SUCCESSION AND BIOACCUMULATION STUDIES OF BLOWFLIES

DECOMPOSING PESTICIDES-INTOXICATED RABBIT CARCASS

Vannessa Lawai

Master of Science

(Forensic Chemistry)

2014

Faculty of Resource Science and Technology

SUCCESSION AND BIOACCUMULATION STUDIES OF BLOWFLIES

DECOMPOSING PESTICIDES-INTOXICATED RABBIT CARCASS

VANNESSA LAWAI

A thesis submitted

In fulfillment of the requirements for the degree of Master of Science

(Forensic Chemistry)

Faculty of Resource Science and Technology

UNIVERSITI MALAYSIA SARAWAK

2014

i

Declaration

I hereby declare that no portion of the work referred to in this dissertation has been submitted

of an application for another degree of qualifications of this or any other university or

institution of higher learning.

____________________________

Vannessa Lawai

Department of Chemistry

Faculty of Resource Science and Technology

University Malaysia Sarawak

ii

Acknowledgements

I would like to express my deepest gratitude and appreciation to my supervisor, Assoc. Prof.

Dr. Zainab Ngaini and my co-supervisor Assoc. Prof. Dr. Nor Aliza Abdul Rahim for their

excellent guidance, generous supervision, patience, encouragement and advices. A deepest

appreciation too for providing me with warm atmosphere throughout the completion of this

research.

I would like to thank Dr Alvin Chai Lian Kuet from Agricultural Research Centre,

Semenggoh for providing some guidance and helps during the early study. I would also like

to thank Mr. Joseph Tau ak Katip, medical lab technologist from Faculty of Medicine and

Health Sciences for helping with carcass preparation.

I would like to thank Wan Sharifatun Handayani, who as a good friend, was always willing

to help and give her best suggestions. Great recognition too towards fellow postgraduates in

Organic Chemistry Laboratory, UNIMAS for their helpful cooperation and consideration.

My warmest gratitude to my family (Anne Lupa, Lawai Jalong, Julia Ling, Magdelen Jalong,

Nixon Girang, Nicholas and Steinmacher,) for their love, supports and inspirations.

Finally, I would like to express my appreciation toward Yayasan Tunku Abdul Rahman

Cawangan Sarawak for the scholarship and UNIMAS for the financial support.

iii

Abstract

Forensic entomotoxicology is a study on the usefulness of insects as alternative toxicological

samples when conventional matrices such as blood, urine or internal organs are no longer

available. The use of blowfly maggots as pesticide indicator as well as in diagnosing the

cause of death was investigated. A field experiment was conducted in which rabbits were

poisoned and killed using four pesticides namely dimethoate, malathion, permethrin and

paraquat dichloride. Pesticides were introduced to the rabbits via oral ingestion to imitate the

real suicidal or accidental pesticides poisonings. Decomposition rate of rabbits and insect

succession were investigated in this study. Dimethoate and permethrin ingested carcass

showed a slow rate of decomposition. Both took 10 days and 34 days, respectively, to

completely decompose if compared to the control carcass which took only 8 days. Both

malathion and paraquat dichloride ingested carcass showed similar rate of decomposition to

control carcass with 7 to 8 days to completely decompose. Small numbers of insects were

found in/on dimethoate and permethrin ingested carcass. Meanwhile, Chrysomya

megacephala and Chrysomya rufifacies were found dominant in infesting malathion and

paraquat dichloride ingested carcass. Both species belonged to the family of blowfly. Lucilia

species were also found infesting paraquat dichloride ingested carcass. Therefore, the

detection of pesticides were only carried out for malathion and paraquat. Detection of

pesticides in maggots samples were investigated by using GC-MS. Malathion was detected

at retention time 8.8 min, while, paraquat dichloride was detected at retention time 12.8 min,

indicating that the maggots fed on pesticides-exposed flesh would be a valid samples to

identify the cause of death in pesticide-related death.

