Parasitic and predatory behavior of Alysia manducator (Hymenoptera: Braconidae) on blow fly larvae feeding on an adult pig carcass in the Western Cape Province of South Africa: preliminary observations and forensic implications.

Wasps are part of the entomofauna associated with vertebrate carrion. They are known to parasitize and prey on specific life stages of insect hosts such as eggs, larvae, pupae, and/or adults associated with vertebrate carrion. However, reports of parasitic behavior of wasps on carrion-associated insect life stages and their possible forensic implications are non-existent in the Western Cape Province of South Africa. This study is part of ongoing research investigating the entomofauna and their pattern of succession on an adult pig carcass in Cape Town, South Africa. During this study, the parasitic wasp Alysia manducator was noted parasitizing and preying on blow fly larvae associated with the decomposing carcass. The arrival of A. manducator coincides with the occurrence of blow fly eggs and/or larvae on the carcass. These wasps were seen in close association with the eggs and larvae of blow flies on various parts of the carcass and some wasps were seen dragging fly larvae attached to their ovipositors away from one part of the carcass to another. Some A. manducator were also observed walking over several larvae on the carcass while exhibiting a stabbing behavior presumably in search of a host for oviposition. We suggest that the observations recorded in this study are of considerable forensic importance as the dragging effect and predatory and stabbing behavior exhibited by A. manducator could potentially disrupt the feeding and development of the fly larvae on the carcass. This could subsequently alter the process of carcass decomposition and/or affect minimum post-mortem interval estimations.

Incidental observation of bone modification by Crematogaster cf. liengmei (Hymenoptera: Formicidae) in Cape Town, South Africa.

Different taxa of insects have been reported to modify the skeletal remains of vertebrates during feeding or the creation of their larval and pupal chambers. Anecdotal reports on the modification of skeletal remains by ants exist but are rare. In this paper, a case of modification of bone due to the feeding activity of the ant, Crematogaster cf. liengmei (Hymenoptera: Formicidae), is reported. In November 2022, a bone was encountered incidentally in the vicinity of the Table Mountain National Park. The bone had been colonized by several individuals of Crematogaster cf. liengmei. Several ants were observed feeding on and removing bone particles and soft tissue remnants. A closer observation revealed striae/furrows on the epiphyseal ends of the bone. A 60-kg pig cadaver used in a winter decomposition trial in the same area was similarly colonized by this ant species, resulting in soft-tissue modification. Due to the similarity in the striae/furrows observed on the bone and the bitemarks inflicted by the ants on the right ear of the pig cadaver in the vicinity, it was concluded that the striae/furrows seen on the bone were created by the ants. Our observations show that Crematogaster cf. liengmei is an important taphonomic bio-agent that can contribute to the modification of skeletal remains in terrestrial environments.

Caught in the act: impact of Crematogaster cf. liengmei (Hymenoptera: Formicidae) necrophagous behavior on neonate pigs (Sus scrofa domesticus L.) in the Western Cape Province of South Africa.

Ants have been documented as part of the entomo-sarcosaprophagous community. They have been known to alter the process of carcass decomposition due to their ability to feed on fly eggs/larvae and create post-mortem skin injuries. However, studies on the impact of ants on decomposing carcasses are scarce, especially within the Western Cape Province of South Africa. This study was part of a research project that utilized two (2) neonate pig carcasses in each month of the year to establish baseline data on the insect species associated with decomposing carcasses in the Western Cape Province of South Africa. In the early spring (September 2020), mid-autumn (May 2021), and mid-winter (July 2021) trials respectively, several individuals of Crematogaster cf. liengmei colonized the pig carcasses shortly after deployment. There, they fed on the flesh of the carcasses and further inflicted bite marks and conspicuous post-mortem skin lesions. Following the reduction in the presence of Crematogaster cf. liengmei specifically in the mid-winter trial, non-vital bleeding was observed as a consequence of the skin lesions. In the early spring, mid- and late autumn (May 2021), and early (June 2021) and mid-winter trials respectively, Crematogaster cf. liengmei prevented the formation of large maggot masses, principally through the predation of fly eggs, larvae, and adults. The observations recorded in this study are of considerable importance in forensic investigations as the effect of the necrophagous behavior of Crematogaster cf. liengmei on decomposing remains can be misinterpreted by inexperienced investigators during crime scene investigations and may alter post-mortem interval estimations.

The effect of NucleoSpin® Forensic Filters on DNA recovery from trace DNA swabs.

