STEM I at MAMS is taught by Dr. Crowthers. In STEM I, we do class activities and projects, building up our skills and knowledge to aid our 6-month long science fair.
Maximizing the Effectiveness of Mosquito Chemical Attractants Using Drosophila as a Model
The goal of this project was to determine the chemical most effective at attracting Drosophila. A choice chamber was set up to test the attractancy of two chemicals: 1-octen-3-ol and lactic acid. 1-octen-3-ol was found to attract more Drosophila within the time frame, but the p-value of a one-tailed paired t-test was greater than the significance value.
Hundreds of millions of humans are affected by mosquitoes annually and millions lose their lives to mosquito-borne diseases. To prevent as many cases as possible, several mosquito-controlling devices are. The difference between each method is each method targets a different aspect of a sensory system of a mosquito. However, no design has yet to implement any attractive odorant. DEET and natural odors aim to repel mosquitoes and zapping traps lure mosquitoes with light. As this project aims at examining odors, Drosophila will be used, since they been proven to be a possible model organism for mosquitoes. 15 Drosophila were put in a choice chamber for 10 minutes. At the ends of the choice chamber, a chemical solution and the control solution, ethanol, both at a quantity of 100 microliters, were dropped onto a cotton ball, giving the Drosophila a choice between the two odors. At the 0, 5, 7.5, and 10 minute mark, the amount of Drosophila was counted and recorded. 1-octen-3-ol was found to attract 6 Drosophila on average while lactic acid only attracted 3, on average. In addition, 1-octen-3-ol also attracted 1 additional Drosophila, on average, within the first 5 minutes. A one-tailed paired t-test was used to calculate the p-value and was found to be 0.102416. However, a hypothetical odor-based mosquito trap should prioritize examining the effectiveness of 1-octene-3-ol as a chemical attractant before testing the effectiveness of lactic acid.
Does utilizing a chemical attractant, like 1-octen-3-ol, of mosquitoes, make mosquito-killing devices more or less effective at attracting and killing mosquitoes?
The hypothesis of this project is that since 1-octen-3-ol was found to have the highest individual attractancy rating, it will also be the most attractive when present in an environment with other odors.
Mosquitoes are one of the deadliest organisms in the world. They are known as vectors: organisms that transmit a disease into a host body (Mathew et al., 2013). Every year, mosquitoes affect, on average, half a billion of humans (Bustamante, 2019). Of that half a billion, 2.7 million are fatalities (Bustamante, 2019). Certain species of mosquitoes need to blood feed in order to reproduce; one source of blood comes from humans (Mathew et al., 2013). Although mosquitoes are pests, they are also vital for the ecosystem; therefore, the extinction of mosquitoes would have severe repercussions on the balance of ecosystems (Peach, 2019). There are thousands of mosquito species in the world, many of which do not target humans (Peach, 2019). There are two vital roles mosquitoes play in ecosystems: pollination and food chains (Peach, 2019). The reason why some species of mosquitoes are attracted to animals is a combination of many factors. Each step of the host-seeking process attracts a mosquito closer and closer to the meal, until finally making contact and sucking the host. The first stimulus that draws the attention of a mosquito is odor and CO2, which occurs at the farthest distance (Caltech, 2015). As there are thousands of species of mosquitoes, this first phase occurs at around 10 to 50 meters (Caltech, 2015). Next, a combination of CO2, as well as the odor, and sight continue to lure the mosquito to its host (Caltech, 2015). This middle step occurs at around 5 to 15 meters (Caltech, 2015). Finally, when the mosquito is close enough, CO2, odor, visual signals, and the host’s body heat, taste, and moisture cues if the host can be fed on (Mathew et al., 2013). The last phase happens within 1 meter of the host or during physical contact (Caltech, 2015). By targeting a mosquito’s olfactory senses, the first phase, a hypothetical trap should be the most effective, as the attractant functions at the farthest range during the mosquito’s host-seeking process. Numerous inventions have been developed, and numerous more are being invented, each with minor differences: the biological aspect of a mosquito being targeted. However, there has not been a method guaranteeing 100% safety from mosquito bites. Although scientists have found DEET to be the most effective repellent at preventing mosquito bites, there are still several flaws. Firstly, the effectivity of DEET is not permanent. DEET must be applied repeatedly to ensure safety from mosquito bites, and even then, unlucky hosts may still be bitten. Secondly, the more pressing issue is DEET negatively affects the environment. Because DEET in insect repellents (has other uses too) functions as a spray, it is physically introduced into the environment. DEET must be treated at wastewater treatment plants (WWTP) because the chemical is not biodegradable. However, these WWTP are not able to remove 100% of all DEET, causing some DEET to be polluted back into the environment; studies have found DEET to be damaging to aquatic insects as well as microorganisms when present in high enough concentrations (Aronson et al., 2011). Other odors have been tested and found to be more eco-friendly, but these odors tend to not be as effective at repelling mosquitoes. An experiment conducted by Mathew et al in 2013 used Aedes Aegypti and a choice chamber to test different compounds for their attractancy or repellency. The choice chamber was comprised of an initial box where the mosquitoes were released, a final chamber where the chemical was released, and two separate tubes connecting the two components. To decide the attractancy or repellency of the compound, Mathew et al implemented the attractancy and repellency index (include formula?). They found that 1-octen-3-ol had the highest attractancy rating for Aedes Aegypti individually, at around 58, while a combination of myristic acid, lactic acid, and carbon dioxide had the highest attractancy rating as a synthesized compound, at around 62. In addition to the combination stated previously, three other synthesized compounds also had a higher attractancy rating compared to 1-octen-3-ol individually (Mathew et al., 2013). A research paper done by Vosshall in 2000 indicated Drosophilae, more commonly known as fruit flies, are great model organisms to use for odor experiments. Drosophilae are able to recognize hundreds of different odorants (Vosshall 2000). In addition, Drosophilae has been proven to adapt to simple olfactory-based tests (Vosshall 2000). Finally, because Drosophilae are able to be easily mutated, they may be easily adapted for an experiment. Another study conducted by Raji and Degennaro in 2017 found that mosquitoes and Drosophilae are attracted to similar chemicals. A mosquito’s and Drosophila’s brain was found to incorporate both odor and taste when feeding (Raji & Degennaro, 2017). Also, both organisms can sense heat through Ionotropic Receptors (Raji & Degennaro, 2017). They also found that combining a chemical attractants with others the resulted compound usually is more attractive.
