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Taxonomy of thicket rats (Grammomys) is highly provisional and the genus is in a critical need of a thorough revision. We compared G. cometes from Eastern Cape Province (n = 150) with G. ibeanus, G. macmillani, and the southern African G. dolichurus, applying analyses of a partial cytochrome-b (Cytb) sequence (375 base pairs), karyotypes, and cranial morphology. Genetically, G. cometes appeared to be very close to G. dolichurus (mean sequence divergence of 3.4% ± 0.8% SE), whereas G. ibeanus and G. macmillani were separated by a mean sequence divergence of 5.4% ± 1.2%. Nucleotide diversity among haplotypes was higher in G. dolichurus (π = 0.0080 ± 0.0010 SD) than in G. cometes (π = 0.0040 ± 0.0009). G. cometes and G. dolichurus showed the same diploid chromosome number (2N = 52) of mostly acrocentric autosomes. None of the karyotypes reported so far for various Grammomys species match the chromosomal sets we found in Eastern Cape Province. Discriminant function analysis on 5 cranial measurements that are not affected by age variation was successful in separating G. cometes and G. ibeanus, but G. dolichurus appeared very similar to the former. In spite of their close genetic and morphological proximity, G. cometes and G. dolichurus tend toward ecological segregation and behave as distinct biological species. G. cometes is endemic to the southern African subregion and the 4 Eastern Cape Province localities are possibly isolates. Specimens were caught in the Afromontane forest above 1,000 m elevation and the lowland riverine forests dominated by Combretum caffra. [ABSTRACT FROM AUTHOR]