cost huge economic losses in soybean production worldwide. Due to the duplicated genome, clustered and highly homologous nature of R genes, as well as recalcitrant to transformation, soybean disease resistance studies is largely lagging compared with other diploid crops. In this review, we focus on the major advances that have been made in identifying both the virulence/avirulence factors of SMV and mapping of SMV resistant genes in soybean. In addition, we review the progress in dissecting the SMV resistant signaling pathways in soybean, with a special focus on the studies using virus-induced gene silencing (VIGS. The soybean genome has been fully sequenced, and the increasingly saturated SNP markers have been identified. With these resources available together with newly developed genome editing tools, and more efficient soybean transformation system, cloning SMV resistant genes, and ultimately generating cultivars with a broader spectrum resistance to SMV are becoming more realistic than ever.
Alishiri, Athar; Rakhshandehroo, Farshad; Zamanizadeh, Hamid-Reza; Palukaitis, Peter
The incidence and distribution of Tobacco mosaic virus (TMV) and related tobamoviruses was determined using an enzyme-linked immunosorbent assay on 1,926 symptomatic horticultural crops and 107 asymptomatic weed samples collected from 78 highly infected fields in the major horticultural crop-producing areas in 17 provinces throughout Iran. The results were confirmed by host range studies and reverse transcription-polymerase chain reaction. The overall incidence of infection by these viruses in symptomatic plants was 11.3%. The coat protein (CP) gene sequences of a number of isolates were determined and disclosed to be a high identity (up to 100%) among the Iranian isolates. Phylogenetic analysis of all known TMV CP genes showed three clades on the basis of nucleotide sequences with all Iranian isolates distinctly clustered in clade II. Analysis using the complete CP amino acid sequence showed one clade with two subgroups, IA and IB, with Iranian isolates in both subgroups. The nucleotide diversity within each sub-group was very low, but higher between the two clades. No correlation was found between genetic distance and geographical origin or host species of isolation. Statistical analyses suggested a negative selection and demonstrated the occurrence of gene flow from the isolates in other clades to the Iranian population.
Full Text Available The incidence and distribution of Tobacco mosaic virus (TMV and related tobamoviruses was determined using an enzyme-linked immunosorbent assay on 1,926 symptomatic horticultural crops and 107 asymptomatic weed samples collected from 78 highly infected fields in the major horticultural crop-producing areas in 17 provinces throughout Iran. The results were confirmed by host range studies and reverse transcription-polymerase chain reaction. The overall incidence of infection by these viruses in symptomatic plants was 11.3%. The coat protein (CP gene sequences of a number of isolates were determined and disclosed to be a high identity (up to 100% among the Iranian isolates. Phylogenetic analysis of all known TMV CP genes showed three clades on the basis of nucleotide sequences with all Iranian isolates distinctly clustered in clade II. Analysis using the complete CP amino acid sequence showed one clade with two subgroups, IA and IB, with Iranian isolates in both subgroups. The nucleotide diversity within each sub-group was very low, but higher between the two clades. No correlation was found between genetic distance and geographical origin or host species of isolation. Statistical analyses suggested a negative selection and demonstrated the occurrence of gene flow from the isolates in other clades to the Iranian population.
Vitti, Antonella; Pellegrini, Elisa; Nali, Cristina; Lovelli, Stella; Sofo, Adriano; Valerio, Maria; Scopa, Antonio; Nuzzaci, Maria
Understanding the induction of plant defenses against viruses using biocontrol agents is essential for developing new strategies against these pathogens, given the ineffectiveness of chemical treatments. The ability of Trichoderma harzianum, strain T-22 (T22) to control Cucumber mosaic virus (CMV) in Solanum lycopersicum var. cerasiforme plants and the changes in the physiology of tomato treated/infected with T22/CMV were examined. Plant growth-promoting effects, photosynthetic performance, reactive oxygen species scavenging enzymes, and phytohormones were investigated. T22 improved tomato growth in terms of plant height and improved photosynthesis, total chlorophyll content and plant gas exchange. In contrast, CMV induced a negative effect on dry matter accumulation and inhibited the photosynthetic capacity. The analysis of plant hormones demonstrated that treating with T22 before or simultaneously to CMV infection, led to a systemic resistance by jasmonic acid/ethylene and salicylic acid signaling pathways. Conversely, systemic resistance was abscissic acid-dependent when T22 treatment was administered after the CMV infection. In conclusion, the data reported here indicate that the T22-based strategy may be the most effective measure against CMV.
Rhee, Sun-Ju; Hong, Jin-Sung; Lee, Gung Pyo
Three isolates of cucumber fruit mottle mosaic virus (CFMMV) were collected from melon, cucumber, and pumpkin plants in Korea. A full-length cDNA clone of CFMMV-Cm (melon isolate) was produced and evaluated for infectivity after T7 transcription in vitro (pT7CF-Cmflc). The complete CFMMV genome sequence of the infectious clone pT7CF-Cmflc was determined. The genome of CFMMV-Cm consisted of 6,571 nucleotides and shared high nucleotide sequence identity (98.8 %) with the Israel isolate of CFMMV. Based on the infectious clone pT7CF-Cmflc, a CaMV 35S-promoter driven cDNA clone (p35SCF-Cmflc) was subsequently constructed and sequenced. Mechanical inoculation with RNA transcripts of pT7CF-Cmflc and agro-inoculation with p35SCF-Cmflc resulted in systemic infection of cucumber and melon, producing symptoms similar to those produced by CFMMV-Cm. Progeny virus in infected plants was detected by RT-PCR, western blot assay, and transmission electron microscopy.
Moreno, I M; Thompson, J R; García-Arenal, F
Systemic movement of Cucumber green mottle mosaic virus (CGMMV) in cucumber plants was shown to be from photoassimilate source to sink, thus indicating phloem transport. Nevertheless, CGMMV was not detected by immunocytochemical procedures in the intermediary cell-sieve element complex in inoculated cotyledons, where photoassimilate loading occurs. In stem internodes, CGMMV was first localized in the companion cells of the external phloem and subsequently in all tissues except the medulla, therefore suggesting leakage of the virus from, and reloading into, the transport phloem during systemic movement. In systemically infected sink leaves, CGMMV was simultaneously detected in the xylem and phloem. Interestingly, CGMMV accumulated to high levels in the differentiating tracheids of young leaves implying that the xylem could be involved in the systemic movement of CGMMV. This possibility was tested using plants in which cell death was induced in a portion of the stem by steam treatment. At 24 degrees C, steam treatment effectively prevented the systemic movement of CGMMV, even though viral RNA was detected in washes of the xylem above the steamed internode suggesting that xylem circulation occurred. At 29 degrees C, CGMMV systemically infected steam-treated cucumber plants, indicating that CGMMV can move systemically via the xylem. Xylem transport of CGMMV was, however, less efficient than phloem transport in terms of the time required for systemic infection and the percentage of plants infected.
Lin, Chih-Hui; Sheu, Fuu; Lin, Hsin-Tang; Pan, Tzu-Ming
Cucumber mosaic virus (CMV) has been identified as the causal agent of several disease epidemics in most countries of the world. Insect-mediated virus diseases, such as those caused by CMV, caused remarkable loss of tomato (Solanum lycopersicon) production in Taiwan. With expression of the CMV coat protein gene (Cmvcp) in a local popular tomato cultivar L4783, transgenic tomato line R8 has showed consistent CMV resistance through T(0) to T(8). In this report, the allergenicity of the CMV coat protein (CMV cp) expressed in transgenic tomato R8 was assessed by investigation of the expression of the transgene source of protein, sequence similarity with known allergens, and resistance to pepsin hydrolysis. There is no known account for either the CMV or its coat protein being an allergen. The result of a bioinformatic search also showed no significant homology between CMV cp and any known allergen. The pepsin-susceptible property of recombinant CMV cp was revealed by a simulated gastric fluid (SGF) assay. Following the most recent FAO/WHO decision tree, all results have indicated that CMV cp was a protein with low possibility to be an allergen and the transgenic tomato R8 should be considered as safe as its host.
Full Text Available Understanding the induction of plant defenses against viruses using biocontrol agents is essential for developing new strategies against these pathogens, given the ineffectiveness of chemical treatments. The ability of Trichoderma harzianum, strain T-22 (T22 to control Cucumber mosaic virus (CMV in Solanum lycopersicum var. cerasiforme plants and the changes in the physiology of tomato treated/infected with T22/CMV were examined. Plant growth-promoting effects, photosynthetic performance, reactive oxygen species (ROS scavenging enzymes, and phytohormones were investigated. T22 improved tomato growth in terms of plant height and improved photosynthesis, total chlorophyll content and plant gas exchange. In contrast, CMV induced a negative effect on dry matter accumulation and inhibited the photosynthetic capacity. The analysis of plant hormones demonstrated that treating with T22 before or simultaneously to CMV infection, led to a systemic resistance by jasmonic acid/ethylene and salicylic acid signaling pathways. Conversely, systemic resistance was abscissic acid-dependent when T22 treatment was administered after the CMV infection. In conclusion, the data reported here indicate that the T22-based strategy may be the most effective measure against CMV.
Vitti, Antonella; Pellegrini, Elisa; Nali, Cristina; Lovelli, Stella; Sofo, Adriano; Valerio, Maria; Scopa, Antonio; Nuzzaci, Maria
Understanding the induction of plant defenses against viruses using biocontrol agents is essential for developing new strategies against these pathogens, given the ineffectiveness of chemical treatments. The ability of Trichoderma harzianum, strain T-22 (T22) to control Cucumber mosaic virus (CMV) in Solanum lycopersicum var. cerasiforme plants and the changes in the physiology of tomato treated/infected with T22/CMV were examined. Plant growth-promoting effects, photosynthetic performance, reactive oxygen species scavenging enzymes, and phytohormones were investigated. T22 improved tomato growth in terms of plant height and improved photosynthesis, total chlorophyll content and plant gas exchange. In contrast, CMV induced a negative effect on dry matter accumulation and inhibited the photosynthetic capacity. The analysis of plant hormones demonstrated that treating with T22 before or simultaneously to CMV infection, led to a systemic resistance by jasmonic acid/ethylene and salicylic acid signaling pathways. Conversely, systemic resistance was abscissic acid-dependent when T22 treatment was administered after the CMV infection. In conclusion, the data reported here indicate that the T22-based strategy may be the most effective measure against CMV. PMID:27777581
Polischuk V. P.
Full Text Available The aim of the present study is to compare the biological and molecular properties of Ukrainian soybean mosaic virus (SMV isolates with those of known strains or isolates from other countries, and to trace their possible origin. The methods of mechanical inoculation, reverse transcription polymerase chain reaction, DNA sequencing and phylogenetic analysis have been used. Results. Five SMV isolates have been collected and biologically purified from breeding plots in Vinnitsa region of Ukraine. It has been found that all these isolates show the same reaction patterns when infecting 11 differential soybean cultivars. Phylogenetic analysis of sequences of the coat protein coding region and P1 coding region revealed strong genetic relationships between representative Ukrainian (UA1Gr and SMV-VA2 isolates which together were sorted in one clade with G2 strain. The investigation of sequence identity showed that different genomic regions of SMV were under different evolutionary constraints. Conclusions. SMV, isolated in Ukraine for the first time, belongs to the G2 strain group that is widespread in North America. The SMV isolates obtained in this work may be employed in the Ukrainian national breeding programs to create soybean with durable virus resistance.
Full Text Available Abstract RNA recombination is one of the two major factors that create RNA genome variability. Assessing its incidence in plant RNA viruses helps understand the formation of new isolates and evaluate the effectiveness of crop protection strategies. To search for recombination in Soybean mosaic virus (SMV, the causal agent of a worldwide seed-borne, aphid-transmitted viral soybean disease, we obtained all full-length genome sequences of SMV as well as partial sequences encoding the N-terminal most (P1 protease and the C-terminal most (capsid protein; CP viral protein. The sequences were analyzed for possible recombination events using a variety of automatic and manual recombination detection and verification approaches. Automatic scanning identified 3, 10, and 17 recombination sites in the P1, CP, and full-length sequences, respectively. Manual analyses confirmed 10 recombination sites in three full-length SMV sequences. To our knowledge, this is the first report of recombination between distinct SMV pathotypes. These data imply that different SMV pathotypes can simultaneously infect a host cell and exchange genetic materials through recombination. The high incidence of SMV recombination suggests that recombination plays an important role in SMV evolution. Obtaining additional full-length sequences will help elucidate this role.
Lizotte, P. H.; Wen, A. M.; Sheen, M. R.; Fields, J.; Rojanasopondist, P.; Steinmetz, N. F.; Fiering, S.
Nanotechnology has tremendous potential to contribute to cancer immunotherapy. The ‘in situ vaccination’ immunotherapy strategy directly manipulates identified tumours to overcome local tumour-mediated immunosuppression and subsequently stimulates systemic antitumour immunity to treat metastases. We show that inhalation of self-assembling virus-like nanoparticles from cowpea mosaic virus (CPMV) reduces established B16F10 lung melanoma and simultaneously generates potent systemic antitumour immunity against poorly immunogenic B16F10 in the skin. Full efficacy required Il-12, Ifn-γ, adaptive immunity and neutrophils. Inhaled CPMV nanoparticles were rapidly taken up by and activated neutrophils in the tumour microenvironment as an important part of the antitumour immune response. CPMV also exhibited clear treatment efficacy and systemic antitumour immunity in ovarian, colon, and breast tumour models in multiple anatomic locations. CPMV nanoparticles are stable, nontoxic, modifiable with drugs and antigens, and their nanomanufacture is highly scalable. These properties, combined with their inherent immunogenicity and demonstrated efficacy against a poorly immunogenic tumour, make CPMV an attractive and novel immunotherapy against metastatic cancer.
P Rodríguez Pardina
Full Text Available Minthostachys mollis (Kunth. Griseb., "peperina", un miembro de la familia Lamiaceae, es una especie aromática que se emplea en la farmacología moderna y en medicina. Está ampliamente distribuida en los Andes, desde Venezuela y Colombia hasta Argentina. En el último país, la principal área de explotación de peperina incluye el área serrana de la provincia de Córdoba, donde la especie es arrancada indiscriminadamente, lo que conlleva una pérdida irreversible de germoplasma. A los fines de preservar este recurso nativo y fuente regional de ingresos, la especie está siendo domesticada. Durante este proceso, se observó la aparición de síntomas de un conspicuo mosaico amarillo, típico de infección viral. Análisis biológicos, serológicos y moleculares (RT-PCR, RFLP, clonado y secuenciación pusieron de manifiesto la presencia del subgrupo IA de Cucumber mosaic virus en las plantas domesticadas de peperina. El aislamiento viral estudiado está íntimamente relacionado con la raza Y previamente informada en Japón. Éste es el primer informe de un virus que infecta a la peperina.
Ruth M. Castro
Full Text Available Leaf samples of Solanum lycopersicum, Capsicum annuum, Cucurbita moschata, Cucurbita pepo, Sechium edule and Erythrina spp. were collected. All samples were positive for begomoviruses using polymerase chain reaction and degenerate primers. A sequence of ∼1,100 bp was obtained from the genomic component DNA-A of 14 samples. In addition, one sequence of ∼580 bp corresponding to the coat protein (AV1 was obtained from a chayote (S. edule leaf sample. The presence of Squash yellow mild mottle virus (SYMMoV and Pepper golden mosaic virus (PepGMV were confirmed. The host range reported for SYMMoV includes species of the Cucurbitaceae, Caricaceae and Fabaceae families. This report extends the host range of SYMMoV to include the Solanaceae family, and extends the host range of PepGMV to include C. moschata, C. pepo and the Fabaceae Erythrina spp. This is the first report of a begomovirus (PepGMV infecting chayote in the Western Hemisphere.
Castro, Ruth M; Moreira, Lisela; Rojas, María R; Gilbertson, Robert L; Hernández, Eduardo; Mora, Floribeth; Ramírez, Pilar
Leaf samples of Solanum lycopersicum, Capsicum annuum, Cucurbita moschata, Cucurbita pepo, Sechium edule and Erythrina spp. were collected. All samples were positive for begomoviruses using polymerase chain reaction and degenerate primers. A sequence of ∼1,100 bp was obtained from the genomic component DNA-A of 14 samples. In addition, one sequence of ∼580 bp corresponding to the coat protein (AV1) was obtained from a chayote (S. edule) leaf sample. The presence of Squash yellow mild mottle virus (SYMMoV) and Pepper golden mosaic virus (PepGMV) were confirmed. The host range reported for SYMMoV includes species of the Cucurbitaceae, Caricaceae and Fabaceae families. This report extends the host range of SYMMoV to include the Solanaceae family, and extends the host range of PepGMV to include C. moschata, C. pepo and the Fabaceae Erythrina spp. This is the first report of a begomovirus (PepGMV) infecting chayote in the Western Hemisphere.
Ability of Aphis gossypii and Myzus persicae to Transmit Cucumber mosaic virus in Single and Mixed Infection with Two Potyviruses to Zucchini Squash Eficiência dos afídeos Aphis gossypii e Myzus persicae na transmissão do Cucumber mosaic virus em infecção simples e mista com dois Potyvirus para abobrinha de moita
Zayame Vegette Pinto; Jorge Alberto Marques Rezende; Valdir Atsushi Yuki; Sônia Maria de Stefano Piedade
The main objective of this work was to investigate the ability of Aphis gossypii and Myzus persicae to transmit Cucumber mosaic virus (CMV) singly and mixed with two potyviruses (Papaya ringspot virus - type W, PRSV-W and Zucchini yellow mosaic virus, ZYMV), to zucchini squash plants (Cucurbita pepo). The results showed that the potyviruses in general were more efficiently transmitted by both species of aphids as compared to CMV. The transmission of PRSV-W, ZYMV and CMV separately was more ef...
HAN He-ping; SUN Ri-fei; ZHANG Shu-jiang; LI Fei; ZHANG Shi-fan; NIU Xin-ke
Turnip mosaic virus (TuMV) which has several strains causes the most important virusdisease in Chinese cabbage in terms of crop damage. In China, Chinese cabbage is infectedby a mixture of strains, breeding of cultivar for the TuMV resistance has become themajor aim. Screening the molecular marker linked to the TuMV-resistance gene formolecular assisted selection is the major method to improve the breeding efficiency. Inthis study, we used AFLP technique and the method of bulked segregant analysis(BSA) tostudy the progeny of Brp0058 x Brp0108, and identified two DNA molecular marker linked toTurnip mosaic virus-resistance gene with a recombination frequency 7.5 cM and 8.4 cM.
Morin, Hélène; Tremblay, Marie-Hélène; Plante, Edith; Paré, Christine; Majeau, Nathalie; Hogue, Richard; Leclerc, Denis
Papaya mosaic virus (PapMV) like particles (VLPs) were used as a platform for fusion of affinity peptides binding to resting spores of Plasmodiophora brassicae-a major pathogen of crucifers. Three peptides with specific affinity to the target were isolated and cloned at the C-terminus of the PapMV coat protein (CP), generating three different high avidity VLPs. The peptides were exposed at the surface of the VLPs and their avidity to resting spores of P. brassicae was measured by flow cytometry. NLP-A, with the peptide DPAPRPR, showed the highest avidity. The binding avidity of NLP-A to P. brassicae spores was comparable to that of a polyclonal antibody. NLP-A was also shown to be more specific than the antibody. Fusion of the affinity peptide to a monomeric form (mCP) of the CP [Lecours, K., Tremblay, M.-H., Laliberté Gagné, M.-E., Gagné, S.M., Leclerc, D., 2006. Purification and biochemical characterization of a monomeric form of papaya mosaic potexvirus coat protein. Protein Express. Purific. 47, 273-280] generated a fusion protein that was unable to assemble into VLPs, and mCP-A fusions failed to bind resting spores. The avidity of VLP-A was increased by adding a glycine spacer between the C-terminus of the PapMV CP and the peptide, and improved even further by using a duplicated A peptide in the fusion protein. The use of high avidity VLPs has advantages over polyclonal antibodies because of target specificity. VLPs offers the specificity of monoclonal antibodies but can be more easily generated using the powerful selection of phage display.
Full Text Available There is increasing interest in the immune response induced by plant viruses since these could be used as antigen-expressing systems in vaccination procedures. Cowpea severe mosaic virus (CPSMV, as a purified preparation (300 g of leaves, 2 weeks post-inoculation, or crude extract from cowpea (Vigna unguiculata leaves infected with CPSMV both administered by gavage to Swiss mice induced a humoral immune response. Groups of 10 Swiss mice (2-month-old females were immunized orally with 10 daily doses of either 50 µg viral capsid protein (boosters of 50 µg at days 21 and 35 after immunization or 0.6 mg protein of the crude extract (boosters of 0.6 mg at days 21 and 35 after immunization. Anti-CPSMV antibodies were quantified by ELISA in pooled sera diluted at least 1:400 at days 7, 14, 21, 28, 35 and 42 after the 10th dose. IgG and IgA against CPSMV were produced systemically, but IgE was not detected. No synthesis of specific antibodies against the proteins of leaf extracts from V. unguiculata, infected or not with CPSMV, was detected. The use of CPSMV, a plant-infecting virus that apparently does not induce a pathogenic response in animals, induced a humoral and persistent (at least 6 months immune response through the administration of low antigen doses by gavage. These results raise the possibility of using CPSMV either as a vector for the production of vaccines against animal pathogens or in quick and easy methods to produce specific antisera for viral diagnosis.
Rao, Li-Xia; Guo, Yushuang; Zhang, Li-Li; Zhou, Xue-Ping; Hong, Jian; Wu, Jian-Xiang
Cucumber green mottle mosaic virus (CGMMV) is a single-stranded, positive sense RNA virus infecting cucurbitaceous plants. In recent years, CGMMV has become an important pathogen of cucurbitaceous crops including watermelon, pumpkin, cucumber and bottle gourd in China, causing serious losses to their production. In this study, we surveyed CGMMV infection in various cucurbitaceous crops grown in Zhejiang Province and in several seed lots purchased from local stores with the dot enzyme-linked immunosorbent assay (dot-ELISA), using a CGMMV specific monoclonal antibody. Seven CGMMV isolates obtained from watermelon, grafted watermelon or oriental melon samples were cloned and sequenced. Identity analysis showed that the nucleotide identities of the seven complete genome sequences ranged from 99.2 to 100%. Phylogenetic analysis of seven CGMMV isolates as well as 24 other CGMMV isolates from the GenBank database showed that all CGMMV isolates could be grouped into two distinct monophyletic clades according to geographic distribution, i.e. Asian isolates for subtype I and European isolates for subtype II, indicating that population diversification of CGMMV isolates may be affected by geographical distribution. Site variation rate analysis of CGMMV found that the overall variation rate was below 8% and mainly ranged from 2 to 5%, indicating that the CGMMV genomic sequence was conservative. Base substitution type analysis of CGMMV showed a mutational bias, with more transitions (A↔G and C↔T) than transversions (A↔C, A↔T, G↔C and G↔T). Most of the variation occurring in the CGMMV genome resulted in non-synonymous substitutions, and the variation rate of some sites was higher than 30% because of this mutational bias. Selection constraint analysis of CGMMV ORFs showed strong negative selection acting on the replication-associated protein, similar to what occurs for other plant RNA viruses. Finally, potential recombination analysis identified isolate Ec as a
Parente, A; Short, M N; Self, R; Parsley, K R
Eleven of the fourteen tryptic peptides of papaya mosaic virus protein have been sequenced by electron ionization mass spectrometry using chemical and enzymic hydrolyses and mixture analysis as required. Mid-chain cleavages of N-C bonds produced secondary ion series which allowed up to 16 residues to be sequenced without further hydrolysis. Mixture analysis on hydrolysis products enabled a 24 residue tryptic peptide to be sequenced from the data recorded in a single mass spectrum.
Kaido, M; Mori, M; Mise, K; Okuno, T; Furusawa, I
Transgenic tobacco plants (V123 plants) expressing a set of full-length brome mosaic virus (BMV) genomic RNAs from the cauliflower mosaic virus 35S promoter were produced. The accumulation level of BMV RNAs in V123 plant cells was approximately 1% of that in nontransgenic tobacco protoplasts inoculated with BMV RNAs. The level of BMV RNA in V123 protoplasts did not increase after inoculating the protoplasts with BMV RNAs, whereas V123 protoplasts supported the accumulation of cucumber mosaic virus (CMV) RNAs to a level similar to that in non-transgenic tobacco protoplasts after inoculation with CMV RNA. Such BMV-specific resistance was also observed in protoplasts from V12 plants expressing full-length BMV RNA1 and RNA2, both of which are required and sufficient for BMV RNA replication. On the other hand, protoplasts from M12 plants, expressing truncated BMV RNA1 and RNA2 in which the 3' 200 nucleotides required for BMV RNA replication were deleted, exhibited weaker resistance to infection with BMV RNA than V12 protoplasts, although the accumulation level of truncated BMV RNA1 and RNA2 in M12 protoplasts was higher than that of BMV RNA1 and RNA2 in V12 protoplasts. These results suggest that expression of BMV RNA replicons is involved in the induction of resistance, rather than high-level accumulation of BMV RNAs and/or their encoded proteins.
Full Text Available The ND18 strain of Barley stripe mosaic virus (BSMV infects several lines of Brachypodium distachyon, a recently developed model system for genomics research in cereals. Among the inbred lines tested, Bd3-1 is highly resistant at 20 to 25 °C, whereas Bd21 is susceptible and infection results in an intense mosaic phenotype accompanied by high levels of replicating virus. We generated an F(6:7 recombinant inbred line (RIL population from a cross between Bd3-1 and Bd21 and used the RILs, and an F(2 population of a second Bd21 × Bd3-1 cross to evaluate the inheritance of resistance. The results indicate that resistance segregates as expected for a single dominant gene, which we have designated Barley stripe mosaic virus resistance 1 (Bsr1. We constructed a genetic linkage map of the RIL population using SNP markers to map this gene to within 705 Kb of the distal end of the top of chromosome 3. Additional CAPS and Indel markers were used to fine map Bsr1 to a 23 Kb interval containing five putative genes. Our study demonstrates the power of using RILs to rapidly map the genetic determinants of BSMV resistance in Brachypodium. Moreover, the RILs and their associated genetic map, when combined with the complete genomic sequence of Brachypodium, provide new resources for genetic analyses of many other traits.
Full Text Available ZYMV is one of the most important plant viruses that cause economical damage in cucurbits. The symptoms of ZYMV in different cucurbits include stunting, yellowing, mottling, severe mosaic, leaf and fruit deformation, blistering and shoe string. Investigation on occurrence of this virus, in Khuzestan province was carried out in November 2009, April and May 2010 by collecting cucurbits samples from different cucurbits fields. After DAS-ELISA test, ZYMV was maintained in squash. Then total RNA were extracted and were tested by RT-PCR. Using RT-PCR, fragments belonging to N-terminal of coat protein and C-terminal of nuclear inclusion bodies were replicated. PCR product for investigation of replication was loaded in 1% agarose gel. From seven regions in Khuzestan, 175 leaf samples showing different symptoms (yellowing, mosaic, deformation and blistering were collected. Seventy one samples out of total samples (175 samples showed ZYMV infection. Occurrence of Zucchini Yellow Mosaic Virus in Khuzestan province was confirmed, using serological and RT-PCR tests. Infection of ZYMV in Khuzestan province (40.5% is higher than the average of Iran’s infection (38%. This article is first report of occurrence ZYMV in different regions of Khuzestan province except Dezful.
Full Text Available During antagonistic coevolution between viruses and their hosts, viruses have a major advantage by evolving more rapidly. Nevertheless, viruses and their hosts coexist and have coevolved, although the processes remain largely unknown. We previously identified Tm-1 that confers resistance to Tomato mosaic virus (ToMV, and revealed that it encodes a protein that binds ToMV replication proteins and inhibits RNA replication. Tm-1 was introgressed from a wild tomato species Solanum habrochaites into the cultivated tomato species Solanum lycopersicum. In this study, we analyzed Tm-1 alleles in S. habrochaites. Although most part of this gene was under purifying selection, a cluster of nonsynonymous substitutions in a small region important for inhibitory activity was identified, suggesting that the region is under positive selection. We then examined the resistance of S. habrochaites plants to ToMV. Approximately 60% of 149 individuals from 24 accessions were resistant to ToMV, while the others accumulated detectable levels of coat protein after inoculation. Unexpectedly, many S. habrochaites plants were observed in which even multiplication of the Tm-1-resistance-breaking ToMV mutant LT1 was inhibited. An amino acid change in the positively selected region of the Tm-1 protein was responsible for the inhibition of LT1 multiplication. This amino acid change allowed Tm-1 to bind LT1 replication proteins without losing the ability to bind replication proteins of wild-type ToMV. The antiviral spectra and biochemical properties suggest that Tm-1 has evolved by changing the strengths of its inhibitory activity rather than diversifying the recognition spectra. In the LT1-resistant S. habrochaites plants inoculated with LT1, mutant viruses emerged whose multiplication was not inhibited by the Tm-1 allele that confers resistance to LT1. However, the resistance-breaking mutants were less competitive than the parental strains in the absence of Tm-1. Based on
Krupovic, Mart; Forterre, Patrick; Bamford, Dennis H
Tailed double-stranded DNA viruses (order Caudovirales) represent the dominant morphotype among viruses infecting bacteria. Analysis and comparison of complete genome sequences of tailed bacterial viruses provided insights into their origin and evolution. Structural and genomic studies have unexpectedly revealed that tailed bacterial viruses are evolutionarily related to eukaryotic herpesviruses. Organisms from the third domain of life, Archaea, are also infected by viruses that, in their overall morphology, resemble tailed viruses of bacteria. However, high-resolution structural information is currently unavailable for any of these viruses, and only a few complete genomes have been sequenced so far. Here we identified nine proviruses that are clearly related to tailed bacterial viruses and integrated into chromosomes of species belonging to four different taxonomic orders of the Archaea. This more than doubled the number of genome sequences available for comparative studies. Our analyses indicate that highly mosaic tailed archaeal virus genomes evolve by homologous and illegitimate recombination with genomes of other viruses, by diversification, and by acquisition of cellular genes. Comparative genomics of these viruses and related proviruses revealed a set of conserved genes encoding putative proteins similar to virion assembly and maturation, as well as genome packaging proteins of tailed bacterial viruses and herpesviruses. Furthermore, fold prediction and structural modeling experiments suggest that the major capsid proteins of tailed archaeal viruses adopt the same topology as the corresponding proteins of tailed bacterial viruses and eukaryotic herpesviruses. Data presented in this study strongly support the hypothesis that tailed viruses infecting archaea share a common ancestry with tailed bacterial viruses and herpesviruses.
Garcia-Ruiz, Hernan; Carbonell, Alberto; Hoyer, J Steen; Fahlgren, Noah; Gilbert, Kerrigan B; Takeda, Atsushi; Giampetruzzi, Annalisa; Garcia Ruiz, Mayra T; McGinn, Michaela G; Lowery, Nicholas; Martinez Baladejo, Maria T; Carrington, James C
In eukaryotes, ARGONAUTE proteins (AGOs) associate with microRNAs (miRNAs), short interfering RNAs (siRNAs), and other classes of small RNAs to regulate target RNA or target loci. Viral infection in plants induces a potent and highly specific antiviral RNA silencing response characterized by the formation of virus-derived siRNAs. Arabidopsis thaliana has ten AGO genes of which AGO1, AGO2, and AGO7 have been shown to play roles in antiviral defense. A genetic analysis was used to identify and characterize the roles of AGO proteins in antiviral defense against Turnip mosaic virus (TuMV) in Arabidopsis. AGO1, AGO2 and AGO10 promoted anti-TuMV defense in a modular way in various organs, with AGO2 providing a prominent antiviral role in leaves. AGO5, AGO7 and AGO10 had minor effects in leaves. AGO1 and AGO10 had overlapping antiviral functions in inflorescence tissues after systemic movement of the virus, although the roles of AGO1 and AGO10 accounted for only a minor amount of the overall antiviral activity. By combining AGO protein immunoprecipitation with high-throughput sequencing of associated small RNAs, AGO2, AGO10, and to a lesser extent AGO1 were shown to associate with siRNAs derived from silencing suppressor (HC-Pro)-deficient TuMV-AS9, but not with siRNAs derived from wild-type TuMV. Co-immunoprecipitation and small RNA sequencing revealed that viral siRNAs broadly associated with wild-type HC-Pro during TuMV infection. These results support the hypothesis that suppression of antiviral silencing during TuMV infection, at least in part, occurs through sequestration of virus-derived siRNAs away from antiviral AGO proteins by HC-Pro. These findings indicate that distinct AGO proteins function as antiviral modules, and provide a molecular explanation for the silencing suppressor activity of HC-Pro.
Full Text Available A Cucumber green mottle mosaic virus (CGMMV was used to present a truncated dengue virus type 2 envelope (E protein binding region from amino acids 379 to 423 (EB4. The EB4 gene was inserted at the terminal end of the CGMMV coat protein (CP open reading frame (ORF. Read-through sequences of TMV or CGMMV, CAA-UAG-CAA-UUA, or AAA-UAG-CAA-UUA were, respectively, inserted in between the CP and the EB4 genes. The chimeric clones, pRT, pRG, and pCG+FSRTRE, were transcribed into full-length capped recombinant CGMMV transcripts. Only constructs with the wild-type CGMMV read-through sequence yielded infectious viruses following infection of host plant, muskmelon (Cucumis melo leaves. The ratio of modified to unmodified CP for the read-through expression clone developed was also found to be approximately 1:1, higher than what has been previously reported. It was also observed that infectivity was not affected by differences in pI between the chimera and its wild counterpart. Analysis of recombinant viruses after 21-days-postinculation (dpi revealed that deletions occurred resulting in partial reversions of the viral population to near wild type and suggesting that this would be the limiting harvest period for obtaining true to type recombinants with this construct.
