Field evaluation of attractive lures for the fruit fly Bactrocera minax (diptera: tephritidae) and their potential use in spot sprays in Hubei province (China)
Field Evaluation of Attractive Lures for the Fruit Fly Bactrocera minax (Diptera: Tephritidae) and Their Potential use in Spot Sprays in Hubei Province (China) Author(s): Xiao-Wei Zhou, Chang-Ying Niu, Peng Han, and Nicolas Desneux Source: Journal of Economic Entomology, 105(4):1277-1284. Published By: Entomological Society of America https://doi.org/10.1603/EC12020 URL: http://www.bioone.org/doi/full/10.1603/EC12020
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FIELD AND FORAGE CROPS
Field Evaluation of Attractive Lures for the Fruit Fly Bactrocera minax (Diptera: Tephritidae) and Their Potential Use in Spot Sprays in Hubei Province (China) XIAO-WEI ZHOU,1 CHANG-YING NIU,1,2 PENG HAN,1,3
J. Econ. Entomol. 105(4): 1277Ð1284 (2012); DOI: http://dx.doi.org/10.1603/EC12020
ABSTRACT The Chinese citrus fruit ßy, Bactrocera minax (Enderlein) is a univoltine Tephritidae pest that infests Citrus species. Field trials were conducted in 2010 to determine the potential use of a lure based on enzymatical-hydrolyzed beer yeast as liquid bait (hereafter named H-protein bait) for B. minax in the Hubei province, China. In a citrus orchard, we compared the attractiveness among aqueous solutions of H-protein bait, GF-120 fruit ßy bait, sugar-vinegar-wine mixture, torula yeast, and Jufeng attractant when used in traps and in spot sprays, that is, lures used in combination with the insecticide trichlorphon. The H-protein bait was the most attractive lure in traps, ensnaring signiÞcantly more adults than sugar-vinegar-wine mixture, torula yeast, and Jufeng attractant, in decreasing efÞciency order. In spot sprays those with H-protein bait killed signiÞcantly more female and male ßies within 40 min than those with sugar-vinegar-wine mixture, GF-120, Jufeng attractant, and the control. In addition, the total number of ßies killed by H-protein bait during the spot spray duration was higher than other treatments. Our results demonstrated that the H-protein bait may be a useful tool in citrus orchards in China to monitor B. minax populations as well as to manage this pest when used in spot sprays. KEY WORDS hydrolyzed-protein bait, GF-120, phenology, citrus orchard, monitoring
The Chinese citrus fruit ßy, Bactrocera minax (Enderlein) (Diptera: Tephritidae) has been a serious pest on citrus fruits in China, India (West Bengal and Sikkim), and Bhutan for more than half a century (Drew 1979, Wang and Luo 1995, Dorji et al. 2006). Among ßies of the Dacinae subfamily (Tephritidae family) B. minax has the particularity of being a univoltine species (Dorji et al. 2006), that is, one generation per year. The host range of B. minax is restricted to the Citrus species and adult emergence usually occurs from late April to early May, with adults present in the Þeld until
August in China. Adults lay eggs on immature citrus fruits from mid-June to July. The phenology varies among infested regions and appears to vary according to local temperatures (Wang and Luo 1995). In the Hubei province, B. minax is considered one of the major fruit ßy pests on citrus trees, causing considerable economic damage by making the fruits improper for merchandizing (Yang et al. 1994). Therefore, effective Integrated Pest Management (IPM) programs are required for areas where the citrus production is a major economic issue. 1 Hubei Insect Resources Utilization and Sustainable Pest Management Key Laboratory, College of Plant Science & Technology, Huazhong Agricultural University, Wuhan 430070, China. 2 Corresponding author, e-mail: firstname.lastname@example.org. 3 French National Institute for Agricultural Research (INRA), 400 route des chappes, 06903 Sophia-Antipolis, France.
