The influence of Methylphenidate Hydrochloride on the development of the forensically significant blow fly Chrysomya chloropyga (Diptera: Calliphoridae) in the Western Cape, South Africa by Hartwig Visser VSSHAR002 SUBMITTED TO THE UNIVERSITY OF CAPE TOWN In partial fulfilment of the requirements for the degree MPhil (Biomedical Forensic Science) Faculty of Health Sciences Division of Forensic Medicine and Toxicology UNIVERSITY OF CAPE TOWN 2016 Supervisor: Dr Marise Heyns Co-supervisor: Ms Bronwen Davies University ofape Town Division of Forensic Medicine and Toxicology University of Cape Town The copyright of this thesis vests in the author. No quotation from it or information derived from it is to be published without full acknowledgement of the source. The thesis is to be used for private study or non- commercial research purposes only. Published by the University of Cape Town (UCT) in terms of the non-exclusive license granted to UCT by the author. University of Cape Town University ofape Town ii iii iv v vi vii Table of Contents Title page ......................................................................................................................... i Declaration ...................................................................................................................... ii TurnItIn report ................................................................................................................. iii Table of contents .......................................................................................................... viii List of figures ................................................................................................................ xiii List of tables ................................................................................................................. xvi Study summary ........................................................................................................... xvii Chapter 1. Research Proposal .................................................................................... 1 1.1.Purpose of the study ........................................................................................... 4 1.2.Background. ......................................................................................................... 4 1.3.Methodology ......................................................................................................... 8 1.3.1.Study design ................................................................................................ 8 1.3.2.Characteristics of the study population ........................................................ 8 1.3.3.Spiking of the food substrate ......................................................................... 8 1.3.4.Rearing of blow fly larvae .............................................................................. 9 1.3.5.Sampling of larvae ........................................................................................ 9 viii 1.3.6.Killing of blow fly larvae ............................................................................... 10 1.3.7.Data analysis and measurement ................................................................. 10 1.3.8.Quantitation of MPH from sampled larvae .................................................. 10 1.3.9.Disposal of end-products ............................................................................ 11 1.4.Statistical analysis .......................................................................................... 11 1.5.Work plan ........................................................................................................... 11 1.6.Ethics. ................................................................................................................ 12 1.7.Benefits of the study ........................................................................................... 12 1.8.References ......................................................................................................... 13 1.9.Addendum to proposal ....................................................................................... 17 Chapter 2. Literature Review ..................................................................................... 20 2.1.Background ....................................................................................................... 21 2.2.Literature review ................................................................................................. 27 2.2.1.Historic foundation ...................................................................................... 27 2.2.2.Post-mortem interval (PMI) ......................................................................... 28 2.2.3.Forensic entomology ................................................................................... 29 2.2.4.Chrysomya chloropyga ............................................................................... 33 ix 2.2.5.Entomotoxicology ........................................................................................ 34 2.2.6.Methodological inconsistencies ................................................................... 40 2.2.7.Methylphenidate hydrochloride ................................................................... 47 2.3.Summary ............................................................................................................ 50 2.4.References ......................................................................................................... 52 Chapter 3. Manuscript ................................................................................................ 67 3.1.Abstract .............................................................................................................. 69 3.2.Introduction ........................................................................................................ 70 3.3.Methodology ....................................................................................................... 73 3.3.1.Study population ......................................................................................... 73 3.3.2.Spiking of the food substrate ....................................................................... 74 3.3.3.Rearing of blow flies and larvae .................................................................. 74 3.3.4.Sampling of larvae ...................................................................................... 75 3.3.5.Larval identification and size measurement. ............................................... 76 3.3.6.Duration of developmental stages ............................................................... 76 3.3.7.Qualitative analysis of MPH from sampled larvae ....................................... 77 3.3.8.Statistical analysis ....................................................................................... 78 3.4.Results ............................................................................................................... 79 x 3.4.1.Species composition ................................................................................... 79 3.4.2.Larval growth patterns ................................................................................. 80 3.4.3.Duration of developmental stages ............................................................... 88 3.4.4.MPH detection............................................................................................. 89 3.5.Discussion .......................................................................................................... 91 3.5.1.Mixed populations ....................................................................................... 91 3.5.2.The effect of MPH on larvae growth rates and sizes ................................... 92 3.5.3.The effect of MPH on the duration of developmental stages ....................... 94 3.5.4.The interaction of variables ......................................................................... 97 3.5.5.Detection of MPH ........................................................................................ 99 3.6.Limitations ........................................................................................................ 100 3.6.1.Mixed population and laboratory cultures .................................................. 100 3.6.2.Sample size .............................................................................................. 101 3.6.3.Data collection........................................................................................... 102 3.6.4. MPH concentration investigated............................................................... 102 3.6.5. Toxicological analysis and drug distribution ............................................. 102 3.6.6 Replication of findings ............................................................................... 103 xi 3.7.Conclusion ....................................................................................................... 103 3.8.References ....................................................................................................... 105 Chapter 4. Appendices ............................................................................................. 114 Appendix A - Highlights ......................................................................................... 115 Appendix B - Acknowledgements: Manuscript ....................................................... 116 Appendix C - Author contributions .........................................................................