iv

Abstrak

Forensik entomotoksikologi merupakan kajian mengenai kegunaan serangga sebagai pilihan

alternatif untuk sampel toksikologi apabila matriks konvensional seperti darah, air kencing

atau organ-organ dalaman tidak dapat diperolehi. Penggunaan berenga langau sebagai

penunjuk racun perosak dan juga untuk mendiagnosis punca kematian diselidik. Satu kajian

dijalankan di mana arnab diracun dan dibunuh menggunakan empat racun perosak iaitu

dimethoate, permethrin, malathion dan parakuat diklorida. Racun perosak diberikan kepada

arnab secara oral bagi menyamai kes membunuh diri dengan mengambil racun perosak

samada secara sengaja atau tidak sengaja. Kadar pereputan arnab dan turutan serangga telah

dikaji. Bangkai yang menelan dimethoate dan permethrin menunjukkan kadar pereputan

yang perlahan. Kedua-dua bangkai masing-masing mengambil masa selama 10 hari dan 34

hari untuk mereput jika dibandingkan dengan bangkai kawalan yang mana hanya mengambil

masa 8 hari. Bangkai yang menelan malathion dan parakuat diklorida menunjukkan kadar

pereputan yang sama dengan bangkai kawalan dengan hanya 7 hingga 8 hari. Serangga

didapati sedikit samada di dalam atau pada bangkai yang menelan dimethoate dan

permethrin. Spesies Chrysomya megacephala dan Chrysomya rufifacies didapati dominan

dalam menggerumuni bangkai yang menelan malathion dan parakuat diklorida. Kedua-dua

spesies ini adalah daripada keluarga langau. Spesies Lucilia juga didapati menggerumuni

bangkai yang menelan parakuat diklorida. Oleh yang demikian, pengesanan racun perosak

hanya dilakukan pada malathion dan parakuat diklorida. Pengesanan racun perosak di dalam

sampel berenga disiasat mengunakan GC-MS. Malathion dikesan pada masa 8.8 minit.

Manakala, parakuat diklorida dikesan pada masa 12.8 minit menunjukkan bahawa berenga

yang memakan daging bangkai yang terdedah kepada racun perosak tersebut sesuai

v

dijadikan sebagai sampel untuk mengenal pasti punca kematian di dalam kematian yang

berkaitan dengan racun perosak.

vi

Table of Contents

Page

Declaration i

Acknowledgements ii

Abstract/ Abstrak iii

Table of Contents vi

List of Abbreviations ix

List of Tables x

List of Figures xi

List of Scheme xvi

Chapter 1: Introduction 1

1.1 Forensic science 1

1.2 Forensic entomology 2

1.2.1 Arthropods 3

1.3 Toxicology 4

1.3.1 Entomotoxicology 5

1.4 Problem statement 6

1.5 Objectives 7

Chapter 2: Literature Review 8

2.1 Forensic entomology 8

2.2 Estimation of post-mortem interval (PMI) 9

2.2.1 Decomposition stages of a body 9

2.3 Carrion insect succession 11

2.4 Diptera (Flies) 12

vii

2.4.1 Blowflies 13

2.5 Fly larvae identification 14

2.5.1 Identification characteristic of Chrysomya megacephala 20

2.5.2 Identification characteristic of Chrysomya rufifacies 21

2.6 Entomotoxicology 22

2.6.1 Extraction of the toxins and controlled substances from insects 27

2.7 Deaths from pesticide poisonings 30

2.8 Pesticides 31

2.8.1 Dimethoate 31

2.8.2 Malathion 33

2.8.3 Paraquat dichloride 34

2.8.4 Permethrin 37

Chapter 3: Material and Methods 40

3.1 Material 40

3.2 Instrumentation 40

3.3 Carcass preparation 41

3.4 Preparation of larvae for species identification 42

3.5 Preparation of stock solutions 43

3.5.1 Malathion 43

3.5.2 Paraquat 43

3.6 Toxicological analysis 44

3.6.1 Maggots from carcass ingesting malathion (CMT) 44

3.6.2 Maggots from carcass ingesting paraquat dichloride (CPQ) 44

3.7 Specificity test

45

viii

Chapter 4: Results and Discussion 46

4.1 Entomotoxicological study on carcass ingesting dimethoat(CDM) and

control carcass (CC1)

46

4.1.1 Decomposition stages and flies activity of CDM and CC1 46

4.2 Entomotoxicological study on carcass ingesting permethrin (CPM) and

control carcass (CC2)

51

4.2.1 Decomposition stages and flies activity of CPM and CC2 52

4.3 Entomotoxicological study on carcass ingesting malathion (CMT) and

control carcass (CC3)

57

4.3.1 Decomposition stages and flies activity of CMT and CC3 58

4.3.2 Species identification of maggots collected from CMT 61

4.3.3 Toxicological analysis of maggots from CMT 66

4.4 Entomotoxicological study on carcass ingesting paraquat dichloride

(CPQ) and control carcass (CC4)

