Control of Eutypa Dieback of Grapes

AVF Proj. ID: 9408 / /Cat.: ,
Primary Investigator(s): James J Marois, Douglas Gublerby

The objective of our research is to find an effective control for Eutypa dieback, caused by the fungus Eutypa lata. Specific objectives were to test efficacy of the fungicides Benlate 50 WP, Rally 40 WP, and Nustar 20 DF, to determine whether multiple applications provide greater control than one application, and to test efficacy of the fungi Cladosporium herbarum, Fusarium lateritium, Aureobasidium pullulans, Trichoderma viride as biological control agents. Other objectives were to study colonization of wounds by applied fungi, to determine how the fungi inhibit infection by Eutypa, to test the combination of Benlate and Fusarium applied together, and to monitor the natural flora of pruning wounds of grapevines. We are testing the fungicides mentioned above and biological control agents in vineyards for their ability to prevent infection by Eutypa. This disease is difficult to study, because several years can elapse between the time of infection and the appearance of symptoms. By reisolating Eutypa from treated spurs after only nine months, we can assess efficacy long before symptoms appear. Our studies have shown that all four fungi can successfully colonize the wound surface when spore suspensions are applied immediately after pruning. Colonization is less successful when application of the spore suspensions is delayed for a period of one or two weeks. All four fungi are effective in reducing infection by Eutypa, but those studies must be conducted over many years to observe results over a variety of weather conditions. Studies on the natural wound colonization showed that numbers of filamentous fungi, yeasts, and bacteria are initially zero, and increase rapidly for 7-14 days after pruning, then remain constant.

Control of Eutypa Dieback of Grapes

AVF Proj. ID: 9307 / /Cat.: ,
Primary Investigator(s): James J Marois, Douglas Gublerby

The objective of our research is to find an effective control for Eutypa dieback, caused by the fungus Eutypa lata. This disease is difficult to study, because several years can elapse between the time of infection and the appearance of symptoms. Specific objectives this year were to continue testing efficacy of the fungicides Benlate 50 WP (multiple applications). Rally 40 WP, and Nustar 20 DF, to study colonization of wounds by applied fungi, to determine how the biological control agents inhibit infection by Eutypa, to test the combination of Benlate and Fusarium applied together, and to monitor the natural flora of pruning wounds of grapevines. The fungicide experiments are still in progress, but data obtained thus far indicate that applications of benlate at 1 and 14 days after pruning is more effective than only one application at pruning. The second application appears to protect the wound during the later half of the wound healing process which ultimately protects the plant from infection. Thus far, none of the inoculated vines that recieved two applications of benomyl have become infected with Eutypa. The biological control agents are spore suspensions of the fungi Fusarium, Clado.sporium, Aureobasidium, Trichoderma, and PeniciIlium. Our studies have shown that these fungi can successfully colonize the wound surface when we apply them as spore suspensions. These fungi also are effective in reducing infection by Eutypa, but those studies must be conducted over many years to observe results over a variety of weather conditions Studies on the natural wound colonization showed that numbers of filamentous fungi, yeasts, and bacteria are initially zero, and increase rapidly for 7-14 days after pruning, then remain constant.

The Effect of Grapevine Latent Viruses on Non-AXR Rootstocks

AVF Proj. ID: 9329 / /Cat.: ,
Primary Investigator(s): Deborah Golinoby

1994 Final Report – Severe latent virus symptoms have been observed in new vineyards planted to non-AXR rootstocks throughout the state. The symptoms may include severe stunting, internode shortening, leaf discoloration, leaf rolling, stem pitting, disorders of the graft union and death. It is suspected that growers have empirically selected scion materials that have included virus strains which cause only mild symptoms on AXR-1. When these scions are grafted to certified rootstocks, the latent virus symptoms appear. Furthermore, Central Valley field selections which have been propagated on their own roots also seem to be affected. Twenty-six samples were collected from affected vineyards throughout the state during 1992-1993, propagated and planted in the Davis Grapevine Virus Collection, and ELISA tested for eight viruses including grapevine corky bark-associated virus (GCBaV), grapevine leafroll-associated viruses (GLRaV) types I,II, III, and IV, grapevine virus A (GVA), grapevine fanleaf virus (GFLV) and tomato ringspot virus (TmRSV). GCBaV isolate 100 was detected in an unusually large number of samples; 13 of 2 6 tested strongly positive. GLRaV II was detected in 4 of 26 samples. Two samples were positive for GLRaV III, which is considered to be the most common leaf roll type in California. Two selections tested positive for GVA; one selection tested positive for GFLV; and one selection tested positive for GLRaV I. Seven samples tested positive for more than one virus. No samples tested positive for GLRaV IV or TmRSV. Woody indexing and in vitro grafting is in progress. Five additional samples were collected in winter 1993-1994 and are being propagated.

