Location: Tropical Pest Genetics and Molecular Biology Research Unit
Project Number: 2040-30400-003-011-I
Project Type: Interagency Reimbursable Agreement
Start Date: Jun 1, 2026
End Date: Dec 30, 2027
Objective:
Objective 1. Determine the prevalence and geographic distribution of insecticide resistance alleles in wild Hawaii melon fly.
Objective 2. Calculate changes in the frequency and distribution of resistance alleles across growers/fields with varying Integrated pest management (IPM) practices.
Objective 3. Determine genetic basis for insecticide resistance in Bactrocera dorsalis and Ceratitis capitata through genomic analysis and comparative analysis of insecticide-resistance-associated genes.
Approach:
The second year of this project will focus on collecting additional wild Z. cucurbitae from across the Hawaiian archipelago and performing whole genome-sequencing to determine the change in allele frequencies of insecticide resistance-associated alleles from the previous collection. Additionally, local land managers and growers will be surveyed to determine insecticide application history and which other IPM practices are currently in place. This can range from sites that are not managed where no selection pressure is applied, to heavily managed farms which rely heavily on application of insecticides. Repeat sampling will be conducted throughout the year to determine within-year temporal and spatial fluctuations of insecticide resistance-associated alleles. By using collection methods established in year 1, we will additionally be able to use collection data to determine population distribution and density over time that can be associated with abiotic factors or host availability. Lastly, we propose a new objective to identify homologous genes in closely related Tephritidae such as C. capitata, B. dorsalis, Anastrepha ludens, and Zeugodacus tau to determine sequence homology and divergence to assess the application of this work to other species. We will additionally perform a population genetic analysis on wild B. dorsalis and C. capitata with comparisons to analogous lab-reared colony data. Calculation of SNP ratios in resistance-associated genes relative to other genes will reveal historical selection for insecticide resistance.
Whole-genome sequencing, pooled sequencing, and data analysis
Whole-genome sequencing will be performed on all flies collected in year 2. All flies will be sequenced individually or as pools by geographic location of the collection. This will enable post-hoc pooling of genotype data to estimate allele frequencies and analysis of allele frequency from the pool. Flies and fly pools will be screened for the high-effect loci and Spinosad resistance associated alleles identified from year 1 Objective 1.
Comparative gene analysis
Gene orthologs for Z. cucurbitae genes identified in year 1 Objective 1 will be identified in A. ludens, B. dorsalis, C. capitata, and Z. tau to determine sequence homology and genetic distance. Similar sequence and low genetic distance between Z. cucurbitae and the other species will indicate their potential for use in associating their function with Spinosad resistance. Low sequence similarity and high genetic distance will indicate low association with Spinosad resistance. Population level whole genome sequencing data for B. dorsalis and C. capitata will be obtained from wild populations and SNP ratios will be estimated in these gene orthologs to determine SNP densities. High SNP densities in these genes will reveal past pressure for resistance and demonstrates utility as diagnostic loci that can be assessed for insecticide resistance management.