Determination of drug levels in larvae of Lucilia sericata (Diptera: Calliphoridae) reared on rabbit carcasses containing morphine

This study concerns the determination of morphine concentrations in fly larvae reared on rabbits administered different concentrations of morphine and a correlation between concentrations of the drug in larvae and tissues. Three rabbits (R1, R2 and R3) were given dosages of 12.5, 25.0 and 50.0 mg/h of morphine over a 3 h period via continuous ear artery perfusion. These dosages and time of perfusion were calculated to create tissue concentrations of morphine similar to those encountered in human death due to overdose. Morphine blood level plateau was attained after 1 h of perfusion. A fourth rabbit was used as a control. To evaluate drug concentrations, tissues were sampled using a coelioscopic technique. Approximately 400 eggs of Lucilia sericata, all of the same age category, were placed in eyes, nostrils and mouth of each rabbit carcass. Larvae and puparia were regularly collected from each rabbit for toxicological analysis. The concentrations of the drug in the tissues sampled were determined to be similar to those normally encountered in human overdoses and were correlated with the dosage of morphine that had been administered. Morphine was detected in all larvae and pupae fed on tissues from carcasses administered morphine, except for puparia from the colony fed on the R1 animal which received 12.5 mg/h dosage of morphine. All samples from the control rabbit were negative for morphine. Concentrations of morphine in larvae reared on rabbit carcasses containing morphine were 30 to 100 times lower than the concentrations found in the tissues. A correlation between the tissue concentrations and larval concentrations was found in only 3rd instar larvae (80 to 140 h following hatching). No correlations were found between administered dosages, tissue concentrations and younger larvae, prepuparial larvae or puparia.     

Determination of drug levels in larvae of Protophormia terraenovae and Calliphora vicina (Diptera: Calliphoridae) reared on rabbit carcasses containing morphine

Two species of blow flies (Diptera: Calliphoridae) were reared on tissues from rabbits administered different dosages of morphine. These species, Protophormia terraenovae and Calliphora vicina are among the first wave of insects colonizing a dead body. Two series of 3 rabbits were given dosages of 10, 20, and 40 mg/h of morphine over a 3 h period via ear artery perfusion. A morphine blood level plateau was attained after 1 h of perfusion. Two other rabbits were used as controls. Samples of tissues collected from rabbits using a coelioscopic technique were determined to have morphine concentrations similar to those encountered in human overdoses and were correlated with dosages of morphine administered. All samples from control rabbits were negative for morphine. Larvae and puparia of both species were regularly collected from each rabbit for toxicological analysis. Concentrations of morphine in larvae reared on rabbit carcasses containing morphine were significantly lower than concentrations found in the tissues. There was a decrease in concentration in morphine observed in transition from feeding 3rd instar larva to puparium. A correlation between larval concentration and tissue concentration was found only in feeding 3rd instar larvae.     

Determination of drug levels and the effect of diazepam on the growth of necrophagous flies of forensic importance in southeastern Brazil.

Larvae of Chrysomya albiceps (Wiedemann) and Chrysomya putoria (Wiedemann) (Diptera: Calliphoridae) were reared on tissues from rabbits administered twice the lethal dosage of diazepam in order to study the effects of this drug on the development of these two species. The rabbits were given 50mg of diazepam via ear vein infusion. From 18 to 54h, larvae feeding on tissues containing the drug developed more rapidly than larvae from the control colony for both fly species. The time required for pupariation and adult emergence was significantly greater for colony fed on tissues from diazepam dosed rabbits than for the control ones. These differences are significant for they are large enough to alter the estimate of postmortem interval based on fly development. The presence of diazepam could be detected through gas chromatography-mass spectrometer (GC-MS) in all rabbit samples and in almost all diptera samples in this experiment.     

