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WERA_OLD020: Virus and Virus-Like Diseases of Fruit Trees, Small Fruits, and Grapevines.

Annual/Termination Reports (SAES-422): [07/26/2007] [10/22/2008] [09/11/2009] [11/19/2010] [07/25/2011]

Date of Annual Report: 07/26/2007

Report Information:
  • Annual Meeting Dates: 05/30/07 to 06/01/07
  • Period the Report Covers: 01/2006 to 12/2006

    • Participants:
    Brief Summary of Minutes of Annual Meeting:
    WERA-020: Virus and Virus-Like Diseases of Fruit Trees, Small Fruits, and Grapevines Committee - Annual Meeting May 29 to June 2, 2007 in Leamington, ON

    Chair and Local Organizer: Roberto Michelutti, Canadian Clonal Genebank, Agriculture & Agri-Food Canada, Harrow ON; Secretary: Ken Eastwell, Washington State University, Prosser, WA

    Meeting convened at 08:30 on May 30, 2007:

    Chair Michelutti welcomed the participants to the meeting; Cavalieri (Administrative Advisor) reviewed the renewed structure of WERA's to bring research and extension personnel together.

    Attendees represented the States and Provinces of British Columbia, California, Colorado, Hawaii, Manitoba, Michigan, Ontario, Oregon, Pennsylvania, and Washington

    STATE/PROVINCE REPORTS: Eastwell (WA - report attached on WERA-20 Homepage - Additional Documents section) reported on the development of a more robust assay Little cherry virus-1. There is growing concern that the milder symptoms caused by LChV-1 on current biological indicators may allow the virus to escape detection. On-going research on the cherry isolate of Cherry leafroll virus suggests that this virus can be transmitted to the fruit bearing tree by virus-infected pollen. Serological reagents are being developed to detect members of the genus Foveavirus that affect sweet cherry. Genetic analysis reveals a complex relationship between the members of this genus. The National Research Support Project-005 located at WSU-Prosser, continues to be the primary source of new virus tested fruit tree propagation material that serves both import and export of fruit tree propagation material.

    Guerra (WA) indicated that Cherry leafroll virus had been detected in a certified seed block and increased monitoring is in place to minimize threats from this virus in the certification program. The tree fruit certification program is auditing the certification program to determine the status of Apple chlorotic leafspot virus and is finding some unexpected positives.

    Halbrendt (PA - report attached on WERA-20 Homepage - Additional Documents section) reported on the status of the Plum pox virus eradication program initiated after the virus was confirmed in PA in 1999. The mandatory quarantines in large areas of the State have been rescinded since no plum pox virus has been detected in these areas for 3 years. Prunus species can now be replanted in these areas, although nursery activity is still prohibited. In 2006, six positive plants were identified and only one was in a commercial orchard. Trap plants and yellow traps continue to be analyzed for viruliferous aphids and possible unknown reservoirs of the virus.

    Hu (HI - report attached on WERA-20 Hompage - Additional Documents section) indicated that a survey of citrus trees on the five major islands revealed an overall incidence of Citrus tristeza virus of 74%. Some infections could be detected with either serological or molecular assays, but not by both. Further analyses indicate that most trees are simultaneously infected with multiple strains of the virus. From this study, a strain that can be used as a cross protection agent was identified; field trials are in progress to evaluate this potential. Viruses associated with a citrus blight-like wilt disease in HI are under investigation to determine the nature and etiology of this disease. The long term goal is to develop a management strategy to allow the citrus industry to expand.

    James (B.C.) has characterized Peach virus Agua (formerly identified as 4N6). This virus was identified through its reaction in ELISA with antiserum prepared to Plum pox virus. The virus also reacts in EM decoration assays with antiserum to Apple stem pitting virus. PcVA shares sequence and genome organization similar to ASPV, and share many peptide sequences in the C-terminus of the coat protein region. However, PcVA is a distinct member of genus Foveavirus in the Flexiviridae family.

    Kalinina (ON - report attached on WERA-30 Homepage - Additional Documents section) examined the susceptibility of ornamental Prunus species to Plum pox virus. Generally, ELISA results of self-rooted plants were less definitive than those from tree grafted onto GF305. Inoculated P. americana, P. cistena, P. glandulosa, P. tomentosa developed distinct symptoms and the infection was confirmed by immunocapture RT-PCR, ELISA and by back inoculations. P. triloba yielded weak reactions whereas P. virginiana 'Schubert', P. serrulata 'Kwanzan' yielded inconsistent results that need further clarification. These are important potential host plants for PPV in North America.