71

4.4.1 Decomposition stages and flies activity of CPQ and CC4 73

4.4.2 Species identification of maggots collected from CPQ 76

4.4.3 Toxicological analysis of maggots from CPQ 80

Chapter 5: Conclusion and Recommendations 86

Chapter 6 : References 88

Appendix A : Pictorial Key 102

Appendix B : Temperature and humidity data 103

Appendix C : Conference proceedings 108

Appendix D : Achievement 109

Appendix E : Publication 110

ix

List of Abbreviations

ACS Accessory sclerite

ADP Anterodorsal process

CC1 Control carcass 1

CC2 Control carcass 2

CC3 Control carcass 3

CC4 Control carcass 4

CDM Carcass ingested dimethoate

ChE Cholinesterase

CMT Carcass ingested malathion

CPM Carcass ingested permethrin

CPQ Carcass ingested paraquat dichloride

DA Dorsal arm

EPA Environmental Protection Agency

GC Gas chromatography

GC-MS Gas chromatography – mass spectrometry

LD50 Lethal dose 50

LLE Liquid-liquid extraction

min minutes

NaBH4 Sodium borohydride

NaOH Sodium hydroxide

PMI Postmortem interval

PS parastomal sclerite

SIM Selected ion monitoring

VA Ventral arm

x

List of Tables

Table 2.1: List of drugs that have been detected in flies larval and puparia

tissue

25

Table 4.1: Decomposition stages of CC1 and number of maggots

collected

50

Table 4.2: Decomposition stages of CDM and number of maggots

collected

50

Table 4.3: Decomposition stages of CC2 and number of maggots

collected

53

Table 4.4: Decomposition stages of CPM and number of maggots

collected

54

Table 4.5: Decomposition stages of CC3 and number of maggots

collected

59

Table 4.6: Decomposition stages of CMT and number of maggots

collected

59

Table 4.7: Decomposition stages of CC4 and number of maggots

collected

74

Table 4.8: Decomposition stages of CPQ and number of maggots

collected

74

xi

List of Figures

Figure 2.1: Adult flies of the species of (a) Chrysomya megacephala

(b) Chrysomya rufifacies

14

Figure 2.2: Head of fly larva (a) with definite, hard and sclerotized head

capsule (b) without definite, hard and sclerotized head

capsule

15

Figure 2.3: The appearance of larva’s body with (a) round and smooth

or short spine (b) spine-like flesh processes

16

Figure 2.4: Types of spine processes (a) with branches and feathery or

(b) without branches and spiny

16

Figure 2.5: The cephalopharyngeal skeleton which is either (a) equal (b)

unequal where the DA longer than VA

17

Figure 2.6: Posterior end of a larva with a (a) peg-like tubercles or

cones (b) flat tubercles (c) extended posterior end

17

Figure 2.7: Peritreme with (a) thin and (b) thick lining 18

Figure 2.8: Peritreme which is (a) complete and have an enclosed button

area and (b) incomplete and does not enclose button

18

Figure 2.9: Slits of posterior spiracles which is (a) sinuous and (b)

straight

19

Figure 2.10: Anterior spiracles with (a) ten marginal branches or finger-

like processes and (b) five marginal branches or finger-like

processes

19

xii

Figure 2.11: Identification characteristic of Chrysomya megacephala

with (a) view of whole larva (b) cephalopharyngeal skeleton

which the DA is longer than VA and the presence of ACS,

PS and ADP (c) twelve marginal branches or finger-like

processes (d) incomplete and thick peritreme with three

straight slits

20

Figure 2.12: Identification characteristic of Chrysomya rufifacies with (a)

view of whole larva (b) DA is equal in size with VA (c)

twelve marginal branches or finger-like processes (d) spines

with dark pointed tips (e) incomplete and thick peritreme

with three straight slits

22

Figure 2.13: Structure of dimethoate 32

Figure 2.14: Structure of malathion 33

Figure 2.15: Structure of paraquat dichloride 35

Figure 2.16: Structure of permethrin 38

Figure 3.1: Experimental setup (a) view from distance and (b) closed up

view as seen in circled area.

