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In this study, the maternal genetic diversity, phylogenetic relationship and effect of natural selection on indigenous chickens from Nigeria were assessed. A total of 397-bp fragment of the mitochondrial DNA (mtDNA) D-loop region of 171 indigenous chickens from four populations of Nigeria and four commercial egg line strains (two Anak titan, one Giriraja and one Yaffa) as out-groups were analysed. Thirty-one haplotypes (28 from Nigerian chickens and three from commercial strains) and 34 polymorphic sites were identified. The mean haplotypic and nucleotide diversity were found to be 0.39 ± 0.05 and 0.02 ± 0.02, respectively. Majority of Nigerian chicken haplotypes observed were grouped into haplogroup D which originated from Indian subcontinent, suggesting a single maternal lineage. Genetic variation within and between populations accounted for 97.30 and 2.70% of the total genetic variation, respectively, which is in agreement with a recent and maternal founding effect. High number (4) of negatively selected sites observed based on single likelihood ancestral counting (SLAC) model indicated that the sampled Nigerian chicken populations were undergoing purifying selection. This study concluded that there was relatively high genetic diversity and differentiation, thus, this information will probably paveway for further evaluation studies, preservation and improvement of Nigerian chickens as genetic resources towards ensuring food security.