Toulassi Atiama-Nurbel1, Amandine Ligonière1, Lucile Müller1, Jean-Philippe Deguine1,
Anne Bialecki2 and Serge Quilici1
1
UMR Peuplements Végétaux et Bioagresseurs en Milieu Tropical (CIRAD- Université de La Réunion), 97410 Saint-Pierre, Réunion, France
2
Laboratoire de Chimie des Substances Naturelles et des Sciences des Aliments, Université de La Réunion, 97 744 Saint-Denis cedex 9, La Réunion, France
Abstract
The melon fly Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae) is a serious pest of tropical cucurbit crops in many parts of the world. Damages are caused by female ovipositing in fruits. Nowadays no powerful attractant is available to monitor or control these female population. The use of kairomones (plant volatiles) might be a promising solution for controlling B. cucurbitae. In this study, we evaluated the attractiveness of ten volatiles compounds from Cucurbitaceae for the females of B. cucurbitae using olfactometry bioassays. Ten compounds (alcohols, aldehydes and a terpene) have been tested individually at two doses, and in blends of 2,3,4 and all individual-compounds. The study of responses to individual compounds enabled us to select the four most attrative compounds at the optimal dose: the nonanol, the (Z)-non-6-enal, the (Z)-non-6-en-1-ol and the (E,Z)-nona-2,6-dienal (each of them at least 40% response). Then, three blends induced a response of females equivalent to the odor of fresh cucumber (around 60 % response). The two-component blend with (Z)-non-6-en-1-ol and (E,Z)-nona-2,6-dienal constitutes a promising solution for practical use in trapping systems for B. cucurbitae.
Introduction
The melon fly, Bactrocera cucurbitae (Coquillett), is a serious pest of tropical agricultural crops, whose current distribution ranges from South Asia, from where it originates, to many Pacific Islands (including Hawaii) (Dhillon et al., 2005), some Indian Ocean islands (including Mauritius and Seychelles) (White et al., 2000) and many African countries (Vayssières et al., 2007). This pest is causing damage at least to 81 host species worldwide (Dhillon et al., 2005) with a strong host association with most species of the Cucurbitaceae family (Allwood et al., 1999). In La Réunion, damage may sometimes affect the total cucurbit crop when no control method is applied (Ryckewaert et al., 2010). As chemical control of fly populations appears of limited efficacy and constiutes a non-sustainable approach, it is thus necessary to develop more environmental friendly control methods such as mass-trapping of females based on plant semiochemicals as a source of attractants. While several attractants for males are currently available (raspberry ketone and its derivatives cuelure and melolure), chemicals that attract females are limited to food-type attractants which lack of effectiveness, have limited field life, are difficult to handle and also attract non-target species (Jang & Light, 1996 ; Siderhurst & Jang, 2010). Attracting females is critical in population management because reducing female population by trapping prevents future offspring (Jacobson et al., 1971). A comprehensive understanding of resource-foraging behavior (host finding and oviposition) of fruit fly females as well as the identification of chemical stimuli eliciting this behavior is therefore central for the development of effective trapping systems to monitor and /or control its population. Host fruit are of particular interest since some tephritid females preferentially use host fruit volatiles when searching for oviposition sites (Jang & Light, 1996 ; Light & Jang, 1996). Volatile fruit odors have been used successfully as attractants for the stenophagous apple maggot fly, Rhagoletis pomonella (Walsh) (Reissig et al., 1985 ; Jones, 1988 ; Jones & Davis, 1989 ; Agnello et al., 1990) and volatile fruit odors have been investigated as potential attractants for other oliphagous and polyphagous tephritid species (Robacker et al., 1992 ; Nigg et al., 1994 ; Prokopy & Vargas, 1996 ; Robacker & Heath, 1996 ; Warthen et al., 1997 ; Prokopy et al., 1998). As to B. cucurbitae, previous investigations have focused principally on female response to entire host fruit odor (cucumber, zucchini, bitter melon, kabocha, cantaloupe and ivy gourd) (Miller et al., 2004 ; Piñero et al., 2006). Few studies have identified volatile compounds which can be attractive for the females of the Melon fly. Jacobson et al. (1971) when evaluating the attractiveness of different derivates of nonenyl acetates found that trans-6-nonen-1-ol acetate was attractive for the Melon fly female. More recently, Siderhurst et al. (2010) using coupled GC-EAD studies,
identified 31 compounds in fresh and aged puréed of cucumber that elicited EAD response of females, and found a synthetic nine-component blend attractive for Melon fly female in olfactometer and field tests. As Melon fly had a strong association with a variety of cucurbits, we investigated the volatile composition of many species within this plant-family (Atiama-Nurbel et al., 2014 (in press)) (Atiama-(Atiama-Nurbel, unpub.data) and chose 10 volatile compounds (alcohols, aldehydes and a terpene) according to their occurrence in the odor bouquets within this family and the knowledge on their potential role in insect-plant interactions. The practical objective of this study was to find an attractant for Melon fly females based on volatile compounds of representative of the Cucurbitaceae family. For this, we evaluated the attractiveness of ten volatile compounds for B. cucurbitae females using olfactometry bioassays. Ten compounds (alcohols, aldehydes and a terpene) have been tested individually at two doses, and in blends of 2,3,4 and all individual-compounds.
