Browsing by Author "Pearce Brendon"

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    Assessment of the suitability of blood samples collected for toxicology for subsequent genetic analysis: A follow-up study after five years
    (2023) Grobbelaar, Jana; Davies, Bronwen; Pearce Brendon
    Therapeutic and recreational drug use is a common occurrence across the world. However, substance use may sometimes result in adverse drug reactions and death even when typically non-fatal drug doses are administered. This phenomenon may be caused by variants in the genes encoding drug-metabolising enzymes, which leads to altered drug metabolism and at times, toxicity. Cause or manner of death may not be apparent in these cases, even after conducting a standard autopsy and ancillary toxicological and histological investigations. A molecular autopsy may then be performed to identify an underlying genetic cause. Genetic testing is however not routinely conducted in forensic mortuaries, and historic backlogs within the National Forensic Chemistry Laboratories may delay the processing of toxicology samples by months or even years. As such, specimens that have undergone toxicological testing and were stored long-term are sometimes the only samples available to conduct subsequent molecular autopsies, should it be necessary for the cause of death determination. This study therefore aimed to assess whether blood specimens that were used for toxicological analyses could provide suitable DNA for downstream genetic analyses after an extended storage period of five years. In 2017, DNA was analysed from blood samples collected into vials containing sodium fluoride/potassium oxalate preservatives or vials without preservatives (grey and red top tubes, respectively). A subset of these vials underwent preparation for toxicological analyses at the time, prior to DNA extraction, while the remaining tubes underwent DNA extraction immediately and were stored in a molecular laboratory as controls. DNA analysis was then repeated one year later in a separate study, as well as five years later as part of the current study. DNA quantity and quality scores were significantly lower in red top tubes compared to grey top tubes, and toxicological processing did not significantly influence results. DNA concentration and quality also significantly decreased over time for all sample types. PCR amplification and Sanger sequencing results were mostly poor for red top tubes, but grey top tubes showed overall improvements in sequence quality. However, all DNA analysis results generally improved when DNA was extracted using a modified salting out method. Based on these results, it is suggested that forensic laboratories that often experience delays in sample processing should perform molecular autopsies using blood stored in sodium fluoride/potassium oxalate preservative coupled with a salting out DNA extraction method.
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    Open Access
    Investigation into X-STR haplotype frequencies for forensic human identification in South Africa
    (2023) Whittaker, Amy-Leigh; Mole, Calvin; Pearce Brendon
    The utilisation of X-chromosome short tandem repeats (X-STRs) for DNA profiling has been demonstrated to be particularly useful in resolving distant familial relations and deficiency paternity testing. The implementation of X-STRs within a medico-legal context requires baseline frequency data for the general population to allow for appropriate statistical interpretations of results. This study aimed to generate the first X-STR data for the South African population andinternally validate the Qiagen Investigator Argus X-12 QS kit. Biological samples from 781 South African individuals (517 males and 264 females) with either African, mixed, European, or Indian/Asian ancestry were processed. Statistical analyses were performed using StatsX and Arlequin. Herein, allele and haplotype frequencies and forensic parameters for the South African population are reported, as well as data related to the reproducibility, sensitivity, limit of detection, and concordance of the Investigator Argus X-12 QS kit. DXS10135 was the most informative locus, while DXS7423 was the least informative locus. The combined power of discrimination for both males and females was greater than 0.999999999. The haplotype diversity of all four linkage groups exceeded 0.993. Linkage group 1 was the most informative, with 421 unique haplotypes. Possible linkage disequilibrium was detected in five loci pairs in male samples and three loci pairs in female samples. However, it is expected that the effects of false linkage disequilibrium were present, and only loci pairs within the same linkage group may be in true linkage disequilibrium. All loci in female samples were in Hardy-Weinberg equilibrium, except DXS10148. Additionally, a total of 59 off-ladder alleles were identified. The discriminatory power of these results suggests X-STRs may be beneficial for forensic casework in South Africa. The availability of this data could allow this method to be used locally to assist with civil inheritance disputes and the identification of unknown individuals.