Jocelyn G. Millar, J. Steven McElfresh and Richard E. Rice1
Department of Entomology, University of California, Riverside, CA 92521, USA
1Department of Entomology, University of California, Davis, CA 95616, USA
Key words - sex pheromone, mating disruption, controlled release, dispenser, Anarsia lineatella, Ectomyelois ceratoniae, Cydia pomonella
The first identifications of insect pheromones several decades ago generated tremendous interest and speculation in both the scientific and agricultural communities, due to the perceived potential of insect pheromones as "magic bullets" which could be used in insect control. Indeed, insect pheromones are now used worldwide as monitoring tools, but with few exceptions, the potential for insect control with pheromones has not been realized. However, a nucleus of researchers have persisted with the development of applications for pheromones over the years, and the resulting body of empirical and experimental knowledge has resulted in the elucidation of factors which are critical to the successful use of pheromones. For example, it is now generally recognized that pheromone-based mating disruption will work best in relatively flat, isolated blocks, with low initial populations of moths. Furthermore, a relatively uniform "blanket" of pheromone above an empirically-defined critical concentration is usually provided by large numbers of point source dispensers. These guidelines have been developed more often than not as a result of control failures, and we now have a greater appreciation of the complexities of successful application of mating disruption. This complexity can be summed up simply: with mating disruption, we are attempting to modify the behavior of several generations of a population of organisms for their entire lifespans. Unlike insecticide useage, where a single contact with the active agent removes the insect from the system, behavior modification requires that pheromone coverage be continuous and complete for periods of several months.
In the following pages, we describe some of the technological problems we have encountered in the development of applications of insect pheromones, both for monitoring and mating disruption, and the solutions which have been developed.
Pheromones for monitoring
Peach twig borer, Anarsia lineatella
Peach twig borer (PTB) is a major pest of stone fruit and nut crops worldwide. The pheromone for this insect was identified by Roelofs et al. (1975) as a blend of E5-10Ac and E5-10OH (ca. 4:1), and it has since been used widely as a monitoring tool. However, there were two anomolies in the initial report of the pheromone. First, the blends of synthetic pheromone used in the initial studies were minimally attractive, and it was not until an entirely new batch of pheromone was used that traps began catching moths. The cause of this problem was never identified (W. Roelofs, pers. comm.). Second, there appeared to be geographical differences in the response to pheromone, with a Washington population responding best to the pure alcohol, while California populations preferred blends of the acetate with the alcohol.
We reexamined the pheromone (Millar & Rice 1992), and found several minor components in both gland extracts and effluvia, none of which had any effect on the attractiveness of baits. As part of the same study, we synthesized and screened approximately 20 analogs of the pheromone components, and found that E6-10Ac and E7-10Ac strongly antagonized the pheromone. While these studies were in progress, we received reports of failures of commercial PTB lures from a California company. Careful analysis revealed that the faulty lures were contaminated with several percent of E6-10Ac, accounting for the poor lure performance. It should be stressed that up to this point, there had been no reason to suspect that this compound was inhibitory, and furthermore, it is not trivial to detect small amounts of E6-10:Ac in E5-10:Ac.
In 1993, lures from two commercial distributors again failed. Analysis followed by synthesis and screening of trace contaminants revealed a second antagonist, 5-decyn-1-yl acetate (Millar & Rice 1996). Again, this component was present in trace amounts, but it had very significant effects on lure performance. Finally, in 1996, a third failure of commercial lures occurred. Analysis determined that this pheromone, although of high chemical purity overall (>98%), was again contaminated with small amounts of E6-10Ac.
There are several points which can be drawn from this. First, if pheromone products are not reliable, their use will decline, and all the work invested in their development will be wasted. Second, synthetic pheromones used for monitoring lures, particularly for pheromones with histories of problems, should be field tested before marketing. Third, pheromone purity may be critical for efficacy, but the level of purity required will vary depending on the nature of the impurities and the insect species. Overall, researchers must be realistic with regard to the quality of pheromones which can be economically produced. However, simple precautions such as efficacy testing will be beneficial to both distributors (in terms of maintaining their markets) and to the continued development of pheromone technology.
Carob moth, Ectomyelois ceratoniae
Carob moth is the major pest of dates in California, and it is a pest of dates and almonds in other countries. The pheromone of the carob moth consists of an 8:1:1 blend of Z9,E11,13-14Ald, Z9,E11-14Ald, and Z9-14Ald (Baker et al. 1989). However, the major triene aldehyde readily degrades in light and air, making it difficult to work with, and is of questionable reliability for extended periods as a trap bait, particularly under the harsh conditions in California date gardens. The degradation problem has been addressed in two ways. First, a pheromone mimic (Z7,E9,11-dodecatrienyl formate) was developed, in which a formate ester replaced the aldehyde function. This compound proved to be as good or better as a trap bait than the synthetic pheromone blend (Todd et al. 1992).
