Hard ticks (Ixodidae) are important vectors of infectious diseases worldwide. Because of its geographical location, Georgia is a bridge between different biogeographical and climate zones and is rich in biodiversity. Different species of hard ticks are distributed in Georgia and have not yet been barcoded. Understanding the spatial distribution of disease vectors is important for health risk assessment. Cases of tick-borne diseases (borreliosis, Crimean-Congo hemorrhagic fever) have also been reported. In this study, we perform molecular-genetic characterization and barcoding of hard ticks distributed in Georgia. Mitochondrial cytochrome C oxidase I (COI-5P) was the molecular marker. Samples of hard ticks were collected from different regions of Georgia (Khashuri, Ambrolauri, Aspindza, Akhaltsikhe), DNA extraction and PCR amplification protocols were selected and optimized; Specimen were sequenced with the Sanger sequencing method and results were analyzed in Geneious Prime. All sample species were identified with high reliability (average query coverage 99.83 %, average percent Identity 99.82 %): Hyalomma scupense (=detritum), Hyalomma marginatum, Dermacentor marginatus, Rhipicephalus annulatus, and Rhipicephalus bursa. All of the listed species pose a threat to human health (including Crimean-Congo hemorrhagic fever, etc.), except for the species Rhipicephalus annulatus, which mostly infects cattle. The sequences are uploaded and published in GenBank and BOLD (Barcode of Life Data Systems), with indications of the samples’ geographical locations. To our knowledge, this is the first DNA barcoding results of hard ticks distributed in Georgia. It allows further phylogenetic analysis and comparison of species distribution across the Caucasus region and the globe. Future research is important to collect and analyze Ixodidae samples from all regions of Georgia in order to obtain more comprehensive data.