Long-term genomic adaptation in a natural Drosophila melanogaster population from central Anatolia

Gaining a clear picture of how microevolutionary change unfolds in natural populations demands high-resolution genomic data sampled repeatedly over time. While seasonal adaptation has been documented in Drosophila melanogaster , long-term directional genomic changes within single natural populations remain poorly resolved, particularly in understudied regions such as the Eastern Mediterranean and Anatolia. Here, we present a seven-year (2015–2021) temporal Pool-Seq analysis of a natural D. melanogaster population sampled from a single locality (Yeşilöz, Ankara, Turkey). We analyzed Pool-Seq samples collected in June, August, September, and October, representing post-winter emergence, peak summer temperatures, and autumn transition shocks. Using multiple ANCOVA models across 1,526,725 SNPs, we identified 9,055 SNPs showing significant directional allele frequency shifts across years (Benjamini–Hochberg q < 0.05). These SNPs mapped to 395 genes harboring HIGH or MODERATE functional impact variants. The sensitivity model excluding the low-coverage September 2015 cohort yielded ~ 8,200 year-significant SNPs with highly consistent results, demonstrating that this cohort did not bias our primary findings. A second, highly conservative joint-correction model identified a core set of 627 SNPs. Across all models, seasonal allele frequency variation within years was negligible, and climatic principal components (temperature and humidity-related) explained no additional variance beyond the overall year effect. Gene Ontology enrichment analysis revealed a pronounced over-representation of genes associated with zinc ion binding, transition metal ion binding, and metallopeptidase activity (Molecular Function). Length-adjusted logistic regression confirmed these associations were independent of gene size, whereas an initially significant cellular component term ( plasma membrane ) did not survive length control and was not interpreted further. Overall, our findings provide compelling evidence for sustained directional genomic change in a natural D. melanogaster population, with primary functional targets concentrated in transcriptional regulation (zinc-finger, metal-ion binding) and proteolytic processes.