Depredation is a human-wildlife conflict over resource which often includes a combination of socio-economical, ecological and conservation issues. However, estimating the amount of depredated resource can be especially challenging when depredation occurs on fish in the marine environment. This is the case for killer whales (Orcinus orca) and sperm whales (Physeter macrocephalus) depredation on the demersal Patagonian toothfish (Dissostichus eleginoides) longline fishery operating within the Crozet Islands EEZ (southern Indian Ocean). As the two species remove the entire fish from the hooks whether they depredate longline hooks separately or simultaneously, this study aimed at providing two indirect methods of assessment of depredated biomass over a 11-year period (from 2003 to 2013), both accounting for spatial variations of depredation levels. In the first method, we used fishing data from 6 525 longline sets to calculate the difference between Catch Per Unit Effort (CPUE) of non-depredated and depredated hooks. From this difference and using the fishing effort (number of hooks), we estimated that 575 ± 35 t and 739 ± 87 t of Patagonian toothfish were respectively depredated by killer whales and sperm whales when the two species occurred separately, and 1 679 ± 74 t were depredated from 2003 to 2013 when the two species co-occurred around vessels. The second method was new in a way that we used the differences of proportion of by-catch species (i.e. grenadiers – Macrourus sp.) between non-depredated and depredated longline sets to estimate the number of depredated Patagonian toothfish. This approach, which can only be implemented when a sufficient level of bycatch species occurs, provides strong support to the CPUE method. From these two methods, depredation rates were estimated to range from 27,3% to 29.1% % of the total capture (landed + depredated), that is one of the highest among all similar depredation situations reported elsewhere in the world. In addition to providing methodology insights that could be used in other areas with such depredation issues, these findings emphasize the critical importance for fishery managers and researcher to account for depredation when assessing fish stocks, fishery economy and/or conservation of odontocetes.