Greenness assessment and phototoxicity of rose bengal and methylene blue on immature aquatic stages of malaria vector Anopheles pharoensis

This study systematically evaluated the efficiency of rose bengal and methylene blue as photosensitizers against the immature aquatic stages of Anopheles pharoensis. Genetic identification using the COI partial sequence confirmed the species, and the obtained sequence was submitted to GenBank (Accession No. PQ346929). Both photosensitizers exhibited 100% mortality in larvae I within 24 h at their highest concentrations, demonstrating strong biocidal activity. LC₅₀ values for rose bengal increased from 1.50 ppm (24 h) and 1.34 ppm (48 h) in larvae I to 3.83 ppm (24 h) and 3.12 ppm (48 h) in pupae. Similarly, methylene blue showed LC₅₀ values rising from 1.14 ppm (24 h) and 0.90 ppm (48 h) in larvae I to 2.91 ppm (24 h) and 2.51 ppm (48 h) in pupae, indicating stage-dependent susceptibility. Enzymatic responses revealed a progressive increase in acetylcholinesterase (AChE) and glutathione S-transferase (GST) activity in the developmental stage, suggesting a physiological adaptation to the photosensitizers. Molecular docking against the AChE protein (PDB ID: 6xyu) confirmed insecticidal bioactivity, with methylene blue exhibiting superior binding affinity, aligning with the in-vitro larvicidal results. Furthermore, a Complex GAPI assessment confirmed the environmental sustainability of both photosensitizers, supporting their potential as eco-friendly alternatives for mosquito control. The use of Complex GAPI in assessing the environmental sustainability of photosensitizers in mosquito control represents a novel approach in the field of integrated pest management. This advancement not only aligns with the principles of green chemistry but also addresses the growing need for sustainable alternatives to traditional chemical insecticides. These findings highlight the feasibility of utilizing light-activated photosensitizers for sustainable vector management.