Teoh, Pak-Guan; Ooi, Aik-Seng; AbuBakar, Sazaly; Othman, Rofina Yasmin
A Cucumber green mottle mosaic virus (CGMMV) was used to present a truncated dengue virus type 2 envelope (E) protein binding region from amino acids 379 to 423 (EB4). The EB4 gene was inserted at the terminal end of the CGMMV coat protein (CP) open reading frame (ORF). Read-through sequences of TMV or CGMMV, CAA-UAG-CAA-UUA, or AAA-UAG-CAA-UUA were, respectively, inserted in between the CP and the EB4 genes. The chimeric clones, pRT, pRG, and pCG+FSRTRE, were transcribed into full-length capped recombinant CGMMV transcripts. Only constructs with the wild-type CGMMV read-through sequence yielded infectious viruses following infection of host plant, muskmelon (Cucumis melo) leaves. The ratio of modified to unmodified CP for the read-through expression clone developed was also found to be approximately 1:1, higher than what has been previously reported. It was also observed that infectivity was not affected by differences in pI between the chimera and its wild counterpart. Analysis of recombinant viruses after 21-days-postinculation (dpi) revealed that deletions occurred resulting in partial reversions of the viral population to near wild type and suggesting that this would be the limiting harvest period for obtaining true to type recombinants with this construct.
Ding, Xin Shun; Schneider, William L; Chaluvadi, Srinivasa Rao; Mian, M A Rouf; Nelson, Richard S
Virus-induced gene silencing (VIGS) is used to analyze gene function in dicotyledonous plants but less so in monocotyledonous plants (particularly rice and corn), partially due to the limited number of virus expression vectors available. Here, we report the cloning and modification for VIGS of a virus from Festuca arundinacea Schreb. (tall fescue) that caused systemic mosaic symptoms on barley, rice, and a specific cultivar of maize (Va35) under greenhouse conditions. Through sequencing, the virus was determined to be a strain of Brome mosaic virus (BMV). The virus was named F-BMV (F for Festuca), and genetic determinants that controlled the systemic infection of rice were mapped to RNAs 1 and 2 of the tripartite genome. cDNA from RNA 3 of the Russian strain of BMV (R-BMV) was modified to accept inserts from foreign genes. Coinoculation of RNAs 1 and 2 from F-BMV and RNA 3 from R-BMV expressing a portion of a plant gene to leaves of barley, rice, and maize plants resulted in visual silencing-like phenotypes. The visual phenotypes were correlated with decreased target host transcript levels in the corresponding leaves. The VIGS visual phenotype varied from maintained during silencing of actin 1 transcript expression to transient with incomplete penetration through affected tissue during silencing of phytoene desaturase expression. F-BMV RNA 3 was modified to allow greater accumulation of virus while minimizing virus pathogenicity. The modified vector C-BMV(A/G) (C for chimeric) was shown to be useful for VIGS. These BMV vectors will be useful for analysis of gene function in rice and maize for which no VIGS system is reported.
AKM Mahbubul Alam
Full Text Available This work was conducted to identify mungbean genotypes showing yield stability and resistance to mungbean yellow mosaic virus (MYMV disease. Sixteen genotypes were evaluated in a randomized complete block design with two replications for two years (2011 and 2012 at three locations (Gazipur, Ishurdi and Madaripur of the Bangladesh Agricultural Research Institute. An analysis of variance exhibited significant effects of genotype (G, environment (E, and genotype × environment (G×E on grain yield. Among eight agronomic characters, the principal component 1 (PC1 was always higher than the PC2. Considering G×E interaction, BM6 was the best genotype at all three locations in both years. Based on grain yield and stability performance, BM6 ranked first while the worst performing genotypes were BM1 and G10. Based on discrimination and representation, Gazipur was identified as an ideal environment for these mungbeans. Relationship between soil-plant analysis developments (SPAD value was positive with yield but negative with MYMV severity. BM6, G1 and G2 were considered as promising sources of resistance for low disease score and stable response across the environments. The environment proved to have an influence on MYMV infection under natural infestation. A positive correlation was observed between disease score and the temperature under natural growing condition.
Alam, Akm Mahbubul; Somta, Prakit; Jompuk, Choosak; Chatwachirawong, Prasert; Srinives, Peerasak
This work was conducted to identify mungbean genotypes showing yield stability and resistance to mungbean yellow mosaic virus (MYMV) disease. Sixteen genotypes were evaluated in a randomized complete block design with two replications for two years (2011 and 2012) at three locations (Gazipur, Ishurdi and Madaripur) of the Bangladesh Agricultural Research Institute. An analysis of variance exhibited significant effects of genotype (G), environment (E), and genotype × environment (G×E) on grain yield. Among eight agronomic characters, the principal component 1 (PC1) was always higher than the PC2. Considering G×E interaction, BM6 was the best genotype at all three locations in both years. Based on grain yield and stability performance, BM6 ranked first while the worst performing genotypes were BM1 and G10. Based on discrimination and representation, Gazipur was identified as an ideal environment for these mungbeans. Relationship between soil-plant analysis developments (SPAD) value was positive with yield but negative with MYMV severity. BM6, G1 and G2 were considered as promising sources of resistance for low disease score and stable response across the environments. The environment proved to have an influence on MYMV infection under natural infestation. A positive correlation was observed between disease score and the temperature under natural growing condition.
SandraPérezAlvarez; 薛朝阳; 周雪平
The cucumber mosaic virus (CMV) isolate P1 caused very mild symptoms on many plant species.After serial passages by mechanical inoculation over five years, CMV P1 caused severe symptoms on several tobacco cultivars and tomato. A specific band of approximately 0.3 kb in length was amplified by RT-PCR with primers synthesized based on reported CMV satellite RNA (satRNA) sequences. Sequence analysis showed there were two satRNAs (Sat-Pl-1 and Sat-P1-2). Sat-Pl-1 contained 335 nucleotides, and Sat-P1-2 contained 394 nucleotides. These two satRNAs shared 64% overall nucleotide sequence homology, and differences between the two satRNAs included mutations as well as deletions. Sat-Pl-1 was identical to a satRNA (Z96099) reported in 1995 in CMV P1. Based on differences in the sequence and secondary structure between these two satRNAs, we conclude that Sat-P1-2 represents the emergence of a new satellite ( necrotic satellite) from attenuated satRNA populations. The possible effect of the emergence of this new satRNA is discussed.
The cucumber mosaic virus (CMV) isolate P1 caused very mild symptoms on many plant species. After serial passages by mechanical inoculation over five years, CMV P1 caused severe symptoms on several tobacco cultivars and tomato. A specific band of approximately 0.3 kb in length was amplified by RT-PCR with primers synthesized based on reported CMV satellite RNA (satRNA) sequences. Sequence analysis showed there were two satRNAs (Sat-P1-1 and Sat-P1-2). Sat-P1-1 contained 335 nucleotides, and Sat-P1-2 contained 394 nucleotides. These two satRNAs shared 64% overall nucleotide sequence homology, and differences between the two satRNAs included mutations as well as deletions. Sat-P1-1 was identical to a satRNA (Z96099) reported in 1995 in CMV P1. Based on differences in the sequence and secondary structure between these two satRNAs, we conclude that Sat-P1-2 represents the emergence of a new satellite (necrotic satellite) from attenuated satRNA populations. The possible effect of the emergence of this new satRNA is discussed.
Bink, Hugo H J; Schirawski, Jan; Haenni, Anne-Lise; Pleij, Cornelis W A
The RNA genome of turnip yellow mosaic virus (TYMV) consists of more than 6,000 nucleotides. During a study of the roles of the two hairpins located in its 90-nucleotide 5' untranslated region, it was observed that stabilization of the 5'-proximal hairpin leads to a delay in the development of symptoms on plants. This delay in symptom development for both locally and systemically infected leaves was found to be dependent on a change in the free energy of the hairpin caused by introduced mutations. A protoplast transfection assay revealed that the accumulation of plus-strand full-length RNA and subgenomic RNA, as well as protein expression levels, was affected by hairpin stability. Stabilization of this hairpin inhibited translation. A model is proposed in which a destabilized 5'-proximal hairpin allows maximal translation of the viral proteins. It is suggested that this hairpin may exist in close proximity to the 5' cap as long as its stability is low enough to enable translation. However, at an acidic pH, the hairpin structure becomes more stable and is functionally transformed into the initiation signal for viral packaging. Slightly acidic conditions can be found in chloroplasts, where TYMV assembly is driven by a low pH generated by active photosynthesis.
Kundu, Subrata; Chakraborty, Dipjyoti; Pal, Amita
The role of salicylic acid (SA) in inducing resistance to MYMIV infection in Vigna mungo has been elucidated by proteomics. Twenty-nine proteins identified by MALDI-TOF/TOF, predicted to be involved in stress responses, metabolism, photosynthesis, transport and signal transduction, showed increased abundance upon SA treatment. Susceptible plants showed characteristic yellow mosaic symptoms upon MYMIV infection. A concentration dependent decrease in physiological symptoms associated with MYMIV was observed upon exogenous SA treatment prior to viral inoculation; and no visible symptom was observed at 100 μM SA. SA treatment stimulated SOD and GPX activity and inhibited CAT activity thus preventing ROS mediated damage. Significant increase in chlorophyll, protein, carbohydrate, phenolic content and H(2)O(2) were observed. Involvement of calmodulin for transmission of defense signal by SA is suggested. A metabolic reprogramming leading to enhanced synthesis of proteins involved in primary and secondary metabolisms is necessary for SA mediated resistance to MYMIV. Identification of proteins showing increased abundance, involved in photosynthetic process is a significant finding which restores virus-induced degradation of the photosynthetic apparatus and provides enhanced metabolites required for repartition of resources towards defense.
The promoter region of cauliflower mosaic virus (CaMV) 35s RNA was employed to construct an intermediate expression vector which can be used in Ti plasmid system of Agrobacterium iumefaciens.The original plasmid,which contains a polylinker between CaMV 35s RNA and its 3' termination signal in pUC18 was modified to have another antibiotic resistance marker (kanamycin resistance gene Kmr) to facilitate the selection of recombinant with Ti plasmid.Octopine synthase (ocs) structural gene was inserted into this vector downstream of CaMV 35s RNA promoter.This chimaeric gene was introduced into integrative Ti plasmid vector pGV 3850,and then transformed into Nicotiana tobaccum the chimaeric gene into tobacco cells.In both cases,the expression of ocs gene was demonstrated.The amount of octopine was much more than the nopaline synthesized by nopaline synthase (nos) gene transferred at the same time with Ti plasmid vector.This demonstrated that CaMV 35s RNA promoter is stronger in transcriptional function than the promoter of nos in tobacco cells.
ZHI Hai-jian; GAI Jun-yi
A sample composed of 96 soybean accessions was evaluated for their diseased rate (I),diseased rank (S), latent period (LP) and rate of disease development (R) in order tostudy the quantitative resistance to soybean mosaic virus (SMV) in soybeans. The resultsshowed that the performances of the above four resistance components were significantlydifferent among accessions and that some of the accessions, such as Zhongzihuangdou,Peixian Tianedan, Youbian30 could be infected by four SMV strains, Sa, SC8, N1 and N3,but their I, S, and R were lower and LP longer than most other accessions. These resultsdemonstrated the existence of quantitative resistance to SMV in soybeans. It was foundthat some soybean accessions, such as AGS19 and Lishui Zhongzihuangdou, previouslyidentified as resistant to SMV infection, performed some infection but resistant toexpansion in the present study. In addition, the resistance in Pixian Chadou and HuaiyinQiuheidou might be either qualitative or quantitative. Furthermore, the present studyalso indicated that the resistance spectrum and durability of accessions with quantitativeresistance might be wider and longer than those with qualitative resistance.
Kitayama, Masahiko; Hoover, Haley; Middleton, Stefani; Kao, C Cheng
Brome mosaic virus (BMV) (the Russian strain) infects monocot plants and has been studied extensively in barley and wheat. Here, we report BMV can systemically infect rice (Oryza sativa var. japonica), including cultivars in which the genomes have been determined. The BMV capsid protein can be found throughout the inoculated plants. However, infection in rice exhibits delayed symptom expression or no symptoms when compared with wheat (Triticum aestivum). The sequences of BMV RNAs isolated from rice did not reveal any nucleotide changes in RNA1 or RNA2, while RNA3 had only one synonymous nucleotide change from the inoculum sequence. Preparations of purified BMV virions contained RNA1 at a significantly reduced level relative to the other two RNAs. Analysis of BMV RNA replication in rice revealed that minus-strand RNA1 was replicated at a reduced rate when compared with RNA2. Thus, rice appears to either inhibit RNA1 replication or lacks a sufficient amount of a factor needed to support efficient RNA1 replication.
Safaeizadeh, M; Saidi, A; Palukaitis, P
Here we identified four isolates, MS, 3H, 50A, and 2K of cucumber mosaic virus (CMV) infecting tomato, on the basis of their non-coding intergenic region and a part of the coat protein (CP) sequence in the CMV genomic RNA3. The sequences from the four isolates were compared with other previously characterized isolates of CMV isolated from different plant species across the globe. Sequence comparisons revealed that the two CMV isolates from Hamedan province (MS and 3H) had the highest sequence identity with CMV-G10 (98%), which was previously reported as a severe Hellenic tomato isolate of CMV, while the CMV isolates from Tehran province, including CMV-2K (isolated from Karaj region) and CMV-50A (isolated from Varamin region), had the highest sequence identity with that of CMV-ALF (99%). Phylogenetic analysis of the nucleotide sequences showed that CMV-MS and CMV-3H belong to group IB, while CMV-2K and CMV-50A belong to group IA. This is the first report on the molecular characterization of novel isolates of CMV infecting tomato plants in Iran.
Shevchenko T. P.
Full Text Available Aim. In current work, we proceeded with the strain attribution of Ukrainian isolates CMV based on the phylogenetic analysis of the partial sequences of the coat protein gene. Methods. ELISA, RT-PCR, DNA sequencing and phylogenetic analysis. Results. Cucumber mosaic virus (CMV is widespread among the variety of crops from the Cucurbitaceae and Solanaceae families in Ukraine. The symptomatic samples from different regions of Ukraine were collected and tested for the presence of CMV. The coat protein (CP gene of two isolates was amplified and sequenced. The partial nucleotide sequences of CP gene were determined and compared to those of other CMV strains belonging to the IA, IB and II subgroups. Comparison of the nucleotide sequences of Ukrainian isolates showed their similar identity percentages and close relationships with the subgroup IB strains from other countries. The highest nucleotide homology was shared with the strains ABI (Korea and SD (China. Conclusions. Based on the highest identities of the coat protein gene sequences and close phylogenetic relationships with the subgroup IB members of CMV, the Ukrainian isolates under study were identified as belonging to the subgroup IB. Our findings show for the first time an occurrence of the IB subgroup isolates of CMV in Ukraine.
Sokhandan Bashir, N; Nematollahi, S; Torabi, E
To monitor genetic variation between Cucumber mosaic virus (CMV) isolates of northwest Iran, samples of cucurbitaceous plants expressing symptoms similar to those caused by CMV were collected. The samples were first screened by ELISA to detect CMV and to determine its subgroup. All detected CMV isolates appeared to be subgroup I (S-I). When total RNA from the samples was subjected to RT-PCR with a pair of primers corresponding to the CMV coat protein (CP) flanking regions, the expected ~870 bp DNA fragment was amplified at 18 samples of 34 tested. MspI restriction analysis of 18 amplified products produced two DNA fragments with sizes about 530 and 330 bp corresponding to MspI profile of CMV S-I. The amplification products of four representative samples were cloned and nucleotide sequences of 1-5 clones from each isolate were determined. The clones from each isolate were over 99% identical and also the isolates themselves were only up to 2% divergent. These isolates clustered in subgroup IA clade on a consensus phylogenetic tree and formed a distinct subclade suggesting that the isolates have originated from a common source.
Full Text Available Field survey of the cucurbit crops revealed a high incidence of Cucumber green mottle mosaic virus (CGMMV in Khyber Pakhtunkhwa Province (KPK, Pakistan. Among the seven districts surveyed, average percent incidence of CGMMV was recorded up to 58.1% in district Nowshera, followed by 51.1% in district Charsada, 40.5% in district Swabi and 37.3% in district Mardan. In Swat and Dir districts average incidence CGMMV was recorded upto 31.2% and 29.4%, respectively. Among the different crops highest incidence in plain areas of KPK was recorded in bottle gourd (59.3% followed by 56.3% in Squash, 54.5% in Pumpkin, 45.5% in Melon, 41.7% in Cucumber and 29.9% in Sponge gourd. In Northern hilly areas highest incidence of CGMMV (52.9% was observed in pumpkin, followed by 49.6% in bottle gourd, 47.3% in squash, 45.1% in Melon 42.3% in cucumber and 41.6% in sponge gourd. Little variability was observed in the coat protein amino acid sequence identities of CGMMV Pakistan isolate, when compared with other reported isolates.
Mohsen Mohamed Elsharkawy
Full Text Available Trichoderma asperellum SKT-1 is a microbial pesticide that is very effective against various diseases. Our study was undertaken to evaluate T. asperellum SKT-1 for induction of resistance against yellow strain of Cucumber mosaic virus (CMV-Y in Arabidopsis plants. Disease severity was rated at 2 weeks post inoculation (WPI. CMV titre in Arabidopsis leaves was determined by indirect enzyme-linked immunosorbent assay (ELISA at 2 WPI. Our results demonstrated that among all Arabidopsis plants treated with barley grain inoculum (BGI of SKT-1 NahG and npr1 plants showed no significant reduction in disease severity and CMV titre as compared with control plants. In contrast, disease severity and CMV titre were significantly reduced in all Arabidopsis plants treated with culture filtrate (CF of SKT-1 as compared with control plants. RT-PCR results showed increased expression levels of SA-inducible genes, but not JA/ET-inducible genes, in leaves of BGI treated plants. Moreover, expression levels of SA- and JA/ET-inducible genes were increased in leaves of CF treated plants. In conclusion, BGI treatment induced systemic resistance against CMV through SA signaling cascade in Arabidopsis plants. While, treatment with CF of SKT-1 mediated the expression of a majority of the various pathogen related genes, which led to the increased defense mechanism against CMV infection.
刘俊君; 彭学贤; 莽克强
cDNA of soybean mosaic virus (Beijing isolate, SMV-BJ) has been synthesized, using viralgenomic RNA as template and random hexanucleotides as primers. Based on the sequences of SMV-BJ coat protein (CP) gene as well as SMV- and WMV-II-related regions, oligonucleotides were made as primers for polymerase chain reaction (PCR). NIb gene of SMV-BJ was amplified by PCR, and cloned into pBluescript SK. The complete sequence was determined. The comparison of NIb genes between SMV-BJ and WMV-II . (USA) shows that similarities for nucleotide sequence reach 80.3%, and the deduced amino acid sequence. 91 3%. In consideration of the high identities in between the CP gene and the 3’-non-coding region between them, WMV-II might be considered as a watermelon strain of SMV Besides, some unexpected sequences were found in the 3’-region of 2 NIb gene clones. Following modification and splicing, a binary vector of NIb gene has been constructed for its expression in higher plant for the purpose of studying the possible repl
程宁辉; 方荣祥; 濮祖芹; 方中达
A new strain (TN strain) of cucumber mosaic virus (CMV) was isolated from tomato plants with necrotic symptoms and proved to carry a necrogenic satellite RNA (TN-Sat RNA). Double-strand cDNA of the TN-Sat RNA was synthesized by reverse transcription and polymerase chain reaction using primers designed according to the conserved terminal sequences of known CMV satellite RNAs. Sequence analysis indicated that the TN-Sat RNA consisted of 390 nucleotides (nt). Comparison of the sequence of the TN-Sat RNA with those of other CMV satellite RNAs revealed four homologous regions ( I . 1-81 nt; II . 216-261 nt; III. 278-338 nt; IV . 349-390 nt) and one hypervarible domain in the region of 82-215 nt. Moreover, the TN-Sat RNA contained a characteristic necro-genic consensus sequence at the 3’ end (339-367 nt) as reported in the known necrosis-inducing CMV satellite RNAs.
Xiangling L(U); Xinhai LI; Chuanxiao XIE; Zhuanfang HAO; Hailian JI; Liyu SHI; Shihuang ZHANG
The development of genomics and bioinfor-matics offers new tools for comparative gene mapping. In this paper, an integrated QTL map for sugarcane mosaic virus (SCMV) resistance in maize was constructed by compiling a total of 81 QTL loci available, using the Genetic Map IBM2 2005 Neighbors as reference. These 81 QTL loci were scattered on 7 chromosomes of maize, and most of them were clustered on chromosomes 3 and 6. By using the method of meta-analysis, we identified one "consensus QTL" on chromosome 3 covering a genetic distance of 6.44 cM, and two on chromosome 6 covering genetic distances of 16 cM and 27.48 cM, respectively. Four positional candidate resistant genes were identified within the "consensus QTL" on chromosome 3 via the strategy of comparative genomics. These results suggest that application of a combination of meta-analysis within a species with sequence homology comparison in a related model plant is an efficient approach to identify the major QTL and its candidate gene(s) for the target traits. The results of this study provide useful information for iden-tifying and cloning the major gene(s) conferring resistance to SCMV in maize.
A Rouhibakhsh; Q M I Haq; V G Malathi
Mungbean yellow mosaic India virus (MYMIV) is a whitefly-transmitted begomovirus with a bipartite genome. We investigate the functions of the MYMIV-AV2 protein, the open reading frame present upstream of the coat protein gene in DNA A component. The ability of MYMIV-AV2 mutants to replicate, spread and cause symptoms in legume hosts, blackgram, cowpea and French bean was analysed. Plants agroinoculated with mutants K73R, C86S and the double mutant C84S, C86S showed increase in severity of symptoms compared with the wild type. However, mutants W2S and H14Q,G15E caused marked attenuation of symptoms. While the double mutants C84S,C86S caused a 50-fold increase in double-stranded supercoiled and single-stranded DNA accumulation, the mutations W2S and H14Q,G15E showed a decrease in double-stranded supercoiled and single-stranded viral DNA accumulation. Because AV2 mutants affect the ratio between open circular and supercoiled DNA forms, we hypothesize that these mutations may modulate the functions of the replication initiation protein.
Lim, S H; Ko, M K; Lee, S J; La, Y J; Kim, B D
The nucleotide sequence of the 3'-terminal region of the Korean isolate of cymbidium mosaic virus (CyMV-Ca) from a naturally infected cattleya was determined. The sequence contains an open reading frame (ORF) coding for the viral coat protein (CP) at the 3'-end and three other ORFs (triple gene block or movement protein) of CyMV. The CP gene encodes a polypeptide chain of 220 amino acids with a molecular mass of 23,760 Da. The deduced CP sequence showed a strong homology with those of two CyMVs reported. A construct of the CyMV-Ca CP gene in the antisense orientation in the plant expression vector pMBP1 was transferred via Agrobacterium tumefaciens-mediated transformation into Nicotiana occidentalis which is a propagation host of CyMV. The T1 progeny of the transgenic plants were inoculated with CyMV and found to be highly resistant to CyMV infection.
Full Text Available 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.
Trucco, Verónica; de Breuil, Soledad; Bejerman, Nicolás; Lenardon, Sergio; Giolitti, Fabián
The complete nucleotide sequence of an Alfalfa mosaic virus (AMV) isolate infecting alfalfa (Medicago sativa L.) in Argentina, AMV-Arg, was determined. The virus genome has the typical organization described for AMV, and comprises 3,643, 2,593, and 2,038 nucleotides for RNA1, 2 and 3, respectively. The whole genome sequence and each encoding region were compared with those of other four isolates that have been completely sequenced from China, Italy, Spain and USA. The nucleotide identity percentages ranged from 95.9 to 99.1 % for the three RNAs and from 93.7 to 99 % for the protein 1 (P1), protein 2 (P2), movement protein and coat protein (CP) encoding regions, whereas the amino acid identity percentages of these proteins ranged from 93.4 to 99.5 %, the lowest value corresponding to P2. CP sequences of AMV-Arg were compared with those of other 25 available isolates, and the phylogenetic analysis based on the CP gene was carried out. The highest percentage of nucleotide sequence identity of the CP gene was 98.3 % with a Chinese isolate and 98.6 % at the amino acid level with four isolates, two from Italy, one from Brazil and the remaining one from China. The phylogenetic analysis showed that AMV-Arg is closely related to subgroup I of AMV isolates. To our knowledge, this is the first report of a complete nucleotide sequence of AMV from South America and the first worldwide report of complete nucleotide sequence of AMV isolated from alfalfa as natural host.
Full Text Available Abstract Background Cucumber green mottle mosaic virus (CGMMV, a member of the genus Tobamovirus, can be transmitted by seeds and infects many cucurbit species, causing serious yield losses in cucumber and watermelon plants. In this paper, five serological methods including antigen-coated plate enzyme-linked immunosorbent assay (ACP-ELISA, triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA, Dot-immunobinding assay (DBIA, direct tissue blot immunoassay (DTBIA and immunocapture reverse transcriptase polymerase chain reaction (IC-RT-PCR were described for detection and diagnosis of CGMMV. Results Using the purified CGMMV particles as immunogens, six murine monoclonal antibodies (MAbs were produced. Five serological methods were established using the MAb 4H1 and detection sensitivity was compared using purified preparations and infected-plant tissue extracts. The detection sensitivity of ACP-ELISA was 0.16 ng of purified CGMMV, whereas TAS-ELISA was more sensitive than ACP-ELISA with a minimum detection of 0.04 ng of purified CGMMV. The sensitivities of TAS-ELISA and DBIA were similar for detecting CGMMV in infected-plant tissue extracts, and were four times higher than ACP-ELISA. The IC-RT-PCR was the most sensitive method, which could detect as little as 0.1 pg of purified virus. The detection sensitivity of IC-RT-PCR for CGMMV-infected plant tissues was about 400 times higher than that of TAS-ELISA and DBIA. Conclusions The established ACP-ELISA, TAS-ELISA, DBIA and DTBIA are suitable for routine CGMMV detection of large-scale samples in the field survey, while IC-RT-PCR is more sensitive and suitable for acquiring information about the viral genome.
Zahid, Kiran; Zhao, Jian-Hua; Smith, Neil A; Schumann, Ulrike; Fang, Yuan-Yuan; Dennis, Elizabeth S; Zhang, Ren; Guo, Hui-Shan; Wang, Ming-Bo
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.
José Evando Aguiar Beserra Júnior
Full Text Available Foram avaliadas 20 linhagens de melancia, provenientes do cruzamento da cultivar comercial suscetível Crimson Sweet e da introdução PI 595201 resistente ao Watermelon mosaic virus (WMV e Papaya ringspot virus (PRSV-W. As linhagens, e os parentais foram inoculados com o WMV ou com o PRSV-W em casa-de-vegetação distintas. Aos 35 e 49 dias após a primeira inoculação (DAI, as plantas foram avaliadas por meio de uma escala de notas, em que 1 (ausência de sintomas a 5 (intenso mosaico e deformações foliares. Pelos resultados infere-se que, aos 35 DAI, as linhagens 1, 2 e 20 apresentaram resistência tanto para o WMV como para o PRSV-W, com médias de 1,95, 1,80 e 2,25 para o WMV, e de 2,50, 2,30 e 2,50 para o PRSV-W, respectivamente. As linhagens 5, 7 e 13 foram resistentes somente ao WMV e as plantas das linhagens 3, 10 e 18 para o PRSV-W. A reação das linhagens permaneceu em geral pouco alterada aos 49 DAI. A existência de linhagens resistentes somente ao WMV e somente ao PRSV-W, ao lado de linhagens resistentes a ambos os vírus, é indicativo de que as resistências ao WMV e ao PRSV-W não são controladas pelos mesmos genes.Twenty advanced watermelon breeding lines, derived from the cross between cv. Crimson Sweet (susceptible and PI 595201 (resistant to WMV and PRSV-W, were screened for resistance to both potyviruses. The twenty lines, among with Crimson Sweet and PI 595201, were inoculated with either WMV or PRSV-W, in two different greenhouse trials. Plants were evaluated for symptoms 35 and 49 days after the first inoculation (DAI, using a scale from 1 (no symptoms to 5 (severe mosaic and foliar distortion. Evaluations at 35 DAI indicated that lines 1, 2 and 20 had good levels of resistance to both WMV and PRSV-W, with ratings of 1,95, 1,80 and 2,25 for WMV, and of 2,50, 2,30 and 2,50 for PRSV-W, respectively. Lines 5, 7 and 13 were resistant to WMV only, whereas lines 3, 10 and 18 were resistant to PRSV-W only. The reaction of
Maria Tereza Grombone-Guaratini
Full Text Available Studies about reproductive biology of weed species can have implications on the establishment of controlling practices that minimize the effects of these weed populations on agricultural fields. The pollination biology of Bidens alba (L. DC., B. pilosa L., and Bidens subalternans DC., was studied at different sites and climatic seasons. Bidens pilosa and B. subalternans are widely distributed in agricultural areas, in disturbed habitats, and along road sides. Bidens alba occur only along the coast. The three species are self-compatible and non agamospermous. The composition of the pollinator community changes during the year and between sites. Hymenopterans and lepidopterans are the most frequent visitors to Bidens species in both areas studied. Although the species are self-compatible, the presence of pollinators may affect the levels of inbreeding. The attraction of insects by Bidens species may be benefical to agricultural crop and may also have important implications for conservation biology.Estudos de biologia reprodutiva de espécies invasoras podem ter implicações sobre o estabelecimento de práticas de controle que minimizem o efeito das populações destas espécies em áreas agrícolas. A biologia da polinização de Bidens alba (L. DC., B. pilosa L. e Bidens subalternans DC. foi estudada em diferentes locais e estações climáticas. Bidens pilosa e B. subalternans são espécies amplamente distribuídas em áreas agrícolas, em habitats perturbados e em margens de estradas. Bidens alba ocorre somente em regiões litorâneas. As três espécies são auto-compatíveis e não são agamospérmicas. A composição da comunidade de polinizadores apresenta diferenças durante o ano e entre locais. Himenópteros e lepidópteros são os visitantes mais freqüentes nas espécies de Bidens. Embora as espécies sejam auto compatíveis, a presença de polinizadores pode afetar os níveis de endocruzamento. A atração de insetos por esp
Mazus reptans N.E. Br (creeping mazus) is a perennial flowering groundcover plant in the family Scrophulariaceae. A plant of M. reptans ‘Alba’ with mild mosaic symptoms was obtained from a Maryland nursery in 2010. Electron microscopy revealed the presence of slightly flexuous particles of 595-674...
Bald-Blume, N.; Bergervoet, J.H.W.; Maiss, E.
A nucleic acid based test for the detection of the economically important plant virus Cucumber mosaic virus (CMV) based on the Luminex xTAG technology was developed. This technology has the advantage of allowing the simultaneous detection of various targets. Applying this method, we prove the presen
Rogers Stephanie M
Full Text Available Abstract Background The September 11, 2001 attacks on the World Trade Center and the Pentagon increased the concern about the potential for terrorist attacks on many vulnerable sectors of the US, including agriculture. The concentrated nature of crops, easily obtainable biological agents, and highly detrimental impacts make agroterrorism a potential threat. Although procedures for an effective criminal investigation and attribution following such an attack are available, important enhancements are still needed, one of which is the capability for fine discrimination among pathogen strains. The purpose of this study was to develop a molecular typing assay for use in a forensic investigation, using Wheat streak mosaic virus (WSMV as a model plant virus. Method This genotyping technique utilizes single base primer extension to generate a genetic fingerprint. Fifteen single nucleotide polymorphisms (SNPs within the coat protein and helper component-protease genes were selected as the genetic markers for this assay. Assay optimization and sensitivity testing was conducted using synthetic targets. WSMV strains and field isolates were collected from regions around the world and used to evaluate the assay for discrimination. The assay specificity was tested against a panel of near-neighbors consisting of genetic and environmental near-neighbors. Result Each WSMV strain or field isolate tested produced a unique SNP fingerprint, with the exception of three isolates collected within the same geographic location that produced indistinguishable fingerprints. The results were consistent among replicates, demonstrating the reproducibility of the assay. No SNP fingerprints were generated from organisms included in the near-neighbor panel, suggesting the assay is specific for WSMV. Using synthetic targets, a complete profile could be generated from as low as 7.15 fmoles of cDNA. Conclusion The molecular typing method presented is one tool that could be
Wang, D; Maule, A J
Seed transmission of an isolate of pea seed-borne mosaic virus (PSbMV) in several pea genotypes has been studied. Cross-pollination experiments showed that pollen transmission of PSbMV did not occur and accordingly, virus was not detected in pollen grains by ELISA or electron microscopy. Comparative studies between two pea cultivars, one with a high incidence of seed transmission and one with none, showed that PSbMV infected the floral tissues (sepals, petals, anther and carpel) of both cultivars, but was not detected in ovules prior to fertilization. Virus was detected equally well in seed coats of the progeny in both cultivars. Analysis of virus incidence and concentration in pea seeds of different developmental stages demonstrated that in the cultivar with a high incidence of seed transmission, PSbMV directly invaded immature embryos, multiplied in the embryonic tissues and persisted during seed maturation. In contrast, the cultivar without seed transmission did not show invasion of immature embryos by the virus; there was no evidence for virus multiplication or persistence during embryo development and seed maturation. Hence seed transmission of PSbMV resulted from direct invasion of immature pea embryos by the virus and the block to seed transmission in the non-permissive cultivar probably occurred at this step.
