Ihsan ul Haq (corresponding author), Insect Pest Control Laboratory, FAO/IAEA Agriculture and Biotechnology Laboratories A-2444, Seibersdorf, Austria. E-mail: email@example.com; firstname.lastname@example.org
Enhanced male melon fly, Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae) mating success due to application of a juvenile hormone analogue, methoprene (M+) and feeding on a sugar–protein diet (P+) (hydrolysed yeast), supports their incorporation into operational area-wide programmes integrating the sterile insect technique. But enhanced mating success may have a cost in terms of shortened sterile male longevity as has been reported under certain situations for other tephritid pest species. The objective of this study was to analyse the effect of methoprene treatment and addition of protein to the adult diet and then switching from a sugar–protein diet to sugar only (dietary switched after first 3 days; DS), on male longevity in laboratory. The treatments were as follows: (i) M-P+; (ii) M-P+ DS; (iii) M+P+ DS; and (iv) M-P- (fed only sugar and water continuously since emergence). All treatments were significantly different from each other in their survival response in the following order, M-P+ > M+P+ DS > M-P+ DS > M-P-. Access to sugar–protein diet and then switching after 3 days had no negative effect on male longevity compared with males fed only sugar throughout. The results are discussed in the context of methoprene treatment and a sugar–protein diet as a pre-release feeding protocol for maturing sterile flies in fly emergence/release facilities.
The sterile insect technique (SIT), applied as a component of area-wide integrated pest management, is a well established environment-friendly technique used for melon fly suppression (Jang et al. 2008) or eradication (Kakinohana 1994). For successful SIT application, the sterile mass-reared males must be able to survive in the nature to compete with wild males (Hendrichs et al. 2002). Diet supplementation with protein can increase the male competitiveness but may have a cost in terms of shortened male longevity.
Feeding on protein diet has a variable effect on survival in different tephritid species. Carey et al. (1999) found higher mortality rates in Ceratitis capitata males first provided with protein in their adult diet and then deprived of it than in males only with access to sugar. Some other studies in C. capitata showed that protein-fed males died significantly faster than only sugar-fed males when deprived of food after releasing them in field cages (Kaspi and Yuval 2000; Maor et al. 2004). However, other studies showed no negative effect of adding protein to the diet on C. capitata male short-term survival (2–4 days) in field cages (Shelly and Kennelly 2002; Shelly and McInnis 2003). Similarly, Haq et al. (2013) also found no adverse effect on starvation survival in protein-fed Bactrocera cucurbitae males when compared with protein-deprived males. On the other hand, a positive effect of dietary protein in terms of increased longevity was observed in Bactrocera tryoni (Froggatt) (Pérez-Staples et al. 2008) and in Anastrepha serpentina (Wiedemann) (Jácome et al. 1995).
In operational SIT programme releases, ideally the sterile flies should be kept in the fly emergence and release facility until close to sexual maturity (particularly in male-only genetic sexing strains). Conversely, if sterile flies are released before they attain sexual maturity, many males may die before being able to mate with wild females.
Recent studies on B. cucurbitae showed that combining the application of juvenile hormone analogue methoprene with access to a diet including hydrolysed yeast, accelerated male sexual maturity with males reaching sexual maturity at 5 days of age compared with 14 days in control, while methoprene application to only sugar-fed males had no effect on maturity acceleration (Haq et al. 2010). Application of methoprene to males with access to hydrolysed yeast had no adverse effect on male starvation survival (Haq et al. 2013). Nevertheless, the deprivation in these studies was quite drastic since it included not only all food but also water. However assuming that released flies also will not be able to find water after release does not apply in many field situations, where sterile flies readily forage for water.
The objective of the current study was therefore to compare the survivability of methoprene-treated vs. untreated males switched to only sugar and water after 3 days of feeding on a diet including hydrolysed yeast, and comparing them with males continuously fed a diet including hydrolysed yeast or sugar and water only.
Materials and Methods
The maintenance of the B. cucurbitae genetic sexing strain (McInnis et al. 2004) and the application of methoprene follow the methodology of earlier studies (Haq et al. 2010). The whole laboratory experiment was run in a single room with a photoperiod of 14L : 10D at 24 ± 1°C, and 60 ± 5% RH. The males were separated by sex at the pupal stage by separating the brown male pupae from the white female pupae. The males only status was insured by removing any female emerged from the brown pupae within 1 day after emergence. The males were allowed access to water and to feed on sugar only or a diet consisting of hydrolysed yeast (MP Biomedicals LLC, Solon, OH; http://www.mpbio.com) and sugar (1 : 3) [even though hydrolysed yeast contains also minerals and vitamins besides amino acids, the major constituent is protein (60%), so hereafter called sugar–protein diet]. The males fed on the sugar–protein diet either continuously or switched (dietary switch; DS) to sugar and water only. The treatments were as follows: (i) Males not methoprene treated, fed the sugar–protein diet and water continuously (M-P+); (ii) Males not methoprene treated, fed the sugar–protein diet for the first 3 days and then switched to sugar and water only (M-P+ DS); (iii) Males treated with methoprene, fed the sugar–protein diet for the first 3 days and then switched to sugar and water only (M+P+ DS); (iv) Males not methoprene treated, fed only sugar and water continuously since emergence (M-P-).
