Swordfish, striped marlin and sailfish are the dominant billfish species caught in the Indian Ocean. These species are currently assessed and managed as single stocks. Evidence from population genetics studies to date has not provided evidence that a revision to this approach is warranted. Exploring and understanding the level of population heterogeneity is a priority for sustainable management of these fisheries. This paper presents results from a recent investigation of population structure of swordfish, striped marlin and sailfish using cutting-edge sequencing technology as part of a larger collaborative project “Population Structure of IOTC species and sharks of interest in the Indian Ocean (PSTBS-IO)”. Sampling of striped marlin was challenging with only three individuals collected within the Indian Ocean, permitting only a rudimentary population comparison to a single outlier location in the Coral Sea, SW Pacific Ocean. Similarly, sailfish also proved difficult to obtain, however, sufficient samples were obtained to permit analysis from two Indian Ocean locations. No population structure was revealed for this comparison, which was consistent with sailfish representing a highly mixed Indian Ocean population. For both striped marlin and sailfish, sampling coverage was insufficient for the current study to unequivocally conclude whether there was evidence of sufficiently restricted gene flow to consider whether the Indian Ocean basin contains stocks of these species that are effectively isolated population(s) from the Atlantic and/or Pacific Oceans for fisheries management purposes. In contrast, swordfish samples permitted a broad spatial analysis that included 6 sampling locations within the Indian Ocean and an outlier sample location from the southwest Pacific in the Coral Sea. Differences in gene-frequencies of swordfish between the Indian and south-west Pacific Ocean, in the current study, and results from earlier studies from the south-east Atlantic Ocean support the assessment and management of swordfish within the Indian Ocean as an independent unit. Notwithstanding this conclusion, the potential for connectivity between the NE Indian and western Pacific Oceans requires further investigation. The indication of different northern and southern genetic groupings of swordfish, which may represent different reproductive populations, warrants targeted investigation via additional structured sampling and analysis that includes and expands the areas covered in this project and, ideally, includes sampling of spawning adults. For the three billfish species examined, insufficient sampling coverage was obtained for both sailfish and striped marlin to enable adequate assessment of population genetic structure. However, results for swordfish indicate consideration of management strategy refinement may be warranted, and highlights the need for targeted research to confirm the temporal stability of the results presented here, and in particular, further extending of sample coverage in the northern regions of the IO. Additional sampling both within and from locations outside the Indian Ocean should also be completed in order to provide a more comprehensive understanding of inter-ocean population boundaries for all three Indian Ocean billfish species.