Comparative evaluation of an original microhaplotype panel and its expanded version for human identification and...

Microhaplotypes are an emerging kind of genetic markers composed of SNPs in closely linked allelic combinations, typically spanning up to 300 bp. When analyzed using next generation sequencing, they are considered a promising alternative to STRs in complex forensic casework involving mixed samples and degraded DNA. This study presents the first application of MPS to analyze microhaplotypes in 1165 individuals from a highly admixed urban Brazilian population (SABE cohort, São Paulo city). These data were derived from a WGS dataset, generated with a target coverage of 30 × using Illumina HiSeq X sequencing platforms. We evaluated the performance of the original panel of 130 microhaplotypes (MH-Base) and developed an expanded panel (MH-Plus) by incorporating additional SNPs into MH-Base loci to enhance marker informativeness. Forensic parameters and ancestry inference accuracy were assessed for both panels and compared with other genomic marker sets previously applied to the same cohort, including high-density SNP datasets. Additionally, we investigated the performance of both MH panels in AMR populations from the 1000 Genomes Project (ACB, ASW, CLM, MXL, PEL, PUR). MH-Plus improved all forensic parameters obtained with MH-Base. They showed greater potential for mixture deconvolution in Brazilian and recently admixed American populations, with CPD and CPE values exceeding those obtained with traditional STRs. The MH-Base panel provided ancestry estimates that were slightly better aligned with WGS reference values in the SABE cohort (4-parental model) and in AMR populations (3-parental model). In addition, the results obtained with the MH-Base panel in the SABE cohort validated the forensic and ancestry inference findings from a previous study conducted in another admixed Brazilian population (Ribeirão Preto - São Paulo), in which the same loci were genotyped using array-based methods and imputation tools. This study, therefore, contributes to consolidating a robust microhaplotype-based framework for forensic and anthropological applications in admixed Latin American populations.