A range of naturalised harmful plant storage pests including beetles, psocids, moths and mites habitually threaten the food safety, market access, trade and the overall profitability and sustainability of the Australian grain industry. Some of these pests have already been detected with resistance to phosphine and the spread of resistance is on the rise annually. In addition, new resistances are also being developed, a recent example being the detection of strong resistance to phosphine in several populations of flat grain beetles.

What is the biosecurity problem?

Australian export regulations require that all grain exported from Australia is free from insect infestation. This ‘nil tolerance’ standard is also adopted by domestic markets.

The Australian grain industry relies on chemicals, particularly phosphine fumigant, as the key tools used to meet the ‘nil tolerance’ standard. A major drawback, however, with this strategy is the threat of resistance in target species.

A major difficulty for the industry is that there is no practical replacement for phosphine and there are very few contact insecticides suitable for application to stored grain. Faced with this scenario, the industry has no choice but to maintain the tools that it has and must adopt a resistance management strategy to achieve this.

The main outputs of this project are to:

• improve understanding of the factors involved in the development of resistance to phosphine
• validate phosphine resistance management tactics
• improve understanding of distribution of species of flat grain beetle species and the role of gene flow in selection for resistance to phosphine 
• validate molecular resistance diagnostic tools and analysis of resistance gene frequencies.  

Who will be the end-users of this research?

The primary end-users will be:

• grain companies, farmers and other grain storers. In particular, individuals with responsibility for managing grain biosecurity
• officers with responsibility for biosecurity research and policy within government and non-government organisations.

Phosphine is the main fumigant used in Australia to control insect pests in grain storages; both bulk grain handlers and farmers rely on phosphine for the control of insects and more than 80% of grain is fumigated with phosphine during storage.

However, insect resistance to phosphine is increasing in most grain growing areas. To manage this resistance, a rapid and sensitive method for identifying phosphine resistance is required. The current detection method for phosphine resistance relies on time-consuming laboratory bioassay procedures (more than seven days required). A previous study (Park et al., 2008*) had suggested that certain proteins, displayed by two-dimensional electrophoresis (2D-PAGE), from whole Rhyzopertha dominica, differed between resistant and susceptible insects and might be developed as a rapid diagnostic tool.

Research outcomes

This project initially aimed to identify the genes encoding those differing proteins as a first step towards developing a diagnostic tool. However, when the project team used a larger number of strains of R. dominica and a more robust method the results of the previous study were not supported. The project team then proposed that this approach to biomarker discovery might yet be successful if they were to concentrate a more appropriate subset of proteins for proteomic analysis.

Various lines of evidence suggested that mitochondria are a site of important differences associated with phosphine resistance. A proteomic comparison of mitochondrial proteins from susceptible and resistant Tribolium castaneum (chosen for its completed genome sequence) was therefore conducted. The study did not reveal any significant differences in the expression of the more abundant mitochondrial proteins between resistant and susceptible insects. The aim of identifying differentially expressed proteins that are diagnostic for phosphine resistance has been shown to be beyond the scope of this project.

Research Implications

No changes to the use of phosphine for fumigation nor the management of resistance can be recommended as a result of this work.

Acknowledgements

David Schlipalius and Pat Collins (Queensland Primary Industries and Fisheries) supplied some of the insect strains.

*Park, B.-S., Lee, B.-H., Kim, T.-W., Ren, Y.L., Lee, S.-E., 2008. Proteomic evaluation of adults of Rhyzopertha dominica resistant to phosphine. Environmental Toxicology and Pharmacology 25, 121–126.