石河子春季常见尸食性昆虫种类及丝光绿蝇发育观察

目的观察新疆石河子市区尸食性昆虫的种类及温度对优势种丝光绿蝇生长发育的影响。方法 2007年和2008年的4月15日至6月15日在石河子市（E44.18°、N86.00°）3个不同地点诱集尸食性昆虫,并鉴定分类。将优势种丝光绿蝇在不同温度下饲养,观察其体长、体重、羽化率、雌雄比、历期等生长发育特点。结果石河子地区春季3个不同地点共诱集到尸食性昆虫6目14科16种,其中以膜翅目、双翅目和鞘翅目为主;丝光绿蝇幼虫体长、体重随着温度升高而增长、增加,化蛹时又有所缩短、减轻;不同的温度对幼虫的羽化率和雌雄比影响不明显。结论本文所得数据可为新疆石河子地区的法医昆虫学及相关研究和实践提供一定参考。     

青岛地区常见嗜尸性蝇种及其季节性分布研究

目的观察青岛地区嗜尸性蝇种季节性演替规律,为死亡时间推断提供参考。方法在室外设立田间观察场所,2012年4月1日开始至2013年4月1日,每月第1天6时放置猪尸,20min后开始观察,至18时,期间每小时观察1次;第3天以后,分别于7时、12时、18时各观察1次。观察时记录尸体上出现的昆虫种类、虫态、数量并采集相应标本,进行分类鉴定。结果可以观察到有嗜尸性蝇类侵袭尸体的月份为4月至11月,共发现3科23种蝇类,其中丝光绿蝇、大头金蝇和亮绿蝇数量位列前三,分别占全部观察数量的40.71%、31.95%和7.12%。丝光绿蝇、大头金蝇中午时段出现的数量显著多于早上和傍晚。一年中各种蝇类在中午出现的数量均多于早上及晚上。4月上旬,蝇类早、晚与中午出现的数量相差显著;而在7、8月份差别较小。大部分嗜尸性蝇种集中在新鲜期和肿胀期。结论本文观察数据可为利用嗜尸性蝇类演替规律进行法医学死亡时间推断,以及建立地区嗜尸性蝇种类数据提供参考。     

贵阳地区常见尸食性蝇类mtDNA分子标记的检测

目的应用mt DNA中COI(348bp)、COII(637bp)及16Sr DNA(513bp)基因序列鉴定贵阳地区常见尸食性蝇类的种属。方法收集贵阳地区常见尸食性蝇类标本,经形态学分类鉴定后提取胸肌DNA,扩增COI(348bp)、COII(637bp)及16Sr DNA(513bp)基因片段,测序后用DNAMAN 4.0序列分析软件进行序列拼接,NCBI中的BLAST、MEGA 5.2软件包对所得序列分析建立种内及种间进化分歧率,并构建系统发育树。结果 COI种间平均进化分歧率为12.55%,种间范围在3.0%~18.6%,种内范围在0%~0.7%;COII种间平均进化分歧率为13.73%,种间范围在5.3%~21.6%,种内范围在0%~0.6%;16Sr DNA种间平均进化分歧率为4.54%,种间范围在0.4%~7.6%,种内范围在0%~0.4%。结论 COI、COII及16Sr DNA可用于尸食性蝇类的种属鉴定,该方法可作为传统分类方法的补充手段。     

The effect of antifreeze (ethylene glycol) on the survival and the life cycle of two species of necrophagous blowflies (Diptera: Calliphoridae)

Entomotoxicology involves the analysis of the presence and the effects of toxicological substances in necrophagous insects. Results obtained by entomotoxicological studies may assist in the investigation of both the causes and the time of death of humans and animals. Ethylene glycol (EG) is easy to purchase, sweet and extremely toxic. It may be consumed accidentally or purposefully, in an attempt to cause death for suicidal or homicidal intent. Several cases report fatalities of humans and animals. The present study is the first to examine the effects of EG on the survival, developmental rate and morphology of two blowfly species, (Diptera: Calliphoridae) typically found on corpses and carcasses: Lucilia sericata (Meigen) and L. cuprina (Wiedemann). Both species were reared on substrates (beef liver) spiked with three different concentrations of EG that could cause death in either a human or cat: 1/2LD₅₀ (T1), LD₅₀ (T2), 2LD₅₀ (T3), in addition to a control treatment (C) with no EG.Results of this research show that: a) both species are unable to survive if reared on a food substrate spiked with the highest concentration of EG (T3), while lower and medium concentrations (T1, T2) affect, but not prevent, the survival and the completion of the life cycle of such species; b) adults of L. sericata eclose only in C and T1, while adults of L. cuprina in both C, T1, T2; however, c) the developmental time of both species reared in T1 and T2 is statistically slower than the control; d) the body length of the immatures of both of the species reared in T1 and T2 is statistically smaller than the control.     

