Laboratory test of the Dirhinus giffardii (Silvestri) (Hymenoptera: Chalcididae) against the pupae of Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae)

Fruit flies being serious pest of fruits and vegetables are responsible for huge economic losses in the world. Pupal parasitoid, Dirhinus giffardii is one of the most significant biological control agents that has been used effectively in controlling the fruit flies. Laboratory studies were conducted to investigate the parasitism, emergence and post emergence sex ratio of D. giffardii through different exposure times and parasitoid density against the pupae of melon fruit fly, Bactrocera cucurbitae. The studies manifested that the exposure time and parasitoid density had a significant effect on the parasitism and emergence ratio of parasitoids. Results revealed that mean rate of parasitism (45.66) was the highest in case of five pair of parasitoids and mean rate of emergence (42.66) of D. giffardii was also the highest on exposure period of six days. Further, it was observed that, exposure time and parasitoid density had no significant effect on the post emergence sex ratio of male and female parasitoids. The mean per female parasitism was increasing with the increase in number of pairs of D. giffardii and reached to its peak after exposure time of six days. These findings suggest that D. giffardii has a great parasitizing potential against the pupae of B. cucurbitae and can effectively suppress its populations.


Introduction
The melon fruit fly, Bactrocera cucurbitae (Coquillett) is the most damaging pest of vegetables. It is distributed all over the world but mostly cause significant damage in Asian countries like China, Pakistan, India, New Guinea, Nepal, Hawaiian Islands and Philippines [1]. According to a survey B. cucurbitae harms over eighty one host plants, however it is the main pest of cucurbitaceous vegetables [2]. In general, the fruit flies cause massive losses to fruits and vegetables in the entire world and are known as key insect pest of the horticultural fruit plants. Approximately, they have 4,000 species which are well identified in all over the world [3]. In Pakistan, the B. cucurbitae causes 50-90% losses in fruits like melon, guava, citrus, mango and cucurbits [4]. Melon fruit fly can also injure non-cucurbit host plants by damaging their stem, flowers and root tissues [5]. The adult melon fruit fly lay 300-1000 eggs, during egg laying process ovipositor pierces the skin of fruits, after the eggs hatch, maggots begin to start feeding on the pulp of fruits by making tunnels inside the fruits [6]. During feeding, maggots consume food to store energy for the upcoming pupal stage which cause post-harvest losses and due to this reason market value of fruits become reduced [7]. In general, farmers use enormous amount of pesticides for the suppression of melon fruit fly which is declared as a quarantine pest. Agrochemicals and insecticides are notorious elements that become the prominent fragment of worldwide agriculture systems from the last century. Pesticidal leftovers did scatter in the environment, causing shocking defacement of land ecosystems and contaminating the human foods [8,9]. The human health is on real threat and facing certain difficult problems because in our agro-ecosystem, agrochemicals deposits are present at every time [10]. In order to control pests, growers adopt the habit of frequent and unjudicious application of pesticides as a result, insect pests develop resistance that make vulnerable their competence and also depreciate the environment [11]. Different human fitness related concerns are correlated with pesticides, such as cancers, nausea, headaches, birth defects, endocrine disruption and infertility [12]. Particularly due to constant exposure of pesticides children health is at more risk [13].
Therefore, reduce fruit fly losses by using non-chemical methods will be implemented. There are some substitutes to pesticides that do not cause the same environmental problems. One of these replacements include biological control methods which provide one of the most effective, obviously safe, and feasible mechanisms against insect pests [14]. By using biological control tactic, living organisim such as parasite, predator and disease causing organism is being introduced in to the environment of pests to reduce their population. Parasitoid, Dirhinus giffardii (Silvestri) (Hymenoptera: Chalcididae) is a useful biological control agent which can be used as a surrogate of pesticides hazards to lessening the population of fruit flies [15]

Rearing of Bactrocera cucurbitae
The fruit flies, B. cucurbitae were mass reared on pumpkin and artificial diet containing wheat bran (26%), sugar (12%), dried troula yeast (3.6%), Sodium benzoate (0.1%), Methyl-p-hydroxybenzoate (0.1%) and water (58%) [22]. Eggs of fruit flies were placed directly on the diet trays having artificial diet. These eggs were collected in plastic glasses having 0.5 mm holes around them smeared internally with guava juice and put in adult fruit fly cages. The hatched larvae feed on the diet till complete maturation. After the full fed larvae jumped out of the trays and fell on the substrate (sand/saw dust) for pupation, the pupae were collected through sieving and used for maintaining the culture and experiments. The adult fruit flies were provided protein hydrolysate and sugar.

Rearing of Dirhinus giffardii
The colony of parasitoids D. giffardii being well maintained at (NIA, Tando jam) biocontrol agents rearing lab from the last several years. Parasitoids were reared in glass cages on pupae of B. cucurbitae and artificial diet, a fresh diet solution (30% honey and 70% water) were offered to the parasitoids through soaked cotton wigs which were impregnated with honey and water.

Data analysis
After mentioned exposure periods rate of parasitism, sex ratio and parasitoids emergence was recorded and results were analyzed by using the software statistics 8.1. One-way analysis of variance (ANOVA) and Tukey's honestly significant difference (HSD) test were used for comparisons of means among different treatments.

Results
The investigations on pupae of melon fruit fly (B. cucurbitae) were conducted by using various exposure periods and parasitoid pairs in order to check the parasitism, emergence and post emergence sex ratio of D. giffardii. The results obtained from the experiments showed that after the exposure time of 24 hour, the maximum mean parasitism of 24.33±1.20 was recorded by using 5 pairs of D. giffardii followed by 17     The relationship between parasitization and emergence ratio of parasitoids strongly supports the phenomena that as we increase the parasitoid pairs, the number of emerged parasitoids also increase. It was observed that maximum mean emergence was in case of five pair of parasitoids while exposure time also had significant effect on the emergence of D. giffardii. Results revealed that based on exposure periods, the rate of mean parasitoid emergence trend was observed in following order 6 day>5 day>4 day>3 day>2 day> and 1 day, respectively.
The trend of post emergence sex ratio depicted that male and female emerged parasitoids were not in equal numbers and exposure time and number of parasitoids have no significant effect on them. Some factors are known to influence the sex ratio of parasitoid progeny, such as parental sex ratio, host size, and host age, while parasitoids (Itoplectis naranyae and Pimpla nipponica) could change the sex ratio of their offspring in response to host age [27][28][29]. In contrast [30] reported that the age of the host had no effect on the progeny sex ratio of the wasp Brachymeria lasus. In our study, we found that the male and female emerged parasitoids were almost in different proportion but in some observations males were dominant regarding sex ratio [31]. Further, it was concluded that number of parasitoids, exposure time and per female parasitism significantly interlinked with each other (Fig.  1).

Conclusion
In conclusion, parasitism rate and emergence percentage of the pupal parasitoid, D. giffardii was satisfactory on the host (Melon fruit fly pupae). The results suggest that D. giffardii could be a suitable candidate for the biological control of B. cucurbitae. The present study will also be helpful in reduction of the fruit flies population through mass production and release of pupal parasitoid D. giffardii.