Forensic DNA profiling is a globally accepted method for human identification, however, obtaining full DNA profiles from trace DNA can be challenging. The optimal recovery of DNA from trace DNA swabs is therefore crucial. Methods for extracting DNA from swabs often make use of a spin basket combined with a centrifugation step, to enhance the release of cells from the swab prior to DNA extraction. The NucleoSpin® Forensic Filter (Macherey-Nagel, Düren) is a type of spin basket, but it has not been thoroughly assessed on trace DNA samples. This study aimed to assess if the inclusion of the NucleoSpin® Forensic Filter significantly improved DNA recovery and DNA profiling success from cotton and flocked swabs used to collect trace DNA and buccal cells (control). Buccal cells and trace DNA samples were collected from 25 volunteers using each swab type (cotton and flocked) in duplicate. DNA was extracted from the samples using the NucleoSpin® DNA Forensic kit, one set with, and the other set without, NucleoSpin® Forensic Filters. DNA concentration was assessed using real time PCR, and DNA profiling was done using the PowerPlex® ESX 16 system. The inclusion of the NucleoSpin® Forensic Filters significantly improved DNA concentration for buccal cells that were collected using flocked swabs (p = 0.035). However, no significant differences were noted for trace DNA samples for either swab type. There was also no significant difference in DNA profiling success when NucleoSpin® Forensic Filters were used, regardless of swab and sample type. These results may be helpful for laboratories that are considering the NucleoSpin® Forensic Filters in the DNA extraction workflow, particularly for trace DNA samples.

Towards molecular autopsies: Development of a FFPE tissue DNA extraction workflow.

Sudden unexpected death (SUD) is a devastating event and forms a substantial proportion of the cases investigated at forensic mortuaries each year. Despite post-mortem investigations, the cause of death may remain undetermined. There is potential for these unresolved cases to benefit from retrospective molecular autopsies for investigation into genetic mutations which may have contributed towards death. Often, formalin fixed paraffin embedded tissues (FFPET) are the only archival sources of DNA available for retrospective analyses. However, extracting usable DNA from FFPET is challenging as current methods yield poor quality and quantity DNA. Thus, this study aimed to optimise DNA recovery from FFPET by investigating several variables within the DNA extraction workflow, including the selection of tissue type, number and thickness of tissue sections, deparaffinisation method, and DNA extraction kit. The quantity and quality of DNA recovered were assessed using spectrophotometry, real time PCR, digital capillary electrophoresis and DNA profiling. This study was the first to implement a nuclei quantification using microscopy to guide the selection of the best tissue type to use for DNA analysis. The use of a greater number of thinner tissue sections (100 sections, each 1 µm) significantly improved DNA concentration, purity and fragment length. Additionally, the combination of Deparaffinization Solution with the QIAamp® DNA FFPE Tissue Kit proved most favourable with a median DNA yield of 320 ng and 55% of DNA fragments greater than 400 bp. Isolated DNA was of single source, indicating no contamination in the workflow, and FFPET blocks that were stored for up to 3.5 years did not significantly affect DNA degradation (p = 0.1764). These results are especially informative for designing library preparation and sequencing workflows for determining cause of death in unresolved SUD cases.

Forensic human identification: Investigation into tooth morphotype and DNA extraction methods from teeth.

When a body is decomposed, hard tissues such as teeth may provide the only DNA source for human identification. There is currently no consensus as to the best DNA extraction method, and there is a lack of empirical data regarding tooth morphotype and condition that may impact DNA recovery. Therefore, this study sought to investigate which variables significantly improved DNA concentration, integrity and profiling success. A total of 52 human teeth were assessed, representing all tooth morphotypes from three deceased individuals. DNA was extracted using both the QIAamp® DNA Investigator Kit and the phenol-chloroform method. DNA concentration and degradation index were assessed using real time PCR, prior to conventional DNA profiling. Contrary to international guidelines promoting the use of molars, DNA profiling from molars was the least successful, with premolars, followed by canines, performing the best. The presence of fillings reduced the DNA quantity and quality obtained and may explain the poor performance of molars. DNA from the maxillae were significantly less degraded when the QIAamp® was used, although this did not influence DNA profiling success. A significant increase in DNA concentration, integrity and profiling success was observed in diseased teeth (periodontitis) compared to those without disease. This may be due to increased white blood cell presence at the site. There was no significant difference in DNA profiling success between the two DNA extraction methods. However, different teeth yielded failed DNA profiles for each extraction method, suggesting that repeated attempts, using alternative DNA extraction methods, is recommended. The recovery of additional DNA profiling information from degraded samples may help to ultimately reduce the burden of unidentified human remains.

