The dalatiid genus Isistius Gill (1864) has three valid species currently recognized in the literature: Isistius brasiliensis Quoy & Gaimard (1824), I. plutodus Garrick & Springer (1964), and I. labialis Meng, Zhu & Li (1985). The most common species, I. brasiliensis, has a wide geographic distribution and is found in subtemperate and tropical seas circumglobally. A comparative analysis of specimens from different localities throughout its range, however, had never been undertaken. In the present paper, the morphological variation of this species along its entire distribution has been thoroughly analyzed, corroborating that it represents a single widespread species and that I. labialis is its junior synonym. The other congeneric species, I. plutodus, is known from only a few specimens and is also distributed worldwide. A detailed comparative analysis of available material of I. plutodus was conducted verifying its validity as a single widespread species. The present study analyzed in detail the external morphology (coloration, dentition, dermal denticles), internal morphology (skeleton, musculature), lateral-line canals, and morphometric and meristic characters of species of Isistius in order to better define the genus and its included valid species.