A comparison of vertical and horizontal trap orientations for attracting male Bactrocera spp. as fruit fly pest

Fruit flies, such as Bactrocera spp., are significant agricultural pests focusing on crop yields in Indonesia. One way of environmentally friendly control is the application of attractants to the trap. This study aims to ascertain the trap model that efficiently captures Bactrocera spp. This survey was conducted in Malang’s Dau Subdistrict between December 2022 to February 2023. The traps used were modified Steiner traps, namely plastic bottle traps with vertical and horizontal models. Any traps were baited with 0.125 milliliters of methyl eugenol, which was dripped on cotton and suspended inside the trap. To determine the effective trap model for capturing fruit flies, the study randomly installed vertical (3 pieces) and horizontal (3 pieces) traps in each citrus orchard. The findings within the Friedman testing analysis suggested that the attraction of fruit flies to vertical and horizontal trap models had significant differences. Horizontal traps had 58% effectiveness compared to vertical traps. The types of fruit flies in the vertical model are Bactrocera dorsalis, B. carambolae, and B. umbrosa. Meanwhile, in the horizontal model, B. umbrosa was absent. Modifying this trap model increases the ability as a potential fruit fly trap and a fruit fly pest monitoring tool.


Introduction
The taxonomic classification of Bactrocera, belonging to the order Diptera and family Tephritidae, encompasses approximately 500 species, with around 4000 species being identified currently.These species are predominantly found in New-east Asia and the Southern Ocean islands, exhibiting a significant level of ecological endemism [1].A large number of species occur in subtropical or tropical rainforests and are not considered a threat to other species.Approximately 70 species of Bactrocera pose a significant threat to global trade and agricultural productivity due to their larval stage infestation of crucial fruit and vegetable crops.The severity of the impact varies depending on factors such as the 2 specific Tephritidae species, the type of fruit crop or plant-based affected, and the area of concern [2][3][4][5][6][7][8].Although the geographic distribution of most Bactrocera species is restricted, several species exhibit invasive characteristics, prompting numerous countries to implement continuous trapping initiatives with the purpose of detecting infestations.
One way to control fruit flies is male culling techniques (MAT) [9][10].Heavy traps with attractants are one of the environmentally friendly controls that can be used to control fruit flies.Attractants can lure male fruit flies into the trap, monitor fruit fly populations, and disrupt fruit fly mating activities [11].This specific method has proven its most efficacy in populations that exhibit restricted distribution, and it is widely employed in fruit fly eradication initiatives on a global scale.To attract and control male pests, bait and insecticide formulations can be strategically positioned within traps or selectively distributed at specific locations; those items include lampposts, phone posts, and highway plantations.The selection of the MAT execution mechanism is typically influenced by factors such as the specific species being targeted, the geographical and terrain characteristics of the area, and the available resources.The application referred to as Sterile Insect Technique (SIT) can be advantageous in "areawide" initiatives when the complete eradication of fruit flies is unattainable.[12].This method is a constituent of an Integrated Pest Management, or IPM strategy, which integrates several methods, such as technical, mechanical, chemical, and biological controls [13].
Methyl eugenol (ME), also referred to as 4-Allyl-1,2-dimethoxybenzene, represents a highly effective bait.Furthermore, it has the ability to elicit compulsive feeding behavior in male fruit flies [14,15].Although methyl eugenol ether (ME) is not a pheromone, the mode of action of ME is almost similar to that of pheromones, which attracts fruit fly images so that the male fruit fly population is reduced, leading to decreased mating frequency.Methyl eugenol (ME) is an organic substance that occurs naturally in an extensive list of plant species totaling approximately 450, belonging to 80 families across 38 distinct orders.These plants are primarily found in tropical regions [16].Methyl eugenol, commercialized under the trademark petrogenol, is a frequently used attractant by citrus farmers, particularly in the Dau District of Malang Regency.In addition to its user-friendly nature, the product's accessibility is facilitated by its widespread availability in the market.An additional factor to consider is the significant volatility of this male bait, its efficacy in modest quantities, and its compatibility with simple trapping mechanisms.
An affordable and straightforward trap design that achieves widespread use among farmers in Indonesia is a modified Steiner trap built with plastic bottles.So far, the traps used by citrus farmers, especially in the Dau Sub-district, Malang Regency, are vertical.In interviews with local farmers, they stated that many fruit flies find it challenging to enter the trap even though it has been given quite a lot of holes in the neck of the bottle so that the trapped male fruit flies are few.This study aims to modify the trap model and compare its effectiveness so that the orientation of this fruit fly trap becomes potential and can be used as a monitoring tool for fruit fly pests.

