Optimizing Tissue Culture Techniques for Virus Elimination in Grapevines

Microshoot tip tissue culture is the method of choice to eliminate virus infection from valuable grapevine clones. However, survival rates are often low or highly variable. The objectives of this research were 1) to optimize tissue culture protocols to increase shoot tip survival and growth rate, increasing efficiency of tissue culture treatment; and 2) to investigate the use of small single node cuttings for virus elimination. A new protocol was found which was a clear improvement over those used in the past, greatly increasing survival and efficiency of tissue culture treatment for virus elimination in grapevines. Four media were used in various sequence for a total of five different protocols. The effect of virus status on survival was studied utilizing the new protocol. We found no significant difference between healthy or virus-infected source materials in percent survival or percent root development in culture, although incidence of disease symptoms was significantly higher in plants cut from virus-infected source plants. The virus elimination rate in this study was 97%. For objective #2, we have found a significant percentage of “escapes” among relatively large explants cultured from virus infected plants. This suggests that as virus detection techniques become more sensitive it may be possible to regenerate clean stock far more efficiently using relatively large explants. We have found that plants continue to test negative for viruses by ELISA. This is further indication that the virus has been successfully eliminated. A woody index test is in progress to confirm this.

A Study of Anagrus Egg Parasitoids Important for Biological Control of Erythroneura Leafhopper Pests of Wine Grapes in California

In many of California wine grape vineyards, especially in San Joaquin and Sacramento Valleys, the variegated leafhopper (VLH) has become the dominant pest since it was first reported there in 1980, replacing the western grape leafhopper (GLH) in importance. While the two species of mymarid wasps of the genus Anagrus, previously known as A. epos, provided more or less satisfactory control of GLH before 1980, parasitism of VLH is lower and does not currently provide sufficient control. Cornerstones of biological control are the proper identification of natural enemy species as well as evaluation of their effectiveness in the field following release. From data we have collected, we believe that several Anagrus species that were imported to California and released in selected vineyards in San Joaquin Valley during 1986-1991 either have not established or perhaps interbred with the local populations. One of the species native to California, A. erythroneurae, is the most abundant natural enemy of both leafhopper species throughout California, whereas lesser numbers (ca 10-20%of all samples) of a yet undescribed species (A. sp. 1) have been recognized from collections made in the Napa, Sonoma, and San Joaquin Valleys. Overall, more than 5,000 specimens of Anagrus were examined and identified during our study including those reared from potential overwintering refuges, i.e., almond, apple, blackberry, prune and wild grape plants. The conclusions resulting from this two-year study and our recommendations for further research and biological control programs are presented.

A Study of Anagrus Egg Parasitoids Important for Biological Control

During year one of research, collections of Anagrus egg parasitoids of western grape and variegated leafhoppers were made on winegrapes in Napa Valley, Sonoma Valley, Alexander Valley, Russian River Valley (Guerneville-Trenton-Healdsburg-Asti areas), El Dorado Co., Mendocino Co., San Luis Obispo Co. (Paso Robles), Merced Co., and during September-November 1995 in Riverside County (Temecula area). Additional material, at no cost to this project, was available for study from vineyards in Sacramento Valley, Santa Barbara, and San Luis Obispo Counties (California) as well as from Arizona, Colorado, New Mexico, New York (USA), and Mexico. Four or five different species of Anagrus wasps which are associated with cultivated grapes in North America have been recognized. Two of these species occur on winegrapes in California as follows: Anagrus erythroneurae S. Trjapitzin & Chiappini and Anagrus new species (A. Sp.), both parasitic on western grape and variegated leafhoppers eggs. In most locations in California winegrape-growing areas where A. sp. occurs, A. erythroneurae is also present. These localities are in Sonoma Valley and Alexander Valleys (Sonoma Co.), Oakville (Napa Co.), Mondavi and El Rio Vineyards at Lodi (San Joaquin Co.), and Davis (Yolo Co.). In these areas A. erythroneurae is a dominant species, comprising more than 50-70%of all collected parasitoids per sample. In most of the other winegrape-growing areas of California, such as El Dorado Co., Santa Barbara Co., and Temecula wine country (Riverside Co.), we found only one species, A. erythroneurae. The same species dominates in the San Joaquin Valley (California, USA), Mexicali Valley and other areas of Baja California, Mexico but is rare in collections from Colorado, New Mexico and New York (USA). In September 1995, S. Triapitsyn made collections in Centralia, Illinois, which is the type locality of Anagrus epos Girault, a species which was previously mistakenly considered to be a major biological control agent of grape-infesting leafhoppers in North America. Two specimens similar to A. epos were collected in Centralia and compared with the type specimens of this species. As a result, it was determined that A. epos is not known to occur in California and subsequently plays no role in the natural control of leafhoppers on grapes in this state.

Spiders in Vineyard Agro-Ecosystems

The overall goal of this research project is to continue to elucidate the ecological roles and potential economic value of spiders in vineyard agro-ecosytems. Specific objectives include: 1) To determine key behavioral habits and to study prey selection of both Cheiracanthium and Trachelas spiders, 2) To further investigate the diversity and abundance of spiders in vineyards, and 3) To develop a color field guide to important spiders in California vineyards. During 1995, spider research was conducted both in vineyards near Ripperdan (Grenache) and at CSU Fresno (Barbera, conventional and organic plots). Direct observation sampling in the grapevine canopy was conducted during evening hours (peak hunting activity periods for Cheiracanthium and Trachelas) using headband-mounted lights with red filters to avoid disturbing normal spider behavior. Another spider sampling technique used in 1995 involved placing corrugated cardboard bands around the base of vine trunks (particularly effective for Trachelas). Near the band locations, canopy shake samples were also conducted. Direct observation sampling involved a total of 26 hours viewing time in 1995. Only 8.3% of the total sampling time resulted in successful sighting of a Cheiracanthium or Trachelas spider. Frequency of occurrence data for the two spider genera mirrored results from 1994, in that 92% spider sightings were Cheiracanthium with the remaining 8% being Trachelas. Also similar to 1994 patterns, juvenile spiders were observed more frequently than were adults. Behavioral activities of Cheiracanthium spiders were rather diverse: adults hunting (25%), adults resting (25%), adults with egg sac/molting sac (8.3%), and juveniles hunting (54.2%). No Cheiracanthium were directly observed in the act of feeding on prey in 1995. Trachelas spiders were found in only two behavioral categories: juveniles resting (50%), and juveniles hunting (50%). Band trapping results at CSUF revealed the highest numbers of Trachelas adults in conventional plots in late January 1995, while Trachelas juveniles peaked in organic plots on the same date. Both adult and juvenile Cheiracanthium reached highest densities in late January in the organic plots. Band traps at the Ripperdan Ranch produced peak Trachelas adults and juveniles in late February 1995. Cheiracanthium adults at Ripperdan peaked throughout February, while juveniles reached highest densities in late February. Joint CSUF/UC collaboration (co-authors Mark Mayse, Michael Costello, Billy O’Keefe, Kent Daane, Curtis Sisk) on a color-illustrated field guide entitled, “Spiders in San Joaquin Valley Grape Vineyards” came to fruition in late 1995 with the guide’s publication. In addition, we are completing a translation of the guide’s text and caption information into Spanish to further expand the circle of effective contact for this publication; the translation will ultimately be included as a simple insert included in the basic color field guide.

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.

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.