IDENTIFICATION, CHARACTERIZATION AND TAXONOMY OF PLANT VIRUSES

IDENTIFICATION, CHARACTERIZATION AND TAXONOMY OF PLANT VIRUSES

Sponsoring Institution

National Institute of Food and Agriculture

Project Status

COMPLETE

Funding Source

HATCH

Reporting Frequency

Annual

Accession No.

0204480

Grant No.

(N/A)

Cumulative Award Amt.

(N/A)

Proposal No.

(N/A)

Multistate No.

(N/A)

Project Start Date

Jul 1, 2005

Project End Date

Jun 30, 2011

Grant Year

(N/A)

Program Code

[(N/A)]- (N/A)

Recipient Organization
MISSISSIPPI STATE UNIV
(N/A)
MISSISSIPPI STATE,MS 39762

Performing Department
Entomology & Plant Pathology

Non Technical Summary
Virus infections reduce the quality and quantity of agricultural or horticultural crops. The extent to which yield is reduced will depend in any particular year and locality on many factors including: (i) virus species or strain; (ii) host species/variety; (iii) incidence and activity of any vector; (iv) time of infection; (iv) environmental conditions, etc. Up-to-date knowledge on biological, epidemiological and molecular traits of virus species/isolates and availability of reliable, quick and cheap detection methods are the basis for establishment of proper control methods in order to limit their spread. Aim of this project is to identify major viruses infecting crops in Mississippi; to develop reliable and quick methods for virus diagnosis; to generate the very first data on viruses infecting ferns and muscadine grapes; to carry out taxonomic study on some grapevine viruses. The project objectives will be pursued mainly (but not only) by molecular approaches.

Animal Health Component

(N/A)

Research Effort Categories

Basic

(N/A)

Applied

(N/A)

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
212	4030	1040	50%
212	4030	1101	40%
212	4030	1170	10%

Knowledge Area
212 - Pathogens and Nematodes Affecting Plants;

Subject Of Investigation
4030 - Viruses;

Field Of Science
1040 - Molecular biology; 1101 - Virology; 1170 - Epidemiology;

Goals / Objectives
The project intends: (i) to identify major viruses infecting sweet potato, soybean, vegetables and ornamentals in Mississippi; (ii) to develop sensitive and quick detection methods for routine virus diagnosis; (iii) to generate first data on viruses infecting ferns and muscadine grapes; (iv) to characterize taxonomically some grapevine viruses.

Project Methods
1. Identification of viruses in Mississippi: Identification of viruses will include both field surveying and laboratory analysis. PCR will be the preferred method for detection. Considering that the number of virusesis extensive, the work in first phase will focus on the limited number of the most important viruses (3-5/plant species) reported to be present in the USA. Primers will be designed on conserved regions of published sequences available in the DataBank. Amplified products will be selected and cloned in a plasmid vector and sequenced. Sequence data will be analyzed and compared with reference isolates/strains available in DataBank. Both, variability of viruses/variants from different locations as well as variability of the virus within single host plant will be evaluated. In case of symptomatologies not associated with known viruses, the research approach will be as for the objective 3. In case of suspectred cases, investigation will also include 'virus-like agents' (i.e. viroids, phytoplasmas, etc). 2. Development and/or improvement of diagnostic methods: The following approaches will be applied: (i) Design and test degenerate primers for broad-range detection of related viruses (e.g. begomoviruses, tobamoviruses, tospoviruses, tymoviruses, etc); (ii) Develop multiplex RT-PCR methods in order to identify multiple virus infections in a single test; (iii) Develop one-step RT-PCR methods to simplify detection procedure; (iv) Design host-specific primers (for soybean and sweet potato) to be used as internal control in RT-PCR assays as already applied for other crops (Nassuth et al., 2000; Menzel et al., 2002; Abou Ghanem-Sabanadzovic et al., 2003); (v) produce non-radioactive riboprobes containing cRNA sequences of various viruses for large-scale assays by molecular hybridization tests. 3. Investigation on fern and muscadine viruses: Due to the complete lack of background information, the study will rely on virus-aspecific tests (general tests) such as (i) dsRNA analysis; (ii) sap transmission onto a range of artificial host plants and (iii) electron microscopy. Extracted dsRNAs will be used for random-primed cloning and sequencing. Obtained sequence data will be processed and analyzed by specific computer programs. In case of positive results in mechanical transmission trials, isolated virus(es) will be purified and completely characterized from biological, morphological, physico-chemical and molecular point of view. 4. Study on viruses of fam. Tymoviridae: This part of the project will focus on (i) taxonomic identification of a new virus of the fam. Tymoviridae reported from grapevine through genome cloning and sequencing as well as cytopathological and serological characterization; (ii) characterization of two 3'end co-terminal ORFs with unknown function of Grapevine fleck virus through their expression in vitro, production of specific antisera, immunoelectron microscopy study on their expression in vivo, sub-cellular localization and possible role(s).