Determination of Coleoptera fauna on carcasses in Ankara province,Turkey

In this study, 40 species from Staphylinidae, Histeridae, Dermestidae, Silphidae, Nitidulidae and Cleridae families of Coleoptera which were found in 12 pig (Sus scrofa L.) carcasses were identified and recorded during a one-year period at the Hacettepe University Beytepe Campus located in Ankara, Turkey. According to the duration of their presence on the carcasses, 22 of these species were accepted to be important in decomposition. Their distribution over the months and the duration of their presence in the various decomposition stages over the seasons were determined.     

Bionomics of two forensically important blowfly species Chrysomya megacephala and Chrysomya putoria (Diptera: Calliphoridae) reared on four types of diet

The use of heterogeneous animal tissues for the rearing of necrophagous insect species can produce uneven biological data, which can compromise the determination of larval age and, consequently, estimates for post-mortem intervals. We investigated the development of two species, Chrysomya megacephala and Chrysomya putoria (Diptera: Calliphoridae), reared on four substrates: minced beef (control) and semi-synthetic diets with the addition of sardine, rumen or chicken eggs. No differences in total developmental times were detected among larvae reared on different diets. Length and width of larvae were partially affected by the type of food. Third instar larvae and pupae of both species were heavier on beef treatment when compared with other substrates. Overall mortality was lower when beef was used as food. Longevity of adults and sex ratio were not negatively affected by the use of diets. Egg-based diet was the least effective for both species. Given the fact that several bionomical parameters of larvae reared on diets were close to those obtained when minced beef was offered, and considering the putrid odour, frequency of contamination and lack of homogeneity of animal tissue, semi-synthetic diets can be used for rearing C. megacephala and C. putoria.     

吸毒案件中的尿吗啡检测

目的　研究吸毒人员停止使用毒品海洛因后 ,尿吗啡含量自然递减的一般规律。方法　对确证的 1 82名吸毒 (海洛因 )人员在停止使用毒品后连续 9天采集尿样 ,用放射免疫分析法检测尿吗啡含量。结果　 1 82名吸毒 (海洛因 )人员在停止吸毒后第 8天 ,尿吗啡平均含量降至 30ng/ml。 结论　在办案中 ,吸毒 (海洛因 )人员的尿样检测 ,建议最好在末次使用海洛因 8天之内     

四季尸食性蝇类参与尸体软组织分解过程初探

目的观察了解四季尸食性蝇类参与尸体软组织分解过程的规律。方法在室外自然环境中设置尸体,分别在春、夏、秋、冬季节,用肉眼观察尸体每一时段的变化及尸食性蝇类参与尸体分解的现象。结果在4个不同的季节中,尸体软组织分解过程中尸食性蝇类参与种类及虫态各不相同。通过观察获得尸体变化各阶段的大体历期,尸体变化各阶段常见的尸食性蝇种及虫态资料。结论观察所得资料可为推测晚期尸体的死亡时间提供参考。     

Forensic-medical diagnostic implication of morphine levels in the blood and urine

Toxicological characteristics are presented for 198 cases of acute parenteral poisoning with morphine and heroin. The range of their metabolites concentration in the blood and urine encountered in practice are analysed. Principal causes of death due to opiate poisoning in inpatients are shown. Opiates toxicity was assessed by the method of logit-regression and dose-effect curves for analysis of probability of death depending on opiate metabolite concentration in blood and urine. Relations between probability of death and detection of morphine in biological media of the victims are considered. Morphine concentrations in the blood and urine undoubtedly indicating morphine poisoning are determined.     

