Settling, Oviposition and Reproduction Response of Peach Fruit Fly, Bactrocera zonata (Saunders) to the Plant Extracts of Native Species of Kalat Division in Balochistan

1Department of Agriculture Extension, Kalat, Balochistan, Pakistan 2Department of Entomology, Sindh Agriculture University, Tandojam, Hyderabad, Pakistan 3Department of Plant Sciences, Sardar Bahadur Khan (SBK) Women’s University Quetta, Pakistan 4Department of Entomology, Faculty of Agriculture, Lasbela University of Agriculture, Water & Marine Sciences (LUAWMS), Lasbela, Pakistan Article Information Received 04 June 2021 Revised 15 September 2022 Accepted 08 October 2022 Available online 23 December 2022 (early access)


INTRODUCTION
T he demand for fruits has increased in many folds especially in the form of canned or fresh fruits due to increasing population. However, disease and insect pest problems affect both quality and quantity of fruits. Among insect pest fruit flies of family Tephritidae (Diptera) are most harmful insect pests of fruits and vegetables. About 4000 species of fruit flies were reported out of which 700 species found throughout the world (Fletcher, 1987). Approximately, 250 species of Bactrocera fruit flies are widely found in most tropical and temperate areas throughout the world. Nearly, 11 fruit fly species including B. cucurbita, B. zonata and B. dorsalis are examined to cause losses of fruit and vegetable production in Pakistan.
Peach fruit fly is one of the most harmful species of Tephritidae. It is a serious pest of peach and custard apple in neighboring country (Butani, 1976;Grewal and Malhi, 1987), as well as guava and mango in Pakistan (Syed et al., 1970). It is a polyphagous species attacking some 40 species of fruit and vegetables (White and Elson-Harris, 1992).
The presence of male adults of B. zonata in the orchards of mango and guava was about 74.66% and O n l i n e

F i r s t A r t i c l e
46.62%, respectively in Pakistan (Khan et al., 2005). In semi-desert and coastal areas of Balochistan and Punjab the fruit fly B. zonata has been found as a serious threat for peach orchards, although these pests were rarely found in hilly areas of Pakistan (Marwat et al., 1992;Sarwar, 2006). There is urgent need to adopt eco-friendly technologies such as Integrated Pest Management to enhance productivity of fruits and vegetables. Number of control methods used to reduce damage of the fruit flies to vegetation. For this purpose, many cultural, biological, chemical practices including insecticidal and pesticide sprays have been used. The pesticide and insecticide sprays are able to kill the larvae of fruit flies and other insects to some extent but it affects the quality and taste of the fruits. Additionally, chemical sprays also affect environment and surrounding vegetation. Due to lack of awareness, local farmers use many insecticide sprays without knowing the effect of those sprays, which further causes toxic effects to environment and fruit quality.
A wide range of natural botanical extracts against plant diseases (Matrose et al., 2020;Shamsullah et al., 2020) and insect pests (Schmutterer and Singh, 2002) have become popular to achieve their environment friendly management. These plant extracts have been used against peach fruit fly (Siddiqi et al., 2011;Ilyas et al., 2017) as well as other fruit flies (Riaz et al., 2018;Ugwu et al., 2021).
Among natural insect repellents, neem has a great commercial potential. It has an advantage over synthetic insecticides in a way that it has no or less toxic effect on man, animals and useful insects (Schumutterrer, 1985). As many as 540 insect species including all key agriculture insect pests have already been reported to be vulnerable and exhibit various behavioral and physiological effects of neem (Schmutterer and Singh, 2002). Neem plant extract affects the insect potency, long life and productivity (Arora and Dhaliwal, 1994). The secondary metabolites present in neem extract prevent the growth and productivity to insects (Prakash and Rao, 1997). Different experiments proved that the neem extracts are the best sprays to check the insects and pests. However, little is known about the efficacy of similar insect repelling properties of Zataria multiflora (Karim and Yousefi, 2013) and Achillea santolina (Yonus et al., 2016).
The upper and lower highlands of Kalat division in Balochistan have a diverse resource of endemic plants with ethnomedicinal properties including Z. multiflora and A. santolina (Tareen et al., 2010;Bibi et al., 2016), which can also be explored for their potential to coup with agricultural insect pests and diseases. Therefore, present research work was carried out to determine the effect of neem along with two other plant extracts, with similar properties, on settling and reproduction response of B. zonata peach fruit fly.

Laboratory studies
This study was undertaken in the Department of Entomology, Faculty of Agriculture, Lasbela University of Agriculture, Water and Marine Sciences, Lasbela, Pakistan. The experiment, to study the potential of different plant extracts at different doses, included two factors; different plant extracts (with 3 levels) and concentrations (with 4 levels including control) and each treatment factor was replicated four times.
Water extract of 'Neem' seed (Azadirachta indica A. Juss.) Meliaceae, stem and leaves extract of 'Izghand' (Zataria multiflora Boiss.) Lamiaceae, and extract from leaves and flower of 'Boh-e-Madran' (Achillea santolina L.) Asteraceae, were used in this study. These plant materials were brought from Ayurvedic shop and ground to fine powder in a grinding machine. From each plant sample, 30 g of powder was added in 500 ml of water and shaken after 24 h to mix evenly. These samples were used at 0.5, 1 and 2% concentrations in laboratory experiments. The culture of B. zonata was maintained under controlled laboratory conditions at 28 ± 2°C and 60 ± 5% R.H.

