Cladistics of Rhipicephalus microplus and laboratory assessment of acaricidal efficacy of different Cassia fistula extracts against R. microplus larvae.

The present study aimed to analyze the cladistics and population structure analysis of Rhipicephalus microplus ticks infesting buffaloes in Haryana, India, as well as the assessment of the anti-tick efficacy of the ethanolic extracts of Cassia fistula (bark, pod pulp, and flowers) against R. microplus larvae. The molecular characterization and population structure analysis were performed by targeting the amplification of the partial mitochondrial cytochrome C oxidase subunit 1 (cox1) gene, whereas anti-tick efficacy was evaluated using a larval packet test. The sequences generated herein revealed a homology of 98.26-100% to the GenBank-archived R. microplus sequences. In population structure analysis, high haplotype (0.500 ± 0.265) and low nucleotide (0.002 ± 0.001) diversities were recorded for the sequences generated in this study. Significantly negative neutrality indices were recorded for the overall dataset. The extracts were found to significantly affect mortality rates in a dose-dependent manner, and the ethanolic extracts of the bark, pod pulp, and flowers of C. fistula exhibited median lethal concentration (LC₅₀) values of 27.989, 40.457, and 49.43 mg/mL, respectively. The LC₅₀ value recorded for the combination of the ethanolic extracts of the bark, flower, and pod pulp of C. fistula was 19.724 mg/mL, whereas the synthetic acaricide ivermectin had an LC₅₀ value of 351.56 mg/mL. In conclusion, R. microplus populations infesting cattle and buffalo hosts in India exhibited negligible genetic differentiation and high gene flow between them. Additionally, the combination of all C. fistula extracts could serve as a potential substitute for the synthetic acaricide.