Investigation of morphine and morphine glucuronide levels and cytochrome P450 isoenzyme 2D6 genotype in codeine-related deaths

Compared to morphine and morphine-6-glucuronide (M6G), codeine and its other major metabolites codeine-6-glucuronide and norcodeine have weak affinity to opioid μ-receptors. Analgesic effects of codeine are thus largely dependent on metabolic conversion to morphine by the polymorphic cytochrome P450 isoenzyme 2D6 (CYP2D6). How this relates to toxicity and post-mortem whole blood levels is not known. This paper presents a case series of codeine-related deaths where concentrations of morphine, M6G and morphine-3-glucuronide (M3G), as well as CYP2D6 genotype, are taken into account. Post-mortem toxicological specimens from a total of 1444 consecutive forensic autopsy cases in Central Norway were analyzed. Among these, 111 cases with detectable amounts of codeine in femoral blood were identified, of which 34 had femoral blood concentrations exceeding the TIAFT toxicity threshold of 0.3mg/L. Autopsy records of these 34 cases were retrieved and reviewed. In the 34 reviewed cases, there was a large variability in individual morphine to codeine concentration ratios (M/C ratios), and morphine levels could not be predicted from codeine concentrations, even when CYP2D6 genotype was known. 13 cases had codeine concentrations exceeding the TIAFT threshold for possibly lethal serum concentrations (1.6 mg/L). Among these, 8 individuals had morphine concentrations below the toxic threshold according to TIAFT (0.15 mg/L). In one case, morphine as well as M6G and M3G concentrations were below the limit of detection. A comprehensive investigation of codeine-related fatalities should, in addition to a detailed case history, include quantification of morphine and morphine metabolites. CYP2D6 genotyping may be of interest in cases with unexpectedly high or low M/C ratios.     

Effect of murine retroviral infection on hair and serum levels of cocaine and morphine.

LP-BM5 retrovirally infected female C57BL/6J mice were administered cocaine, morphine or both by daily intraperitoneal injection for 9 weeks. Drug concentrations were measured by radioimmunoassay in serum and in hair extracts. Hair samples obtained from all drug-treated mice were positive for the drug injected, while none of the saline-treated mice had detectable drug levels in hair or serum. The average morphine concentration obtained from non-infected mice was 11 ng/mg hair whereas the amount found in the LP-BM5-infected mice was significantly higher (20 ng/mg hair). Mice injected with both morphine and cocaine were given 50% of the regular dose of each drug and drug levels in the hair of these animals were approximately half that of mice injected with the full dose of the single drug. Non-infected mice treated with both drugs had a mean value of 7 ng morphine/mg hair and 374 ng cocaine/mg hair while retrovirus-infected mice had significantly higher concentrations, 10 ng morphine/mg hair and 1160 ng cocaine/mg hair (P less than 0.001). Serum concentrations of cocaine and morphine were significantly higher (P less than 0.01) in the retrovirus-infected animals from 40 min to 1.5 h. The increased concentrations of cocaine and morphine in serum during retrovirus infection are accompanied by a significant increase in the amount of drug incorporated into the hair matrix. This change indicates that retroviral infection may influence the disposition of these drugs in the systemic circulation.     

温度对4种常见尸食性蝇类生长发育的影响及其法医学意义

为查明温度对尸食性蝇类生长发育的影响，便于推断死亡时间，在实验室恒温条件下，以常见的4种尸食性蝇类，即巨尾阿丽蝇Aldrichinagrahami、丝光鲜蝇Luciliasericata、大头金蝇Chrysomyamegacephala和肥须亚麻蝇Parasarcophagacrassipalpis为材料进行研究。结果表明，在实验温度范围内，除巨尾阿丽蝇二、三龄幼虫和丝光绿蝇三龄幼虫之外，各种类各发育阶段的历期均随温度增高而缩短，巨尾阿丽蝇蛹在12℃和30℃时停止发育。根据试验结果，利用线性回归方程对发育速率与温度间的关系进行了模拟，结果均有良好的相关性。研究结果具有推断死亡时间的法医学意义。     

