Our oceans are under increasing pressure from human activity, including fishing, pollution and global warming. Marine Protected Areas (MPAs) can be used to provide respite for ecosystems and preserve biodiversity for the future, and there is momentum to conserve 30% of the world’s oceans in MPAs by 2030. However, to effectively preserve biodiversity, the MPA network should contain a representative sample of all unique bioregions in the oceans. Identifying bioregions in the pelagic ocean is difficult to do by direct surveying, as it is the largest habitat on Earth, and biological sampling is expensive and slow. Here we use multiple large datasets, including many from the Integrated Marine Observing System, and advanced multivariate statistical techniques, to develop bioregionalisations for a range of trophic levels, including microbes, phytoplankton, zooplankton, and pelagic fish. We then compare these bioregions, assessing the number of bioregions for each group and whether their borders align. Finally, we evaluate the representativeness of the current Australian MPA network by calculating whether all pelagic bioregionalisations are currently protected, and to what extent.