Progress 07/01/05 to 06/30/11

Outputs
OUTPUTS: All originally planned objectives of this project have been fulfilled: we conducted a survey on viruses of soybeans, corn and sweet potatoes and identified main viruses infecting these crops in Mississippi. We have also characterized several new viruses infecting ornamental plants (i.e petunia, Japanese holly fern, calibrachoa, etc), grapevines, muscadines and blackberries. For each of these novel viruses we have developed molecular tools for their reliable and sensitive detection. Finally, we have characterized several members of the family Tymoviridae from muscadines/blackberries and contributed to better overall knowledge of this taxon of plant viruses.Permutation of RdRp motifs, discovered in one of marafiviruses characterized during this project, represents an undescribed feature for plant viruses. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: This project targeted scientists (plant virologists) involved in research on plant viruses as well as diagnosticians and extension staff. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Results obtained during this project have contributed to the overall knowledge on plant viruses present in Mississippi. Several viruses identified in this project are associated with economically important diseases in their respective hosts (i.e grapevines, blackberry, etc). The discovery of two similar viruses in tomato (Southern tomato virus) and rhododendron (Rhododendron virus A) added to the overall knowledge about evolutionary relationships between partitiviruses and totiviruses. Diagnostic tools developed during this project allow detection and identification of several new viruses. We have also developed "universal", taxon-specific, primers for detection of viruses belonging to the family Tymoviridae.

Publications

Valverde, R. A., and S. Sabanadzovic, 2011. A strain of Japanese holly fern mottle virus infecting leatherleaf fern in the United States. Plant Health Progress (doi:10.1094/PHP-2011-0128-01-RS) Published January 28, 2011.
Valverde R.A.. S. Sabanadzovic and M.C. Rush. 2011. Identification of Oryza sativa endornavirus in rice genotypes from breeding programs in the United States. Plant Breeding 130: 271-274.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic and I.E. Tzanetakis, 2011. Blackberry virus E: an unusual flexivirus. Archives of Virology 156: 1665-1669.
Sabanadzovic S. and R.A. Valverde, 2011. Properties and detection of two cryptic viruses from pepper (Capsicum annuum). Virus Genes 43: 307-312.
Tzanetakis I.E., S. Sabanadzovic, D.T. Johnson, H.J. Burrack, R.R. Martin, 2011. The emerging problem of blackberry yellow vein disease. Book of Abstracts X International Rubus and Ribes Symposium, Zlatibor, SERBIA, 65.
Poudel B., W.M. Wintermantel, S. Sabanadzovic, I.E. Tzanetakis, 2011 Epidemiological studies on Blackberry yellow vein associated virus and Blackberry chlorotic ringspot virus. Book of Abstracts X International Rubus and Ribes Symposium, Zlatibor, SERBIA: p75.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, I.E. Tzanetakis, 2011. Two new filamentous viruses of blackberries. Book of Abstracts X International Rubus and Ribes Symposium, Zlatibor, SERBIA: p76.
Villanueva F., S. Sabanadzovic, R. A. Valverde, J. Navas-Castillo, 2011. Evidencia de infeccion por virus con genoma de dsRNA pertenecientes a varios generos en aguacate. XI Congreso Nacional de Virologia, Sociedad Espanola de Virologia, May 29-June 1, 2011, Granada, SPAIN.
Valverde R.A., S. Sabanadzovic, R. Okada, J. Navas-Castillo, T. Fukuhara, 2011. Book of Abstracts 4th Conference of the International Working Group on Legume and Vegetable Viruses, Antequera-Malaga, SPAIN: p 40.
Okada R., E. Kiyota, S. Sabanadzovic, R.A. Valverde, M.J. Roossinck, T. Fukuhara and H. Moriyama, 2011. Structural conservation of the double-stranded RNA genome between plant endornaviruses. Book of Abstracts The Second International Mycovirus Symposium, Otaru, Hokkaido, JAPAN.
Okada R., E. Kiyota, S. Sabanadzovic, R.A. Valverde, M.J. Roossinck, T. Fukuhara and H. Moriyama, 2011. Identification of conserved sequences, motifs and structures between plant endornaviruses. Book of Abstracts XV International Congress of Virology - International Union of Microbiological Societies 2011 Congress, Sapporo, JAPAN.

