Molecular and morphometric divergence of four species of butterflies (Nymphalidae and Pieridae) from the Western Himalaya, India.

Morphometric and molecular divergence among four butterfly species of the families Nymphalidae and Pieridae from the western Himalaya region were investigated using molecular tools, traditional morphometric measures and a truss network system. The considered species were Danaus chrysippus, Vanessa cardui, Pieris brassicae and Pieris canidia. Traditional taxonomy is sometimes unable to discriminate cryptic species or species that have close morphological features. Although taxonomists carefully examine external body features to differentiate the species; however, there is a risk for misidentification during a visual assessment of cryptic species. Therefore, we aimed to use the truss network system of 14 morphological landmarks interconnected to yield 90 variables about molecular taxonomy. Principal component analysis (PCA), discriminant function analysis (DFA) and cluster analysis (CA) were employed to determine morphometric variations. In the traditional analysis, 79, 68, 16 and 5 characters out of 90 were found significant (p < 0.05) for D. chrysippus, V. cardui, P. brassicae and P. canidia, respectively. One to seven principal components were extracted through PCA; they explained 87.5-100% of the total variance in samples. Notably, DFA correctly classified 100% of the original grouped cases and 100% of the cross-validated grouped cases. However, the variations were not the same for the two different methods (truss and traditional) employed for the analysis. We correctly identified all the species; the interspecies sequence divergence was between 0.1034 and 0.1398, and the intra-species sequence divergence range was 0.0001 to 0.0128 using the Cytochrome c oxidase subunit-I (COI) gene. The present study provides useful information about the application and complementary role of traditional with truss morphometric analysis for the precise identification and classification of the selected species.