Control of Eutypa Dieback

Objectives: The overall objective of this research is to develop more effective methods for the management of Eutypa dieback. There are several specific aspects to this broad objective, including: 1. Evaluation of various fungicides for pruning wound protection. 2. Determination of the duration of susceptibility of wounds and the mechanism by which they become less susceptible with time. 3. Evaluation of natural pruning wound colonizers as wound protectants (biological control). 4. Investigation of the relationship between vineyard age and disease incidence. Samjuaxy.: In four field experiments, only two fungicides, Benlate and Nustar, were consistently very effective. In general, the fungicides were more effective when wounds were inoculated one day after fungicide treatment, compared with inoculation after two weeks. Wound colonizing microorganisms were screened in the laboratory and the field for their potential to inhibit Eutypa. The ten isolates tested in the field were less effective in 1991 than in 1990. However, the biological control agents were generally more effective when inoculation was delayed. Two of the biological control agents, Fusarjum lateritium and Cladosporium herbarum. were as effective as benomyl when inoculated with Eutypa two weeks after treatment. Wound susceptibility experiments from 1991 indicated that wounds made in December were susceptible much longer than one month. Wounds made later in the dormant season were susceptible for a shorter duration, depending primarily on temperature. In both 1990 and 1991, accumulation of degree-days after pruning was strongly related to xylem wound response and populations of epiphytic microorganisms on the wounds. These factors led to decreased susceptibility. Degree-day accumulation may be used to predict wound susceptibility. These data support the practice of pruning when warm weather is likely to follow (late pruning). Different grape cultivars appear to have different levels of susceptibility to Eutypa in the field. We have used ascospore inoculations and mycelial inoculations, but have not detected any strong differences in susceptibility among cultivars. In order to characterize the relationship between vineyard age and incidence of disease, we surveyed, in 1991, 12 Chenin blanc vineyards ranging in age from 8 to 24 years. Results were very similar to those obtained for French Colombard in 1990. Vines less than 10 years of age had little disease. Disease incidence increased rapidly between 10 and 20 years, and stabilized after 20 years of age, with nearly all vines, but less than half of the spurs diseased.

Cultural Control of Phomopsis Cane and Leaf Spot

The overall goal of this project is to evaluate the potential for control of Phomopsis cane and leaf spot caused by Phomopsis viticola using cultural methods to reduce initial inoculum. Phomopsis is cane and leaf spot is a regular problem in grape production in certain areas of California and can be a widespread problem when disease is accentuated by spring rains. A standard control measure in many areas was a dormant spray of dinoseb, however registration was lost last year. The only other dormant spray available is sodium arsenite, an extremely toxic compound that many wineries and packing sheds will not allow the growers to use. The first and second objectives were to determine the relationship of the intensity of overwintering infections and disease severity and the potential of reducing disease by reducing the number of pycnidia through careful pruning. The source of primary inoculum which initiates the epidemic is from mature pycnidia which form on second year wood. Two vineyards in the Russian River Valley and two in the Lodi area where Phomopsis is a consistent and severe problem were surveyed for the incidence of infected second year wood both before and after pruning. Three treatments were established in one vineyard. First, all of the diseased spurs were left, next 1/2 of the disease spurred were removed, and third, all of the diseased spurs were removed. This was replicated 4 times in a 5 vine by 5 vine blocks to minimize interplot interference. At two weeks prebloom, bloom, and two weeks post bloom, the number of leaves with lesions and the number of lesions per leaf were examined. No disease was observed in any of the plots. Although we could readily isolate Phomopsis from diseased spurs in the laboratory, we concluded that the drought had reduced the inoculum to such low levels that disease was no longer present. These experiments were repeated the next year with the same results. Therefor, as mentioned by phone and letter, we are not continuing the request for funding at this time, but will continue to monitor for the disease.

