Clearing technique to examine the cephalopharyngeal skeletons of blow fly larvae.

Blow fly larvae found in human corpses at death scenes are important in forensic investigations. These larvae are not only useful as an entomological means to estimate the time elapsed between death and the discovery of the corpse (i.e., postmortem interval or PMI), but they can also be used to detect drugs and other toxic substances which might prove to be the cause of In this regard, the accurate identification of blow fly larvae is a principal requirement for subsequent forensic analysis. Identification of blow fly larvae can be accomplished with taxonomic keys that rely on a variety of distinct morphological features. External morphological features include overall body appearance, anterior spiracles, posterior spiracles, and spine bands along the body. Internal organs, particularly the structure of the cephalopharyngeal skeleton and tracheal trunks, are also valuable information for identifying blow fly larvae Some components of the cephalopharyngeal skeleton have been used to differentiate blow fly and flesh fly species in Japan This important structure is embedded within the head region of the larva and is concealed by a thick sclerotized larval integument (Chapman 1982). A clearing method is thus required to reveal the exact structure of this organ. Recently, Zeng et al. (2001) proposed a simple technique for clearing chitosan from crab or lobster shells by using hydrochloric acid (HCl) as a decalcification agent. Hence, we applied this method to see if it would clear the bodies of blow fly larvae sufficiently for the cephalopharyngeal skeletons to be critically examined. A total of 355 third instar Chrysomya megacephala (Fabricius) larvae preserved in 70% alcohol for three months was used in this study. The anterior part of each larva was separated from the whole body under a dissecting microscope using a scalped blade. Anterior parts were then divided into seven groups (50 larvae per group) – one control and six experimental groups. The specimens in the control group were subjected to a clearing method previously described (Smith 1986) in which the larvae were macerated in 10% potassium hydroxide (KOH) for at least 15 min, then transferred to a tube containing 1 ml of glacial acetic for at least 15 min. An equal amount of Berlese fluid was added to the tube and the tube was then covered with a cap and left for 24 h at room temperature. Following the incubation period, each specimen was transferred using a small camel hair brush onto …