Hemoglobin in the diet modulates post-blood feeding behavioral rhythms and gene expression in Aedes aegypti

Female Aedes aegypti mosquitoes rely on blood meals to acquire nutrients essential for egg development. However, blood ingestion also introduces physiological stressors--including thermal, osmotic, and oxidative stress--particularly during the digestion of heme-containing proteins. In Drosophila melanogaster, oxidative stress response genes are regulated by the circadian clock, and core circadian transcription factors are redox-sensitive. Additionally, sufficient iron ingestion is necessary to maintain normal circadian behaviors, as iron metabolism genes influence circadian behaviors, and flies lacking iron storage and transport are arrhythmic. However, whether similar interactions between iron metabolism and circadian rhythms exist in mosquitoes remains unclear. Here, we leverage the known alteration of mosquitoes activity rhythms following a bloodmeal to investigate whether dietary hemoglobin contributes to post-feeding activity suppression and gene regulation in the female Ae. aegypti. Using vertebrate blood and artificial blood mimic diets of equal nutritional values but with and without hemoglobin, we compared mosquito egg production, locomotor activity, sleep profiles, and transcript abundance of genes involved in circadian regulation and host-seeking behavior. Hemoglobin intake significantly reduced post-feeding activity, increased sleep amounts, and suppressed transcription of the core circadian gene period in mosquito heads. Short periods of sleep deprivation during the post-blood feeding period of inactivity did not alter egg production, timing of deposition, or viability. Our findings reveal that hemoglobin-derived heme influences behavioral and molecular responses in Ae. aegypti after a blood meal, pointing to a complex regulatory network linking heme acquisition, oxidative stress, daily rhythms, and behavior.