Short tandem repeat (STR) markers located on the X chromosome (X-STRs) may help to resolve complex kinship cases, as they can sometimes provide more information than autosomal STRs, depending on the genetic relationship being investigated.

A multi-step study involving twelve laboratories from the Italian working group (GeFI) of the International Society for Forensic Genetics (ISFG) was conducted to examine attitudes towards and proficiency in the use of X-STRs in kinship testing. The study also aimed to gather new research data to enhance the practical application of these markers in Italian laboratories.

According to a preliminary survey, the majority of laboratories routinely performed X-STR analysis in kinship cases, but only in conjunction with autosomal STRs. Only two laboratories reported using X-STRs alone in selected pedigrees. The vast majority of laboratories employed staff who had received specific training in X-STR testing, primarily after completing ISFG/GeFI courses. Regarding the Italian population haplotype frequency database used for likelihood ratio (LR) calculations, the majority of laboratories referenced the study by Bini et al. (2015), which included 200 haplotypes.

Participating laboratories were provided with the haplotypic data of the 12 X-STRs included in the Investigator Argus X-12 kit (QIAGEN), for both the Bini 2015 Italian dataset and an expanded Italian dataset comprising an additional 1,042 haplotypes collected in a previous GeFI collaborative study. These data, together with specific information on the recombination fraction and mutation rates of the selected X-STR markers, were used to calculate LR values for a simulated pedigree provided by the organisers, using the dedicated FamlinkX software. All laboratories returned LR values within the expected order of magnitude, with minimal deviations due exclusively to typing errors in the FamlinkX input data. These errors were identified and corrected immediately.

In the second phase of the study, the effect of performing FamlinkX LR calculations using either the smaller Bini 2015 haplotype frequency database or the larger 1,242 haplotype reference database was investigated in a large set of relevant pedigrees (n > 150), including full sisters, paternal half-sisters, grandmother / granddaughter and paternal aunt / niece pairs.

Authors

• Selena Cisana (Centro Avanzato di Diagnostica "A. Bertinaria", Italy)
• Alessia Riem (Department of Public Health Sciences and Pediatrics, University of Turin, Italy)
• Serena Aneli (Department of Public Health Sciences and Pediatrics, University of Turin, Italy)
• Carla Bini (Department of Medical and Surgical Sciences, Section of Legal Medicine, University of Bologna, Italy)
• Elena Chierto (Department of Public Health Sciences and Pediatrics, University of Turin, Italy)
• Arianna Delicati (Legal Medicine Unit, Department of Cardiac, Thoracic, Vascular Sciences and Public Health, University of Padova, Italy)
• Ciro Di Nunzio (CEINGE Researcher at the Biological and Genetic Individual Typing Laboratory, University of Naples Federico II, Italy)
• Michele Di Nunzio (Department of Medicine, Legal Medicine Unit, University of Barcelona, Spain)
• Matteo Fabbri (Dipartimento di Medicina Traslazionale, Sezione Dip. di Medicina Legale, University of Ferrara, Italy)
• Paolo Fattorini (Department of Medicine, Surgery and Health, University of Trieste, Italy)
• Denise Gianfreda (Institute of Legal Medicine, Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, Italy)
• Giorgia Leone (Section of Legal Medicine, Department of Health Surveillance and Bioethics, Catholic University of the Sacred Heart, Italy)
• Martina Onofri (Forensic Sciences Laboratory, Section of Legal Medicine, Department of Medicine and Surgery, Santa Maria Hospital, University of Perugia, Italy)
• Giulia Soldati (Unit of Forensic Medicine, Department of Diagnostics and Public Health, University of Verona, Italy)
• Carlo Robino (Department of Public Health Sciences and Pediatrics, University of Turin, Italy)