Calò, Annalisa; Eiben, Sabine; Okuda, Mitsuhiro; Bittner, Alexander M.
Virus particles and proteins are excellent examples of naturally occurring structures with well-defined nanoscale architectures, for example, cages and tubes. These structures can be employed in a bottom-up assembly strategy to fabricate repetitive patterns of hybrid organic-inorganic materials. In this paper, we review methods of assembly that make use of protein and virus scaffolds to fabricate patterned nanostructures with very high spatial control. We chose (apo)ferritin and tobacco mosaic virus (TMV) as model examples that have already been applied successfully in nanobiotechnology. Their interior space and their exterior surfaces can be mineralized with inorganic layers or nanoparticles. Furthermore, their native assembly abilities can be exploited to generate periodic architectures for integration in electrical and magnetic devices. We introduce the state of the art and describe recent advances in biomineralization techniques, patterning and device production with (apo)ferritin and TMV.
Zhou, X; Robinson, D J; Harrison, B D
Complete nucleotide sequences of the DNA-A-like molecules of three East African cassava mosaic virus (EACMV) isolates from Kenya (-K, 2801 nt) and Malawi (-MH and -MK, both 2804 nt) were determined. These sequences were compared with that published for a Tanzanian isolate (-T, 2801 nt) and the partial sequence of a third Malawian isolate. Intergenic region sequences of all isolates, and deduced amino acid sequences of their AC1 (Rep) proteins, each formed a tightly related cluster that was distinct from the comparable components of other begomoviruses. Other complementary-sense genes (AC2, AC3, AC4) differed between EACMV isolates in a way consistent with the accumulation of point mutations. In contrast, virus-sense genes (CP, AV2) of isolates -MH and -MK differed (substantially for AV2) from those of other EACMV isolates but somewhat resembled those of tomato yellow leaf curl virus-Israel, suggesting they had been acquired by recombination with an unidentified begomovirus.
Full Text Available Raman spectroscopy provides many advantages compared to other common analytical techniques due to its ability of rapid and accurate identification of unknown specimens as well as simple sample preparation. Here, we described potential of Raman spectroscopic technique as an efficient and high throughput method to detect plants infected by economically important viruses. To enhance the detection sensitivity of Raman measurement, surface enhanced Raman scattering (SERS was employed. Spectra of extracts from healthy and Turnip yellow mosaic virus (TYMV infected Chinese cabbage leaves were collected by mixing with gold (Au nanoparticles. Our result showed that TYMV infected plants could be discriminated from non-infected healthy plants, suggesting the current method described here would be an alternative potential tool to screen virus-infection of plants in fields although it needs more studies to generalize the technique.
Ohshima, Kazusato; Matsumoto, Kosuke; Yasaka, Ryosuke; Nishiyama, Mai; Soejima, Kenta; Korkmaz, Savas; Ho, Simon Y W; Gibbs, Adrian J; Takeshita, Minoru
Cucumber mosaic virus (CMV) is a damaging pathogen of over 200 mono- and dicotyledonous crop species worldwide. It has the broadest known host range of any virus, but the timescale of its evolution is unknown. To investigate the evolutionary history of this virus, we obtained the genomic sequences of 40 CMV isolates from brassicas sampled in Iran, Turkey and Japan, and combined them with published sequences. Our synonymous ('silent') site analyses revealed that the present CMV population is the progeny of a single ancestor existing 1550-2600 years ago, but that the population mostly radiated 295-545 years ago. We found that the major CMV lineages are not phylogeographically confined, but that recombination and reassortment is restricted to local populations and that no reassortant lineage is more than 251 years old. Our results highlight the different evolutionary patterns seen among viral pathogens of brassica crops across the world.
Lígia Maria Lembo Duarte
Full Text Available The culture and commercialization of ornamental plants have considerably increased in the last years. To supply the commercial demand, several Hemerocallis and Impatiens varieties have been bred for appreciated qualities such as flowers with a diversity of shapes and colors. With the aim of characterizing the tobamovirus isolated from Hemerocallis sp. (tobamo-H and Impatiens hawkeri (tobamo-I from the USA and São Paulo, respectively, as well as to establish phylogenetic relationships between them and other Tobamovirus species, the viruses were submitted to RNA extraction, RT-PCR amplification, coat-protein gene sequencing and phylogenetic analyses. Comparison of tobamovirus homologous sequences yielded values superior to 98.5% of identity with Tomato mosaic virus (ToMV isolates at the nucleotide level. In relation to tobamo-H, 100% of identity with ToMV from tomatoes from Australia and Peru was found. Based on maximum likelihood (ML analysis it was suggested that tobamo-H and tobamo-I share a common ancestor with ToMV, Tobacco mosaic virus, Odontoglossum ringspot virus and Pepper mild mottle virus. The tree topology reconstructed under ML methodology shows a monophyletic group, supported by 100% of bootstrap, consisting of various ToMV isolates from different hosts, including some ornamentals, from different geographical locations. The results indicate that Hemerocallis sp. and I. hawkeri are infected by ToMV. This is the first report of the occurrence of this virus in ornamental species in Brazil.O cultivo e comercialização de plantas ornamentais têm aumentado consideravelmente nos últimos anos. Para suprir a demanda comercial, diversas variedades de Hemerocallis sp. e Impatiens hawkeri têm sido desenvolvidas pelas qualidades apreciáveis como flores com diversidade de formas e cores. Com o objetivo de caracterizar o tobamovirus isolado de Hemerocallis sp. (tobamo-H e Impatiens hawkeri (tobamo-I provenientes dos EUA e São Paulo
Full Text Available Lettuce mosaic virus (LMV causes disease of plants in the family Asteraceae, especially lettuce crops. LMV isolates have previously been clustered in three main groups, LMV-Yar, LMV-Greek and LMVRoW. The first two groups, LMV-Yar and LMV-Greek, have similar characteristics such as no seed-borne transmission and non-resistance-breaking. The latter one, LMV-RoW, comprising a large percentage of the LMV isolates contains two large subgroups, LMV-Common and LMV-Most. To date, however, no Korean LMV isolate has been classified and characterized. In this study, LMV-Muju, the Korean LMV isolate, was isolated from lettuce showing pale green and mottle symptoms, and its complete genome sequence was determined. Classification method of LMV isolates based on nucleotide sequence divergence of the NIb-CP junction showed that LMV-Muju was categorized as LMV-Common. LMV-Muju was more similar to LMV-O (LMV-Common subgroup than to LMV-E (LMV-RoW group but not LMV-Common subgroup even in the amino acid domains of HC-Pro associated with pathogenicity, and in the CI and VPg regions related to ability to overcome resistance. Taken together, LMV-Muju belongs to the LMV-Common subgroup, and is expected to be a seed-borne, non-resistance-breaking isolate. According to our analysis, all other LMV isolates not previously assigned to a subgroup were also included in the LMV-RoW group.
RNA silencing is a post-transcriptional genesilencing phenomenon induced by double-stranded RNA (dsRNA).In an attempt to generate dsRNA-mediated transgenic maize plants resistant to sugarcane mosaic virus (SCMV),we cloned SCMV Nib gene-specificsequences and inserted it into the binary vector p3301 in the sense and antisense orientations (named SCMVir-Nib),which could produce RNAs capable of duplex formation in plant cells.Maize immature embryos were co-cultured with Agrobacterium carrying two vectors,one marker-free vector harboring the SCMVirNIb and one vector harboring bar gene as the selective marker.Resistant calli were recovered by selection on medium containing Biolaphos.Among the regenerated plantlets from resistant calli,14 plants have been certified to contain SCMVirNIb by PCR amplification and DNA dot blot.T1 plants derived from the 14 plants were challenged in a greenhouse with SCMV inoculums and the percentages of resistant plants in 11 T1 lines were higher than 60%.One plant in the T1 line was found to carry SCMVirNIb without bar gene by PCR assay.T2 plants derived from T1 SCMV resistant transgenic plants were challenged with SCMV inoculums in field.The percentages of resistant plants from 3 lines,including the line derived from the marker-free transgenic plant,were higher than 85%.The non-transgenic control plants were all susceptible.Further molecular analysis confirmed that the resistant plants from the marker-free transgenic line contained SCMVirNIb but not the bar gene.
Full Text Available Francisella tularensis is a facultative intracellular pathogen, and is the causative agent of a fatal human disease known as tularemia. F. tularensis is classified as a Category A Biothreat agent by the CDC based on its use in bioweapon programs by several countries in the past and its potential to be used as an agent of bioterrorism. No licensed vaccine is currently available for prevention of tularemia. In this study, we used a novel approach for development of a multivalent subunit vaccine against tularemia by using an efficient tobacco mosaic virus (TMV based delivery platform. The multivalent subunit vaccine was formulated to contain a combination of F. tularensis protective antigens: OmpA-like protein (OmpA, chaperone protein DnaK and lipoprotein Tul4 from the highly virulent F. tularensis SchuS4 strain. Two different vaccine formulations and immunization schedules were used. The immunized mice were challenged with lethal (10xLD100 doses of F. tularensis LVS on day 28 of the primary immunization and observed daily for morbidity and mortality. Results from this study demonstrate that TMV can be used as a carrier for effective delivery of multiple F. tularensis antigens. TMV-conjugate vaccine formulations are safe and multiple doses can be administered without causing any adverse reactions in immunized mice. Immunization with TMV-conjugated F. tularensis proteins induced a strong humoral immune response and protected mice against respiratory challenges with very high doses of F. tularensis LVS. This study provides a proof-of-concept that TMV can serve as a suitable platform for simultaneous delivery of multiple protective antigens of F. tularensis. Refinement of vaccine formulations coupled with TMV-targeting strategies developed in this study will provide a platform for development of an effective tularemia subunit vaccine as well as a vaccination approach that may broadly be applicable to many other bacterial pathogens.
Banik, Sukalyani; Mansour, Ahd Ahmed; Suresh, Ragavan Varadharajan; Wykoff-Clary, Sherri; Malik, Meenakshi; McCormick, Alison A; Bakshi, Chandra Shekhar
Francisella tularensis is a facultative intracellular pathogen, and is the causative agent of a fatal human disease known as tularemia. F. tularensis is classified as a Category A Biothreat agent by the CDC based on its use in bioweapon programs by several countries in the past and its potential to be used as an agent of bioterrorism. No licensed vaccine is currently available for prevention of tularemia. In this study, we used a novel approach for development of a multivalent subunit vaccine against tularemia by using an efficient tobacco mosaic virus (TMV) based delivery platform. The multivalent subunit vaccine was formulated to contain a combination of F. tularensis protective antigens: OmpA-like protein (OmpA), chaperone protein DnaK and lipoprotein Tul4 from the highly virulent F. tularensis SchuS4 strain. Two different vaccine formulations and immunization schedules were used. The immunized mice were challenged with lethal (10xLD100) doses of F. tularensis LVS on day 28 of the primary immunization and observed daily for morbidity and mortality. Results from this study demonstrate that TMV can be used as a carrier for effective delivery of multiple F. tularensis antigens. TMV-conjugate vaccine formulations are safe and multiple doses can be administered without causing any adverse reactions in immunized mice. Immunization with TMV-conjugated F. tularensis proteins induced a strong humoral immune response and protected mice against respiratory challenges with very high doses of F. tularensis LVS. This study provides a proof-of-concept that TMV can serve as a suitable platform for simultaneous delivery of multiple protective antigens of F. tularensis. Refinement of vaccine formulations coupled with TMV-targeting strategies developed in this study will provide a platform for development of an effective tularemia subunit vaccine as well as a vaccination approach that may broadly be applicable to many other bacterial pathogens.
Berrie, L C; Rybicki, E P; Rey, M E
Complete nucleotide sequences of the DNA-A (2800 nt) and DNA-B (2760 nt) components of a novel cassava-infecting begomovirus, South African cassava mosaic virus (SACMV), were determined and compared with various New World and Old World begomoviruses. SACMV is most closely related to East African cassava mosaic virus (EACMV) in both its DNA-A (85% with EACMV-MH and -MK) and -B (90% with EACMV-UG2-Mld and EACMV-UG3-Svr) components; however, percentage sequence similarities of less than 90% in the DNA-A component allowed SACMV to be considered a distinct virus. One significant recombination event spanning the entire AC4 open reading frame was identified; however, there was no evidence of recombination in the DNA-B component. Infectivity of the cloned SACMV genome was demonstrated by successful agroinoculation of cassava and three other plant species (Phaseolus vulgaris, Malva parviflora and Nicotiana benthamiana). This is the first description of successful infection of cassava with a geminivirus using Agrobacterium tumefaciens.
Full Text Available Abstract Background Although tobacco mosaic virus (TMV coat protein (CP has been isolated from virus particles and its crystals have grown in ammonium sulfate buffers for many years, to date, no one has reported on the crystallization of recombinant TMV-CP connecting peptides expressed in E. coli. Methods In the present papers genetically engineered TMV-CP was expressed, into which hexahistidine (His tags or glutathione-S-transferase (GST tags were incorporated. Considering that GST-tags are long peptides and His-tags are short peptides, an attempt was made to grow crystals of TMV-CP cleaved GST-tags (WT-TMV-CP32 and TMV-CP incorporated His-tags (WT-His-TMV-CP12 simultaneously in ammonium sulfate buffers and commercial crystallization reagents. It was found that the 20S disk form of WT-TMV-CP32 and WT-His-TMV-CP12 did not form high resolution crystals by using various crystallization buffers and commercial crystallization reagents. Subsequently, a new experimental method was adopted in which a range of truncated TMV-CP was constructed by removing several amino acids from the N- or the C-terminal, and high resolution crystals were grown in ammonium sulfate buffers and commercial crystallization reagents. Results The new crystallization method was developed and 3.0 Å resolution macromolecular crystal was thereby obtained by removing four amino acids at the C-terminal of His-TMV-CP and connecting six His-tags at the N-terminal of His-TMV-CP (TR-His-TMV-CP19. The Four-layer aggregate disk structure of TR-His-TMV-CP19 was solved. This phenomenon showed that peptides at the C-terminus hindered the growth of high resolution crystals and the peptides interactions at the N-terminus were attributed to the quality of TMV-CP crystals. Conclusion A 3.0 Å resolution macromolecular crystal of TR-His-TMV-CP19 was obtained and the corresponding structure was solved by removing four amino acids at the C-terminus of TMV-CP and connecting His-tags at the N
Butler, P J
The tobacco mosaic virus (TMV) particle was the first macromolecular structure to be shown to self-assemble in vitro, allowing detailed studies of the mechanism. Nucleation of TMV self-assembly is by the binding of a specific stem-loop of the single-stranded viral RNA into the central hole of a two-ring sub-assembly of the coat protein, known as the 'disk'. Binding of the loop onto its specific binding site, between the two rings of the disk, leads to melting of the stem so more RNA is availa...
Slomka, M J; Buck, K W; Coutts, R H
When callus tissue was cultured from leaf pieces taken from a Nicotiana tabacum cv. Xanthi nc. plant systemically infected with tomato golden mosaic virus (TGMV), TGMV-specific DNA persisted for up to 6 months in culture. Analysis of TGMV-specific intracellular DNA forms indicated a decrease in double-stranded relative to single-stranded forms and an increase in sub-genomic relative to genomic single-stranded DNA species in the callus tissue compared to those in the original leaf explant. The implications of the results with regard to TGMV replication are discussed.
Bink, Hugo H. J.; Schirawski, Jan; Haenni, Anne-Lise; Pleij, Cornelis W. A.
The RNA genome of turnip yellow mosaic virus (TYMV) consists of more than 6,000 nucleotides. During a study of the roles of the two hairpins located in its 90-nucleotide 5′ untranslated region, it was observed that stabilization of the 5′-proximal hairpin leads to a delay in the development of symptoms on plants. This delay in symptom development for both locally and systemically infected leaves was found to be dependent on a change in the free energy of the hairpin caused by introduced mutat...
Lam, Patricia; Gulati, Neetu M.; Stewart, Phoebe L.; Keri, Ruth A.; Steinmetz, Nicole F.
The 2014 Ebola epidemic is the largest to date. There is no cure or treatment for this deadly disease; therefore there is an urgent need to develop new diagnostics to accurately detect Ebola. Current RT-PCR assays lack sensitive and reliable positive controls. To address this critical need, we devised a bio-inspired positive control for use in RT-PCR diagnostics: we encapsulated scrambled Ebola RNA sequences inside of tobacco mosaic virus to create a biomimicry that is non-infectious, but stable, and could therefore serve as a positive control in Ebola diagnostic assays. Here, we report the bioengineering and validation of this probe.
Full Text Available Mungbean yellow mosaic India virus (MYMIV, a bipartite legume infecting geminivirus that causes considerable yield losses in South-East Asia. Pathogen derived resistance (PDR in plants is a very effective approach to acquire resistance against viral infections. Extrinsic expression of RNAi constructs targeting viral infective proteins is one of the effective scenarios to silence viral infectivity. In the present study, we tested the efficacy of three intron-spliced hairpin RNAi constructs which prepared by targeting the Coat Protein (CP/AV1, Replication initiation protein (Rep/AC1 and Intergenic region (IR of Soybean isolate of MYMIV (MYMIV-Sb in respect of reducing the virus DNA accumulation. In planta transient assay method were used to introduce the RNAi constructs in cowpea seedlings. This approach gave up to 80 % of protection to cowpea plants against virus infection. Only 15-20 % disease symptoms were observed in RNAi constructs inoculated cowpea plants. Among three constructs, RNAi-Rep construct showed maximum efficacy when compared with RNAi-CP and RNAi-IR. Results obtained in this study confirmed that at transient level, introduction of virus gene in form of hairpin RNAi construct (against the virus emerged as an effective strategy to control spreading the virus.
Sasaki, Nobumitsu; Ogata, Takuya; Deguchi, Masakazu; Nagai, Shoko; Tamai, Atsushi; Meshi, Tetsuo; Kawakami, Shigeki; Watanabe, Yuichiro; Matsushita, Yasuhiko; Nyunoya, Hiroshi
Tomato mosaic virus (ToMV) encodes a movement protein (MP) that is necessary for virus cell-to-cell movement. We have demonstrated previously that KELP, a putative transcriptional coactivator of Arabidopsis thaliana, and its orthologue from Brassica campestris can bind to ToMV MP in vitro. In this study, we examined the effects of the transient over-expression of KELP on ToMV infection and the intracellular localization of MP in Nicotiana benthamiana, an experimental host of the virus. In co-bombardment experiments, the over-expression of KELP inhibited virus cell-to-cell movement. The N-terminal half of KELP (KELPdC), which had been shown to bind to MP, was sufficient for inhibition. Furthermore, the over-expression of KELP and KELPdC, both of which were co-localized with ToMV MP, led to a reduction in the plasmodesmal association of MP. In the absence of MP expression, KELP was localized in the nucleus and the cytoplasm by the localization signal in its N-terminal half. It was also shown that ToMV amplified normally in protoplasts prepared from leaf tissue that expressed KELP transiently. These results indicate that over-expressed KELP interacts with MP in vivo and exerts an inhibitory effect on MP function for virus cell-to-cell movement, but not on virus amplification in individual cells.
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Korber, Bette T.; Fischer, William; Liao, Hua-Xin; Haynes, Barton F.; Letvin, Norman; Hahn; Beatrice H.
The present invention relates to mosaic clade M HIV-1 Env polypeptides and to compositions comprising same. The polypeptides of the invention are suitable for use in inducing an immune response to HIV-1 in a human.
程晔; 陈炯; 郑滔; 杨建平; 陈剑平
从杭州地区呈现玉米矮花叶典型症状的玉米病组织中提纯得到大量线状病毒粒子，大多数长度为750?nm。病组织中含有大量风轮状内含体和板状集结体。病毒外壳蛋白为33.6?kD。病毒RNA1 3’端序列(1.8?kb)与甘蔗花叶病毒(SCMV)同源性最高，达71.5%～99.1%，与高梁花叶病毒(SrMV)同源性次之，为67.8%～68.5%，与玉米矮花叶病毒(MDMV)同源性最低，仅为38.4%～48.4%，从而初步认为此病害由SCMV引起。根据已发表的SCMV外壳蛋白氨基酸序列作亲缘性分析，表明SCMV可分为美国、南非、澳大利亚；德国和中国三大类。%Recently maize dwarf mosaic disease was occurred on maize crop seriously in large scale in Hangzhou district. Purified preparations from the infected maize leaves contained numerous filamentous virus particles of c.750 nm in length. Cells of infected plants contained typical pinwheels and laminated aggregates. The coat protein of the virus was 33.6 kD. A 1.8 kb fragment of 3'-terminus of the viral RNA was amplified by RT-PCR, cloned and its sequence was determined. Sequence comparisons showed that it shared 71.5%～99.1% homology with isolates of sugarcane mosaic virus, 67.8%～68.5% with sorghum mosaic virus and 38.4%～48.4% with maize dwarf mosaic virus, indicating that the pathogen of this disease on maize in Hangzhou was sugarcane mosaic virus. In addition, the relationships of sugarcane mosaic virus isolates from different origins all over world were discussed based on coat protein sequences.
何煜波; 桂明; 李永伟; 唐爱菊; 陈集双
利用32p标记的cDNA探针对采集于湖南省永州等地的10种天南星科观赏植物和大田作物绿帝王(Philodendron sodiroi)、半夏(Pinellia ternata)、海芋(Alocasia macrorhiza)、马蹄莲(Zantedeschia aethiopica)、龟背竹(Monstera deliciosa)、尖尾芋(Alocasia cucullata)、白蝶合果芋(Sygonium podophyllum)、羽裂蔓绿绒(Philodendron selloum)、广东万年青(Aglaonema modestum)、芋(Colocasia esculenta)进行了芋花叶病毒(Dasheen mosaic virus,DsMV)和黄瓜花叶病毒(Cucumber mosaic virus,CMV)的检测,同时对部分样品进行病毒提纯、病毒粒子观察和内含体结构检查.结果显示:DsMV在10种植物上普遍存在,是最主要的病毒病原;同时首次在芋、绿帝王和广东万年青等3种植物上检测到CMV和DsMV的复合侵染.
Long, Chengwen; Li, Pei; Chen, Meihang; Dong, Liangrun; Hu, Deyu; Song, Baoan
A series of novel 1,4-pentadien-3-one derivatives containing 4-thioquinazoline moiety were designed and synthesized. Antiviral bioassay results indicated that most of the title compounds exhibited excellent antiviral activities against tobacco mosaic virus (TMV) and cucumber mosaic virus (CMV) in vivo. Among the title compounds, 7j exhibited the best curative activity against TMV, with a half-maximal effective concentration (EC50) value of 213.5 μg/mL, which was better than that of ningnanmycin (270.9 μg/mL). Meanwhile, 7a showed remarkable protection activity against TMV and curative activity against CMV, with EC50 values of 124.3 and 365.5 μg/mL, respectively, which were superior to those of ningnanmycin (195.1 and 404.9 μg/mL, respectively). Comparative molecular field analysis (CoMFA) and comparative molecular similarity index analysis (CoMSIA) models were generated on the basis of the curative activities against TMV and exhibited good predictive abilities with cross-validated q(2) and non-cross-validated r(2) values for CoMFA and CoMSIA of 0.548, 0.647 and 0.994, 0.993, respectively. These results provided a practical tool for guiding the design and synthesis of novel and more potent 1,4-pentadien-3-one derivatives containing 4-thioquinazoline moiety.
de Wispelaere, Mélissanne; Gaubert, Stéphane; Trouilloud, Séverine; Belin, Christophe; Tepfer, Mark
In order to better understand the role of recombination in creating the diversity of viral genomes that is acted on by selection, we have studied in detail the population of recombinant RNA3 molecules occurring in tobacco plants coinfected with wild-type strains of cucumber mosaic virus (CMV) and tomato aspermy virus (TAV) under conditions of minimal selection pressure. Recombinant RNA3s were observed in 9.6% of the samples. Precise homologous recombination predominated since it was observed at 28 different sites, primarily in six hot spots. Imprecise homologous recombination was observed at two sites, particularly within a GU repeat in the 5' noncoding region. Seven of the eight aberrant homologous recombination sites observed were clustered in the 3' noncoding region. These results have implications on the role of recombination in host adaptation and virus evolution. They also provide essential baseline information for understanding the potential epidemiological impact of recombination in transgenic plants expressing viral sequences.
The plasmid containing the promoter Act1, the coat protein (cp) gene of wheat yellow mosaic virus (WYMV) and the selectable bar gene, was delivered via particle bombardment, directly into immature embryos of a wheat cultivars. PCR and PCR-RFLP were employed to screen the existence of the cp gene in T0 and T1 generations. Seeds from the positive T1 plants were sowed in fields heavily contaminated with WYMV to detect their resistance. In field trial of virus infection, one of the transgenic wheat lines, P8-T2, exhibited highly disease-resistance. Western blot and RT-PCR analysis showed that the expression level of cp gene in the resistant transgenic line was reduced greatly compared to those susceptible to WYMV infection. This provided evidence to presume that the resistance obtained by the transgenic wheat line was stimulated by the mechanism of the virus induced gene silencing.
Comparative spatial spread overtime of Zucchini Yellow Mosaic Virus (ZYMV) and Watermelon Mosaic Virus (WMV) in fields of transgenic squash expressing the coat protein genes of ZYMV and WMV, and in fields of nontransgenic squash.
Klas, Ferdinand E; Fuchs, Marc; Gonsalves, Dennis
The spatial and temporal patterns of aphid-vectored spread of Zucchini Yellow Mosaic Virus (ZYMV) and Watermelon Mosaic Virus (WMV) were monitored over two consecutive years in plantings of nontransgenic and transgenic squash ZW-20H (commercial cv. Freedom II) and ZW-20B, both expressing the coat protein genes of ZYMV and WMV. All test plants were surrounded by nontransgenic plants that were mechanically inoculated with ZYMV or WMV, and served as primary virus source. Across all trials, none of the transgenic plants exhibited systemic symptoms upon infection by ZYMV and WMV but a few of them developed localized chlorotic dots and/or blotches, and had low mixed infection rates [4% (6 of 139) of ZW-20H and 9% (13 of 139) of ZW-20B], as shown by ELISA. Geostatistical analysis of ELISA positive transgenic plants indicated, (i) a lack of spatial relationship on spread of ZYMV and WMV for ZW-20H with flat omnidirectional experimental semivariograms that fitted poorly theoretical models, and (ii) some extent of spatial dependence on ZYMV spread for ZW-20B with a well structured experimental semivariogram that fitted poorly theoretical models during the first but not the second growing season. In contrast, a strong spatial dependence on spread of ZYMV and WMV was found for nontransgenic plants, which developed severe systemic symptoms, had prevalent mixed infection rates (62%, 86 of 139), and well-defined omnidirectional experimental semivariograms that fitted a spherical model. Geostatistical data were sustained by virus transmission experiments with Myzus persicae in screenhouses, showing that commercial transgenic squash ZW-20H alter the dynamics of ZYMV and WMV epidemics by preventing secondary plant-to-plant spread.
Cláudia Roberta Ribeiro de Oliveira
Full Text Available O rendimento do feijão-caupi pode ser afetado por diversos fatores, em especial as viroses. As principais espécies de vírus que infectam o feijão-caupi, no Brasil, são: Cucumber mosaic virus (CMV, Cowpea aphid-borne mosaic virus (CABMV, Cowpea severe mosaic virus (CPSMV e o Bean golden mosaic virus (BGMV. Este trabalho foi realizado em duas etapas e teve como objetivo avaliar a reação de genótipos de feijão-caupi quanto à resistência à infecção simples pelo CMV e mista nas combinações CMV+CABMV, CMV+CPSMV-I e CMV+CABMV+CPSMV-I. Inicialmente, foram incluídos 57 genótipos, sendo três avaliações em gaiolas com tela antiafídeos sob infecção controlada, e uma em condição de campo sob infecção natural. Em seguida, foram selecionados 18 genótipos para serem desenvolvidos em nove ensaios, oito em gaiolas com tela antiafídeos sob infecção controlada, e um em campo sob infecção natural. Nesses ensaios, avaliaram-se os efeitos qualitativos e quantitativos resultantes das infecções. No ensaio de campo, foram avaliados o número de plantas assintomáticas, comprimento de vagem, número de grãos por vagem, massa de cem grãos e produtividade. As coinfecções reduziram a altura da planta e a massa seca. Além disso, nas infecções envolvendo os três vírus ocorreu a morte prematura de alguns genótipos. Os genótipos BR17-Gurguéia, Epace V-96, TE97-309G-9, TE97-309G-22, TE97-309G-24 e Patativa, além de bom comportamento diante das coinfecções virais, têm sementes com padrão comercial, podendo ser empregadas diretamente em programas de melhoramento.
Neeraj Verma; B K Mahinghara; Raja Ram; A A Zaidi
A viral disease was identified on geraniums (Pelargonium spp.) grown in a greenhouse at the Institute of Himalayan Bioresource Technology (IHBT), Palampur, exhibiting mild mottling and stunting. The causal virus (Cucumber mosaic virus, CMV) was identified and characterized on the basis of host range, aphid transmission, enzyme linked immunosorbent assay (ELISA), DNA-RNA hybridization and reverse transcription polymerase chain reaction (RTPCR). A complete coat protein (CP) gene was amplified using degenerate primers and sequenced. The CP gene showed nucleotide and amino acid homology up to 97%–98% and 96%–99%, respectively with the sequences of CMV subgroup II. The CP gene also showed homologies of 75%–97% in nucleotide and 77%–96% in amino acid with the CMV Indian isolates infecting various crops. On the basis of sequence homology, it was concluded that CMV-infecting geraniums in India belong to subgroup II.
Longchamp, Jean-Nicolas; Escher, Conrad; Fink, Hans-Werner
Modern structural biology relies on NMR, X-ray crystallography and cryo-electron microscopy for gaining information on biomolecules at nanometer, sub-nanometer or atomic resolution. All these methods, however, require averaging over a vast ensemble of entities and hence knowledge on the conformational landscape of an individual particle is lost. Unfortunately, there are now strong indications that even X-ray free electron lasers will not be able to image individual molecules but will require nanocrystal samples. Here, we show that non-destructive structural biology of single particles has now become possible by means of low-energy electron holography. Individual tobacco mosaic viruses deposited on ultraclean freestanding graphene are imaged at one nanometer resolution revealing structural details arising from the helical arrangement of the outer protein shell of the virus. Since low-energy electron holography is a lens-less technique and since electrons with a deBroglie wavelength of approximately 1 Angstrom ...
Zhang, Chunquan; Grosic, Sehiza; Whitham, Steven A; Hill, John H
Soybean mosaic virus (SMV) is a major viral pathogen of soybean. Among the three SMV resistance genes, Rsv1 mediates extreme resistance (ER) against most SMV strains, including the β-glucuronidase-tagged G2 isolate that was previously used in studies of Rsv1. Using virus-induced gene silencing (VIGS), we screened 82 VIGS constructs to identify genes that play a role in Rsv1-mediated ER to SMV infection. The target genes included putative Rsv1 candidate genes, soybean orthologs to known defense-signaling genes, and 62 WRKY transcription factors. We identified eight VIGS constructs that compromised Rsv1-mediated resistance when the target genes were silenced, including GmEDR1, GmEDS1, GmHSP90, GmJAR1, GmPAD4, and two WRKY transcription factors. Together, our results provide new insight into the soybean signaling network required for ER against SMV.
Full Text Available Abstract Amino acid sequence analyses indicate that the Soilborne wheat mosaic virus (SBWMV 19K protein is a cysteine-rich protein (CRP and shares sequence homology with CRPs derived from furo-, hordei-, peclu- and tobraviruses. Since the hordei- and pecluvirus CRPs were shown to be pathogenesis factors and/or suppressors of RNA silencing, experiments were conducted to determine if the SBWMV 19K CRP has similar activities. The SBWMV 19K CRP was introduced into the Potato virus X (PVX viral vector and inoculated to tobacco plants. The SBWMV 19K CRP aggravated PVX-induced symptoms and restored green fluorescent protein (GFP expression to GFP silenced tissues. These observations indicate that the SBWMV 19K CRP is a pathogenicity determinant and a suppressor of RNA silencing.
Chen, Yun-Xia; Wu, Mian; Ma, Fang-Fang; Chen, Jian-Qun; Wang, Bin
Soybean mosaic virus (SMV) is a devastating plant virus classified in the family Potyviridae, and known to infect cultivated soybeans (Glycine max). In this study, seven new SMVs were isolated from wild soybean samples and analyzed by whole-genome sequencing. An updated SMV phylogeny was built with the seven new and 83 known SMV genomic sequences. Results showed that three northeastern SMV isolates were distributed in clade III and IV, while four southern SMVs were grouped together in clade II and all contained a recombinant BCMV fragment (~900 bp) in the upstream part of the genome. This work revealed that wild soybeans in China also act as important SMV hosts and play a role in the transmission and diversity of SMVs.