42

Figure 4.1: Temperature data on the study of CC1 47

Figure 4.2: Temperature data on the study of CDM 47

Figure 4.3: Fresh condition of CDM in day 3. 50

Figure 4.4: Dead blowflies at the scene. 51

Figure 4.5: Temperature data on the study of CC2 52

Figure 4.6: Temperature data on the study of CPM 53

xiii

Figure 4.7: The condition of CC2 which were (a) fresh stage in day 1

(b) bloated in day 3 (c) advanced decay in day 7 (d)

skeletonised in day 8

55

Figure 4.8: The decomposition stage of CPM which were (a) bloated in

day 3 (b) remained bloated in day 5 (c) sagged and lacerated

in day 12 (d) skeletonisation in day 33

56

Figure 4.9: Dead flies at CPM experiment area 57

Figure 4.10: Temperature data on the study of CC3 58

Figure 4.11: Temperature data on the study of CMT 58

Figure 4.12: The condition of CMT during decomposition stages which

were (a) bloated in day 2 (b) advanced decay in day 4 (c)

advanced decay in day 5 (d) skeletonised in day 6

60

Figure 4.13: Adult flies at the scene which was (a) alive (b) dead 60

Figure 4.14: Third instar of Chrysomya megacephala 61

Figure 4.15: Third instar larva of Chrysomya megacephala which having

(a) ACS and PS (b) longer DA than VA (c) ten marginal

branches or fingerlike processes (d) thick peritreme and

three straight slits

62

Figure 4.16: Third instar of Chrysomya rufifacies 63

Figure 4.17: Third instar larva of Chrysomya rufifacies with (a) equal

DA and VA (b) ten marginal branches or finger-like

processes (c) spines with dark pointed tips (d) thick

peritreme and three straight slits

63

xiv

Figure 4.18: Third instar larva of Sarcophaga species with (a) thin and

incomplete peritreme as well as narrow slits (b) no oral hook

(c) DA longer than VA (d) anterior spiracles have fourteen

marginal branches or finger-like processes

64

Figure 4.19: Third instar larva of Musca domestica with (a) strongly

sinuous slits with distinct and thick peritreme (b) seven

marginal branches or finger-like processes (c) no oral hook

(d) DA longer than VA

65

Figure 4.20: The total ion chromatogram for standard malathion (5 mg/L) 66

Figure 4.21: Mass spectrum obtained from the response peak of standard

malathion

66

Figure 4.22: The total ion chromatogram for spiked blank maggots 68

Figure 4.23: Mass spectrum of malathion in spiked blank maggots 68

Figure 4.24: The total ion chromatogram for blank maggots from CC3 69

Figure 4.25: Total ion chromatogram for maggots from CMT collected at

(a) stomach/upper body (b) lower body

70

Figure 4.26: Mass spectrum of the response peaks for maggots from

CMT

70

Figure 4.27: Temperature data on the experiment of CC4 72

Figure 4.28: Temperature data on the experiment of CPQ 73

Figure 4.29: The condition of CPQ during decomposition stages which

were (a) bloated in day 2 (b) body sagged in day 3 (c)

advanced decay in day 4 (d) advanced decay in day 5

(e) skeletonisation in day 6

75

Figure 4.30 Adult flies on CPQ 76

xv

Figure 4.31: Third instar larva of Chrysomya rufifacies with (a) spine-

like flesh processes (b) darken tips on the spine (c) no oral

hook (d) equal DA and VA (e) nine anterior spiracles (f)

incomplete peritreme with straight and broad slits

77

Figure 4.32: Third instar larva of Chrysomya megacephala with (a)

presence of PS (b) DA longer than VA, and ten marginal

branches of anterior spiracles (c) thick peritreme and three

straight slits

78

Figure 4.33: Third instar larva of Lucilia species with (a) no oral hook

(b) DA longer than VA (c) eight marginal branches or

finger-like processes (d) complete peritreme enclosed the

button area with narrow slits

79

Figure 4.34: Third instar larva of Sarcophaga species with (a) no oral

hook (b) DA longer than VA (c) thin and incomplete

peritreme with narrow slits that did not pointed towards the

opening (d) fourteen marginal branches or finger-like

processes

80

Figure 4.35: Total ion chromatogram of diene product 5 81

Figure 4.36: Mass spectra obtained from (a) analysed standard paraquat

dichloride (b) NIST08 mass spectral library

82

Figure 4.37: Total ion chromatogram of (a) spiked blank maggots (b)