Traps, attractive lures, and mass trapping techniques are common strategies for management of fruit ßies (Hendrichs et al. 1995, Stonehouse et al. 2002, Vargas et al. 2009). For example, McPhail traps baited with a fermenting mixture of citrus juice and brown sugar were successfully used to trap fruit ßies in early studies in Florida (Newell 1936). In addition, an approach combining olfactory attractants and baits sprayed with insecticides, that is, lures ϩ insecticides has been used to combine detection and monitoring in the management of fruit ßies. Protein baits sprayed with insecticides were Þrst used for Caribbean fruit ßy control (Steiner 1952). Since then, protein bait sprays have become a major method of suppressing or eradicating fruit ßy populations in many parts of the world. Female tephritids require a protein meal for ovarian development and egg production (Christenson and Foote 1960, Mangan 2003, Perez-Staples et al. 2007) and protein sources, such as bacteria in bird feces, are thought to be scarce in natural conditions (Drew et al. 1983). Therefore, protein sources are highly attractive to tephritid females and thus protein bait sprays have been successfully used to manage tephritid species (Yee and Chapman 2005; Mangan et al. 2006; Vargas and Prokopy 2006; McQuate 2009; Pin˜ ero et al. 2009a,b). Two commonly used protein baits for fruit ßy detection, monitoring, management, and trapping are the GF-120 Fruit Fly Bait (Dow AgroSciences, Indianapolis, IN) and torula yeast (ERA International,
0022-0493/12/1277Ð1284$04.00/0 ᭧ 2012 Entomological Society of America
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Freeport, NY) (Burns et al. 2001; Vargas et al. 2002; Prokopy et al. 2003; Stark et al. 2004; Mangan et al. 2006; McQuate 2009; Pin˜ ero et al. 2009a,b). To our knowledge, potential use of protein baits to manage B. minax in China has not been documented. In most places where B. minax is present in this country, farmers frequently rely on spraying sugar-vinegar-wine mixture with insecticides to manage the pest (Wang and Luo 1995). Though most fruit ßies can be monitored by traps baited with sexual pheromone and parapheromones, B. minax is not known to be attracted to any male lures (Drew et al. 2006). How the GF-120 and torula yeast can be effective against B. minax in China has not yet been documented. By contrast, the Jufeng attractant is registered and commercially available as lure for B. minax in China though its efÞcacy has never been clearly demonstrated. In our laboratory, a new protein bait has recently been developed to control B. minax. It is based on an enzymatically hydrolyzed protein produced by the industrial processing of beer yeast. This hydrolyzed protein bait (hereafter named H-protein bait) also includes feeding stimulants, orange juice and brown sugar, as well as a chemical attractant ammonium acetate. In 2010, in Þeld conditions we assessed the effectiveness of the H-protein bait and several other commercially available lures for potential monitoring and management of B. minax. In a citrus orchard, two experiments were performed: 1) we compared the efÞcacy of the H-protein bait, torula yeast, sugar-vinegar-wine (SVW) mixture, and Jufeng as attractant for B. minax, and 2) we studied how various lures (Hprotein bait, SVW mixture, GF-120, and Jufeng as an attractant) are effective against B. minax when used in spot sprays, that is, when combined with an insecticide. As a hand sprayer was needed to create the spot sprays, torula yeast that did not completely dissolve in water (it is provided as pellets) could not be used in this experiment. GF-120 was used only in the second experiment as a positive control (it is commonly used for spot sprays in various crops or orchards). Materials and Methods Field Site. The study was conducted from early May to early August 2010, during the fruiting season, in a 0.4 ha citrus orchard in Yichang, in the Hubei province, China. The orchard was composed of mainly mandarin (Citrus reticulata), navel orange (Citrus sinensis), and pomelo (Citrus maxima) trees. Density of trees in the orchard was Ϸ1,000 trees per ha. No management practice for B. minax or other pest control was carried out in the orchard during the experimental periods, except for routine management such as grass-mowing and pruning. The average daily temperature and relative humidity during the experiments were 27.2 Ϯ 0.3ЊC, 67.5 Ϯ 4.3%. Lures. The lure treatments were prepared using the following formulations: 1) H-protein bait: 20% vol:vol solution in water; 2) SVW mixture: 3% sugar solution with vinegar and wine mixture; 3) torula yeast: two pellets per 300 ml of water; 4) GF-120 fruit ßy bait: 1:3
