Previous studies provided no unequivocal evidence demonstrating that field populations of Lumbricus rubellus Hoffmeister (1843), exhibit genetically inherited resistance to As-toxicity. In this study F1, F2 and F3 generation offspring derived from adults inhabiting As-contaminated field soil were resistant when exposed to 2000 mg kg−1 sodium arsenate. The offspring of uncontaminated adults were not As-resistant. Cocoon viability was 80% for F1 and 82% for F2 offspring from As-contaminated adults and 59% in the F1 control population. High energy synchrotron analysis was used to determine whether ligand complexation of As differed in samples of: resistant mine-site adults, the resistant F1 and F2 offspring of the mine-site earthworms exposed to the LC25 sodium arsenate (700 mg kg−1) of the F1 parental generation; and adult L. rubellus from an uncontaminated site exposed to LC25 concentrations of sodium arsenate (50 mg kg−1). XANES and EXAFS indicated that As was present as a sulfur-coordinated species. As-resistance in F1, F2 and F3 offspring of the earthworm Lumbricus rubellus.