BACKGROUND: The Colorado potato beetle (CPB) is the most harmful pest of potato in potato cultivation regions globally. Although it is an economically important agricultural pest, the population structure and colonization route of this species in Turkey are uncertain. We used microsatellite and mitochondrial DNA (mtDNA) markers to obtain information about the population source, structure and bio-invasion route of CPB populations in Turkey. RESULTS:The common single mtDNA haplotype in European CPB populations was obtained in all Turkish CPB populations based on mtDNA data analysis. However, microsatellites revealed a low level of genetic variation in CPB populations. The results of microsatellite analysis [factorial correspondence analysis (FCA), Bayesian analysis of genetic population structure (BAPS), unweighted pair group method with arithmetic mean (UPGMA) dendrogram, F-statistics and Nei's distances] indicated three groups for invasive CPB: Thrace-Marmara and Aegean; Black Sea, Central Anatolia and Mediterranean; Northeastern Anatolia. Region-specific alleles have been identified in regions, where commercial potato cultivation and insecticide use are intensive.CONCLUSION: The detection of a single fixed European haplotype in all Turkish populations has proved that CPB in Turkey originated from Europe as a result of a founder event occurred in European populations. Low genetic variation was due to the short time period since the spread of CPB from America to Europe. The highest number of private alleles were found in the top commercial potato cultivation region-Central Anatolia from where the CPB populations spread to other parts of Turkey.