Progress 01/01/10 to 12/31/10

Outputs
OUTPUTS: Objective 1: Survey activities in 2010 identified the presence of Tobacco ring spot virus in Joe-pye weed. This weed is common in Mississippi and represents a reservoir for this virus. The genome of Pokeweed mosaic virus, one of the most widespread viruses in the Eastern United States was completed in 2010. Additionally, we identified novel viruses in kudzu (no viruses were reported from this host in the USA) and ligustrum, currently in advanced stages of characterization. Survey on blackberry viruses in the state revealed the widespread presence of viruses, in both wild and cultivated species. The most widespread viruses appear to be Tobacco ring spot virus (TRSV), Blackberry yellow vein associated virus (BYVaV), Blackberry virus Y (BVY) and Blackberry virus E (BVE). Study on dsRNA viruses in different pepper (Capsicum spp) species has been initiated while characterizations of Blackberry virus E, Grapevine leafroll associated Carnelian virus - GLRaCV and Rhododendron virus A have been concluded in 2010. Objective 2: Specific reverse transcription polymerase chain reaction tests was developed for several recently characterized (i.e. Rhododendron virus A, Grapevine leafroll-associated Carnelian virus, BVE) or viruses as yet in characterization (i.e. viruses identified in Kudzu, ligustrum, blackberries, etc) and used to gain knowledge on their incidence/importance in the region. A multiplex RT-PCR detection method was developed for detection and discrimination between 2 cryptic viruses of pepper. Additionally a non-radioactive DIG-labeled probe has been synthesized for the detection of Rhododendron virus A. Objective 3: The cloning of reverse-transcribed dsRNA patterns isolated from several wild grapevine specimens revealed the presence of viruses distinct from those described in cultivated species (Vitis vinifera). Characterization of oryza-like and enamo-like viruses, as well as multiple cryptic viruses, isolated from wild grapevines/muscadines is currently on-going. Regarding ferns, a dsRNA pattern reminiscent of Japanese holly fern mottle virus (JHFMoV) has been found in symptomatic specimens of leatherleaf fern (Rumohra adiantiformis L) indicating that this (or a very similar) virus may infect this economically important ornamental plant species. Objective 4: An undescribed marafi-like virus was identified from Ranunculus sp collected in the Great Smoky Mountains National Park. Molecular and cytopathological data indicate that this virus is likely a new species in the genus Marafivirus (fam Tymoviridae). PARTICIPANTS: Training: R.C. Stephenson - Graduate student; R. Strangman - Undergraduate student. Scientific collaborations: Dr. N. Abou Ghanem-Sabanadzovic; Drs. T. Allen, W. Moore, D. Ingram & A. Lawrence - Mississippi State University, MS; Dr. I. E. Tzanetakis - University of Arkansas, AR; Dr. R.A. Valverde - Louisiana State University, LA; Drs. A. Rowhani & D.A. Golino - University of California Davis, CA; Drs. J.K. Uyemoto & W.M. Wintermantel - USDA/ARS; Dr. J. Navas-Castillo - IHSM-UMA-CSIC, Spain; Dr. M.J. Roossinck - The Noble Foundation. OK; Drs R. Okada, E. Kyota, C.K. Young, H. Moriyama, T. Fukuhara - Tokyo University of Agriculture and Technology, Japan. TARGET AUDIENCES: This project targets farmers, diagnosticians/extension experts and scientists involved in basic and applied research of plant viruses. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Results obtained in the framework of this project contribute to overall knowledge of the presence and importance of plant viruses in the region. Some viruses identified during 2010 could have significant economic importance in respective crop/ornamental host species (i.e. blackberry viruses, GLRaCV, fern virus). Diagnostic methods, developed in 2010, allow reliable detection of several novel viruses in a single or multiple RT-PCR reactions.

