Novel Attractants of Galleria Mellonella L (Lepidoptera Pyralidae Galleriinae)
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Original article Novel attractants of Galleria mellonella L (Lepidoptera Pyralidae Galleriinae) L Türker I2 Togan S Ergezen3 M Özer1 1 Middle East Technical University, Department of Chemistry, Ankara; 2 Middle East Technical University, Department of Biology, Ankara; 3 Marmara Universitesi, Egitim Fakültesi Bioloji Bölümü, Istanbul, Turkey (Received 20 January 1993; accepted 14 April 1993) Summary — Various synthetic esters were tested for their attractiveness to females of Galleria mel- lonella in comparison with n-nonanal, a component of the sex pheromone of Galleria mellonella. It has been shown that methyl octanoate, ethyl octanoate and methyl decanoate attract females. Parti- cularly, methyl octanoate is as effective as n-nonanal in bioassay and could be a potential pest control agent. Galleria mellonella / sex pheromone / attractant / chemical control INTRODUCTION Burges, 1977, 1978). Further investiga- tions are needed to improve some of these techniques (Spangler, 1986, 1987) The greater wax moth, Galleria mellonella or to more effective and suitable L, is the most serious pest of honey bee explore colonies and stored combs. The rapidly new methods. growing larvae of Galleria feed upon all the Sex pheromones of many Lepidopteran hive products as they burrow through the species are used as pest-management comb and cause heavy damage. There- agents. The sex pheromone of the greater fore, the control of moths without harming wax moth is secreted from the wing the bees and bee products has great eco- glands of males (Barth, 1937) in response nomic importance for bee keepers. to a low frequency sound produced by the There are several physical, chemical wingbeat of females (Spangler, 1986, and biological control methods applied 1987) and has been identified as a 7:3 ad- against the greater wax moth invasion of mixture of 2 aldehydes, n-nonanal and n- honey bee colonies, but all of these have undecanal (Röller et al, 1968; Leyrer and certain disadvantages that limit their use Monroe, 1973). In laboratory and field bio- (Cantwell and Smith, 1970; Ali et al, 1973; assays live males were found to be signifi- cantly more attractive than a mixture of the Experimental chemicals above-mentioned aldehydes in pheromone baits, which did not attract a few beyond The synthesis of n-nonanal was carried out by meters (Finn and Payne, 1977; Flint and the periodic acid oxidation of 9,10-dihydroxy Merkle, 1983). stearic acid (Ernest, 1957). Methyl, ethyl, propyl, Since the above-mentioned aldehydes butyl, allyl esters of octanoic and decanoic acids were synthesized via the reaction of the corre- are easily oxidized in air and lose their ac- sponding acid chlorides and alcohols. Decyl and their effect cannot reach tivity, stimulating octyl acetates, methyl, ethyl, propyl, butyl and al- females unless are conti- effectively they lyl esters of nonanoic acid were all synthesized nously supplied by living males. Hence, via direct esterification reactions of the corre- the aldehydes have a limited applicability sponding alcohols and acids. All the chemicals in population control of Galleria mellonella. were purified repeatedly until spectroscopic puri- Esters are more stable under natural con- ty was reached. ditions. In the we tested the ef- present study, The test chamber fectiveness of several synthetic ester molecules on the females of the greater wax moth. The test-apparatus for the bioassay was de- signed to observe the attractiveness or stimula- tory potential of the synthesized chemicals. It consisted of 2 chambers each MATERIALS AND METHODS spherical glass with a volume of 1 I. The chambers were con- nected via a glass tube 3 cm in diameter and 30 Experimental insects cm in length. To avoid saturation of the chamber where the Larvae of Galleria mellonella were cultured at insects were placed, 2 short plug holes in the 29° ± 1°C and at ≈ 70% relative humidity in an chamber were left open. The apparatus was artificial diet prepared as previously described cleaned thoroughly, dried and reassembled be- by Schmidt and Monroe (1976). The cultures fore each test. The test compounds (table I) were kept in complete darkness. The pupae were introduced into chamber B in adsorbed were collected in individual vessels and newly- form on a supended strip of filter paper (fig 1). emerged female and male moths (6-24 h after Ten female moths were introduced into the oth- adult emergence) were collected in separate er chamber, which was left open to air via the groups. plug holes. Bioassay for stimulatory activity The behavior of these insects has been de- of synthetic substances scribed as "inactive response". If a high degree of mobility within the flask in which they were placed was initially observed, this behavior was called Prior to with the test experimentation chemicals, "agitative activity". If, on the other