blank maggots

83

Figure 4.38: Mass spectrum of spiked blank maggots 83

Figure 4.39: Total ion chromatograms for maggots from CPQ 84

Figure 4.40: Mass spectrum of maggots from CPQ 84

xvi

List of Scheme

Scheme 4.1: Reduction process of paraquat 45

Scheme 4.2 : Major fragments of malathion in electron ionisation mode 67

Scheme 4.3 : Major fragments of reduced paraquat in electron ionisation mode 81

1

CHAPTER 1

INTRODUCTION

1.1 Forensic science

Forensic science is an application of scientific techniques and principles of biology, physics

and chemistry in providing evidence to legal or related investigations in crime cases. The

word forensic is derived from Latin word forum, which means public. The term was

originally applied in ancient Rome where the senate met in a forum which is a public place

to discuss and debate political and policy issues. Technically, forensic meaning is to be

applied to public or legal concerns. Meanwhile, science is the collection of systematic

methodologies used to understand the physical world. In other words, forensic science is an

apt term for a scientist whose works are to answer questions for court through report and

testimony (Houck & Siegel, 2006).

Forensic science begins at the crime scene where an incident or crime took place. Forensic

scientists rely on the scientific method for processing and evaluating evidence (Brown &

Davenport, 2012) and their role is to bring answer to judicial questions both in criminal and

civilian affair (Chadly, 2004). Forensic science is best described as the science of linking

people, things and places that involved in criminal activities by processing the evidences

collected such as deoxyribonucleic acid (DNA), fingerprints, soils and any traces around the

crime scene (Houck & Siegel, 2006).

2

Forensic science covers broad disciplines such as criminology, pathology, anthropology,

odontology, engineering, behavioural sciences, documentations, entomology, toxicology

and many more (Houck & Siegel, 2006). All those disciplines are employed in solving crime

and information gained from the knowledge within these disciplines may lead to suspects or

new clues. A death that is unexpected or is thought to have been caused by injury or poison

is always investigated for the purpose of determining whether it was homicide or suicide

(Wright, 2005). The first two commonly asked questions are the time and the cause of death.

Due to these circumstances, forensic pathologist is the best candidates to answer the

questions. Forensic pathologist is primarily employed to investigate the deaths of persons

who died suddenly and unexpectedly or as a result of injury (Wright, 2005). However, in

some cases, some questions regarding the time of death and cause of death are difficult to be

answered by the forensic pathologist. The closest discipline that can provide answers to such

questions is forensic entomology.

1.2 Forensic entomology

Forensic entomology is the application of the study of insects and their relative arthropods

to criminal or legal cases. The entomologist studies the species identification, insect’s growth

rates and insect succession in order to determine both the location and approximate time of

victim’s death (Byrd & Cina, 2013). The knowledge of insects distribution, behaviour and

biology can also provide information on when (post-mortem interval), where and how a

person died or a crime was committed (Heo et al., 2007; Syamsa et al., 2010). The

information is vital in investigation related to humans and wildlife (Gennard, 2007). Human

corpses, whether they have been produced naturally or by foul play, are processed by insect

decomposers in the same manner as any other carrion (Syamsa et al., 2010). The insect’s

3

colonisation can appear minutes after death and persist for a long time until a body is

skeletonised (Heo et al., 2007).

Forensic entomology was first documented in the 13th century by the Chinese lawyer and

death investigator Sung Tzu in the text book “The Washing Away of Wrongs”. He described

the case of stabbing near a rice field and blowflies were drawn toward an invisible trace of

blood to a single sickle where the owner finally confessed to his crime (Benecke, 2001). In

the 18th and 19th century, medico-legal doctors observed that buried bodies were colonized

by many arthropods during mass exhumations in France. It was then concluded that maggots

play an important role in decomposition process (Benecke, 2008). In 1855, the first modern

entomology was reported by a French doctor. He introduced the knowledge on insect’s life

cycles that can provide information on the estimation of post-mortem interval (PMI)

(Benecke, 2001). Since then, several studies were focused on the life cycle of various species

of insects in various case reports. Between 1960s until 1980s, the study of insect’s life cycle

was still recognized and used by medical doctors in solving casework. Up until now, many

researches have reported on entomology and its utilization in criminal investigations

including murder and other high profile cases (Benecke, 2001).

