我国法医昆虫学的研究进展

法医昆虫学是应用昆虫学知识解决有关法律问题的法医学分支学科。在推断死亡时间等方面具有明显优势。本文在阐述法医昆虫学定义和研究内容基础上,综述尸体上的昆虫生态群落演替现象、昆虫发育形态学、DNA分析技术的应用、法医昆虫毒理学等法医昆虫学问题在我国的研究进展。     

A forensic entomology case from the Amazon rain forest of Brazil

The first case of application of forensic entomology in the Brazilian Amazonia is described. The corpses of 26 men were found in the rainforest in Rondonia State, Brazil. Fly larvae collected on the bodies during autopsy were identified as Paralucilia fulvinota (Diptera, Calliphoridae). No data or specimens were collected at the crime scene. At the laboratory, the larvae developed into pupae in 58 h and into adults in 110.5 h. The total development time for P. fulvinota was measured in field experiments inside the forest. The age of the larvae when collected from the bodies was estimated as the difference between the time required for them to become adults and the total development time for this species. The estimated age of the maggots and the minimum postmortem interval was 5.7 days.     

物证鉴定的技术方法确认和标准操作规程

物证鉴定中使用的技术方法直接影响和决定了物证检验结果的质量,因此对技术方法的管理是物证鉴定实验室质量保证体系的核心因素之一。开展技术方法的开发确认和内部确认并建立方法的标准操作规程,是保证实验室能够正确选择和使用科学可靠的技术方法,并获得一致结果的主要手段。运行高水平质量保证体系的物证鉴定实验室,应该在案件物证检验工作中使用经过实验室内部确认的技术方法,而且重要新技术方法还应在开发确认基础上进行内部确认,根据方法确认结果建立该方法的标准操作规程。     

Generalized additive models and Lucilia sericata growth: assessing confidence intervals and error rates in forensic entomology

Forensic entomologists use blow fly development to estimate a postmortem interval. Although accurate, fly age estimates can be imprecise for older developmental stages and no standard means of assigning confidence intervals exists. Presented here is a method for modeling growth of the forensically important blow fly Lucilia sericata, using generalized additive models (GAMs). Eighteen GAMs were created to predict the extent of juvenile fly development, encompassing developmental stage, length, weight, strain, and temperature data, collected from 2559 individuals. All measures were informative, explaining up to 92.6% of the deviance in the data, though strain and temperature exerted negligible influences. Predictions made with an independent data set allowed for a subsequent examination of error. Estimates using length and developmental stage were within 5% of true development percent during the feeding portion of the larval life cycle, while predictions for postfeeding third instars were less precise, but within expected error.     

Gene expression during blow fly development: improving the precision of age estimates in forensic entomology

Forensic entomologists use size and developmental stage to estimate blow fly age, and from those, a postmortem interval. Since such estimates are generally accurate but often lack precision, particularly in the older developmental stages, alternative aging methods would be advantageous. Presented here is a means of incorporating developmentally regulated gene expression levels into traditional stage and size data, with a goal of more precisely estimating developmental age of immature Lucilia sericata. Generalized additive models of development showed improved statistical support compared to models that did not include gene expression data, resulting in an increase in estimate precision, especially for postfeeding third instars and pupae. The models were then used to make blind estimates of development for 86 immature L. sericata raised on rat carcasses. Overall, inclusion of gene expression data resulted in increased precision in aging blow flies.     

Applicability of DNA analysis on adhesive tape in forensic casework

Adhesive tape is commonly used in crimes and is often the subject of forensic evaluation. DNA analysis of adhesive tape can provide DNA profiles of suspects. The object of this study was to evaluate the applicability of DNA analysis on adhesive tape samples in forensic casework. We retrospectively reviewed all cases involving adhesive tape or similar items received by our institute for DNA analysis during the past 11 years. From 100 forensic cases reviewed, 150 adhesive tape samples were examined. A total of 98 DNA profiles were obtained from these samples. Sixty-two of the profiles provided feasible case-relevant information. In conclusion, DNA profiling of adhesive tape samples can be useful in a variety of forensic cases.     