Epidemiology and Control of Grapevine Powdery Mildew in California

Powdery mildew can overwinter in cleistothecia lodged in bark. Cleistothecia become lodged in the bark when they wash off of leaves during fall rains. If cleistothecia fall to the soil instead of the bark, they do not usually remain viable. In the spring, cleistothecia in the bark release ascospores during mild prolonged wetness periods. These same temperatures (15 to 25 C) favor germination and infection of grape leaves. Cleistothecia do not remain viable for prolonged periods of 30 C. Canopies which do not have the majority of the leaves growing over the bark are less likely to have as many cleistothecia lodged in the bark. Most infections occur within 10 cm of the bark. Cleistothecia were less viable if they originated from the Central Valley or Southern California than from Coastal regions. This may be why most early infections in the southern areas are the result of infected dormant buds. However, prolonged temperatures over 30 C kill the fungus at any point in its life cycle. Cleistothecia can form as early as July and produce viable ascospores as early as August. Therefore, diligent disease control is important, including post harvest. Spring applications of wettable sulfur (5 lb / 100 gal / acre) were most effective in controlling early season mildew. Spraying should start when shoots are at the 0 to 2 inch stage. Applications should be repeated at ten day intervals when rains, soaking fogs and dews keep grape leaves wet for 12 hours or more.

Genetics of Fungicide Resistance to Bayleton, Rally, and Rubigan

Objective: To determine the genetics of fungicide resistance in U^. necator to the triazole and pyrimidine DMI fungicides currently registered on grapes in California. Twenty isolates were collected from Lone Oak in Monterey County and twenty isolates were collected from the Renaissance Vineyards in Lassen County in 1990. Some Lone Oak isolates have previously exhibited DMI resistance to a high degree. Renaissance isolates have never been exposed to DMI’s and do not demonstrate resistance. Thirty single conidia were transferred to separate isolated plants from the most resistant Lone Oak isolate and thirty single conidia from the most sensitive Renaissance isolate. The resulting colonies were allowed to proliferate on young Carignane plants to test for sexual mating type. Fifteen isolates from each vineyard (Renaissance and Lone Oak) were tested for sexual mating type by rubbing conidia from a colony of New York type ‘F’ or New York type ‘G’ directly onto a leaf infected with the colony to be tested and adjacent to the California test colony. The plants containing both colonies were grown in isolation tubes for 30 to 60 days and then observed by microscope for cleistothecia formation. If an isolate produced cleistothecia when crossed with New York ‘F’ it was determined to be type ‘G’ and if it produced cleistothecia when crossed with New York ‘G’ it was determined to be type ‘F’. At this time, two California type ‘G’ sensitive isolates, one California ‘F’ sensitive isolate, and one California ‘F’ resistant isolate has been identified. However, viability of the ascospores has not yet been verified and is currently being tested. Because of the difficulty in obtaining successful crosses this report is preliminary. During the next year, we will screen the crosses and their progeny for DMI sensitivity. The results of these screenings will be reported as they occur.

Grapevine fanleaf virus: detection and distribution

The first three objectives of this study are dependent upon the fourth. That objective has been initiated with the establishment of the plant material in containers and the partial completion of the graft inoculations. We saved time establishing the plant materials by using rooted dormant cuttings supplied by the Department of Viticulture and Enologys field crew. These plants were potted into 1 gal containers and established. We also have ample numbers of additional established plants for re-grafting and to use as uninoculated controls. The majority of the rootstocks have been graft inoculated (see the table below) and we will soon begin another round of chip-budding. We will also use ELISA to assess the GFLV levels in the sample tissues both above and below the site of graft inoculation. The Muscadinia rotundifoliaand the Vitis berlandieri are now being propagated under mist conditions and may be graftable by Fall 1991. We sampled for fanleaf and tomato ringspot virus during the summer and fall of 1990 and found fanleaf widely scattered, but also found tomato ringspot in Napa, Sonoma and San Joaquin counties. We have resampled San Joaquin county this spring and with these results began to conclude this initial survey (the rough draft of a article for submission to California Agriculture article is included in Appendix 1). The incidence of TomRSV was higher than we expected, but does not pose a direct threat to the industry. TomRSV causes grapevine yellow vein disease, a disease which causes substantial yield reductions (on the order of fanleaf degeneration) on the east coast of the US. We suspect that this disease mimics fanleaf in California, but does not cause severe yield reductions. It is important for researchers and California Department of Food and Agriculture inspectors to recognize TomRSVs incidence and symptom expression to avoid confusion with fanleaf degeneration. Dr. Walker and Rowhani have completed a related research project determining which sample tissue produces the highest and most reliable ELISA reading over the course of the growing and dormant season (Appendix 2). Shoot tips and young leaves are the best sample tissue when growth is active, cambial scrapings of young phloem, cambium and young xylem are best after growth stops and before dormancy. During the dormant season actively growing tissues (shoots, callus, roots) forced from canes gave the highest values. We will use these results to best quantify the level of GFLV in the infected rootstocks. Elizabeth Frantz, a graduate student of Dr. Walker’s, is researching sampling strategies for GFLV detection in the vineyard. She completed sampling three 1225 vine plots with varying levels of GFLV incidence (low, medium and high) and is now testing each vine for GFLV with ELISA. We can then take the data and apply sampling strategies to it to determine how to best sample GFLV-infected vineyards.