One vial of wild-type Drosophila was bred in order to accumulate a sufficient amount of flies for multiple test trials. The Drosophila purchased from Carolina were funneled into an empty tube, where they then were anesthetized using fly-nap. The anesthetized Drosophila were placed under a microscope to identify the sex of each fly. 1 female for every 4 males was placed into the culture tube to breed. Culture tubes were filled with media: potato flakes, yeast, and water. Choice Chamber The choice chamber was created using 3 Kirkland water bottles in order to create an environment in which the Drosophila could navigate around and see the Drosophila at all times. The ends of the bottles were cut, at the farthest ridge, and were hot glued and taped together, in order to prevent the flies from escaping as well as providing structural integrity to the choice chamber. The choice chamber was modeled to form a t-shape. Testing 15 awake Drosophila were transferred from the culture tube to the choice chamber using a funnel. After all 15 Drosophila were in the choice chamber, the cap of the entrance bottle was closed. At one end of the choice chamber, 100 microliters of ethanol at a 99% concentration was dropped onto a cotton ball. At the other end of the choice chamber, 1-octen-3-ol or lactic acid was dropped onto another cotton ball. Both 1-octen-3-ol and lactic acid were at a 1% stock solution for consistency purposes. At the 0, 5, 7.5, and 10 minute-mark, the amount of Drosophila was recorded on a spreadsheet. The number of Drosophila at the 0 minute-mark was recorded because during the time when the Drosophila were in the choice chamber, the time it took to prepare the cotton balls and placing them in the choice chamber enabled enough time for the Drosophila to fly throughout the choice chamber. Cotton balls were used in order to minimize the amount of Drosophila dying to the chemical. Statistical Test A paired t-test was used to compare 1-octen-3-ol and lactic acid. A paired t-test was implemented because the hypothesis of the project is that 1-octen-3-ol will attract the most Drosophila, and a paired t-test will determine whether the data of the two tested chemicals is significant or not. The test is right-tailed as the expected number of Drosophila reaching each end was greater than the observed number of Drosophila at each end. The null hypothesis is that there is not significant difference between the effectiveness of attractancy between 1-octen-3-ol and lactic acid. The significancy level is 0.05.
Two trials were conducted for each chemical: 1-octen-3-ol and lactic acid. At the 0, 5, 7.5, and 10 minute mark, the number of Drosophila at the ends of the choice chamber were counted. 1-octen-3-ol attracted more Drosophila over 10 minutes, at an average of 6 out of the total 15 Drosophila being attracted towards the end with the 1-octen-3-ol solution. Lactic acid attracted 3 Drosophila, on average, over the 10 minute time period.The p-value of the one-tailed paired t-test was p=0.102416. Therefore, the null hypothesis is accepted; it is not certain whether 1-octen-3-ol will attract more Drosophila than lactic acid can.
As the p-value of the conducted one-tailed paired t-test was greater than the significance level, the null hypothesis is accepted; it is not certain whether 1-octen-3-ol will attract more Drosophila than lactic acid can.
More trials should be conducted to achieve more significant data. Once the p-value of the one-tailed paired t-test is below the significance level, the null hypothesis may be rejected, and therefore would indicate which chemical attracted is the most effective at luring Drosophila. This chemical may then be implemented into mosquito electrocution traps. This chemical should also be made widely available. However, more testing would need to be done for this hypothetical more effective attractant in terms of safety for recreational use as well as for the environment. Finally, an electrocution trap utilizing a chemical attractant would also have to be compared to the effectiveness of other mosquito-controlling methods.
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Feb Fair Poster