SPE-LC-MS/MS法测定尿液与血液中海洛因3种代谢物

目的采用SPE-LC-MS/MS方法,同时检测尿液与血液中海洛因主要代谢物3-β-D-葡萄糖醛酸吗啡(M3G)、吗啡和O6-单乙酰吗啡(O6)。方法采用BAKERBONDTMspe Octadecyl(C18)进行提取,应用LC-MS/MS方法检测并通过MRM及内标法进行量化。结果尿液中M3G、吗啡、O6-单乙酰吗啡的最低检测限(LOD)分别为1.24pg、6.71pg、0.47pg;回收率依次为82.25±12.25%、93.75±13.25%、88.70±11.90%。血液中M3G、吗啡、O6-单乙酰吗啡的最低检测限分别为1.50pg、8.21pg、0.52pg。回收率依次为89.85±21.15%、73.70±17.90%、90.10±3.90%。结论本文所建方法同时适用于尿液与血液中海洛因主要代谢物M3G、吗啡、O6-单乙酰吗啡的提取、净化、分析。     

The illicit preparation of morphine and heroin from pharmaceutical products containing codeine: 'homebake' laboratories in New Zealand

Since 1983 a large number of small-scale illicit laboratories producing morphine and heroin from commercially available, codeine-based pharmaceutical products have been encountered in New Zealand. The codeine demethylation procedure is based on the use of pyridine hydrochloride. Very simple laboratory equipment and reagents are required and these can be utilised by people with little or no chemical background, following a recipe-like procedure. The process yields a characteristic product known as 'homebake'. This process is fully described.     

Evaluation of two immunoassay procedures for drug testing in hair samples

A preliminary initial enzyme-linked immunosorbent assay (LUCIO-Direct ELISA kit) and a preliminary DRI enzyme immunoassay were evaluated for drug detection in head hair with respect to lowered cutoff values recommended in Germany for the control of abstinence in cases of re-granting of drivers' licences. Following drug classes were included: cannabinoids, opiates, cocaine like substances, amphetamine, methamphetamine (and methylenedioxyamphetamines), methadone, and benzodiazepines. 759 analyses were performed using LUCIO-Direct ELISA kits and 936 analyses using DRI enzyme immunoassay tests. Sample size for each drug group and immunoassay test reached from 74 to 178. The LUCIO-Direct ELISA kit revealed a sensitivity of 91% for amphetamine up to 98% for methadone (methamphetamine 92%, cocaine 94%, opiates 94%, benzodiazepines 96%) and values of specificity of 72% for methadone up to 89% for amphetamine and benzodiazepines. The test was not useful for a preliminary screening for tetrahydrocannabinol (sensitivity of 65%) in consideration of a suggested cutoff of 0.02 ng/mg. The DRI enzyme immunoassay test was only useful for morphine and cocaine testing at low recommended new cutoff values (0.1 ng/mg) revealing sensitivities of 94% and 99%, respectively.     

洛非西定对大鼠吗啡药代动力学的影响研究

用放射免疫测定方法 ,研究大鼠皮下注射盐酸吗啡及盐酸吗啡合并灌服盐酸洛非西定后 ,大鼠吗啡的药代动力学。结果表明 ,在合并用洛非西定后 ,大鼠血清吗啡的T1/2Ka由 0 2 5 7h增加到 0 349h ,T1/2K由 2 5 0 2h减少到 2 2 5 0h ,Cmax由 3330 μg·L-1减少到 16 48μg·L-1,AUC由 12 390h·μg·L-1减少到 72 80 6h·μg·L-1,CL由 0 80 7L·h-1·kg-1增加到 1 374L·h-1·kg-1。洛非西定可降低吗啡的吸收及血液中吗啡的浓度 ,加快吗啡在大鼠体内的总体清除率     

Some notes on the nocturnal larviposition by two species of Sarcophaga (Diptera: Sarcophagidae)

Experiments in the month of September 2005 were performed to study nocturnal larviposition by the flesh flies (Diptera: Sarcophagidae). Two species namely Sarcophaga albiceps Meigen and Sarcophaga hirtipes Wiedemann were observed to deposit their larvae during night time as well. Therefore, possibility of night larviposition by flesh flies should always be taken into consideration while determining age of maggots.