Impact of singular versus combinatorial environmental stress on RONS generation in Drosophila melanogaster larvae.

We investigated environmentally correlated abiotic stressor desiccation (D), heat (H), and starvation (S) in the generation of reactive oxygen and nitrogen species (RONS) using Drosophila melanogaster larvae as an experimental model, subjected to either individual stressors or exposed to a combinatorial form of stressors (D + H, H + S, and D + S). The study was also extended to find synergistic endpoints where the impacts of all three stressors (D + H + S) were exerted simultaneously. We estimated the lethal time (LT20) at specific doses using regression and probit analyses based on the larval survival. LT20 values were used as the base-level parameter for further oxidative stress experimental analysis work. First, all stressors led to the activation of a typical common oxidative stress-mediated response irrespective of the mode of exposure. As envisaged, D. melanogaster larvae exhibited a homeostatic stress tolerance mechanism, triggering an antioxidant defense mechanism, indicated by an elevated level of total antioxidant capacity and enhanced activities of superoxide dismutase, catalase, glutathione reductase, and glutathione peroxidase. In all types of stress-exposed regimes, we found a negative impact of stressors on the activity of mitochondrial enzyme aconitase. Elevated levels of other oxidative stress markers, viz., lipid peroxidation, protein carbonyl content, and advanced oxidative protein products, were obvious although the increment was treatment-specific. Desiccation stress proved to be the most dominant stressor compared to heat and starvation. Among the combination of stressors, rather than a single stressor, D + H impacted more than other binary stress exposures. Focusing on the impact of singular versus combinatorial stress exposure on RONS generation, we observed an increase in the RONS level in both singular and combinatorial forms of stress exposure although the magnitude of the increment varied with the nature of stressors and their combinations. The present study indicated an "additive" effect when all three stressors (D + H + S) operate simultaneously, rather than a "synergistic" effect.

Differential manifestation of RONS and antioxidant enzymes in response to singular versus combinatorial stress in Chironomus ramosus.

In nature, organisms face multiple abiotic stress concurrently. Our previous study has indicated how threshold level of lethality depends on the type and combination of stressors. Many mechanisms exist by which organisms respond to stressors and maintain homeostasis. We examined the homeostatic pliability in an extremophilic oriental midge Chironomus ramosus larvae under various combinatorial stress conditions of desiccation (DS), heat (HS) and starvation (SS). Exposure to these stressors led to activation of a common response pathway of oxidative stress. Abundance of antioxidant enzymes like superoxide dismutase, catalase, glutathione reductase and glutathione peroxidase along with selective as well as stressor specific increase in total antioxidant capacity were reflected from the corresponding level of reactive oxygen and nitrogen species (RONS) in larvae exposed to various combinatorial stress. Additionally, we found stressor specific increment in lipid peroxidation level, protein carbonyl content and advanced oxidative protein products during the stress regime. Further investigation revealed a sharp decline in the activity of mitochondrial aconitase enzyme activity in response to abiotic stress induced oxidative stress. The combinatorial stressor specific comparative study based on biochemical and fluorescence based redox-endpoint assays confirmed that the generation of oxidative stress is the consequential convergent pathway of DS, HS and SS, but the quantum of RONS decides the redox potential of homeostatic response and survival rate.