The influence of altitude on the abundance and occurrence of species of the family Calliphoridae (Diptera) in corpses in a Mediterranean area.

A carcass is defined as a temporary resource that can support high levels of diversity compared to other resources. The level of diversity often depends on the environmental conditions in which the corpse is found. Calliphoridae (Diptera) are the most important necrophagous insects used in forensic investigations because this family is common, widespread, abundant, and usually the first to colonize a corpse. However, understanding the processes responsible for variation in calliphorid species abundance and richness along gradients in ecology remains a key challenge. In this study, we analyzed the influence of altitude on the abundance and diversity of Calliphoridae species in corpses at three different altitudes (400, 900, and 1500 m a.s.l.) in a mountainous area of southeastern Spain. The results revealed a gradient of decreasing abundance with increasing altitude. An altitudinal substitution of species was observed, with Chrysomya albiceps being found at the lowest altitude and Calliphora vicina at the highest altitude. Seasonal variation was also noted, with Ch. albiceps being the dominant species in the warmer months and C. vicina being the dominant species in winter. Our results confirm the importance of Calliphorids as seasonal, altitudinal, and environmental indicators, given the wide distribution and abundance of this family. This information is of great interest on the interpretation of data in forensic practice.

Exploring the ethical dimensions in forensic entomology: From codes of conduct to professional accountability.

This article investigates a critical part of professional careers: adhering to a code of ethics that defines proper behavior within the context of one's specialized practice. It aims to improve the setting of forensic entomology work by highlighting the ethical issues that frequently emerge. The primary goal is to uncover common ethical issues involved in this field's everyday activities and provide informed advice on appropriate resolution options. This investigation dives into the complex interaction of religious beliefs, cultural aspects, and issues of ethics, evaluating their relevance in forensic entomology. It examines a wide range of ethical issues, from the ethics involved in collecting samples at crime scenes and morguesto the ethics created into scientific investigations, report writing, and court testimony. These guiding principles are critical in setting scientific standards and building public trust in forensic entomology's ability to provide exceptional services. Notably, regardless of whether the forensic entomology service provider is accredited by any validating organization, these ethical requirements remain critical, emphasizing their global application.

Morphometric analysis for determination of larval instars in Dermestes frischii Kugelann and Dermestes undulatus Brahm (Coleoptera: Dermestidae).

Dermestes frischii Kugelann, 1792 and Dermestes undulatus Brahm, 1790 are the most abundant species worldwide at outdoor or indoor crime scenes during the dry and skeletal stages of decomposition. The attribution of larval age in these beetles is problematic due to the variable number of instars, which is influenced by environmental factors. In this study, a morphometric approach was used to look for potential morphological features as evidence of larval stages. Breeding and monitoring were performed for both species in an incubator with a preset temperature of 28°C ± 0.5 without a photoperiod. Morphometric measurements were made on 10 larvae per instar for each species using length, width, and thickness parameters. Linear discriminant analysis was then used to generate decision boundaries that clearly separated larval stages. The cross-validation procedure demonstrated that the morphometric approach successfully discriminated adjacent larval stages in both species with high values of sensitivity and specificity. This less-invasive approach could improve the ability to estimate minPMI in forensic studies of Dermestidae beetles. Future studies may extend this approach to other species and establish good practices for collecting and storing specimens for morphometric analysis.

Deciphering the impact of microenvironmental factors on cuticular hydrocarbon degradation in Lucilia sericata empty Puparia: Bridging ecological and forensic entomological perspectives using machine learning models.

Blow flies (Calliphoridae) play essential ecological roles in nutrient recycling by consuming decaying organic matter. They serve as valuable bioindicators in ecosystem management and forensic entomology, with their unique feeding behavior leading to the accumulation of environmental pollutants in their cuticular hydrocarbons (CHCs), making them potential indicators of exposure history. This study focuses on CHC degradation dynamics in empty puparia of Lucilia sericata under different environmental conditions for up to 90 days. The three distinct conditions were considered: outdoor-buried, outdoor-above-ground, and indoor environments. Five predominant CHCs, n-Pentacosane (n-C25), n-Hexacosane (n-C26), n-Heptacosane (n-C27), n-Octacosane (n-C28), and n-Nonacosane (n-C29), were analyzed using Gas Chromatography-Mass Spectrometry (GC-MS). The findings revealed variations in CHC concentrations over time, influenced by environmental factors, with significant differences at different time points. Correlation heatmap analysis indicated negative correlations between weathering time and certain CHCs, suggesting decreasing concentrations over time. Machine learning techniques Support Vector Machine (SVM), Multilayer Perceptron (MLP), and eXtreme Gradient Boosting (XGBoost) models explored the potential of CHCs as age indicators. SVM achieved an R-squared value of 0.991, demonstrating high accuracy in age estimation based on CHC concentrations. MLP also exhibited satisfactory performance in outdoor conditions, while SVM and MLP yielded unsatisfactory results indoors due to the lack of significant CHC variations. After comprehensive model selection and performance evaluations, it was found that the XGBoost model excelled in capturing the patterns in all three datasets. This study bridges the gap between baseline and ecological/forensic use of empty puparia, offering valuable insights into the potential of CHCs in environmental monitoring and investigations. Understanding CHCs' stability and degradation enhances blow flies' utility as bioindicators for pollutants and exposure history, benefiting environmental monitoring and forensic entomology.

