The giant earthworm, Rhinodrilus priollii Righi 1967, is among the largest terrestrial invertebrates known worldwide, reaching lengths >2 m. To investigate the evolutionary history of the species and aspects of their reproductive biology, we collected R. priollii specimens from several field sites in central Amazonia. Phylogenetic analyses of 16 individuals using a fragment of cytochrome c oxidase subunit 1 (CO1) identified seven haplotypes that diverged between 2–8%. Population structures indicate episodes of gene flow between populations and their divergence within the past 1–2 million years. Histological examination of clitella from sexually mature specimens identified cocoon secretory cells throughout the dorsal and dorsoventral epidermis. Unlike previously described secretory cells, those in R. priollii contained granules with a proteinaceous core covered by external glycosylation. Further, collagenous matrices formed the bulk of swollen clitella while albumin-secreting cells were noticeably absent, collectively suggesting a mechanism of cocoon production somewhat different from that described in other clitellate megadriles.