Cold storage effects on immature calliphoridae: Survival limits and forensic consequences for PMI estimation

This study investigated the effects of continuous refrigeration (4°C ± 2°C and 50% ± 5% RH) on the development and survival of immature stages (eggs, first, second, and third instar larvae, and pupae) of four forensically important blowfly species (Chrysomya megacephala, Chrysomya putoria, Hemilucilia segmentaria, and Lucilia cuprina) compared to a control group (27°C ± 2°C and 50% ± 5% RH). Groups of approximately 500 individuals at each stage were exposed to refrigeration and the control treatment, with periodic verifications to assess development and survival. Statistical analysis (two-way ANOVA and Šidák posthoc test) revealed significant differences (p < .0001) in development time and survival between the groups. Eggs and first instar larvae did not survive longer than 36 h under refrigeration. Third instar larvae showed greater resistance, with L. cuprina surviving for up to 612 h, entering quiescence without molting. The results highlight the significant impact of refrigeration on the development of these warm-climate species, warning against potential imprecisions in time of colonization (TOC) estimations and, consequently, in postmortem interval (PMI) if refrigeration effects are not considered. The research emphasizes the need for species-specific data and caution when applying linear development models in forensic entomology, especially in cases involving refrigeration.