Véronique Alunni

Difficulties of sex determination from forensic bone degraded DNA: A comparison of three methods

Sex determination is of paramount importance in forensic anthropology. Numerous anthropological methods have been described, including visual assessments and various measurements of bones. Nevertheless, whatever the method used, the percentage of correct classification of a single bone usually varies between 80% and 95%, due to significant intra- and inter-population variations, and sometimes variations coming from secular trends. DNA is increasingly used in a forensic context. But forensic DNA extraction from bone raises several issues, because the samples are very often badly altered and/or in very small quantity. Nuclear DNA is difficult to get from degraded samples, according to low copy number, at least in comparison with mitochondrial DNA. In a forensic context (as in a paeleoanthropological context) DNA sex determination is usually complicated by the weak amount of DNA, the degraded nature of nucleic acids, the presence of enzymatic inhibitors in DNA extracts, the possible faint amplification of Y band and the risk of contamination during either excavation or manipulation of samples. The aim of this work was to compare three methods of DNA sex determination from bones: procedure #1 using a single PCR amplification, procedure #2 using a double PCR amplification, and procedure #3 adding bleaching for decontamination of the bone, instead of simply rubbing the bone. These processes were applied to samples of bones (49 samples coming from 39 individuals) that were in various states of post mortem alteration. The main results are the following. (i) No DNA could be extracted from three skulls (parietal bones, mastoid process), the compact bone of one rib, and the diaphysis of one femur; (ii) there was a contamination in three skulls; and (iii) the Y band did not appear in two male cases, with one of the three procedures (male tibia, procedure #2) and with procedures #2 and #3 (male femur). This study emphasises the main issue while working with altered bones: the impossibility to extract DNA in some cases, and, worth of all, the contamination of the sample or the faint amplification of Y band which leads to a wrong sex answer. Multiple and significant precautions have to be taken to avoid such difficulties.

Comparing discriminant analysis and neural network for the determination of sex using femur head measurements

The measurement of the femoral head is usually considered an interesting variable for the sex determination of skeletal remains. To date, there are few published reference measurements of the femoral head in a modern European population for the purpose of sex determination. In this study, 116 femurs from 58 individuals of the South of France (Nice Bone Collection, Nice, France) were studied. Three measurements of the femoral head were taken: the vertical head diameter (VHD), the transversal head diameter (THD) and the head circumference (HC). The results show that: (i) there is no statistical difference between the right and left femurs for each of the three measurements (VHD, THD and HC). Therefore we arbitrarily chose to use the measures from the right femurs (N=58) to pursue our experiments; (ii) the measurements of the femoral head are similar to those of contemporary American populations; (iii) the dimensions of the femoral head place the measurements of the French population somewhere between Germany or Croatia, and Spain; (iv) there is no significant secular trend (in contrast with the femoral neck diameter); (v) the femoral head measurement as a single variable is useful for sex determination: a 96.5% rate of accuracy was obtained using THD and HC measurements with the artificial neural network; and a 94.8% rate of accuracy using VHD, both with the discriminant analysis and the neural network.

Forensic aspect of cremations on wooden pyre

Three cases of cremation on open-air pyres are described. One was classified as a suicide and two as homicides. Fire duration was estimated at approximately 1 h, close to 2 h and more than 3 h, respectively. The position of the remains, the colour alteration of bone and the burned bone fractures biomechanics are discussed. Knowledge of normal burn patterns in fire and detection of perimortem lesions are essential. These three cases highlight the specific thermal alterations and burning processes in accordance with fire duration. In each case, careful investigation yielded clues as to the manner of death. Close cooperation between law enforcement and forensic pathology investigators is required in order to correctly identify the circumstances of death.

Positive identification by X-rays bone trabeculae comparison

Positive (certain, absolute) identification of human remains needs a scientific comparison between ante mortem and post-mortem biologic features, as fingerprint, odontological, radiological or DNA comparisons. X-rays comparison has been extensively used, usually comparing some peculiarities such as outlines of the bones, degenerative evolution or pathological conditions. Trabeculae comparisons are sparsely underlined in the forensic literature. We report on a case of decomposed body where fingerprint, DNA and odontological comparisons were not possible. After dissecting the leg and preparing the bones, comparison of ante mortem and postmortem trabeculae led to a positive identification. It was observed that tens of radiolucencies and radiodensities drawn by the trabeculae were useful for comparison, within a very small part of bone. In the case reported here the positive identity could have been assessed only by the comparison of the first metatarsal. The statement of positive identification needs scientific criteria that will be discussed in this article.