Four hundred males, one hundred from each treatment, were allowed to feed for 3 days in five 12 × 10 × 20 cm Plexiglass screened cages (20 males per cage) and were then transferred to similar sized clean cages and supplied with respective diets ad libitum and water. To minimize the effect of microbial growth in the different types of diets and their effect on longevity, the diet was replaced every 7 days, and water was replenished as necessary. Each cage contained a strip of tent-shaped filter paper to serve as a lek site. Five replications each involving 20 males from each treatment were tested for their survival. The experiment was stopped when all males in three out of the four treatments of males reached 100% mortality (day 70). Mortality was recorded each morning at the same time (5 days a week) and dead males were removed after every observation. The Cox regression model was fitted to analyse the variance in survival response at 95% confidence intervals and complementary pair-wise comparisons were run by Kaplan–Meier test by using R software (http://www.r-project.org).
The cumulative survival curves over time are shown in (fig. 1). The Cox regression hazard model showed that all treatments were significantly different from each other (χ2 = 228.4, d.f. = 3, P < 0.001). The pair-wise comparisons among treatments showed significant differences in male survival in the following order [M-P+ > M+P+ DS (Z = 8.91, P < 0.001) > M-P+ DS (Z = 3.18, P < 0.001) > M-P- (Z = 3.11, P < 0.001)]. The M-P+ males showed maximum median longevity (median value +/- interquartile range; 70+/− 70–55) followed by M+P+ DS (56 + /− 61–52.25) and M-P+ DS (53+/− 59.25–46) males, while M-P- (37.5+/− 53–32) males had the lowest longevity.
Bactrocera cucurbitae males feeding on the sugar–protein diet throughout (M-P+) showed highest survival compared to only sugar-fed males (M-P-) and to both types of dietary switched males (M+P+ DS, M-P+ DS). This response is similar to that of C. capitata males under a similar feeding protocol (Barry et al. 2007). But it differs from the findings of Davies et al. (2005), who observed no difference in survival between C. capitata males fed on the sugar–protein diet and those fed only on sugar.
This study has implications for SIT application in area-wide control programmes against B. cucurbitae. The advantages of M+P+ treatment in terms of accelerated sexual maturity and enhanced mating success of B. cucurbitae males (Haq et al. 2010), as well as the significant positive effects of these treatments on male longevity, support their application as pre-release treatments to teneral sterile males. In the successful SIT programme in Japan, the mass-reared sterile B. cucurbitae flies (both sexes) were fed only on sugar and water and kept after emergence in the release facility only for 3 days (Nakamori and Kuba 1990). Our study, however, suggests that feeding sterile males the sugar–protein diet for 3 days before release may even increase their longevity, particularly in cases where nitrogenous sources are scarce in nature. Furthermore, the finding that methoprene-treated males showed higher survival than non-treated males confirms the benefit of methoprene application in combination with feeding on the sugar–protein diet as pre-release treatments.
Barry et al. (2007) concluded that the post-release food foraging ability of C. capitata sterile males will have a larger effect on sterile male survivorship than the pre-release diet. Post-release food foraging capacity of sterile males remains of course important (Maor et al. 2004), however, unlike in C. capitata males, the chief effect of the M+P+ treatment in B. cucurbitae males is the accelerated sexual maturation, shortening their long pre-maturation period (Haq et al. 2010). Mass reared C. capitata males mature faster (4–6 days), and methoprene application has no significant effect on accelerating male maturity (Faria et al. 2008). Thus, a pre-release protocol M+P+ may contribute significantly to the effectiveness of the SIT for B. cucurbitae, but less so in C. capitata, where only the sugar–protein diet is sufficient to increase sterile male performance (Yuval et al. 2007). However, this B. cucurbitae study was done in small cages under laboratory conditions. Similar studies in field cages should be carried to decide on pre-release treatments for sterile males to be used in SIT programmes.
We are thankful to Todd Shelly (APHIS, USDA) and two anonymous referees for critical reviews of an earlier version of this manuscript. We also thank David Soriano for advice in statistical analysis.
This article was published online on 10 March 2011. The reference for Haq et al. (2013) has now been updated to show the correct citation details for J. Appl. Entomol. Vol. 137, Suppl. 1.