Ethics statement
An experiment was undertaken at the Biology Department.The Department of Biology, Faculty of Mathematics and Natural Science, Brawijaya University, Indonesia, does not require ethical approval for manipulation and handling because B. dorsalis, B. carambolae, and B. umbrosa are insect pests.They are neither classified as endangered nor afforded legal protection-Brawijaya University's approved protocol conducted all procedures in this study.All of the experimental procedures involving pests were conducted by the Department of Plant Protection, Faculty of Agriculture, guidelines of Gadjah Mada University, Yogyakarta, Indonesia, and approved by the Department of Biology, Faculty of Mathematics and Natural Science, Brawijaya University, Indonesia.

Site of Interest
The research has been conducted in two orchards in Banjartengah sub-village, Sumbersekar village, Dau Sub-District, Malang Regency, from December 2022 to February 2023.The first citrus orchard is located at coordinates S07 o 55.115' E112 o 34.037', while the second is at S07 o 55.134' E112 o 34.160' (Figure 1).

Trap Making
Two types of traps were used in this study, namely vertical and horizontal trap models.These two traps modified the Steiner trap model and were adjusted for placement at the edge of each orchard.In each orchard, six traps (three vertical models and three horizontal models) were installed in a row at the edge of the orchards are evenly distributed around ten meters apart in and out of traps.
The vertical model traps were constructed using plastic bottles with a capacity of 1500 ml.At the top of the bottle body, there are three rows of holes around the bottle body in a zigzag pattern with a diameter of r 0.7 cm as a place for fruit flies to enter.A 50 cm long galvanized zinc wire was inserted through the bottle cap that had been given a hole.A cylindrical cotton roll, measuring 1 cm in diameter, was coated with 0.125 ml of methyl eugenol, commercialized as petrogenol.The cotton roll was placed carefully to facilitate the dispersion of its flavor, allowing fruit flies to perceive and detect it.The cotton was then tied to the end of a 2.10 mm diameter wire parallel to the three rows of holes.The established trap bottles were filled with 200 milliliters of distilled water and 5 ml of 1% formalin before installation.An essential component of monitoring and controlling fruit fly pest species is using male fruit fly attractants, such as Methyl Eugenol.Moreover, utilizing these lures is essential for implementing pest management strategies involving male culling methods and detecting invasive pest populations using traps [17][18].The trap model is presented in Figure 2a.In the second model, horizontal traps were also made from plastic bottles with a size of 1500 ml.The neck of the bottle was cut to 10 cm, and the mouth was inserted into the other cut bottle.Then, both were glued using tape.Next, in the middle of the bottle, a hole measuring r 2.10 mm was given as a place to enter the galvanized zinc wire, and a cotton roll that had been dabbed with 0.125 ml of petrogenol was tied to the end of the wire.The opposite end of the rope was secured to a tree branch 1.5 to 2 meters off a b the ground.Before installation, 200 ml of distilled water and 5 ml of 1% formalin were added to the trap.Selection of trees using purposive sampling method, with lush branches and abundant citrus fruit production (Figure 2b).

