Subleathal toxic effects of monocrotophos and carbamazepine on chrysomya megacephala (Fabricius, 1794): a multifaceted approach employing transcriptomics and geometric morphometrics, with forensic implications

Abstract

Chrysomya megacephala, a blowfly species of the Calliphoridae family, plays a crucial role in forensic entomology by aiding in the estimation of post-mortem interval (PMI). This study aims to investigate the impact of sublethal doses of monocrotophos, an organophosphate insecticide, and carbamazepine, an anticonvulsant drug, on the developmental stages of C. megacephala, from embryogenesis to emergence. The life cycle of C. megacephala consists of four distinct stages: egg, larva, pupa, and adult, with the duration of each stage influenced by temperature and humidity. To assess the effects of these toxicants, the EC50 values for monocrotophos and carbamazepine were estimated. Subsequently, the impact of sublethal doses on various developmental stages was examined. The molecular basis of these effects was further elucidated through the analysis of differentially expressed genes and transcriptomic data. The findings of this study contribute to the understanding of the influence of toxicants on the development of C. megacephala, which is essential for accurate PMI estimation in forensic entomology. Moreover, the results provide insights into the potential consequences of monocrotophos and carbamazepine exposure on the biology and development of this forensically important blowfly species. Toxicological analysis is crucial for detecting harmful substances in organisms and assessing their impact on human health. This study investigated the chronic in vivo toxicity of monocrotophos and carbamazepine on C. megacephala using enzymatic biochemical assays and transcriptomic analysis. The activities of superoxide dismutase (SOD), acetylcholinesterase (AChE), glutathione reductase (GR), and glutathione S-transferase (GST) were evaluated in the third instar larvae of C. megacephala under control conditions and in the presence of sub-lethal concentrations of the toxicants. Biochemical assays, which are reliable and commonly employed techniques, were used to identify targets and understand biomolecular functions. The study aimed to elucidate the detoxification mechanisms involved in the response to monocrotophos and carbamazepine exposure. Additionally, differential gene expression (DGE) analysis was performed to investigate the molecular basis of the detoxification mechanism. The findings of this study contribute to the understanding of the toxic effects of pollutants on various molecular pathways and provide insights into the potential impacts of drugs on human physiology. The integration of omics technologies, particularly transcriptomics, has significantly advanced the field of forensic entomology. This study presents a de novo transcriptomic profiling of the third instar larvae of C. megacephala to investigate the potential genes and pathways altered by sublethal concentrations of Carbamazepineand Monocrotophos, and their effects on development and various biological activities. The larvae were selected at the point of transformation from the feeding to the non-feeding phase, assuming that transcripts present in both the larval and upcoming pupal stages could be detected. Due to the lack of a previously published genome for this species or any closely related species, de novo profiling was performed. Differentially expressed genes (DEGs) were matched against the EuKaryotic Orthologous Group (KOG) database to categorize them into molecular families, highlighting the most prevalent functional categories. Gene Ontology (GO) enrichment analysis was employed to interpret the high-throughput molecular data and formulate hypotheses about the underlying biological mechanisms. KEGG pathway enrichment analysis was used to functionally annotate DEGs, providing insights into their associated functions and pathways. The results of this study contribute to a better understanding of the altered genes and pathways due to the effects of Carbamazepine and Monocrotophos on the development and biological activities of C. megacephala larvae, ultimately enhancing the accuracy of postmortem interval estimation in forensic entomology. Insect wings exhibit remarkable diversity in shape, size, and structure, reflecting their crucial role in insect evolution and radiation. The aerodynamic performance of insects is fundamentally tied to the specific attributes of their wings, and the interplay between shape and movement patterns determines both flying abilities and energy requirements. Geometric morphometric (GMM) analysis is a powerful tool for quantifying wing shape, offering efficient and statistically robust analyses that can connect abstract, multivariate outcomes to the physical structure of the original specimens. This study investigates the effects of two sublethal concentrations of monocrotophos, an organophosphate insecticide, and carbamazepine, an anticonvulsant, on wing development in C. megacephala, a forensically important dipteran species. Significant alterations in wing dimensions and morphology were observed using GMM analysis. Additionally, differential gene expression (DGE) associated with wing development was analysed, complementing and reinforcing the GMM results. The higher sublethal dose (1/3rd of EC50) was used for DGE analysis, as it showed more promising results in basic entomotoxicological analyses. The study of the effects of these intoxicants on C. megacephala can also be critical from a forensic perspective, as the species is of significant forensic importance. The findings of this study contribute to our understanding of the impact of environmental toxicants on insect wing development and morphology, with potential implications for insect evolution, ecology, and forensic entomology.