Morphological and molecular identification of forensically important insect species (Diptera) in Kampala, Uganda using Mitochondrial Cytochrome Oxidase I Gene

Abstract

Insects attracted to corpses may provide important indications of the period post death otherwise called post-mortem interval (PMI), criminal event reconstruction, solving criminal cases, corpse movement and cases of neglect. The use of insects (Diptera) for PMI estimation in Uganda is of limited use since blowfly immature stages are not morphologically distinct. However, this has been solved by the use of molecular techniques which help identify insect species regardless of stage of development. They are quick, accurate and cost-effective. This study was conducted to identify insect species of forensic importance during decomposition stages of pigs (Sus scrofa domesticus) carcasses in Kampala, Uganda. Forensically important adult insect specimens were collected from outdoor (n=150) and indoor (n=120) carcasses and identified using taxonomic keys. Molecular identification of insect larvae was also done using Polymerase Chain Reaction (PCR) amplification of the mitochondrial cytochrome oxidase I partial gene followed by Restriction Fragment Length Polymorphism (RFLP), sequencing and phylogenetic analysis. The five stages of decomposition of the pig carcasses were categorized as: fresh, bloated, active, advanced and dry. The rate of decomposition in outdoor was faster than that in indoor environment. Three different families of Diptera; Calliphoridae, Sarcophagidae, and Muscidae were identified morphologically from both indoor and outdoor environments. The most abundant adult insect species was Chrysomya chloropyga (34.6%) and Chrysomya putoria (28.3%) for the outdoor and indoor environments, respectively. Sarcophaga bercaea was the least abundant adult insect species for both environments. All the species were present in the bloated and decay stages. Only Chrysomya chloropyga was found in the post decay stage. Other species were not observed in the post decay stage. Sarcophaga bercaea was not present in the bloated indoor.No insects were in the dry stage. Among insect larvae collected, Chrysomya putoria (32%) and Chrysomya chloropyga (48.1%) were the most abundant insect larvae from the indoor and outdoor environments respectively. Chrysomya albiceps (20%) and Hemipyrellia fernandica (10.9%) were the least abundant from the indoor and outdoor environments, respectively.Phylogenetic analysis of mitochondrial COI gene of insect larvae clearly clustered together and separated into four genetic clades of Calliphoridae flies (Chrysomya putoria, Chrysomya chloropyga, Chrysomya albiceps and Hemipyrellia fernandica). A fragment of the mitochondrial COI partial gene in this study displayed that DNA-based method can be used as supplemental means to morphological method for identification of the Calliphoridae as insect species of forensic importance in the different environments in Uganda. This concept should be extended to human corpses so that if they are consistent with results from Sus scrofa domesticus carcasses then this technique should be explored for homicide investigations in Uganda.