Effects of population variations and temperature on Chrysomya megacephala (Diptera: Calliphoridae) development: implications for estimating the postmortem interval.

Forensic entomology requires knowledge of the developmental rates of the species that colonize a body after death to estimate the postmortem interval (PMI). These developmental rates may vary depending not only on the species but also on the geographic location due to population differences. Therefore, the objectives of this work were to determine the developmental duration of the forensically important fly Chrysomya megacephala under constant controlled and field condition temperatures and to compare these results, through a meta-analysis, with data reported by other authors on populations from different localities. For this, C. megacephala colonies were established in the laboratory, and the duration of the life cycle was studied at two controlled temperatures (25 °C and 27 °C) and field conditions (27.5 ± 3.2 °C). Analysis of variance was performed to determine differences in developmental time and larval length between constant laboratory temperatures and field conditions. A generalized linear model was performed with predictor variables extracted from the literature (diet, relative humidity, latitude, longitude) to evaluate the effect of population variation on developmental times. The results showed significant differences in developmental times between 25 and 27 °C. As expected, the complete life cycle of C. megacephala was shorter at 27 °C. Finally, the meta-analysis suggested differences between the developmental times of different populations, based on temperature and geographic location. The results of this study provide fundamental developmental data to use C. megacephala in PMI estimations. Finally, we suggest that, when making expert reports, information from local populations should be used to determine a more accurate and reliable PMI.

First observation of Afromorgus chinensis (boheman, 1858) (Coleoptera: Trogidae) on a rabbit (Oryctolagus cuniculus L., 1758) carcass and its implications in forensic entomology.

Beetles (Coleoptera) are known to constitute forensic evidence in medico-legal investigations as their presence can be used to date human remains in almost all decomposition stages. Many forensic studies focus on the successional colonization pattern of flies (Diptera); however, beetles have not so far been studied extensively for this aspect. A beetle of the genus Afromorgus Scholtz, 1986, A. chinensis (Boheman, 1858) (Scarabaeoidea: Trogidae), was found beneath a late decaying rabbit carcass at Paya Indah Wetland, Dengkil, Malaysia, for the first time. Both genus and species are already known to occur in Malaysia from literature.

Direct relationship between evapotranspiration rate (ETO) and vertebrate decomposition rate.

When vertebrate scavenging is excluded, the Evapotranspiration Rate (ETo) of a given geographic region directly regulates the decomposition rate of unclothed vertebrate carrion, with any deviation attributed to insect activity. We conducted four decomposition experiments using pig carrion (Sus scrofa domesticus) over the span of two years (2018-2020) at a location in Davis, California. We used ETo, a variable that accounts for five climatic parameters (wind, temperature, humidity, solar radiation, and altitude) as the rate-determining variable of the decomposition process. We found ETo to have a strong (R2 = 0.98) predictive relationship with the decomposition rate. To account for maggot activity decomposing the carrion, we measured maggot weight in 2019 and 2020 using a novel method, and in 2020 we used FLIR imagery to measure maggot mass temperatures as a surrogate measurement of total maggot activity. Maggot activity was a significant predictor (p < 0.0001) of the decomposition rate, while maggot weight was not (p > 0.1). We hope to show the forensic entomology community the potential of using ETo. Future projects can incorporate ETo as a baseline to decomposition studies to determine if ETo remains the most accurate descriptor of decomposition and ultimately increase certainty in the Postmortem Interval (PMI).

Application of omics techniques in forensic entomology research.

