MIEKA: Development and evaluation of a large-scale microhaplotype panel for extended kinship analyses

In recent years, microhaplotypes have emerged as versatile markers in forensic genetics, with applications including mixture deconvolution, individual identification, biogeographical ancestry inference, and kinship analysis. Microhaplotypes are short DNA regions (<300 bp) comprising closely linked SNPs. While their utility in forensics has been demonstrated, most studies have focused on relatively small marker sets and have not systematically evaluated the impact of locus and allelic dropout. Here, we present the MIcrohaplotype panel for Extended Kinship Analysis (MIEKA), a novel panel combining 286 previously developed microhaplotype markers. MIEKA is designed to complement established STR-based kinship inference by providing additional discriminatory power in cases where STR results alone are insufficient, particularly for distinguishing full from half siblings and offering additional potential to resolve third-degree relationships under challenging sample conditions. The performance of MIEKA was evaluated in pairwise kinship analyses and compared with 32 standard STRs, the FORensic Capture Enrichment (FORCE) SNP panel, and the combination of MIEKA and STRs (MIEKA+STR panel) using simulations of first- to fifth-degree relationships generated with FamLink2. Particular emphasis was placed on full versus half sibling discrimination investigated using likelihood ratio (LR) calculations in Merlin. Robustness was assessed through simulations of locus and allelic dropout up to 50%, as well as analysis of low DNA quantity and quality samples. Our findings show that the combined MIEKA + STR panel enables reliable discrimination of second-degree and, to a substantial extent, third-degree relationships, with clearly separated LR distributions for full versus half siblings. Classification remains robust under high locus dropout (>50%), and only high allelic dropout (>30%) results in decreased accuracy. The MIEKA + STR panel proves useful for the analysis of poor-quality and low-template DNA samples and for kinship inference up to third-degree relationships. Collectively, these results establish MIEKA, particularly in combination with STRs, as a robust and practical tool for extended kinship analysis in challenging forensic contexts.