Phosphine toxicity in Caenorhabditis elegans: synergy and cross-resistance with other pest control treatments, including gamma radiation

Abstract

Hydrogen phosphide (PH3), also known as phosphine, is an ideal fumigant to control insect pest infestation of stored grain, as it is inexpensive, easy to use and leaves little or no chemical residue. As no other general use fumigant is available, phosphine is used to protect 80% of the Australian grain harvest, with the remainder consisting primarily of animal feed and seed grain for planting. Heavy reliance on phosphine has resulted in the development of resistance among insect pests of stored products. In this project, the model organism, Caenorhabditis elegans, was utilized for exploring the mechanisms of phosphine toxicity and interaction with other treatments including gamma radiation as well as testing for synergistic actions between these treatments and phosphine.By looking into the effect of oxygen, I found that hyperoxia synergistically enhances the toxicity of phosphine against wild type C. elegans at 15, 20 or 25 °C, but it only marginally increases the effectiveness of phosphine against phosphine-resistant animals at 20 °C. The sub-lethal concentration of phosphine (70 ppm) with 80% oxygen under 15, 20 and 25 °C gave 60%, 96%, and 99% mortality respectively, in the wild type nematodes. Interestingly, the nematodes of both strains consume significantly more oxygen at 20 °C comparing to the other temperatures. However, the wild type worms consume significantly more oxygen than dld-(wr4) at all three temperatures. The toxicity of arsenite, on the other hand, was negatively correlated with phosphine toxicity. The phosphine-resistant mutant exhibited sensitivity to arsenite, which was close to an arsenite-sensitive mutant. Combining 4 mM of arsenite (~LC50) with 70 ppm phosphine resulted in elevated mortality of 89% in the phosphine-resistant mutant, whereas the combination was not lethal to wild type animals.One method of grain disinfestation is gamma irradiation; a treatment that can co-exist with phosphine in the grain storage system. I tested the toxicity of two distinct forms of irradiation on C. elegans, UV and gamma irradiation. By utilizing mutant lines that are sensitive or resistant to either phosphine or radiation, I found hypersensitivity to phosphine of mutations originally selected for hypersensitivity to either UV or gamma radiation. The phosphine-resistant and the radiation-resistant mutants were each significantly more resistant to UV and ionizing radiation than wild type C. elegans. UV and gamma radiation-sensitive mutant exhibited hypersensitivity to phosphine, considerably higher than the wild type in most cases. Unexpectedly, a gamma and UV radiation-resistant mutant was also hypersensitive to phosphine.The effect of pre-exposure to UV, ionizing radiation, and heat-shock was investigated, and I observed that these pre-treatments induced tolerance against phosphine in C. elegans. Heat-shock increased phosphine tolerance in the wild type strain by 3-fold, but no significant induction was observed in the phosphine-resistant mutant (dld-1(wr4)). On the other hand, mild exposure to UV and gamma radiation doubled phosphine resistance in the dld-1(wr4) mutant, but this effect was only observed with gamma radiation in the wild type strain.The interaction between phosphine and the other treatments in my work demonstrates the involvement of phosphine toxicity with oxidative respiration, where temperature, oxygen, and arsenite have synergized phosphine. Also, the cross-resistance between phosphine and gamma radiation supports that oxidative damage is involved in the mode of action of phosphine. Finally, the observation that heat shock induces phosphine resistance in wild type, but not resistant animals provides a focus for future molecular studies.