Various species of Coleoptera such as Scolytidae, Curculionidae, Dermestidae, Nitidulidae, Scarabeidae, Dynastidae, and Cerambycidae have been reported as pests (Rochat el al. 1991, Rochat et al. 2000a, Rochat et al. 2000b; Finnegan & Chambers 1993; Jaffe et al. 1993; Fettöther et al. 1995; Malosse et al. 1995; Cosse & Bartelt 2000). Steirastoma breve (Sulzer) (Coleoptera: Cerambycidae), known as the “cocoa beetle”, is one of the main pests in cocoa (Theobroma cacao, Sterculiaceae) plantations in many countries, and its presence has been reported in South America from Argentina to Venezuela, in addition to some Caribbean islands such as Trinidad, Grenada, Martinique, Puerto Rico, and Jamaica among others (Entwistle 1972; Sánchez & Capriles 1979). According to the literature on S. breve, the egg phase lasts an average of 4.2 days, the larval phase 54.9 days, the pupal phase 10.9 days, and the adult phase 35 days for males and 69 days for females (Mendes & García 1984).

Adults feed on epidermal tissues of the main stem and branches of young plants, producing a characteristic gnawed area, which produces damage to the floral clusters due to the fact that flowers grow in tight clusters on the stem and branches. Moreover, female adults lay eggs inside slits cut into the bark with their mandibles. Usually, both sexes attack young trees aged from six months to four years (Entwistle 1972; Sánchez & Capriles 1979).

Larvae cause severe damage in stem and branches. After eclosion, larvae bore into the bark where they feed on the cambial tissues and the bark itself. First, larvae make a round chamber, progressively enlarging and elongating it until it forms a tunnel or irregular spiral-like galleries. This results in a ringed stem or ringed branch. Depending on the age and location of the damage, these events can kill the apical area. If the main stem is attacked, they can quickly kill the entire plant (Entwistle 1972; Sánchez & Capriles 1979).

Because the use of insecticides has progressively decreased, it is very important to search for alternative pest control methods that are safe for the environment and highly efficient in the management of S. breve populations. In most cases, ethological control has been applied successfully (Nakamuta et al. 1997; Howse et al. 1998; Seybold et al. 2000). This entails previous study of olfactory behavior, as well as the identification and evaluation of the chemical compounds involved in insect communication (Hernández et al. 1992; Howse et al. 1998). An olfactometer has been used to study the olfactory behavior of many coleopteran pests (Rochat et al. 1991; Jaffe et al. 1993; Cerda et al. 1996, Cerda et al. 1999). Electroantennography (EAG) is a technique that has been used for the analysis of biologically active compounds (Marion-Poll & Thiéry 1996). In Coleoptera, EAG and gas chromatography coupled with electroantennographic detection (GC-EAD) have been used to identify the chemical structure and stereochemistry of the sex pheromone of another cerambicid Anaglyptus subfaciatus Nakamuta et al. (1994). Since nothing is known about the olfactory behavior of S. breve towards odors derived from its host plant or from odors emitted by adult insects, the objectives of this study are evaluation of olfactory behavior and electroantennographic responses of S. breve when stimulated by odors derived from cocoa plant tissues and by volatiles produced by males, females, and n-hexane extracts of insect body parts.

Materials and Methods

Results

The responses of S. breve in olfactometric bioassays when stimulated with various odor sources are shown in Table 1. Only three sets of experiments showed statistically significant differences. For example, when CPB vs. male volatiles were tested as odor sources, 55% of the insects showed a statistically significant orientation towards CPB while 29% responded to male odors, with 16% remaining undecided or inactive. There was a clear preference of S. breve towards CPB (67%, P < 0.05 Binomial test) versus a clean air stimulus.

When results from orientation of both sexes towards female vs. male odors are compared, we observed that 67% of the evaluated females oriented toward male odors (Binomial test, P < 0.05). Male to male, male to female, and female to female orientations were not statistically different. The treatments with other odor source combinations (♀ + CPB, ♂ + CPB, ♂ + ♀) did not show significant differences (Binomial test, P > 0.05) (Table 1).