Grapevine Viroids as Possible Factors in Disease, Clonal Variation and Wine Quality

The general overview of this project is to provide an understanding of the importance of the grapevine viroids to vine growth, productivity, and wine quality. In addition, viroids may provide a novel yet practical approach to “customize” vine growth and development to achieve such goals as the reduction in excessive vegetative vigor. VIROIDS were first identified in the 1970’s as causal agents of plant disease. It has been recognized that in some cases these sub-viral RNA molecules can be readily transmitted into receptive plant species without producing any apparent host plant reaction viewed as an expression of plant disease . Thus, the viroid-RNA can be considered as a transmissible yet non-infectious entity. Because of this condition, it is possible that the biological activity of a viroid may be expressed by altering a normal growth response of a plant. With our observation of a virtual ubiquitous occurrence of 1-3 viroids in all vines in California, the role of the viroids in grapevine tissue has become of interest. This unusual property of the widespread association of viroids with grapevines indicates that eyery vine characteristic including viticultural and enological property is viewed through a viroid background. Many of the accomplishments that have been achieved in this project have been published (enclosure copy of Szychowski et al., Vitis 30, 25-36, 1991) or presented in Sept. 1990 at the 10th Meeting b7 the ICVG (International Council for the Study of Viruses and Virus Diseases of the Grapevine) and at the Annual Meeting of the ASEV in June, 1990. These reports provide information concerning: 1) The first field trial of viroid-free grapevines now in the fourth growing season at the Oakville Experiment Station. 2) The commonality among the grapevine viroids from California and Europe. 3) The production and propagation of viroid-free varietals and rootstocks. 4) The comparative properties of viroids from grapevines which relate to the incidence of yellow speckle and vein banding diseases.

Meristem tip culturing for the elimination grapevine viruses

Young grapevines have been successfully propagated from virus infected grapes. Ten different virus isolates have been included in- these experiments. We have found that it is easy to culture plants from explants which are 2-3 mm in size or larger. Although we have cultured plants from explants which are l mm or smaller, the survival rate is low. We hope to improve that survival rate by further refinement of technique. Early data using the ELISA test on the plants which are out of tissue culture and in soil indicate that a significant number of explants have been freed of virus. This is very encouraging data which suggest that the meristem tip culture is successful for virus elimination. However, retesting over the next few years as the vines mature is needed to be sure that the virus is completely eliminated from the plants, not just in low concentrations. A large number of explants have been sucsessfully cultured and transferrsd to soil in the 1991-1992 funding year but due to staff and funding shortages they have not been ELISA tested. Grapevine virus collections essential to further studies of virus elimination techniques have been established and are thriving. Experiments are underway to determine the optimum explant size for grapes infected with leafroll, fanleaf, Rupestris stem pitting and corky bark. The progress which has been made in developing antisera for leafroll and corky bark should facilitate this work. In addition, efforts are being made to use antiviral chemotherapeutics as a step in the tissue culture process. Wood from four leafroll-infected cabernet sauvignon selections from Napa Valley has been collected and the selections are undergoing therapy; these materials will be used for evaluations of the effectiveness of the therapy and for subsequent evaluation of the effects of the virus diseases on clonal characteristics.

