Chlorine dioxide enhances lipid peroxidation through inhibiting calcium-independent cellular PLA2 in larvae of the Indianmeal moth, Plodia interpunctella.

Polyunsaturated fatty acids usually undergo lipid peroxidation induced by reactive oxygen species (ROS). Calcium-independent cellular phospholipase A2 (iPLA2) can maintain fatty acid compositions in phospholipids depending on physiological conditions. An insect iPLA2 (Pi-iPLA2) was predicted from the transciptome of the Indianmeal moth, Plodia interpunctella. It encodes 835 amino acids. It possesses five ankyrin repeats in the N terminal and patatin lipase domain in the C terminal. Pi-iPLA2 was expressed in all developmental stages of the Indianmeal moth. In the larval stage, it was expressed in all tissues tested. RNA interference (RNAi) specific to Pi-iPLA2 was performed using specific double-stranded RNA (dsRNA). It resulted in almost 70% of reduction in gene expression. Under such RNAi condition, P. interpunctella exhibited significant accumulation of lipid peroxidation based on the amount of malondialdehyde. RNAi of Pi-PLA2 expression also impaired cellular immune response of P. interpunctella. Chlorine dioxide (ClO2), an insecticidal agent by generating ROS, increased lipid peroxidation in a dose-dependent manner. However, the addition of vitamin E (an antioxidant) reduced the formation of lipid peroxidation. ClO2 treatment significantly reduced expression of Pi-iPLA2 but increased lipid peroxidation in larval fat body of P. interpunctella. Furthermore, larvae treated with dsRNA specific to Pi-iPLA2 were significantly susceptible to ClO2 treatment. These results suggest that Pi-iPLA2 plays a crucial role in repairing damaged fatty acids from phospholipids. Our results also suggest that ClO2 can elevate lipid peroxidation through inhibiting Pi-iPLA2 expression in addition to direct ROS production.

Heat Tolerance Induction of the Indian Meal Moth (Lepidoptera: Pyralidae) Is Accompanied by Upregulation of Heat Shock Proteins and Polyols.

The Indian meal moth, Plodia interpunctella, causes massive damage to stored grains and processed foods. Heat treatment has been widely used to control insect pests infesting stored grains. However, heat treatment may result in unsatisfactory control owing to heat tolerance of target insects. This study quantified the heat tolerance and analyzed its induction in P. interpunctella. Susceptibility of P. interpunctella to different high temperatures was assessed in all developmental stages. Heat treatment at 44 °C for 1 h caused significant mortalities to all developmental stages, with late-instar larvae exhibiting the highest tolerance. However, the survivorship to heat treatment was significantly increased by pre-exposure to 37 °C for 30 min. The induction of heat tolerance was accompanied by upregulation of two heat shock proteins of Hsc70 and Hsp90. Trehalose and glycerol concentrations in the hemolymph also increased after pre-exposure to 37 °C for 30 min. RNA interference (RNAi) by specific double-stranded RNAs effectively suppressed the inducible expressions of both Hsc70 and Hsp90 in response to 37 °C for 30 min. Either RNAi of Hsc70 or Hsp90 significantly impaired the heat tolerance induction of P. interpunctella. These results suggest that the induction of heat tolerance in P. interpunctella involves the upregulation of these heat shock proteins and hemolymph polyol levels.

Oxidative stress induced by chlorine dioxide as an insecticidal factor to the Indian meal moth, Plodia interpunctella.

A novel fumigant, chlorine dioxide (ClO2) is a commercial bleaching and disinfection agent. Recent study indicates its insecticidal activity. However, its mode of action to kill insects is yet to be understood. This study set up a hypothesis that an oxidative stress induced by ClO2 is a main factor to kill insects. The Indian meal moth, Plodia interpunctella, is a lepidopteran insect pest infesting various stored grains. Larvae of P. interpunctella were highly susceptible to ClO2 gas, which exhibited an acute toxicity. Physiological damages by ClO2 were observed in hemocytes. At high doses, the larvae of P. interpunctella suffered significant reduction of total hemocytes. At low doses, ClO2 impaired hemocyte behaviors. The cytotoxicity of ClO2 was further analyzed using two insect cell lines, where Sf9 cells were more susceptible to ClO2 than High Five cells. The cells treated with ClO2 produced reactive oxygen species (ROS). The produced ROS amounts increased with an increase of the treated ClO2 amount. However, the addition of an antioxidant, vitamin E, significantly attenuated the cytotoxicity of ClO2 in a dose-dependent manner. To support the oxidative stress induced by ClO2, two antioxidant genes (superoxide dismutase (SOD) and thioredoxin-peroxidase (Tpx)) were identified from P. interpunctella EST library using ortholog sequences of Bombyx mori. Both SOD and Tpx were expressed in larvae of P. interpunctella especially under oxidative stress induced by bacterial challenge. Exposure to ClO2 gas significantly induced the gene expression of both SOD and Tpx. RNA interference of SOD or Tpx using specific double stranded RNAs significantly enhanced the lethality of P. interpunctella to ClO2 gas treatment as well as to the bacterial challenge. These results suggest that ClO2 induces the production of insecticidal ROS, which results in a fatal oxidative stress in P. interpunctella.