Publications

Garlapati R.B., D.C. Cross, M.A. Caprio, S. Sabanadzovic and J. Chen, 2010. Phenotypic and genotypic characters associated with polygyne hybrid imported fire ant. Midsouth Entomologist 3: 26-33.
Sabanadzovic S., D.M. Ingram and A. Lawrence, 2010. First report on Tobacco ringspot virus in Joe-pye weed (Eupatorium purpureum L.) in Mississippi. Plant Disease 94: 126.
Abou Ghanem-Sabanadzovic N., S. Sabanadzovic, J.K. Uyemoto, D.A. Golino and A.K. Rowhani, 2010. A putative new Ampelovirus associated with grapevine leafroll disease. Archives of Virology 155: 1871-1876.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic and R.A. Valverde, 2010. Novel monopartite dsRNA virus from rhododendron. Archives of Virology 155: 1859-1863.
Okada R., E. Kiyota, C-K Yong, H. Moriyama, T. Fukuhara, S. Sabanadzovic and R.A. Valverde, 2010. Common molecular properties among plant endornaviruses. 29th Annual Meeting of the American Society of Virology: p266.
Valverde R.A., M.R. Roossinck, S. Sabanadzovic and J.Navas-Castillo, 2010. Biological and molecular properties of an endornavirus infecting bell pepper (Capsicum annuum). Proceedings 2nd Meeting of the Spanish Network of Plant Virology, Puerto di Santa Maria, Cadiz, Spain: p70.
Sabanadzovic S. and R. Valverde, 2010. Genomic sequences and simultaneous detection of two cryptic viruses from pepper. Phytopathology 100: S112.
Sabanadzovic S. and I.E. Tzanetakis, 2010. A new ilarvirus from subgroup 1 infects ligustrum. Phytopathology 100: S112.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2010. Viruses of plants in the Great Smoky Mountains National Park. Phytopathology 100: S112.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2010. Another marafivirus infecting blackberries. Phytopathology 100: S112.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, W.F. Moore, T.W. Allen, R.C. Stephenson and A. Lawrence, 2010. First virus infecting Kudzu in USA. Phytopathology 100: S112.
Stephenson R.C. and S. Sabanadzovic, 2010. Investigations on population and hosts of Bean pod mottle virus in Mississippi. Phytopathology 100: S123.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2010. Two undescribed viral species isolated from native grapevines. Phytopathology 100: S2.
Bindu P., W.M. Wintermantel, S. Sabanadzovic and I.E. Tzanetakis, 2010. Epidemiological studies on Blackberry yellow vein-associated virus. Phytopathology 100: S103

Progress 01/01/09 to 12/31/09

Outputs
OUTPUTS: Objective 1. The survey on viruses infecting soybeans was completed in 2009. Bean pod mottle virus (BPMV) is the prevalent virus and was present in 65% of tested samples. Tobacco ring spot virus was present in ca 3.5% specimens. Peanut mottle virus was found in several fields and was associated with severe symptomatology. The virus was isolated on herbaceous hosts and partially sequenced. Soybean mosaic virus, Bean yellow mosaic virus and Southern bean mosaic virus were detected in a few geographically distinct locations. The study on the BPMV population in MS has been initiated and is on-going. Several new viruses (i.e. a marafivirus from giant ragweed) or new hosts for known viruses (i.e. calibrachoa for TVCV) have also been studied. Multi-lab collaborative research on characterization of a new virus from tomato (Southern tomato virus - STV) has been completed. A study on several new viruses of blackberries, as well as on viruses in non-agronomic ecosystems has been initiated. Objective 2. Research on development of general tools for endornavirus detection has been initiated in collaboration with LSU with very encouraging preliminary results. Additionally, a set of new primers for "universal" tymovirid detection has been published. Objective 3. Characterization of new viruses from fern (Japanese holly fern mottle virus, JHFMoV) and native grapevines (Grapevine virus Q = Grapevine Syrah virus 1, GVQ = GSyV-1) has been completed. Both viruses possess some unique features and appear to be new viral species. The virus from fern may be a prototype of a new taxon of plant viruses. Preliminary results on viral community in native grapevine germplasm show the presence of a number of as yet undescribed species. Objective 4. Two new species in the family Tymoviridae were characterized in 2009. One of them, Blackberry virus S (BlVs) has a genome organization typical for marafiviruses and represents the first member of this taxon described from small fruits. The second virus (GVQ = GSyV-1) is characterized by a unique (permuted) arrangement of motifs of the viral RdRp unreported in plant viruses and/or in viruses belonging to Alphavirus-like Supergroup. Furthermore, in the framework of international collaboration, a virus from Cypriot grapevine was identified as an isolate of a newly described virus GLRaV-Pr. PARTICIPANTS: Training: R.C. Stephenson - Graduate student. Scientific collaborations: Dr. N. Abou Ghanem-Sabanadzovic, Dr. A. Henn - (MSU), Dr. T. Allen - (MSU), Dr. D. Ingram - (MSU), A.M. Lawrence - (MSU), Dr. I. E. Tzanetakis - Universiy of Arkansas, Dr. R.A. Valverde - Louisiana State University, Dr. H.R. Pappu - Washington State University, Dr. R. Martin - USDA/ARS - Corvallis OR, Dr. A.E. Gorbalenya - Leiden University - The Netherlands, Dr. G.P. Martelli - University of Bari - Italy, Dr. M. Digiaro - Medit Agronomic Institute of Bari - Italy, Dr. T. Elbeaino - Medit Agronomic Institute of Bari - Italy, Dr. A. Rowhani - University of California Davis TARGET AUDIENCES: Our research targets farmers, diagnosticians/extension experts and scientists involved in basic research/taxonomy of plant viruses. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Our study in 2009 has contributed to a better understanding of problems caused by viruses in soybean fields in MS. BPMV is the most prevalent virus and should be the primary target of control efforts. Diagnostics developed for endornavirus and tymo/marafivirus detection facilitate their detection and can be transferred/applied in any plant virology lab. From a purely scientific viewpoint, our discoveries of unusual features in the JHFMoV and GVQ/GSyV-1 genomes represent significant contributions towards general knowledge on virus origin, evolution and taxonomy.