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(vol:vol) solution at the recommended application rate; and 5) Jufeng attractant: 25 g of lure in 300 ml water as recommended. The pH of lures was checked after preparation using a pH meter (PB-10, Sartorius, Germany). Experiment 1: Assessment of Attractiveness of the Lures in Citrus Orchard. The experiment was set up on 4 May 2010 and trapping was carried out periodically, once a week, until 10 August 2010. Four types of lures were tested for attractiveness in traps: H-protein bait, SVW mixture, torula yeast, and Jufeng attractant. The traps were made of modiÞed plastic water bottles (height: 18 cm, diameter: 9 cm, a 4 ϫ 3 cm window 8 cm from the bottom of the bottle). This design ensnares adult ßies which are unable to escape and drown in water or die because of starvation. The traps were baited with 250 ml of aqueous solution and attached to citrus tree branches 1.5 m in height. The traps were located randomly within the orchard with a space of 15 m separating the traps. The trials were set up in a randomized complete-block design using nine replicates (traps) per lure type, that is, 4 ϫ 9 ϭ 36 traps were used every week. Once a week, when traps were checked and ßies were removed, the traps were thereafter washed with water and lures renewed. At the same time, each trap was moved to a new position within the experimental site. The number of male and female B. minax captured was recorded and the ßies were placed in 75% ethanol. The females were subsequently dissected to estimate their sexual maturity under a stereomicroscope (Nikon Inc., SMC-10, Japan) and placed into three categories: immature (ovaries small and no eggs present), semimature (ovaries with eggs developing but not mature), and mature (ovaries with mature eggs present). This classiÞcation was based on preliminary laboratory experiments (C.Y.N., unpublished data). The number of male and female ßies trapped per week was analyzed using a generalized linear model (Proc Genmod; SAS Institute 1999) with ÔLure,Õ ÔSex,Õ ÔDateÕ as factors. Potential interactions among these factors were also tested. In addition, we tested for a potential relationship between the numbers of B. minax female ßies ensnared in H-protein bait traps and the sexual maturity of these ßies, using regression analyses. This analysis was carried out to assess the attractiveness of the H-protein bait in relation to the sexual maturity of B. minax females. Experiment 2: Assessment of Effectiveness of Lures When Used in Spot Sprays. Spot spray tests were conducted from 4 June to 14 June in 2010 in the citrus orchard. The test was designed to study the speed and the overall effectiveness of four different lures when used in spot sprays under Þeld conditions. Five lureinsecticide solutions, that is, spot sprays, were prepared: the H-protein bait, the SVW mixture, the Jufeng attractant, the GF-120 (positive control), and water (control), and all solutions were supplemented with the Trichlorphon 90% WP insecticide (Dacheng Pesticide Co., Ltd., Shandong, China) at the rate of 0.12 g per 100 ml of solution. For each replicate (i.e., spot spray), 150 ml solutions (with lures and insecti-
ZHOU ET AL.: ATTRACTIVE LURES FOR THE FRUIT FLY Bactrocera minax
Table 1. Statistics from the generalized linear model used to analyze the numbers of ﬂies Source of variationa A: Lure attractiveness Lure Sex Date Lure ϫ Date Lure ϫ Sex Date ϫ Sex B: Spot spraysÑimmediate effect Lure Sex Lure ϫ Sex C: Spot spraysÑtotal effect Lure Sex Lure ϫ Sex
3 1 10 30 3 10
143.09 0.09 283.24 654.01 0.91 16.34
Ͻ0.001 0.761 Ͻ0.001 Ͻ0.001 0.823 0.090
4 1 4
63.05 3.30 5.35
Ͻ0.001 0.069 0.253
4 1 4
46.57 10.71 6.21
Ͻ0.001 0.001 0.184
a (A) trapped during the course of the exp, (B) found dead on the spot sprays during the Þrst 40 min after initial spray application (i.e. immediate effect), and (C) killed on the spot sprays until no dead ßies were longer observed on the spots (i.e. persistent effect) among the various lures tested (Lure factor), as function of ßies sex (Sex factor) and also as function of date of sampling (Date factor) in case of the Lure attractiveness experiment.