Fruit fly observations
Observations of vertical and horizontal trapping of fruit flies were made every two weeks for four observation periods (8 weeks).Methyl eugenol (ME) was added weekly, while traps were replaced every two weeks.All captured fruit flies were collected in 100 ml zalp pots containing 70% alcohol and aerated.Fruit fly identification was performed morphologically using a trinocular stereo microscope and Nikon DS-Fi3 camera at the Ecology and Biodiversity Laboratory, Department of Biology, Brawijaya University.Based on morphological characteristics, observations were made regarding the type and number of captured male and female fruit flies.The Australian handbook is reviewed given that classifying [19].

Data analysis
The mean number of species in vertical and horizontal trap models measured fruit fly abundance.Vertical and horizontal trap models were analyzed using Friedman's non-parametric test with SPSS 21 software to determine the difference between the two orchards and the significance of fruit fly abundance.

Comparison of vertical and horizontal trap orientations
A contrast between the vertical and horizontal trap models in the two orchards showed different numbers of fruit flies between the two.In vertical traps, the species trapped were B. dorsalis with 137 individuals, B. carambolae with 79 individuals, and B. umbrosa with four individuals.While in horizontal traps, only two species were obtained, namely B. dorsalis, with as many as 178 individuals, and B. carambolae 138 individuals (Figure 3).The disparity in the number of fruit flies between the two orchards is not solely attributable to differences in biogeography, but it is also thought that the coevolution that takes place between fruit flies and their hosts assists in the formation of fruit fly classes [20].In the first orchard, the number of citrus trees as host plants is more numerous and denser than in the second orchard, affecting fruit flies' presence.The presence of plants can provide shade, nutrients, and hosts for the survival of fruit flies [21].B. dorsalis was the most common species of fruit insect among the two trap models.The potential of B. dorsalis to outcompete others resulted in this species being the most abundant in both orchards in terms of both male and female individuals.The species is highly polyphagous and is considered one of the world's most prominent invaders [22].B. dorsalis innate ability to fly, adaptability, and wide host range mean it can easily invade more significant habitats [23].This species is often cited as an essential host for invasive species in various agricultural fields [24].
Fruit flies' morphological traits, seen on the head, thorax, and abdomen, can be used to identify them (Figure 4).The thorax of B. dorsalis is black to brown, the tibia on the front and rear legs are dark, and the head features a noticeable mark.On terga 4 and 5, the abdomen can be seen to have a T line and a spot on the lateral side.Regarding the wings, it enters the tip of R 2 + 3 and has a line at the top with a small coastal band.A black thorax, a prominent spot on the head, and a little patch inside the post pronotal and surrounding the mental are the distinguishing features of B. carambolae.A black tibia on the legs, a box-shaped patch on the lateral side of tergum 4, and a distinct T line on the abdomen.This species' wings feature a costal band that stretches around R 4+5 and overlaps R2+3.The abdomen of B. umbrosa includes bands on the lateral sides and vertical lines extending from tergum 3 to 5. It also has a black thorax.The wings contain three bands that span them simultaneously [19].The horizontal trap model attracted 58% more fruit flies than the vertical trap model during the 8week collection period.The outcomes of the Friedman Test presented a substantial difference in fruit flies' attraction to vertical and horizontal trap models, as indicated by the Asymp.Sig value of 0.00 <0.05.The study's findings indicate that the horizontal trap contained more captured fruit flies.Thus, the horizontal trap model captures fruit flies better than the vertical trap.Based on observations, one of the causes of more fruit flies trapped in horizontal traps is that they are easier to enter through the mouth of the bottle.Although the number of holes is large in the vertical trap model, fruit flies have difficulty entering the holes provided, so most circle the bottle.In addition, the mouth of the bottle, which is quite large in the horizontal model, allows the aroma of methyl eugenol to invite more fruit flies to come to the trap.

