Integrative Description of Calotropis sp. nov. (Apocynaceae) from Karnataka Based on Morphological, Molecular, and Biochemical Data

During extensive field surveys in the Kalaburagi region of Karnataka, India, a Calotropis population exhibiting a consistent combination of morphological features differing from those typically reported for Calotropis procera and Calotropis gigantea was documented. This population is provisionally referred to here as Calotropis prefina. The present study evaluates its taxonomic position using an integrative framework combining detailed morphological assessment, leaf and stem anatomical analysis, biochemical characterization of latex (milk coagulation activity and temperaturedependent pH profiling), chloroplast DNA barcoding (rbcL), and herbarium-based authentication. Comparative analyses revealed a stable and diagnostic suite of characters involving plant stature, stem pubescence, latex viscosity, leaf morphology, floral architecture, pollinia structure, vascular organization, and laticifer distribution. Calotropis prefina exhibited markedly reduced pollen viability (57.5%) compared with C. procera (82.5%) and C. gigantea (80%), indicating reproductive differentiation. Biochemical assays showed reduced milk clotting activity and a consistently alkaline, thermally stable latex pH profile in C. prefina relative to C. procera and C. gigantea, indicating functional divergence. Molecular characterization using the chloroplast rbcL barcode confirmed the placement of the investigated taxon within the genus Calotropis. NCBI BLAST analysis revealed the highest sequence similarity with Calotropis procera (98.04% - 97.86%), while C. gigantea was also retrieved among the closest matches with comparable identity values. Independent identification through the BOLD Systems database similarly failed to achieve unambiguous species-level discrimination, returning both C. procera (97.83%) and C. gigantea as closest reference taxon with closely comparable percentage similarity. These results indicate strong genetic affinity within Calotropis while highlighting the Phylogenetic interpretation placed C. prefina within the Calotropis clade, closely allied to C. procera and C. gigantea, yet forming a distinct lineage. Taken together, the convergence of morphological, anatomical, biochemical, and molecular evidence supports the interpretation of Calotropis prefina as a distinct taxonomic entity within the genus Calotropis. The provisional designation is applied pending formal nomenclatural validation in accordance with the International Code of Nomenclature, with herbarium authentication currently underway at the Botanical Survey of India.