Table 2 shows the results of EAG experiments when male and female antennae were stimulated with female odors, male odors, CPB, and clean wet air. Female antennae produced significantly higher amplitudes (Mann-Whitney U-test, P < 0.05) than males, and gave stronger responses when stimulated with male odors. Male and female electric potential average responses were similar when exposed to CPB odors. Table 2 shows that male and female antennae stimulated with clean wet air showed slight variations in electrical potentials (mV). EAG responses of female antennae towards male + CPB odor produced a larger antennal depolarization that was significant compared to male antennae (Mann-Whitney U-test, P < 0.05) (Table 2).

Additional EAG results obtained from stimulating male and female antennae with n-hexane extracts of body parts are shown in Table 3. Female antennal responses were stronger than males, especially when male prothorax extract was applied as stimulus. Female antennal responses were higher (Mann-Whitney U-test, P < 0.05) when they were exposed to male extracts. On the other hand, these results also show that female head extract produced a stronger signal from male antennae, even though they are both of low intensity, and that prothorax extract generated a significant, slightly higher electroantennographic response from female antennae.

Discussion

The results from olfatometric and EAG experiments suggest that S. breve olfactory behavior is highly influenced by odors emitted by cocoa plants (kairomones) and also quite possibly by a sex pheromone. The fact that during olfactometric tests S. breve female and male individuals were attracted by CPB volatiles, and that additionally these plant odors produced a significant electrical potential deflection (mV) in antennae of both sexes of S. breve, suggests that volatile compounds emitted by cocoa plant tissues are used by the insects as cues to locate cocoa plants as their hosts. After the insect arrives, the host could be a site for feeding, mating, and oviposition, as has been reported for other coleopteran pests of the Curculionidae and Cerambycidae families (Jaffe et al. 1993; Hanks 1999). The presence of a characteristic superficially gnawed area over the attacked cocoa plant cortex is evidence that S. breve use plants as a substrate for feeding.

In another Cerambycidae, Anaglyptus subfasciatus, the existence of a male-produced sex pheromone was first suspected by the fact that females were attracted to males in wind tunnel bioassays (Nakamuta et al. 1994). This was later confirmed by Leal et al. (1995), who identified the pheromone components, but Nakamuta et al. (1997) showed that a blend of host plant volatiles and the male sex pheromone used as baits in yellow water traps were more attractive than sex pheromone or host attractant alone. In some other Cerambycidae species, the existence of sex pheromones in males and also in females is confirmed, along with their host plant relationships (Schröreder et al.1994; Fettköther et al. 1995; Bento et al. 1993; Fukaya et al. 1996).

The presence of a male sex pheromone in S. breve is suggested by olfactometric test results which indicate that females are significantly more attracted to males. In addition, female antennae showed a very strong deflection in EAG experiments only when stimulated by male odors. Female antennae generated strong amplitude signals with all n-hexane male extracts. Male prothorax extract was the source that caused the strongest response.

Sources for pheromone production in Coleoptera vary according to the kind of tissues involved. In Carpophilus freemani (Nitidulidae) the pheromone gland has been located in the abdomen (Dowd & Bartelt 1993). However, in other coleopterans the gland is located in the prothorax. In the case of Anthonomus grandis Boheman (Curculionidae), the aggregation pheromone is produced in the fat bodies associated with the digestive tract (Wiygul et al. 1982), and in Rhynchophorus palmarum L. (Curculionidae), the pheromone glands are located in the male prothorax (Sánchez et al. 1996). In the cerambycid Hylotrupes bajulus the pheromone is produced in a gland situated in the male prothorax (Fettköther et al. 1995). According to the strong evidence provided by the olfactometric and electroantennographic experiments of the present study, it is highly probable that the S. breve pheromone production system is also located in the prothorax. That each of the n-hexane body extracts elicited EAG responses implies that the S. breve digestive system could be associated with pheromone dispersion, as has been reported for other coleopteran insects. Therefore, once male insects arrive at a plant having been attracted by volatile host compounds, it is very probable that males start releasing a pheromone in order to enhance female searching behavior for mating.

In conclusion, the results of the present study when combined with the findings reported in the literature indicate that the chemical communication system and olfactory behavior of S. breve is probably similar to that described for the cerambycid A. subfasciatus. However, it is necessary to continue research, currently in progress, in order to identify the chemical compounds involved in the S. breve communication system. This may enable their use as safe tools for the control of this important neotropical pest.