Forensic DNA extraction methods for human hard tissue: A systematic literature review and meta-analysis of technologies and sample type.

DNA identification of human remains has a valuable role in the field of forensic science and wider. Although DNA is vital in identification of unknown human remains, post-mortem environmental factors can lead to poor molecular preservation. In this respect, focus has been placed on DNA extraction methodologies for hard tissue samples, as these are the longest surviving. Despite decades of research being conducted on DNA extraction methods for bone and teeth, little consensus has been reached as to the best performing. Therefore, the aim of this study was to conduct a thorough systematic literature review to identify potential DNA extraction technique(s) which perform optimally for forensic DNA profiling from hard tissue samples. PRISMA guidelines were used, by which a search strategy was developed. This included identifying databases and discipline specific journals, keywords, and exclusion and inclusion criteria. In total, 175 articles were identified that detailed over 50 different DNA extraction methodologies. Results of the meta-analysis conducted on 41 articles - meeting further inclusion criteria - showed that statistically significant higher DNA profiling success was associated with solid-phase magnetic bead/resin methods. In addition, incorporating a demineralisation pre-step resulted in significantly higher profiling successes. For hard tissue type, bone outperformed teeth, and even though dense cortical femur samples were more frequently used across the studies, profiling success was comparable, and in some cases, higher in cancellous bone samples. Notably, incomplete data sharing resulted in many studies being excluded, thus an emphasis for minimum reporting standards is made. In conclusion, this study identifies strategies that may improve success rates of forensic DNA profiling from hard tissue samples. Finally, continued improvements to current methods can ensure faster times to resolution and restoring the identity of those who died in obscurity.

Pressing need for national governmental recognition of forensic anthropology in South Africa as illustrated in a medico-legal case.

Forensic anthropology in South Africa is well developed in the higher education sector, with advanced training and research programmes. Despitethis and decades of academic involvement in casework, forensic anthropology still lacks a defined framework and mandate ata governmentallevel. Therefore, the involvement of forensic anthropologists' expertise varies markedly between cases, provinces, and among various stakeholderswithinthe country, to the detriment of dispensation of social and criminal justice. The lack ofclearly defined guidelines for the renderingof the service was exemplified and demonstrated through a recent forensic case. Here,contextual information was absent, and the remains posed a challenge to analyse, ostensibly due tomissing information. Numerous questions were raised during the analysis of the remains, and broader concerns about the investigative involvement of a forensic anthropologist within South African casework were brought to the fore. Through the analysis of this case, we describe the deductive processes that led to the formation of an opinion that the skeletal linear defects were the result of taphonomic changes. In addition, we highlighthow these efforts where constrained and each step in theprocess unnecessarily hindered. Finally, we demonstrate the capacity andwillingness offorensic anthropology practitioners to be involved, and how, withoutgovernmentalsupport, it is a great potential lamentably untapped.

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.

A first for forensic genetics in Africa: successful identification of skeletal remains from the marine environment using massively parallel sequencing.

In unrelated circumstances, two young adult males allegedly went missing off the coast of Cape Town, South Africa, within two months of each other. Weeks after the second disappearance, a decomposed human lower limb was recovered from a beach in Cape Town, followed by a washed-up decomposed hand three days later. An item of female clothing was found with the remains, and preliminary analysis of the skeleton indicated a female, leading to confusion regarding the possible identity of the decedent. Consequently, DNA analyses were requested to determine the biological sex of the remains, and whether the two sets of remains originated from the same individual. Various samples were collected, including bone, nails and swabs of soft tissue. DNA quantity and quality varied between sample types, with better results obtained from metacarpal bone and swab lysates. DNA profiling revealed a male sex, which suggests cognitive bias may have played a role in initial sex estimations. In addition, massively parallel sequencing confidently matched the two sets of remains (random match probability: 1 in 2.70 x 1031). These results were a first for Africa where massively parallel sequencing was successfully used and assisted in the identification of human remains, thus, affording closure to the next-of-kin. Moreover, this constitutes the first global report where soft tissue lysates from a marine decomposition case yielded full DNA profiles with a massively parallel sequencing approach.