Publications

Sabanadzovic S., R.A. Valverde, J.K. Brown, R.R. Martin and I.E. Tzanetakis, 2009. Southern tomato virus: The link between the families Totiviridae and Partitiviridae. Virus Research 140: 130-137.
El Beaino T., F. Numic, M. Digiaro, S. Sabanadzovic and G.P. Martelli, 2009. Partial characterization of a grapevine leafroll-associated virus isolate from Cyprus. Journal of Plant Pathology 91: 479-484.
Valverde R.A. and S. Sabanadzovic, 2009. A new plant virus with unique properties infecting Japanese holly fern. Journal of General Virology 90, 2542-2549.
Sabanadzovic S, N. Abou Ghanem-Sabanadzovic and A.E. Gorbalenya, 2009. Permutation of the active site of putative RNA-dependent RNA polymerase in a newly identified species of plant Alpha-like virus. Virology 394, 1-7.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2009. Identification and molecular characterization of a marafivirus in Rubus spp. Archives of Virology 154, 1729-1735.
Sabanadzovic S., A. Henn, N. Abou Ghanem-Sabanadzovic and A. Lawrence, 2009. First report on Tobacco mild green mottle virus in calibrachoa plants (Calibrachoa x hybrida) in Mississippi. Plant Disease 93:1354
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, A. Henn, T.W. Allen, D.M. Ingram and A.M. Lawrence, 2009. Sequencing the Pokeweed mosaic virus genome, the final act of a century long characterization. Phytopathology 99(6): S112.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2009. Unraveling the phytovirus world of the Great Smoky Mountains National Park. Phytopathology 99(6): S112.
Stephenson R.C. and S. Sabanadzovic, 2009. Population of Bean pod mottle virus in Mississippi. Phytopathology 99(6): S124.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2009. Grapevine virus Q: The first phytovirus with inverted RdRp motifs. Phytopathology 99(6): S112.
Sabanadzovic S. and N. Abou Ghanem-Sabanadzovic, 2009. Association of a new marafivirus with ring spot symptoms in giant ragweed (Ambrosia trifida L.) in Mississippi. Phytopathology 99(6): S112.
Sabanadzovic S. and R.A. Valverde, 2009. A detection method for endornaviruses from various plant species. Phytopathology 99(6): S113.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic and H.R. Pappu, 2009. Identification and molecular characterization of a new member of the genus Petuvirus (family Caulimoviridae) from rhododendron. Phytopathology 99(6): S112.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, D.M. Ingram and A.M. Lawrence, 2009. Hypervariability of badnavirus-like sequences in Canna indica L. Phytopathology 99(6): S113.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic and I.E. Tzanetakis, 2009. Nucleotide sequences and detection of a new flexivirus from blackberry. Phytopathology 99(6): S112.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic and A.E. Gorbalenya, 2009. Grapevine virus Q: the first plant virus with permuted active site of RNA-dependent RNA polymerase. Extended Abstracts 16th Meeting of ICVG, Dijon, France, 31 Aug-4 Sept 2009: 42-43.
Sabanadzovic S., 2009. Viruses of native Vitis germplasm in the Southeastern United States. Extended Abstracts 16th Meeting of ICVG, Dijon, France, 31 Aug-4 Sept 2009: 32-35.

