Urban soils are subject to intense environmental pressures, e.g. physical disturbance, surface transformation and pollution, which greatly impact the activity of soil organisms. The epigeic earthworm Lumbricus castaneus (Savigny, 1826) is known to be tolerant to trace elements contamination of soils and was found to be abundant in the urban context. In order to investigate how urban environmental stressors shape the population genetic variation of this species, we developed 8 microsatellite loci from a microsatellite-enriched genomic library. Polymorphism was explored in 6 populations (153 individuals) from the Paris region. The number of alleles per locus varied from 7 to 29. The average expected heterozygosity within populations ranged from 0.596 to 0.705. These new microsatellite markers revealed a significant genetic structure at fine spatial scale, highlighting that they could be particularly useful for genetic studies of populations in urban soils.