    Ferguson (ON) reported on the status of efforts in Canada to contain Plum pox virus. No PPV has been detected in two sites in Nova Scotia or three areas in Ontario for three years, so these areas are now removed from the quarantine area. However, the major quarantined area in Ontario remains. Results of the 2006 survey revealed 18 positives in the St. Catherines area. Eradication will only be possible with aggressive testing and enforcement. A group discussion followed regarding strategies that might be feasible to reduce the aphid transmission of PPV.

    Larsen (CO - report attached on WERA-20 Homepage - Additional Documents section) established in-pot trials to determine the potential resistance of new rootstocks to transmission of Cherry raspleaf virus. Soil containing Bing trees on mazzard and 'Citation/Z-stem' rootstocks were infested with viruliferous nematodes in spring 2006. No symptoms are evident yet on the inoculated trees. Grape fanleaf virus was detected in a block of 'Cabernet franc' in 2006. The infection appeared to be localized although significant populations of dagger nematodes were detected. Studies will continue to determine if the CO populations of nematodes are vectors of GFLV.

    Tiffany (Agdia) reported that a new ELISA test is available that recognizes all known strains of Plum pox virus.

    Mock (USDA-NGRL-PDRU - report attached on WERA-20 Homepage - Additional Documents section) indicated that reorganization of the unit is complete and they are now working in close associated with USDA-APHIS to develop methods for the safe introduction of germplasm. A molecular assay has been reported to supplement the biological indicators currently used for the detection of Gooseberry vein banding virus. This will shorten time requirements for virus testing. A molecular assay for Cherry green ring mottle virus and Cherry necrotic rusty mottle was also developed. Some isolates of Cherry necrotic rusty mottle do not produce symptoms on the standard biological indicator 'Sam'. A RT-PCR assay for black currant reversion has been developed but validation is still required.

    Lemmon (ON) indicated that audits of programs in France and Germany cause concern about phytoplasma diseases and led to the detection of phytoplasma (Bois noir) in a single shipment of vines imported into the Okanagan Valley. It is believed that grapevines are a dead end host for the phytoplasma, and that it moves into grapevines from weedy reservoirs. The known vector of the disease does not occur in B.C. Therefore, it is believed that removal of the two infected vines effectively eradicated the Bois noir. Future importations require hot water treatment to eliminate potential Bios noir infestations.

    Results from Rowhani (CA - report attached on WERA-20 Homepage - Additional Documents section) indicate that Plum bark necrosis and stem pitting associated virus shares 97% with Apricot stem pitting virus in Italy, and has been found in cherry and plum. Sequence analysis indicates that this virus has a close relationship to Pineapple mealybug wilt associated virus and some of the grapevine leafroll associated viruses.

    Msikita (PA - report attached on WERA-20 Homepage - Additional Documents section) is investigating the occurrence of Tomato ringspot virus in apple trees where it causes apple decline and eventual death. ELISA detection of ToRSV has been inconsistent and the more sensitive RT-PCR has revealed a much higher incidence of ToRSV. This underscores the need to re-examine current virus detection methods and the need for robust detection methods.

    Anderson (AC Diagnostics) indicated that ELISA reagents are available for PVY-n and that AC Diagnostics will also perform contract research.

    Schilder (MI) detailed the discovery of Plum pox virus in July 2006 at the South West Michigan Research and Extension Center. The infected 5-year old tree was an European plum on Pumiselect® rootstock and did not exhibit any symptoms. All Pumiselect trees in the NC-140 rootstock trial have been removed. Testing of all hosts within a 5 mile radius did not reveal any other infected trees. Based on these results, fruit production is allowed in the regulated area, but no nursery production within the 7 mile area. This included advanced selection of the breeding program and 58 elite selections were sent to NRSP-005 for virus testing and retention.

    Nurseries in the MI blueberry certification programs are testing for shoestring, mottle, tobacco ringspot virus and tomato ringspot virus, and will include blueberry scorch.

    Thompson (BC - report attached on WERA-20 Homepage - Additional Documents section) is evaluating the use of the 'TriFoCap' primers for the detection of tricho-, fovea-, and capilloviruses. These primers successfully detected 20 virus isolates from these genera and, with only one exception, detected them throughout the year. Cherry necrotic crook disease could only be detected in dormant wood. It was reported that of 247 Prunus samples, 3 false positive reactions were obtained from cherry, and from 181 pome fruit samples, 3 false positive were obtained. Therefore, caution in interpretation of results is necessary, the case for all PCR assays.

    Tian (ON) is investigating the use of transgenic plants for resistance to Plum pox virus. The HC-Pro segment is an effective activator of post transcriptional gene silencing in Nicotiana benthamiana, and transformation of plum indicates that a high percentage of transformants are resistant to infection by the D strain of PPV. The advanced selections are sent to France for evaluation for resistance to other strains.

