Geographical distribution and seasonality of insect species and gut microbiota of fly larvae from 65 real cases in China.

Forensic entomology can be pivotal for minimum postmortem interval (PMImin) estimation. Case-based studies can support forensic investigations and guide laboratory research, but systematic data from real cases in China remain limited. In this study, 65 forensic entomology cases collected from 2017 to 2024 in two regions of China, Beijing and Hunan, were analyzed to characterize necrophagous insect occurrence patterns and larval gut microbiota. A total of 30 colonizing insect species from 11 families were identified. Lucilia sericata, Chrysomya megacephala, and Sarcophaga crassipalpis were the most frequently recorded species. Diptera, especially Calliphoridae and Sarcophagidae, were mainly associated with shorter postmortem intervals, whereas coleopteran taxa were more often observed in longer-interval cases. In addition, insect occurrence showed clear geographical distribution and seasonality: L. sericata and S. crassipalpis predominated in Beijing cases, whereas C. megacephala and Sarcophaga peregrina were more frequent in Hunan cases. Overall species diversity was highest in summer, whereas only Calliphora vicina and Aldrichina grahami were observed in winter. 16S rRNA gene sequencing of larval gut microbiota revealed a stable core microbiome across diverse cases and species, including Ignatzschineria, Clostridium, Peptostreptococcus, Vagococcus, and Peptoniphilus. This study provides regional forensic entomology reference data and characterizes the gut microbiota of fly larvae collected directly from human remains in police casework, offering a useful basis for future forensic investigations and laboratory studies.