Cytogenetic characteristics and genome size are powerful tools for species characterization and identification of cryptic species, providing critical insights into phylogenetic and evolutionary relationships. Sitophilus Linnaeus, 1758 grain weevils can benefit from such tools as key pest species of stored products and also as sources of archeological information on human history and past urban environments. Moreover, the phylogenetic relationship among these weevil species remains controversial and is largely based on single DNA fragment analyses. Therefore, cytogenetic analyses and genome size determinations were performed for four Sitophilus grain weevil species, namely the granary weevil Sitophilus
granarius (Linnaeus, 1758), the tamarind weevil S.
linearis (Herbst, 1797), the rice weevil S.
oryzae (Linnaeus, 1763), and the maize weevil S.
zeamais Motschulsky, 1855. Both maize and rice weevils exhibited the same chromosome number (2n=22; 10 A + Xyp). In contrast, the granary and tamarind weevils exhibited higher chromosome number (2n=24; 11 A + Xyp and 11 A + neo-XY, respectively). The nuclear DNA content of these species was not proportionally related to either chromosome number or heterochromatin amount. Maize and rice weevils exhibited similar and larger genome sizes (0.730±0.003 pg and 0.786±0.003 pg, respectively), followed by the granary weevil (0.553±0.003 pg), and the tamarind weevil (0.440±0.001 pg). Parsimony phylogenetic analysis of the insect karyotypes indicate that S.
zeamais and S.
oryzae were phylogenetically closer than S.
granarius and S.
linearis, which were more closely related and share a more recent ancestral relationship.