“Monitoring is essential in phosphine use. When it comes to monitoring, monitoring of the gas concentration is not the only thing that has to be monitored. Other parameters, such as temperature etc. should be monitored in parallel, as phosphine concentration has interesting diurnal circles according to the key abiotic conditions prevailing. Distribution of the gas is also important. The traditional monitoring techniques are able to provide measurements for specific time intervals and at specific locations within the fumigated facilities. However, the measurements refer to the average concentrations, which are not always representative. Extreme dosing and absence of monitoring are key elements that are directly related with resistance development.”

Christos G. Athanassiou Laboratory of Entomology and Agricultural Zoology Department of Agriculture, Crop Production and Rural Environment University of Thessaly, Greece athanassiou@agr.uth.gr

INTRODUCTION Despite the newer developments with liquid insecticides in stored product protection, e.g. introduction of new active ingredients to the market, the fumigant phosphine remains the main insecticide that is used at the post-harvest stages of durable agricultural commodities, due to its effectiveness and the convenience in application. The development of resistance to insecticides is an evolutionary phenomenon, that has been thoroughly studied from both molecular and population genetics. It is thus a complicated phenomenon, that involves basic gene mutations and changes in chromosome structure, and is directly related with fitness cost and insect behavioral characteristics. Although the degree on which human involvement is related with the development of resistance has not been clarified in detail, there are specific indicators that can be used when the presence of resistance is present. These can be summarized in the following:

What happens to the genes if the fumigant is not applied? There are certain active ingredients that can be used with success towards this direction. Rotation of phosphine with other insecticides is the key element in resistance management. These other insecticides are referred often as “resistance breakers”, and can be applied in a carefully planned strategy on the basis of resistance mitigation practices. There are numerous paradigms, for different insecticides that are currently in use in stored product protection, that the avoidance in the use of specific active ingredients for some intervals may “reverse” resistance development through different resistance gene frequencies. This “breaker” approach has yielded good results in several parts of the world, and can be further expanded. For example, liquid insecticides, that can be effective for a long period on grains and related durable commodities have been proposed for this purpose, as their persistence on the commodity is able to lead to the drastic reduction of the fumigations that are performed for a certain period of time.

A FITNESS COST IS INVOLVED Different factors such as insect movement and mating may also impact the frequency of resistance. Considering the availability of new research tools in studying insect behavior, it is now well established that, in some cases, there is a considerable fitness cost of the resistant populations. For example, respiration may be reduced, as well as progeny production capacity, while development may be slower than that of the susceptible populations. This is directly related with frequency and occurrence of resistance, and is likely to cause a more gradual increase in resistance cases, instead of a rapid peak in the presence of the resistance genes. Still, differences in behavior may result in differential response of the resistant individuals to sampling and trapping methods, e.g. reduced movement and attraction towards a pheromone source in trapping protocols.

THERE ARE MOLECULAR MARKERS THAT CAN BE USED FOR EVALUATION The sequence information of the rph2 phosphine resistance gene has been used to develop molecular markers. This has been done in Australia, India, USA and Turkey. Although multiple alleles in DLD that are related with phosphine resistance have been recorded in Australia, there was only one allele from other countries (P45/49S). From a practical point of view, these markers can be further examined to separate weak from strong resistance, which is a key issue is some areas, as strongly resistant individuals should be treated in different way. Overall, there are certain advantages on the use of molecular/genetic markers for resistance, over the use of the classical laboratory evaluations, such as the dose-bioassay protocols (i.e. exposure of the insects at different concentrations in jars). The most important advantage is that the molecular markers can provide early detection, and thus the potential of a population to develop resistance can be seen earlier than using bioassays.

THERE ARE RAPID DIAGNOSTICS AVAILABLE Apart from molecular markers that underline the presence of phosphine resistance genes, there are quick diagnostics that can provide accurate data in a very short period of time, usually hours or even minutes. These diagnostics are based on deviation of insects from normal movement and evaluate time to knockdown or immobilization. This is particularly important, as some of these diagnostics can be operated on site, e.g. by a fumigator in a flour mill prior the initiation of the fumigation. There are important advantages over the use of these diagnostics, as they are much less laborious than dose-response studies that last for days, and require specialized laboratories. It remains unclear, however, if these diagnostics can identify populations that are strongly resistant to phosphine. Nevertheless, these rapid diagnostics are important in the case of routine treatment in given facilities and commodities that are often fumigated.

ABIOTIC CONDITIONS ARE IMPORTANT Monitoring is essential in phosphine use. When it comes to monitoring, monitoring of the gas concentration is not the only thing that has to be monitored. Other parameters, such as temperature etc. should be monitored in parallel, as phosphine concentration has interesting diurnal circles according to the key abiotic conditions prevailing. Distribution of the gas is also important- even more important is monitoring of distribution. The traditional monitoring techniques are able to provide measurements for specific time intervals and at specific locations within the fumigated facilities. However, the measurements refer to the average concentrations, which are not always representative; in this sense it is the extreme values that are responsible for resistance development and not the average. Extreme dosing and absence of monitoring are key elements that are directly related with resistance development.

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