    Tzanetakis (OR - report attached on WERA-20 Homepage - Additional Documents section) summarized the determination of the viruses associated with the rapid decline of strawberry in CA (predominantly Strawberry pallidosis virus). This disease situation was resolved through aggressive management techniques. Similar disease symptoms occur in B.C. but the aphid-borne Strawberry mild yellow edge virus is the underlying cause of decline. Laboratory tests for more than 20 viruses affecting strawberry have been developed to aid in the management of strawberry diseases. Raspberry bushy dwarf is re-emerging as an important disease in raspberry and more recently, in 'Marion' blackberry which develops severe fruit loss in response to infection. Transgenic plants of 'Meeker' red raspberry have been developed and preliminary testing showed that several lines offer robust field resistance. Blueberry scorch has become an important virus, particularly with the discovery that cranberry and huckleberry are latent hosts of the virus. Virus elimination programs are underway in OR. Status of several blackberry viruses was also described.

    Michelutti (ON) expanded the indexing program of the Canadian Clonal Genebank collection of more than 3300 unique accessions. Indexing on 'Kwanzen' gave 14 positive reactions last year and 25 this year. In addition, 224 Prunus and 134 Fragaria were tested for nepoviruses. In parallel, 6 Fragaria were found to be infected with Strawberry mild yellow edge virus. Rubus accessions in the screenhouse (91) were tested for nepoviruses, Raspberry bushy dwarf virus and Tobacco streak virus. Three accessions were positive for RBDV and one was positive for TSV. Eighty-five Rubus accessions in the field were tested and 4 were infected with RBDV and 1 for TSV. Nineteen representatives of the 'Jesuit pears' have been transferred to the germplasm collection for genetic analysis at a later date. Last year, symptoms developed on Ribes similar to aphid damage or Gooseberry vein banding virus. Virus infection was confirmed by PCR, and a second virus was also detected. Tissue print blots in cooperation with Palles and Flores are being used to evaluate the viroid status of the collection.

    Reports (attached on WERA-20 Homepage - Additonal Documents section) were submitted by WERA-020 participants unable to attend: Deborah Golino, Ray Johnson, Simon Scott, and Aiming Wang.

    Photographs attached to WERA-20 Homepage area.

    Business meeting:

    Cavalieri indicated that the project was revision was submitted in 2005 and authorized in 2006. The project will submit its next revision in 2010.

    Eastwell gave a brief summary of the planning meeting of the National Clean Plant Network that was held in Greenbelt, MD on May 9-10. The successful creation and funding of the network would be an important step forward in controlling virus diseases of perennial fruit crops in the U.S. However, it will require several fundamental changes in the way industries and government agencies interact. WERA-020 participants may have a larger role in developing some of the guidelines for network operations.

    Larsen offered to host the nest meeting of WERA-020 in Grand Junction, CO. Michelutti moved and Rowhani seconded to hold the meeting in Grand Junction during the middle two weeks of June 2008 - motion passed unanimously.

    Larsen moved and Hu seconded that the WERA-020 meeting be held in MI in 2009 - motion passed unanimously.

    Larsen moved and Eastwell seconded that WERA-020 participants formally extend appreciation for the efforts made by the hosts at Agriculture & Agri-Food Canada and the local organizer Roberto Michelutti for making the meeting both informative and enjoyable - motion passed unanimously.

    Meeting adjourned.

    Accomplishments:
    Plum bark necrosis and stem pitting associated virus causes severe necrosis of bark tissues, and necrotic pitting on the woody cylinders of Prunus salicina. PBNSPaV was alsdo identified in other Prunus species and cultivars: almond cv. 'Filippo Ceo', peach cvs. 'Springcrest' and 'May Crest', plum cv. 'Black Beaut', apricot cv. 'Castlebrite', and sweet cherry cv. 'Ferrovia'. The virus was detected in samples from Italy, Morocco, and Turkey.

    A robust molecular assay for Little cherry virus-1 from disperse geographic locations was developed and verified.

    Several ornamental Prunus species were identified as hosts for Plum pox virus. Plants on their own roots supported a lower virus titer and therefore may elude detection.

    Group specific primers for the detection of tricho-, fovea-, and capilloviruses were evaluated and found to be a reliable and valuable tool for rapid detection of any one of many viruses that affect temperate fruit trees.

    It was demonstrated that the rapid decline of strawberries in CA and BC was caused by two different viruses emphasizing the need to develop site-specific disease management tools.

