This study is conducted to develop a simple and cost-effective feeding assay method for evaluating the effect of various natural molecules, including phytochemicals for their antifeedant and insecticidal properties against chewing insects, and identified potential molecules from these assay can be evaluated in field condition for pest management. Multiple assays have been developed to evaluate the effect of synthetic and natural molecules on insect feeding, growth, development, and behavior. Further, assays have been designed.
such as clip case tool, diet covering assay, contact assay, droplet feeding assay, choice feeding assay, and lip dish test for insecticide evaluation, Maintaining uniform experimental conditions, addressing issues like infection and cannibalism, ensuring even phytochemical distribution in larval diet, tracking feeding behavior accurately, and determining optimal phytochemical concentrations pose challenges. Batch variation and experimental design complexities must be managed. Additionally, transitioning from the lab assist to the field application, considering formulation and scalability is demanding.
In the obligatory feeding assay, a large number of molecules can be tested simultaneously with a large number of insect sample sizes. The advantage of this feeding assay is that the test compound is mixed with an artificial diet. Thus, we have control over diet composition, which provides robust and reproducible results.
To begin, weigh the desired components of Fractions A, B, and C separately in a beaker, and prepare a homogenous mixture using a spatula or magnetic stir. Boil Fraction C at around 100 degrees Celsius in a microwave for five minutes. Then add it to Fraction A, and mix thoroughly.
Once the mixture cools down, add Fraction B, and pour the prepare to control diet mixture into a transparent polystyrene Petri dish. To prepare quercetin containing an artificial diet, dissolve quercetin hydrate and dimethyl sulfoxide. Then add the dissolved quercetin into Fraction B, followed by a mixture of Fractions A and C.Add an equivalent dimethyl sulfoxide to the prepared controlled diet.
To begin, keep Helicoverpa armigera neonates in the plastic breeding chamber covered with a muslin cloth in the insect culture room. Then gently transfer newly emerged neonates using a fine paintbrush on a freshly prepared, chickpea-based artificial diet for growth. For the feeding assay, collect 21 second-instar larvae for the control and treatment set.
Cut previously prepared control and quercetin-containing diet into small pieces, and record the weight of the diet and the insect's body. Carefully transfer the insect into the culture vial, add the respective diet into the vial, and let the insect feed. From the second day till the 10th day, record the weight of the insect body, followed by a given diet, uneaten diet, and frass.
Count the live and dead larvae and pupae on day 10. To record and analyze the data, compare the insect body weight between the control and treatment groups using a student T-test. Plot a Kaplan-Meier curve for survival percentage using the graphing software using live/dead larvae and pupae count.
Then calculate the percent pupation, and compare larval development and nutritional indices. In the present study, the insect larvae fed with quercetin-containing diet showed a significant decrease compared to the control group, thereby resulting in reduced body size. Also, a notable reduction was observed in the feeding rate of quercetin-fed larvae compared to the control.
Furthermore, there was a significant decrease in percent pupation in quercetin-fed larvae, suggesting developmental retardation upon treatment. For insects fed on the quercetin-containing diet, the efficiency of conversion of ingested food to body matter was reduced by approximately 9%compared to the control. The efficiency of conversion of digested food was reduced by approximately 49%compared to the control.
The approximate digestibility in quercetin-fed insects was increased by 5%compared to the control.
