The implementation of molecular data for the analysis of nemertean diversity has unraveled the taxonomic status of several species and many higher taxa within the group. Nowadays, a large proportion of novel putative species are being discovered and it is necessary to add molecular data to the morphological description to obtain a correct identification. In this study, we used mitochondrial cytochrome oxidase I gene (COI) as molecular marker to investigate the diversity of nemerteans from a marine Sub-Antarctic environment. We used Automatic Barcode Gap Discovery (ABGD), Poisson Tree Processes (PTP) and Bayesian implementation of the PTP model (bPTP) as well as reciprocal monophyly on neighbor-joining (NJ) and maximum likelihood (ML) trees for species delimitation. ABGD showed a clear barcoding gap (6 to 10%) and the presence of 15 different putative nemertean species in the dataset of 54 COI sequences from the marine protected area “Namuncurá–Burdwood Bank” (MPA-BB), expanding the known biodiversity for the Sub-Antarctic region. Ten monostiliferan and five heteronemertean species were found. Our results also confirm the presence of two Parborlasia corrugatus cryptic species in the Antarctic and Sub-Antarctic region. This work highlights the importance of the MPA-BB as a biodiversity hotspot and provide molecular data for the Phylum Nemertea in the Southwestern Atlantic Ocean.