Persistence of blood (DNA/RNA) on shoe soles under varying casework related conditions

Blunt force traumas by footwear can result in severe and even fatal head and upper body injuries. Oftentimes, footwear impressions are only partially available and evidential value is limited. DNA evidence on shoe soles could provide crucial evidence helping to solve crimes by linking target DNA to the activity of interest. Little is known about the persistence and detectability of biological material post such offenses and the interplay of factors affecting the analytical success. In this study, we assessed the persistence of blood on shoe soles under varying parameters such as blood location, different sneakers, weather condition, gait, amount of blood, underground and step count. We applied an optimized DNA/RNA workflow adapted to micro-traces without constraints for the primary DNA pipeline. There is a high probability to link donor DNA to the shoe sole for up to 300-400 steps, regardless of the underground, blood location, and amount of blood. Depending on the sole material and the degree of abrasion of the sole, a longer blood persistence can be observed. Considering blood, 98.2% of the initial DNA amount (1 μl initial blood volume) was lost after 100 steps walked on sole areas that are in constant contact with the ground. Proportion of foreign DNA was marginal (avg. 4.4 alleles), minimizing the probability of unintentional DNA transfer in this context. RNA typing showed high specificity but lower sensitivity than presumptive tests used for body fluid identification (BFI). Luminol is essential for targeted sampling on shoe soles, as latent blood traces (>100-200 steps) provided sufficient biological material for DNA/RNA typing. The generated data help to address the activity of interest and evaluate probabilities about prevalence of target DNA important for casework implications and assessments on activity level.