Jun-Li FENG; Shao-Ning CHEN; Xiang-Shan TANG; Xian-Feng DING; Zhi-You DU; Ji-Shuang CHEN
A real-time RT-PCR procedure using the green fluorescent dye SYBR Green I was developed for determining the absolute and relative copies of cucumber mosaic virus (CMV) genomic RNAs contained in purified virions. Primers specific to each CMV ORF were designed and selected. Sequences were then amplified with length varying from 61 to 153 bp. Using dilution series of CMV genome RNAs prepared by in vitro transcription as the standard samples, a good linear correlation was observed between their threshold cycle (Ct)values and the logarithms of the initial template amounts. The copies of genomic RNA 1, RNA 2,RNA 3 and the subgenomic RNA 4 in CMV virions were quantified by this method, and the ratios were about Our work is the first report concerning the relative amounts of different RNA fragments in CMV virions as a virus with tripartite genome.
Hongyun, Chen; Wenjun, Zhao; Qinsheng, Gu; Qing, Chen; Shiming, Lin; Shuifang, Zhu
A real time reverse-transcription polymerase chain reaction (RT-PCR) was developed for efficient detection of Cucumber green mottle mosaic virus (CGMMV). The method was designed to use a duo-primer system with a TaqMan probe targeting the conserved sequence in 3' noncoding region (NCR) of CGMMV to detect isolates of this virus collected in China. The sensitivity of the real time RT-PCR assay was 0.13 pg of total RNA or 50 molecules of RNA transcripts. This level of sensitivity indicated that the one step real time RT-PCR developed in the present study could be used for routine testing assays. The real time RT-PCR method could assist in the implementation of quarantine measures for prevention and control of the disease caused by CGMMV.
Tang, Shu-Lun; Linz, Lucas B; Bonning, Bryony C; Pohl, Nicola L B
Pea enation mosaic virus (PEMV)--a plant RNA virus transmitted exclusively by aphids--causes disease in multiple food crops. However, the aphid-virus interactions required for disease transmission are poorly understood. For virus transmission, PEMV binds to a heavily glycosylated receptor aminopeptidase N in the pea aphid gut and is transcytosed across the gut epithelium into the aphid body cavity prior to release in saliva as the aphid feeds. To investigate the role of glycans in PEMV-aphid interactions and explore the possibility of viral control through blocking a glycan interaction, we synthesized insect N-glycan terminal trimannosides by automated solution-phase synthesis. The route features a mannose building block with C-5 ester enforcing a β-linkage, which also provides a site for subsequent chain extension. The resulting insect N-glycan terminal trimannosides with fluorous tags were used in a fluorous microarray to analyze binding with fluorescein isothiocyanate-labeled PEMV; however, no specific binding between the insect glycan and PEMV was detected. To confirm these microarray results, we removed the fluorous tag from the trimannosides for isothermal titration calorimetry studies with unlabeled PEMV. The ITC studies confirmed the microarray results and suggested that this particular glycan-PEMV interaction is not involved in virus uptake and transport through the aphid.
Bartels, Melissa; French, Roy; Graybosch, Robert A; Tatineni, Satyanarayana
An infectious cDNA clone of Triticum mosaic virus (TriMV) (genus Poacevirus; family Potyviridae) was used to establish three independent lineages in wheat to examine intra-host population diversity levels within protein 1 (P1) and coat protein (CP) cistrons over time. Genetic variation was assessed at passages 9, 18 and 24 by single-strand conformation polymorphism, followed by nucleotide sequencing. The founding P1 region genotype was retained at high frequencies in most lineage/passage populations, while the founding CP genotype disappeared after passage 18 in two lineages. We found that rare TriMV genotypes were present only transiently and lineages followed independent evolutionary trajectories, suggesting that genetic drift dominates TriMV evolution. These results further suggest that experimental populations of TriMV exhibit lower mutant frequencies than that of Wheat streak mosaic virus (genus Tritimovirus; family Potyviridae) in wheat. Nevertheless, there was evidence for parallel evolution at a synonymous site in the TriMV CP cistron.
Fang, Xin; Waghela, Suryakant D.; Bray, Jocelyn; Njongmeta, Leo M.; Herring, Andy; Abdelsalam, Karim W.; Chase, Christopher; Mwangi, Waithaka
Bovine viral diarrhea virus (BVDV) plays a key role in bovine respiratory disease complex, which can lead to pneumonia, diarrhea and death of calves. Current vaccines are not very effective due, in part, to immunosuppressive traits and failure to induce broad protection. There are diverse BVDV strains and thus, current vaccines contain representative genotype 1 and 2 viruses (BVDV-1 & 2) to broaden coverage. BVDV modified live virus (MLV) vaccines are superior to killed virus vaccines, but they are susceptible to neutralization and complement-mediated destruction triggered by passively acquired antibodies, thus limiting their efficacy. We generated three novel mosaic polypeptide chimeras, designated NproE2123; NS231; and NS232, which incorporate protective determinants that are highly conserved among BVDV-1a, 1b, and BVDV-2 genotypes. In addition, strain-specific protective antigens from disparate BVDV strains were included to broaden coverage. We confirmed that adenovirus constructs expressing these antigens were strongly recognized by monoclonal antibodies, polyclonal sera, and IFN-γ-secreting T cells generated against diverse BVDV strains. In a proof-of-concept efficacy study, the multi-antigen proto-type vaccine induced higher, but not significantly different, IFN-γ spot forming cells and T-cell proliferation compared to a commercial MLV vaccine. In regards to the humoral response, the prototype vaccine induced higher BVDV-1 specific neutralizing antibody titers, whereas the MLV vaccine induced higher BVDV-2 specific neutralizing antibody titers. Following BVDV type 2a (1373) challenge, calves immunized with the proto-type or the MLV vaccine had lower clinical scores compared to naïve controls. These results support the hypothesis that a broadly protective subunit vaccine can be generated using mosaic polypeptides that incorporate rationally selected and validated protective determinants from diverse BVDV strains. Furthermore, regarding biosafety of using a
Lindomar Maria da Silveira
Full Text Available Entre as doenças que ocorrem na cultura da melancia (Citrullus lanatus, a virose ocasionada por Watermelon mosaic virus (WMV se destaca entre as principais, sendo a resistência genética a forma mais indicada de controle. Dessa forma, é importante o conhecimento do controle genético da resistência que se pretende trabalhar. Objetivando estudar a herança da resistência ao WMV em melancia, foram realizados cruzamentos entre o cultivar Crimson Sweet (CS suscetível e a linha L26 resistente. Populações segregantes e não segregantes obtidas dos cruzamentos foram inoculadas com um isolado de WMV e avaliadas quanto ao aparecimento de sintomas e à presença do vírus por testes de ELISA indireto contra antissoro específico para WMV. A hipótese de herança monogênica foi avaliada em diferentes graus médios de dominância e pelo método da máxima verossimilhança. Foram obtidas variâncias genética (σ²G, ambiental (σ²E, fenotípica (σ²F2, aditiva (σ²A e de dominância (σ²D, herdabilidades nos sentidos amplo (h²a e restrito (h²r. A herança monogênica foi rejeitada. O grau médio de dominância indicou efeito de dominância completa. As herdabilidades no sentido amplo foram baixas; portanto, constatou-se que o controle da resistência a WMV nas populações de melancia estudadas é do tipo oligogênica, com presença de efeitos aditivos e não aditivos e presença de genes maiores e poligenes.
Lauffer, M A; Shalaby, R A
The effect of the dipolar ions, glycine, glycylglycine, and glycylglycylglycine on the polymerization of tobacco mosaic virus (TMV) protein has been studied by the methods of light scattering and ultracentrifugation. All three dipolar ions promote polymerization. The major reaction in the early stage is transition from the 4 S to the 20 S state. As in the absence of dipolar ions, the polymerization is enhanced by an increase in temperature; it is endothermic and therefore entropy-driven. The effect of the dipolar ions can be understood in terms of their action as salting-out agents; they increase the activity coefficient of TMV A protein, the 4 S material, and thus shift the equilibrium toward the 20 S state. The salting-out constants, K, for the reaction in 0.10 ionic strength phosphate buffer at pH 6.7 was found by the light scattering method to be 1.6 for glycine, 2.5 for glycylglycine, and 2.5 for glycylglycylglycine. A value of 2.7 was obtained by the ultracentrifugation method for glycylglycine in phosphate buffer at 0.1 ionic strength and pH 6.8 at 10 degrees C. For both glycine and glycylglycine, K increases when the ionic strength of the phosphate buffer is decreased. This result suggests that electrolytes decrease the activity coefficient of the dipolar ions, a salting-in phenomenon. However, the salting-in constants evaluated from these results are substantially higher than those previously determined by solubility measurements. The effect of glycine and glycylglycine on polymerization was studied at pH values between 6.2 and 6.8. The effectiveness of both dipolar ions is approximately 50% greater at pH 6.8 than at pH 6.2. The variation of the extent of polymerization with pH in the presence of the dipolar ions is consistent with the interpretation that approximately one hydrogen ion is bound for half of the polypeptide units in the polymerized A protein.
Coutts, B A; Kehoe, M A; Webster, C G; Wylie, S J; Jones, R A C
Between 2006 and 2010, 5324 samples from at least 34 weed, two cultivated legume and 11 native species were collected from three cucurbit-growing areas in tropical or subtropical Western Australia. Two new alternative hosts of zucchini yellow mosaic virus (ZYMV) were identified, the Australian native cucurbit Cucumis maderaspatanus, and the naturalised legume species Rhyncosia minima. Low-level (0.7%) seed transmission of ZYMV was found in seedlings grown from seed collected from zucchini (Cucurbita pepo) fruit infected with isolate Cvn-1. Seed transmission was absent in >9500 pumpkin (C. maxima and C. moschata) seedlings from fruit infected with isolate Knx-1. Leaf samples from symptomatic cucurbit plants collected from fields in five cucurbit-growing areas in four Australian states were tested for the presence of ZYMV. When 42 complete coat protein (CP) nucleotide (nt) sequences from the new ZYMV isolates obtained were compared to those of 101 complete CP nt sequences from five other continents, phylogenetic analysis of the 143 ZYMV sequences revealed three distinct groups (A, B and C), with four subgroups in A (I-IV) and two in B (I-II). The new Australian sequences grouped according to collection location, fitting within A-I, A-II and B-II. The 16 new sequences from one isolated location in tropical northern Western Australia all grouped into subgroup B-II, which contained no other isolates. In contrast, the three sequences from the Northern Territory fitted into A-II with 94.6-99.0% nt identities with isolates from the United States, Iran, China and Japan. The 23 new sequences from the central west coast and two east coast locations all fitted into A-I, with 95.9-98.9% nt identities to sequences from Europe and Japan. These findings suggest that (i) there have been at least three separate ZYMV introductions into Australia and (ii) there are few changes to local isolate CP sequences following their establishment in remote growing areas. Isolates from A-I and B
Bong K. Choi
Full Text Available Soybean mosaic virus (SMV-CN18 is an Rsv resistance-breaking (RB isolate to overcome soybean resistance genes Rsv1, Rsv3 and Rsv4. The aim of this study was to characterize nuclear inclusion protein a (NIa protein of RB isolate at the molecular level and demonstrate its processing into genome-linked protein (VPg and NIa-Pro domains in Esherichia coli containing a bacterial expression pET vector inserted with NIa gene. The full-length of NIa gene was synthesized by reverse transcription-polymerase chain reaction (RT-PCR and its 1298 nucleotides (nt and 432 amino acids (aa were deduced. The nt and aa sequences of NIa gene of SMV-CN18 shared high identities with the corresponding sequences of the NIa gene of the known SMV isolates, suggesting that the NIa is a highly conserved protein. The NIa-Pro domain contains a highly conserved structural motif for proteolysis, while the VPg domain contains a nuclear localization signal (NLS, a putative NTP-binding site and cellular factor-binding sites. The phylogenetic tree revealed that less divergence of NIa protein exists among twelve SMV isolates, which can be supported by a low bootstrap value between clades. In addition, the full-length of NIa gene, amplified by RT-PCR, was ligated into pET-28b E. coli expression vector with an N-terminal His6-tag. Optimal conditions for expression were at 1mM treatment of IPTG at 25°C for 5 hr. The released protein from bacterial lysates remained soluble and proved the processing form of the NIa polyprotein. E. coli expression system shows the processed product of 29 kDa VPg in SDS-PAGE confirmed by western blot analysis in both crude extracts and purified elution products, using Ni2+-NTA resin. The present study indicates that the N-terminal region of NIa which is processed and expressed in bacteria.
Andrew J Love
Full Text Available Cauliflower mosaic virus (CaMV encodes a multifunctional protein P6 that is required for translation of the 35S RNA and also acts as a suppressor of RNA silencing. Here we demonstrate that P6 additionally acts as a pathogenicity effector of an unique and novel type, modifying NPR1 (a key regulator of salicylic acid (SA- and jasmonic acid (JA-dependent signaling and inhibiting SA-dependent defence responses We find that that transgene-mediated expression of P6 in Arabidopsis and transient expression in Nicotiana benthamiana has profound effects on defence signaling, suppressing expression of representative SA-responsive genes and increasing expression of representative JA-responsive genes. Relative to wild-type Arabidopsis P6-expressing transgenics had greatly reduced expression of PR-1 following SA-treatment, infection by CaMV or inoculation with an avirulent bacterial pathogen Pseudomonas syringae pv tomato (Pst. Similarly transient expression in Nicotiana benthamiana of P6 (including a mutant form defective in translational transactivation activity suppressed PR-1a transcript accumulation in response to Agrobacterium infiltration and following SA-treatment. As well as suppressing the expression of representative SA-regulated genes, P6-transgenic Arabidopsis showed greatly enhanced susceptibility to both virulent and avirulent Pst (titres elevated 10 to 30-fold compared to non-transgenic controls but reduced susceptibility to the necrotrophic fungus Botrytis cinerea. Necrosis following SA-treatment or inoculation with avirulent Pst was reduced and delayed in P6-transgenics. NPR1 an important regulator of SA/JA crosstalk, was more highly expressed in the presence of P6 and introduction of the P6 transgene into a transgenic line expressing an NPR1:GFP fusion resulted in greatly increased fluorescence in nuclei even in the absence of SA. Thus in the presence of P6 an inactive form of NPR1 is mislocalized in the nucleus even in uninduced plants
Full Text Available BACKGROUND: Sugarcane mosaic virus (SCMV is an important virus pathogen in crop production, causing serious losses in grain and forage yields in susceptible cultivars. Control strategies have been developed, but only marginal successes have been achieved. For the efficient control of this virus, a better understanding of its interactions and associated resistance mechanisms at the molecular level is required. METHODOLOGY/PRINCIPAL FINDINGS: The responses of resistant and susceptible genotypes of maize to SCMV and the molecular basis of the resistance were studied using a proteomic approach based on two-dimensional polyacrylamide gel electrophoresis (2-DE and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS/MS analysis. Ninety-six protein spots showed statistically significant differences in intensity after SCMV inoculation. The classification of differentially expressed proteins showed that SCMV-responsive proteins were mainly involved in energy and metabolism, stress and defense responses, and photosynthesis. Most of the proteins identified were located in chloroplasts, chloroplast membranes, and the cytoplasm. Analysis of changes in phytohormone levels after virus inoculation suggested that salicylic acid, abscisic acid, jasmonic acid, and azelaic acid may played important roles in the maize response to SCMV infection. CONCLUSIONS/SIGNIFICANCE: Among these identified proteins, 19 have not been identified previously as virus-responsive proteins, and seven were new and did not have assigned functions. These proteins may be candidate proteins for future investigation, and they may present new biological functions and play important roles in plant-virus interactions. The behavioural patterns of the identified proteins suggest the existence of defense mechanisms operating during the early stages of infection that differed in two genotypes. In addition, there are overlapping and specific phytohormone
Li, Li; Wang, Li; Xiao, Ruijing; Zhu, Guoguo; Li, Yan; Liu, Changxuan; Yang, Ru; Tang, Zhiqing; Li, Jie; Huang, Wei; Chen, Lang; Zheng, Xiaoling; He, Yuling; Tan, Jinquan
The ability of human cells to defend against viruses originating from distant species has long been ignored. Owing to the pressure of natural evolution and human exploration, some of these viruses may be able to invade human beings. If their 'fresh' host had no defences, the viruses could cause a serious pandemic, as seen with HIV, SARS (severe acute respiratory syndrome) and avian influenza virus that originated from chimpanzees, the common palm civet and birds, respectively. It is unknown whether the human immune system could tolerate invasion with a plant virus. To model such an alien virus invasion, we chose TMV (tobacco mosaic virus) and used human epithelial carcinoma cells (HeLa cells) as its 'fresh' host. We established a reliable system for transfecting TMV-RNA into HeLa cells and found that TMV-RNA triggered autophagy in HeLa cells as shown by the appearance of autophagic vacuoles, the conversion of LC3-I (light chain protein 3-I) to LC3-II, the up-regulated expression of Beclin1 and the accumulation of TMV protein on autophagosomal membranes. We observed suspected TMV virions in HeLa cells by TEM (transmission electron microscopy). Furthermore, we found that TMV-RNA was translated into CP (coat protein) in the ER (endoplasmic reticulum) and that TMV-positive RNA translocated from the cytoplasm to the nucleolus. Finally, we detected greatly increased expression of GRP78 (78 kDa glucose-regulated protein), a typical marker of ERS (ER stress) and found that the formation of autophagosomes was closely related to the expanded ER membrane. Taken together, our data indicate that HeLa cells used ERS and ERS-related autophagy to defend against TMV-RNA.
Zhong, M; Zhao, X; Liu, Y; Wang, Y; Cao, K
Cucumber green mottle mosaic virus (CGMMV) is an important and widespread seed-borne virus that infects Cucurbitaceous plants. It is a member of the genus Tobamovirus in the family Virgaviridae with a monopartite (+) ssRNA genome. Here we report the complete genome sequence, construction and testing of the infectious clones of a chb isolate of CGMMV. Full-length CGMMV cDNA was cloned into the vector pUC19. The linearized vector containing full-length cDNA was used as template for in vitro transcription, and the synthesized capped transcript was highly infectious in Chenopodium amaranticolor and cucumber (Cucumis sativus). Inoculated plants showed symptoms typical of CGMMV infection. The infectivity was confirmed by mechanical transmission to new plants, RT-PCR and western blot. Progeny virus derived from infectious transcripts had the same biological and biochemical properties as wild-type virus. To our knowledge, this is the first detailed report of a biologically active transcript from CGMMV.
Mauck, Kerry E; De Moraes, Consuelo M; Mescher, Mark C
Plant viruses can profoundly alter the phenotypes of their host plants, with potentially far-reaching implications for ecology. Yet few studies have explored the indirect, host-mediated, effects of plant viruses on non-vector insects. We examined how infection of Cucurbita pepo plants by Cucumber mosaic virus (CMV) impacted the susceptibility of aphids (Myzus persicae) to attack by the parasitoid wasp Aphidius colemani. In semi-natural foraging assays, we observed higher rates of aphid parasitism on infected plants compared to healthy plants. Subsequent experiments revealed that this difference is not explained by different attack rates on plants differing in infection status, but rather by the fact that parasitoid larvae successfully complete their development more often when aphid hosts feed on infected plants. This suggests that the reduced nutritional quality of infected plants as host for aphids--documented in previous studies--compromises their ability to mount effective defenses against parasitism. Furthermore, our current findings indicate that the aphid diet during parasitoid development (rather than prior to wasp oviposition) is a key factor influencing resistance. These findings complement our previous work showing that CMV-induced changes in host plant chemistry alter patterns of aphid recruitment and dispersal in ways conducive to virus transmission.
Previous work had demonstrated the presence of a unique low-molecular-weight RNA component (LMC) in extracts of tobacco mosaic virus (TMV) infected tissue. Enough of this component has been isolated during the past year to ascertain that it has a molecular weight of 250,000 daltons and that it acts as an in vitro messenger for the synthesis of TMV capsid protein. Thus, we conclude that at least one monocistronic messenger RNA for a virion coded product is generated during TMV infection. Strains of TMV were classified according to nucleotide sequence homology of their RNAs. The strains fall into groups by the test employed. No differences were observed between strains within a group, whereas no homology was detected between groups. Using this information, it was possible, in part, to relate differences in capsid protein amino acid sequences to the degree of nomology of their nucleotide coding sequences. A study was initiated into the Pot Y virus group infection mechanism. In contrast to TMV infection, it was determined that for both tobacco etch and potato virus Y that: viral RNA synthesis is inhibited by actinomycin B and synthesis by virus-related proteins is inhibited by chloramphenicol.
Soybean mosaic virus (SMV), a member of the family Potyviridae, is an important viral pathogen affecting soybean production in Korea. The variability in helper component proteinase (HC-Pro) sequence and pathogenicity of SMV isolates from seven provinces of Korea was investigated and compared with th...
In 2014, we performed a nationwide survey in Korean radish fields to investigate the distribution of Turnip mosaic virus (TuMV). Brassica chinensis sap-inoculated with TuMV-infected radish tissue showed different symptom severity with three isolates. In order to investigate variation among Korean Tu...
de Paula, Nayhanne T; de Faria, Josias C; Aragão, Francisco J L
The RNAi concept was explored to silence the rep gene from the bean golden mosaic virus (BGMV) and a genetically modified (GM) bean immune to the virus was previously generated. We investigated if BGMV-viruliferous whiteflies would reduce viral amount after feeding on GM plants. BGMV DNA amount was significantly reduced in whiteflies feeding in GM-plants (compared with insects feeding on non-GM plants) for a period of 4 and 8 days in 52% and 84% respectively.
Korber, Bette T; Fischer, William; Liao, Hua-Xin; Haynes, Barton F; Letvin, Norman; Hahn, Beatrice H
The present invention relates to nucleic acids encoding mosaic clade M HIV-1 Env polypeptides and to compositions and vectors comprising same. The nucleic acids of the invention are suitable for use in inducing an immune response to HIV-1 in a human.
Inoue-Nagata, A.K.; Fonseca, M.E.N.; Resende, de R.O.; Boiteux, L.S.; Monte, D.C.; Dusi, A.N.; Ávila, de A.C.; Vlugt, van der R.A.A.
A potyvirus was found causing yellow mosaic and veinal banding in sweetpepper in Central and Southeast Brazil. The sequence analysis of the 3' terminal region of the viral RNA revealed a coat protein of 278 amino acids, followed by 275 nucleotides in the 3'-untranslated region preceding a polyadenyl
Guiu-Aragonés, Cèlia; Sánchez-Pina, María Amelia; Díaz-Pendón, Juan Antonio; Peña, Eduardo J; Heinlein, Manfred; Martín-Hernández, Ana Montserrat
Cucumber mosaic virus (CMV) has the broadest host range among plant viruses, causing enormous losses in agriculture. In melon, strains of subgroup II are unable to establish a systemic infection in the near-isogenic line SC12-1-99, which carries the recessive resistance gene cmv1 from the accession PI 161375, cultivar 'Songwhan Charmi'. Strains of subgroup I overcome cmv1 resistance in a manner dependent on the movement protein. We characterized the resistance conferred by cmv1 and established that CMV-LS (subgroup II) can move from cell to cell up to the veins in the inoculated leaf, but cannot enter the phloem. Immunogold labelling at transmission electron microscopy level showed that CMV-LS remains restricted to the bundle sheath (BS) cells in the resistant line, and does not invade vascular parenchyma or intermediary cells, whereas, in the susceptible line 'Piel de Sapo' (PS), the virus invades all vein cell types. These observations indicate that the resistant allele of cmv1 restricts systemic infection in a virus strain- and cell type-specific manner by acting as an important gatekeeper for virus progression from BS cells to phloem cells. Graft inoculation experiments showed that CMV-LS cannot move from the infected PS stock into the resistant cmv1 scion, thus suggesting an additional role for cmv1 related to CMV transport within or exit from the phloem. The characterization of this new form of recessive resistance, based on a restriction of virus systemic movement, opens up the possibility to design alternative approaches for breeding strategies in melon.
Chinese evergreen plants (Aglaonema commutatum) with symptoms of mild stunting, chlorosis, leaf distortion and mosaic, were observed in Southern California. Flexuous rods (ca. 750 nm) were detected in leaf dip and partially purified preparations. Dasheen mosac virus (DMV) was identified as the causal agent on the basis of host range, morphology and reaction with DMV antiserum in immunodouble diffusion and immunosorbent electron microscopy (ISEM) tests. Tetragonia expansa was found to be a new host of this virus. Surveys indicate that DMV is not widespread in cultivars of A. commutatum in Southern California. The virus was purified from leaves of seedling Philodendron selloum by clarification with CCl/sub 4/, CHCl/sub 3/, and Triton X-100, precipitation with PEG-8000 and centrifugation in either Cs/sub 2/SO/sub 4/-sucrose cushion gradients or Cs/sub 2/SO/sub 4/ equilibrium density gradients. Purified virions formed a single UV-absorbing infectious band with densities of 1.31 and 1.245 g/ml in CsCl/sub 2/ and Cs/sub 2/SO/sub 4/ equilibrium density gradients, respectively, and a sedimentation coefficient of 154 S as determined by a linear-log sucrose density gradient centrifugation. Dasheen mosaic virus has a plus-sense ssRNA with the M.W. of 3.2 x 10/sup 6/ under denaturing conditions. Molecular hybridization analysis using /sup 3/H-complementary DNA specific to DMV-Ca RNA showed that DMV-Ca isolate was more closely related to DMV-Fiji isolate than to DMV-Fla isolate, and was very distantly related to ZYMV, TEV. PeMoC and PVY.
高新国; 高宇溥; 杨正生
Objective] To study the epidemic characteristics of sesame yellow mosaic virus disease in fields to provide references for the pre-vention and control of the disease.[Method] In 2014, the appearance and epidemic characteristics of sesame yellow mosaic virus disease in fields were investigated.[ Result] The appearance and epidemic characteristics of sesame yellow mosaic virus disease in fields was associated with the combination effect of the growing stage of sesame and aphid occurrence .Sesame plants were susceptible to sesame yellow mosaic virus in the seedling and early flowering stages.The morbidity peak occurred 15 d after the aphid peak.[Conclusion] Regulating the sowing date of sesame to prevent the aphid peak from occurring in the seedling and early flowering stages can effectively prevent the occurrence of sesame yellow mosaic virus disease.%[目的]明确芝麻黄花叶病毒病田间流行规律，为防治该病提供参考。[方法]于2014年调查芝麻黄花叶病毒病的田间发生、流行规律。[结果]芝麻黄花叶病毒病的发生、流行与芝麻的生育期和蚜虫发生密切相关。苗期、蕾期是感染芝麻黄花叶病毒病的敏感期。蚜虫发生高峰15 d后，病害出现发病高峰。[结论]调节芝麻播种期，使苗期、蕾期，特别是蕾期错过蚜虫发生高峰能有效预防芝麻黄花叶病毒病的发生。
Liang, Chaoqiong; Meng, Yan; Luo, Laixin; Liu, Pengfei; Li, Jianqiang
The replicase genes of five isolates of Cucumber green mottle mosaic virus from Jiangsu, Zhejiang, Hunan and Beijing were amplificated, sequenced and analyzed. The similarities of nucleotide acid sequences indicated that 129 kD and 57 kD replicase genes of CGMMV-No. 1, CGMMV-No. 2, CGMMV-No. 3, CGMMV-No. 4 and CGMMV-No. 5 were 99.64% and 99.74%, respectively. The similarities of 129 kD and 57 kD replicase genes of CGMMV-No. 1, CGMMV-No. 3 and CGMMV-No. 4 were 99.95% and 99.94%, while they were lower between CGMMV-No. 2 and the rest of four reference sequences, just from 99.16% to 99.27% and from 99.04% to 99.18%. All reference sequences could be divided into six groups in neighbor-joining (NJ) phylogenetic trees based on the replicase gene sequences of 129 kD, 57 kD protein respectively. CGMMV-No. 1, CGMMV-No. 3 and CGMMV-No. 4 were clustered together with Shandong isolate (Accession No. KJ754195) in two NJ trees; CGMMV-No. 5 was clustered together with Liaoning isolate (Accession No. EF611826) in two NJ trees; CGMMV-No. 2 was clustered together with Korea watermelon isolate (Accession No. AF417242) in phylogenetic tree of 129 kD replicase gene of CGMMV; Interestingly, CGMMV-No. 2 was classified as a independent group in phylogenetic tree of 57 kD replicase gene of CGMMV. There were no significant hydrophobic and highly coiled coil regions on 129 kD and 57 kD proteins of tested CGMMV isolates. Except 129 kD protein of CGMMV-No. 4, the rest were unstable protein. The number of transmembrane helical segments (TMHs) of 129 kD protein of CGMMV-No. 1, CGMMV-No. 2, CGMMV-No. 3 and CGMMV-No. 5 were 6, 6, 2 and 4, respectively, which were 13, 13 and 5 on the 57 kD protein of CGMMV-No. 2, CGMMV-No. 4 and CGMMV-No. 5. The glycosylation site of 129 kD protein of tested CGMMV isolates were 2, 4, 4, 4 and 4, and that of 57 kD protein were 2, 5, 2, 5 and 2. There were difference between the disorders, globulins, phosphorylation sites and B cell antigen epitopes of 129 kD and 57
Ability of Aphis gossypii and Myzus persicae to Transmit Cucumber mosaic virus in Single and Mixed Infection with Two Potyviruses to Zucchini Squash Eficiência dos afídeos Aphis gossypii e Myzus persicae na transmissão do Cucumber mosaic virus em infecção simples e mista com dois Potyvirus para abobrinha de moita
Zayame Vegette Pinto
Full Text Available The main objective of this work was to investigate the ability of Aphis gossypii and Myzus persicae to transmit Cucumber mosaic virus (CMV singly and mixed with two potyviruses (Papaya ringspot virus - type W, PRSV-W and Zucchini yellow mosaic virus, ZYMV, to zucchini squash plants (Cucurbita pepo. The results showed that the potyviruses in general were more efficiently transmitted by both species of aphids as compared to CMV. The transmission of PRSV-W, ZYMV and CMV separately was more efficient than in mixture.O objetivo desse trabalho foi estudar a eficiência de Aphis gossypii e Myzus persicae na transmissão do vírus do mosaico do pepino (Cucumber mosaic virus, CMV, isoladamente e em mistura com duas espécies de potyvirus (Vírus do mosaico do mamoeiro = Papaya ringspot virus - type W, PRSV-W e Vírus do mosaico amarelo da abobrinha = Zucchini yellow mosaic virus, ZYMV, para planta-testes de abobrinha de moita (Cucurbita pepo. Os dois potyvirus em geral foram transmitidos com mais eficiência pelas duas espécies de afídeos do que o CMV. A transmissão do PRSV-W, ZYMV e CMV, separadamente, foi mais eficiente do que em mistura.
Full Text Available Plants of Capsicum annuum cv. Magali R, resistant to Pepper yellow mosaic virus (PepYMV, which showed severe yellow mosaic, leaf malformation and stunting were observed during the 2003/04 growing season in Lins, São Paulo State, Brazil. Potyvirus-like particles observed in leaf sap from infected plants under the electron microscope reacted with an antiserum against PepYMV in PTA-ELISA. In addition to C. annuum cv. Magali R, this potyvirus also infected systemically the resistant C. annuum cv. Rubia R. The nucleotide sequence of part of the CP gene of this potyvirus shared 96-98% identity with that of other PepYMV isolates. The partial nucleotide sequence of the 3' NTR showed 94-96% identity with that of PepYMV. These data indicate that this potyvirus is a resistance-breaking isolate of PepYMV.Plantas de Capsicum annuum cv. Magali R, resistentes ao Pepper yellow mosaic virus (PepYMV, exibindo sintomas severos de mosaico amarelo, malformação foliar e subdesenvolvimento foram encontradas em plantios na região de Lins, SP, Brasil, em 2003/04. Partículas semelhantes àquelas do gênero Potyvirus foram observadas em extrato foliar de planta infectada examinado em microscópio eletrônico de transmissão. O extrato foliar também reagiu com anti-soro contra o PepYMV em PTA-ELISA. Além de C. annuum cv. Magali R, esse potyvirus também infectou sistemicamente C. annuum cv. Rubia R, que é resistente ao PepYMV. A seqüência de nucleotídeos de parte do gene da proteína capsidial (CP desse potyvirus apresentou 96-98% de identidade com a de outros isolados do PepYMV. A seqüência parcial de nucleotídeos da região 3' não traduzida (3' NTR apresentou 94-96% de identidade com a do PepYMV. Esses resultados são indicativos de que o potyvirus que quebrou a resistência em pimentão é um isolado do PepYMV.
Pinheiro, Patrícia V; Quintela, Eliane D; Junqueira, Ana Maria R; Aragão, Francisco J L; Faria, Josias C
Genetically modified (GM) crops is considered the fastest adopted crop technology in the history of modern agriculture. However, possible undesirable and unintended effects must be considered during the research steps toward development of a commercial product. In this report we evaluated effects of a common bean virus resistant line on arthropod populations, considered as non-target organisms. This GM bean line (named M1/4) was modified for resistance against Bean golden mosaic virus (BGMV) by expressing a mutated REP protein, which is essential for virus replication. Biosafety studies were performed for a period of three years under field conditions. The abundance of some species was significantly higher in specific treatments in a particular year, but not consistently different in other years. A regular pattern was not observed in the distribution of insects between genetically modified and conventional treatments. Data analyses showed that minor differences observed can be attributed to random variation and were not consistent enough to conclude that the treatments were different. Therefore the present study indicates that the relative abundance of species are similar in transgenic and non-transgenic fields.
Babajide Odu O.