Progress 01/01/08 to 12/31/08

Outputs
OUTPUTS: Objective 1. Regular scouting on viruses in soybean fields has been continued also in 2008. As in previous years, Bean pod mottle virus was predominating one and was found in ca 55% of tested samples. The incidence of this virus in surveyed fields varied from 10-15% up to 100%. Contrary to 2007, when no Soybean mosaic virus-infected samples were found, this year we detected this virus in a commercial field in Noxubee Co and in Washington Co. Incidence in the tested samples was ca 15%. Majority of SMV-positive samples displayed severe distortion and dwarfing and were found co-infected by BPMV. Another aphid-transmitted virus, Bean yellow mosaic virus (gen. Potyvirus), was found in 12 tested samples collected from 2 different plots. It was also detected in 2007, indicating that it is probably established in our area and may be a serious problem in the future. Few additional viruses were found at much lower frequency in tested fields. A new virus on tomato (Southern tomato virus = Tomato yellow stunt virus) has been completely characterized in a collaborative work with several scientific institutions nation-wide. The study on pararetrovirus-like sequences in canna plants has been initiated and preliminary data indicate a variety of possibly new viruses present in these plants. Objective 2. In 2008, the routine RT-PCR detection method has been devised for a newly described virus of Japanese holly fern. Additionally, another project aimed at development of quick and reliable methods for identification of endornaviruses in plants has been initiated. Objective 3. Novel plant viruses have been further characterized from Japanese holly fern (Japanese holly fern mottle virus - JHFMoV) and muscadine (Grapevine virus Q - GVQ). Japanese holly fern virus is likely a prototype of a novel genus of plant viruses yet-to-be established. GVQ was found in several samples of native grapevine germplasm and in a wild blackberry. Objective 4. Two new marafiviruses have been detected in muscadine and other native grapes and blackberry (GVQ) and in wild and cultivated blackberries (Blackberry virus S- BlVS). While BlVS has a canonical type of organization of RdRp motifs, this enzyme in GVQ is organized in a unique manner, yet unreported for plant viruses. PARTICIPANTS: Training/professional development: Mr. Ronald Christian Stephenson, PhD student. Collaborators: Dr. Nina Abou Ghanem-Sabanadzovic, Dr. Alan Henn, A. Lawrence (MSU), Dr. Rodrigo Valverde (Louisiana State University), Dr. Ioannis E. Tzanetakis (University of Arkansas), Dr. Robert R. Martin (USDA-ARS, Corvallis, OR), Dr. Judith K. Brown (The University of Arizona), Dr. R. Gergerich (University of Arkansas - retired) TARGET AUDIENCES: Our research is aimed to produce results useful to farmers/producers, diagnosticians and scientists involved in study/taxonomy of plant viruses. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Results of our activities in 2008 have both, basic scientific and practical impacts: discovery of peculiar RdRp organization in GVQ will lead in deeper study of the overall biological implications of this phenomenon in terms of replication efficiency/pathogenicity, etc. Practical impact is reflected in development of detection methodologies that may be used for study of particular virus/group of viruses, especially for early screening and other tactics, part of the efficient management strategies.

Publications

SABANADZOVIC S., N. ABOU GHANEM-SABANADZOVIC, 2008. Molecular characterization and detection of a tripartite cryptic virus from rose. Journal of Plant Pathology 90(2), 285-291.
SABANADZOVIC AND N. ABOU GHANEM-SABANADZOVIC, 2008. First Report of Black raspberry necrosis virus in Rubus canadensis in Tennessee. Plant Disease 92(8), 1254.
SABANADZOVIC S., N. ABOU GHANEM-SABANADZOVIC, A. HENN, A. LAWRENCE, 2008. Characterization of a petunia strain of Turnip vein-clearing virus. Journal of Plant Pathology 90(3), 489-493.
SABANADZOVIC S., N. ABOU GHANEM-SABANADZOVIC, 2008. A novel Marafivirus from Rubus spp.Phytopathology 98: S137
SABANADZOVIC S., N. ABOU GHANEM-SABANADZOVIC, 2008. An undescribed dsRNA virus from Rhododendron. Phytopathology 98:S138
SABANADZOVIC S., N. ABOU GHANEM-SABANADZOVIC, 2008. Detection and identification of an umbravirus from Ageratina altissima. Phytopathology 98: S138
VALVERDE R.A., S. SABANADZOVIC, 2008. A new virus species causing a disease of Japanese holly fern (Cyrtomium falcatum). Phytopathology 98: S162
SABANADZOVIC S., R.A. VALVERDE, J.K. BROWN, R.R. MARTIN, I.E. TZANETAKIS, 2008. Southern tomato virus: Plant Pathogen or Just a Scientific Curiosity 23rd Annual Tomato Disease Workshop, October 15-16, 2008, Raymond (MS)
VALVERDE R.A., S. SABANADZOVIC, 2008. An emerging viral disease of Japanese holly fern and leather leaf fern in the United States caused by a new plant virus. Proceedings Annual Conference Costa Rica, May 2008.
MARTIN R.R., J. SUSAIMUTHU, S. SABANADZOVIC, R.C. GERGERICH, I.E. TZANETAKIS, 2008. Blackberry yellow vein disease is caused by multiple virus complexes. Proceedings 8th International Congress of Plant Pathology, Turin (IT), August 24-29, 2008.

