Determination of human identity from Anopheles stephensi mosquito blood meals using direct amplification and massively parallel sequencing.

DNA obtained from biological evidence can link individuals to a crime scene. DNA is typically obtained from body fluids deposited on various substrates such as fabric or common household objects. However, other unusual sources of human biological material can also be used to generate DNA profiles. Here we show that short tandem repeat (STR) DNA profiles can also be obtained from single source and mixtures of human DNA in the blood meals of Anopheles stephensi mosquitoes. Using direct amplification with the PowerPlex® Fusion 6C System, we have determined that full and partial profiles can be obtained by assessing degradation of DNA at various times post-feed up to 20-24 h post-blood meal. Moreover, we can assign donor identity through both STR profiles, as well as through single nucleotide polymorphisms (SNPs) detected using massively parallel sequencing (MPS) with the Precision ID Identity Panel and Ion Chef™/Ion S5™ System up to 24-48 h post-blood meal. Based on the results from a total of 490 mosquitoes fed on 11 different sources of human blood, we conclude that both STR and SNP technologies can be applied to mosquito blood meals as effective forensic approaches to determine the identity of specific individuals and establish the timing of their presence at a crime scene.