Full Text Available Resistance to Yam mosaic virus (YMV, genus Potyvirus was studied in 10 populations of selected white Guinea yam (Dioscorea rotundata. Plants of resistant genotypes: TDr 35, TDr 1621, TDr 93-1, TDr 93-32, TDr 95-107, TDr 93-23, and susceptible ones: TDr 87/00211, TDr 87/00571 and TDr 95-127 were screened for their reaction to the pathogen by symptom severity scoring scale of 1-5, and by quantifying virus multiplication by triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA. Controlled crosses were made among the genotypes within and between the groups according to reactions to the pathogen. The resultant F1 progenies were evaluated for the infection by disease symptom development and by TAS ELISA to detect a symptomless infection in an insect-proof screenhouse for the assessment of inheritance of resistance to YMV. A genetic analysis of the reactions of progenies derived from the D. rotundata genotypes to inoculation with YMV strongly suggests that resistance to the virus is a dominantly inherited trait. Segregation ratios obtained from the families indicate that at least two dominant genes are involved.
Tarora, Kazuhiko; Shudo, Ayano; Kawano, Shinji; Yasuda, Keiji; Ueno, Hiroki; Matsumura, Hideo; Urasaki, Naoya
In this study, we confirmed that Vasconcellea cundinamarcensis resists Papaya leaf distortion mosaic virus (PLDMV), and used it to produce intergeneric hybrids with Carica papaya. From the cross between C. papaya and V. cundinamarcensis, we obtained 147 seeds with embryos. Though C. papaya is a monoembryonic plant, multiple embryos were observed in all 147 seeds. We produced 218 plants from 28 seeds by means of embryo-rescue culture. All plants had pubescence on their petioles and stems characteristic of V. cundinamarcensis. Flow cytometry and PCR of 28 plants confirmed they were intergeneric hybrids. To evaluate virus resistance, mechanical inoculation of PLDMV was carried out. The test showed that 41 of 134 intergeneric hybrid plants showed no symptoms and were resistant. The remaining 93 hybrids showed necrotic lesions on the younger leaves than the inoculated leaves. In most of the 93 hybrids, the necrotic lesions enclosed the virus and prevented further spread. These results suggest that the intergeneric hybrids will be valuable material for PLDMV-resistant papaya breeding.
Cillo, Fabrizio; Roberts, Ian M; Palukaitis, Peter
The replication-associated proteins encoded by Cucumber mosaic virus (CMV), the 1a and 2a proteins, were detected by immunogold labeling in two host species of this virus, tobacco (Nicotiana tabacum) and cucumber (Cucumis sativus). In both hosts, the 1a and 2a proteins colocalized predominantly to the vacuolar membranes, the tonoplast. While plus-strand CMV RNAs were found distributed throughout the cytoplasm by in situ hybridization, minus-strand CMV RNAs were barely detectable but were found associated with the tonoplast. In both cucumber and tobacco, 2a protein was detected at higher densities than 1a protein. The 1a and 2a proteins also showed quantitative differences with regard to tissue distributions in tobacco and cucumber. About three times as much 2a protein was detected in CMV-infected cucumber tissues as in CMV-infected tobacco tissues. In tobacco, high densities of these proteins were observed only in vascular bundle cells of minor veins. In contrast, in cucumber, high densities of 1a and 2a proteins were observed in mesophyll cells, followed by epidermis cells, with only low levels being observed in vascular bundle cells. Differences were also observed in the distributions of 2a protein and capsid protein in vascular bundle cells of the two host species. These observations may represent differences in the relative rates of tissue infection in different hosts or differences in the extent of virus replication in vascular tissues of different hosts.
Li, Jin-yu; Wei, Qi-wei; Liu, Yong; Tan, Xin-qiu; Zhang, Wen-na; Wu, Jian-yan; Charimbu, Miriam Karwitha; Hu, Bai-shi; Cheng, Zhao-bang; Yu, Cui; Tao, Xiao-rong
Cucumber green mottle mosaic virus (CGMMV) has caused serious damage to Cucurbitaceae crops worldwide. The virus is considered one of the most serious Cucurbitaceae quarantine causes in many countries. In this study, a highly efficient and practical one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed for the detection of CGMMV. The total RNA or crude RNA extracted from watermelon plants or seeds could be detected easily by this RT-LAMP assay. The RT-LAMP assay was conducted in isothermal (63°C) conditions within 1h. The amplified products of CGMMV could be detected as ladder-like bands using agarose gel electrophoresis or visualized in-tube under UV light with the addition of a fluorescent dye. The RT-LAMP amplification was specific to CGMMV, as no cross-reaction was observed with other viruses. The RT-LAMP assay was 100-fold more sensitive than that of reverse-transcription polymerase chain reaction (RT-PCR). This is the first report of the application of the RT-LAMP assay to detect CGMMV. The sensitive, specific and rapid RT-LAMP assay developed in this study can be applied widely in laboratories, the field and quarantine surveillance of CGMMV.
Minicka, Julia; Otulak, Katarzyna; Garbaczewska, Grażyna; Pospieszny, Henryk; Hasiów-Jaroszewska, Beata
This paper presents studies on an ultrastructural analysis of plant tissue infected with different pathotypes of Pepino mosaic virus (PepMV) and the immunolocalization of viral coat proteins. Because the PepMV virus replicates with a high mutation rate and exhibits significant genetic diversity, therefore, isolates of PepMV display a wide range of symptoms on infected plants. In this work, tomato plants of the Beta Lux cultivar were inoculated mechanically with three pathotypes representing the Chilean 2 (CH2) genotype: mild (PepMV-P22), necrotic (PepMV-P19) and yellowing (PepMV-P5-IY). The presence of viral particles in all infected plants in the different compartments of various cell types (i.e. spongy and palisade mesophyll, sieve elements and xylem vessels) was revealed via ultrastructural analyses. For the first time, it was possible to demonstrate the presence of crystalline inclusions, composed of virus-like particles. In the later stage of PepMV infection (14 dpi) various pathotype-dependent changes in the structure of the individual organelles (i.e. mitochondria, chloroplasts) were found. The strongest immunogold labeling of the viral coat proteins was also observed in plants infected by necrotic isolates. A large number of viral coat proteins were marked in the plant conductive elements, both xylem and phloem.
Full Text Available The Cucumber mosaic virus (CMV suppressor 2b co-localizes with AGO4 in cytoplasmic and nuclear fractions of Arabidopsis thaliana. Biochemical fractionation of A. thaliana cellular extracts revealed that 2b and AGO4 coexist in multiple size exclusions. 2b transgenic A. thaliana exhibited an enhanced accumulation of 24nt siRNAs from flowering wageningen (FWA and other heterochromatic loci. These plants also exhibited hypo-methylation of an endogenous- as well as transgene-FWA promoter at non-CG sites. In corroboration, both transgenic 2b and CMV infection affected the regulation of transposons which mimics the ago4 phenotype. In conclusion, 2b perturbs plant defense by interfering with AGO4-regulated transcriptional gene silencing.
Abouhaidar, M G
The nucleotide sequences of cDNA clones corresponding to the 3' OH end of papaya mosaic virus RNA have been determined. The 3'-terminal sequence obtained was 900 nucleotides in length, excluding the poly(A) tail, and contained an open reading frame capable of giving rise to a protein of 214 amino acid residues with an Mr of 22930. This protein was identified as the viral capsid protein. The 3' non-coding region of PMV genome RNA was about 121 nucleotides long [excluding the poly(A) tail] and homologous to the complementary sequence of the non-coding region at the 5' end of PMV RNA. A long open reading frame was also found in the predicted 5' end region of the negative strand.
Hamera, Sadia; Yan, Youngsheng; Song, Xiaoguang; Chaudhary, Safee Ullah; Murtaza, Iram; Su, Lei; Tariq, Muhammad; Chen, Xiaoying; Fang, Rongxiang
The Cucumber mosaic virus (CMV) suppressor 2b co-localizes with AGO4 in cytoplasmic and nuclear fractions of Arabidopsis thaliana Biochemical fractionation of A. thaliana cellular extracts revealed that 2b and AGO4 coexist in multiple size exclusions. 2b transgenic A. thaliana exhibited an enhanced accumulation of 24nt siRNAs from flowering wageningen (FWA) and other heterochromatic loci. These plants also exhibited hypo-methylation of an endogenous- as well as transgene-FWA promoter at non-CG sites. In corroboration, both transgenic 2b and CMV infection affected the regulation of transposons which mimics the ago4 phenotype. In conclusion, 2b perturbs plant defense by interfering with AGO4-regulated transcriptional gene silencing.
徐莉; 顾国华; 葛红; 季桦; 高小红
大豆花叶病毒(Soybean mosaic virus,SMV)病是在世界范围内广泛分布并普遍发生的病毒病害之一,可导致大豆严重减产和种质衰退.从大豆花叶病的症状、危害、SMV的特性、基因组学研究、株系分化、病害流行规律及预测模型等方面综述了近年来国内外SMV的科研方向和进展,旨在为进一步的研究提供依据.
Full Text Available The triple-gene-block protein 3 (TGBp3 of Bamboo mosaic virus (BaMV is an integral endoplasmic reticulum (ER membrane protein which is assumed to form a membrane complex to deliver the virus intracellularly. However, the virus entity that is delivered to plasmodesmata (PD and its association with TGBp3-based complexes are not known. Results from chemical extraction and partial proteolysis of TGBp3 in membrane vesicles revealed that TGBp3 has a right-side-out membrane topology; i.e., TGBp3 has its C-terminal tail exposed to the outer surface of ER. Analyses of the TGBp3-specific immunoprecipitate of Sarkosyl-extracted TGBp3-based complex revealed that TGBp1, TGBp2, TGBp3, capsid protein (CP, replicase and viral RNA are potential constituents of virus movement complex. Substantial co-fractionation of TGBp2, TGBp3 and CP, but not TGBp1, in the early eluted gel filtration fractions in which virions were detected after TGBp3-specific immunoprecipitation suggested that the TGBp2- and TGBp3-based complex is able to stably associate with the virion. This notion was confirmed by immunogold-labeling transmission electron microscopy (TEM of the purified virions. In addition, mutational and confocal microscopy analyses revealed that TGBp3 plays a key role in virus cell-to-cell movement by enhancing the TGBp2- and TGBp3-dependent PD localization of TGBp1. Taken together, our results suggested that the cell-to-cell movement of potexvirus requires stable association of the virion cargo with the TGBp2- and TGBp3-based membrane complex and recruitment of TGBp1 to the PD by this complex.
Tuo, D; Shen, W; Yan, P; Li, Ch; Gao, L; Li, X; Li, H; Zhou, P
Papaya leaf distortion mosaic virus is highly destructive to commercial papaya production. Here, the complete genome sequence was determined for an isolate of papaya leaf distortion mosaic virus, designated PLDMV-DF, infecting the commercialized papaya ringspot virus (PRSV)-resistant transgenic papaya from China. Excluding the 3'-poly (A) tail, the sequence shares high sequence identity to several PLDMV isolates from Taiwan and Japan and is phylogenetically most closely related to the isolate from Japan. Infection of PLDMV-DF in transgenic PRSV-resistant papaya may indicate emergence of this disease in genetically engineered plants. The reported sequence for this isolate may help generate bi-transgenic papaya resistant to PRSV and PLDMV.
Lim, Hyoun-Sub; Vaira, Anna Maria; Bae, Hanhong; Bragg, Jennifer N; Ruzin, Steven E; Bauchan, Gary R; Dienelt, Margaret M; Owens, Robert A; Hammond, John
Cell-to-cell movement of potexviruses requires coordinated action of the coat protein and triple gene block (TGB) proteins. The structural properties of Alternanthera mosaic virus (AltMV) TGB3 were examined by methods differentiating between signal peptides and transmembrane domains, and its subcellular localization was studied by Agrobacterium-mediated transient expression and confocal microscopy. Unlike potato virus X (PVX) TGB3, AltMV TGB3 was not associated with the endoplasmic reticulum, and accumulated preferentially in mesophyll cells. Deletion and site-specific mutagenesis revealed an internal signal VL(17,18) of TGB3 essential for chloroplast localization, and either deletion of the TGB3 start codon or alteration of the chloroplast-localization signal limited cell-to-cell movement to the epidermis, yielding a virus that was unable to move into the mesophyll layer. Overexpression of AltMV TGB3 from either AltMV or PVX infectious clones resulted in veinal necrosis and vesiculation at the chloroplast membrane, a cytopathology not observed in wild-type infections. The distinctive mesophyll and chloroplast localization of AltMV TGB3 highlights the critical role played by mesophyll targeting in virus long-distance movement within plants.
Chávez-Calvillo, Gabriela; Contreras-Paredes, Carlos A; Mora-Macias, Javier; Noa-Carrazana, Juan C; Serrano-Rubio, Angélica A; Dinkova, Tzvetanka D; Carrillo-Tripp, Mauricio; Silva-Rosales, Laura
Antagonism between unrelated plant viruses has not been thoroughly described. Our studies show that two unrelated viruses, papaya ringspot virus (PRSV) and papaya mosaic virus (PapMV) produce different symptomatic outcomes during mixed infection depending on the inoculation order. Synergism occurs in plants infected first with PRSV or in plants infected simultaneously with PRSV and PapMV, and antagonism occurs in plants infected first with PapMV and later inoculated with PRSV. During antagonism, elevated pathogenesis-related (PR-1) gene expression and increased reactive oxygen species production indicated the establishment of a host defense resulting in the reduction in PRSV titers. Polyribosomal fractioning showed that PRSV affects translation of cellular eEF1α, PR-1, β-tubulin, and PapMV RNAs in planta, suggesting that its infection could be related to an imbalance in the translation machinery. Our data suggest that primary PapMV infection activates a defense response against PRSV and establishes a protective relationship with the papaya host.
Denis, Jérôme; Majeau, Nathalie; Acosta-Ramirez, Elizabeth; Savard, Christian; Bedard, Marie-Claude; Simard, Sabrina; Lecours, Katia; Bolduc, Marilène; Pare, Christine; Willems, Bernard; Shoukry, Naglaa; Tessier, Philippe; Lacasse, Patrick; Lamarre, Alain; Lapointe, Réjean; Lopez Macias, Constantino; Leclerc, Denis
Plant-virus-based vaccines have emerged as a promising avenue in vaccine development. This report describes the engineering of an innovative vaccine platform using the papaya mosaic virus (PapMV) capsid protein (CP) as a carrier protein and a C-terminal fused hepatitis C virus (HCV) E2 epitope as the immunogenic target. Two antigen organizations of the PapMV-based vaccines were tested: a virus-like-particle (VLP; PapMVCP-E2) and a monomeric form (PapMVCP(27-215)-E2). While the two forms of the vaccine were both shown to be actively internalized in vitro in bone-marrow-derived antigen presenting cells (APCs), immunogenicity was demonstrated to be strongly dependent on antigen organization. Indeed, C3H/HeJ mice injected twice with the multimeric VLP vaccine showed a long-lasting humoral response (more than 120 days) against both the CP and the fused HCV E2 epitope. The antibody profile (production of IgG1, IgG2a, IgG2b, IgG3) suggests a Th1/Th2 response. Immunogenicity of the PapMV vaccine platform was not observed when the monomer PapMVCP-E2 was injected. These results demonstrate for the first time the potential of the PapMV vaccine platform and the critical function of multimerization in its immunogenicity.
Acharya, Sefali; Sengupta, Soumika; Patro, Sunita; Purohit, Sukumar; Samal, Sabindra K; Maiti, Indu B; Dey, Nrisingha
We developed an efficient chimeric promoter, MUASMSCP, with enhanced activity and salicylic acid (SA)/abscisic acid (ABA) inducibility, incorporating the upstream activation sequence (UAS) of Mirabilis mosaic virus full-length transcript (MUAS, -297 to -38) to the 5' end of Mirabilis mosaic virus sub-genomic transcript (MSCP, -306 to -125) promoter-fragment containing the TATA element. We compared the transient activity of the MUASMSCP promoter in tobacco/Arabidopsis protoplasts and in whole plant (Petunia hybrida) with the same that obtained from CaMV35S and MUAS35SCP promoters individually. The MUASMSCP promoter showed 1.1 and 1.5 times stronger GUS-activities over that obtained from MUAS35SCP and CaMV35S promoters respectively, in tobacco (Xanthi Brad) protoplasts. In transgenic tobacco (Nicotiana tabacum, var. Samsun NN), the MUASMSCP promoter showed 1.1 and 2.2 times stronger activities than MUAS35SCP and CaMV35S(2) promoters respectively. We observed a fair correlation between MUASMSCP-, MUAS35SCP- and CaMV35S(2)-driven GUS activities with the corresponding uidA-mRNA level in transgenic plants. X-gluc staining of transgenic germinating seed-sections and whole seedlings also support above findings. Protein-extracts made from tobacco protoplasts expressing GFP and human-IL-24 genes driven individually by the MUASMSCP promoter showed enhanced expression of the reporters compared to that obtained from the CaMV35S promoter. Furthermore, MUASMSCP-driven protoplast-derived human IL-24 showed enhanced cell inhibitory activity in DU-145 prostate cancer cells compared to that obtained from the CaMV35S promoter. We propose chimeric MUASMSCP promoter developed in the study could be useful for strong constitutive expression of transgenes in both plant/animal cells and it may become an efficient substitute for CaMV35S/CaMV35S(2) promoter.
Pramesh, D; Mandal, Bikash; Phaneendra, Chigurupati; Muniyappa, V
Croton yellow vein mosaic virus (CYVMV) is a widely occurring begomovirus in Croton bonplandianum, a common weed in the Indian subcontinent. In this study, CYVMV (genus Begomovirus, family Geminiviridae) was transmitted by whiteflies (Bemisia tabaci) to as many as 35 plant species belonging to 11 families, including many vegetables, tobacco varieties, ornamentals and weeds. CYVMV produced bright yellow vein symptoms in croton, whereas in all the other host species, the virus produced leaf curl symptoms. CYVMV produced leaf curl in 13 tobacco species and 22 cultivars of Nicotiana tabacum and resembled tobacco leaf curl virus (TobLCV) in host reactions. However, CYVMV was distinguished from TobLCV in four differential hosts, Ageratum conyzoides, C. bonplandianum, Euphorbia geniculata and Sonchus bracyotis. The complete genome sequences of four isolates originating from northern, eastern and southern India revealed that a single species of DNA-A and a betasatellite, croton yellow vein mosaic betasatellite (CroYVMB) were associated with the yellow vein mosaic disease of croton. The sequence identity among the isolates of CYVMV DNA-A and CroYVMB occurring in diverse plant species was 91.8-97.9 % and 83.3-100 %, respectively. The CYVMV DNA-A and CroYVMB generated through rolling-circle amplification of the cloned DNAs produced typical symptoms of yellow vein mosaic and leaf curling in croton and tomato, respectively. The progeny virus from both the croton and tomato plants was transmitted successfully by B. tabaci. The present study establishes the etiology of yellow vein mosaic disease of C. bonplandianum and provides molecular evidence that a weed-infecting monopartite begomovirus causes leaf curl in tomato.
Hasiów-Jaroszewska, Beata; Paeleman, Anneleen; Ortega-Parra, Nelia; Borodynko, Natasza; Minicka, Julia; Czerwoniec, Anna; Thomma, Bart P H J; Hanssen, Inge M
Recently, Pepino mosaic virus (PepMV) infections causing severe yellowing symptoms in tomato plants have been reported in glasshouse tomato crops. When studying this phenomenon in commercial glasshouses, two different types of yellowing symptoms, occurring in adjacent plants, were distinguished: interveinal leaf yellowing and yellow mosaics. After several weeks, the interveinal leaf yellowing symptoms gradually disappeared and the plant heads became green again, with yellow mosaic patterns on the leaves as an intermediate stage. The sequencing of multiple isolates causing interveinal leaf yellowing identified two point mutations, occurring in positions 155 and 166 of the coat protein (CP), as unique to the yellowing pathotype. Site-directed mutagenesis of infectious clones confirmed that both CP mutations are determinants of the interveinal leaf yellowing symptoms. Sequencing of CP clones from plants or plant parts with the yellow mosaic symptoms resulted in a mixture of wild-type and mutated sequences, whereas sequencing of CP clones from the green heads of recovered plants resulted in only wild-type sequences. Yellow mosaic symptoms could be reproduced by inoculation of an artificial 1:1 mixture of RNA transcripts from the wild-type and mutated infectious clones. These results show that the ratio of mutated versus wild-type sequences can determine the nature and severity of symptom development. The gradual recovery of the plants, which coincides with the disappearance of the yellowing mutations, suggests that selection pressure acts to the advantage of the wild-type virus. Experiments with wild-type and mutated infectious clones showed that reverse mutation events from mutant to wild-type occur and that the wild-type virus does not have a replicative advantage over the mutant. These results suggest that reverse mutation events occur, with subsequent selection pressure acting in favour of the wild-type virus in the growing plant parts, possibly related to a lower
Full Text Available The Cmr1 gene in peppers confers resistance to Cucumber mosaic virus isolate-P0 (CMV-P0. Cmr1 restricts the systemic spread of CMV strain-Fny (CMV-Fny, whereas this gene cannot block the spread of CMV isolate-P1 (CMV-P1 to the upper leaves, resulting in systemic infection. To identify the virulence determinant of CMV-P1, six reassortant viruses and six chimeric viruses derived from CMV-Fny and CMV-P1 cDNA clones were used. Our results demonstrate that the C-terminus of the helicase domain encoded by CMV-P1 RNA1 determines susceptibility to systemic infection, and that the helicase domain contains six different amino acid substitutions between CMV-Fny and CMV-P1(. To identify the key amino acids of the helicase domain determining systemic infection with CMV-P1, we then constructed amino acid substitution mutants. Of the mutants tested, amino acid residues at positions 865, 896, 957, and 980 in the 1a protein sequence of CMV-P1 affected the systemic infection. Virus localization studies with GFP-tagged CMV clones and in situ localization of virus RNA revealed that these four amino acid residues together form the movement determinant for CMV-P1 movement from the epidermal cell layer to mesophyll cell layers. Quantitative real-time PCR revealed that CMV-P1 and a chimeric virus with four amino acid residues of CMV-P1 accumulated more genomic RNA in inoculated leaves than did CMV-Fny, indicating that those four amino acids are also involved in virus replication. These results demonstrate that the C-terminal region of the helicase domain is responsible for systemic infection by controlling virus replication and cell-to-cell movement. Whereas four amino acids are responsible for acquiring virulence in CMV-Fny, six amino acid (positions at 865, 896, 901, 957, 980 and 993 substitutions in CMV-P1 were required for complete loss of virulence in 'Bukang'.
Bashir, Nemat Sokhandan; Kalhor, Mohammad Rasaei; Zarghani, Shaheen Nourinejhad
One hundred and twenty three cucurbit samples with one or more symptoms of leaf mosaic, leaf distortion, fruit mosaic, stunting, mottling and yellowing were collected from several locations in the northwest region of Iran. Screening by double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with a cucumber mosaic virus (CMV) polyclonal antibody, produced positive reactions from 13 samples. However, none of these positive samples reacted with a CMV subgroup-II (S-II)-specific monoclonal antibody in a triple antibody sandwich (TAS)-ELSIA. When total RNA from the CMV-infected samples was subjected to reverse transcription polymerase chain reaction (RT-PCR) with a pair of primers corresponding to the flanking regions of the virus coat protein (CP) gene, an expected DNA fragment of about 872 bp was amplified from 10 of the 13 isolates. This fragment covered the CP open reading frame (ORF) plus 92 and 123 bp of the 5' and 3' flanking regions, respectively. Restriction analysis with MspI (HpaII) was done on 9 of the PCR products and revealed a previously described CMV subgroup I (S-I) specific profile (537 and 335 bp fragments) for the isolates B13, B23, B5, SH5, SH17, S342 and S337, and an additional fragment, suggestive of combined profiles, was present for B13, SH5 and S342. Two other isolates, SH12 and B7 had a CMV S-II MspI profile (four visible fragments and a predicted non-visible 28-bp fragment on 2% agarose). Also, BsuRI (HaeIII) did not cut the PCR products characteristic of the CMV S-I specific MspI profile, whereas for the S-II isolates, BsuRI gave two fragments with sizes of approximately 559 and 313 bp. Nucleotide (nt) sequences of clones from the isolates B13, B23, SH5, SH17, S337 and SH12 were determined and aligned with those of previously published CMV strains and isolates. Consensus parsimonious trees constructed on the basis of the whole amplified region (841 nt excluding the primer sequences), CP ORF (nt or deduced amino acid data), or
Butterbach, Patrick; Verlaan, Maarten G; Dullemans, Annette; Lohuis, Dick; Visser, Richard G F; Bai, Yuling; Kormelink, Richard
Tomato yellow leaf curl virus (TYLCV) and related begomoviruses are a major threat to tomato production worldwide and, to protect against these viruses, resistance genes from different wild tomato species are introgressed. Recently, the Ty-1 resistance gene was identified, shown to code for an RNA-dependent RNA polymerase and to be allelic with Ty-3. Here we show that upon TYLCV challenging of resistant lines carrying Ty-1 or Ty-3, low virus titers were detected concomitant with the production of relatively high levels of siRNAs whereas, in contrast, susceptible tomato Moneymaker (MM) revealed higher virus titers but lower amounts of siRNAs. Comparative analysis of the spatial genomic siRNA distribution showed a consistent and subtle enrichment for siRNAs derived from the V1 and C3 genes in Ty-1 and Ty-3. In plants containing Ty-2 resistance the virus was hardly detectable, but the siRNA profile resembled the one observed in TYLCV-challenged susceptible tomato (MM). Furthermore, a relative hypermethylation of the TYLCV V1 promoter region was observed in genomic DNA collected from Ty-1 compared with that from (MM). The resistance conferred by Ty-1 was also effective against the bipartite tomato severe rugose begomovirus, where a similar genome hypermethylation of the V1 promoter region was discerned. However, a mixed infection of TYLCV with cucumber mosaic virus compromised the resistance. The results indicate that Ty-1 confers resistance to geminiviruses by increasing cytosine methylation of viral genomes, suggestive of enhanced transcriptional gene silencing. The mechanism of resistance and its durability toward geminiviruses under natural field conditions is discussed.
Full Text Available RNA silencing is an evolutionarily conserved antiviral mechanism, through which virus-derived small interfering RNAs (vsiRNAs playing roles in host antiviral defence are produced in virus-infected plant. Deep sequencing technology has revolutionized the study on the interaction between virus and plant host through the analysis of vsiRNAs profile. However, comparison of vsiRNA profiles in different tissues from a same host plant has been rarely reported. In this study, the profiles of virus-derived small interfering RNAs (vsiRNAs from leaves and fruits of Lagenaria siceraria plants infected with Cucumber green mottle mosaic virus (CGMMV were comprehensively characterized and compared. Many more vsiRNAs were present in infected leaves than in fruits. vsiRNAs from both leaves and fruits were mostly 21- and 22-nt in size as previously described in other virus-infected plants. Interestingly, vsiRNAs were predominantly produced from the viral positive strand RNAs in infected leaves, whereas in infected fruits they were derived equally from the positive and negative strands. Many leaf-specific positive vsiRNAs with lengths of 21-nt (2,058 or 22-nt (3,996 were identified but only six (21-nt and one (22-nt positive vsiRNAs were found to be specific to fruits. vsiRNAs hotspots were only present in the 5’-terminal and 3’-terminal of viral positive strand in fruits, while multiple hotspots were identified in leaves. Differences in GC content and 5'-terminal nucleotide of vsiRNAs were also observed in the two organs. To our knowledge, this provides the first high-resolution comparison of vsiRNA profiles between different tissues of the same host plant.
José Albersio Araujo Lima
Full Text Available Cowpea (Vigna unguiculata is an important crop of the traditional agriculture system in the Northeast of Brazil. It can be infected by more than 20 virus species and Cowpea severe mosaic virus (CPSMV is one of the most important pathogens that naturally infect cowpea in Brazil. Several CPSMV isolates were obtained and characterized in the Plant Virus Laboratory at the Federal University of Ceará: CPSMV-CE - the first characterized isolate of the virus obtained from cowpea in the State of Ceará; CPSMV-AL - isolated from cowpea in Alagoas; CPSMV-PE - isolated from cowpea in Pernambuco; CPSMV-PR - obtained from soybean (Glycine max in Paraná and CPSMV-CROT - isolated from Crotalaria paulinea, in Maranhão. An isolate of CPSMV with the property to infect the cv. Macaibo, a cowpea cultivar immune to most of CPSMV isolates was also biologically and serologically characterized as a new strain of the virus (CPSMV-MC. The CPSMV-MC was isolated in January 1990 and has been evaluated over 20 years by host range studies and maintenance in vivo by periodical mechanical inoculations in cowpea. The results of this periodical evaluation revealed that the biological integrity and the serological properties of CPSMV-MC were preserved over 20 years, indicating that the genetic preservation of a virus strain could occur over the years. Molecular studies involving part of the coat protein (CP gene of CPSMV-MC and five other Brazilian CPSMV isolates indicated a high degree of conservation, with 92-100% nucleotide sequence identity among the isolates.O feijão-caupi (Vigna unguiculata é uma cultura do sistema tradicional do Nordeste do Brasil, que pode ser infetada por mais de 20 espécies de vírus, sendo o vírus do mosaico severo do caupi (Cowpea severe mosaic virus, CPSMV um dos mais importantes patógenos que infeta naturalmente essa leguminosa no Brasil. Vários isolados do CPSMV foram obtidos e caracterizados no Laboratório de Virologia Vegetal da UFC
Full Text Available The potato/tomato psyllid, Bactericera cockerelli (B. cockerelli, is an important plant pest and the vector of the phloem-limited bacterium Candidatus Liberibacter psyllaurous (solanacearum, which is associated with the zebra chip disease of potatoes. Previously, we reported induction of RNA interference effects in B. cockerelli via in vitro-prepared dsRNA/siRNAs after intrathoracic injection, and after feeding of artificial diets containing these effector RNAs. In order to deliver RNAi effectors via plant hosts and to rapidly identify effective target sequences in plant-feeding B. cockerelli, here we developed a plant virus vector-based in planta system for evaluating candidate sequences. We show that recombinant Tobacco mosaic virus (TMV containing B. cockerelli sequences can efficiently infect and generate small interfering RNAs in tomato (Solanum lycopersicum, tomatillo (Physalis philadelphica and tobacco (Nicotiana tabacum plants, and more importantly delivery of interfering sequences via TMV induces RNAi effects, as measured by actin and V-ATPase mRNA reductions, in B. cockerelli feeding on these plants. RNAi effects were primarily detected in the B. cockerelli guts. In contrast to our results with TMV, recombinant Potato virus X (PVX and Tobacco rattle virus (TRV did not give robust infections in all plants and did not induce detectable RNAi effects in B. cockerelli. The greatest RNA interference effects were observed when B. cockerelli nymphs were allowed to feed on leaf discs collected from inoculated or lower expanded leaves from corresponding TMV-infected plants. Tomatillo plants infected with recombinant TMV containing B. cockerelli actin or V-ATPase sequences also showed phenotypic effects resulting in decreased B. cockerelli progeny production as compared to plants infected by recombinant TMV containing GFP. These results showed that RNAi effects can be achieved in plants against the phloem feeder, B. cockerelli, and the TMV
Ying Wen Huang
Full Text Available Host factors play crucial roles in the replication of plus-strand RNA viruses. In this report, a heat shock protein 90 homologue of Nicotiana benthamiana, NbHsp90, was identified in association with partially purified replicase complexes from BaMV-infected tissue, and shown to specifically interact with the 3' untranslated region (3' UTR of BaMV genomic RNA, but not with the 3' UTR of BaMV-associated satellite RNA (satBaMV RNA or that of genomic RNA of other viruses, such as Potato virus X (PVX or Cucumber mosaic virus (CMV. Mutational analyses revealed that the interaction occurs between the middle domain of NbHsp90 and domain E of the BaMV 3' UTR. The knockdown or inhibition of NbHsp90 suppressed BaMV infectivity, but not that of satBaMV RNA, PVX, or CMV in N. benthamiana. Time-course analysis further revealed that the inhibitory effect of 17-AAG is significant only during the immediate early stages of BaMV replication. Moreover, yeast two-hybrid and GST pull-down assays demonstrated the existence of an interaction between NbHsp90 and the BaMV RNA-dependent RNA polymerase. These results reveal a novel role for NbHsp90 in the selective enhancement of BaMV replication, most likely through direct interaction with the 3' UTR of BaMV RNA during the initiation of BaMV RNA replication.
Full Text Available Abstract Background In order to obtain an initial and preliminary understanding of host and nonhost resistance in the initial step of potyvirus replication, both positive and negative Sugarcane mosaic virus (SCMV strands where traced in inoculated and systemic leaves in host and nonhost resistant maize and sugarcane for one Mexican potyviral isolate (SCMV-VER1. Intermediary replication forms, such as the negative viral strand, seem to only move a short distance as surveyed by RT-PCR analysis and ELISA in different leaves. Virus purification was also done in leaves and stems. Results Susceptible maize plants allowed for viral SCMV replication, cell-to-cell, and long distance movement, as indicated by the presence of the coat protein along the plant. In the host resistant maize plants for the SCMV-VER1 isolate, the virus was able to establish the disease though the initial steps of virus replication, as detected by the presence of negative strands, in the basal area of the inoculated leaves at six and twelve days post inoculation. The nonhost sugarcane for SCMV-VER1 and the host sugarcane for SCMV-CAM6 also allowed the initial steps of viral replication for the VER1 isolate in the local inoculated leaf. SCMV-VER1 virions could be extracted from stems of susceptible maize with higher titers than leaves. Conclusion Nonhost and host resistance allow the initial steps of potyvirus SCMV replication, as shown by the negative strands' presence. Furthermore, both hosts allow the negative viral strands' local movement, but not their systemic spread through the stem. The presence of larger amounts of extractable virions from the stem (as compared to the leaves in susceptible maize lines suggests their long distance movement as assembled particles. This will be the first report suggesting the long distance movement of a monocot potyvirus as a virion.
