*3.1. ROS and Arthropod's Innate Immunity*

To ensure their own survival and existence, arthropods use ROS to eliminate invading pathogens as well as to mount a better immune response during infection (Figure 1A) [52–54]. *Anopheles gambiae,* for example, can survive better at higher levels of systemic ROS when challenged with *Micrococcus* and *Escherichia*. Furthermore, the supplementation of antioxidants in the diet results in significantly higher mortality during bacterial infection [55], indicating that ROS and oxidative stress play a critical role in the arthropods' survival during the acquisition and transmission of infections. On the other hand, a *Plasmodium* refractory strain of *Ano. gambiae* was observed to be in a chronic state of oxidative stress, while the same parasite would survive if antioxidants were provided in the diet [8,40]. The same effects of oxidative stress are observed in the *Rh. microplus* (BME26) cell line. During infection with *Rickettsia ricketsii* or exposure to heat-killed microorganisms, upregulation of genes encoding for ROS production was observed, while antioxidant genes were downregulated [56]. Oxidative burst by macrophages efficiently eliminates pathogens basically by ROS, which are either toxic to the pathogen or work together with hydrolases, reactive nitrogen species, and the NADPH oxidase system (NOX). Supporting the above notion, bacterial infections have been shown to increase ROS in the ticks' hemocytes [39]. ROS play a role to block pathogen transmission by melanotic encapsulation, where invading pathogens are encapsulated to help the prevention of transmission. Melanotic encapsulation of *Plasmodium* has been shown in *Ano. gambiae.* The melanocytic capsule of the refractory strains of *Ano. gambiae* constructed around *Plasmodium* can block parasite development in the mosquitoes' midgut and the strain was observed to have higher levels of ROS [40]. On the other hand, in mosquitoes, ROS also act as a signaling molecule for the mitogen-activated protein kinase (MAPK)–dependent cascade and phosphatidylinositol 3-kinase (PI3K)/Akt-dependent pathway, which has been shown to regulate innate immunity and affects the physiology and development of the malarial parasite [41,42].

**Figure 1.** Schematic diagram of the life cycle of the pathogen (**A**) after infection of the arthropod versus the arthropod-borne pathogen's (**B**) life cycle and its interaction with ROS. Created with Biorender.com (accessed on 10 May 2022).