Materials and Methods
Flies
The strains of reared flies used in the study, came from infested pumpkins, Cucurbita maxima D. (Cucurbitaceae) collected in June 2000 at three locations in La Réunion (Petite Ile, Bassin Martin, and Piton Saint-Leu). Adult flies obtained from these samples were reared under controlled conditions: 25 ± 2°C, 70 ± 20% RH, and a photoperiod of 12:12 (L: D) h. They were given free access to granulated sugar, enzymatic yeast hydrolysate (ICN Biomedicals, Aurora, OH), and water. Zucchini (Cucurbita pepo L.) (Cucurbitaceae) was used as an egg- laying substrate for the adult females and the larvae were fed with pumpkins. New cohorts of adult flies were reared in the same conditions but they were allowed no contact with host plant material and females had no prior ovipositional experience until the start of the bioassays. Flies were approximately 25-30 days old when tested and were presumed to be mated (95% of mated females were found in cages with adults of both sexes when 7 days old (Jang et al., 1997)).
Chemicals
The ten chemicals tested were purchased from Sigma-Aldrich (Saint Louis, MO, USA). Compound purities were >98% based on GC-MS analysis. In all bioassays, chemical compounds were presented at a given dose (1 or 5 µL) on a cotton disc (Quilici et al., 2013).
Experimental procedure
Laboratory olfactometers were used to evaluate the relative attractiveness of single compounds and blend of cucurbit volatiles. The one-arm olfactometers derived from the model previously described by (Katsoyannos et al., 1980) were used to evaluate the relative attractiveness of single compounds and blends of cucurbit volatiles. It consists of a clear polycarbonate (Nalgene®, Thermo Scientific, New York) cylindrical tube (84 cm length and 11.5cm wide) divided into 3 consecutive chambers ended by a fan (4 x 4 x 2 cm, 12V, Sunon, China) providing an airstream (0.35 m s.-1) along the system. The first chamber (test chamber) containing the females is limited by a wire screen funnel leading to the second chamber (catch chamber) followed by the last chamber (bait chamber) containing the chemical applied on a cotton disk. The olfactometers were placed on a lab bench in a series of 4 units (3 with chemicals and 1 control) that operate simultaneously. Neon light strips on top of the four olfactometers provide an homogeneous light of 2000 lux.
In each olfactometer, 30 females were placed in the test chamber and then exposed to the tested chemical(s) for 1 hour. Females responding to the odor stream oriented themselves toward the odor source, landed on the wire-screen cone and were funneled into the catch chamber, where they were counted after 1 hour of exposure.
We first evaluated the attractiveness of 10 individual compounds at 2 doses (1 and 5 µl) for B.
cucurbitae female. As a result of this first experiment, we decided to use the compounds at
their optimal dose (ie the dose with significantly better response rates or minimal dose (1μl) (when no significant difference)) for composing blends for further tests. Twelve different lures were composed: one blend with all the single compounds, one blend with the four single compounds with best female response and 10 other blends combining these last 4 compounds in two or three-component blends (Table V-1). In order to compare the relative attractiveness of blends, 1µl of each blend was used in each olfactometer. The relative attractiveness of the 4 best individual compounds and of the 6 blends was compared with the attractiveness of a fresh cucumber cut in pieces (cucumber of 800 g, Var. ‘Vantage’, 35.6 x 7.9 cm). The amount of cucumber presented per olfactometer was 30g. Each treatment (single compounds, blends