Giovanni Arcudi

Whole genome amplification and real-time PCR in forensic casework

BackgroundWGA (Whole Genome Amplification) in forensic genetics can eliminate the technical limitations arising from low amounts of genomic DNA (gDNA). However, it has not been used to date because any amplification bias generated may complicate the interpretation of results. Our aim in this paper was to assess the applicability of MDA to forensic SNP genotyping by performing a comparative analysis of genomic and amplified DNA samples. A 26-SNPs TaqMan panel specifically designed for low copy number (LCN) and/or severely degraded genomic DNA was typed on 100 genomic as well as amplified DNA samples.ResultsAliquots containing 1, 0.1 and 0.01 ng each of 100 DNA samples were typed for a 26-SNPs panel. Similar aliquots of the same DNA samples underwent multiple displacement amplification (MDA) before being typed for the same panel. Genomic DNA samples showed 0% PCR failure rate for all three dilutions, whilst the PCR failure rate of the amplified DNA samples was 0% for the 1 ng and 0.1 ng dilutions and 0.077% for the 0.01 ng dilution. The genotyping results of both the amplified and genomic DNA samples were also compared with reference genotypes of the same samples obtained by direct sequencing. The genomic DNA samples showed genotype concordance rates of 100% for all three dilutions while the concordance rates of the amplified DNA samples were 100% for the 1 ng and 0.1 ng dilutions and 99.923% for the 0.01 ng dilution. Moreover, ten artificially-degraded DNA samples, which gave no results when analyzed by current forensic methods, were also amplified by MDA and genotyped with 100% concordance.ConclusionWe investigated the suitability of MDA material for forensic SNP typing. Comparative analysis of amplified and genomic DNA samples showed that a large number of SNPs could be accurately typed starting from just 0.01 ng of template. We found that the MDA genotyping call and accuracy rates were only slightly lower than those for genomic DNA. Indeed, when 10 pg of input DNA was used in MDA, we obtained 99.923% concordance, indicating a genotyping error rate of 1/1299 (7.7 × 10-4). This is quite similar to the genotyping error rate of STRs used in current forensic analysis. Such efficiency and accuracy of SNP typing of amplified DNA suggest that MDA can also generate large amounts of genome-equivalent DNA from a minimal amount of input DNA. These results show for the first time that MDA material is suitable for SNP-based forensic protocols and in general when samples fail to give interpretable STR results.

Identification from chest x-rays: reliability of bone density patterns of the humerus

Abstract:  A critical review of Kahana and Hiss’ study on identification from bone trabecular pattern and a test of their method conducted on the humerus are presented. Bone trabecular pattern was studied through the generation of a numerical file representing the gray scale. Using the correlation coefficient, several pairwise comparisons between numerical files were performed. The test gave nearly 30% of incorrect exclusions (the method did not recognize couples of radiographs belonging to the same subject) and 50% of misidentifications (the method recognized couples of radiographs belonging to different subjects, as belonging to the same subject); therefore, this research shows that at the present time, it is not possible to safely quantify identification through bone density patterns, of the proximal humerus taken from thoracic X‐rays. Thus, an “easy”—but dangerous—use of trabecular density patterns on this specific type of radiogram as an identification method should be currently avoided.

Differential diagnosis between cranial fractures of traumatic origin and explosion fractures in burned cadavers.

Interpreting the damages on a cadaver which has been exposed to the action of fire is a challenge for the forensic pathologist. The finding of a burned body gives in fact the opportunity to make a series of inquiries which deal with the whole medical-legal practice. In our case, the victim has been hit over the head with a blunt object and has been then carbonized. The medical-legal problems implied lie in the definition of the exact manner and cause of death, and therefore in the analysis of vital, perimortal and postmortal injuries, as to formulate a differential diagnosis between the former traumatic originated and the latter caused by combustion.

Aortic dissection and cocaine use

Most of the cocaine - deaths are said to be related to cardiovascular complications. This paper addresses a rather infrequent complication of chronic cocaine use, represented by the aortic dissection. The case in point pertains to a 45-year-old, caucasian male, substance abuser who suffered an aortic dissection following the use of cocaine. Blood concentrations of cocaine and benzoylecgonine were considered not to be within a potentially toxic range.

Frequency assessment of 25 SNPs in five different populations

Allele and genotype frequencies of 25 SNPs previously selected and validated for forensic purposes were assessed in 250 unrelated individuals originating from five different countries of Europe (Spain, Croatia, Bulgaria, Turkey and Serbia). All the SNPs generated extremely low F(st) values confirming our previous results on Italian, African (Benin) and Asian (Mongolian) populations. As a consequence of such F(st) values we observed similar values of random match probability across the populations: 2.26 x 10(-10) in the Spanish population, 2.13 x 10(-10) in the Croatian population, 4.21 x 10(-10) in the Bulgarian population, 2.52 x 10(-10) in the Serbian population and 1.46 x 10(-10) in the Turkish population.

Spontaneous corification: a case of anomalous decomposition

We report a case of a natural corification of a woman found dead in her house after two years from her disappearance. The subject was a 63-year-old woman found in an apartment of a condominium located in the outskirts of Rome. Forensic autopsy showed a very well preserved corpse through a corification process with no external injuries observed. Corification is a special transformation process observed in corpses closed in metal lined coffins. This case report highlights the unusual characteristics of this special form of decomposition found in a domestic setting. The process producing this previously never encountered phenomenon is discussed.

An unusual case of suicidal double ligature strangulation

As suicide by ligature strangulation is rare, the correct medico-legal diagnosis may be difficult to establish if there is no clear supporting circumstantial evidence. We report an unusual case of suicide by strangulation with a double ligature. Investigation of the scene of death and pathological findings at autopsy are reported and discussed. We emphasise the importance of an external examination and an on-the-spot investigation.

Genetic Analysis of Ten Gonosomal STR Loci in an Italian Population Using the Elucigene QSTR-XY Amplification Kit

Genotyping of X-chromosomal short tandem repeats (X-STRs) is an emerging tool in forensic genetics because of its inheritance pattern, and a large number of markers has been characterized. Quantitative fluorescence polymerase chain reaction (QF-PCR) analyses of STR markers on the X-chromosome are performed routinely in medical genetics laboratories for the rapid detection of aneuploidy in chromosome X. In this study, 595 Italian participants were genotyped at 10 gonosomal STRs (DXS680, DXS98, DXS6807, DXS1187, XHPRT, DXS742, DXS6809, DXYS267, DXYS218, and DYS448) using a commercially available QF-PCR kit. Here, we report the allele architecture of DXS1187 and DXYS218, which have not previously been characterized for forensic use. The presence and extent of genetic linkage and linkage disequilibrium between all X-STRs were estimated. Allele and haplotype frequencies in the Italian population were assessed and reported together with statistical parameters.