Optimal Management of Lepidopteran Pests of Grapes

AVF Proj. ID: 9325 / /Cat.: ,
Primary Investigator(s): Mark A. Mayse, Simon Gravesby

The primary objectives of this research project are: 1) To evaluate and refine monitoring programs for western grapeleaf skeletonizer (WGLS) and omnivorous leafroller (OLR) in grapes, and 2) To determine spatial and temporal patterns of WGLS and OLR distribution in order to develop a non-preventive, true IPM program for dealing with these two serious vineyard pests. Several significant results have already been produced during the two years of this project (1992, 1993). Sex pheromone bucket traps with insecticide strips caught more WGLS moths at peak flight than did traps with ethylene glycol. OLR bucket traps with insecticide strips were somewhat more efficient than sticky bottom pheromone traps. With respect to monitoring OLR larvae, the UC-recommended bunch count technique (developed mostly on Thompson seedless), appeared to have questionable applicability for detecting OLR in tight-clustered grape cultivars (e.g., Barbera). Thus, early season OLR infestations, readily evident before berry sizing in such cultivars, later became “invisible” as the clusters developed. Generally low population levels of both WGLS and OLR during 1993 made collection of sufficient numbers in field trials a major challenge. A study comparing releases of Trichogramma parasiotoids and applications of Bacillus thuringiensis microbial insecticide against OLR which involved careful examination of 600 clusters at harvest produced only 4 OLR larvae and 5 OLR pupae. However, these same counts also revealed from 1 to 6 clubionid spiders (prob. Chiracanthium inclusum) in each grape cluster, suggesting that these predators may have been a factor in the low OLR numbers. For all the commercial vineyard blocks which were sampled in 1992 and 1993, the growers felt that chemical control was unnecessary, based on the pest monitoring data which we compiled. Thus, we have clearly demonstrated that preventive treatment for these lepidopteran pests of grapes is by no means an essential activity in San Joaquin Valley vineyards.

Meristem tip culturing for the elimination of grapevine viruses

AVF Proj. ID: 9324 / /Cat.: ,
Primary Investigator(s): Deborah Golinoby

The most important technical problem which limits the importation of winegrape clones and rootstocks is the difficulty with which grapevine viruses are eliminated using the old heat therapy techniques. These techniques are slow and inefficient. A number of laboratories worldwide have reported that meristem tip culturing is an effective technique for eliminating grapevine viruses. We have developed those those techniques in our laboratories and are evaluated their effectiveness. This technology should result in streamlining of procedures for the elimination of grapevine viruses from valuable grape propagating materials. As a result of these streamlined procedures, the length of time required for entry of infected materials should be substantially reduced. In turn, this should provide greater accessibility to foreign materials which include European wine clones, improved rootstocks which are needed for many reasons including their potential phylloxera resistance and other products of worldwide grapevine breeding programs. Furthermore, these techniques should further the efficient therapy for virus elimination of important California field selections suspected or proven to be infected with grapevine latent viruses. This will be the final report on this project. Nearly all of the objectives of this project have been accomplished. In the cases where the work is complete or nearly complete, I have attached the scientific papers which document the research. My laboratory will be completing the final experiments in the next few months to fill in the last of the data needed for publication. Treated explants will be monitored to insure that virus does not reappear. Additional future work will focus on improving survival rates and efficiency. The technique of shoot tip culture seems to be successful. Foundation Plant Materials Service is now using this technique to perform therapy on both diseased vines from quarantine and on valuable California field selections which are not available as certified stock.