Chen, Shaoning; Gu, Hao; Wang, Xiaoming; Chen, Jishuang; Zhu, Weimin
A multiplex reverse-transcription polymerase chain reaction (RT-PCR) protocol was developed for simultaneous detection and discrimination of subgroups of Cucumber mosaic virus (CMV), including its satellite RNA, Tomato mosaic virus (ToMV) and Tobacco mosaic virus (TMV), using 18S rRNA as an internal control. Species- and subgroups-specific primers designed to differentiate CMV subgroups I and II, ToMV and TMV, were assessed using the cDNA clones of viral genomes, CMV satellite RNA and 18S rRNA gene from tomato (Solanum lycopersicum L.) or tobacco (Nicotiana tobacum). Using total RNA extracted from artificial mixture of tomato leaf tissues infected by each virus, the reaction components and cycling parameters were optimized and a multiplex RT-PCR procedure was established. Six fragments of 704, 593, 512, 421, 385, 255 bp, specific to CMV subgroup II, CMV subgroup I, ToMV, TMV, satellite RNA and 18S rRNA, respectively, were simultaneously amplified. The sensitivity of the multiplex RT-PCR method for detecting CMV was 100 times higher than that of double-antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA). This method was successfully used for field detection. Among 141 samples collected from East China through tomato growth seasons, 106 single infections with one of the above isolates were detected and 13 mixed infections were found. The results showed the potential use of this method for investigating the epidemiology of viral diseases infecting tomato.
Guerra-Peraza, O; Kirk, D; Seltzer, V; Veluthambi, K; Schmit, A C; Hohn, T; Herzog, E
Transport of the viral genome into the nucleus is an obligatory step in the replication cycle of plant pararetro- and geminiviruses. In both these virus types, the multifunctional coat protein (CP) is thought to be involved in this process. Here, a green fluorescent protein tagging approach was used to demonstrate nuclear import of the CPs of Rice tungro bacilliform virus (RTBV) and Mungbean yellow mosaic virus--Vigna (MYMV) in Nicotiana plumbaginifolia protoplasts. In both cases, at least two nuclear localization signals (NLSs) were identified and characterized. The NLSs of RTBV CP are located within both N- and C-terminal regions (residues 479KRPK/497KRK and 744KRK/758RRK), and those of MYMV CP within the N-terminal part (residues 3KR and 41KRRR). The MYMV and RTBV CP NLSs resemble classic mono- and bipartite NLSs, respectively. However, the N-terminal MYMV CP NLS and both RTBV CP NLSs show peculiarities in the number and position of basic residues. In vitro pull-down assays revealed interaction of RTBV and MYMV CPs with the nuclear import factor importin alpha, suggesting that both CPs are imported into the nucleus via an importin alpha-dependent pathway. The possibility that this pathway could serve for docking of virions to the nucleus is discussed.
Bhat, A I; Siljo, A; Deeshma, K P
The loop-mediated isothermal amplification (LAMP) assay for Piper yellow mottle virus and the reverse transcription (RT) LAMP assay for Cucumber mosaic virus each consisted of a set of five primers designed against the conserved sequences in the viral genome. Both RNA and DNA isolated from black pepper were used as a template for the assay. The results were assessed visually by checking turbidity, green fluorescence and pellet formation in the reaction tube and also by gel electrophoresis. The assay successfully detected both viruses in infected plants whereas no cross-reactions were recorded with healthy plants. Optimum conditions for successful amplification were determined in terms of the concentrations of magnesium sulphate and betaine, temperature, and duration. The detection limit for both LAMP and RT-LAMP was up to 100 times that for conventional PCR and up to one-hundredth of that for real-time PCR. The optimal conditions arrived at were validated by testing field samples of infected vines of three species from different regions.
Thompson, Jeremy R; Langenhan, Jamie L; Fuchs, Marc; Perry, Keith L
In the early 2000s an epidemic of cucumber mosaic virus (CMV) spread within the Midwestern and Eastern US affecting snap and dry bean (Phaseolus vulgaris L.) cultivation. Fifty one CMV isolates from this period were partially characterized from varied hosts by sequencing a section from each of the three genomic RNAs. Aside from one subgroup II strain from pepper, all isolates, including those from snap bean, fell within the IA subgroup. The nucleotide sequence diversity of virus populations sampled at multiple sites and at different years was significantly higher than that of a population from single site in a single year, although in general the number of polymorphisms was low (virus infection on plant growth. Inoculations with pseudorecombinants derived from Bn57 and the non-bean infecting strain Fny confirmed RNA2 as a specific determinant for snap bean infection. Bn57, along with almost all isolates identified in this study contained the Y631 locus in the 2a protein, a determinant for systemic infection in bean. The presence of this locus extended to all non-bean hosts except two pepper infecting isolates. Infection by Bn57 in snap bean had a significant effect on pod number and mass with a 55 and 41 percent reduction in greenhouse assays, respectively. To our knowledge Bn57 is the first CMV strain isolated from P. vulgaris to be fully sequenced and cloned, providing a useful tool for analyses of CMV-host interactions.
A seed-transmitted virus has consistently caused significant economic damage to barley crops in Korea in recent years, and may be increasing because many farmers save seed for replanting. Because some barley seed is imported, there is the potential for introduction of new seed-transmitted viruses, c...
Many RNA viruses rely on internal ribosome entry site (IRES) elements to deviate from the canonical cap-dependent translation mechanism. In contrast to the well-defined IRES elements found in animal viruses, plant viral IRESes identified to date reportedly consist of relatively short, ill-defined se...
Li, Junmin; Zheng, Hongying; Zhang, Chenhua; Han, Kelei; Wang, Shu; Peng, Jiejun; Lu, Yuwen; Zhao, Jinping; Xu, Pei; Wu, Xiaohua; Li, Guojing; Chen, Jianping; Yan, Fei
RNA silencing is an evolutionarily conserved antiviral mechanism, through which virus-derived small interfering RNAs (vsiRNAs) playing roles in host antiviral defense are produced in virus-infected plant. Deep sequencing technology has revolutionized the study on the interaction between virus and plant host through the analysis of vsiRNAs profile. However, comparison of vsiRNA profiles in different tissues from a same host plant has been rarely reported. In this study, the profiles of vsiRNAs from leaves and fruits of Lagenaria siceraria plants infected with Cucumber green mottle mosaic virus (CGMMV) were comprehensively characterized and compared. Many more vsiRNAs were present in infected leaves than in fruits. vsiRNAs from both leaves and fruits were mostly 21- and 22-nt in size as previously described in other virus-infected plants. Interestingly, vsiRNAs were predominantly produced from the viral positive strand RNAs in infected leaves, whereas in infected fruits they were derived equally from the positive and negative strands. Many leaf-specific positive vsiRNAs with lengths of 21-nt (2058) or 22-nt (3996) were identified but only six (21-nt) and one (22-nt) positive vsiRNAs were found to be specific to fruits. vsiRNAs hotspots were only present in the 5'-terminal and 3'-terminal of viral positive strand in fruits, while multiple hotspots were identified in leaves. Differences in GC content and 5'-terminal nucleotide of vsiRNAs were also observed in the two organs. To our knowledge, this provides the first high-resolution comparison of vsiRNA profiles between different tissues of the same host plant.
Full Text Available This study was conducted to evaluate the effectiveness of plant growth promoting rhizobacteria (PGPR in protecting chillipepper plant from infection of cucumber mosaic virus (CMV and chilli veinal mottle virus (ChiVMV. Seven isolates of PGPR, i.e. BC1, BTP2H, BTP3G, BTP3O BTP1, BTP2D, and T1F were applied as seed treatment and soil drench. Plants height, number of branch, and fruits weight were measured every one and ten weeks after virus inoculation. Virus concentration in plants and disease incidence were confirmed by enzyme-linked immunosorbant assay (ELISA. Results showed that inoculation with PGPR improved the seed germination. Eight days after sowing, the percentage of PGPR treated seed germination reached 50-84%; whereas those of untreated seed reached only 18%. In general, PGPR treatment significantly reduced (p < 0.05 the effect of virus infection on plant growth. Two PGPR isolates, i.e. BTP1 and BTP2H, maintained fruit weight of infected plants as good as those of healthy plants. Based on ELISA, PGPR was able to inhibit the disease incidence. The BTP3O and BTP2D isolates even protected the plant from ChiVMV infection. Concentration of salicylic acid and peroxidase were relatively higher on plants treated with PGPR than those without PGPR treatment. This gave an indication that PGPR may act as induction agents for systemic acquired resistance. Therefore, PGPR treatment is a promising strategy to control viral diseases on chillipepper.
Cheng, Yu-Qin; Liu, Zhong-Mei; Xu, Jian; Zhou, Tao; Wang, Meng; Chen, Yu-Ting; Li, Huai-Fang; Fan, Zai-Feng
Symptom development of a plant viral disease is a result of molecular interactions between the virus and its host plant; thus, the elucidation of specific interactions is a prerequisite to reveal the mechanism of viral pathogenesis. Here, we show that the chloroplast precursor of ferredoxin-5 (Fd V) from maize (Zea mays) interacts with the multifunctional HC-Pro protein of sugar cane mosaic virus (SCMV) in yeast, Nicotiana benthamiana cells and maize protoplasts. Our results demonstrate that the transit peptide rather than the mature protein of Fd V precursor could interact with both N-terminal (residues 1-100) and C-terminal (residues 301-460) fragments, but not the middle part (residues 101-300), of HC-Pro. In addition, SCMV HC-Pro interacted only with Fd V, and not with the other two photosynthetic ferredoxin isoproteins (Fd I and Fd II) from maize plants. SCMV infection significantly downregulated the level of Fd V mRNA in maize plants; however, no obvious changes were observed in levels of Fd I and Fd II mRNA. These results suggest that SCMV HC-Pro interacts specifically with maize Fd V and that this interaction may disturb the post-translational import of Fd V into maize bundle-sheath cell chloroplasts, which could lead to the perturbation of chloroplast structure and function.
Laliberté Gagné, M E; Lecours, K; Gagné, S; Leclerc, D
Papaya mosaic virus (PapMV) coat protein (CP) in Escherichia coli was previously showed to self-assemble in nucleocapsid-like particles (NLPs) that were similar in shape and appearance to the native virus. We have also shown that a truncated CP missing the N-terminal 26 amino acids is monomeric and loses its ability to bind RNA. It is likely that the N-terminus of the CP is important for the interaction between the subunits in self-assembly into NLPs. In this work, through deletion and mutation analysis, we have shown that the deletion of 13 amino acids is sufficient to generate the monomeric form of the CP. Furthermore, we have shown that residue F13 is critical for self-assembly of the CP subunits into NLPs. The replacement of F13 with hydrophobic residues (L or Y) generated mutated forms of the CP that were able to self-assemble into NLPs. However, the replacement of F13 by A, G, R, E or S was detrimental to the self-assembly of the protein into NLPs. We concluded that a hydrophobic interaction at the N-terminus is important to ensure self-assembly of the protein into NLPs. We also discuss the importance of F13 for assembly of other members of the potexvirus family.
Kao, C Cheng; Ni, Peng; Hema, Masarapu; Huang, Xinlei; Dragnea, Bogdan
The Brome mosaic virus (BMV) coat protein (CP) accompanies the three BMV genomic RNAs and the subgenomic RNA into and out of cells in an infection cycle. In addition to serving as a protective shell for all of the BMV RNAs, CP plays regulatory roles during the infection process that are mediated through specific binding of RNA elements in the BMV genome. One regulatory RNA element is the B box present in the 5' untranslated region (UTR) of BMV RNA1 and RNA2 that play important roles in the formation of the BMV replication factory, as well as the regulation of translation. A second element is within the tRNA-like 3' UTR of all BMV RNAs that is required for efficient RNA replication. The BMV CP can also encapsidate ligand-coated metal nanoparticles to form virus-like particles (VLPs). This update summarizes the interaction between the BMV CP and RNAs that can regulate RNA synthesis, translation and RNA encapsidation, as well as the formation of VLPs.
Chen, Yanfei; Chen, Jishuang; Zhang, Huarong; Tang, Xiangshan; Du, Zhiyou
Cucumber mosaic virus (CMV) is a tripartite RNA virus and has been divided into three subgroups, named IA, IB, and II. Some studies have found a few natural reassortants between CMV subgroups, although reassortment between CMV subgroups is infrequent. In our present work, a CMV reassortant, named CMV-Tsh, was obtained from a tomato plant. The complete sequence of CMV-Tsh genomic RNAs has been determined and analyzed. The results of sequence comparisons and phylogenetic analyses revealed that CMV-Tsh RNAs 1 and 3 are derived from one or two CMV subgroup II strain(s), while RNA2 is derived from a CMV subgroup IA strain. A PCR and restriction enzyme analysis-based method was developed to analyze the possibility of mixed infection by CMV strains of different subgroup in the CMV-Tsh-infected tomato plant. The results of the restriction enzyme analysis proved that CMV-Tsh is the unique strain in the tomato plant. Taken together, CMV-Tsh is a natural reassortant having CMV subgroup IA RNA2 and subgroup II RNAs 1 and 3.
Cuenca, Sol; Mansilla, Carmen; Aguado, Marta; Yuste-Calvo, Carmen; Sánchez, Flora; Sánchez-Montero, Jose M; Ponz, Fernando
Elongated flexuous plant viral nanoparticles (VNPs) represent an interesting platform for developing different applications in nanobiotechnology. In the case of potyviruses, the virion external surface is made up of helically arrayed domains of the viral structural coat protein (CP), repeated over 2000 times, in which the N- and C-terminal domains of each CP are projected toward the exterior of the external virion surface. These characteristics provide a chemical environment rich in functional groups susceptible to chemical conjugations. We have conjugated Candida antarctica lipase B (CALB) onto amino groups of the external surface of the potyvirus turnip mosaic virus (TuMV) using glutaraldehyde as a conjugating agent. Using this approach, TuMV virions were transformed into scaffolds for CALB nanoimmobilization. Analysis of the resulting structures revealed the formation of TuMV nanonets onto which large CALB aggregates were deposited. The functional enzymatic characterization of the CALB-bearing TuMV nanonets showed that CALB continued to be active in the nanoimmobilized form, even gaining an increased relative specific activity, as compared to the non-immobilized form. These novel virus-based nanostructures may provide a useful new approach to enzyme nanoimmobilization susceptible to be industrially exploited.
Ooi, Aikseng; Tan, Sianghee; Mohamed, Rosmawati; Rahman, Noorsaadah Abdul; Othman, Rofina Yasmin
A cucumber green mosaic mottle virus (CGMMV) full-length clone was developed for the expression of Hepatitis B surface antigen (HBsAg). The expression of the surface displayed HBsAg by the chimeric virus was confirmed through a double antibody sandwich ELISA. Assessment of the coat protein composition of the chimeric virus particles by SDS-PAGE analysis showed that 50% of the coat proteins were fused to the HBsAg. Biological activity of the expressed HBsAg was assessed through the stimulation of in vitro antibody production by cultured peripheral blood mononuclear cells (PBMC). PBMC that were cultured in the presence of the chimeric virus showed up to an approximately three-fold increase in the level of anti HBsAg immunoglobulin thus suggesting the possible use of this new chimeric virus as an effective Hepatitis B vaccine.
The 5'-nontranslated leader(omega sequence) of Tobacco mosaic virus(TMV) was used as a translational enhancer sequence in the expression of the hepatitis B surface antigen(HBsAg) gene in transgenic ginseng callus cultures.The adr subtype HBsAg gene was placed under the control of the Cauliflower mosaic virus(CaMV) 35S promoter linking to the TMV leader sequence. The antisense omega sequence was used in a control construct. The resulting constructs cloned in the binary vector pBI121 were used to transform the ginseng callus tissue via the Agrobacterium-mediated procedure. The integration and expression of the HBsAg gene were evaluated by PCR and western blot, respectively. Enzyme-linked immunoassays(ELISA) using a monoclonal antibody directed against human serum-derived HBsAg revealed a three to four-fold enhanced expression of HBsAg in ginseng cells conferred by the TMV omega element.
Mi Sa Vo Phan
Full Text Available Molecular and biological characteristics of an isolate of Cucumber mosaic virus (CMV from Glycine soja (wild soybean, named as CMV-209, was examined in this study. Comparison of nucleotide sequences and phylogenetic analyses of CMV-209 with the other CMV strains revealed that CMV-209 belonged to CMV subgroup I. However, CMV-209 showed some genetic distance from the CMV strains assigned to subgroup IA or subgroup IB. Infectious full-genome cDNA clones of CMV-209 were generated under the control of the Cauliflower mosaic virus 35S promoter. Infectivity of the CMV-209 clones was evaluated in Nicotiana benthamiana and various legume species. Our assays revealed that CMV-209 could systemically infect Glycine soja (wild soybean and Pisum sativum (pea as well as N. benthamiana, but not the other legume species.
Ntui, Valentine Otang
Cassava ranks fifth among the starch producing crops of the world, its annual bioethanol yield is higher than for any other crop. Cassava cultivar KU50, the most widely grown cultivar for non-food purposes is susceptible to Sri Lankan cassava mosaic virus (SLCMV). The objective of this work was to engineer resistance to SLCMV by RNA interference (RNAi) in order to increase biomass yield, an important aspect for bioethanol production. Here, we produced transgenic KU50 lines expressing dsRNA homologous to the region between the AV2 and AV1 of DNA A of SLCMV. High level expression of dsRNA of SLCMV did not induce any growth abnormality in the transgenic plants. Transgenic lines displayed high levels of resistance to SLCMV compared to the wild-type plants and no virus load could be detected in uninoculated new leaves of the infected resistant lines after PCR amplification and RT-PCR analysis. The agronomic performance of the transgenic lines was unimpaired after inoculation with the virus as the plants presented similar growth when compared to the mock inoculated control plants and revealed no apparent reduction in the amount and weight of tubers produced. We show that the resistance is correlated with post-transcriptional gene silencing because of the production of transgene specific siRNA. The results demonstrate that transgenic lines exhibited high levels of resistance to SLCMV. This resistance coupled with the desirable yield components in the transgenic lines makes them better candidates for exploitation in the production of biomass as well as bioethanol.
Yang, Ting; Meng, Yao; Chen, Li-Juan; Lin, Hong-Hui; Xi, De-Hui
Alpha-momorcharin (α-MMC) is a type-I ribosome inactivating protein with a molecular weight of 29 kDa that is found in Momordica charantia, and has been shown to be effective against a broad range of human viruses as well as having anti-tumor activities. However, the role of endogenous α-MMC under viral infection and the mechanism of the anti-viral activities of α-MMC in plants are still unknown. To study the effect of α-MMC on plant viral defense and how α-MMC increases plant resistance to virus, the M. charantia-cucumber mosaic virus (CMV) interaction system was investigated. The results showed that the α-MMC level was positively correlated with the resistance of M. charantia to CMV. α-MMC treatment could alleviate photosystem damage and enhance the ratio of glutathione/glutathione disulfide in M. charantia under CMV infection. The relationship of α-MMC and defense related phytohormones, and their roles in plant defense were further investigated. α-MMC treatment led to a significant increase of jasmonic acid (JA) and vice versa, while there was no obvious relevance between salicylic acid and α-MMC. In addition, reactive oxygen species (ROS) were induced in α-MMC-pretreated plants, in a similar way to the ROS burst in JA-pretreated plants. The production of ROS in both ibuprofen (JA inhibitor) and (α-MMC+ibuprofen)-pretreated plants was reduced markedly, leading to a greater susceptibility of M. charantia to CMV. Our results indicate that the anti-viral activities of α-MMC in M. charantia may be accomplished through the JA related signaling pathway.
Full Text Available Alpha-momorcharin (α-MMC is a type-I ribosome inactivating protein (RIP with a molecular weight of 29kDa that is found in Momordica charantia, and has been shown to be effective against a broad range of human viruses as well as having anti-tumor activities. However, the role of endogenous α-MMC under viral infection and the mechanism of the anti-viral activities of α-MMC in plants are still unknown. To study the effect of α-MMC on plant viral defense and how α-MMC increases plant resistance to virus, the M. charantia–cucumber mosaic virus (CMV interaction system was investigated. The results showed that the α-MMC level was positively correlated with the resistance of M. charantia to CMV. α-MMC treatment could alleviate photosystem damage and enhance the ratio of glutathione/glutathione disulfide (GSH/GSSH in M. charantia under CMV infection. The relationship of α-MMC and defense related phytohormones, and their roles in plant defense were further investigated. α-MMC treatment led to a significant increase of jasmonic acid (JA and vice versa, while there was no obvious relevance between salicylic acid (SA and α-MMC. In addition, reactive oxygen species (ROS were induced in α-MMC-pretreated plants, in a similar way to the ROS burst in JA-pretreated plants. The production of ROS in both ibuprofen (JA inhibitor and (α-MMC+ibuprofen-pretreated plants was reduced markedly, leading to a greater susceptibility of M. charantia to CMV. Our results indicate that the anti-viral activities of α-MMC in M. charantia may be accomplished through the JA related signaling pathway.
Valentine Otang Ntui
Full Text Available Cassava ranks fifth among the starch producing crops of the world, its annual bioethanol yield is higher than for any other crop. Cassava cultivar KU50, the most widely grown cultivar for non-food purposes is susceptible to Sri Lankan cassava mosaic virus (SLCMV. The objective of this work was to engineer resistance to SLCMV by RNA interference (RNAi in order to increase biomass yield, an important aspect for bioethanol production. Here, we produced transgenic KU50 lines expressing dsRNA homologous to the region between the AV2 and AV1 of DNA A of SLCMV. High level expression of dsRNA of SLCMV did not induce any growth abnormality in the transgenic plants. Transgenic lines displayed high levels of resistance to SLCMV compared to the wild-type plants and no virus load could be detected in uninoculated new leaves of the infected resistant lines after PCR amplification and RT-PCR analysis. The agronomic performance of the transgenic lines was unimpaired after inoculation with the virus as the plants presented similar growth when compared to the mock inoculated control plants and revealed no apparent reduction in the amount and weight of tubers produced. We show that the resistance is correlated with post-transcriptional gene silencing because of the production of transgene specific siRNA. The results demonstrate that transgenic lines exhibited high levels of resistance to SLCMV. This resistance coupled with the desirable yield components in the transgenic lines makes them better candidates for exploitation in the production of biomass as well as bioethanol.
Ntui, Valentine Otang; Kong, Kynet; Khan, Raham Sher; Igawa, Tomoko; Janavi, Gnanaguru Janaky; Rabindran, Ramalingam; Nakamura, Ikuo; Mii, Masahiro
Cassava ranks fifth among the starch producing crops of the world, its annual bioethanol yield is higher than for any other crop. Cassava cultivar KU50, the most widely grown cultivar for non-food purposes is susceptible to Sri Lankan cassava mosaic virus (SLCMV). The objective of this work was to engineer resistance to SLCMV by RNA interference (RNAi) in order to increase biomass yield, an important aspect for bioethanol production. Here, we produced transgenic KU50 lines expressing dsRNA homologous to the region between the AV2 and AV1 of DNA A of SLCMV. High level expression of dsRNA of SLCMV did not induce any growth abnormality in the transgenic plants. Transgenic lines displayed high levels of resistance to SLCMV compared to the wild-type plants and no virus load could be detected in uninoculated new leaves of the infected resistant lines after PCR amplification and RT-PCR analysis. The agronomic performance of the transgenic lines was unimpaired after inoculation with the virus as the plants presented similar growth when compared to the mock inoculated control plants and revealed no apparent reduction in the amount and weight of tubers produced. We show that the resistance is correlated with post-transcriptional gene silencing because of the production of transgene specific siRNA. The results demonstrate that transgenic lines exhibited high levels of resistance to SLCMV. This resistance coupled with the desirable yield components in the transgenic lines makes them better candidates for exploitation in the production of biomass as well as bioethanol.
Silva, Fabiana Lima; Fischer, Dominique Corinne Hermine; Tavares, Josean Fechine; Silva, Marcelo Sobral; de Athayde-Filho, Petronio Filgueiras; Barbosa-Filho, Jose Maria
Bidens pilosa L. is a cosmopolitan annual herb, known for its traditional use in treating various diseases and thus much studied for the biological activity of its extracts, fractions and isolated compounds. Polyacetylenes and flavonoids, typical metabolite classes in the Bidens genus, predominate in the phytochemistry of B. pilosa. These classes of compounds have great taxonomic significance. In the Asteraceae family, the acetylene moiety is widely distributed in the Heliantheae tribe and some representatives, such as 1-phenylhepta-1,3,5-triyne, are noted for their biological activity and strong long-wave UV radiation absorbance. The flavonoids, specifically aurones and chalcones, have been reported as good sub-tribal level markers. Natural products from several other classes have also been isolated from different parts of B. pilosa. This review summarizes the available information on the 198 natural products isolated to date from B. pilosa.
Petronio Filgueiras de Athayde-Filho
Full Text Available Bidens pilosa L. is a cosmopolitan annual herb, known for its traditional use in treating various diseases and thus much studied for the biological activity of its extracts, fractions and isolated compounds. Polyacetylenes and flavonoids, typical metabolite classes in the Bidens genus, predominate in the phytochemistry of B. pilosa. These classes of compounds have great taxonomic significance. In the Asteraceae family, the acetylene moiety is widely distributed in the Heliantheae tribe and some representatives, such as 1-phenylhepta-1,3,5-triyne, are noted for their biological activity and strong long-wave UV radiation absorbance. The flavonoids, specifically aurones and chalcones, have been reported as good sub-tribal level markers. Natural products from several other classes have also been isolated from different parts of B. pilosa. This review summarizes the available information on the 198 natural products isolated to date from B. pilosa.
Shanmugapriya, Gnanasekaran; Das, Sudhanshu Sekhar; Veluthambi, Karuppannan
Mungbean yellow mosaic virus (MYMV) is a bipartite begomovirus that infects many pulse crops such as blackgram, mungbean, mothbean, Frenchbean, and soybean. We tested the efficacy of the transgenically expressed intron-spliced hairpin RNA gene of the transcriptional activator protein (hpTrAP) in reducing MYMV DNA accumulation. Tobacco plants transformed with the MYMV hpTrAP gene accumulated 21–22 nt siRNA. Leaf discs of the transgenic plants, agroinoculated with the partial dimers of MYMV, di...
Fiallo-Olivé, Elvira; Martínez-Zubiaur, Yamila; Moriones, Enrique; Navas-Castillo, Jesús
The complete genome sequence of two isolates of the bipartite begomovirus (genus Begomovirus, family Geminiviridae) Sida golden mosaic Florida virus (SiGMFV) is presented. We propose that both isolates, found infecting Malvastrum coromandelianum (family Malvaceae) in Cuba, belong to a new strain of SiGMFV. Phylogenetic analysis showed that SiGMFV DNA-A is located in a monophyletic cluster that includes begomoviruses infecting malvaceous weeds from the Caribbean.
Full Text Available Eukaryotic cells restrain the activity of foreign genetic elements, including viruses, through RNA silencing. Although viruses encode suppressors of silencing to support their propagation, viruses may also exploit silencing to regulate host gene expression or to control the level of their accumulation and thus to reduce damage to the host. RNA silencing in plants propagates from cell to cell and systemically via a sequence-specific signal. Since the signal spreads between cells through plasmodesmata like the viruses themselves, virus-encoded plasmodesmata-manipulating movement proteins (MP may have a central role in compatible virus:host interactions by suppressing or enhancing the spread of the signal. Here, we have addressed the propagation of GFP silencing in the presence and absence of MP and MP mutants. We show that the protein enhances the spread of silencing. Small RNA analysis indicates that MP does not enhance the silencing pathway but rather enhances the transport of the signal through plasmodesmata. The ability to enhance the spread of silencing is maintained by certain MP mutants that can move between cells but which have defects in subcellular localization and do not support the spread of viral RNA. Using MP expressing and non-expressing virus mutants with a disabled silencing suppressing function, we provide evidence indicating that viral MP contributes to anti-viral silencing during infection. Our results suggest a role of MP in controlling virus propagation in the infected host by supporting the spread of silencing signal. This activity of MP involves only a subset of its properties implicated in the spread of viral RNA.
de la Iglesia, Francisca; Martínez, Fernando; Hillung, Julia; Cuevas, José M; Gerrish, Philip J; Daròs, José-Antonio; Elena, Santiago F
A potential drawback of recent antiviral therapies based on the transgenic expression of artificial microRNAs is the ease with which viruses may generate escape mutations. Using a variation of the classic Luria-Delbrück fluctuation assay, we estimated that the spontaneous mutation rate in the artificial microRNA (amiR) target of a plant virus was ca. 6 × 10(-5) per replication event.
The 5′-terminal (RTn) and 3′-terminal (RTc) halves of the coat protein readthrough domain and the 19 ku cysteine-rich protein of Chinese wheat mosaic virus (CWMV) were amplified by RT-PCR, cloned and expressed in E. coli. Antisera and monoclonal antibodies against these proteins were prepared by immunising these purified proteins to mice. Detection of RTn, RTc and 19 ku proteins in CWMV infected wheat sap and leaf tissue indicated that the RTn and RTc proteins were distributed on the surface of virus particles whereas the 19 ku protein was in the cytoplasm of the infected wheat cells.
Decai Tuo; Wentao Shen; Pu Yan; Xiaoying Li; Peng Zhou
Papaya leaf distortion mosaic virus (PLDMV) is becoming a threat to papaya and transgenic papaya resistant to the related pathogen, papaya ringspot virus (PRSV). The generation of infectious viral clones is an essential step for reverse-genetics studies of viral gene function and cross-protection. In this study, a sequence- and ligation-independent cloning system, the In-Fusion® Cloning Kit (Clontech, Mountain View, CA, USA), was used to construct intron-less or intron-containing full-length ...
Maneechoat, Phoowanarth; Takeshita, Minoru; Uenoyama, Misa; Nakatsukasa, Maki; Kuroda, Atsuko; Furuya, Naruto; Tsuchiya, Kenichi
Host responses to infection by a mild strain of cucumber mosaic virus, termed CMV-m1, were re-examined in several plant species in comparison with those by a severe strain CMV-Y. Mild systemic symptoms were developed on the six plant species inoculated with CMV-m1. Virus titer in the Nicotiana benthamiana plants infected with CMV-m1 was significantly lower than those infected with CMV-Y, although infection by CMV-m1 interfered with further infection by CMV-Y in the plants. Subsequently, the attenuated virulence of CMV-m1 was analyzed by reassortment and recombination analyses between CMV-m1 and CMV-Y RNAs. The results suggested that the 2b protein of CMV-m1 (m1-2b) is involved in the formation of mild symptoms in N. benthamiana. Furthermore, site-directed mutagenesis demonstrated that Thr18 of m1-2b is responsible for formation of mild symptoms. Local RNA silencing suppressor activity of m1-2b was a little lower than that of severe strain CMV-Y. We discuss the relationship between attenuation of CMV-m1 and the features of m1-2b.
Seo, Jang-Kyun; Kwon, Sun-Jung; Chaturvedi, Sonali; Choi, Soon Ho; Rao, A L N
Satellite RNAs (satRNA) associated with Cucumber mosaic virus (CMV) have been shown to generate multimers during replication. We have discovered that multimers of a CMV satRNA generated in the absence of its helper virus (HV) are characterized by the addition of a hepta nucleotide motif (HNM) at the monomer junctions. Here, we evaluated the functional significance of HNM in HV-dependent replication by ectopically expressing wild type and mutant forms of satRNA multimers in planta either in (+) or (-)-strand polarity. Comparative replication profiles revealed that (-)-strand multimers with complementary HNM (cHNM) are the preferred initial templates for HV-dependent replication than (-)-strand monomers and multimers lacking the cHNM. Further mutational analyses of the HNM accentuate that preservation of the sequence and native length of HNM is obligatory for efficient replication of satRNA. A model implicating the significance of HNM in HV-dependent production of monomeric and multimeric forms of satRNA is presented.
Ntui, Valentine Otang; Kynet, Kong; Azadi, Pejman; Khan, Raham Sher; Chin, Dong Poh; Nakamura, Ikuo; Mii, Masahiro
Cucumber mosaic virus is an important plant pathogen with a broad host range encompassing many plant species. This study demonstrates the production of transgenic potato lines exhibiting complete resistance to cucumber mosaic virus strain O and Y by post transcriptional gene silencing. Two constructs were used, one, pEKH2IN2CMVai, contains inverted repeat of 1,138 bp fragment of a defective CMV replicase gene derived from RNA2 of cucumber mosaic virus strain O (CMV-O), while the other, TRV-based VIGS vector (pTRV2CMVai), contains the same fragment of the replicase gene, but without inverted repeat. These constructs were used to produce transgenic potato lines of cultivar 'Danshaku', a susceptible genotype to CMV. Transgenic lines derived from pEKH2IN2CMVai accumulated small interfering RNA (siRNA) before and after virus challenge, whereas those derived from pTRV2CMVai showed siRNA expression after virus challenge. When transgenic lines were challenged with CMV-O or CMV-Y, four lines exhibited complete (100%) resistance to both strains, whereas the other lines had high levels of resistance. Infectivity of CMV-O was lower than that of CMV-Y in the highly resistant plants. There were no significant differences with regard to resistance between plants derived from pEKH2IN2CMVai and those obtained from pTRV2CMVai. The presence of CMV-specific siRNA in the resistant phenotypes indicates that the resistance was acquired through RNA silencing.