Testing and evaluation of 43 "noncore" Y chromosome markers for forensic casework applications

A developmental validation study was performed on three Y-STR multiplex systems, Multiplex III (MPIII), Multiplex IV (MPIV), and Multiplex V (MPV), to ascertain their potential applicability to forensic casework. MPIII contains eight Y-STRs, including DYS426, DYS435, DYS436, DYS441, DYS442, DYS446, DYS462, and Y-GATA-A10, and one InDel, YAP (DYS287). MPIV contains 21 Y-STR loci, including DYS443, DYS444, DYS445, DYS447, DYS448, DYS449, DYS452, DYS453, DYS454, DYS455, DYS456, DYS458, DYS463, DYS464, DYS468, DYS484, DYS522, DYS527, DYS531 DYS557, and DYS588. MPV contains 13 Y-STR loci, including DYS459, DYS476, DYS488, DYS513, DYS549, DYS561, DYS570, DYS575, DYS576, DYS590, DYS594, DYS598, and DYS607. Full genetic profiles were consistently obtained for all three multiplexes with 25-50 pg of male DNA. No significant amplification was observed with 1 mug of female DNA. Each multiplex permitted the determination of the number of male donors in male:male DNA admixtures. Species specificity studies demonstrated some cross-reactivity with some primate samples. Environmentally compromised blood samples produced full or partial profiles after exposure to various conditions for up to 1 year. Full profiles were recovered from simulated casework specimens including cigarette butts and postcoital cervicovaginal swabs. Population data were collected to determine individual loci gene diversity and multiplex discriminatory capacity.     

Quantifiler observations of relevance to forensic casework

The Quantifiler (QF) kit is regularly used by forensic scientists for DNA quantitation. We performed in-house validation studies which revealed some interesting observations. The QF standard displayed a two-fold difference between two different lot numbers which suggests that every standard should be tested prior to use. The Promega K562 DNA standard works well with the QF kit. c. 41% of samples that inhibited the internal PCR control (IPC) system within the QF kit still produced good Profiler Plus reactions. QIAquick was effective at removing inhibitors. The presence of dyes within casework samples were observed not to inhibit QF amplifications. Template DNA greater than 100 ng/muL appeared to inhibit the IPC. Close to identical concentration results were obtained when alternative analysis settings were used. These validation findings will assist DNA processes involved in forensic casework.     

法医昆虫学的发展与主要成就

本文介绍了国内外法医昆虫学的发展历史和取得的主要成就，并对其今后的发展趋势进行了探讨。     

Analysis of condom lubricants for forensic casework

The detection of DNA is inhibited in cases of sexual assault involving condom use. Trace evidence, including condom lubricant residues, provides crucial associative evidence in such cases. The existing Fourier transform infrared spectroscopy (FTIR) methods for lubricant analysis and detection are limited with regard to sensitivity and discrimination. The aim of this research was to establish a new method as an alternative to FTIR for the analysis of condom lubricant residues. Pyrolysis gas chromatography-mass spectrometry (PyGC-MS) and GC-MS are highly sensitive methods of analysis for a wide range of chemical substances. PyGC-MS and GC-MS were used to analyze condom lubricants in standard solution, from clean swabs and from postcoital swabs. Pyrolysis of polydimethylsiloxane (PDMS) lubricant forms cyclic products known as cyclic dimethyl siloxanes (DMS), which are separated and detected by the GC-MS. The polyethylene glycol (PEG) lubricant can be analyzed by GC-MS directly from solution. The methods of extraction and analysis presented in this paper were shown to be significantly more sensitive than FTIR for the analysis of PDMS and PEG condom lubricants. PDMS was detected as low as 1 mug in standard solution and from clean swabs using the PyGC-MS method. PEG was detected as low as 0.5 microg from standard solution and 50 mug from clean swabs using the GC-MS method. Unfortunately, we were unable to provide further discrimination between condom brands and subbrands. The methods established throughout the research were used successfully to detect condom lubricants from donated postcoital swabs. Lubricants were detected in abundance on swabs 12 h postcoitus. Recommendations are made regarding implementation of new methods for routine analysis of casework samples along with strict pyrolysis interpretation criteria to minimize the possibility of misinterpretation of false positives.     