Male fruit fly attraction
Treating methyl eugenol (ME) in vertical and horizontal trap models can attract male fruit fly individuals by 479 individuals or 88% more than females during the 8-week observation duration (Figure 5).ME is a volatile substance that evaporates and emits an aromatic odour when it does so.This substance is a food attractant or necessary for male-sexed fruit flies' reproduction [25].The chemical formula for methyl eugenol is C 12H2402.This substance is a hormone that is essential for the intake and mating behaviour of male fruit flies [26].Furthermore, the metabolic process occurs in the body of male fruit flies to produce attractant substances (sex pheromone).Male fruit flies will figure out the primary source of the scent and eat it if they detect the ME scent.The radius of the attractant and ME scent is up to 3 km.This substance is produced by exocrine glands and used by living things to identify their own species, other individuals, and groups and aid reproduction [27].In contrast to hormones, pheromones travel outside the body and can only affect and be recognized by individuals of the same species.Insect pheromones can be utilized in the management of insect pests both directly and indirectly, namely used in terms of monitoring insect pests (monitoring), mass trapping, mating disruption, or a combination of pheromones as attractants with insecticides or insect pathogens as killers [28].Based on the results of [29], the high catch of male fruit flies in ME traps can cause suppression or cut off the male fruit fly population in the orchard, which will significantly have a negative outcome on the regeneration of the following fruit insect population.

Host plant aviability
The availability of hosts impacts the abundance of species in fruit fly densities [30].The more availability of hosts in the orchards, the greater the possibility of increasing the population of fruit flies on these plants.Figure 4 shows the fluctuating number of fruit flies around the interval of the eight-week monitoring period.The prevalent pattern of expanding and decreasing fruit fly catches is related to fruit availability in the orchard.The high catch of fruit flies during the third collection period was due to the ripening phase of the citrus fruits.Although many fruit flies prefer ripe discards, some prefer rotten fruit.According to [31], decaying fruits attract female fruit flies to throw away eggs, resulting in an increase in the number of them within the plantation.
In addition, the high yield in the third observation was also related to the high humidity conditions, compared to the previous observation period, which was lower.Bactrocera spp.populations will increase when the relative humidity of the air is high and decrease when it is low [3].Similarly, temperature measurements in both farms showed that the second and third observations were lower than the last.An increase in temperature reduces the length of the early life stages of Bactrocera spp., including both the immature and embryonic stages, compared to the larval stage [32,33].The existence of fruit-fly species can be inferred in a given habitat and is closely correlated with temperature and humidity levels.
Meanwhile, the decline in Bactrocera spp.during the last observation period was due to harvesting ripe fruit, so the availability of hosts in the orchard continued to decline every week.The impact of harvesting affects the decline in abundance of a population of the presence of fruit flies observed on the crop because the host source where eggs are laid has decreased and will increase again along with the ripening of the following fruit [34].This shows that the host plants' availability affects the fruit fly's catch (Figure 6).

Conclusions
The horizontal trap model was more effective at trapping fruit flies than the vertical trap model.Horizontal traps have an effectiveness of 58% compared to vertical traps, so the orientation of this fruit fly trap is potential and can be used as a fruit insects pest monitoring tool.The consequences of fruit fly surveyance show that the fruit fly species present in citrus plants are B. dorsalis, B. carambolae, and B. umbrosa.Abiotic conditions, including humidity, temperature, and the presence of suitable hosts, mainly influence the abundance of fruit flies in citrus orchards.

Time of Observation
Fruit fly Catch Data (indiv)

Figure 1 .
Figure 1.The study site of research (a) Malang Regency, (b) Dau Sub-District.

Figure 3 .
Figure 3. Bactrocera spp.catch results between vertical and horizontal traps in citrus orchards, Malang District.Fruit fly Catch Data (indiv)

Figure 5 .
Figure 5. Catch rates of males and females between the two traps in citrus orchard, Malang District.

Figure 6 .
Figure 6.Number of individuals trapped at 1 st -4 th time of observation.