Animal Models in Forensic Science Research: Justified Use or Ethical Exploitation?

A moral dilemma exists in biomedical research relating to the use of animal or human tissue when conducting scientific research. In human ethics, researchers need to justify why the use of humans is necessary should suitable models exist. Conversely, in animal ethics, a researcher must justify why research cannot be carried out on suitable alternatives. In the case of medical procedures or therapeutics testing, the use of animal models is often justified. However, in forensic research, the justification may be less evident, particularly when research involves the infliction of trauma on living animals. To determine how the forensic science community is dealing with this dilemma, a review of literature within major forensic science journals was conducted. The frequency and trends of the use of animals in forensic science research was investigated for the period 1 January 2012-31 December 2016. The review revealed 204 original articles utilizing 5050 animals in various forms as analogues for human tissue. The most common specimens utilized were various species of rats (35.3%), pigs (29.3%), mice (17.7%), and rabbits (8.2%) although different specimens were favored in different study themes. The majority of studies (58%) were conducted on post-mortem specimens. It is, however, evident that more needs to be done to uphold the basic ethical principles of reduction, refinement and replacement in the use of animals for research purposes.

Optimizing the use of cadavers by integrating pathology during anatomy dissection.

An accurate knowledge of anatomy, especially natural variation within individuals, is of vital clinical importance. Cadaver dissection during anatomical training may be a valuable introduction to pathology for undergraduate students, which can contribute greatly to a successful medical career. The purpose of this study was to determine the extent and type of pathology lesions in a cadaver population (n = 127) used for medical dissection. This was done to gauge whether sufficient pathology lesions representative of all the organ systems were present in the cadaver population to warrant the use of cadavers as an additional pathology learning resource. This study demonstrated a wide variety of pathology lesions in different organ systems. The respiratory system was most affected with pulmonary tuberculosis (TB) lesions being the most common finding (seen in 76% of cadavers) followed by bronchopneumonia and emphysema. Other common pathology findings included atherosclerosis, colonic diverticula, and chronic pyelonephritis. Skeletal fractures and degenerative joint disease were also noted. This study shows that cadaveric dissection offers a chance to alert and expose students to a wide variety of gross pathology and histopathology. It has been suggested that most medical students will practice in primary health care and as such more attention should be given to the pathology of commonly encountered diseases. This is particularly true for developing countries, where diseases such as TB are commonly encountered. The integration of pathology into the dissection hall may therefore be beneficial to student learning while simultaneously optimizing the use of cadaver material. Anat Sci Educ 9: 575-582. © 2016 American Association of Anatomists.

Making large class basic histology lectures more interactive: The use of draw-along mapping techniques and associated educational activities.

At Stellenbosch University, South Africa, basic histology is taught to a combination class of almost 400 first-year medical, physiotherapy, and dietetic students. Many students often find the amount of work in basic histology lectures overwhelming and consequently loose interest. The aim was to determine if a draw-along mapping activity would focus students during large class lectures. After each lecture on three basic histology tissues, a guided draw-along mapping session covering the work from the lecture was introduced in the form of a click-advance PowerPoint presentation which was used to demonstrate the unfolding of an "ideal" map. The lecturer simultaneously drew a similar map using an overhead projector allowing the students to draw their own maps on blank sheets of paper along with the lecturer. Students remained attentive during the activity and many participated in answering informal questions posed by the lecturer as the map-making session progressed. After the last session, students completed an anonymous, voluntary questionnaire (response rate of 78%). The majority of students found the draw-along maps useful (94%) and believed that its use should be continued in the future (93%). A significant increase (P < 0.001) was found in the test results of student cohorts who were given the current intervention compared to cohorts from previous years who were given mind maps as handouts only or had no intervention. The use of the draw-along mapping sessions were successful in focusing students during large class lectures while also providing them with a useful tool for their studies.

How hard is hard enough? An investigation of the force associated with lateral blunt force trauma to the porcine cranium.