cide) were applied to foliage of two citrus trees using hand-held sprayers (500 ml in capacity) (Farm and Garden Machinery Sales Co., Jinhua, China). The tests were carried out at 10:30 a.m., that is, when B. minax shows its highest diurnal activity (C.Y.N. and X.W.Z., unpublished data). Three replicates were undertaken per lure tested and sprayed trees were always at least 15 m apart from each other. Two parameters were recorded: 1) to assess the speed of the effect of spot sprays on B. minax, dead ßies on the sprayed trees were counted 40 min after the solutions had been
applied, and 2) to evaluate the total number of ßies killed on the sprayed trees, spot sprays were checked for dead ßies every day until none were observed. Overall effectiveness of sprays was assessed according to the lures used. The number of male and female killed on spot sprays were analyzed using a generalized linear model (Proc Genmod; SAS Institute 1999) with Lure and Sex as factors, and interaction between these two factors was also tested. Results Attractiveness of the Lures. The statistical results are summarized in Table 1A. Overall, a total of 8,894 B. minax adults were ensnared during the season (4,340 females and 4,554 males). The sex of ßies trapped was not a driving factor (no signiÞcant Sex factor). The various lures trapped males and females in a similar way (no signiÞcant interaction between Sex and Lure factors) and regardless of date (no signiÞcant interaction between Date and Sex factors). Lures varied strongly in their attractiveness (i.e., signiÞcant Lure factor, P Ͻ 0.001). Overall, traps with the H-protein bait trapped much more B. minax ßies than traps with other lures (Fig. 1). In addition, the SVW mixture proved to be more efÞcient than Torula yeast and Jufeng attractant lures. The number of B. minax ßies also varied according to the date (signiÞcant Date factor) (Table 1A) with ßies being trapped from midMay through early August and a peak was observed on 8 June (Fig. 1). It reßected the seasonal phenology of B. minax in the studied area. However, this peak was more marked in the case of the H-protein bait than for the other lures, although the SVW mixture showed also a peak to a lesser extent. For example, the number
Fig. 1. Mean number of B. minax ßies (mean Ϯ SEM) captured per trap weekly using various lures (H-protein bait, SVW, torula yeast, and Jufeng attractant) in experimental orchards in Hubei province, China.
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Fig. 2. Female ßies of B. minax in different ovarian development situations ensnared in traps by H-protein bait solution by date in citrus orchards in the Hubei province, China.
of ßies trapped by H-protein bait traps increased much more than those trapped by the SVW mixture when ßy density increased in the orchard, hence signiÞcant interaction between the Lure and the Date factors (Table 1A). The torula yeast and Jufeng attractant lures, that is, the two least efÞcient lures, only trapped a few ßies throughout the entire season. B. minax females previously ensnared by H-protein bait traps were dissected and their sexual maturity was estimated. Most females trapped by the H-protein bait were still immature and semimature until mid-June (Fig. 2). The proportion of fully mature females increased from Ϸ5% in mid-June to almost 90% in late June. All females trapped from early July until the end
of the test were sexually mature. There was a positive signiÞcant relationship between the number of female ßies trapped by the H-protein bait for immature (Fig. 3; R2 ϭ 0.91, F9 ϭ 89.88, P Ͻ 0.001), semimature (R2 ϭ 0.61, F9 ϭ 13.60, P ϭ 0.005) but not for fully mature ßies (R2 ϭ 0.01, F9 ϭ 0.12, P ϭ 0.733) (Fig. 3). Effectiveness of the Lures for Spot Sprays. The number of females and males killed within 40 min varied signiÞcantly according to the lures used in spot sprays (Fig. 4, signiÞcant Lure factor, Table 1B). Overall, during the Þrst 40 min after spray applications, a similar number of B. minax males and females were killed on sprayed trees (no signiÞcant Sex factor) and there was no interaction between
Fig. 3. Relationship between sexual maturity stage (immature, semimature, and mature) ovarian development of females of B. minax ßies trapped in H-protein bait traps and the numbers of ßies trapped by these traps. Regression lines are included for the three relationships tested.
ZHOU ET AL.: ATTRACTIVE LURES FOR THE FRUIT FLY Bactrocera minax
Fig. 4. Mean number (ϮSEM) of B. minax ßies killed in spot sprays (lure ϩ trichlorphon) within 40 min after application. Lures tested were H-protein bait, GF-120, SVW mixture, Jufeng attractant, and control (i.e., water).
Lure and Sex factors. Nonetheless, more males than females tend to be killed in the speciÞc case of H-protein bait spot sprays, but the results were not statistically signiÞcant (Fig. 4). The total number of B. minax females and males killed on spot sprays (i.e., until dead ßies were no longer observed on sprayed trees) differed signiÞcantly according to lures used (signiÞcant Lure factor, Fig. 5, Table 1C). Spot sprays with H-protein bait ensnared more females than the GF-120, the SVW
mixture, the Jufeng attractant and control spot sprays. By contrast to what was observed within the Þrst 40 min after initial spot sprays application, signiÞcantly more males than females were trapped (signiÞcant Sex factor). The interaction between the two factors was not signiÞcant, that is, H-protein bait, GF-120 and SVW mixture spot sprays showed the same trend (though the less efÞcient Jufeng attractant and the control sprays did not show this difference in numbers of male and female trapped).