    Impact Statements:
    1. The loss during the 2003-2004 season in California strawberry caused by Strawberry pallidosis associated virus (SPaV) was estimated to $25M. With identification of the pathogen and aggressive pest management the disease has almost been eliminated for the state.
    2. The TriFoCap test was evaluated against a range of viruses mostly obtained from WERA-20 participants. The TriFoCap nested RT-PCR test is a useful tool for screening fruit tree samples for Tricho-, Fovea-, and Capilloviruses and can make the testing of quarantinable material more efficient and more rapid.
    3. Quarantine regulations and the high cost of bringing in new stock from abroad make it critical for growers and researchers to be able to locate existing material already in the US. The National grape registry was created to provide a single comprehensive site listing all grape plant material available within the U.S. and to identify material that has been tested and certified as clean in regard to certain grapevine diseases.
    4. Hawaii is capable of producing high quality citrus fruit that is often preferred over imported citrus whose quality has suffered from the long trip to the islands if it were not for the high incidence of Citrus tristeza virus. The development of citrus resistant to Hawaiian CTV strains would be beneficial to not only Hawaii, but also other citrus-producing regions in the US and abroad.
    5. Results from PDRU research projects will benefit the USDA quarantine program by producing more effective pathogen detection methods, improving knowledge on etiology of poorly described pathogens, and the development of therapeutic methods to eliminate pathogens from potentially valuable germplasm. These improvements will help create a more effective quarantine program that encourages compliance with federal regulations regarding movement of germplasm and diminishes the temptations to smuggle plant material into the United States.
    Last Modified: 01-Aug-2007

    Date of Annual Report: 10/22/2008

    Report Information:
  • Annual Meeting Dates: 08/11/08 to 08/13/08
  • Period the Report Covers: 10/2007 to 09/2008

    • Participants:
    Brief Summary of Minutes of Annual Meeting:
    Minutes: H. Larsen, acting secretary.

    The 2008 annual meeting of WERA-20 was held Aug. 11-13, 2008 at the Holiday Inn in Grand Junction, CO.

    Participants:

    Guerra, Lauri (lguerra@prosser.wsu.edu) - WA State Dept. of Ag
    Halbrendt, John (jmh23@psu.edu) - Penn St. Univ.
    Hu, John (Johnhu@hawaii.edu) - Univ. of Hawaii
    Larsen, Harold (Harold.Larsen@colostate.edu) - Colo. St. Univ.
    Licha, Margarita (Margarita.F.Licha@aphis.usda.gov) - USDA-APHIS-, Beltsville
    Mock, Raymond (Raymond.Mock@ARS.USDA.GOV) - USDA-ARS, Beltsville
    Pokharel, Ramesh (Ramesh.Pokharel@Colostate.edu) - Colo. St. Univ.
    Vrient, Albert (avrient@agdia.com) - AgDia

    The meeting opened at 9:00 am, Monday, Aug. 11 with a welcome by H. Larsen (local host). In the absence of the official chairman for the meeting, the group decided to have H. Larsen serve as the chairperson for the meeting. The projected secretary also was unable to attend, so H. Larsen agreed to record the meeting for the minutes and to prepare the minutes after the meeting conclusion.

    State / Agency reports:

    Pokharel (CO) - See written report: He reported on his work with dagger nematodes and Cherry Rasp Leaf Virus (CRLV), one of the primary virus disease problems facing cherry and apple growers in Colorado. Dagger nematodes are very common in soils within the area, but species identification in Xiphinema is difficult. Collaborative work to look at molecular identification of dagger species from Colorado Soils is underway with A. Szalanski (Univ. of Ark.). Pot-in-pot studies were established in 2006 to examine comparative susceptibility of Bing cherry on Citation / Z-interstem rootstock and on Mazzard rootstock to CRLV infection. No symptoms have been observed in any of the scions or rootstock suckers to date. Observations continue on Van and Bing cherry on Gisela 5 and Gisela 7 rootstocks for susceptibility / resistance to CRLV. Nicotiana benthamiana mechanically inoculated with CRLV using common buffer and carborundum powder in 2007 failed to show any symptoms. Apple and peach inoculated with diseased cherry as the tissue source have not produced any symptoms to date.

    An incidence of Fanleaf virus on Cabernet franc grapes was found in fall 2006. No spread was detected via ELISA tests in adjacent vines nor in an adjacent block of Cabernet sauvignon, despite the presence of a substantial dagger nematode population in these soils. The block was removed in winter 2006-07, fumigated in late summer 2007, and replanted in 2008. It and the neighboring block are being monitored for any sign of spread.

    A lively discussion of nematode associated viruses and vector nematode identification followed. Morphological variability of dagger nematodes found in Colorado soils is a great difficulty for identification. Halbrendt (PA) suggested establishing single female colonies for ID purposes (minimize morphological and genetic variability).

