Abstract
Candidatus Liberibacter solanacearum (CLSo) is an unculturable pathogenic bacteria that infects potato plants and tubers, inducing significant biochemical and metabolic changes for defensive purposes. These changes result in a dark brown striped post-fry colouration in tubers, and commonly referred to as ‘Zebra chip’ in the literature. Potatoes are among the most important food crops in the world and the fourth largest export crop from New Zealand; this means CLSo is an important crop disease which causes millions of dollars of loss to the potato industry and significant food waste. This thesis aimed firstly to broaden the understanding of the biochemical and metabolic effects of CLSo in potato tubers; and secondly, to determine if the post-fry symptoms of infection could be mitigated by the application of the postharvest technology, pulsed electric fields (PEF).
Firstly, it was identified that there were limited quantitative measures of CLSo symptom severity in the literature. Most authors relied upon a subjective visual affirmation of the disease severity with a rating system from 0 to 3, where 0 = no symptoms and 3 = severe symptoms. Chapter 2 of this thesis used image analysis to develop a quantitative colour threshold that could be used to categorise tubers into two groups of severity. This method successfully distinguished two groups of tubers based upon their surface area colouration after frying, and was used as a categorisation tool for Chapters 3, 5 and 6. In addition this method was developed to be cross-cultivar applicable by using two New Zealand grown cultivars, Agria and Russet Burbank, that exhibited very different raw tuber symptoms.
Chapters 3 and 5 were designed to provide a better understanding of CLSo infection at both an early stage of storage and after a three month period. Using gas chromatography (GC) and liquid chromatography with high resolution mass spectrometry (LC–HRMS), the free amino acids, targeted phenolic acids and untargeted metabolites were compared between CLSo negative and positive tubers with reference to the severity categorisation tool from Chapter 2. It was found that there were significant cultivar effects on the upregulation of free amino acids during the early period of storage, where Agria exhibited significantly higher responses to infection compared to Russet Burbank. The importance of the phenylpropanoid pathway was highlighted as a significant defence mechanism in potato tubers in response to CLSo, and still played an important role in defence after three months in storage.
A novel sampling technique was developed to determine the level of antioxidant enzyme activity of CLSo infected tubers in Chapter 4. Small, 5 mm cores were excised from CLSo infected tubers in asymptomatic regions and from regions presenting post fry symptoms. The comparisons between these tissues provided a more in-depth view of the enzymatic antioxidant stress response than what has been previously identified. It was observed that symptomatic tissues had significantly higher activity levels of the enzymes with the function of hydrogen peroxide detoxification compared to asymptomatic tissues. Furthermore, the similarities that were observed between cultivars suggested these responses are a baseline defence mechanism in potato tubers.
The post-fry browning symptoms of CLSo infection in potato tubers is caused by the Maillard reaction between reducing sugars and free amino acids at high heat. The levels of these precursor compounds are higher in CLSo infected tubers for defensive purposes. However, the level of browning that occurs during processing due to their upregulation is unacceptable for consumers. Chapter 6 used PEF to increase the level of glucose and free amino acid leaching from CLSo infected tubers during blanching. The application of PEF reduced the overall level of these compounds in CLSo infected tubers. Using the colour categorisation tool developed in Chapter 2, it was observed that PEF treatment in combination with Blanching treatment produced the greatest number of CLSo infected tubers with acceptable post-fry colouration than either treatment alone. Each cultivar responded slightly differently, where 19% of CLSo infected Agria and 37% of Russet Burbank tubers had an acceptable colour rating after PEF + Blanching treatment. This was an improvement from the 6% of Agria and 3% of Russet Burbank tubers that were considered acceptable before treatment. It was postulated that there were some underlying biochemical or metabolic defensive responses that limited the improvement of the post-fry colour for some tubers even after treatment. It can be suggested that the increased levels of free amino acids and glucose in tubers are not the only factors to consider for the application of this technology to CLSo infected tubers.
Overall, this thesis contributes valuable insight into the biochemical and metabolic changes induced in CLSo infected tubers. The methods developed in this thesis could have wider application to improve future CLSo studies through improving sampling specificity and introducing more quantitative categorisation techniques. These findings can be used to strengthen current CLSo research and provide new insight into how potato tubers respond to disease. However, more research is needed to identify other localized biochemical and metabolic changes in potato tubers, to determine if PEF could prove more effective as a pre-processing tool to mitigate CLSo symptoms in fried potato tubers.