Second, we conducted screening trials with a number of antioxidants and UV stabilizers (Figure 1) with interesting results. First, the best stabilizers of the pheromone mimic proved to be carbon black, Topanol CA® (a cocondensate of 3-methyl-6-t-butylphenol and crotonaldehyde; ICI Americas), and N,N'-diphenyl-1,4-phenylenediamine (data not shown). The latter compound had to be formulated in polyethylene glycol instead of the lauryl acetate diluent used with the other compounds because of solubility problems. Butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA, Sustane IF®), and alpha-tocopherol, all of which have been used extensively in pheromone formulations, gave intermediate or poor results. The results with carbon black were particularly surprising. This compound is thought to act by physically blocking incident radiation, but its excellent efficacy suggests that it may also have another mode of action.
Figure 1 Carob moth pheromone remaining in plastic vials with different stabilizers after aging in sunlight for 13 days (lauryl acetate carrier)
As a result of these studies, we have increased the effective field lifetimes of our carob moth baits (0.5 mg of stabilized pheromone mimic on grey rubber septa) to at least 3-4 weeks. Our baits are currently being used extensively by the California date industry, and we have also supplied researchers in Europe, the Middle East, and Australia with baits or compounds for field trials.
Our results illustrate several points. First, in some cases, pheromone mimics may represent satisfactory alternatives to the true pheromone, at least for monitoring purposes. However, they should be used with caution, because the effects of competition from females on trap catch is unknown. Second, even very fragile molecules can be stabilized by a judicious choice of stabilizers, extending the field lifetimes of formulations to satisfactory levels. This is an area which has remained largely unexplored by academic researchers.
Codling moth, Cydia pomonella
Codling moth is a major pest of apples, walnuts and pears worldwide, and despite several decades of effort, reliable and effective mating disruption of this pest has been elusive. The major component of the pheromone was identified as E8,E10-12OH (Roelofs et al. 1971), but a number of minor components have been identified from both gland extracts and effluvia (Einhorn et al. 1984; Arn et al. 1985). The effects of these components are still controversial, and commercial formulations used in mating disruption have been comprised of the major component alone or in combination with 12OH and 14OH.
One of the key factors which has slowed the development of effective mating disruption for codling moth is unsatisfactory dispenser performance: it has proven difficult to develop dispensers which release adequate quantities of high quality pheromone for periods of several months. In particular, in California, the moth has several generations per year, so that orchards need protection for periods of up to 5 to 6 months.
Despite the fact that codling moth mating disruption has been studied for many years, the release rates of dispensers have been poorly characterized and understood. The problem has been exacerbated by the rapid evolution of dispensers: dispensers have often been modified yearly, making it difficult if not impossible to compare results from year to year, and from one manufacturer to another. All too often, researchers have been left to speculate whether results were due to the pheromone dispensers used, or to environmental circumstances which affected moth populations. The problem was further compounded by the lack of standard protocols for conducting field trials.
Ideally a pheromone dispenser should have a constant release rate for its lifetime. In practice, release rates at a fixed temperature decrease with field aging. However, the decrease may be compensated to some extent by the increase in release rate with increasing temperatures as the season progresses.
In 1990, we began a 5 year study, measuring the release rates of field-aged dispensers under standardized conditions. These measurements did not allow us to determine actual field release rates, but they did allow us to accurately compare the changes in release characteristics of dispensers over time. A representative example of the data is shown in Figure 2.
Figure 2 Release rates of codlemone, E8,E10-12OH from field-aged mating disruption dispensers
Several key points emerged. First, in many cases, release rates dropped to low levels in 60 days or less, even though projected lifetimes of the dispensers were often 90 days or more. Second, considerable degradation of the pheromone occurred with some dispensers, as demonstrated by the buildup of sticky residues on dispenser surfaces, and by the change in the ratios of components released (e.g., for one dispenser type, the ratio of E8,E10-12OH to dodecanol changed from 1.08 initially to 0.66 after 3 months). It must be emphasized that these linked problems of degradation and rapid decline in release rates are not unique to codling moth pheromone.
However, our release rate measurements over a period of 5 years indicated very significant improvements in the performance of several dispensers. For the best dispensers, release rates after 12 weeks of field aging still approached 50% of the initial rates. However, other dispenser types, some of which were marketed, continued to show rapid declines in release rates with time.
In summary, information available to field researchers with regard to performance characteristics of the mating disruption dispensers that they are evaluating has often been sketchy at best. Information which has been available has often been obtained from artificially aged rather than field aged dispensers. Finally, many release rate measurements have been calculated indirectly, for example by subtracting the amount of pheromone remaining in a dispenser from the initial amount present. This fails to take into account degraded pheromone, which would not be detected or quantified by gas chromatography, resulting in actual release rates being lower than calculated. In short, adequate characterization of release rates of dispensers should become a part of the standard protocol of dispenser evaluation trials in order to introduce hard facts rather than "guesstimation" into the interpretation of field trial results.
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