Oxidative Stress in Invertebrate Systems

Invertebrates have been valuable research models in the discovery of many scientific principles owing to the numerous advantages they provide. Throughout the life cycle, many of them thrive in pathogen-rich environments, manage harsh weathers, exposed to a number of allochemicals, and adapt well to both terrestrial and marine ecosystems. Their remarkable ability to cope up with the enormous oxidative stress generated in all these circumstances, make them attractive models in this field of research. Endocrine control of oxidative stress in insects is recently emerging. Adipokinetic hormone, glucagon, ecdysteroids and juvenile hormone have been implicated in antioxidative protective role in insects. Drosophila and Caenorhabditis elegans have provided the largest body of evidence addressing the free radical theory of ageing. Oxidative stress is also induced by pesticides/insecticides. In mollusks, pesticides exert their biological effects via generation of ROS. Oxidative stress has been shown to be associated with exposure to several organophosphorous compounds and different classes of pyrethroids. Malathion is a potential hazard to the environment. Adverse effects induced by malathion in earthworms and insects have been reported. Information is now available in great detail on the role of ROS in modulating insect immunity during parasite invasion and bacterial infection. In Drosophila melanogaster ROS are actively produced in the midgut at a basal level in the presence of commensal microbiota and highly generated upon bacterial challenge. The involvement of reactive oxygen species (ROS) in mosquito immunity against bacteria and Plasmodium was investigated in the malaria vector Anopheles gambiae. The concentration of ROS increased in sand fly midguts after they fed on the insect pathogen Serratia marcescens. Elevated oxidative stress was previously reported for a mosquito line experimentally infected with Wolbachia, indicating that oxidative stress may be important for Wolbachia-mediated antiviral protection. In a nutshell, this chapter highlights the current advances of oxidative stress in invertebrate model systems and its implications

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Not AvailableBacillus thuringiensis (Bt) crystal proteins (Cry) bind to aminopeptidase N (APN) receptors on insect midgut membrane leading to pore formation and subsequent death. However, evolution of insect resistance to Bt toxins threatens their long-term application. Therefore, search for new targets which could function as Cry toxin receptors is an immediate mandate. In the present study, two full-length APN cDNAs were cloned from Malpighian tubule and salivary gland tissues of the moth, Achaea janata. Both these APNs showed 99% and 32% sequence homology with fat body and midgut APNs respectively. Tissue distribution analysis revealed the presence of two different APN isoforms, one predominant in non-gut visceral tissues while the other exclusively expressed in the midgut. Immunofluorescence and western blot analyses showed cross-reactivity in Malpighian tubule and salivary gland when probed with anti-fat body APN antiserum. These results clearly indicated the presence of non-gut (AjAPN1) and gut-specific (AjAPN4) isoforms in this moth. The expression of both the isoforms steadily increased during the larval development. Hormonal studies indicated regulation of the APN genes by the morphogenetic hormones, 20-hydroxyecdyone and juvenile hormone. Further, in vitro ligand-blotting studies demonstrated binding of Cry toxins to APNs in Malpighian tubule and salivary gland indicating their potential as alternate targets.Not Availabl

Gut-specific arylphorin mediates midgut regenerative response against Cry-induced damage in Achaea janata

Development of insect resistance to biopesticides is a current and pertinent global issue. Earlier, it was established that lepidopteran larvae can recover from Bt intoxication via a midgut regenerative response and subsequently generate resistance. Molecular aspects of restoration of the midgut integrity following toxin exposure are emerging recently. In the present study, we bring out the pivotal role of gut arylphorin in mediating the midgut regenerative response following sublethal Bt exposure in Achaea janata. Bt-induced midgut damage was characterized by microscopic analysis using differential interference contrast (DIC) and immunofluorescence (IF). Extensive disruption of brush-border membrane, associated with the underlying cytoskeletal alterations including F-actin, α-actin and β-tubulin was observed. Single-photon fluorescence microscopy combined with fluorescence lifetime imaging (FLIM) established the metabolic state associated with enhanced stem cell proliferation and migration from the basal side towards the luminal side following the damage. In-silico analysis revealed the phylogenetic relationship of gut arylphorin with closely related insect species and indicated the presence of two different subunits. Transient RNAi knockdown of the arylphorin resulted in diminished expression of mitotic Cyclin B mRNA levels. Human monoclonal Cyclin B antibody cross-reactivity with the Cyclin B located in the stem cells further validate the role of arylphorin as the mitogenic factor responsible for stem cell proliferation and epithelial regeneration. An in-depth understanding of resistance mechanisms will aid in the design of new strategies for the long-term usage and efficacy of Bt technology against pest control. © 202