Minimizing Fertilizer Inputs for Variegated Leafhopper Management in Grapes

This research investigated the relationship between host plant (i.e,, grapevine) nitrogen status and population dynamics of the variegated leafhopper (VLH) in a commercia1 vineyard for three years. During each year, replicated fieli plots received the following treatments: 1) control (no fertilizer added), .’) 75 lbs. N / acre (synthetic ammonium nitrate), 3) 150 lbs. N / acre (synthetic ammonium nitrate), and 4) two tons of compost / acre ( = 38 lbs. N / acre). The response of VLH to these treatments was investigated by monitoring nymph densities and conducting oviposition (egg-laying) and nymphal development rate tests. During all three years, VLH nymph populations reached a seasonal peak during the first generation. Nymph densities during the second and third generations were especially low in 1989 and 1990, and reached a moderate level during the second generation of 1991. Oviposition tests conducted in July 1989 and June 1990 demonstrated greater VLH egg-laying in the synthetic fertilizer plots. However, oviposistion during the July 1991 test was greatest in the compost plots. No significant differences in VLH nymphal development rates were observed among treatments.

Production of Antisera to Grapevine Viruses

A good supply of antisera to two isolates (type II and type III) of the long clostroviruses associated with grapevine leafroll (GLRV) has been produced as well as a low tittered antiserum to the type IV GLRV. Monoclonal antibodies are produced to type III of GLRV and the titer of the antibody is quite high when evaluated in ELISA tests. A clostrovirus was mechanically transmitted from leafroll (type III) infected pinot noir to Nicotiana occidentalis. Investigations revealed that this virus is not associated with grapevine leafroll disease. A high-tittered antiserum to grapevine fanleaf virus (GFLV) has been produced and quantities have been increased. A low-tittered antiserum to tomato ringspot virus (TmRV, causal agent of grapevine yellow vein disease) has been produced and efforts are underway to produce a high-tittered antiserum to this virus. 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 also being used by California Department of Food and Agriculture (CDFA), Pest Exclusion branch for testing the registered grapevine material in their registration and certification program. An antiserum to grapevine corky bark virus (GCBV) has also been produced and conditions for optimizing the reactivity of the antiserum in ELISA tests were done. Another polyclonal antiserum to GCBV has been produced. The reason for producing another antiserum for GCBV was to get a cleaner antiserum (with lower healthy background in ELISA test) by doing more rigorous purification of the virus particles. Evaluation of this antiserum is underway. Some monoclonal antibody lines were produced for GCBV 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. Double stranded RNA has been consistently extracted from a grapevine infected with rupestris stempitting and we are planning to make a cDNA clone from the dsRNA and use it in a cDNA hybridization system as a diagnostic tool. A source for GLRV-type I has been identified. Amounts of virus has been purified and within next few weeks we will start immunizing a rabbit to produce specific polyclonal antibody to this virus.

Spiders as Beneficials in Grape Agro-ecosystems

Spiders represent an entirely predacious group of organisms whose potential importance as beneficials in agro-ecosystems has generally been overlooked. The goal of this research was to investigate the potential importance of spiders as a mortality factor for grape insect pests. Spider populations were sampled at frequent intervals in a number of Fresno County vineyards. Except for one site, all vineyards utilized “soft” pesticide alternatives. Furthermore, the sites represented a range of environments including vineyards near and distant from riparian habitats. Spiders were abundant in vineyards managed with the use of “soft” pesticides (e.g., B.t. & soaps) in the Fresno County area. Three species in the families Agelenidae and Clubionidae were particularly common. Species in the family Salticidae were less abundant, although consistently present. Spider populations “built” in the canopy and persisted through the growing season with little fluctuation over short time intervals. The relative abundance of spiders in vineyard canopies suggests that they are of considerable potential importance as biological control agents in vineyards, having been grossly overlooked. It is suggested that the negative impact of pesticide use on spider populations may be extensive. Our findings indicate the need for additional research directed at defining the impact that spiders have on specific insect pests of vineyards. In addition, studies are needed to investigate the effects of ground cover management, pesticide use, and other cultural factors on spider population dynamics.