Patrizia Giangregorio, Italian Institute For Environmental Protection and Research, ISPRA

Genetic monitoring of small, isolated populations with extremely low genetic variability is essential for effective conservation management, as precise and reliable individual identification is critical to avoid biased population estimates and ensure robust demographic inference. The Apennine brown bear, a critically endangered and genetically depauperate population, poses significant challenges for individual genotyping using conventional short tandem repeat (STR) panels due to low polymorphism, genotyping errors, and limited comparability across laboratories and time.
We evaluated the genotyping performance of a high-throughput sequencing (HTS) microsatellite panel comprising 43 loci, including the ZFX/Y for sexing, on 46 invasive samples representing 46 individuals previously genotyped with capillary STR methods.
After quality filtering, 36 polymorphic loci were retained, showing high amplification success (mean=96%) and low genotyping error rates. Individual discrimination power was high, with a probability of identity (PID) of 5.0×10⁻¹⁶ and a probability of identity among siblings (PIDsib) of 4.1×10⁻⁸. The low PID and PIDsib values, combined with a high number of mismatches among genotypes, ensure reliable individual identification and provide redundancy to accommodate missing data across multiple loci. Genetic diversity metrics reflected the population’s low variability, yet several loci exhibited multi-allelic patterns.
This HTS-STR panel overcomes the limitations of capillary electrophoresis-based genotyping by offering a greater number of markers, along with unambiguous allele identification, thereby improving the informativeness and reliability of traditional STR analyses. Our results demonstrate the utility of this approach for precise individual identification, with additional potential for parentage inferences, pedigree reconstructions, and monitoring demographic and genetic parameters over time.