Fig. 5. Mean total number (ϮSEM) of B. minax ßies killed in spot sprays (lure ϩ trichlorphon) until dead ßies found dropped to zero (daily). Lures tested were H-protein bait, GF-120, SVW mixture, Jufeng attractant, and control (i.e., water).
JOURNAL OF ECONOMIC ENTOMOLOGY Discussion
Many ßies in the Bactrocera genus are major pests around the world (Vargas et al. 2009, Daane and Johnson 2010, Pascual et al. 2010, Han et al. 2011, Benelli et al. 2012, Canale and Benelli 2012) but B. minax is considered a major pest particularly in Asia. The Sterile Insect Technique (SIT) was used in the 1980s to manage this pest in the Guizhou province in China; this method achieved some success (Wang and Luo 1995). However, it relied mainly on government heavy investment in mass-rearing facilities and the widespread release of sterile insects. In the current study, we demonstrated that the H-protein bait may be a promising lure to monitor this ßy in Citrus orchards in China and moreover to manage this pest when used in spot sprays. The H-protein bait was the most attractive lure to both sexes of B. minax; not only in traps but also in spot sprays. Other lures tested, including the wellknown GF-120, proved to be less efÞcient than the H-protein bait in attracting B. minax. Various lures including fruit-derived, food-based, visual, and pheromone will continue to be an integral part of the management of tephritid pests. For a given tephritid pest, the attraction and responses to the lures are completely different. For example, ßies in the Bactrocera genus depend much more on olfactory lures compared with Rhagoletis ßies, which seem to rely on visual cues (Prokopy and Papaj 2000). The hydrolyzed proteins were used as baits to attract fruit ßies, but comparisons of several proteins indicated that hydrolyzed torula yeast was superior in attracting Anastrepha spp. (Epsky et al. 1993). In this case, it is necessary to assess the effect of the protein bait lures on each target species before commercial use. Unfortunately, very few studies are available about the response of lures in B. minax. Drew et al. (2006) reported the attractiveness of various combinations of colors and shapes to B. minax in Bhutan. Our study is the Þrst to conÞrm that B. minax can have signiÞcant response to an enzymatical-hydrolyzed protein bait. By quantifying attraction and responses of B. minax compared with other commercially used lures, a breakthrough has been made in understanding the H-protein bait against B. minax. However, many components of the protein baits (e.g., protein concentrations, pH, and ammonia concentrations) also likely greatly inßuence attractiveness for B. minax. Thus, bearing in mind the aforementioned factors, it would be worth developing improved lures when furthering work on the Chinese citrus fruit ßy. In this Þeld study, we showed that the H-protein bait was very effective on B. minax both when used in traps and for spot sprays. However, it is worth mentioning that proteins-based lures used in traps may perform differently than when sprayed on plant foliage. For example, some proteins sprayed on plants may be phytotoxic and thus may have to be diluted before potential use as spot sprays (Mangan et al. 2006), and protein mixtures containing ammonium acetate may not always work well for trapping purposes (Robacker et al. 1996).
