The development of an immunoassay to identify and quantify the two species of the potato Cyst Nematode, G.rostochiensis & G.pallida.

Abstract

The potato cyst nematodes (PCN) G.rostochiemis (Woll.) and G.pallida (stone) are sedentary endoparasites and serious economic pests of potato crops throughout the temperate world. Therefore there is an agricultural need for a quick, cheap and user- friendly method to assess the specific population density in the field. This thesis describes the development of just such a diagnostic immunoassay procedure. Firstly two monoclonal antibodies (Mabs), MR8/4 and MR8/5 were selected from a screen of hybridoma lines for their specificity to G.pallida and G.rostochiensis respectively. The specific nature of the antigen and antibody binding was then determined before a procedure was standardized to identify and quantify PCN in both pure and mixed populations. Further tests proved that the Mabs only recognized live eggs. A procedure was then developed to extract the nematode antigen direct from soil samples. The robustness of the extraction procedure and immunoassay was confirmed by comparison with the lengthier traditional flotation and counting by microscope method. The traditional counting procedure does not take into account the viability of the cysts therefore the new extraction procedure and immunoassay was used to determine the efficiency of nematicide treatment from a field fumigation trial. This confirmed the influence of soil type and conditions at time of application on fumigation efficiency. Finally a methodology was evolved to convert the assay procedures into a marketable commercial technique.