Genetics of phosphine resistance in the rice weevil Sitophilus oryzae (L.) (Coleoptera: Curculionidae)

Abstract

The rice weevil, Sitophilus oryzae (Linnaeus) is a cosmopolitan insect pest that can cause severe damage to stored grain with regard to both quality (loss of nutritional and aesthetic value) and quantity (loss of grain weight). Heavy reliance on the fumigant phosphine to disinfest stored products has led to resistance to phosphine in many insect species including S. oryzae. The frequency and level of phosphine resistance in S. oryzae has increased in the last fifty years in Asian and Africa countries and recently in Australia as well. A strongly phosphine resistant strain of S. oryzae was first detected in 2009 in Australia. I have determined the strong resistance trait in this strain to be incompletely recessive, autosomal and controlled by at least two major genes. The weakly resistant Australian strain shares a common resistance factor with the strongly resistant strain (So_rph1), which is responsible for weak resistance. I also compared the resistance trait of the Australian strain to that of collected from Vietnam and China. Complementation analysis revealed a shared genetic mechanism of phosphine resistance between the three strains that I tested. I then identified and sequenced the genes conferring phosphine resistance in this species. This process began with simply sequencing transcripts of the two genes known to be responsible for phosphine resistance in Rhyzopertha dominica and Tribolium castaneum (rph1/tc_rph1 and rph2/tc_rph2). rph2/tc_rph2 had been published and was known to encode the metabolic enzyme, dihydrolipoamide dehydrogenase (DLD). Variants of DLD (So_rph2) also cause strong resistance in S. oryzae when interacting with resistance variants of So_rph1. Therefore, rph2/DLD is responsible for strong resistance in each of the three species. A specific substitution mutation (N505T) in DLD is likely responsible for phosphine resistance as it is found in all strongly resistant strains of S. oryzae from Australia, Vietnam and China. A fitness cost associated with the resistance allele of So_rph2 was observed in the absence of selection in populations derive from the F1 progeny between a strongly resistant strain and strains lacking a resistance allele at So_rph2. As DLD is one of three subunits (E1, E2, E3) of a-ketoacid dehydrogenase complexes, I hypothesized that suppression of the activity of subunits other than DLD might also result in resistance to phosphine. Thiamine pyrophosphate is critical cofactor of the E1 subunit and is required by very few other enzymes. I reasoned that insects cultured on a diet of white rice, which is deficient in thiamine, would experience inhibition of the activity of E1. I expected to observe the change in response to phosphine in both the susceptible and resistance strains of S. oryzae when fed on white rice diet compared with the thiamine containing diet of whole grain (brown rice or whole wheat). Against my expectation, I found no significant affect of a thiamine deficient diet toward phosphine resistance in S. oryzae. A diet of white rice, however, did result in slowed developmental and a reduced number of progeny, but this effect was similar in both the phosphine resistant and susceptible strains. My work has demonstrated that the genetic mechanism for phosphine resistance is similar between S. oryzae and other pest insects studied so far. It will be straightforward to develop a molecular diagnostic to detect resistance alleles of S. oryzae in insect populations globally, facilitating resistance management.