Blunt force trauma forms a substantial portion of deaths worldwide. However, few studies have attempted to determine the force involved with blunt force trauma to the lateral part of the head. Nor have many studies been conducted at velocities exceeding 10 m/s. The acquisition of human tissue for experimental studies is becoming increasingly difficult. As such, the current study investigates the trauma and the force involved with cranial blunt force trauma in a porcine model. Thirty whole porcine heads were subjected to single impact tests on the fronto-parietal region at velocities ranging from 10 m/s to 25 m/s. Half the specimens were subjected to impact by a short projectile resembling a hammer head and the other half were subjected to impact with a Hopkinson pressure bar (HPB). Both implements had the same impact diameter and were machined from the same material. The HPB is an apparatus commonly used in material testing. Its use to determine fracture force in whole cranial specimens is novel. Fractures appeared similar in both the hammer tests and HPB tests. Lacerations and fractures resembled the shape of the striker surface with the most common fracture observed being a semi-circular depressed fracture. The mean peak fracture force was 7760 N (± 4150 N), with a mean displacement of 3.1mm (± 1.1mm). Peak fracture forces concur well with previous studies although no clear trend appears to exist between level of trauma and peak impact force.

The translucent cadaver: a follow-up study to gauge the efficacy of implementing changes suggested by students.

In a study conducted in 2011, the use of full body digital X-ray images (Lodox(®) Statscan(®)) and drawings were described for surface anatomy education during which suggestions were made by students on how to improve the method. Educational innovations should continuously be adjusted and improved to provide the best possible scenario for student learning. This study, therefore, reports on the efficacy of implementing some of these suggestions. Suggestions incorporated into the follow-up study included: (1) The inclusion of eight strategically placed labeled digital X-ray images to the dissection halls, (2) The placement of both labeled and unlabeled digital X-ray images online, (3) The inclusion of informal oral questions on surface anatomy during dissection, (4) The requirement of students to submit individual drawings in addition to group drawings into their portfolios, and (5) Integrating information on how to recognize anatomical structures on X-rays into gross anatomy lectures given prior to dissection. Students were requested to complete an anonymous questionnaire. The results of the drawings, tests and questionnaires were compared to the results from the 2011 cohort. During 2012, an increased usage of the digital X-rays and an increase in practical test marks in three out of the four modules (statistically significant only in the cardiovascular module) were reported. More students from the 2012 cohort believed the images enhanced their experience of learning surface anatomy and that its use should be continued in future. The suggested changes, therefore, had a positive effect on surface anatomy education.

Talking about death: implementing peer discussion as a coping mechanism to overcome fears about dissection, death, and dying.

Many studies have reported on the perceptions of medical students toward dissection. It is important to understand the feelings and symptoms experienced during dissection so that they can be adequately handled. Prior to dissection, first year students are given lectures on aspects of dissection, death and dying, and death rituals in various cultures. Two separate questionnaires, one given during the first week of dissection and another given one month into the program were then completed anonymously by dissection groups. The questions were designed to be open-ended, thereby encouraging group discussion amongst students. The questionnaires were used to determine the perception of students to dissection and to discover if these perceptions change during the dissection program. The first questionnaire revealed that students do experience fears and anxiety prior to and at the beginning of dissection; however, most of these fears dissipated by the time of the second questionnaire. One month into dissection students cited talking to peers as their main coping mechanism and fewer students mentioned emotional detachment from their cadaver as a coping mechanism, as was the case in the first questionnaire. Dissection was perceived as a positive experience by our student cohort and most students cited the main advantage of dissection as the ability to visualize organs in three dimensions. The comprehensive answers received from the students indicated that thorough discussion of feelings amongst peers occurred, introducing students to an important coping mechanism at an early stage of their learning.

The translucent cadaver: an evaluation of the use of full body digital X-ray images and drawings in surface anatomy education.

It has been noted by staff at the Faculty of Health Sciences, Stellenbosch University that medical students neglect the study of surface anatomy during dissection. This study reports on the novel use of Lodox(®) Statscan(®) images in anatomical education, particularly the teaching of surface anatomy. Full body digital X-ray images (Lodox Statscan) of each cadaver (n = 40) were provided to second year medical students. During dissection students were asked to visualize landmarks, organs, and structures on the digital X-ray and their cadaver, as well as palpate these landmarks and structures on themselves, their colleagues, and the cadaver. To stimulate student engagement with surface anatomy, dissection groups were required to draw both the normal and actual position of organs on a laminated image provided. The accuracy of the drawings was subsequently assessed and students were further assessed by means of practical identification tests. In addition, students were asked to complete an anonymous questionnaire. A response rate of 79% was obtained for the student questionnaire. From the questionnaire it was gathered that students found the digital X-ray images beneficial for viewing most systems' organs, except for the pelvic organs. Although it appears that students still struggle with the study of surface anatomy, most students believed that the digital X-rays were beneficial to their studies and supported their continued use in the future.