    Hu (HI) - See written report. Pineapple mealybug wilt (PMW) research update: PMW has now been shown to be associated with four viruses (PMWaV-1, PMWaV-2, PMWaV-3, and PMWaV-4; all Ampeloviruses). Based on sequence available for PMWaV-4, it is distinctly different from the other three, PMWaV-1 toPMWaV-3. Badnavirus studies update: Cloning and sequencing of badnavirus-like particles in pineapple revealed the presence of four distinct clades. The phylogenetic relatedness of these clades to other badnaviruses, other Caulimoviridae members, and retro-like elements was examined. Grey pineapple mealybug had 100% transmission efficiency after a 3-day acquisition period and the PMWaV-2 virus persisted in these viruliferous mealybug vectors up to 3 days. Thus PMWaV-2 is transmitted by the grey pineapple mealybug in a semi-persistent manner. Longtailed mealybug also was shown to be a vector. Reverse transcription-polymerase chain reaction (RT-PCR) and PCR assays for PMWaV's and badnaviruses were used to determine the distribution and incidence of these viruses in two new commercially important pineapple hybrids and the germplasm accessions maintained at the ARS-USDA National Clonal Germplasm Repository in Hilo, HI. Differences were found within pineapple cultivars and hybrids with regard to incidence of PMWaV-2 and -3 and badnavirus clades A, C, & D. The high incidences of some PMWaVs and badnaviruses in newly-imported pineapple hybrids verify the importance of using virus screening assays to select virus-free plant material prior to mass propagation.

    Halbrendt (PA) - See written report. Reported on the Plum Pox virus (PPV) situation. The PPV eradication project in PA seems to be working at this time. The regulated areas in PA have decreased from a high of 349 sq. miles in 2003 to 51 sq. miles in 2008. The 2007 survey produced negative results for the first time since the detection of PPV in 1999. Two small areas in Adams County and one in Cumberland County remain under regulation for PPV, but no positive or suspect samples were found in 2007. In New York, samples positive for PPV have been reported in Niagara County and suspects in Wayne County. A total of 26.7 acres of orchard were destroyed in 2007 because of the positive detections in 2006 and 2007. NY currently is using a 50-meter minimum removal buffer around any positive detection due to the close proximity of the infections to the Canadian PPV infected area and the uncertain risk of reinfection via the natural movement of aphids between the two areas. The PPV infection found in Michigan was on a Pumiselect rootstock from OR. One aspect of concern was that trees on Pumiselect rootstock were also sold to WalMart which will make tracking more difficult.

    Guerra (WA) - See written report (from K. Eastwell, WA). Washington Dept. of Agric. work: Recently encountered a problem with CLS virus in some WA nursery material; he is looking at possibility of field transmission. There is a need for better serological detection methods for cherry mottle leaf (peaches can serve as symptomless hosts), little cherry (becoming an increasingly frequent problem), and western-X disease (seems to be on the increase now). He also is looking at peach latent mosaic viroid and prune dwarf virus (PDV, specifically interactions with other Prunus viruses).

    K. Eastwell's work: He has successfully produced monoclonal antibodies against Little cherry virus 1 (enhancing detection of this virus for quarantine / certification programs). Cherry leafroll virus (CLRV) is pollen-borne (successfully transmitted CLRV from pollen to the developing fruit), but controlled pollination of several thousand blossoms has not demonstrated transmission of the virus to the fruit-bearing tree; the virus gets as far as the sub-epidermal cell layer of the pedicel, but no further. Work on Foveaviruses in fruit trees and grapevines has focused on characterization, examination of molecular similarities, and development of polyclonal antibodies to the bacterial-expressed coat protein of a representative virus on one of the four distinct clades; this antibody reacts with all viruses within the same cluster. Another polyclonal antibody was developed for Rupestris stem pitting associated virus (RSPaV). Hop stunt viroid (HSVd) is of growing concern to hop producers; a hop root certification program has been revitalized to provide HSVd and virus-free roots to nurseries. The Washington Hop Commission has invested heavily in equipment and facilities for molecular detection of HSVd.

    Main activities of NRSP-005 included: 1) rescuing the Mich. St. Univ. Peach breeding program after discovery of PPV at an MSU research station; 2) assisting USDA-APHIS-PPQ by providing therapy to virus-infected clones; & 3) distributing over 17,700 virus-tested buds to researchers & industry; & 4) virus testing and therapy of 185 new acquisitions of stone and pome fruit cultivars from nine foreign countries and eight U.S. states.

