Because of its importance as a pollinator and its potential economic usefulness for the biodegradation of organic animal waste, the genetic and phenotypic diversity of the drone fly, Eristalis tenax L. (Diptera: Syrphidae), was studied in both wild and captive populations from southeastern Europe. Wild specimens from a natural protected habitat (with low human impact), field crop habitat (semisynanthropic condition), and intensive pig farming habitat (synanthropic condition) were compared with a laboratory colony reared on artificial media. An integrative approach was applied based on allozyme loci, cytochrome c oxidase I mitochondrial DNA, wing traits (size and shape), and abdominal color patterns. Our results indicate that the fourth and eighth generations of the laboratory colony show a severe lack of genetic diversity compared with natural populations. Reduced genetic diversity in subsequent generations (F4 and F8) of the laboratory colony was found to be linked with phenotypic divergence. Loss of genetic variability associated with phenotypic differentiation in laboratory samples suggests a founder effect, followed by stochastic genetic processes and inbreeding. Hence, our results have implications for captive bred Eristalis flies, which have been used in crop pollination and biodegradation of organic waste under synanthropic conditions.