Microbial interactions in the kiwifruit phyllosphere

The plant microbiome is well recognised for its beneficial roles in promoting plant growth and enhancing resistance to biotic stresses, including pathogen attack. Antagonistic interactions between beneficial microbes and pathogens may occur via antibiotic production, competition for nutrients and space, and the activation of plant defence mechanisms. The bacterial communities inhabiting the phyllosphere are acquired through both horizontal transmission from the environment and vertical transmission from parent plants, often via seeds or reproductive structures, making environmental factors crucial determinants of microbial community composition. Pseudomonas syringae pv. actinidiae (Psa) is the causal agent of bacterial canker disease in kiwifruit plants. The kiwifruit export industry represents a significant component of New Zealand’s economy. Although the 2010 Psa outbreak was mitigated primarily by developing more resistant kiwifruit varieties, the pathogen remains widespread in many orchards. Current control measures rely on unsustainable practices, raising concerns over the emergence of antibiotic-resistant Psa strains and potential environmental impacts. Bacterial canker disease initiates with an epiphytic phase in which Psa colonises the leaf surface. Upon reaching a critical population density, Psa penetrates the plant through natural openings such as stomata. Chemotaxis, the directed movement of bacteria in response to chemical gradients, is used to find these openings. In the context of plants, this movement is often guided by plant-derived compounds, allowing pathogens to locate entry points such as stomata or wounds. The link between chemotaxis and pathogenicity is well established in numerous host-pathogen systems, including Pseudomonas syringae. The overall aim of this thesis was to investigate the bacterial communities on kiwifruit leaves (phyllosphere) and their interactions with the pathogen Pseudomonas syringae pv. actinidiae. In Chapter 3, I examined the impact of adjacent land use on phyllosphere bacterial communities by comparing orchards bordered by maize fields to those surrounded by native bush. Using sequencing on the Oxford Nanopore Technologies’ MinION platform, I detected a statistically significant but modest difference in community composition, with orchards bordered by maize fields harbouring more homogenic bacterial communities compared to those surrounded by native bush. Chapter 4 focused on isolating and characterising antagonistic bacteria. Phyllosphere bacteria from kiwifruit leaves were isolated and assessed for their ability to inhibit Psa growth in swim medium assays. Five antagonistic isolates were selected for further study of their inhibitory effects. The results revealed diverse inhibitory mechanisms, with some isolates acting in a cell density-dependent manner and others being effective at low densities. In Chapter 5, whole-genome sequencing of the selected antagonistic isolates was performed to assess their genetic potential for plant protection and microbial interactions. Analysis of gene clusters involved in secondary metabolite biosynthesis, quorum sensing, and motility confirmed that all strains shared key traits associated with their antagonistic activity, despite wider genomic differences. Together, these findings demonstrate that the kiwifruit phyllosphere hosts highly diverse bacterial communities, with members capable of inhibiting Psa. These microbial communities hold promise as a reservoir for developing novel biocontrol strategies against bacterial canker disease.