BSMV infection caused a pronounced increase in the frequency of triploid and aneuploid seeds in eleven barley varieties, but with considerable variation in frequency among varieties. In some of the varieties triploids exceeded three per cent. In virus-free material a few triploids were found in m...
Pumpkin (Cucurbita moschata Duch.), Cucumber (Cucumis sativus Linn.) and Zucchini (Cucurbita pepo Linn.) are important crops in tropical and subtropical regions in the world, and they are popular vegetable crops in China. There are currently 59 viruses known infecting cucurbit plants which including...
Klarzynski, Olivier; Descamps, Valérie; Plesse, Bertrand; Yvin, Jean-Claude; Kloareg, Bernard; Fritig, Bernard
Sulfated fucans are common structural components of the cell walls of marine brown algae. Using a fucan-degrading hydrolase isolated from a marine bacterium, we prepared sulfated fucan oligosaccharides made of mono- and disulfated fucose units alternatively bound by alpha-1,4 and alpha-1,3 glycosidic linkages, respectively. Here, we report on the elicitor activity of such fucan oligosaccharide preparations in tobacco. In suspension cell cultures, oligofucans at the dose of 200 microg ml(-1) rapidly induced a marked alkalinization of the extracellular medium and the release of hydrogen peroxide. This was followed within a few hours by a strong stimulation of phenylalanine ammonia-lyase and lipoxygenase activities. Tobacco leaves treated with oligofucans locally accumulated salicylic acid (SA) and the phytoalexin scopoletin and expressed several pathogenesis-related (PR) proteins, but they displayed no symptoms of cell death. Fucan oligosaccharides also induced the systemic accumulation of SA and the acidic PR protein PR-1, two markers of systemic acquired resistance (SAR). Consistently, fucan oligosaccharides strongly stimulated both local and systemic resistance to tobacco mosaic virus (TMV). The use of transgenic plants unable to accumulate SA indicated that, as in the SAR primed by TMV, SA is required for the establishment of oligofucan-induced resistance.
Shivaprasad, P V; Thomas, M; Balamani, V; Biswas, D; Vanitharani, R; Karthikeyan, A S; Veluthambi, K
Mungbean yellow mosaic virus-Vigna (MYMV) sequences cloned as partial dimers within the T-DNA of a binary vector were deleted at a high frequency upon conjugal mobilization from Escherichia coli into Agrobacterium tumefaciens. This deletion involving the genome-length viral DNA did not occur when the binary plasmid was inside E. coli and when the binary plasmid was introduced into Agrobacterium by electroporation. Deletions occurred in both DNA A and DNA B partial dimers. A minimum of 500-nt continuity on either side of the nonanucleotide in the duplicated common region is required for deletion. A. tumefaciens cells in which deletion was complete, grew as larger colonies reflecting a growth advantage. The small, slow-growing colonies eventually lost the genome-length viral sequences after a few more cycles of growth. Partial dimers in binary plasmids pGA472 and pBin19 with RK2 replicon underwent deletion while those in pPZP with pVS1 replicon did not undergo deletion. Deletion was observed in A. tumefaciens strains C58, A136, A348 and A281 with C58 chromosome background, but not in Ach5 and T37. Interestingly, deletion did not occur in A. tumefaciens strain AGL1 with a recA mutation in C58 chromosome, implying a clear role for recombination in deletion. These observations suggest the choice of Agrobacterium strains and binary vectors for agroinoculation of geminiviruses.
Karthikeyan, A S; Vanitharani, R; Balaji, V; Anuradha, S; Thillaichidambaram, P; Shivaprasad, P V; Parameswari, C; Balamani, V; Saminathan, M; Veluthambi, K
One DNA A (KA30) and five different DNA B components (KA21, KA22, KA27, KA28 and KA34) of a geminivirus, Mungbean yellow mosaic virus-Vigna (MYMV-Vig) were cloned from a pooled sample of field-infected Vigna mungo plants from Vamban, South India. MYMV-Vig DNA A (KA30) and one of the DNA B components (KA27) exhibited 97% and 95% sequence identities, respectively, to those of MYMV reported from Thailand. However, the DNA B components KA21, KA22, KA28 and KA34 exhibited only 71 to 72% sequence identity to MYMV DNA B. Co-existence of multiple DNA B components in field-infected V. mungo was proved by Southern and PCR analyses. Each of the five DNA B components was infective together with the DNA A upon agroinoculation. Agroinoculation with mixed cultures of Agrobacterium with partial dimers of DNA A and all five DNA Bs proved that all five DNA B components can co-infect a single V. mungo plant.
Bink, Hugo H. J.; Schirawski, Jan; Haenni, Anne-Lise; Pleij, Cornelis W. A.
The RNA genome of turnip yellow mosaic virus (TYMV) consists of more than 6,000 nucleotides. During a study of the roles of the two hairpins located in its 90-nucleotide 5′ untranslated region, it was observed that stabilization of the 5′-proximal hairpin leads to a delay in the development of symptoms on plants. This delay in symptom development for both locally and systemically infected leaves was found to be dependent on a change in the free energy of the hairpin caused by introduced mutations. A protoplast transfection assay revealed that the accumulation of plus-strand full-length RNA and subgenomic RNA, as well as protein expression levels, was affected by hairpin stability. Stabilization of this hairpin inhibited translation. A model is proposed in which a destabilized 5′-proximal hairpin allows maximal translation of the viral proteins. It is suggested that this hairpin may exist in close proximity to the 5′ cap as long as its stability is low enough to enable translation. However, at an acidic pH, the hairpin structure becomes more stable and is functionally transformed into the initiation signal for viral packaging. Slightly acidic conditions can be found in chloroplasts, where TYMV assembly is driven by a low pH generated by active photosynthesis. PMID:12805444
Arif Muhammad Khan
Full Text Available The citrus mealybug, Planococcus citri, is an important plant pest with a very broad plant host range. P. citri is a phloem feeder and loss of plant vigor and stunting are characteristic symptoms induced on a range of host plants, but P. citri also reduces fruit quality and causes fruit drop leading to significant yield reductions. Better strategies for managing this pest are greatly needed. RNA interference (RNAi is an emerging tool for functional genomics studies and is being investigated as a practical tool for highly targeted insect control. Here we investigated whether RNAi effects can be induced in P. citri and whether candidate mRNAs could be identified as possible targets for RNAi-based P. citri control. RNAi effects were induced in P. citri, as demonstrated by specific target reductions of P. citri actin, chitin synthase 1 and V-ATPase mRNAs after injection of the corresponding specific double-stranded RNA inducers. We also used recombinant Tobacco mosaic virus (TMV to express these RNAi effectors in Nicotiana benthamiana plants. We found that P. citri showed lower fecundity and pronounced death of crawlers after feeding on recombinant TMV-infected plants. Taken together, our data show that actin, chitin synthase 1 and V-ATPase mRNAs are potential targets for RNAi against P. citri, and that recombinant TMV is an effective tool for evaluating candidate RNAi effectors in plants.
Zheng gui-jie; Yang Yong-qing; Ma Ying; Yang Xiao-feng; Chen Shan-yu; Ren Rui; Wang Da-gang; Yang Zhong-lu; ZhI hai-jian
Soybean mosaic virus (SMV) disease is one of the most destructive viral diseases in soybean (Glycine max (L.) Merr.). SMV strain SC3 is the major prevalent strain in huang-huai and Yangtze valleys, China. The soybean cultivar Qihuang 1 is of a rich resistance spectrum and has a wide range of application in breeding programs in China. In this study, F1, F2 and F2:3 from Qihuang 1×nannong 1138-2 were used to study inheritance and linkage mapping of the SC3 resistance gene in Qihuang 1. The secondary F2 population and near isogenic lines (nILs) derived from residual heterozygous lines (RhLs) of Qihuang 1×nannong 1138-2 were separatively used in the ifne mapping and candidate gene analysis of the resistance gene. Results indicated that a single dominant gene (designated RSC3Q) controls resistance, which was located on chromosome 13. Two genomic-simple sequence repeat (SSR) markers BARCSOYSSR_13_1114 and BARCSOYSSR_13_1136 were found lfanking the two sides of the RSC3Q. The interval between the two markers was 651 kb. Quantitative real-time PCR analysis of the candidate genes showed that ifve genes (Glyma13g25730, 25750, 25950, 25970 and 26000) were likely involved in soybean SMV resistance. These results would have utility in cloning of RSC3Q resistance candidate gene and marker-assisted selection (MaS) in resistance breeding to SMV.
TAO; Xiaorong; (陶小荣); ZHOU; Xueping; (周雪平); LI; Guixin; (李桂新); YU; Jialin; (于嘉林)
The hypersensitive response (HR) is one of the most important defense responses during the incompatible interaction between plant and pathogen. The viral determinant of HR on legumes induced by Cucumber mosaic virus (CMV) was studied, and our previous results showed that 243 nucleotides on 2a polymerase gene of CMV were involved in the induction of HR on legumes. With further analysis of the nucleotides and amino acids in this region, the amino acids Phe and Ala at positions 631 and 641 in the 2a polymerase of CMV-Fny, a legume local necrotic strain, were specifically exchanged to Tyr and Ser, respectively and simultaneously, in the 2a polymerase of CMV-P1, a legume systemic infecting strain, and three point mutants were constructed. The point mutant Fny-F/Y (Phe631 to Tyr) induced large necrotic lesions instead of pinpoint lesions, and the size of lesions could enlarge from initial sites. The point mutant Fny-A/S (Ala641 to Ser) induced similar symptoms as CMV-Fny. The double-point mutant Fny-FA/YS (Phe631 to Tyr and Ala641 to Ser) infected the legumes systemically without HR. These data indicate that the induction of HR on legumes is co-determined by two amino acids at positions 631 and 641 in CMV 2a polymerase.
Kato, Masahiko; Ishibashi, Kazuhiro; Kobayashi, Chihoko; Ishikawa, Masayuki; Katoh, Etsuko
Tm-1, the protein product of Tm-1, a semidominant resistance gene of tomato, inhibits tomato mosaic virus (ToMV) replication by binding to ToMV replication proteins. Previous studies suggested the importance of the Tm-1 N-terminal region for its inhibitory activity; however, it has not been determined if the N-terminal region is sufficient for inhibition. Furthermore, the three-dimensional structure of Tm-1 has not been determined. In this study, an N-terminal fragment of Tm-1 (residues 1-431) as a fusion protein containing an upstream maltose-binding protein was expressed in E. coli Rosetta (DE3) cells at 30°C and then purified. The solubility of the fusion protein was greater when the cells were cultured at 30°C than when cultured at lower or higher temperatures. The purified N-terminal Tm-1 fragment from which the maltose-binding protein tag had been removed has inhibitory activity against ToMV RNA replication.
Acosta-Ramírez, Elizabeth; Pérez-Flores, Rebeca; Majeau, Nathalie; Pastelin-Palacios, Rodolfo; Gil-Cruz, Cristina; Ramírez-Saldaña, Maricela; Manjarrez-Orduño, Nataly; Cervantes-Barragán, Luisa; Santos-Argumedo, Leopoldo; Flores-Romo, Leopoldo; Becker, Ingeborg; Isibasi, Armando; Leclerc, Denis; López-Macías, Constantino
Identifying the properties of a molecule involved in the efficient activation of the innate and adaptive immune responses that lead to long-lasting immunity is crucial for vaccine and adjuvant development. Here we show that the papaya mosaic virus (PapMV) is recognized by the immune system as a pathogen-associated molecular pattern (PAMP) and as an antigen in mice (Pamptigen). A single immunization of PapMV without added adjuvant efficiently induced both cellular and specific long-lasting antibody responses. PapMV also efficiently activated innate immune responses, as shown by the induction of lipid raft aggregation, secretion of pro-inflammatory cytokines, up-regulation of co-stimulatory molecules on dendritic cells and macrophages, and long-lasting adjuvant effects upon the specific antibody responses to model antigens. PapMV mixed with Salmonella enterica serovar Typhi (S. typhi) outer membrane protein C increased its protective capacity against challenge with S. typhi, revealing the intrinsic adjuvant properties of PapMV in the induction of immunity. Antigen-presenting cells loaded with PapMV efficiently induced antibody responses in vivo, which may link the innate and adaptive responses observed. PapMV recognition as a Pamptigen might be translated into long-lasting antibody responses and protection observed. These properties could be used in the development of new vaccine platforms.
Ni, Peng; Vaughan, Robert C; Tragesser, Brady; Hoover, Haley; Kao, C Cheng
Brome mosaic virus (BMV) packages its genomic and subgenomic RNAs into three separate viral particles. BMV purified from barley, wheat, and tobacco have distinct relative abundances of the encapsidated RNAs. We seek to identify the basis for the host-dependent differences in viral RNA encapsidation. Sequencing of the viral RNAs revealed recombination events in the 3' untranslated region of RNA1 of BMV purified from barley and wheat, but not from tobacco. However, the relative amounts of the BMV RNAs that accumulated in barley and wheat are similar and RNA accumulation is not sufficient to account for the difference in RNA encapsidation. Virions purified from barley and wheat were found to differ in their isoelectric points, resistance to proteolysis, and contacts between the capsid residues and the RNA. Mass spectrometric analyses revealed that virions from the three hosts had different post-translational modifications that should impact the physiochemical properties of the virions. Another major source of variation in RNA encapsidation was due to the purification of BMV particles to homogeneity. Highly enriched BMV present in lysates had a surprising range of sizes, buoyant densities, and distinct relative amounts of encapsidated RNAs. These results show that the encapsidated BMV RNAs reflect a combination of host effects on the physiochemical properties of the viral capsids and the enrichment of a subset of virions. The previously unexpected heterogeneity in BMV should influence the timing of the infection and also the host innate immune responses.
Kumar, Bhupender; Talukdar, Akshay; Verma, Khushbu; Bala, Indu; Harish, G D; Gowda, Sarmrat; Lal, S K; Sapra, R L; Singh, K P
Yellow Mosaic Virus (YMV) is a serious disease of soybean. Resistance to YMV was mapped in 180 soybean genotypes through association mapping approach using 121 simple sequence repeats (SSR) and four resistance gene analogue (RGA)-based markers. The association mapping population (AMP) (96 genotypes) and confirmation population (CP) (84 genotypes) was tested for resistance to YMV at hot-spot consecutively for 3 years (2007-2009). The genotypes exhibited significant variability for YMV resistance (P Molecular genotyping and population structure analysis with 'admixture' co-ancestry model detected seven optimal sub-populations in the AMP. Linkage disequilibrium (LD) between the markers extended up to 35 and 10 cM with r2 > 0.15, and >0.25, respectively. The 4 RGA-based markers showed no association with YMV resistance. Two SSR markers, Satt301 and GMHSP179 on chromosome 17 were found to be in significant LD with YMV resistance. Contingency Chi-square test confirmed the association (P < 0.01) and the utility of the markers was validated in the CP. It would pave the way for marker assisted selection for YMV resistance in soybean. This is the first report of its kind in soybean.
Khan, Arif Muhammad; Ashfaq, Muhammad; Kiss, Zsofia; Khan, Azhar Abbas; Mansoor, Shahid; Falk, Bryce W
The citrus mealybug, Planococcus citri, is an important plant pest with a very broad plant host range. P. citri is a phloem feeder and loss of plant vigor and stunting are characteristic symptoms induced on a range of host plants, but P. citri also reduces fruit quality and causes fruit drop leading to significant yield reductions. Better strategies for managing this pest are greatly needed. RNA interference (RNAi) is an emerging tool for functional genomics studies and is being investigated as a practical tool for highly targeted insect control. Here we investigated whether RNAi effects can be induced in P. citri and whether candidate mRNAs could be identified as possible targets for RNAi-based P. citri control. RNAi effects were induced in P. citri, as demonstrated by specific target reductions of P. citri actin, chitin synthase 1 and V-ATPase mRNAs after injection of the corresponding specific double-stranded RNA inducers. We also used recombinant Tobacco mosaic virus (TMV) to express these RNAi effectors in Nicotiana benthamiana plants. We found that P. citri showed lower fecundity and pronounced death of crawlers after feeding on recombinant TMV-infected plants. Taken together, our data show that actin, chitin synthase 1 and V-ATPase mRNAs are potential targets for RNAi against P. citri, and that recombinant TMV is an effective tool for evaluating candidate RNAi effectors in plants.
Huh, Sung Un; Choi, La Mee; Lee, Gil-Je; Kim, Young Jin; Paek, Kyung-Hee
WRKY transcription factors regulate biotic, abiotic, and developmental processes. In terms of plant defense, WRKY factors have important roles as positive and negative regulators via transcriptional regulation or protein-protein interaction. Here, we report the characterization of the gene encoding Capsicum annuum WRKY transcription factor d (CaWRKYd) isolated from microarray analysis in the Tobacco mosaic virus (TMV)-P(0)-inoculated hot pepper plants. CaWRKYd belongs to the WRKY IIa group, a very small clade in the WRKY subfamily, and WRKY IIa group has positive/negative regulatory roles in Arabidopsis and rice. CaWRKYd transcripts were induced by various plant defense-related hormone treatments and TMV-P(0) inoculation. Silencing of CaWRKYd affected TMV-P(0)-mediated hypersensitive response (HR) cell death and accumulation of TMV-P(0) coat protein in local and systemic leaves. Furthermore, expression of some pathogenesis-related (PR) genes and HR-related genes was reduced in the CaWRKYd-silenced plants compared with TRV2 vector control plants upon TMV-P(0) inoculation. CaWRKYd was confirmed to bind to the W-box. Thus CaWRKYd is a newly identified Capsicum annuum WRKY transcription factor that appears to be involved in TMV-P(0)-mediated HR cell death by regulating downstream gene expression.
孙浩华; 薛峰; 陈集双
大豆花叶病毒(Soybean mosaic virus,SMV)病是在世界范围内广泛分布并普遍发生的病毒病害之一,导致大豆严重减产和种质衰退.这引起了国内外许多学者的科研兴趣,研究内容涉及SMV株系划分与发生分布、传播流行方式、寄主上的症状和影响因素、基因组结构组成及各基凶功能、植物生理生化抗性、大豆SMV抗性遗传育种等各方面.本文综述了近年来国内外SMV的科研方向和进展,旨在为进一步的研究提供依据.
Jean Michel Hily
Full Text Available Identification of the determinants of pathogen reservoir potential is central to understand disease emergence. It has been proposed that host lifespan is one such determinant: short-lived hosts will invest less in costly defenses against pathogens, so that they will be more susceptible to infection, more competent as sources of infection and/or will sustain larger vector populations, thus being effective reservoirs for the infection of long-lived hosts. This hypothesis is sustained by analyses of different hosts of multihost pathogens, but not of different genotypes of the same host species. Here we examined this hypothesis by comparing two genotypes of the plant Arabidopsis thaliana that differ largely both in life-span and in tolerance to its natural pathogen Cucumber mosaic virus (CMV. Experiments with the aphid vector Myzus persicae showed that both genotypes were similarly competent as sources for virus transmission, but the short-lived genotype was more susceptible to infection and was able to sustain larger vector populations. To explore how differences in defense against CMV and its vector relate to reservoir potential, we developed a model that was run for a set of experimentally-determined parameters, and for a realistic range of host plant and vector population densities. Model simulations showed that the less efficient defenses of the short-lived genotype resulted in higher reservoir potential, which in heterogeneous host populations may be balanced by the longer infectious period of the long-lived genotype. This balance was modulated by the demography of both host and vector populations, and by the genetic composition of the host population. Thus, within-species genetic diversity for lifespan and defenses against pathogens will result in polymorphisms for pathogen reservoir potential, which will condition within-population infection dynamics. These results are relevant for a better understanding of host-pathogen co-evolution, and of
Ali, N; Heslop-Harrison, Js Pat; Ahmad, H; Graybosch, R A; Hein, G L; Schwarzacher, T
Pyramiding of alien-derived Wheat streak mosaic virus (WSMV) resistance and resistance enhancing genes in wheat is a cost-effective and environmentally safe strategy for disease control. PCR-based markers and cytogenetic analysis with genomic in situ hybridisation were applied to identify alien chromatin in four genetically diverse populations of wheat (Triticum aestivum) lines incorporating chromosome segments from Thinopyrum intermedium and Secale cereale (rye). Out of 20 experimental lines, 10 carried Th. intermedium chromatin as T4DL*4Ai#2S translocations, while, unexpectedly, 7 lines were positive for alien chromatin (Th. intermedium or rye) on chromosome 1B. The newly described rye 1RS chromatin, transmitted from early in the pedigree, was associated with enhanced WSMV resistance. Under field conditions, the 1RS chromatin alone showed some resistance, while together with the Th. intermedium 4Ai#2S offered superior resistance to that demonstrated by the known resistant cultivar Mace. Most alien wheat lines carry whole chromosome arms, and it is notable that these lines showed intra-arm recombination within the 1BS arm. The translocation breakpoints between 1BS and alien chromatin fell in three categories: (i) at or near to the centromere, (ii) intercalary between markers UL-Thin5 and Xgwm1130 and (iii) towards the telomere between Xgwm0911 and Xbarc194. Labelled genomic Th. intermedium DNA hybridised to the rye 1RS chromatin under high stringency conditions, indicating the presence of shared tandem repeats among the cereals. The novel small alien fragments may explain the difficulty in developing well-adapted lines carrying Wsm1 despite improved tolerance to the virus. The results will facilitate directed chromosome engineering producing agronomically desirable WSMV-resistant germplasm.
Potschka, M.; Kock, M.H.J.; Adams, M.L.; Schuster, T.M.
The kinetics of assembly and disassembly of tobacco mosaic virus coat protein (TMVP) following temperature jumps have been studied by small-angle X-ray scattering and turbidimetry. The structures of the principal aggregates of TMVP oligomers (A protein), intermediate size (helix I) and large size helical rods (helix II), have been characterized by their average radii of gyration of thickness, cross section, and shape obtained from the corresponding regimes of the small-angle scattering pattern. This structural information was obtained within seconds after the temperature-induced initiation of either polymerization or depolymerization and allowed the authors to detect transient intermediates. This methodology made it possible to observe and characterize the structure of a principal intermediate. Taken together with other kinetic information, these data suggest that polymerization of TMVP under virus self-assembly conditions may proceed via a single-layered helical nucleus that contains about 20 subunits. Previous studies have shown that overshoot polymerization of TMVP can occur and result in metastable long helical viruslike rods which subsequently depolymerize and then form short helical rods, depending on the conditions of the final equilibrium state. The longer rods (helix II) are overshoot polymers which form within seconds and contain 17 1/3 subunits per turn (helix IIB), in contrast to the subunit packing arrangement of 16 1/3 subunits per turn found in the shorter helical rods (helix IA). The latter packing arrangement is the one found in TMV. An overall polymerization scheme is proposed for the formation of these two helical forms of TMVP.
Betancourt, Mónica; Fraile, Aurora; García-Arenal, Fernando
Two groups of Cucumber mosaic virus (CMV) satellite RNAs (satRNAs), necrogenic and non-necrogenic, can be differentiated according to the symptoms they cause in tomato plants, a host in which they also differ in fitness. In most other CMV hosts these CMV-satRNA cause similar symptoms. Here, we analyse whether they differ in traits determining their relative fitness in melon plants, in which the two groups of CMV-satRNAs cause similar symptoms. For this, ten necrogenic and ten non-necrogenic field satRNA genotypes were assayed with Fny-CMV as a helper virus. Neither type of CMV-satRNA modified Fny-CMV symptoms, and both types increased Fny-CMV virulence similarly, as measured by decreases in plant biomass and lifespan. Necrogenic and non-necrogenic satRNAs differed in their ability to multiply in melon tissues; necrogenic satRNAs accumulated to higher levels both in single infection and in competition with non-necrogenic satRNAs. Indeed, multiplication of some non-necrogenic satRNAs was undetectable. Transmission between hosts by aphids was less efficient for necrogenic satRNAs as a consequence of a more severe reduction of CMV accumulation in leaves. The effect of CMV accumulation on aphid transmission was not compensated for by differences in satRNA encapsidation efficiency or transmissibility to CMV progeny. Thus, necrogenic and non-necrogenic satRNAs differ in their relative fitness in melon, and trade-offs are apparent between the within-host and between-host components of satRNA fitness. Hence, CMV-satRNAs could have different evolutionary dynamics in CMV host-plant species in which they do not differ in pathogenicity.
Zhu, Feng; Xi, De-Hui; Yuan, Shu; Xu, Fei; Zhang, Da-Wei; Lin, Hong-Hui
Systemic resistance is induced by pathogens and confers protection against a broad range of pathogens. Recent studies have indicated that salicylic acid (SA) derivative methyl salicylate (MeSA) serves as a long-distance phloem-mobile systemic resistance signal in tobacco, Arabidopsis, and potato. However, other experiments indicate that jasmonic acid (JA) is a critical mobile signal. Here, we present evidence suggesting both MeSA and methyl jasmonate (MeJA) are essential for systemic resistance against Tobacco mosaic virus (TMV), possibly acting as the initiating signals for systemic resistance. Foliar application of JA followed by SA triggered the strongest systemic resistance against TMV. Furthermore, we use a virus-induced gene-silencing-based genetics approach to investigate the function of JA and SA biosynthesis or signaling genes in systemic response against TMV infection. Silencing of SA or JA biosynthetic and signaling genes in Nicotiana benthamiana plants increased susceptibility to TMV. Genetic experiments also proved the irreplaceable roles of MeSA and MeJA in systemic resistance response. Systemic resistance was compromised when SA methyl transferase or JA carboxyl methyltransferase, which are required for MeSA and MeJA formation, respectively, were silenced. Moreover, high-performance liquid chromatography-mass spectrometry analysis indicated that JA and MeJA accumulated in phloem exudates of leaves at early stages and SA and MeSA accumulated at later stages, after TMV infection. Our data also indicated that JA and MeJA could regulate MeSA and SA production. Taken together, our results demonstrate that (Me)JA and (Me)SA are required for systemic resistance response against TMV.
Silva, Rodolpho G G; Vasconcelos, Ilka M; Martins, Thiago F; Varela, Anna L N; Souza, Pedro F N; Lobo, Ana K M; Silva, Fredy D A; Silveira, Joaquim A G; Oliveira, Jose T A
The physiological and biochemical responses of a drought tolerant, virus-susceptible cowpea genotype exposed to drought stress (D), infected by Cowpea severe mosaic virus (CPSMV) (V), and to these two combined stresses (DV), at 2 and 6 days post viral inoculation (DPI), were evaluated. Gas exchange parameters (net photosynthesis, transpiration rate, stomatal conductance, and internal CO2 partial pressure) were reduced in D and DV at 2 and 6 DPI compared to control plants (C). Photosynthesis was reduced by stomatal and biochemical limitations. Water use efficiency increased at 2 DPI in D, DV, and V, but at 6 DPI only in D and DV compared to C. Photochemical parameters (effective quantum efficiency of photosystem II and electron transport rate) decreased in D and DV compared to C, especially at 6 DPI. The potential quantum efficiency of photosystem II did not change, indicating reversible photoinhibition of photosystem II. In DV, catalase decreased at 2 and 6 DPI, ascorbate peroxidase increased at 2 DPI, but decreased at 6 DPI. Hydrogen peroxide increased at 2 and 6 DPI. Peroxidase increased at 6 DPI and chitinase at 2 and 6 DPI. β-1,3-glucanase decreased in DV at 6 DPI compared to V. Drought increased cowpea susceptibility to CPSMV at 2 DPI, as verified by RT-PCR. However, at 6 DPI, the cowpea plants overcome this effect. Likewise, CPSMV increased the negative effects of drought at 2 DPI, but not at 6 DPI. It was concluded that the responses to combined stresses are not additive and cannot be extrapolated from the study of individual stresses.
Musser, Richard O; Hum-Musser, Sue M; Gallucci, Matthew; DesRochers, Brittany; Brown, Judith K
Plants are routinely exposed to biotic and abiotic stresses to which they have evolved by synthesizing constitutive and induced defense compounds. Induced defense compounds are usually made, initially, at low levels; however, following further stimulation by specific kinds of biotic and abiotic stresses, they can be synthesized in relatively large amounts to abate the particular stress. cDNA microarray hybridization was used to identify an array of genes that were differentially expressed in tomato plants 15 d after they were exposed to feeding by nonviruliferous whiteflies or by viruliferous whiteflies carrying Pepper golden mosaic virus (PepGMV) (Begomovirus, Geminiviridae). Tomato plants inoculated by viruliferous whiteflies developed symptoms characteristic of PepGMV, whereas plants exposed to nonviruliferous whitefly feeding or nonwounded (negative) control plants exhibited no disease symptoms. The microarray analysis yielded over 290 spotted probes, with significantly altered expression of 161 putative annotated gene targets, and 129 spotted probes of unknown identities. The majority of the differentially regulated "known" genes were associated with the plants exposed to viruliferous compared with nonviruliferous whitefly feeding. Overall, significant differences in gene expression were represented by major physiological functions including defense-, pathogen-, photosynthesis-, and signaling-related responses and were similar to genes identified for other insect-plant systems. Viruliferous whitefly-stimulated gene expression was validated by real-time quantitative polymerase chain reaction of selected, representative candidate genes (messenger RNA): arginase, dehydrin, pathogenesis-related proteins 1 and -4, polyphenol oxidase, and several protease inhibitors. This is the first comparative profiling of the expression of tomato plants portraying different responses to biotic stress induced by viruliferous whitefly feeding (with resultant virus infection
In order to study the feasibility of Cucumber mosaic virus (CMV) as an expression vector, the full-length cDNA of RNA 3 from strain SD was cloned and the sequence around the start codon of the coat protein (CP) gene was modified to create an NsiⅠ site for insertion of foreign genes. The CP gene was replaced by the green fluorescent protein (GFP) gene. The cDNAs of Fny RNAs 1 and 2 and the chimeric SD RNA 3 were cloned between the modified 35S promoter and terminator. Tobacco protoplasts were transfected with a mixture of the viral cDNAs containing 35S promoter and terminator as a replacement vector and expressed GFP. A complementation system was established when the replacement vector was inoculated onto the transgenic tobacco plants ex-pressing SD-CMV CP. GFP was detected in the inoculated leaves in 5 of 18 tested plants and in the first upper systemic leaf of one of the 5 plants ten days after inoculation. However, no GFP could be detected in all the plants one month after inoculation. Recombination between the CMV vector and the CP transgene was proved by retro-transcriptional polymerase chain reaction (RT-PCR) and verified by DNA sequencing. Our results argue against the feasibility of the CMV-based replace-ment vector trans-complemented by the CP transgene, and at the same time, enlighten ways to improve the CMV-based expression vector and the biosafety of CMV CP-mediated virus resistant transgenic plants.
Kil Hyun Kim
Full Text Available Virus-induced gene silencing (VIGS is an effective tool for the study of soybean gene function. Successful VIGS depends on the interaction between virus spread and plant growth, which can be influenced by environmental conditions. Recently, we developed a new VIGS system derived from the Soybean yellow common mosaic virus (SYCMV. Here, we investigated several environmental and developmental factors to improve the efficiency of a SYCMV-based VIGS system to optimize the functional analysis of the soybean. Following SYCMV: Glycine max-phytoene desaturase (GmPDS infiltration, we investigated the effect of photoperiod, inoculation time, concentration of Agrobacterium inoculm, and growth temperature on VIGS efficiency. In addition, the relative expression of GmPDS between non-silenced and silenced plants was measured by qRT-PCR. We found that gene silencing efficiency was highest at a photoperiod of 16/8 h (light/dark at a growth temperature of approximately 27°C following syringe infiltration to unrolled unifoliolate leaves in cotyledon stage with a final SYCMV:GmPDS optimal density (OD₆₀₀ of 2.0. Using this optimized protocol, we achieved high efficiency of GmPDS-silencing in various soybean germplasms including cultivated and wild soybeans. We also confirmed that VIGS occurred in the entire plant, including the root, stem, leaves, and flowers, and could transmit GmPDS to other soybean germplasms via mechanical inoculation. This optimized protocol using a SYCMV-based VIGS system in the soybean should provide a fast and effective method to elucidate gene functions and for use in large-scale screening experiments.
Kim, Kil Hyun; Lim, Seungmo; Kang, Yang Jae; Yoon, Min Young; Nam, Moon; Jun, Tae Hwan; Seo, Min-Jung; Baek, Seong-Bum; Lee, Jeom-Ho; Moon, Jung-Kyung; Lee, Suk-Ha; Lee, Su-Heon; Lim, Hyoun-Sub; Moon, Jae Sun; Park, Chang-Hwan
Virus-induced gene silencing (VIGS) is an effective tool for the study of soybean gene function. Successful VIGS depends on the interaction between virus spread and plant growth, which can be influenced by environmental conditions. Recently, we developed a new VIGS system derived from the Soybean yellow common mosaic virus (SYCMV). Here, we investigated several environmental and developmental factors to improve the efficiency of a SYCMV-based VIGS system to optimize the functional analysis of the soybean. Following SYCMV: Glycine max-phytoene desaturase (GmPDS) infiltration, we investigated the effect of photoperiod, inoculation time, concentration of Agrobacterium inoculm, and growth temperature on VIGS efficiency. In addition, the relative expression of GmPDS between non-silenced and silenced plants was measured by qRT-PCR. We found that gene silencing efficiency was highest at a photoperiod of 16/8 h (light/dark) at a growth temperature of approximately 27°C following syringe infiltration to unrolled unifoliolate leaves in cotyledon stage with a final SYCMV:GmPDS optimal density (OD)600 of 2.0. Using this optimized protocol, we achieved high efficiency of GmPDS-silencing in various soybean germplasms including cultivated and wild soybeans. We also confirmed that VIGS occurred in the entire plant, including the root, stem, leaves, and flowers, and could transmit GmPDS to other soybean germplasms via mechanical inoculation. This optimized protocol using a SYCMV-based VIGS system in the soybean should provide a fast and effective method to elucidate gene functions and for use in large-scale screening experiments.