Progress 01/01/07 to 12/31/07

Outputs
OUTPUTS: Objective 1: Study on soybean viruses has revealed, besides Bean pod mottle virus (BPMV) which was present in ca 75% of infected samples, infections by Bean yellow mosaic virus (BYMV), Southern bean mosaic virus (SBMV) and Tobacco ringspot virus (TRSV). Curiously, two samples were found infected by Tomato spotted wilt virus (which is the most widespread virus of tomatoes in MS). No Soybean mosaic virus (SMV) or Alfalfa mosaic virus (AMV) infections were observed during this study. Other viruses detected in Mississippi during 2007 were: a tripartite cryptic virus from Rosa spp (Rose cryptic virus-1, RCV-1), a novel putative allexivirus in blackberries (Blackberry virus E, BlVE) and an undescribed dsRNA virus associated with yellowing disease in tomatoes (Tomato yellow stunt associated virus- TYSaV). All three viruses were completely characterized. In ornamentals, Hibiscus ringspot virus (HRSV) was readily detected in several diseased hibiscus specimens collected during 2007. Objective 2: RT-PCR detection methods were developed for the routine diagnosis of newly identified viruses of rose (RCV-1), blackberry (BlVE) and tomatoes (TYSaV). Objective 3: Characterization of the putative virus infecting ornamental ferns in Mississippi and Louisiana was continued. The genome of the virus was almost completely sequenced and study on the identification of possible vectors has been initiated. Complete sequences showed that the virus genome is split in two RNA molecules. According to sequence data of the RNA-1, the virus is related to Raspberry bushy dwarf virus (gen. Idaeovirus), whereas the organization of RNA-2 differs from RBDV and appears, in part, phylogenetically closer to members of the genus Umbravirus. PARTICIPANTS: Collaborators: Dr. Nina Abou Ghanem-Sabanadzovic, Dr. Alan Henn, Dr. Rodrigo Valverde (Louisiana State University), Dr. Adib Rowhani (University of California Davis). Training/professional development: Mr. Ronald Christian Stephenson, PhD student; Ms. Estela Blanca Castillo (El Salvador)- Short training on "Diagnosis of plant viruses and virus-like pathogens" September-October 2007.

Impacts
Results of the 2007 activities showed that there are a number of viruses present in Mississippi that we were unaware of, calling for deeper study on their importance for local agriculture. Accordingly, diagnostic methods developed during the past year will allow study of the incidence and importance of these pathogens in 2008. In addition, these diagnostic methods are easily applicable in any other laboratory interested in these pathogens.

Publications

SABANADZOVIC S. and N. ABOU GHANEM-SABANADZOVIC, 2007. An undescribed virus of blackberry from Mississippi. Phytopathology 97:S102.
SABANADZOVIC S., R. A. VALVERDE, N. ABOU GHANEM-SABANADZOVIC and D. GUTIERREZ, 2007. Mycovirus-like dsRNA molecule from tomato. Phytopathology 97:S103
VALVERDE R.A. and S. SABANADZOVIC, 2007. Association of a virus with a diseases of Japanese holly fern (Cyrtomium falcatum) and leatherleaf fern (Rumohra adiantiformis). Phytopathology 97:S117.
ABOU GHANEM-SABANADZOVIC N., S. SABANADZOVIC and A.K. ROWHANI, 2007. Genomic organization, phylogeny and serology of three grapevine leafroll-associated ampeloviruses. Phytopathology 71:S1

Progress 01/01/06 to 12/31/06

Outputs
Objective 1: Field surveys were continued during 2006 in order to identify viruses present in Mississippi. In case of sweet potato, the incidence of Sweet potato virus G (SPVG) and Ipomoea vein mosaic virus (IVMV), first time reported in 2005, resulted higher than thought. These viruses were frequently detected in mixed infections, together or with Sweet potato feathery mottle virus (SPFMV). Tomato yellow leaf curl virus (TYLCV), so far reported only from 2 locations in Mississppi, was confirmed in samples from Jones County, indication that this serious problem on tomato may be spreading. Peanut production seems to be threatened by the high incidence of Tomato spotted wilt virus and aphid-transmissible potyviruses (incidence in some tested fields was almost 70-80%). In addition, some new plant host species were described for: (i) Cucumber mosaic virus (butterfly bush-Buddleia davidii, fam Loganaceae); (ii) Tobacco ringspot virus (Joe-pye weed- Eupatorium purpureum, fam. Asteraceae), etc. Objective 2: A primer set was designed and successfully applied in detection of new fern virus, which is extensively studied in the Objective 3. Objective 3: Characterization of the putative virus infecting ornamental ferns in Mississippi and Louisiana was continued. Partially purified preparations from infected fern samples showed the presence of quasi-spherical viral particles of c. 30nm. Initial molecular data indicate that this virus is evolutionary close to viruses belonging to fam Bromoviridae and gen Idaeovirus.. Additionally, an extensive survey on fern viruses in the Great Smoky Mountain National Park showed apparent absence of spontaneous virus infections in tested samples.

