Temporal and spatial profiling of Aedes albopictus immune responses to chikungunya virus infection

The global expansion of Aedes albopictus from Southeast Asia to various regions worldwide poses a significant public health concern due to its role as a vector for several pathogens, including chikungunya virus (CHIKV), which infects over one million people annually. In this study, aimed at understanding the molecular interactions between Ae. albopictus and CHIKV, we analyzed by RNA sequencing CHIKV-infected and uninfected control mosquitoes. We focused our attention on key mosquito organs at one- and five-days post-blood meal ingestion, which correspond to viral entry and dissemination, and found specific transcriptional changes involving various pathways during the CHIKV lifecycle. The mosquito midgut plays a crucial role in the early stages, when the virus enters along with human blood components, encounters the resident microbiota, interacts with the developing peritrophic matrix, and counteracts the mosquito's digestive enzymes. We found that RNA interference (RNAi) was predominantly activated in the midgut during the initial virus invasion. Additionally, several key enzymes involved in autophagy and ubiquitination were also more abundant in infected midguts compared to controls. At later time points, after viral dissemination into the hemocoel, key immune responses are triggered in the hemolymph and, accordingly, immune mechanisms such as the activation of leucine-rich repeats (LRRs) proteins, secretion of antimicrobial peptides (e.g., holotricin), and melanization (mediated by phenoloxidase, PO) were the most prominent. RNA-seq results were validated by RT-qPCR on selected candidates in different tissues and a catalogue of Ae. albopictus immune genes (891 contigs) grouped into 24 different immune and immune-related families was compiled. This study explores the molecular interactions between Ae. albopictus and CHIKV across developmental stages, providing key insights into arbovirus transmission dynamics and mosquito vector competence.