    Creation of a National Clean Plant Network was mandated in the 2008 Farm Bill under "Te Food, Conservation, and Energy Act of 2008" and the legislation includes the establishment of clean plant centers for pathogen diagnosis and elimination. These will produce clean material for propagation and will maintain foundation blocks of pathogen-tested plant material throughout the USA. Organizational meetings have been held in Prosser, WA and Davis, CA to create a structural model for the network. The goal is to have the organization in place by Oct., 2008.

    Licha (USDA-APHIS-PGQP) - On Oct. 1, 2005, APHIS assumed responsibility for the plant quarantine program in Beltsville, MD. Program Supervisor - Dr. Joe Foster; Team Leader - Dr. Margarita Licha (Fruit Plant Pathologist - Plant Germplasm Quarantine Program). She reviewed the policy for importing budwood and described the testing protocols used to detect pathogens for both pome and stone fruits and then provided a summary of program activities, goals, and needs for the future.

    Mock (USDA-NGRL-PDRU) - See written report. The National Germplasm Resources Lab - Plant Disease Research Unit (NGRL-PDRU) performs research on viral and sub-viral pathogens of clonally propagated prohibited crop genera, emphasizing deciduous tree and small fruits, sugarcane, grasses, and sweet potoatoes. The mission of the unit is to characterize and investigate the etiology of poorly described diseases and pathogens of quarantine significance and to develop more reliable detection and elimination methods. Five current projects related to deciduous fruits include the following: 1) Development / adaptation of existing protocols for in vitro culture and therapy of infected stone and small fruits; 2) characterization of flowering cherry isolates of Cherry necrotic rusty mottle virus (CNRMV); 3) etiological investigations of several poorly characterized quarantine diseases of pome fruits (including flat limb/rubbery wood); 4) development of a detection method for six viroids that infect pome & stone fruits; and 5) etiological investigations of what is believed to be Stayman Blotch Virus in apple in the Shenandoah Valley, VA (w/ Dr. Keith Yoder, Va. Tech. Ag. Res. & Ext. Ctr., Winchester, VA). Results from PDRU research projects will benefit the USDA quarantine program by producing more effective pathogen detection methods, improving knowledge on etiology of poorly described pathogens, and developing therapeutic methods to eliminate pathogens from potentially valuable germplasm. An adjunct benefit is helping create a more effective quarantine program that encourages compliance with federal regulations regarding movement of germplasm and diminishes the temptations to smuggle plant material into the United States.

    Cavalieri (WERA-20 Administrative Advisor) - Phone conference call, covered the following administrative items:

    1) WERA-20 is approaching the time for the required Mid-Point Assessment of the committee. He will send electronic files of two forms to H. Larsen to forward to all members to fill out in April, 2009. Please give these your attention when you get them. He needs to submit a summary of each of these by June 1, 2009. Specifically, he needs each member to provide a one sentence "Impact Nugget" and a one sentence statement of "Who Cares & Why".

    2) Update on the "Clean Plant Network" - a) National structures have been established for tree fruits and for grapes; & b) funding for NRSP-005 has been set for this year (to bridge to future funding arrangements).

    3) The "Specialty Crops Research Initiative" - connections (multi-state) will be very important in successful submissions for these.

    4) Avoid August for future meetings (participation this time is one reflection of problems for this timing). Face-to-Face meetings (annual) are NOT required for the committee - could be done via conference call if travel expenses should become prohibitive in the future. This might be a possibility for future years.

    5) Hatch funds for next year - Few of the appropriation bills are likely to be passed until the next Congress (do have continuing resolution funding until such, at the same levels as prior budget). [Ray Mock (USDA-NGRL-PDRU) - funding under continuing resolutions until next spring on a month-by-month basis, 1/12 of budget each month.]

    Vrient (AgDia) - Changes at AgDia are on the horizon. Mike Tiffany is contemplating retiring soon; fruit is being given increased importance in the restructuring as plant groups are being shuffled around. Albert Vrient is likely to assume stone fruit and ornamentals areas.
    Experienced elevated PPV suspect detections (possibly elevated background levels) recently. Question to group as to possible causes: Halbrendt  weeds have lots of natural materials within them that react to the system. [AgDia: used antisera from Canada: rabbit source]. Guerra  possibly heatshock protein confounding results.

    Possibilities for future? - Guerra - field detection strips for fruit (Immunostrip w/ dried reagent, immuno test using gold-labeled antibody; 30 min / test). Cost varies, GMO cheaper (viral/bacterial tests $2-5 each). Need for a good, simple tool for growers. TmRSV tests? - not currently available.

    Business Meeting: Chaired by H. Larsen.