With the advent of the post-genome era, omics technologies have developed rapidly and are widely used, including in genomics, transcriptomics, proteomics, metabolomics, and microbiome research. These omics techniques are often based on comprehensive and systematic analysis of biological samples using high-throughput analysis methods and bioinformatics, to provide new insights into biological phenomena. Currently, omics techniques are gradually being applied to forensic entomology research and are useful in species identification, phylogenetics, screening for developmentally relevant differentially expressed genes, and the interpretation of behavioral characteristics of forensic-related species at the genetic level. These all provide valuable information for estimating the postmortem interval (PMI). This review mainly discusses the available omics techniques, summarizes the application of omics techniques in forensic entomology, and their future in the field.

New developmental data of Chrysomya megacephala (Diptera: Calliphoridae) in tropical temperatures and its implications in forensic entomology.

The estimation of the postmortem interval (PMI) is an essential information in death investigations. It is necessary to know the developmental data of the most important necrophagous insect species in every geographical area. Chrysomya megacephala (Fabricius, 1794) (Diptera: Calliphoridae) is one of the most common species associated with human body decomposition, especially in the tropics, so a precise knowledge of its life cycle is crucial. However, despite its ubiquity, developmental data in a range of tropical temperatures is scarce. For this reason, the aim of this study is to provide the developmental data of the blowfly, C. megacephala, in a range of tropical temperatures and to apply these data in forensic entomology. Four temperatures were examined (i.e., 27.0°C 29.5°C, 32.0°C and 34.5°C) and the time of developments from egg to adult were recorded. To build the growth curves, five larvae per day were measured with a digital caliper. Accumulated degree-days were calculated and the isomorphen diagram for this species was plotted. As we predicted, the results showed that the life cycle of this species was affected by the increasing temperature. The implications of these new data for determining the minimum PMI in forensic entomology were discussed.

Descriptive analyses of differentially expressed proteins during intrapuparial stage based on the label-free proteomics technique between Chrysomya megacephala and Synthesiomyia nudiseta.

The lack of rapid and accurate species identification methods on pupae restricts the practical application of forensic entomology. It is a new idea to construct portable and rapid identification kits based on the principle of antigen/antibody interaction. Screening differentially expressed proteins (DEPs) of fly pupae is a basis of solving the problem. Here, we used the label-free proteomics technique to discover the DEPs and further validate using the parallel reaction monitoring technique (PRM) in the common flies. In this study, we reared the Chrysomya megacephala and Synthesiomyia nudiseta at constanttemperature, and then we sampled at least four pupae at 24 h intervals until the end of the intrapuparial stage. We found 132 DEPs between Ch. megacephala, and S. nudiseta groups, with 68 and 64 proteins being up-regulated and down-regulated between the two groups. Among the 132 DEPs, we selected five proteins having potential for further development and utilization, such as C1-tetrahydrofolate synthase, Malate dehydrogenase, Transferrin, Protein disulfide-isomerase, and Fructose-bisphosphate aldolase, for further validation using PRM-targeted proteomics, with the trends of PRM results being consistent with the label-free data for corresponding proteins. The present study investigated DEPs via the label-free technique during the pupal development in the Ch. megacephala, and S. nudiseta and provided reference data for development of rapid and accurate identification kits.

[Injuries after the biological exposure]. / Povrezhdeniya ot biologicheskogo vozdeistviya.

The aim of the scientific work is to justify the need to highlight the topic of injuries after the biological exposure in forensic medicine. The definition of the «biological trauma¼ concept is presented - it means impairment of the body structure and function as a result of a special injury effects typical for wildlife representatives (animals, plants). Biological exposure includes antigenic, toxin, allergic, bioelectric, bioorganic exposures and their combinations. Biological injuries should be distinguished from mechanical injuries caused by small, medium, and large mammals and reptiles. Options of the antemortem and postmortem biological factor effects are considered. The qualitative limits of the postmortem period are defined. A new method of forensic reconstruction of postmortem conditions is proposed. Forensic entomological, forensic microbiological examinations, and their complex combination with forensic examination are isolated as independent methods.

The effect of postmortem penetrative trauma on oviposition behavior of Calliphora vicina.

