Evaluation of the usefulness of insertion-null markers in critical skeletal remains.

Forensic DNA analysis in compromised skeletal remains may pose challenges due to DNA degradation, often resulting in partial or negative autosomal STRs profiles. To address this issue, alternative approaches such as mitochondrial DNA or SNPs typing may be employed; however, they are labour-intensive and costly. Insertion-null alleles (INNULs), short interspersed nuclear elements, have been suggested as a valuable tool for human identification in challenging samples due to their small amplicon size. A commercial kit including 20 INNULs markers along with amelogenin (InnoTyper® 21) has been developed. This study assesses its utility using degraded skeletal remains, comparing the results obtained (the number of detected alleles, RFU values, PHR, and the number of reportable markers) to those obtained using GlobalFiler™. Subsequently, the random match probability of the two profiles for each sample was determined using Familias version 3 to evaluate the power of discrimination of the results obtained from each kit. In every sample, InnoTyper® 21 yielded more alleles, higher RFU values, and a greater number of reportable loci. However, in most cases, both profiles were similarly informative. In conclusion, InnoTyper® 21 serves as a valuable complement to the analysis of challenging samples in cases where a poor or negative profile was obtained.

Performance comparison of four qPCR and three autosomal STR commercial kits from degraded skeletal remains.

This research evaluates the current DNA quantification (Quantifiler™ Trio, PowerQuant®, Investigator® Quantiplex® Pro and InnoQuant® HY Fast) and autosomal STRs amplification kits (GlobalFiler™, PowerPlex® Fusion 6 C, Investigator® 24Plex QS) using 62 degraded skeletal remains from armed conflicts (petrous bone, femur, tibia, and tooth) with several parameters (autosomal small, large, and male target, degradation index, probability of degradation, number of alleles above analytical threshold, number of alleles above stochastic threshold, RFU, peak height ratio, number of reportable loci). The best qPCR/autosomal STRs amplification tandem was determined by comparing quantification results by a DNA quantity estimation based on sample average RFU. InnoQuant® HY Fast was the most sensitive kit, and no significative differences were observed among amplification kits; however, Investigator® 24 Plex QS was found to be the most sensitive in our samples. That is why InnoQuant™ and Investigator® 24Plex QS were determined to be the best tandem.

A comparison between petrous bone and tooth, femur and tibia DNA analysis from degraded skeletal remains.

Skeletal remains are the only biological material that remains after long periods; however, environmental conditions such as temperature, humidity, and pH affect DNA preservation, turning skeletal remains into a challenging sample for DNA laboratories. Sample selection is a key factor, and femur and tooth have been traditionally recommended as the best substrate of genetic material. Recently, petrous bone (cochlear area) has been suggested as a better option due to its DNA yield. This research aims to evaluate the efficiency of petrous bone compared to other cranium samples (tooth) and postcranial long bones (femur and tibia). A total amount of 88 samples were selected from 38 different individuals. The samples were extracted by using an organic extraction protocol, DNA quantification by Quantifiler Trio kit and amplified with GlobalFiler kit. Results show that petrous bone outperforms other bone remains in quantification data, yielding 15-30 times more DNA than the others. DNA profile data presented likeness between petrous bone and tooth regarding detected alleles; however, the amount of DNA extracted in petrous bones allowed us to obtain more informative DNA profiles with superior quality. In conclusion, petrous bone or teeth sampling is recommended if DNA typing is going to be performed with environmentally degraded skeletal remains.

A comparison of five DNA extraction methods from degraded human skeletal remains.

Extracting DNA from degraded human remains poses a challenge for any forensic genetics laboratory, as it requires efficient high-throughput methods. While little research has compared different techniques, silica in suspension has been identified in the literature as the best method for recovering small fragments, which are often present in these types of samples. In this study, we tested five DNA extraction protocols on 25 different degraded skeletal remains. Including the humerus, ulna, tibia, femur, and petrous bone. The five protocols were organic extraction by phenol/chloroform/isoamyl alcohol, silica in suspension, High Pure Nucleic Acid Large Volume silica columns (Roche), InnoXtract™ Bone (InnoGenomics), and PrepFiler™ BTA with AutoMate™ Express robot (ThermoFisher). We analysed five DNA quantification parameters (small human target quantity, large human target quantity, human male target quantity, degradation index, and internal PCR control threshold), and five DNA profile parameters (number of alleles with peak height higher than analytic and stochastic threshold, average relative fluorescence units (RFU), heterozygous balance, and number of reportable loci) were analysed. Our results suggest that organic extraction by phenol/chloroform/isoamyl alcohol was the best performing method in terms of both quantification and DNA profile results. However, Roche silica columns were found to be the most efficient method.

GITAD 2020: quality assurance test through 20 years of experience.

GITAD (Grupo Iberoamericano de Trabajo en Análisis de DNA) was founded in 1998 as the first operational group of AICEF (Academia Iberoamericana de Criminalística y Estudios Forenses), formally created in 1999. The mission and the vision of GITAD are to promote the development of forensic genetics in Ibero-American countries and to achieve the maximum level of innovation and quality in each country, and with that aim, a proficiency test was developed. Since 1999, the member laboratories receive four reference samples with the objective of obtaining the genetic profile with their routine protocols, a theoretical exercise since 2003, and since 2007, it was incorporated a forensic sample, which changes every year. The consensus results and the different discrepancies are discussed in an annual meeting. This article illustrates the evolution of the proficiency test through 20 years from different points of view: the increase of participant laboratories, the evolution of the different DNA typing techniques reported by the Ibero-American participant laboratories, the challenges that the proficiency test have met, and future perspectives for a continuous improvement of the proficiency test, especially regarding its accreditation under ISO 17043.

Usefulness of Microbiome for Forensic Geolocation: A Review.

Forensic microbiomics is a promising tool for crime investigation. Geolocation, which connects an individual to a certain place or location by microbiota, has been fairly well studied in the literature, and several applications have been found. The aim of this review is to highlight the main findings in this field, including the current sample storage, DNA extraction, sequencing and data analysis techniques that are being used, and its potential applications in human trafficking and ancient DNA studies. Second, the challenges and limitations of forensic microbiomics and geolocation are emphasised, providing recommendations for the establishment of this tool in the forensic science community.