    * Next years location (for 2009 meeting): MSP (Mock/Hu) - to accept invitation from Michigan St. Univ. to host the meeting.
    * 2010 meeting location: MSP (Mock/Hu) - to go to Pennsylvania (WSU-Prosser as a backup).
    * Date for 2009: MSP - Recommendation to look at May / June (virus symptoms better for field trips).
    * Chairperson for 2009: MSP - to have local host or his/her designee serve as chairperson.
    * Secretary for 2009: MSP - Leave to decide at the meeting.

    New Business:

    - L Guerra: There is a need for a manual of pome / stone fruit virus / virus-like diseases in order to capture info from people who are retiring. It needs to be peer reviewed and might best be in electronic media format. It is something to think about for next year. He will prepare a format suggestion to consider at the 2009 meeting.

    Meeting was adjourned at 4:30 pm.

    Field trips:

    Tues: Field trip to Delta County orchards; return via the Grand Mesa scenic highway.

    Wed: Field trip to Mesa County orchards (morning); tour of the Colorado National Monument (afternoon).

    Reports submitted by WERA-20 members unable to attend 2008 meeting:
    See Meeting Minutes Attachment section for full texts of individual reports.

    Almeida (CA; UC-Berkeley)> - Transmission of grapevine leafroll-associated virus 3 by the vine mealybug: Grapevine leafroll disease is caused by grapevine leafroll-associated viruses (GLRaVs). Within this virus complex, GLRaV-3 is the predominant species in the world. Several GLRaVs have been shown to be transmitted from vine to vine by mealybugs although a detailed characterization of transmission biology is lacking. The introduction of the vine mealybug (Planococcus ficus) in California and other regions of the world may result in increasing disease incidence of established GLRaVs. We studied the characteristics of GLRaV-3 transmission by the vine mealybug. Our results indicate that the vine mealybug transmits GLRaV-3 in a semipersistent manner. First instars were more efficient vectors than adult mealybugs. GLRaV-3 transmission lacked a latent period in the vector. Virus transmission occurred with a 1-h acquisition access period (AAP) and peaked with a 24-h AAP. Mealybugs inoculated GLRaV-3 with a 1-h inoculation access period (IAP), and transmission efficiency increased with longer plant access period up to 24 h, after which transmission rate remained constant. After an AAP of 24 h, mealybugs lost GLRaV-3 and infectivity 4 days after virus acquisition. In addition, GLRaV-3 was not transovarially transmitted from infected females to their progeny as detected by reverse transcription polymerase chain reaction. In summary, we systematically analyzed transmission parameters of GLRaV-3 by the vine mealybug and showed that transmission of this virus occurs in a semipersistent manner. This research fills in important gaps in knowledge of leafroll virus transmission, which is critical for development of leafroll disease management practices.

    Rowhani (CA; UC-Davis) - Real time TaqMan® RT-PCR (TaqMan® RT-PCR) assays were developed to detect the viruses associated with Rugose wood complex of grapevines. The viruses detected were Rupestris stem pitting-associated virus (RSPaV) in the genus Foveavirus, Grapevine virus A (GVA), Grapevine virus B (GVB) and Grapevine virus D (GVD) in the genus Vitivirus. The coat protein was found to be the most conserved gene within the viral species, therefore, the primers and probes for TaqMan® RT-PCR assays were designed from the multiple alignment of the coat protein sequence of various isolates of each virus. Comparisons were also made between the conventional one step RT-PCR and TaqMan® RT-PCR for the detection of these viruses using four fold serial dilutions of both purified RNA and crude extract prepared from grapevine tissue. Results showed that TaqMan® RT-PCR was more sensitive and could detect viruses at 32 and 256 fold higher dilutions for purified RNA and crude extract, respectively, compared to RT-PCR.

    In a different experiment, Low Density Arrays (LDA) were designed based on the real-time RT-PCR (TaqMan®) assays for the specific detection of 13 viruses that infect Grapevines. The viruses included in the study were Grapevine leafroll associated viruses (GLRaV) types 1, 2, 3, 4, 5, and 9, Grapevine leafroll associated virus 2-Redglobe strain (GLRaV-2RG), RSPaV, GVA, GVB, Grapevine Fanleaf Virus (GFLV), Tomato Ringspot virus (ToRSV), and Grapevine Fleck virus (GFkV). The LDAs were evaluated against a wide range of geographically distributed isolates. Geographical locations included Africa, Europe, Australia, Asia, Latin America and the United States. High throughput detection of these viruses using LDAs was compared to RT-PCR and real time TaqMan® RT-PCR. The efficiency of different RNA extraction methodologies and buffers were compared for use in low density array detection. In addition improving the RNA extraction technique and testing the quality of the RNA using the 18S ribosomal RNA TaqMan® assay as an RNA specific internal control proved to generate better diagnostic assays. This is the first report on use of LDA for the detection of plant viruses.