Flies in the family Calliphoridae are purported to demonstrate a strong attraction to and preferential colonization of wounds when present on human corpses or carrion. This well-circulated concept in Forensic Entomology is based on surprisingly few empirical studies that have examined the oviposition behavior of necrophagous Diptera toward wounds of vertebrate animals. In the present study, the oviposition behavior of Calliphora vicina toward piglets inflicted with postmortem sharp force trauma was examined during a 10-h test period under controlled laboratory conditions and in an outdoor urban environment. Three species of flies (C. vicina, C. livida and Cynomya cadaverina) deposited eggs on wounded and non-wound piglets during the field studies, although more than 80% of all eggs laid were by C. vicina regardless of wound status. For all species, oviposition occurred predominately on the head, including in eyes, ears, nose and mouth, and less frequently in other locations on the body. Eggs were never found in or near wounds. Similar oviposition patterns were displayed by C. vicina under laboratory conditions, with the exception for limited egg deposition in wounds under overcrowded conditions. In this latter scenario, egg deposition represented only 0.14% of the total reproductive output of adult flies. The findings do not support the contention that calliphorids preferentially oviposit in wounds resulting from sharp force penetrating trauma.

Estimating the intra-puparial period of Hydrotaea spinigera (Stein,1910) (Diptera: Muscidae) with morphological and gene expression changes.

Hydrotaea spinigera (Stein, 1910) (Diptera: Muscidae) is a forensically important sarcosaprophagous species widely distributed throughout the Oriental and Australasian regions. At the advanced decomposition stage or the skeletonize stage, the immature stages of H. spinigera, especially the pupae, can still be found in large quantities and could be used as important indicators to estimate the minimum postmortem interval (PMImin). However, there have been no studies on the intra-puparial period of this species. Herein, we studied morphological and differential gene expression changes during the intra-puparial development of H. spinigera, aiming to accurately estimate the intra-puparial age of H. spinigera. The intra-puparial morphological changes of H. spinigera were observed at seven constant temperatures ranging from 16 °C to 34 °C and divided into 12 sub-stages. Structures that could be used to estimate the intra-puparial age, such as compound eyes, mouthparts, antennae, thorax, legs, wings, and abdomen, were observed in detail, and the developmental process of each structure was divided into 5 to 10 stages. The time range of each sub-stage, or when a structure appeared, was recorded. For the gene expression section, the most suitable reference genes were screened by geNorm, NormFinder, BestKeeper, and ΔCt methods. Based on the selected reference genes, real-time quantitative PCR (RT qPCR) was used to detect the expression changes of the white and hsp90 genes with developmental time at 19 °C, 25 °C, and 31 °C. Results showed that the trend of hsp90 gene expression under different temperatures was not consistent, while white genes exhibited regular changes during development, and could thus be used for age estimation of H. spinigera. This study provides an important basis for forensic entomology to use morphological and differential gene expression for estimating the age of H. spinigera during the intra-puparial period. Moreover, the combination of the two methods can produce a more accurate minimum postmortem interval (PMImin) estimate compared to when each method is used separately.

Colonization of Artificial Substrates by Invertebrate Macrofauna in a River Ecosystem-Implications for Forensic Entomology.

Forensic entomology includes the analysis of organisms colonizing various parts of the body in order to determine the circumstances of an incident, mainly the time, place, and cause of death. The presence of insects and other arthropods on carcasses can be a source of knowledge for the judicial system. However, this type of research (on submerged bodies) is less published. The aim of our study was to analyse the qualitative and quantitative structure of macroinvertebrates colonizing potential evidence in an upland river. The experimental research involved an eight-week exposure to articles of clothing made of different materials: natural materials (bottom sediments with plants from a river), synthetic (socks), and cotton (t-shirts). Control samples of water after 2, 4, 6, and 8 weeks were taken from experiment locations in the River Bystrzyca with a tube apparatus and hand net. The results indicated that the abundance of organisms on a given substrate depended on the period of development of invertebrate macrofauna and the time of exposure of the substrates. The abundance of aquatic macrofauna on the exposed items increased in direct proportion to the duration of the experiment, which may indicate the adaptability of these organisms to new habitat conditions. Among the taxonomic groups used in forensic entomology, Diptera, Coleoptera, and Odonata were the most abundant. The remaining taxa (including Heteroptera), though not widely used in judicial proceedings, can also provide valuable information about the circumstances of an incident.

Thick quilt may severely impact the estimation of postmortem interval using forensic entomology-based methods -two case reports.

