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Multivariate analysis of codon and amino acid usage was performed for three Leishmania species,including L.donovani,L.infantum and L.major.It was revealed that all three species are under mutational bias and translational selection.Lower GC12 and higher GC3s in all three parasites suggests that the ancestral highly expressed genes (HEGs),compared to lowly expressed genes (LEGs),might have been rich in AT-content.This also suggests that there must have been a faster rate of evolution under GC-bias in LEGs.It was observed from the estimation of synonymous/non-synonymous substitutions in HEGs that the HEG dataset of L.donovani is much closer to L.major evolutionarily.This is also supported by the higher dN value as compared to ds between L.donovani and L.major,suggesting the conservation of synonymous codon positions between these two species and the role of translational selection in shaping the composition of protein-coding genes.