1.2.1 Arthropods

Arthropods are the most abundant animal on the planet and easily found in a wide range of

ecosystems on earth. Arthropods are one of the most important biological group on earth

(Benecke, 2001) and play a main role as ecosystem builders and also in breaking down and

recycling organic matters in the soil (Giribet & Edgecombe, 2012). Arthropods are divided

4

into five distinct classes which are Pycnogonida (marine arthropods), Euchelicerata (spiders

and scorpions), Myriapoda (centipedes and millipedes), Crustacea (crabs and shrimps) and

Hexapoda (insects) (Giribet & Edgecombe, 2012). The largest group of arthropods is insects,

which consist of more than a million species (Myers, 2001). Insects are divided into four

dominate species which are Coleoptera (beetles), Diptera (flies), Hymenoptera (bees and

ants) and Lepidoptera (moths and butterflies). There are thousands of new species of insects

being studied and described each year (Davies, 1988). Hundreds of arthropods species are

attracted to decomposing cadaver especially flies (Diptera) and beetles (Coleoptera)

(Benecke, 2001). The most common insects found are from the order Diptera and

Coleoptera. Insects feed, live and breed in and on the carrion depending on the biological

preferences (Benecke, 2001). Therefore, insects are very useful in crime investigations to

provide information on the PMI. Apart from estimating the PMI, insects can also be used in

the determination of chemical compound presence in insect’s body such as drugs, toxins,

heavy metals and poisons which are commonly practiced by entomotoxicologist.

1.3. Toxicology

Toxicology derives from the Greek words “toxicos’ and ‘logos’ which both means poisonous

and study, respectively. It is a broad field of study which involved a multidisciplinary science

related to negative effect of chemicals to living organisms including human. In ancient days,

arsenic, lead, antimony, mandrake, hemlock, opium, aconite, animal venoms and some other

plant products were used as poison (Fenton, 2002). Nowadays, most of the toxin and poisons

are synthetic products such as pesticides and drugs (Fenton, 2002).

5

Toxicology involves two main fields which are toxicokinetics and toxicodynamics.

Toxicokinetics deals with absorption, distribution, biotransformation, metabolism and

excretion or elimination of toxic substances from the body. Whereas toxicodynamics on the

other hand, deals with the effects of toxic substance on an organism such as irritant,

corrosive, teratogenic or sterilizing agent, asphyxiation or suffocation, carcinogen, mutagen

and anaesthetic or narcotic. Toxicology can be subdivided into many subdisciplines such as

aquatic toxicology, ecotoxicology, environmental toxicology, medical toxicology,

toxicgenomics, chemical toxicology, forensic toxicology and entomotoxicology (Fenton,

2002).

1.3.1 Entomotoxicology

Entomotoxicology is a study which involves entomology and toxicology. Insects are used to

detect the presence of drugs or toxins in a decomposing tissue (Introna et al., 2001). Insect

are secondary detectors used for toxicological analysis when the conventional matrices such

as blood, urine and internal organs are no longer available (Gosselin et al., 2011a). Larvae

of flies that are found consuming a body may ingest, incorporate and bio-accumulate

chemical metabolite of drugs from the corpse into their own tissues (Wolff et al., 2001).

Entomotoxicology also investigates the effects of drugs or toxins on the development of

insects and relative arthropods (Introna et al., 2001). The major interest of entomotoxicology

is to determine the intake of drugs or toxins prior to deaths especially in skeletonised remains

(Gosselin et al., 2011a).

6

1.4 Problem statement

Suicide cases involving poison intake are not rare in Malaysia. It has become a common

method in suicidal in most developing country (Gunnel & Eddleston, 2003). Sometimes,

deaths connected to suicide mission by intake of poisons are not discovered for a period of

time ranging from few days to months. This situation usually occurs especially when the

victim lives alone and do not socialised with the neighbours. By the time the body is

discovered, the body decomposition has advanced. The loss of body fluids and internal

organs would complicate the forensic investigations which is vital in toxicological analysis

(Musvasva et al., 2001).

Necrophagous insects are the most reliable sources for toxicological analysis since these

insects feed on the body and live or breed in and on the corpse, depending on their biological

preferences and on the state of body decomposition (Kumara et al., 2010). Insects can be

analysed quite easily after homogenisation of the most representative specimens by common

toxicological procedures such as radio-immune assay (RIA), gas chromatography (GC), thin

layer chromatography (TLC), high pressure liquid chromatography mass spectrometer

(HPLC-MS) and gas chromatography mass spectrometer (GC-MS) (Introna et al., 2001).

The analyses of the community of insects encountered on decomposing tissues combined

with the knowledge of biology, ecology and local environmental conditions can provide

valuable forensic insights (Goff & Lord, 2001).