Effect of plant extract on the oviposition of peach fruit fly
The water extracts of the plants were used to determine its effect on oviposition of fruit fly. Extracts were mixed with chikoo juice at 0.5%, 1 and 2 concentrations. This treated juice was applied inside plastic glasses having pin-holes all around the wall of glass. The glasses applied with treated juice were exposed to 15 days old 20 fruit fly adults (mixed population) for 24 h for egg laying in the pin-holes. Untreated chikoo juice was used as control. The eggs laid in different treatment were collected separately with a camel hairbrush in water filled petri dishes. The number of eggs laid in different treatments were compared for determining the effect of extracts.

Effect of plant extracts on the progeny of peach fruit fly
Water extract of A. indica, Z. multiflora, and A. santolina were mixed with the diet of fruit fly at 0.5, 1 and 2 concentrations. The flies were separately offered untreated chikoo fruit for oviposition for three days. Five pairs of fruit flies were fed with the treated and control (untreated) diet at each concentration for 15 days. The fruits were labelled and then kept in glass jars having oneinch layer of sand at the bottom. After 15 days, number of pupae were counted and emerged adults were also counted and compared.

Settling and post settling reproduction response of peach fruit fly on chikoo fruit treated with different plant extracts
For the purpose, individual fruits were treated by dipping them in various concentrations of the plant extracts and were air dried in the laboratory. Untreated fruits were used as control. These treated fruits were placed in rearing cages. About 20 flies were introduced in the cages and their settling response was recorded after each 6 h for three days. Development of pupae and emergence of adults were also recorded from the fruits exposed to treatments.

Statistical analysis
The collected data was statistically analyzed using MSTAT-C software and means were compared by Duncan's Multiple Range Tests (DMRT) at P ≤ 0.05.

Effect on the oviposition
The results showed that exposure of fruit flies to the water extracts of tested plants, reduced the number of eggs laid. It indicates that these plant extracts acted as oviposition deterrents against fruit fly. The interaction effect of plant extracts with its doses was non-significant. However, various doses of these extracts had significant difference on reducing oviposition of peach fruit fly B. zonata (Table 1a). Minimum significant (average 8 number of eggs) oviposition was at 2% concentration. Comparatively, the treatment of three plant extracts did not exhibit any significant variance on the number of eggs laid. Table 1b shows the number of pupae developed from the eggs laid by female peach fruit flies fed on diet treated with different concentrations of plants extracts. An interaction of plants extracts and their concentration, applied to artificial diet of peach fruit flies, did not show any significant effect. However, lowest number of pupae 18.25 recovered at 2% concentration of A. indica extract which were lower than 1, 0.5% and control of same plant extract but pupae recovered from controls were significantly higher than those recovered at all other concentrations of plant extracts. Similarly, 22.50 in Z. multiflora and 20.50 in A. santolina extract at 2% concentration were lower than those of all concentrations and their controls. The comparison of plants extract shows that there was no significant difference in mean number of pupae recovered from all three extracts. However, the different concentrations of plants extract showed significant difference that the significantly lowest number of 20.42 pupae were recovered at highest concentration. Table 1c shows the number of adults emerged from the developed pupae. The plant extracts alone and its interaction with its various concentrations had no significant difference on the number of adults emerged from the developed pupae. In contrast, the concentration factor had significant effect on the number of adults emerge from developed pupae. Minimum significant adults 15.25 emerged at 2% followed by 23.50 and 28.50 at 1 and 0.5%, respectively. Table II shows the number of fruit flies settled on chikoo fruits treated with various concentrations of plant extracts after given age intervals. Chikoo fruits were dipped in different concentration of plants extract, dried under shade and exposed to fruit fly in order to determine the settling response of fruit fly. Overall, there was no significant difference in interaction effect of plant extracts, its doses and age intervals. However, the lowest settling observed was 0.25 flies settled fruit -1 after 18 h at 0.5, 1 and 2% concentrations of A. indica and Z. multiflora extracts and while at same time interval in control treatments the settled flies were 1.5 and 2.0 fruit -1 , respectively which were higher than those treated. This lowest value was also observed after 36 and 54 h at 2% concentration of A. Indica and after 36 h at 1% concentration of Z. multiflora.

