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Metabolic-stress disinfection and disinfestation (MSDD) is a Methyl Bromide (MeBr) alternative disinfestation technology that utilises the combination of pressure change, hypercarbia, hypoxia and ethanol treatments. The thesis describes the investigation of the insecticidal efficacy of MSDD against various insect pests; longtailed mealybug (Pseudococcus longispinus), 5th instar light brown apple moth (LBAM, Epiphyas postvittana) larvae and 5th instar codling moth larvae (Cydia pomonella). MSDD consists of 2 phases, namely a physical phase and followed by a chemical phase. The physical phase is characterised by cycles of pressure changes (90-110 kPa) carried out by drawing air from the treatment chamber and replacing it with ballast gases (CO2 or N2). The physical phase usually lasts for 30 min. At the end of the physical phase, the chemical phase is initiated by drawing the pressure down to 10 kPa and ethanol is introduced into the chamber (held for 60 min). This 90 min MSDD treatment protocol was shown to be effective in controlling surface pests. Further investigation showed parameters associated to both the physical phase (the length of the physical phase) and the chemical phase (the concentration of ethanol, the length of chemical phase and the pressure of chemical phase) were required to ensure high mortality of pests. The efficacy of MSDD was limited to surface pests. Inoculation of 5th instar codling moth larvae into the apples severely limited the capacity of MSDD to disinfest. Low mortality was shown to be correlated with the inability of ethanol to penetrate into apple flesh. Ethanol only penetrated 1 cm (from the skin) into the apple flesh. Investigation into the protein changes in the haemolymph of LBAM larvae revealed that the insecticidal efficacy of MSDD is strongly linked to the chemical phase and its ability to disrupt melanisation regulation. MSDD prevented the up-regulation of alaserpin and induced the over-expression of serine protease. Serpin family has been shown to negatively regulate melanisation, whereas serine protease has been shown to promote melanisation. By disrupting this regulation, MSDD induced excessive melanisation in the larvae which was evident in the physical appearance of the treated larvae. Melanisation is an innate immune response against pathogens in most arthropods. However, excessive melanisation has been shown to be cytotoxic to insects. Research into the effect of MSDD on fruit physiology and quality showed that MSDD had adverse effect on 'Hayward' and 'Hort 16A' kiwifruit. The most significant result was the increase in rot incidence in both cultivars. MSDD also resulted in disorders such as flesh breakdown and uneven ripening in the 'Hort 16A' fruit. Ripening (and softening) was also accelerated in the non-cool stored 'Hayward' kiwifruit. Conversely, MSDD treatments did not have noticeable effect on the physiology and quality of 'Cripps Pink' apple and 'Hass' avocado. Ripening physiology and quality parameters, such as rot incidence, internal browning (apple) and flesh greying (avocado) were not affected by MSDD treatments. Packaging materials restricted the efficacy of MSDD. Materials with high ethanol sorption such as cardboard (kiwifruit boxes) reduced the mortality of insects treated with MSDD significantly. Exposure to ethanol was also demonstrated to be a critical factor to ensure high mortality. 'Bulk' treatment using a harvest crate showed that MSDD only managed to disinfest the first layer of fruit. Mortality steeply decreased in the subsequent layers of fruit. Therefore, a single layer fruit tray was needed to ensure high mortality. In conclusion, MSDD treatments were shown to be effective in controlling insect pests. Both the physical and chemical phases were important in achieving high insect mortality. The MSDD mode of action is proposed to be involving the disruption of the melanisation regulation in the form of up-regulation and down-regulation of serine protease and its inhibitor (serpin), respectively. The efficacy of MSDD was limited to surface pests, non-absorbing and single layered packaging materials. 'Cripps Pink' apple and 'Hass' avocado were MSDD tolerant. Conversely, 'Hayward' and 'Hort 16A' were susceptible to over- production of ethanol and flesh breakdown disorder in response to MSDD treatments, respectively. |