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The response of fruit ßies to protein baits is known to depend greatly on their feeding status and sexual development (Robacker 1991, Cornelius et al. 2000, Miller et al. 2004). Our Þeld trials suggested that most of the B. minax adult ßies ensnared by H-protein bait traps occurred before late June, that is, about 1 mo after adult ßies emerged according to the phenology of B. minax in Hubei province (C.Y.N., unpublished data). This need for proteins in female ßies may be exploited as they are likely to be particularly attracted by protein sources early in the season (Health et al. 1993). The highest number of female ßies trapped per week occurred on 8 June and it drastically decreased on 6 July. Five percent of females trapped on 15 June were sexually mature whereas almost 96% of females in traps with H-protein bait solutions in early July were fully sexually mature (many mature eggs). When females become sexually mature they need proteins for the development of eggs and therefore Þnding protein sources is of primary importance (Mangan 2003, Perez-Staples et al. 2007). Studies reported that females deprived of nitrogenous food are more attracted by protein bait (Prokopy et al. 1992, 2003; Vargas et al. 2002; Rousse et al. 2005; Miller et al. 2004). Our results support the hypothesis that responses of B. minax females were affected by protein requirement as demonstrated in other Bactrocera species (Robacker 1991, Miller et al. 2004, Perez-Staples et al. 2007, Mangan 2009). Investigations of ovarian development on B. minax females revealed that ovarian development appears to be a reliable indicator of sexual maturity, and protein was a vital component of ovarian development for female B. minax. In laboratory conditions, when B. minax females were fed a full diet (including proteins provided ad libitum), they reach sexual maturity and complete the reproduction phase in Ϸ25 d (C.Y.N., unpublished data). It is also consistent with the low number of sexually mature females trapped from late June as ßies no longer needed much protein (when oogenesis was Þnished). In our Þeld trials, traps baited with protein lures captured more nontarget insects than other lures because these insects are attracted to the odors of protein bait (Thomas 2003). The nontarget insects captured were mainly the species belonging to Diptera, Hymenoptera, and few Coleoptera and Lepidoptera, such as scarabs and moths. For Diptera, a number of Muscidae, Calliphoridae, and Sarcophagidae were trapped. Some beneÞcial insects, like honey bees and other wild bees, were trapped in small numbers. When protein bait was used in sprays, these pollinators may feed on the bait and be affected or even killed by insecticides (Thomas and Mangan 2005, Desneux et al. 2007). This problem associated with the use of pesticides in bait spray can be solved by using insecticides which are less toxic on nontarget organisms. Bait sprays with new types of insecticide that are less harmful to nontarget organisms have been used successfully in Þeld trials against Anastrepha ludens (Loew), A. obliqua (Macquart), A. suspensa (Loew), Bactrocera cucurbitae (Coquillett), Bactrocera dorsalis (Hendel),
ZHOU ET AL.: ATTRACTIVE LURES FOR THE FRUIT FLY Bactrocera minax
and Ceratitis capitata (Wiedemann) (Burns et al. 2001, Moreno et al. 2001, Vargas et al. 2003, Prokopy et al. 2003, Stark et al. 2004, Thomas and Mangan 2005, Pin˜ ero et al. 2009b). In fact, the new toxicants that are noticeably different to traditional pesticides are less likely to kill adults through contact, but instead must be ingested before producing maximum toxic effect (DowElanco 1994, Vargas et al. 2002). The results of the current study may be useful in optimizing IPM programs for the control of B. minax in China. Previous IPM programs developed against Bactrocera ßies hinted at the importance of efÞcient lures in IPM packages for successful management. For example, a wide-scale successful IPM program against B. dorsalis was implemented in Hawaii in 2000 and use of protein bait sprays and spot sprays (protein baitbased) targeting female ßies was proven to be a keystone component of such IPM (Mau et al. 2007; Vargas and Prokopy. 2006; Pin˜ ero et al. 2009a,b). Using Hprotein bait has great efÞciency potential when implemented in IPM programs against B. minax, for two main reasons. Firstly, H-protein bait showed great efÞciency in mass trapping as well as in spot spray targeting B. minax adults. In practice, adults are caught or killed before they became sexually mature, that is, before the ßies effectively start to lay eggs on or in fruits and thus it could prevent any adverse effects on fruits by larvae. Secondly, the speciÞc phenology of B. minax, that is, univoltine species, may allow IPM programs to focus most of management efforts for a relatively short time span in the year; from the emergence of ßies and until they become sexually mature. The main sanitation measures in Citrus orchards, for example, collection and removal of all fallen infested fruits at the end of the fruit season combined with H-protein bait mass trapping and spot sprays can largely suppress the B. minax populations and therefore ensure a successful IPM program. In addition, the implementation of such a strategy may be cost-effective and environmentally sound, it would particularly help to reduce pesticide applications and possible negative effects on beneÞcial arthropods (Desneux et al. 2006, 2007; Han et al. 2010) and on associated ecosystem services (Lu et al. 2012). The data presented here strongly showed that Hprotein bait can be an effective lure for both female and male B. minax in citrus orchards during the fruiting season. Protein bait attracted more B. minax adults than other attractants in this study. Further studies are required to enhance the formulation and the spray concentration; moreover, this protein bait can be used in combination with attract-and-kill devices.
Acknowledgments The authors thank the International Atomic Energy Agency Research (Grant no. 16015) for funding to C.Y.N. and the Plant Health & Environment Department of INRA for funding to N.D.
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