Metaphase karyotypes of 9 species of the Myzomyia Series show intra- and interspecific differences based on quantitative variation and distribution of constitutive heterochromatin in the sex chromosomes or the centromeric regions of the autosome pairs or both. Anopheles jeyporiensis and Anopheles aconitus each exhibit 4 forms of mitotic karyotypes, which may reflect interspecific differences within each taxon. The well-defined genetic species within the Anopheles minimus and the Anopheles culcifacies complexes clearly exhibit distinctive metaphase karyotypes that can be used as diagnostic characters for separating these sibling species, which are difficult to identify by morphological criteria alone. Our analysis on metaphase karyotypes of Anopheles pampanai, Anopheles varuna, and Anopheles flavirostris also confirms their morphological identification based on heterochromatin differences in the sex chromosomes and autosome 2.