Effect on settling response of peach fruit fly
Similarly, lowest settling response was observed 0.25 flies settled fruit -1 after 18 and 36 h at 2% concentration in A. santolina extract. The highest settling response on treated chikoo fruits was observed after 42 h at 0.5% in Z. multiflora (2.25 flies) which was higher than 2 and 1% concentrations. Moreover, the values of control were higher than values of each concentration showing 3.0 flies after 24 h in the control of Z. multiflora (Table II).
Unlike interaction of all three factors, settling response was significantly different at different concentrations. Minimum settling was observed at 2% and maximum settling was observed at control treatment (Table II). However, the interaction of different concentration of plant extract with various time intervals was non-significant by DMRT at P ≤ 0.05 (Table IIIb).
In accordance with concentration factor, time interval also showed a significant effect by DMRT at P ≤ 0.05 on settling response of peach fruit fly in this experiment (Table  II). Significantly lowest number of flies 0.58 and 0.60 settled on fruits after 18 and 36 h, respectively. Whereas, highest significant number of flies 1.73 settled after 42 h. However, the interaction of various time intervals with different plant extract had no significant effect on settling response of peach fruit flies (Table IIIa).  Accordingly, the interaction of different plant extracts with its various concentrations had no significant difference on settling response of fruit fly (Table 1d). Nevertheless, 2% concentration of A. indica and Z. multiflora showed lower number of fruit flies settled than at 0.5, 1% and control. The comparison of the effects of plant extracts on mean settling response showed no significant difference, however different doses of plant extracts had a significant difference on settling response of fruit fly, showing inverse trend to the increasing concentration of extracts. The lowest number of 0.60 fruit fly fruit -1 settled at 2% concentration fallowed by 0.73 at 1% and 0.87 at 0.5% dose as compared to 1.64 in control, which were significantly highest. Table 1e shows the number of pupae developed after settling on the fruits treated with different concentrations of plant extracts. As the fruit flies lay eggs beneath the skin of fruit and counting of eggs is not possible, therefore, the oviposition response of the test insect would either be noted by counting the larvae in the infested fruit by dissecting them or by counting the number of pupae recovered. The results indicate that both different plant extracts and its various doses alone had significant effect on the number of pupae developed on the fruits treated but their interaction had no significant difference. In case of A. indica seed extract, the lowest number of 22.00 pupae were recovered from 2% and most pupae 56.50 were recovered in control. Similar trend was seen while treated with other two plant extracts. However, plants extract results show that significantly lowest number of pupae 35.44 recovered from A. indica seed extract treated fruits followed by 37.50 from Z. multiflora and 43.50 from A. santolina extract. Likewise, at various doses lowest significant number of pupae 25.92 were recovered from the fruits treated with 2% concentration which was significantly lower than those 54.08 recovered at control. Table 1f shows the number of adults emerged from pupae developed after settling on the fruits treated with different concentrations of plant extracts. The results indicate that both factors plant extracts and its various doses had significant effect on the number of adults emerged from collected pupae on the fruits treated but the interaction of these two factors had no significant difference (

DISCUSSION
Neem contains an array of chemicals having different complex mode of action on insects. Azadirachtin, a chemical compound from the neem seeds, is the main component responsible for the toxic effects in insects. Azadirachtin from neem effects insects in a variety of different ways: as an antifeedant, insect growth regulator and sterilant (Mordue and Nisbet, 2000).
Results of our study are in accordance with a study which implies neem seed kernel extract, for oriental fruit fly, as an ovipositional deterrent (Chen et al., 1996). Similarly, neem seed kernel extract had effect on the fecundity and post-embryonic development of fruit flies B. cucurbitae and B. dorsalis (Singh, 2003). This idea was further supported the ingestion of neem can significantly reduce the longevity and fertility of melon fly O n l i n e F i r s t A r t i c l e N. Shah et al. and the oriental fruit fly (Khan et al., 2007). Accordingly, significant repellency of the Mexican fruit fly was observed in terms of oviposition at 3 and 5% aqueous neem extract and 4.5% neem oil treatment (Valencia-Botin et al., 2004). Lower number of B. zonata adults settled on fruits treated with petroleum ether extract of sweet flag followed by neem acetone extract and sweet flag extract. The number of pupae and adults obtained from fruits decreased with increase in the extract dosage (Naheed et al., 2004). Effect of crud extracts of A. santolina determined for feeding toxicity against adult of Oryzaephilus surinamensis. The results of the study showed that Hexane extract of A. santolina was effective reaching survival rate of adult to 66.2% at concentration of 10% (Yonus et al., 2016). Antifeedant activity of Z. maltiflora oil was reported to be more effective on feeding deterrence index than Thymus daenensis (Karim and Yousefi, 2013).
Looking at the settling response our findings agreed with the similar study, where settling of melon fruit fly was adversely affected by neem seed water extract and neem oil (Khattak et al., 2009). The study further reported that significantly lower number of pupae was recovered from the fruit treated with 2% concentration of neem oil and 3% of neem seed water extract as compared to control.

CONCLUSION
All the plant extracts exhibited insecticidal properties and resulted in reduction of fruit fly population in laboratory condition and decreasing trend of fruit fly population continued with increasing concentration. A. indica showed most promising results followed by A. santolina. These plants studied in this research work, contain an array of chemicals having different complex mode of action on insects, hence further investigation is needed to evolve strategies for integrated pest management techniques. Furthermore, the increasing dosage of these plant extracts had increasing degree of fruit fly repellence. Hence, more research is needed to standardize the dosage. Moreover, these findings are based on laboratory trials and may differ in field trials due to the various biotic and abiotic conditions in different agro-ecological zones.

O n l i n e F i r s t A r t i c l e
Response of Peach Fruit Fly to the Plant Extracts