hand, the ob- a set of control was on experiments performed served behavior among the test insects was very test of moths. The females were groups placed similar to that observed during exposure to n- in the and the hole was chamber, plug opened nonanal, it was termed "active response". This to air for 2 h. Within that of time no moths period last type of activity can be described as follows: passed into the other chamber and they re- first, movement ot the antennae, then rapid fan- mained still. The control experiment was repeat- ning of wings and dancing of each individual ed 5 times. around itself were observed. These patterns of All the experiments were carried out at at- activity were accompanied by the transport of mospheric pressure in daylight and between 9 some or all the stimulated insects into the second am and 3 pm. flask where the test chemicals were placed. All the insects which moved into the second flask ex- Three classes of female responses were ob- hibited and served during the experiments. Upon exposure to ovipositioning fanning. some of the esters, the test insects were sessile, In the control groups, the effectiveness of sol- just as the insects in the control experiments vents used for cleaning purposes such as ace- were at all tested volumes of the compounds. tone, ethyl alcohol and water were tested on fe- male behavior and found to be ineffective. After- RESULTS AND DISCUSSION wards, the synthesized compounds were tested for a 2-h period. Firstly, 1 μl of the compound was introduced into chamber B (fig 1). If no re- The pattern of the "active response" type sponse was observed within ca 2 h, then the of behavior was very similar to the sexual volume was increased up to 50 μl, followed by responses of females to the sex phero- increments of 10 to 400 For each vol- μl up μl. mone n-undecanal as described by Röller ume of the fresh of Galleria compounds, groups et al (1968). Furthermore, all the tested mellonella individuals were used. Ten females were found to be ineffective on were used in each test (5 replicates). compounds males. The behavioral thresholds (the volume of chemical required to initiate the response in the Types of female responses to the tested females, the equivalent weights) and the latency compounds are given in table I, along with periods (the time elapsed until the response was the volumes and weights of the chemicals observed) were recorded (table I). Similar tests for the behavioral thresholds and latency were carried out on Galleria mellonella males. periods. Expression of the "active response" type Statistical methods of behavior by females but not males to methyl and ethyl octanoates and methyl decanoate as well as n-nonanal suggests Analysis of variance (ANOVA) was used to that the first 3 esters constitute novel sex compare the means of the percent age of at- tracted females. To satisfy the assumptions of attractants in the greater wax moth. The the method, the percentages were subjected to percentage of females attracted by these improved angular transformation (Sokal and compounds in each of the replicates (sets) Rohlf, 1981). Homogeneity of group variances has been presented in table II. were tested by Fmax (Sokal and Rohlf, 1981). Pairwise of means of the When the ANOVA F-statistic was significant at comparision least at the 5% level, T-test was then used to transformed percentages by a subsequent partition means into significant ranges. T-test (Sokal and Rohlf, 1981) revealed that the percent attraction by n-nonanal rasite important des colonies d’abeilles do- and methyl octanoate were not significant- mestiques et des réserves entreposées ly different at the 5% level and that these 2 dans les rayons. Aussi, une méthode de compounds were significantly more attrac- lutte non dangereuse pour les abeilles est- tive to the insects than ethyl octanoate or elle d’une grande importance économique methyl decanoate (table II). pour les apiculteurs. Il existe divers procé- The amounts of test chemicals used in dés de lutte contre la teigne de la ruche, the present study should not be directly physiques, chimiques et biologiques, mais compared with the amount of pheromone ils présentent tous des inconvénients. La sexuelle du mâle de la produced by the insects under natural con- phéromone teigne ditions because the signal is conveyed by est un mélange de n-nonanal et de n- the amount of chemical transferred into the undécanal dans le rapport de concentra- vapor phase which should be considerably tion de 7:3. Parce que ces aldéhydes peu- much less (all have high boiling points) vent facilement s’oxyder à l’air et qu’ils per- than the amount used in the liquid phase dent alors leur activité, leur application est of these compounds. If ventilation is not limitée. C’est pourquoi des composés ap- adequate, material transferred from one parentés ont été produits par synthèse, et chamber into the other is mainly due to leur pouvoir attractif testé dans un appareil passive diffusion.