Estimating the preappearance interval from temperature in Creophilus maxillosus L. (Coleoptera: Staphylinidae)

Abstract: Some carrion insects appear on carcasses late in decomposition. While using these late‐arriving taxa to estimate postmortem interval, it is vital to accurately estimate an interval preceding their appearance on a cadaver called the preappearance interval (PAI). This paper tests the possibilities of a PAI estimation from temperature in case of late‐arriving, predatory carrion beetle Creophilus maxillosus. The relationship between temperature and PAI of adult and larval C. maxillosus was modeled using the results of a large‐scale experiment on succession in forests of Central Europe. In both stages, PAI was found to be strongly, inversely related to temperature averaged for the duration of PAI. The PAI estimation from mean degree‐day accumulations (mDDA), estimated DDA, and “PAI*T” functions were tested using external successional data. The greatest accuracy of PAI estimates was achieved using an exponential “PAI*T” function.     

Effect of Investigator Disturbance in Experimental Forensic Entomology: Carcass Biomass Loss and Temperature

Abstract: Often carrion decomposition studies are conducted using a single carcass or a few carcasses sampled repeatedly through time to reveal trends in succession community composition. Measurements of biomass and other abiotic parameters (e.g., temperature) are often collected on the same carcasses but are rarely a focal point of the studies. This study investigated the effects that repeated sampling during experiments have on the decomposition of carrion, measured as both gross biomass (carcass plus fauna) and net biomass (carcass only), on carcasses disturbed on every visit (with weighing only or also with the collection of fauna) and on carcasses disturbed only once. Each trial lasted at least 21 days, with samples taken in triplicate. Rat carcasses used in this study were placed in the field on the same day and either weighed on every visit or ignored until a given day. Internal and ambient air temperatures were recorded on each carcass at the time of sampling and on undisturbed carcasses using temperature loggers. The presence of succession fauna did not result in significant biomass loss on most days; however, there were individual days early in decomposition (days 3 through 6) when the succession fauna comprised a large portion of the gross biomass. With the exception of biomass loss by the emigration of maggots on days 4 and 5, neither repeated weighing of the carcasses nor repeated weighing and faunal sampling of the carcasses statistically affected the rate of biomass loss. Internal temperatures of carcasses sampled repeatedly were frequently 2–5°C lower than those that had not been disturbed, and ambient temperatures differed significantly depending on the location of measurement device. Results indicate that methods used historically for biomass loss determination in experimental forensic entomology studies are adequate, but further refinements to experimental methodology are desirable.     

The effect of time after body discovery on the accuracy of retrospective weather station ambient temperature corrections in forensic entomology

Weather station data are used by forensic entomologists to estimate ambient temperatures at body discovery sites. Correlation data from sites may also be collected to correct for site and station differences. This experiment tested the accuracy over time of retrospective weather data correction using linear regression between stations and sites. Temperatures were logged at six hypothetical body discovery sites during a seven-day period for which a hypothetical body lay in situ, and a further four correlation periods. The accuracy of weather data for hypothetical body in situ periods improved after correction in 22 of 24 correlations; however, mean predicted body in situ temperatures for sites differed significantly between correlation periods. Predicted temperatures generally rose with time after body removal, which correlated with rising site temperatures accompanying seasonal change. Therefore, practitioners should be cautious in making correlations if weather patterns during correlation differ greatly from those while the body was in situ.     

Spread and habitat selection of Chrysomya albiceps (Wiedemann) (Diptera Calliphoridae) in Northern Italy: forensic implications

Abstract: Habitat selection exploited by Chrysomya albiceps during its initial spread in Northern Italy was analyzed in relation to landscape structure. The results of two short studies and a case report are here discussed. C. albiceps was not found on experimental pig carcasses in the urban area of Pavia. It was missing in the woody mountains surrounding Lecco, but it was found in the same area, at a lower altitude, within the typical “urban sprawl” landscape. It was then recorded in a natural reserve, among a rich carrion‐fly population. Indications coming from habitat selection suggest that C. albiceps has not yet saturated its potential ecological niche in newly colonized areas of Northern Italy. Factors like temperature, altitude, and interspecific competition can act as limiting factors, affecting habitat selection and distribution in newly colonized areas; the influence of those factors has to be taken into account for forensic purposes.     

