Preliminary Study on the Possibility to Detect Virus Nucleic Acids in Post-Mortem Blood Samples.

BACKGROUND: In many forensic cases, the medical records of the deceased are not available at the time of the autopsy; therefore, no information about the deceased's state of health, including any infectious diseases contracted during life, is accessible. The detection of some of the principal viral infections, such as hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus type 1 (HIV-1), could contribute to determining causes of death and interesting applications could be found in medico-legal practice, such as occupational risk assessment. To date, accurate and sensitive serological and molecular assays capable of detecting these viruses have been validated on biological samples taken from living beings, while their efficiency on forensic post-mortem biological samples has yet to be thoroughly assessed. To further this aim, this study evaluated whether the nucleic acid amplification techniques (NAATs) for the detection of viral genomes that are applied in clinical settings can be used, with the same success rate, for these latter samples. METHODS: Manual viral nucleic acid extraction processes and fully-automated amplification-based detection techniques developed in-house were evaluated on blood samples taken during the routine autopsies of 21 cadavers performed 2 to 9 days after death. Information on HBV, HCV, and HIV-1 seropositive status was previously known for only four of these cadavers. RESULTS: Using automated quantitative real-time PCR (qPCR) and qualitative PCR (end-point) analyses, it was possible to confirm the presence of viral genomes in the four post-mortem whole blood samples with previously reported specific serological positivity. In addition, the genomes of HCV and/or HIV-1 genomes were detected in three other blood samples with unknown serological status at the time of autopsy. CONCLUSIONS: Therefore, our findings suggest that molecular assays may detect the presence of viral genomes in forensic post-mortem blood samples up to five days after death. This provides an additional means of investigation that can contribute to the determination of the deceased's cause of death.

Monozygotic twins: Identical or distinguishable for science and law?

Monozygotic twins, also known as monovular twins, share an identical genetic heritage because they are two individuals who derive from the same zygote. For this reason, they have been considered indistinguishable. They represent a limit for the application of markers and analytical methods that are routinely used in forensic science because analyses of DNA fragments (short tandem repeats analysed by capillary electrophoresis) are unable to distinguish monozygotic twins. The recent introduction of ultra-deep next generation sequencing in forensic genetics, also known as massively parallel sequencing, has made it possible to identify a number of genetic variations through genome sequencing (such as copy number variations, single nucleotide polymorphisms and DNA methylation) that make it possible to distinguish monozygotic twins. Here, we present a case of ascertaining biological paternity, in which the alleged father had a monozygotic twin brother. This case led to the examination of international law in similar cases in which the only available biological evidence derives from classical forensic genetic analysis, performed with short tandem repeat (autosomal and/or gonosomal) capillary electrophoresis and the probative value, if recognised, of the next generation sequencing technology in the courtroom.

Inhibition of vaginal transmission of HIV-1 in hu-SCID mice by the non-nucleoside reverse transcriptase inhibitor TMC120 in a gel formulation.

OBJECTIVE: The development of drugs that can be used as topical microbicides is currently recognized as a priority area of research. DESIGN: A preclinical evaluation of the potential effectiveness of TMC120, a non-nucleoside reverse transcriptase inhibitor (NNRTI), as a topical microbicide to prevent vaginal HIV-1 transmission in a humanized severe combined immunodeficient (hu-SCID) mouse model. METHODS: Reconstituted mice received an intravaginal application of a TMC120-containing gel 20 min prior to a non-invasive vaginal challenge with cell-associated HIV. The possible cytotoxic effect of TMC120-containing-gel on lymphocytes was assessed and their in vivo migration was followed using fluorescently labelled human lymphocytes. Systemic infection was monitored by p24 antigen detection in culture supernatant from cocultured intraperitoneal cells using antigen capture enzyme-linked immunosorbent assay test and by the presence of integrated proviral HIV-1 DNA in DNA extracted from spleen cells. In vivo migration of labelled lymphocytes was examined by analysis of cells isolated from regional lymph nodes. RESULTS: In this model, systemic infection was successfully inhibited by the presence of TMC120-containing gel at vaginal level. The in vivo migration of human lymphocytes from the vagina to regional lymph nodes, following the deposition of TMC120-containing gel, excluded the possibility that inhibition of systemic infection was a result of NNRTI toxicity. CONCLUSIONS: Vaginal transmission of HIV was successfully prevented by the application of a gel formulation containing TMC120. This is the first evidence of the in vivo effectiveness of a microbicide preparation containing an NNRTI against cell-associated HIV.