Involvement of Viroids in Grapevine Diseases and the Impact on Vine Performance

AVF Proj. ID: 9323 / /Cat.: ,
Primary Investigator(s): J.S. Semancik, J.A. Wolpertby

The role of viroids as agents of plant disease is well established. Control experiments have been initiated to evaluate the role of grapevine viroids as the causal agent of the Yellow Speckle (YS) disease and in association with grapevine fanleaf virus in the expression of Vein Banding (VB) symptoms. YS symptoms which are difficult to evaluate under climatic conditions in California were induced under growth chamber conditions with California GYSVd-1 isolates from Mission and Cabernet franc but not from Zinfandel. Symptom expression of YS is affected by vine physiology and development. These observations confirm the presence of the Yellow Speckle disease agent in California vines. However, the disease expression as well as the viroid-disease relationship appears to be more complex than initially presumed. The first experimental demonstration of synergism between a viroid and a virus has been accomplished with the induction of VB symptoms. Both grapevine viroids and grapevine fanleaf virus (GFLV) were required. Thus, VB can no longer be considered as simply a late season expression of fanleaf and the importance of viroids to vine performance has been demonstrated. Performance trials at Oakville of the first available viroid-free vines in the world have been concluded due to the Phyloxera pressure on the own-rooted Cabernet Sauvignon. Data on vine growth, yield, fruit maturity and wine quality suggest a difference in vine performance especially in the growth parameters. These differences point to a potential for viroids to affect and reduce growth. The first viroid-free rootstocks can now be utilized to expanded these studies. The continued development of additional viroid-free commercial varietals, rootstocks, as well as rootstock germplasm sources is essential to our investigation of the impact of viroids on grapevines. Propagation and maintenance of viroid-free materials in a Foundation Vineyard at UC-Davis will provide greater availability to FPMS for general distribution as well as for indexing purposes in the absence of a viroid background. Monitoring for the incidence of transmission to viroid-free vines interplanted in a commercial planting of viroid-containing vines indicated that no field spread had occurred after six growing seasons.

Development of Methodologies for Rapid Detection of Grapevine Viruses

AVF Proj. ID: 9309 / /Cat.: ,
Primary Investigator(s): Adib Rowhaniby

1994 Final Report – A low tittered antiserum to grapevine leafroll associated virus (GLRaV) type I (TI) has been produced. To produce this antiserum, a partially purified virus was used. The antibody produced from this antiserum worked well when used to coat the plate and trap the virus in ELISA assays. Antiserum to a new type of GLRaV has also been produced. The virus has been isolated from a vine with leafroll disease. The produced antiserum has a high titer and did not react with any of the known types of GLRaV (types I, II, III and IV) or GVA. We are in the process of characterizing and identifying this virus type. To overcome the shortage of polyclonal antibodies for GLRaV Til, III and IV, virus from each type has been isolated and purified. Rabbits have been immunized by Til and TIV and we are in the process of preparing and evaluating antisera to these viruses. A polymerase chain reaction (PCR) technique which is a very sensitive and reliable method for the detection of plant viruses has been developed for the detection of grapevine fanleaf virus (GFLV) in grapevine tissue. This technique was able to detect GFLV in a sample when one infected grapevine leaf was mixed with 200 healthy ones. We have modified and simplified the PCR methodology by combining immunology and PCR (immunocapture-PCR, IC-PCR) for the detection of GFLV and GLRaV Till. In this technique the lengthy process of nucleic acid extraction required for PCR has been eliminated. The results showed that this modification works quite well for the detection of these two viruses.

The Effect of Grapevine Latent Viruses on Alternative Rootstocks

AVF Proj. ID: 9226 / /Cat.: ,
Primary Investigator(s): Deborah Golino, Adib Rowhaniby

1993 Final Report – An epidemic of latent virus disease is suspected in new vineyards planted to alternative rootstocks throughout the state. The working hypothesis which explains this observation is that growers have been propagating scion materials selected empirically for virus strains which cause mild symptoms on AXR-1. This hypothesis is supported by widespread observations of severe latent virus symptoms in Northcoast vineyards where field scion selections, which appear healthy on AXR-1, have been grafted to certified alternative rootstocks. Furthermore, Central Valley field selections which have been propagated on their own roots also seem to be affected. Symptoms include severe stunting, internode shortening,leaf discoloration, leaf rolling, abnormal dormancy, shoot tip necrosis, severe pitting and grooving at the graft union, dead tissue at the graft union, discoloration of the rootstock wood, and death 2 to 3 years after planting. The hypothesis that latent viruses are causing these vineyard failures needs to be tested immediately. We propose to do this by field budding latent virus infected scions to a panel of alternative rootstocks and observing the effects. The effect on bench grafts will also be studied. In addition, survey work needs to be conducted as soon as possible to obtain information on the impact of this problem in the nursery and in the vineyard. If this epidemic is severe, faster virus detection techniques will be needed to screen field scion selections. In addition, many California clonal materials which are not certified may need to be treated for virus infection. Last year, 2 5 sites where vineyard replant failure may have been caused by the presence of latent viruses were selected for further work. Wood was collected from vines in those sites and propagated. Observations thus far show that the samples have poor vigor and show many characteristic virus symptoms. ELISA testing and green-grafting, using a mini-chip graft developed by Golino, is in progress. Results are pending. A history of affected vineyards was started and shows that the most severe problems are on 3 3 09 and Freedom. However, this may be coincidental and is still a preliminary finding.

