Differential gene expression during pupal development of Lucilia sericata (Diptera: Calliphoridae) for improving postmortem interval estimation

Abstract The main objective of forensic entomology is to estimate the postmortem interval (PMI) by analyzing the size and duration of the immature stages of insects colonizing a cadaver. However, this estimation becomes difficult during the pupal stage due to the absence of significant visible morphological changes. This study explores the differential gene expression of Lucilia sericata (Calliphoridae) pupae as a molecular tool for PMI estimation. Pupae were collected daily under controlled conditions at 23°C, and RNA was extracted for transcriptomic analysis. Six target genes (ecdysone receptor, Hsp90, actin, regucalcin, wingless, and fat body protein 1) were identified and correlated with distinct percentages of pupal development. Our results suggests that the combination of the expression levels of at least three genes, such as ecdysone receptor, regucalcin, and wingless, is sufficient to infer the percentage of pupal development throughout the entire pupal stage. Additionally, graphical tools were developed to facilitate the use of individual and grouped genes as markers for L. sericata age estimation. Since the pupal period comprises approximately half of the Calliphoridae developmental cycle, these findings are particularly valuable when pupae represent the most advanced evidence recovered from a crime scene or autopsy.