Ian Dadour

Introduction: In addition to their use in the estimation of the minimum post-mortem interval (minPMI) necrophagous blowflies (Diptera: Calliphoridae) may also represent a reliable specimen for toxicological analyses (Entomotoxicology) especially in the absence of tissues and fluids normally taken for such purpose. Numerous researchers have extracted several toxicological substances and measured their effects in altering the morphology, the rate of development and the survival on blowfly immatures associated with remains. The accumulation of such substances in blowflies may compromise their use for the estimation of the minPMI. At present a modest number of substances and insect species/instars have been studied. Furthermore, many early studies utilized analytical procedures which are now obsolete with little/no validation. Methods and Results/Conclusions: This research presents the method validation and the effects on blowflies for: Nicotine in Calliphora vomitoria (L.) by GC-MS. Nicotine is an alkaloid present in the tobacco plant. Nicotine is one of the most deadly poisons known to man and it’s extremely easy to purchase. Methamphetamine in C. vomitoria by GC-MS. Methamphetamine is a psychostimulant synthetic drug originally used as a medical treatment but now a common recreational drug (meth, crystal, and ice). High doeses can cause death. Endosulfan in C. vomitoriaby GC-MS. Endosulfan is a toxic pesticide, responsible for many fatal poisoning incidents around the world. Endosulfan is acutely neurotoxic to both insects and humans. Coumatetralyl in Lucilia sericata (Meigen) by LC/MS-MS. Cumarine compounds are used in anti-coagulant therapies. Coumatetralyl is a product largely used in poisoned baits.

Forensic entomology is the study of insects within a legal framework. The outstanding variable in determining an accurate post mortem interval (PMI) is the time that adult flies first lay live larvae or eggs onto a body. The predicted order of insects, typically blowflies and flesh flies that are attracted to a decomposing corpse or cadaver is pivotal to determining the PMI in forensic entomology. The predicted order is divided into primary, secondary and tertiary insects. This correlates with the process of decomposition and how it changes over time. Blowflies generally arrive after death and recent research has shown that Calliphora dubia, Calliphora varifrons and sarcophagid adults lay live larvae onto guinea pig carcasses within 1 hour of exposure. These flies are typically the primary visitors to a corpse. Very little is known about ovoviviparous blowflies, which are invariably the first blowflies to visit a corpse in south western Australia and are often the critical species in a PMI determination. New trials exposed 30 guinea pig carcasses throughout the day (0600-2000hrs) during spring and summer in bushland on each of 5 successive days. Replicate carcasses were set up randomly along a kilometre of bushland track and a carcass was removed every 0.5, 1.0, 2.0, 4.0, 8.0hrs throughout the day blowflies layed onto carcasses within 30 minutes (33% of carcasses), and on average on all carcasses within 1.5hrs of death. Eight different oviparous (e.g Lucilia sericata, Chrysomya rufifacies, Chrysomya varipes and Australophyra rostrata), and the oviviviparous fly species above deposited either eggs or live larvae onto the carcasses within 6hrs. This study highlights the previously unknown rapidity with which blowfly species are able to commence laying onto carcasses and from a large number of fly species, some of which are considered in the literature to be late colonisers of carcasses. Two repeat trials of exposure of carcasses again over 5 successive days in summer revealed 5 similar species of blowflies laid onto carcasses. This research has many ramifications for the post mortem interval estimation especially if all flies previously designated secondary and tertiary are in fact primary visitors to a corpse.

Following a homicide the regulation of forensic practice in many countries prevents the pathologist performing an immediate autopsy. Preceding each autopsy the corpse and the insects associated with it are stored in a cool room in a mortuary (4°C) to slow down the decomposition process as well as the development of most of the necrophagous insects associated with the corpse. In late stages of decay, fly immatures may be present in masses. The aim of this research was to investigate the growth and development of different larval masses of the blowfly Lucilia sericata (Meigen) (Diptera: Calliphoridae) when stored in a cold environment (4°C). The experiments were divided into a number of trials comprising different storage conditions (continuous exposure or cyclic exposure) for immature stages (L2 and L3) and included four different densities of larvae (100, 500, 1000, 5000 larvae) each placed on 3kg of liver. L. sericata adults were first established in colony and allowed to complete one generation prior to each experiment. Results show that if the larval mass has a density between 1000 and 5000 larvae and have already reached the third instar, then there is no influence on the development time of larvae. Therefore, the identification of the size of the larval mass as well as the instar of the larvae present are crucial data for any subsequent determination of a correct Post Mortem Interval.

The identification of insects associated with a corpse and their specific insect biology in the context of a crime scene is used to provide the best estimation of the minimum post mortem interval. In many cases when insect material is observed at homicides, suspicious deaths and suicides then the medical examiner, coroner, pathologist or trained forensic field officer will call an entomologist. In some cases, although not ideal, any one of these attending persons may do the collections on behalf of an entomologist. Crime scene protocols suggest that all care must be taken in the collection of entomological evidence and this should be gathered only by individuals that have knowledge and experience and/or accreditation as a practitioner in this activity. Unfortunately, in many countries just a small number of personnel, both pathologist and law enforcement are well trained in collection and preservation of insect material on the crime scene. Moreover, only a small number of forensic teams have forensic entomology information sheets and tools in their crime scene kits. It is for these reasons that we have made this information available via a smartphone application (app). Smartphone adoption is now widespread in both the consumer and professional spheres. A simple survey made by the authors has shown that a high percentage of Italian and Australian forensic officers own a smartphone. At present, despite there being a considerable number of smartphone applications focused on general entomology and general forensics (e.g. crime scene marker, time of death, anthropology-anthropometry), no app dedicated to forensic entomology is available. SmartInsects is divided in sections such as: general concepts, insect pictures, guidelines for sampling and storage of insect material and collection of environmental data. Moreover, useful references and contacts are provided. This app is purely designed to guide the crime scene technician or proxy for collecting and preserving entomological samples. App updates with translation into different languages will be become available after further input from the forensic entomology community.

Forensic entomology is today worldwide accepted as an essential tool to reach the success on a murder investigation and not only. Cases involving stored products and infestations of human environments as offences against livestock and pets are now usually investigated with the scientific support of a forensic entomology expert together with law enforcement and all the other professionals appointed by prosecutor and/or defence. At present Italy can boast about a good number of forensic entomology laboratories scattered along the peninsula and mainly bound to Universities. Since 2007 the city of Turin has been the base of a forensic entomology laboratory (FELab) linked with the Public Health Service (ASL TO1). Because of the world financial crisis on 2010-2011 the priorities and money found of the ASL moved to a different direction. At the same time the University of Turin at the Department of Chemistry showed a great interest in the forensic entomology practice. From the end of 2011 a forensic entomology laboratory is hosted at the of the University of Turin and it is born thanks the collaboration between the Department of Chemistry and the Centre for Forensic Science at University of Western Australia. Beyond the work in foresic cases where insects are involved, the lab activity research is mostly focused on entomotoxicology. Students in particular from the Master program in "Clinical, Forensic Chemistry and Doping Control” can practice and apply for thesis. Moreover the lab has a strictly collaboration with the Regional Antidoping Center and Regional Toxicology Laboratory and with the investigative Police, Court and forensic Pathologists. In this poster we show the preliminary research works in which the lab is now involved. The lab is now open to collaborate with other lab and Insititutions to improve the knowledge in entomotoxicology for forensic purpose.