Microsatellite and mitochondrial DNA genetic monitoring of the western corn rootworm, Diabrotica virgifera virgifera LeConte, was undertaken in Croatia and Serbia from 1996 to 2011 and in the United States in 2011. The seven U.S. populations displayed the greatest allelic diversity. In Europe, the highest number of alleles was found in Rugvica, Croatia, and Surčin, Serbia, the two sites closest to international airports. The highest number of mitochondrial (mt) DNA haplotypes was recorded from Croatia in 1996. From 2009 to 2011, haplotype diversity declined, and Croatia and Serbia had a single fixed haplotype. U.S. continuous maize locations had one haplotype, while three haplotypes were found at crop-rotated locations. Minimal temporal genetic differentiation was found within and between populations in Europe and the United States. Bayesian cluster analysis identified two genetic clusters that grouped western corn rootworm from Croatia and Serbia separately from U.S. populations; however, these clusters were not neat, and numerous U.S. individuals had both European and U.S. ancestry, suggesting bidirectional gene flow. Bottlenecks were identified within most Croatian populations sampled in 1996, only two populations in 2009, and in all populations in 2011. Bottlenecks were not identified from Serbia from 1996 to 2011 or from the United States in 2011. As suspected Serbia was identified as the geographic source of western corn rootworm in Croatia. The temporal genetic monitoring undertaken allowed a deeper understanding of the population genetics of western corn rootworm in Croatia, neighboring Serbia, and its geographic source in the United States. The data obtained can be used to inform western corn rootworm pest management strategies in Croatia and Europe.