Background and objectives: Sudden cardiac death (SCD) is a natural and unexpected death of cardiac origin that occurs within 1 h from the onset of acute symptoms. The major leading causes of SCD are cardiomyopathies and channelopathies. In this review, we focus on channelopathies, inherited diseases caused by mutations affecting genes encoding membrane ion channels (sodium, potassium or calcium channels) or cellular structures that affect Ca²⁺ availability. The diagnosis of diseases such as long QT, Brugada syndrome, short QT and catecholaminergic polymorphic ventricular tachycardia (CPVT) is still challenging. Currently, genetic testing and next-generation sequencing allow us to identify many rare alterations. However, some non-coding variants, e.g., splice-site variants, are usually difficult to interpret and to classify. Methods: In our review, we searched for splice-site variants of genes involved in channelopathies, focusing on variants of unknown significance (VUSs) registered on ClinVar up to now. Results: The research led to a high number of splice-site VUSs of genes involved in channelopathies, suggesting the performance of deeper studies. Conclusions: In order to interpret the correlation between variants and pathologies, we discuss experimental studies, such as RNA sequencing and functional analysis of proteins. Unfortunately, as these in vitro analyses cannot always be performed, we draw attention to in silico studies as future perspectives in genetics. This review has the aim of discussing the potential methods of detection and interpretation of VUSs, bringing out the need for a future reclassification of variants with currently unknown significance.

Background/Objectives: SUDEP is the sudden, unexpected death of someone with epilepsy, and occurs mainly during sleep or at rest, or when the individual does not seem to have experienced a convulsive seizure. The cause of death in SUDEP is still unknown, and it may differ between cases. Cardiac factors are among the most prevalent causes observed in SUDEP. Therefore, within the forensic medicine framework, identifying well-known DNA markers involved in cardiac sudden and unexpected death would aid in understanding the cause of SUDEP, as well as in finding cardiac risk markers in patients with epilepsy. The purpose of this study was to identify any genetic variants by analyzing blood and formalin-fixed paraffin-embedded (FFPE) tissue samples, utilizing next-generation sequencing techniques. Methods: We investigated five cases of SUDEP that were examined at the Legal Medicine department of Ancona (Italy). Peripheral blood or FFPE cardiac tissues were collected, and different DNA extraction methods were performed. In particular, this study underlines a new extraction method from FFPE tissue, adapting the Casework kit for forensic application to our purpose. Later, about one hundred genes correlated to inherited cardiac diseases were sequenced through the Ion PGM System and Ion GeneStudio S5 Systems. Results: Bioinformatic analysis showed some genetic variants of unknown significance (VUS) on genes involved in SUDEP: RYR2, SCN8A, and AKAP9. Conclusions: As expected, very low coverage of the target base was observed for FFPE tissue samples because of the complexity of the biological material. Therefore, the presence of any significant variants in unamplified regions cannot be excluded in the FFPE samples. As suggested by the literature, the variants found in the blood samples are potentially associated with SUDEP.

Background/Objectives: The study of DNA transfer and persistence has become increasingly significant, driven by advancements in DNA detection sensitivity and the need for reliable forensic evidence. In forensic investigations, saliva and saliva-stained materials are recognised as valuable DNA sources, particularly in cases of homicide, sexual assault, and burglary, where saliva can be transferred between individuals during the criminal act. The time between the crime and sample collection is a critical factor that can influence the success of the analysis. The value of the specimens collected from the victim’s skin or mouth (perilabial and labial sites, teeth and tongue) after the crime has not been investigated with currently used highly sensitive and specific molecular methods. Methods: On the assumption that a significant loss of DNA occurred, in our study, 10 voluntary pairs were tested at different time points after intense kissing and samples were taken from the above-mentioned sites to assess the presence of the donor’s DNA. Extracted DNA was quantified using the Plexor HY System kit (Promega), and both autosomal STRs and Y-STRs were analysed. Results: The results reveal a greater persistence of male DNA on the female partner, particularly in the labial and perilabial regions, even up to 120 min after contact, in terms of both concentration and duration. Conclusions: This study emphasises the forensic importance of salivary DNA as a solid source of evidence, particularly in investigations involving mixed DNA profiles.

Background: Left ventricular noncompaction (LVNC) is a myocardial abnormality char-acterized by prominent trabeculations, a thin compacted layer, and deep intertrabecular recesses. LVNC is a heterogeneous myocardial disorder of uncertain etiology, which may occur as a congenital or acquired condition or develop in association with other cardio-myopathies. Genetic factors have also been implicated in its pathogenesis. Objectives: The aim of this study was to investigate the genetic background of patients with LVNC through next-generation sequencing (NGS) analysis and to identify potentially dis-ease-associated variants. Methods: Twenty-five patients with LVNC underwent genetic testing by NGS using the Ion Torrent™ platform. Variants identified by NGS were con-firmed by Sanger sequencing. The interpretation was performed according to ACMG guidelines using multiple bioinformatic and clinical databases. Results: Genetic variants were identified in 13 patients, with 14 variants detected in 9 genes (TTN, MYH7, CTNNA3, DSG2, FLNC, JPH2, NKX2-6, TNNC1, and JUP). Eight variants (57%) were nov-el, four (29%) had been previously reported in clinical cases without scientific discussion, and two (14%) were already described in the literature. According to ACMG criteria, vari-ants were classified as pathogenic (n=1), likely pathogenic (n=7), or variants of uncertain significance (n=5). Conclusions: These findings highlight the marked genetic heterogenei-ty of LVNC and support the role of genetic testing in improving diagnosis, risk stratifica-tion and clinical management.

This Special Issue on advanced research in forensic genetics aims to showcase recent methodological developments, emerging challenges, and future perspectives in a field that is rapidly evolving under the pressure of technological innovation and increasing interpretative complexity

Sudden cardiac death (SCD) is a natural death of cardiac origin and accounts for millions of deaths worldwide each year, representing a major public health issue. Despite its significance, scientific investigation into SCD is often limited. Not all laboratories collect biological samples, such as blood or tissue, for genetic testing, and even when such analyses are performed, pathogenic or clinically relevant genetic variants are not always detected or fully informative. This review aims to highlight biological specimens that are frequently overlooked and underutilized in genetic analyses due to their complex nature: formalin-fixed, paraffin-embedded (FFPE) tissues. Although FFPE samples are suboptimal for traditional genetic investigations, they often represent the only available material when fresh or properly frozen tissue has not been collected. The main limitation in using FFPE tissues arises from formalin-induced modifications that can compromise DNA quality. Recent research, however, has identified novel biomarkers of interest, particularly microRNAs (miRNAs), short non-coding RNAs that remain stable in their expression even under suboptimal environmental conditions. The focus of this review is to emphasize the potential of FFPE tissues, historically used for histological studies, as valuable sources for innovative molecular analyses employing these novel biomarkers. By leveraging miRNAs and other emerging molecular targets, FFPE specimens could provide insights previously inaccessible through conventional DNA-based testing, thereby expanding the toolbox for postmortem investigations in SCD. This review underlines the importance of reconsidering archived FFPE tissues not merely as historical or morphological resources, but as promising matrices for cutting-edge molecular and forensic research.