This paper introduces two indoor cases in China, characterized by bodies lying on a mattress, covered with thick quilts and wearing clothes. There were obvious deviation in the estimated minimum postmortem interval (PMImin) of the corpses using entomological methods. Based on the forensic entomology evidence from the scene, the PMImin estimated using temperature data from the nearest weather station was longer than the actual postmortem interval (PMI) based on the police investigation and the security camera footage. The most probable cause of the errors in PMImin estimation was the hindrance in heat dissipation since the corpses were covered with thick quilts while lying on the mattress. Therefore, the heat generated by the decomposition process and larval activity was hard to lose, resulting in the rapid development of insects. These case reports emphasize the importance of temperature collection in forensic entomological investigations. Our findings call for standardized temperature acquisition procedures, including which temperature measurements (body, microenvironment or ambient temperature) should be used in forensic entomological investigations when handling similar cases.

Identification of Forensically Important Carrion Beetles (Coleoptera: Staphilinidae) in China Based on COI and COII.

Unambiguous and speedy necrophagous insect species identification is common task in forensic entomological study. Carrion beetles (Staphilinidae: Silphinae) belong to a small group of Coleoptera with less than 200 species worldwide. Some species are commonly found on dead body during forensic entomological investigation. Despite some species are hard to be categorized morphologically, present DNA-based technologies offer a potential identification strategy. Here, 37 carrion beetle specimens were collected from 15 locations throughout Chinese mainland. The cytochrome oxidase subunit I (COI) and II (COII) genes among all specimens were successfully sequenced, which provided reliable markers for precise identification. Mostly, the interspecific distance could clarify the capability of these genes for identifying included carrion beetle species. Exceptions existed between close species in Nicrophorus genus (Fabricius 1775). The sequenced gene's phylogenetic analysis revealed that all carrion beetle specimens were correctly classified into eight genera, and most have relatively high supporting values (>90%). Our data gives genetic diversity and a reference for global forensically important carrion beetle species identification, as well as a conductive significance for future application of Chinese carrion beetles in forensic entomology.

Distinguishing Four Calliphoridae Species (Diptera) from Jamaica Using the Cephalopharyngeal Skeleton: Application to Forensic Investigations.

Flies of the family Calliphoridae, commonly called blow flies, are important in the decomposition process. Knowledge on their succession pattern on corpses, species identification and the duration of their life cycle stages can be useful in forensic investigations especially when estimating the post-mortem interval. We performed linear-based morphometrics on the cephalopharyngeal skeleton of four blow fly species found in Jamaica to distinguish species and determine larval development stage. We collected eggs from pigs' heads used as bait in the field and conducted rearing exercises in the laboratory. We used the internal skeletonized structure, the cephalopharyngeal skeleton, to develop a practical and efficient method for species identification. For the first instar, we found species can be differentiated using all the measurements analysed in the study. We found that the mouth hook length may be useful in distinguishing larvae in the second instar. For the larvae in the third instar, the whole length of the skeleton, from mouth hook to length of the dorsal cornue, may be useful for separating species. We provide information on the cephalopharyngeal skeleton of Lucilia lucigerens (James), a blow fly species endemic to Jamaica, for the first time. Our work provides relevant information that could be utilized for species identification and life stage determination if fly evidence is to be incorporated in forensic investigations in Jamaica.

Teaching Forensic Entomology, Forensic Anthropology, and Haematology & Serology during the COVID-19 pandemic: Practical activities for distance learning.

As the COVID-19 pandemic upended college and university instruction throughout the world, instructors were hard-pressed to find suitable alternatives for practical activities typically carried out outside of classrooms-in laboratories, workshops, clinics, and in the field. In response to this unanticipated challenge, they relied on their ingenuity to achieve pre-pandemic goals under pandemic conditions that necessitated the shift to online teaching. The Forensic Science Undergraduate Program housed in the School of Medicine of the National Autonomous University of Mexico was not exempt from this educational upheaval but, due to its interdisciplinary nature, required creating and/or adopting a wide range of activities capable of training students to perform practical tasks associated with subject areas that span the natural and social sciences, the humanities, and the law. This report aims to describe the approaches undertaken in three subjects (Forensic Entomology, Forensic Anthropology, and Hematology & Serology) by interviewing instructors and examining their teaching materials. Also, through online surveys, students' reactions to these approaches were elicited to learn about their suitability and teaching potential. Instructor's experiences during the pandemic have proven to be a rich source of ingenious solutions, with implications well-beyond the current crisis, such as creating blended or fully online courses aimed at larger numbers of students, forensic and legal professionals, and even other instructors. The wide variety of forensic sciences offers the opportunity to innovate and improve the teaching and learning of science, particularly to the benefit of students that must combine their school tasks with professional and/or family duties.