Impacts
Results of the last year activities, in case of sweet potato, have shown that virological situation is more complex than anticipated. Consecutively, rather frequent mixed infections may have higher negative economic impact on production than previously thought. Discovery of "new" viruses calls for better virus control through adoption of proper prevention measures and better management strategies. Accordingly, development and improvement of diagnostic methods, is aimed at early detection and timely control of viral agents.

Publications

ABOU GHANEM-SABANADZOVIC N., S. SABANADZOVIC, P. GUGERLI AND A. ROWHANI, 2006. Molecular characterization of grapevine leafroll-associated viruses 4 and 6. Proceedings of 15th Meeting of ICVG, April 3-7 2006, Stellenbosch, South Africa, 30-31 .
ABOU GHANEM-SABANADZOVIC N., S. SABANADZOVIC, J.K. UYEMOTO AND A. ROWHANI, 2006. A putative new ampelovirus associated with grapevine leafroll disease. Proceedings of the 15th Meeting of ICVG, April 3-7 2006, Stellenbosch, South Africa.
S. SABANADZOVIC, 2006. Study on viruses in the Great Smoky Mountain national Park: a prologue. 10th ATBI/DLIA Conference, Gatlinburg, TN, Dec 6-8, 2006.

Progress 07/01/05 to 12/31/05

Outputs
Objective 1: Several field surveys were carried out during 2005 in order to identify viruses present in Mississippi. In case of tomato and soybean, results were as expected, confirming Tomato spotted wilt virus and Bean pod mottle virus as major problems respectively. In contrary, several viruses, previously unreported to occur in Mississippi, have been discovered in sweet potato and various ornamental species. They are: (i) Turnip vein-clearing virus and Tobacco mild green mosaic in ornamentals, (ii) Peanut mottle virus and Bean common mosaic virus in peanut, (iii) Sweet potato virus G and Ipomoea vein mosaic virus in sweet potato. In addition, an undescribed virus was found in three blackberry samples. The virus is apparently a new viral species and seems to be associated with severe symptoms observed on original plants. Its characterization is ongoing in order to establish proper taxonomic position and study its biological, epidemiological and molecular properties. Objective 2: Development of a multiplex RT-PCR methodology for the detection of four major peanut viruses has been initiated. Preliminary results are very encouraging and research on this topic is in development. Objective 3: Preliminary data generated on the virus infections in ferns showed the presence of a distinct dsRNA pattern (indicative for the virus infections) in several symptomatic fern samples collected from Mississippi and Louisiana. These dsRNAs were reverse-transcribed and cloned. Analysis of selected clones is underway.

Impacts
Results of this project will contribute to better understand the real presence of viruses and their economic impact on production. Identification of some unreported viruses will urge for better virus control through adoption of proper prevention measures and better management strategies, especially in ornamental industry. Improvement of diagnostic methods, allowing detection of several viral species in a single reaction, will have multiple benefits from the financial (i.e. lower costs/sample) and scientific viewpoints (increased efficiency, reduced test time/sample, etc).

Publications

Abou Ghanem-Sabanadzovic N., S. Sabanadzovic, M. Digiaro and G.P. Martelli, 2005. Complete nucleotide sequence of the RNA-2 of Grapevine deformation and Grapevine Anatolian ringspot viruses. Virus Genes 30(3), 335-340.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, A. Henn, C. Balbalian and A. Lawrence, 2005. Tobamovirus infection of petunias in Mississippi. Phytopathology 95:S92.
Sabanadzovic S., N. Abou Ghanem-Sabanadzovic, A. Rowhani and J.K. Uyemoto, 2005. Detection of Cherry virus A, Cherry necrotic rusty mottle virus and Little cherry virus 1 in California orchards. Journal of Plant Pathology 87(3), 173-177.
Dreher T.W., M.C. Edwards, A.J. Gibbs, A-L. Haenni, R.W. Hammond, I. Jupin, R. Koenig, S. Sabanadzovic, N. Abou Ghanem-Sabanadzovic and G.P. Martelli, 2005. Family Tymoviridae. In: Fauquet C.M., M. Mayo, J. Maniloff, U. Desselberger, L.A. Ball (eds.) Virus Taxonomy (VIIIth Report of the ICTV), Elsevier/Academic Press, London, pp 1061-1074.