Spiders in Vineyard Agro-Ecosystems

AVF Proj. ID: 9223 / /Cat.: , ,
Primary Investigator(s): Mark Mayseby

The overall goal of this research project is to continue to elucidate the ecological roles, along with the potential economic value, of spiders in vineyard agro-ecosystems. Key objectives include determining which spider species may be directly associated with vineyard cover crops, and further delineating the patterns of abundance and distribution of important spider species in vineyards. One clear pattern beginning to emerge is an inverse relationship between spider and leafhopper densities in vineyards. Our findings in this and other studies agree with several other vineyard spider researchers in demonstrating that when spiders are abundant, leafhoppers generally tend to stay below economically damaging levels. The two most abundant spiders sampled from mid-June to the end of November in 1992 belonged to the family Clubionidae (two-clawed hunting spiders): Trachelas pacificus and Chiracanthium inclusum. Juveniles for these two species were roughly three times as abundant as adults in samples taken throughout the season. The next most abundant spider was Theridion (family Theridiidae), which was also of particular interest by being detected only in the grapevine canopy (i.e., almost never from the cover crops between vine rows). It should be noted that the most abundant spiders commonly detected in both cover crops and canopy were Trachelas and the micryphantids. Another particularly noteworthy discovery during the 1992 season involves an apparent correlation between western grapeleaf skeletonizer (WGLS) mortality and Trachelas pacificus occurrence. Corrugated cardboard bands wrapped around the base of vines are very effective in concentrating WGLS larvae seeking pupation sites. Bands which individually contained up to 70 WGLS pupae typically were found to be free of spiders. However, it was also not uncommon to find bands with only 5 to 10 WGLS pupae; in virtualy all these cases a large number of Trachelas juveniles were also found residing in those bands. Additional data collected in “round-the-clock” sampling trials during 1992 indicated that in estimating population densities for several important spider species, the actual time of day when samples are taken can be of considerable importance.

Production of Antisera to Grapevine Viruses

AVF Proj. ID: 9221 / /Cat.: ,
Primary Investigator(s): Adib Rowhaniby

A good supply of antisera to two isolates (type II and type III) of the long clostroviruses associated with grapevine leafroll (GLRaV) has been produced as well as a low tittered antiserum to the type IV GLRaV. These antisera have been evaluated and optimized for ELISA test. Monoclonal antibodies are produced to type III of GLRaV and the titer of the antibody is guite high when evaluated in ELISA tests. A high-tittered antiserum to grapevine fanleaf virus (GFLV) has been produced. A low-tittered antiserum to tomato ringspot virus (TmRSV, causal agent of grapevine yellow vein disease) has been produced and works quite well in ELISA tests. The antisera conditions have been optimized for GFLV and GYW detection in ELISA tests and they are routinely being used to test the foundation stocks at FPMS. The same GFLV antiserum is also being used by the California Department of Food and Agriculture (CDFA), Pest Exclusion branch for testing the registered grapevine material in the registration and certification program. An antiserum to grapevine corky bark virus (GCBaV) has also been produced and conditions for optimizing the reactivity of the antiserum in ELISA tests were done. Some monoclonal antibody lines were produced for GCBaV but subsequently lost their ability to produce specific antibodies. Our attempt has failed to purify enough rupestris stempitting associated virus for the production of antiserum. We have attempted to make a cDNA clone from the dsRNA and use it in a cDNA hybridization system as a diagnostic tool, but it failed. The attempt will be continued. A source for GLRaV-type I was identified. Amounts of virus was purified and used to immunize a rabbit, but the titer of produced antiserum was very low and was not useable in ELISA. A new source of GLRaV type I has been found. Quantities of virus have been purified. A rabbit has been immunized with the purified virus. The preliminary tests indicated that the specific antibody titer is building up in the animal. The rabbit will be given boost injections with the same virus to develop a high tittered antibody for GCBaV T I.