Human genomic DNA quantitation system, H-Quant: development and validation for use in forensic casework

The human DNA quantification (H-Quant) system, developed for use in human identification, enables quantitation of human genomic DNA in biological samples. The assay is based on real-time amplification of AluYb8 insertions in hominoid primates. The relatively high copy number of subfamily-specific Alu repeats in the human genome enables quantification of very small amounts of human DNA. The oligonucleotide primers present in H-Quant are specific for human DNA and closely related great apes. During the real-time PCR, the SYBR Green I dye binds to the DNA that is synthesized by the human-specific AluYb8 oligonucleotide primers. The fluorescence of the bound SYBR Green I dye is measured at the end of each PCR cycle. The cycle at which the fluorescence crosses the chosen threshold correlates to the quantity of amplifiable DNA in that sample. The minimal sensitivity of the H-Quant system is 7.6 pg/microL of human DNA. The amplicon generated in the H-Quant assay is 216 bp, which is within the same range of the common amplifiable short tandem repeat (STR) amplicons. This size amplicon enables quantitation of amplifiable DNA as opposed to a quantitation of degraded or nonamplifiable DNA of smaller sizes. Development and validation studies were performed on the 7500 real-time PCR system following the Quality Assurance Standards for Forensic DNA Testing Laboratories.     

Internal validation of the 7500 Real Time PCR System for use in forensic casework in Hellenic Police

According to guidelines of the quality assurance standards for the forensic DNA testing laboratories, prior to introducing an existing DNA analysis procedure, reliability of the procedure has to be demonstrated by carrying out internal validation. In order to introduce a sensitive and accurate DNA quantification method in our laboratory, a validation study of the 7500 Real Time PCR System (Applied Biosystems) using the QUANTIFILER Human DNA Quantification Kit (Applied Biosystems) was performed. Here we report the results and the experience we have gained during this internal validation study.     

The use of Megaselia abdita (Diptera: Phoridae) in forensic entomology

This case study demonstrates the importance of the Phorid, Megaselia abdita (Schmitz), as an indicator for post-mortem interval estimation in criminal investigations involving forensic entomology where it is usually the more frequently occurring Calliphorids that are most useful. A case example is discussed where the temperatures were low for the period of time the deceased was missing.     

室外温度与室内恒温下EC随死后经历时间的变化

目的在室外环境和室内恒温条件下测定昼夜不同时间猪后腿肌肉电导率(electrical conductivity,EC)值,分析并比较两种环境条件下,EC值随死后经历时间(time since death, TSD)变化的规律。方法取5块即刻屠宰的猪后腿肌肉,均分为两份,随机分成两组,分别置于秋季室外环境和室内18℃恒温环境中。在死后10d内分别于早晚8时(每隔12h)取样,测定其浸渍液EC值。结果两种环境条件下,EC随TSD变化的趋势在整体上是一致的,二者相关性均较好(R2室外=0.971,P室外=0.004,R2室内=0.98,P室内=0.002)。室外环境温度下,肌肉的EC值在白天增长明显,夜间增长不明显,尤其环境温度低于13℃时,出现明显的平台现象,与室内恒温条件下肌肉EC值持续上升的变化趋势有显著的差异。结论无论室外环境温度还是室内恒温下,肌肉EC与TSD的相关性均较好,但在具体分析时,应考虑到夜间低温平台期造成的时间延搁。     

Quantifying the Temperature of Maggot Masses and its Relationship to Decomposition

Numerous Calliphoridae species have been observed to form larval aggregations during the feeding stage of development, resulting in localized increases in temperature. This study investigates the relationship between maggot numbers in a mass and heat generation. Single‐species aggregations (Lucilia sericata) of various sizes (50–2500 individuals) were reared in the laboratory at a constant ambient temperature of 22°C. Internal mass temperatures were recorded every 5 min throughout the feeding stage of development. Results showed that mass temperatures increased with mass numbers (p‐value < 0.001), ranging from 2.5 to 14°C above ambient. A minimum mass size of 1200 produced overall temperatures that were significantly warmer than ambient, diverging away from 22°C after c. 26 h. These results indicate that the microclimate of a mass has the potential to differ significantly from ambient, which may be influencing larval development rates and should therefore be factored into mPMI estimates to increase accuracy.