    Scott (SC) - Research on the bloom delay associated with the peach germplasm Ta Tao 5 and characterization of the virus and virus-like agents associated with this phenomenon has continued. Ta Tao 5 is infected with Peach latent mosaic viroid (PLMVd), Apple chlorotic leaf spot virus (ACLSV), and Asian prunus virus (APV). The complete nucleotide sequences of the isolate of PLMVd and ACLSV (GenBank Accession EU223295) have been determined. APV has been partially purified and approx 50% of the sequence of the virus has been determined.

    Ta Tao 5 that had been heat-treated at NRSP-5, Prosser, WA has been shown to contain only APV according to repeated PCR tests. In 2008 trees of heat-treated T a Tao 5 exhibited no delay in bloom compared to other peach cultivars commonly grown in South Carolina. Trees of Ta Tao 5 bloomed at least 10 days after the heat-treated material.

    A trial involving trees of the peach cultivar Springprince inoculated with different sources of ACLSV, PLMVd, and APV (heat-treated Ta Tao 5) and compared with trees that had been inoculated by transferring the mixture contained in Ta Tao 5 has been established. In 2008, only those treatments which contained PLMVd displayed bloom delay.

    The APV infecting Ta Tao 5 appears to be a mixture of at least APV-1 and APV-3 (Marais et al., 2006. Virus Research 120: 176-183) and further research is needed to confirm this. A second trial involving the cultivar Juneprince and refined treatment mixtures based on the preliminary data from the trial with Springprince has been established and will be monitored for the next 5 -7 years.

    See Meeting Minutes Attachment section for full texts of individual reports.


    URL: Copy of minutes

    Accomplishments:
    1. High incidences of some PMWaVs and badnaviruses were found in newly-imported pineapple hybrids; the discovery verifies the importance of using virus screening assays to select virus-free plant material prior to mass propagation.

    2. Systematic analysis of transmission parameters for GLRaV-3 by the vine mealybug showed transmission of this virus occurs in a semipersistent manner. This research fills important knowledge gaps for leafroll virus transmission that are critical for development of leafroll disease management practices.

    3. TaqMan® RT-PCR assays were found to be more sensitive and could detect viruses at 32 and 256 fold higher dilutions for purified RNA and crude extract, respectively, compared to RT-PCR, for viruses associated with Rugose wood complex of grapevines. The viruses detected were Rupestris stem pitting-associated virus (RSPaV) in the genus Foveavirus, Grapevine virus A (GVA), Grapevine virus B (GVB) and Grapevine virus D (GVD) in the genus Vitivirus. The enhanced efficiency of detection will help reduce risk of release of grape materials infected with these viruses to plant nurseries and thus lower incidence of such in plant materials purchased by grape growers.

    Impact Statements:
    1. Regulatory agencies often have difficulty accessing current and accurate scientific information and advice. The WERA-20 provided a forum for accessing this information, which is often unpublished, for use in formulating regulations and making decisions which have a major effect on the fruit industry.
    2. Eradication efforts for Plum Pox Virus in Pennsylvania have greatly reduced the area of incidence from 2004 to 2008. Continued vigilance and lack of positive samples for 3 years will allow producers to replant stone fruit trees and return to production. This demonstrates the value of concerted effort to eradicate such diseases.
    Last Modified: 22-Oct-2008

    Date of Annual Report: 09/11/2009

    Report Information:
  • Annual Meeting Dates: 06/14/09 to 06/17/09
  • Period the Report Covers: 10/2008 to 09/2009

    • Participants:
    Brief Summary of Minutes of Annual Meeting:
    Meeting Program:

    Chair, Local Organizer and Secretary: Annemiek Schilder, Michigan State University, East Lansing, Michigan.

    Sunday June 14: arrival, 8-10 pm: Welcome reception.

    Monday June 15 (8:30 am-5:30 pm): State reports by participants. See the attached full reports under the link for the Meeting Minutes. A special presentation on the National Clean Plant Network was made by Joseph Postman.

    Attendees visited the MSU Southwest Michigan Research and Extension Center to view virus symptoms in grape vineyards, and cherry and apple orchards. A presentation was made detailing the response to discovery of Plum pox virus in the MSU breeding program and the operations to rescue advanced peach selections. Nurseries were toured to observed tissue culture practices to produce blueberry plants; symptoms of several viruses were also observed in neighboring blueberry plantings.

    Tuesday June 16: (9 am-5 pm): MSU Southwest Michigan Research and Extension Center (SWMREC, Benton Harbor, Michigan)