A study was conducted to develop an ecofriendly and sustainable fungicide based on submicron chitosan dispersions using low molecular weight chitosan and locally prepared chitosan. The physicochemical properties of both submicron dispersions were determined and their antimicrobial effect on Colletotrichum gloeosporioides was also investigated. The antifungal effect was further demonstrated by the production of fungal cell wall degrading enzymes and by the production of defence related enzymes in the plants along with the effect of submicron chitosan dispersions on the vegetative growth of plants. The effect of submicron chitosan dispersions on biochemical and physiological responses of dragon fruit during storage at 10 ± 2 °C and 80 ± 5% relative humidity for 28 days was also investigated. In vitro antifungal activity of submicron chitosan dispersions showed that low molecular weight chitosan has better antifungal properties than the locally prepared chitosan and therefore was selected for further studies. However, 600 nm droplets of 1.0% chitosan showed promising results in terms of suppressing mycelial growth (90.2%), conidial germination (93.1%) and reducing dry weight of mycelium (37.9%). Similarly, in vivo studies showed that the 600 nm droplets of 1.0% chitosan helped to reduce the disease incidence by up to 33.0 %. Beside the direct antifungal effect, this study suggests that the submicron chitosan dispersions enhanced host resistance through the production of defence related compounds, such as total phenols, peroxidase (PO), polyphenol oxidase (PPO), phenylalanine ammonia lyase (PAL) and also the activity of pathogenesis related proteins, such as β, 1-3 glucanase and chitinase. Treatments with submicron chitosan dispersions as 600 nm droplets of 1.0% chitosan showed more vigorous growth of plants as compared to the control. The submicron chitosan dispersions with 600 nm droplets of 1.0% chitosan helped to reduce the incidence of disease on fruit by up to 94% as compared to the control. The biochemical and physiological studies of dragon fruit after 28 days of storage showed that weight loss of the fruit treated with 600 nm droplets of 1.0% chitosan compared with the control reduced two fold but this was not significantly different from the conventional chitosan. The same trend was observed in all the parameters tested, including antioxidant activity. Control of disease using submicron chitosan dispersions has the potential to add market value to fresh produce. The findings from all the experiments showed that 600 nm droplets of 1.0% chitosan help to reduce anthracnose in the field and also during postharvest storage. Thus, 600 nm droplets of 1.0% chitosan could be used commercially in fields of dragon fruit as a green fungicide. Being non-toxic and biodegradable, chitosan has the potential to become an alternative to synthetic fungicides for protecting fruit crops and thus assisting sustainable agriculture.

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The ability to perform nucleic acid-based detection of plant pathogens away from conventional laboratory facilities has the potential to be beneficial in situations where results are required very rapidly or where resources and access to laboratory equipment are limited. Methods for use in such situations must combine sensitivity and specificity with rapid and simple workflows. The aim of this project was to investigate aspects of on-site testing for plant pathogens by developing detection methods for a range of target species. Detection methods based on loop-mediated isothermal amplification (LAMP) exhibit characteristics which make them potentially suitable for on-site testing. LAMP-based methods were developed for detection of plant pathogens with three potential non-laboratory testing scenarios in mind: testing during plant health inspection (assays for Phytophthora ramorum, P. kernoviae and Guignardia citricarpa); testing to assess inoculum levels in the processing of plant products (an assay for Botrytis cinerea); and testing in under-resourced settings (assays for Cassava brown streak virus and Ugandan cassava brown streak virus). In developing these detection methods, attempts were made to address some of the specific requirements of potential end-users of the tests in each case. For testing in the context of inspection, a particular emphasis was placed on the need for simple, rapid methods for nucleic acid extraction. As well as investigating the use of rapid extraction methods in conjunction with LAMP, work was also carried out to investigate how on-site nucleic acid extraction using lateral flow devices could be integrated with current field and laboratory testing for P. ramorum.

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