Krezhova, Dora; Hristova, Dimitrina; Iliev, Ilko; Yanev, Tony
Application of multispectral remote sensing techniques to plant condition monitoring has been adopted for various purposes. Remote sensing is a reliable tool for detecting signs of vege-tation stress and diseases. Spectral reflectance and chlorophyll fluorescence are functions of tissue optical properties and biological status of the plants, and illumination conditions. The mean reflectance spectrum depends on the relative composition of all the pigments in the leaf including chlorophylls, carotenoids etc. Chlorophyll fluorescence results from the primary re-actions of photosynthesis and during the last decade it finds widening application as a means for revelation of stress and diseases. The changes in chlorophyll function take place before the alteration in chlorophyll content to occur so that changes in the fluorescence signal arise before any visible signs are apparent. The aim of our investigations was to study the development and spreading out of a viral infection on the leaves of two cultivars tobacco plants (Nicotiana tabacum L.) infected with Tobacco mosaic virus (TMV). We applied two remote sensing tech-niques (spectral reflectance and chlorophyll fluorescence measurements) for evaluation of the changes in the optical properties of the plants in accordance to their physiological status. The serological analyses via the Double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) were made with appropriate kits (Leowe, Germany) for quantitative assessment of the concentration of viruses in the plants. The tobacco plants were grown in green house under controlled conditions. The first cultivar Nevrocop 1146 is known as resistive to the TMV, i.e. it shows hypersensitive response. The second cultivar named Krumovgrad is normally sen-sitive to the TMV. At growth stage 4-6 expanded leaf, up to one leaf from 20 plants for each cultivar were inoculated with TMV. The leaves opposite to the infected ones formed the group of control (untreated) leaves. The
Coat protein mutations in an attenuated Cucumber mosaic virus encoding mutant 2b protein that lacks RNA silencing suppressor activity induces chlorosis with photosynthesis gene repression and chloroplast abnormalities in infected tobacco plants.
Mochizuki, Tomofumi; Yamazaki, Ryota; Wada, Tomoya; Ohki, Satoshi T
In tobacco plants, the Cucumber mosaic virus (CMV) pepo strain induces mosaic symptoms, including pale green chlorosis and malformed tissues. Here, we characterized the involvement of 2b protein and coat protein (CP) in the development of mosaic symptoms. A 2b mutant (R46C) that lacks viral suppressor of RNA silencing (VSR) activity showed an asymptomatic phenotype with low levels of virus accumulation. Tomato spotted wilt virus NSs protein did not complement the virulence of the R46C, although it did restore high-level virus accumulation. However, R46C mutants expressing mutated CP in which the amino acid P129 was mutated to A, E, C, Q, or S induced chlorosis that was associated with reduced expression of chloroplast and photosynthesis related genes (CPRGs) and abnormal chloroplasts with fewer thylakoid membranes. These results suggest that the CP of the CMV pepo strain acquires virulence by amino acid mutations, which causes CPRG repression and chloroplast abnormalities.
Lin, Wenwu; Yan, Wenkai; Yang, Wenting; Yu, Chaowei; Chen, Huihuang; Zhang, Wen; Wu, Zujian; Yang, Liang; Xie, Lianhui
We characterised the virus-derived small interfering RNAs (vsiRNA) of bamboo mosaic virus (Ba-vsiRNAs) and its associated satellite RNA (satRNA)-derived siRNAs (satsiRNAs) in a bamboo plant (Dendrocalamus latiflorus) by deep sequencing. Ba-vsiRNAs and satsiRNAs of 21-22 nt in length, with both (+) and (-) polarity, predominated. The 5'-terminal base of Ba-vsiRNA was biased towards A, whereas a bias towards C/U was observed in sense satsiRNAs, and towards A in antisense satsiRNAs. A large set of bamboo genes were identified as potential targets of Ba-vsiRNAs and satsiRNAs, revealing RNA silencing-based virus-host interactions in plants. Moreover, we isolated and characterised new isolates of bamboo mosaic virus (BaMV; 6,350 nt) and BaMV-associated satRNA (satBaMV; 834 nt), designated BaMV-MAZSL1 and satBaMV-MAZSL1, respectively.
Full Text Available To investigate the plant genes affected by Bamboo mosaic virus (BaMV infection, we applied a cDNA-amplified fragment length polymorphism technique to screen genes with differential expression. A serine/threonine kinase-like (NbSTKL gene of Nicotiana benthamiana is upregulated after BaMV infection. NbSTKL contains the homologous domain of Ser/Thr kinase. Knocking down the expression of NbSTKL by virus-induced gene silencing reduced the accumulation of BaMV in the inoculated leaves but not in the protoplasts. The spread of GFP-expressing BaMV in the inoculated leaves is also impeded by a reduced expression of NbSTKL. These data imply that NbSTKL facilitates the cell-to-cell movement of BaMV. The subcellular localization of NbSTKL is mainly on the cell membrane, which has been confirmed by mutagenesis and fractionation experiments. Combined with the results showing that active site mutation of NbSTKL does not change its subcellular localization but significantly affects BaMV accumulation, we conclude that NbSTKL may regulate BaMV movement on the cell membrane by its kinase-like activity. Moreover, the transient expression of NbSTKL does not significantly affect the accumulation of Cucumber mosaic virus (CMV and Potato virus X (PVX; thus, NbSTKL might be a specific protein facilitating BaMV movement.
Schoelz, James E; Angel, Carlos A; Nelson, Richard S; Leisner, Scott M
The genomes of many plant viruses have a coding capacity limited to inclusion body protein' (IB) present in infected plants. P6 is now referred to in most articles as the transactivator (TAV)/viroplasmin protein, because the first viral function to be characterized for the Caulimovirus P6 protein beyond its role as an inclusion body protein (the viroplasmin) was its role in translational transactivation (the TAV function). This review will discuss the currently accepted functions for P6 and then present the evidence for an entirely new function for P6 in intracellular movement.
Full Text Available The Nicotiana tabacum genome contains four Dicer-like proteins (DCLs and six RNA-dependent RNA polymerase (RDR homologues involved in the RNA silencing mechanism employed against viral infection. DCL1 synthesizes 18-21 nt-long microRNA, whereas DCL2, DCL3 and DCL4 produce 22 nt, 24 nt and 21 nt-long siRNA, respectively, in the RNA silencing process. This study aimed to clarify which components among these are involved in changes in the amount of virus and the development of symptoms in Cucumber mosaic virus (CMV-infected tobacco. Infected transgenic tobacco lines with a single down-regulation of DCL2, DCL4, RDR1 or a double down-regulation of both DCL2 and 4 were analyzed. The amounts of viral RNA in young developing leaves in transgenic tobacco lines were examined by Northern blot analysis. Most transgenic plants inoculated with CMV Pepo, a virulent strain, exhibited cyclic mosaic symptoms. The amount of viral RNA in single down-regulated lines varied based on leaf position in a similar manner to that noted in non-transgenic tobacco, while that of the double down-regulated line did not. Furthermore, the expression of RNA-silencing-related genes during high and low CMV infection did not differ among the transgenic plants. These results suggested that (i changes in the amounts of the virus in the developing leaves of all the single down-regulated lines were associated with cyclic symptom expression in fully expanded leaves, and (ii the lower expression of DCL2, DCL4 and RDR1 may be sufficient to establish cyclic symptom development.
T Jacob; T Jebasingh; M N Venugopal; R Usha
A survey was conducted to study the biological and genetic diversity of Cardamom mosaic virus (CdMV) that causes the most widespread disease in the cardamom growing area in the Western Ghats of south India. Six distinct subgroups were derived based on their symptomatology and host range from the sixty isolates collected. The serological variability between the virus isolates was analysed by ELISA and Western blotting. The 3′ terminal region consisting of the coat protein (CP) coding sequence and 3′ untranslated region (3′UTR) was cloned and sequenced from seven isolates. Sequence comparisons revealed considerable genetic diversity among the isolates in their CP and 3′UTR, making CdMV one of the highly variable members of Potyviridae. The possible occurrence of recombination between the isolates and the movement of the virus in the cardamom tract of south India are discussed.
Robson Marcelo Di Piero
Full Text Available The objective of the present study was to evaluate the protection of passion fruit plants against CABMV by using preparations from Agaricus brasiliensis and Lentinula edodes mushrooms. In experiments carried out in the greenhouse, the fruiting body extracts from some of the isolates of both mushrooms significantly reduced CABMV incidence in passion fruit plants. This protective effect occurred when the plant leaves, pre-treated with extracts, were later inoculated mechanically with the virus. However, the extracts did not protect the plants in experiments involving CABMV transmission by aphid vectors. An inhibitory effect of mushroom extracts on the virus particles was also demonstrated on Chenopodium quinoa, a CABMV local lesion host, by inoculating the plants with a mixture of extracts and virus suspension. Still in C. quinoa, the mushroom extracts from some isolates induced systemic resistance against the virus. These results showed that aqueous extracts from A. brasiliensis and L. edodes fruiting bodies had CABMV infectivity inhibitors, but that was not enough to control the viral disease on passion fruit plants at all, considering they were infected through a vector.O endurecimento dos frutos do maracujazeiro, causado pelo Cowpea aphid-borne mosaic virus (CABMV, é um dos problemas mais sérios que atingem a cultura. Tentativas de se obter plantas resistentes ao vírus ou estirpes fracas premunizantes não apresentaram sucesso até o momento. O objetivo do presente estudo foi o de avaliar a proteção das plantas de maracujá contra o CABMV, utilizando preparações dos cogumelos Lentinula edodes e Agaricus blazei, através da indução de resistência. Em experimentos conduzidos no interior de casa de vegetação, os extratos de basidiocarpos de ambos os cogumelos reduziram significativamente a incidência da virose em plantas de maracujá que tiveram as folhas pré-tratadas com esses extratos e que foram posteriormente inoculadas
This article describes a lesson inspired by Sicilian mosaics. The author first presented a PowerPoint presentation of mosaics from the Villa Romana del Casale and reviewed complementary and analogous colors. Students then created mosaics using a variety of art materials. They presented their work to their peers and discussed the thought and…
玉米矮花叶病(maize dwarf mosaic virus,MDMv)是世界上玉米产区普遍发生的病毒病害之一.自20世纪90年代以来,我国玉米矮花叶病发生严重,山西、甘肃、山东、河北以及北京等省市先后大流行,造成了巨大的农业经济损失.在我国玉米产区造成危害的主要是该病毒的B株系,主要借蚜虫传播和种子传播;在玉米矮花叶病的防治中,种植抗病品种,并辅以合理的栽培管理,可有效防止MDMV.本文主要综述玉米矮花叶病病毒的理化特性、玉米矮花叶病的发生危害、病原及其传播方式、发病条件、流行与防治、品种(自交系)抗性、抗性鉴定、抗性遗传及其抗病基因工程研究等方面的研究进展,以期为以后玉米矮花叶病的有效防治提供一定的参考.
Ishibashi, Kazuhiro; Kezuka, Yuichiro; Kobayashi, Chihoko; Kato, Masahiko; Inoue, Tsuyoshi; Nonaka, Takamasa; Ishikawa, Masayuki; Matsumura, Hiroyoshi; Katoh, Etsuko
The tomato mosaic virus (ToMV) resistance gene Tm-1 encodes a protein that shows no sequence homology to functionally characterized proteins. Tm-1 binds ToMV replication proteins and thereby inhibits replication complex formation. ToMV mutants that overcome this resistance have amino acid substitutions in the helicase domain of the replication proteins (ToMV-Hel). A small region of Tm-1 in the genome of the wild tomato Solanum habrochaites has been under positive selection during its antagonistic coevolution with ToMV. Here we report crystal structures for the N-terminal inhibitory domains of Tm-1 and a natural Tm-1 variant with an I91-to-T substitution that has a greater ability to inhibit ToMV RNA replication and their complexes with ToMV-Hel. Each complex contains a Tm-1 dimer and two ToMV-Hel monomers with the interfaces between Tm-1 and ToMV-Hel bridged by ATP. Residues in ToMV-Hel and Tm-1 involved in antagonistic coevolution are found at the interface. The structural differences between ToMV-Hel in its free form and in complex with Tm-1 suggest that Tm-1 affects nucleoside triphosphatase activity of ToMV-Hel, and this effect was confirmed experimentally. Molecular dynamics simulations of complexes formed by Tm-1 with ToMV-Hel variants showed how the amino acid changes in ToMV-Hel impair the interaction with Tm-1 to overcome the resistance. With these findings, together with the biochemical properties of the interactions between ToMV-Hel and Tm-1 variants and effects of the mutations in the polymorphic residues of Tm-1, an atomic view of a step-by-step coevolutionary arms race between a plant resistance protein and a viral protein emerges.
Yield of varieties of Cucurbita pepo preimmunized with mild strains of Papaya ringspot virus - type W and Zucchini yellow mosaic virus Produção de variedades de Cucurbita pepo premunizadas com estirpes fracas do Papaya ringspot virus - type W e do Zucchini yellow mosaic virus
Full Text Available Papaya ringspot virus - type W (PRSV-W and Zucchini yellow mosaic virus (ZYMV are the most prevalent viruses in cucurbit crops in Brazil and responsible for frequent yield losses. Diseases caused by these viruses are difficult to control. The objective of this work was to evaluate the effects of mild strains PRSV-W-1 and ZYMV-M on the yield of Cucurbita pepo L. cvs. Samira, Novita Plus, AF 2847, and Yasmin, under plastic greenhouse and field conditions. Plants infected with ZYMV-M and grown in a plastic greenhouse did not exhibit typical leaf symptoms or significant alterations in quantitative and qualitative fruit yield. However, when infected with PRSV-W-1, or PRSV-W-1 + ZYMV-M, the plants exhibited severe leaf mosaic symptoms and reduced fruit quality, although there were no changes in the number and mean fruit weight harvested from these plants. When these plants were infected with PRSV-W-1 and studied simultaneously in the field and plastic greenhouse, intensification of symptoms in the fruits and leaves was more pronounced under the greenhouse conditions. Quantitative yield did not change. Environmental factors seem to influence symptoms induced by PRSV-W-1.O Papaya ringspot virus - type W (PRSV-W e o Zucchini yellow mosaic virus (ZYMV são os vírus predominantes em culturas de cucurbitáceas no Brasil, onde geralmente causam danos significativos na produção. As doenças causadas por ambos os vírus são de difícil controle. O objetivo desse trabalho foi avaliar o efeito das estirpes fracas PRSV-W-1 e ZYMV-M na produção de abobrinha de moita (Cucurbita pepo L. cvs. Samira, Novita Plus, AF 2847, and Yasmin em condições de estufa plástica e de campo. Plantas infectadas com a estirpe ZYMV-M sob condições de estufa plástica não exibiram sintomas foliares típicos da doença e alterações na quantidade e qualidade dos frutos produzidos. No entanto, quando infectadas com a estirpe PRSV-W-1, ou PRSV-W-1 + ZYMV-M, as plantas
Full Text Available Tm-2² is a coiled coil-nucleotide binding-leucine rich repeat resistance protein that confers durable extreme resistance against Tomato mosaic virus (ToMV and Tobacco mosaic virus (TMV by recognizing the viral movement protein (MP. Here we report that the Nicotiana benthamiana J-domain MIP1 proteins (NbMIP1s associate with tobamovirus MP, Tm-2² and SGT1. Silencing of NbMIP1s reduced TMV movement and compromised Tm-2²-mediated resistance against TMV and ToMV. Furthermore, silencing of NbMIP1s reduced the steady-state protein levels of ToMV MP and Tm-2². Moreover, NbMIP1s are required for plant resistance induced by other R genes and the nonhost pathogen Pseudomonas syringae pv. tomato (Pst DC3000. In addition, we found that SGT1 associates with Tm-2² and is required for Tm-2²-mediated resistance against TMV. These results suggest that NbMIP1s function as co-chaperones during virus infection and plant immunity.
Maheshwari, Richa; Panigrahi, Gatikrushna; Angappan, K
The present study was carried out to find out the variations present in different isolates of yellow mosaic virus (YMV) causing yellow mosaic disease of pulses in southern parts of India. The coat protein gene of YMV was amplified using gene specific and deng universal primers with DNA isolated from YMV infected samples. Further, cloning and DNA sequencing of CP gene was carried out. CP gene decrypt sequences revealed that YMV infected samples of Black gram, Cowpea and Green gram were similar to the MYMV-Tamil Nadu isolates. Whereas the YMV infected sample of Horse gram was found to be similar with HYMV. Hence, in the present study, two distinct YMV infecting pulses in Tamil Nadu (MYMV and HYMV species) were identified and it was observed that there exists considerable genetic variation among these species. In addition, Cowpea crop which was earlier supposed not to be susceptible for YMV infection also showed the presence of this virus similar to the MYMV. Overall, the findings of the present study indicate that the CP region is efficient enough to provide a simple, rapid, and reliable method for early detection of YMV infections in pulses, which would help to develop proper management strategies to control these viruses.
Bricault, Christine A. [Department of Plant Pathology and Plant-Microbe Biology, 334 Plant Science Building, Cornell University, Ithaca, NY 14850 (United States); Perry, Keith L., E-mail: KLP3@cornell.edu [Department of Plant Pathology and Plant-Microbe Biology, 334 Plant Science Building, Cornell University, Ithaca, NY 14850 (United States)
In the atomic model of Cucumber mosaic virus (CMV), six amino acid residues form stabilizing salt bridges between subunits of the asymmetric unit at the quasi-threefold axis of symmetry. To evaluate the effects of these positions on virion stability and aphid vector transmissibility, six charged amino acid residues were individually mutated to alanine. All of the six engineered viruses were viable and exhibited near wild type levels of virion stability in the presence of urea. Aphid vector transmissibility was nearly or completely eliminated in the case of four of the mutants; two mutants demonstrated intermediate aphid transmissibility. For the majority of the engineered mutants, second-site mutations were observed following aphid transmission and/or mechanical passaging, and one restored transmission rates to that of the wild type. CMV capsids tolerate disruption of acid–base pairing interactions at the quasi-threefold axis of symmetry, but these interactions are essential for maintaining aphid vector transmissibility. - Highlights: ► Amino acids between structural subunits of Cucumber mosaic virus affect vector transmission. ► Mutant structural stability was retained, while aphid vector transmissibility was disrupted. ► Spontaneous, second-site mutations restored aphid vector transmissibility.
André Luiz Lourenção
Full Text Available Há cerca de dez anos vem sendo observada a presença de Sceloenopla bidens (F., 1792 (Coleoptera: Chrysomelidae: Hispinae em folhas de Philodendron spp., em Campinas e outras localidades paulistas. O adulto permanece na face inferior das folhas, onde se alimenta, causando injúrias características. As larvas criam-se nas folhas, minando-as e comprometendo o aspecto ornamental da planta. Em condições de laboratório, o desenvolvimento do inseto desde ovo até emergência do adulto durou aproximadamente 48 dias. Em Campinas, efetuaram-se observações de seus danos na Floricultura Campineira, em cinco espécies de filodendros presentes - P. melinoni, P. bipinnatifidum, P. erubescens, P. selhom e P. wilsoni e no parque do Instituto Agronômico (IAC, onde a maioria das espécies não se encontra identificada. Em ambos os locais, verificou-se comportamento diferenciado de algumas espécies de filodendros em relação a S. bidens. Não se observou sua alimentação ou presença em outras epífitas dessa família (Araceae, situadas próximo a filodendros infestados, sugerindo possível especificidade da espécie com o gênero Philodendron.The occurrence of Sceloenopla bidens (F., 1792 (Coleoptera: Chrysomelidae: Hispinae on Philodendron spp. has been observed in Campinas and other cities of the State of São Paulo, Brazil. The adults stay in the lower surface of the leaves, where they feed, causing typical leaf injury. The larvae are leaf miners and, therefore, impair the ornamental effect of the plant. Under laboratory conditions, the insect development from egg to adult emergence lasted 48 days. In the philodendron germplasm of the Instituto Agronômico, a variation of performance among the species in relation to injury caused by this insect was verified. Two species showed leaves without damage, adults or larvae while others, as P. renauxii, exhibited highly damaged leaves and many adults. The presence or feeding of S. bidens in other epiphytes
Full Text Available Abstract Background Gene silencing vectors based on Barley stripe mosaic virus (BSMV are used extensively in cereals to study gene function, but nearly all studies have been limited to genes expressed in leaves of barley and wheat. However since many important aspects of plant biology are based on root-expressed genes we wanted to explore the potential of BSMV for silencing genes in root tissues. Furthermore, the newly completed genome sequence of the emerging cereal model species Brachypodium distachyon as well as the increasing amount of EST sequence information available for oat (Avena species have created a need for tools to study gene function in these species. Results Here we demonstrate the successful BSMV-mediated virus induced gene silencing (VIGS of three different genes in barley roots, i.e. the barley homologues of the IPS1, PHR1, and PHO2 genes known to participate in Pi uptake and reallocation in Arabidopsis. Attempts to silence two other genes, the Pi transporter gene HvPht1;1 and the endo-β-1,4-glucanase gene HvCel1, in barley roots were unsuccessful, probably due to instability of the plant gene inserts in the viral vector. In B. distachyon leaves, significant silencing of the PHYTOENE DESATURASE (BdPDS gene was obtained as shown by photobleaching as well as quantitative RT-PCR analysis. On the other hand, only very limited silencing of the oat AsPDS gene was observed in both hexaploid (A. sativa and diploid (A. strigosa oat. Finally, two modifications of the BSMV vector are presented, allowing ligation-free cloning of DNA fragments into the BSMV-γ component. Conclusions Our results show that BSMV can be used as a vector for gene silencing in barley roots and in B. distachyon leaves and possibly roots, opening up possibilities for using VIGS to study cereal root biology and to exploit the wealth of genome information in the new cereal model plant B. distachyon. On the other hand, the silencing induced by BSMV in oat seemed too
Reis, Rita Alexandra Feliciano dos
Dissertação de mestrado, Biologia Molecular e Microbiana, Faculdade de Ciências e Tecnologia, Universidade do Algarve, 2015 Grapevine fanleaf virus, vírus do urticado ou nó-curto da videira e Arabis mosaic virus são dois Nepovirus, da família Secoviridae e ambos possuem um genoma bipartido de cadeia simples e sentido positivo. Além destes, foram detetados em alguns isolados de ArMV e GFLV, RNAs satélite de grande tamanho, satRNA do tipo B. Estes vírus encontram-se entre os principais agent...
Shanmugapriya, Gnanasekaran; Das, Sudhanshu Sekhar; Veluthambi, Karuppannan
Mungbean yellow mosaic virus (MYMV) is a bipartite begomovirus that infects many pulse crops such as blackgram, mungbean, mothbean, Frenchbean, and soybean. We tested the efficacy of the transgenically expressed intron-spliced hairpin RNA gene of the transcriptional activator protein (hpTrAP) in reducing MYMV DNA accumulation. Tobacco plants transformed with the MYMV hpTrAP gene accumulated 21-22 nt siRNA. Leaf discs of the transgenic plants, agroinoculated with the partial dimers of MYMV, displayed pronounced reduction in MYMV DNA accumulation. Thus, silencing of the TrAP gene, a suppressor of gene silencing, emerged as an effective strategy to control MYMV.
Zheng, Hongying; Xiao, Caili; Han, Kelei; Peng, Jiejun; Lin, Lin; Lu, Yuwen; Xie, Li; Wu, Xiaohua; Xu, Pei; Li, Guojing; Chen, Jianping; Yan, Fei
The complete 6243-nucleotide sequence of a cucumber green mottle mosaic virus (CGMMV) isolate from bottle gourd in Zhejiang province, China, was determined. A full-length cDNA clone of this isolate was constructed by inserting the cDNA between the 35S promoter and the ribozyme in the binary plasmid pCB301-CH. A suspension of an Agrobacterium tumefaciens EHA105 clone carrying this construct was highly infectious in Nicotiana benthamiana and bottle gourd. Another infectious clone containing the green fluorescence protein (GFP) reporter gene was also successfully constructed. This study is the first report of the efficient use of agroinoculation for generating CGMMV infections.
Pappachan, Anju; Subashchandrabose, Chinnathambi; Satheshkumar, P S; Savithri, H S; Murthy, M R N
A unique feature of several T=3 icosahedral viruses is the presence of a structure called the beta-annulus formed by extensive hydrogen bonding between protein subunits related by icosahedral three-fold axis of symmetry. This unique structure has been suggested as a molecular switch that determines the T=3 capsid assembly. In order to examine the importance of the beta-annulus, a deletion mutant of Sesbania mosaic virus coat protein in which residues 48-59 involved in the formation of the beta-annulus were deleted retaining the rest of the residues in the amino terminal segment (rCP (Delta48-59)) was constructed. When expressed in Escherichia coli, the mutant protein assembled into virus like particles of sizes close to that of the wild type virus particles. The purified capsids were crystallized and their three dimensional structure was determined at 3.6 A resolution by X-ray crystallography. The mutant capsid structure closely resembled that of the native virus particles. However, surprisingly, the structure revealed that the assembly of the particles has proceeded without the formation of the beta-annulus. Therefore, the beta-annulus is not essential for T=3 capsid assembly as speculated earlier and may be formed as a consequence of the particle assembly. This is the first structural demonstration that the virus particle morphology with and without the beta-annulus could be closely similar.
Zhao, Lei; Hao, Xingan; Wu, Yunfeng
Polysaccharides are essential macromolecules present in all living organisms, and have many kinds of biological activities, such as anti-oxidation, hypoglycemic, enhancing immunity, anti-aging, anti-rheumatism, anti-cancer and so on. In this study, the antiviral activity of polysaccharide peptide (PSP) was tested, compared with Ningnanmycin, and firstly found it has a stronger antiviral activity by using half-leaf method and leaf disk method. Subsequently, the mechanism of antiviral activity of PSP was preliminarily studied. As a result, its antiviral effect was better than the commercial agent Ningnanmycin, despite of protective effect, curative effect and inactivation effect. On the other hand, PSP as a commercial anti-cancer drug could easily and rapidly get in large quantities by liquid fermentation, which makes the industrialized production feasible. Also PSP is less toxic, easily biodegradable and ecofriendly. All the results are suggesting that PSP has potential as a pesticide to be used for the control of plant virus in the future.
周雪平; 薛朝阳; 陈青; 戚益军; 李德葆
Based on reported TMV-U1 sequence, primers were designed and fragments covering the entire genome of TMV broad bean strain (TMV-B) were obtained with RT-PCR. These fragments were cloned and sequenced and the 5’ and 3’ end sequences of genome were confirmed with RACE. The complete sequence of TMV-B comprises 6 395 nucleotides (nt) and four open reading frames, which correspond to 126 ku (1 116 amino acids), 183 ku (1 616 amino acids), 30 ku (268 amino acids) and 17.5 ku proteins (159 amino acids). The complete nucleotide sequence of TMV-B is 99.4% identical to that of TMV-U1. The two virus isolates share the same sequence of 5’, 3’ non-coding region and 17.5 K ORF, and 6, 1 and 3 amino acid changes are found in 126 K protein, 54 K protein and 30 K protein, respectively. The possible mechanism on the infection of TMV-B in Vicia faba is discussed.
Complete nucleotide sequences and construction of full-length infectious cDNA clones of cucumber green mottle mosaic virus (CGMMV) in a versatile newly developed binary vector including both 35S and T7 promoters.
Park, Chan-Hwan; Ju, Hye-Kyoung; Han, Jae-Yeong; Park, Jong-Seo; Kim, Ik-Hyun; Seo, Eun-Young; Kim, Jung-Kyu; Hammond, John; Lim, Hyoun-Sub
Seed-transmitted viruses have caused significant damage to watermelon crops in Korea in recent years, with cucumber green mottle mosaic virus (CGMMV) infection widespread as a result of infected seed lots. To determine the likely origin of CGMMV infection, we collected CGMMV isolates from watermelon and melon fields and generated full-length infectious cDNA clones. The full-length cDNAs were cloned into newly constructed binary vector pJY, which includes both the 35S and T7 promoters for versatile usage (agroinfiltration and in vitro RNA transcription) and a modified hepatitis delta virus ribozyme sequence to precisely cleave RNA transcripts at the 3' end of the tobamovirus genome. Three CGMMV isolates (OMpj, Wpj, and Mpj) were separately evaluated for infectivity in Nicotiana benthamiana, demonstrated by either Agroinfiltration or inoculation with in vitro RNA transcripts. CGMMV nucleotide identities to other tobamoviruses were calculated from pairwise alignments using DNAMAN. CGMMV identities were 49.89% to tobacco mosaic virus; 49.85% to pepper mild mottle virus; 50.47% to tomato mosaic virus; 60.9% to zucchini green mottle mosaic virus; and 60.96% to kyuri green mottle mosaic virus, confirming that CGMMV is a distinct species most similar to other cucurbit-infecting tobamoviruses. We further performed phylogenetic analysis to determine relationships of our new Korean CGMMV isolates to previously characterized isolates from Canada, China, India, Israel, Japan, Korea, Russia, Spain, and Taiwan available from NCBI. Analysis of CGMMV amino acid sequences showed three major clades, broadly typified as 'Russian,' 'Israeli,' and 'Asian' groups. All of our new Korean isolates fell within the 'Asian' clade. Neither the 128 nor 186 kDa RdRps of the three new isolates showed any detectable gene silencing suppressor function.
Duarte Keila M.R.
Full Text Available Monoclonal antibodies were obtained against Tomato mosaic tobamovirus (ToMV isolated in Brazil. One antibody (8G7G2 isotyped as IgG2b (kappa light chain showed strong specificity and very low cross reaction with the Tobacco mosaic virus (TMV. It can be used in identification of tomato mosaic virus (ToMV.
Balaji, V; Vanitharani, R; Karthikeyan, A S; Anbalagan, S; Veluthambi, K
Mungbean yellow mosaic virus-Vigna (MYMV-Vig), a Begomovirus that causes yellow mosaic disease, was cloned from field-infected blackgram (Vigna mungo). One DNA A clone (KA30) and five different DNA B clones (KA21, KA22, KA27, KA28 and KA34) were obtained. The sequence identity in the 150-nt common region (CR) between DNA A and DNA B was highest (95%) for KA22 DNA B and lowest (85.6%) for KA27 DNA B. The Rep-binding domain had three complete 11-nt (5'-TGTATCGGTGT-3') iterons in KA22 DNA B (and KA21, KA28 and KA34), while the first iteron in KA27 DNA B (5'-ATCGGTGT-3') had a 3-nt deletion. KA27 DNA B, which exhibited 93.9% CR sequence identity to the mungbean-infecting MYMV, also shared the 3-nt deletion in the first iteron besides having an 18-nt insertion between the third iteron and the conserved nonanucleotide. MYMV was found to be closely related to KA27 DNA B in amino acid sequence identity of BV1 (94.1%) and BC1 (97.6%) proteins and in the organization of nuclear localization signal (NLS), nuclear export signal (NES) and phosphorylation sites. Agroinoculation of blackgram (V. mungo) and mungbean (V. radiata) with partial dimers of KA27 and KA22 DNA Bs along with DNA A caused distinctly different symptoms. KA22 DNA B caused more intense yellow mosaic symptoms with high viral DNA titre in blackgram. In contrast, KA27 DNA B caused more intense yellow mosaic symptoms with high viral DNA titre in mungbean. Thus, DNA B of MYMVVig is an important determinant of host-range between V. mungo and V. radiata.
V Balaji; R Vanitharani; A S Karthikeyan; S Anbalagan; K Veluthambi
Mungbean yellow mosaic virus-Vigna (MYMV-Vig), a Begomovirus that causes yellow mosaic disease, was cloned from field-infected blackgram (Vigna mungo). One DNA A clone (KA30) and five different DNA B clones (KA21, KA22, KA27, KA28 and KA34) were obtained. The sequence identity in the 150-nt common region (CR) between DNA A and DNA B was highest (95%) for KA22 DNA B and lowest (85.6%) for KA27 DNA B. The Rep-binding domain had three complete 11-nt (5′-TGTATCGGTGT-3′) iterons in KA22 DNA B (and KA21, KA28 and KA34), while the first iteron in KA27 DNA B (5′-ATCGGTGT-3′) had a 3-nt deletion. KA27 DNA B, which exhibited 93.9% CR sequence identity to the mungbean-infecting MYMV, also shared the 3-nt deletion in the first iteron besides having an 18-nt insertion between the third iteron and the conserved nonanucleotide. MYMV was found to be closely related to KA27 DNA B in amino acid sequence identity of BV1 (94.1%) and BC1 (97.6%) proteins and in the organization of nuclear localization signal (NLS), nuclear export signal (NES) and phosphorylation sites. Agroinoculation of blackgram (V. mungo) and mungbean (V. radiata) with partial dimers of KA27 and KA22 DNA Bs along with DNA A caused distinctly different symptoms. KA22 DNA B caused more intense yellow mosaic symptoms with high viral DNA titre in blackgram. In contrast, KA27 DNA B caused more intense yellow mosaic symptoms with high viral DNA titre in mungbean. Thus, DNA B of MYMV-Vig is an important determinant of host-range between V. mungo and V. radiata.