Collection and interpretation of “touch DNA” from crime scenes represent crucial steps during criminal investigations, with clear consequences in courtrooms. Although the main aspects of this type of evidence have been extensively studied, some controversial issues remain. For instance, there is no conclusive evidence indicating which sampling method results in the highest rate of biological material recovery. Thus, this study aimed to describe the actual considerations on touch DNA and to compare three different sampling procedures, which were “single-swab”, “double-swab”, and “other methods” (i.e., cutting out, adhesive tape, FTA^® paper scraping), based on the experimental results published in the recent literature. The data analysis performed shows the higher efficiency of the single-swab method in DNA recovery in a wide variety of experimental settings. On the contrary, the double-swab technique and other methods do not seem to improve recovery rates. Despite the apparent discrepancy with previous research, these results underline certain limitations inherent to the sampling procedures investigated. The application of this information to forensic investigations and laboratories could improve operative standard procedures and enhance this almost fundamental investigative tool’s probative value.

The determination of the Post-Mortem Interval (PMI) is an issue that has always represented a challenge in the field of forensic science. Different innovative approaches, compared to the more traditional ones, have been tried over the years, without succeeding in being validated as successful methods for PMI estimation. In the last two decades, innovations in sequencing technologies have made it possible to generate large volumes of data, allowing all members of a bacterial community to be sequenced. The aim of this manuscript is to provide a review regarding new advances in PMI estimation through cadaveric microbiota identification using 16S rRNA sequencing, in order to correlate specific microbiome profiles obtained from different body sites to PMI. The systematic review was performed according to PRISMA guidelines. For this purpose, 800 studies were identified through database searching (Pubmed). Articles that dealt with PMI estimation in correlation with microbiome composition and contained data about species, body site of sampling, monitoring time and sequencing method were selected and ultimately a total of 25 studies were considered. The selected studies evaluated the contribution of the various body sites to determine PMI, based on microbiome sequencing, in human and animal models. The results of this systematic review highlighted that studies conducted on both animals and humans yielded results that were promising. In order to fully exploit the potential of the microbiome in the estimation of PMI, it would be desirable to identify standardized body sampling sites and specific sampling methods in order to align data obtained by different research groups.

Chimerism analysis is a well-established method for monitoring the state of hematopoietic stem cell transplantation (HSCT) over time by analyzing peripheral blood or bone marrow samples of the recipient in several malignant and non-malignant hematologic diseases. From a clinical point of view, a continuous monitoring is fundamental for an effective early therapeutic intervention. This paper provides a comparative overview of the main molecular biology techniques which can be used to study chimerism after bone marrow transplantation, focusing on their advantages and disadvantages. According to the examined literature, short tandem repeats (STR) analysis through simple PCR coupled with capillary electrophoresis (STR-PCR) is the most powerful method which guarantees a high power of differentiation between different individuals. However, other methods such as real-time quantitative PCR (qPCR), digital PCR (dPCR), and next-generation sequencing (NGS) technology were developed to overcome the technical limits of STR-PCR. In particular, these other techniques guarantee a higher sensitivity, which allows for the detection of chimerism at an earlier stage, hence expanding the window for therapeutic intervention. After a comparative evaluation of the various techniques, it seems clear that STR-PCR still remains the gold standard option for chimerism study, even if it is likely that both dPCR and NGS could supplement or even replace the common methods of STR analysis.

DNA methylation is one of the epigenetic marks which has been studied intensively in recent years for age predicting purposes in the forensic area. In order to integrate age prediction into routine forensic workflow, the purpose of this study was to standardize and optimize a DNA methylation-based protocol tailored to the Italian context. A previously published protocol and age-predictive method was implemented for the analysis of 84 blood samples originating from Central Italy. The study here presented is based on the Single Base Extension method, considering five genes: ELOVL2, FHL2, KLF14, C1orf132, now identified as MIR29B2C, and TRIM59. The precise and specific steps consist of DNA extraction and quantification, bisulfite conversion, amplification of converted DNA, first purification, single base extension, second purification, capillary electrophoresis, and analysis of the results to train and test the tool. The prediction error obtained, expressed as mean absolute deviation, showed a value of 3.12 years in the training set and 3.01 years in the test set. Given that population-based differences in DNA methylation patterns have been previously reported in the literature, it would be useful to further improve the study implementing additional samples representative of the entire Italian population.

Forensic age estimation is crucial for identifying unknown individuals and narrowing suspect pools in criminal investigations. Over the past 15 years, significant progress has been made in using biochemical, genetic, and epigenetic markers to estimate chronological age. Methods: From research on PubMed a total of 155 studies, related to advancements in age prediction techniques, were selected following PRISMA guidelines. Studies considered eligible dealt with radiocarbon dating, aspartic acid racemization, mitochondrial DNA analysis, signal joint T-cell receptor excision circles, RNA analysis, telomeres, and DNA methylation in the last 15 years and were summarized in a table. Results: Despite these advancements, challenges persist, including variability in prediction accuracy, sample degradation, and the lack of standardization and reproducibility. DNA methylation emerged as the most promising approach capable of high accuracy across diverse populations and age ranges. Multimodal methods integrating several biomarkers show promise in improving reliability and addressing these limitations. Conclusion: While significant progress has been made, further standardization, validation, and technological integration are needed to enhance forensic age estimation. These efforts are essential for meeting the growing demands of forensic science while addressing ethical and legal considerations.

Background. Sixty years after the launch of the first human into space, different studies on the physiological changes that humans undergo during dynamic flight phases and prolonged weightlessness have been undertaken. Understanding these changes is important for the creation of the preventative measures that are essential to ensuring astronaut health. Among these changes, those of the skin are frequent, despite being rarely treated during missions. The skin is a physical barrier that protects the body from pathogen invasion and environmental changes, and it harbors diverse microbial communities that form the skin microbiota. Methods. A systematic literature review of skin microbiome changes during space flight was conducted using public electronic databases (PubMed and Scopus) selecting studies published from 2015 to 2022. The systematic review was performed according to 2020 PRISMA guidelines. Results. A total of 17 studies were collected and, after screening for inclusion and exclusion criteria, eight studies were included in this review. According to the examined literature, some skin microbiota changes seems to be only temporary, in particular Gamma- and Betaproteobacteria abundance tends to decrease, while the occurrence of the Malassezia species and Firmicutes, including Staphylococcus and Streptococcus, tends to increase. At the same time, there seems to be an exchange of microorganisms between astronauts and between the confined environment and a single astronaut, with alterations in the proportion of microorganisms maintained during the flight, in particular for species such as Corynebacterium spp., Staphylococcus spp., Streptococcus spp. and Cloacibacterium spp. Given that skin contributes both to protecting the body from pathogen invasion and environmental changes and to maintaining human homeostasis, changes in the skin microbiota of astronauts might result in skin diseases. Discussion. The skin microbiota of astronauts seems to influence the microbial composition of the International Space Station, but further studies should be performed to better understand skin microbiota dynamics and to prevent the development of dermatologic conditions during space flight.