An amazing case of fatal self-immolation

We present a surprising case of suicide by self-immolation. A surveillance camera filmed the victims agony. We were able to ascertain that he survived 13 min after ignition. This case was the starting point for a review of the literature of the forensic elements required to correctly analyze cases of suicide by self-immolation and to establish the causal link between the burn lesions and death. The authors will focus on the forensic and medical aspects in favor of suicide by self-immolation and on the forensic aspects required to understand the ignition process, the circumstances surrounding the fire with a particular emphasis on determining whether an accelerant was doused on the victim before ignition.

Saw marks in bones: A study of 170 experimental false start lesions

This experimental study examined false starts because these are seldom discussed in the forensic literature and usually concern animal bones rather than human bones. We created 170 experimental false starts (50 on pig femurs and 120 on human femurs) using five different saws (4 with an alternating set of teeth, and one with a wavy set). Teeth per inch (TPI) ranged from 7 to 24. Saws were classified as either rip saws or universal saws. The bone lesions were studied using stereomicroscopy. This study focused on three features that are easily observed with a stereomicroscope: the minimum width of the kerf, the profile of the lesions (concave or convex), and the shape of the edges (narrow-wide pattern, necking in the middle, or straight pattern). These features proved to be useful clues to recognize the class of the saw. Our study found some variability between lesions but also some repetitive features that allow for the classification the handsaws studied according to class. There were also some significant differences in lesions between pig bones and human bones, suggesting that pig femurs (versus human femurs) are not always a good alternative for studying saw marks.

Comparison of macroscopic and microscopic (stereomicroscopy and scanning electron microscopy) features of bone lesions due to hatchet hacking trauma

This experimental study examined the lesions produced by a hatchet on human bones (tibiae). A total of 30 lesions were produced and examined macroscopically (naked eye) and by stereomicroscopy. 13 of them were also analyzed using scanning electron microscopy. The general shape of the lesion, both edges, both walls, the kerf floor and the extremities were described. The length and maximum width of the lesions were also recorded. The microscopic analysis of the lesions led to the description of a sharp-blunt mechanism. Specific criteria were identified (lateral pushing back, fragmentation of the upraising, fossa dug laterally to the edge and vertical striae) enabling the forensic expert to conclude that a hacking instrument was used. These criteria are easily identifiable using scanning electron microscopy, but can also be observed with stereomicroscopy. Overall, lateral pushing back and vertical striae visible using stereomicroscopy and scanning electron microscopy signal the use of a hacking tool.

Sex determination using the DSP (probabilistic sex diagnosis) method on the coxal bone: Efficiency of method according to number of available variables

The DSP method (probabilistic sex diagnosis) was applied to100 contemporary coxal bones from elderly individuals of the South of France. Ten variables with a posterior probability greater or equal to a 0.95 threshold were used. There was no statistical difference between right side and left side measurements. There was no mistake for sex assignment but the level of indetermination varied a great deal. It was higher in females than in males. The best combinations were obtained when using all 10 variables, some combination of 9 variables (all except SS or SIS or VEAC) or the first 8 variables. We conclude that the DSP method is of great interest in forensic anthropology, thanks to a very weak possibility of mistake when using the software for sex determination of the coxal bone. The main drawback is the level of indetermination that can be high depending on the available variables.

Bone beveling caused by blunt trauma: a case report

The authors report a fatal case of blunt trauma to the skull caused by a rib of a beach umbrella. The skull displayed a round hole in the right temporal bone with typical internal beveling. Blunt trauma mimicking a gunshot wound (round perforation of the skull with internal beveling) is very rarely reported in the forensic literature.

Fatal heat stroke in a child entrapped in a confined space

We report the case of a child succumbing to heatstroke caused by confinement in an icebox. The post mortem examination found cyanosis and hematomas indicating that the child had tried to get out of the container. The temperature of the body was higher than it should have been considering the rigor and delay before post mortem examination. The autopsy showed no significant injury and toxicological tests were negative. A physiological study etablished that death resulted from heatstroke, not a lack of oxygen or CO2 poisoning. We conclude that heat stroke should be considered as a possible mechanism of death even in the absence of context of environmental hyperthermia. We recommend that in these situations involving confinement, establishing the mechanism of death should be done not only on the basis of a detailed post-mortem examination to rule out other causes of death, but also based on complete physiological investigations.