This is the report from a BSPP Undergraduate ‘Vacation’ Bursary.
Click here to read more/apply for one yourself.
Heavy metal hyperaccumulating plants have unique physiology adapted to metalliferous soils. They actively uptake and store the heavy metals at a level toxic to typical plants, which is suggested to kill and be less palatable to herbivores and pathogens. Noccaea caerulescens is studied as a model hyperaccumulator; it accumulates nickel, zinc and cadmium to different degrees depending on the population. The plant pathogen we studied in interaction with N. caerulescens is Pseudomonas syringae. This bacterial pathogen has a great genetic diversity and a wide host range. The strains we used are P. syringae pv. maculicola (Psm) from the UK, a strain isolated from cauliflower (Brassica oleracea var. botrytis), which is also capable of infecting N. caerulescens, and P. syringae pv. syringae B11 (B11) isolated from a natural population of N. caerulescens in an abandoned mining site in Snowdonia.
The Preston lab has provided evidence for the elemental defence hypothesis. It was previously shown that increase in zinc in N. caerulescens is correlated with elevated reactive oxygen species (ROS) levels in leaves. Additionally, it has been found in unpublished work that the combined stress of zinc and hydrogen peroxide (a ROS) is more lethal to P. syringae, and zinc has been shown to cause P. syringae to be less able to recover from oxidative stress. My project aimed to discover how nickel and ROS might affect P. syringae in vitro, and how environmental nickel availability might impact N. caerulescens in coping with P. syringae infection.
I have performed many different techniques combining both in vitro microbiology experiments and in planta microscopy. For the microbiology aspect, I have performed plate tolerance assays studying the growth responses of B11 and Psm under nickel stress; I also used roGFP (reduction-oxidation sensitive green fluorescent protein), a redox biosensor transformed B11 to investigate its oxidative response to the combined stresses of ROS and nickel. Colony counts and phenotyping techniques were also used to determine the characteristics of the bacterial population after the combined stress. Luminescence imaging was used to track the growth and spread of lux-YFP transformed Psm in infected leaves using their bioluminescence. In addition, confocal microscopy was carried out using the fluorescent properties of the bacteria to image with higher resolutions down to individual bacteria.
Results showed no observable differences in oxidative and growth responses between nickel concentrations of 0.4 mM and 0.8 mM when combined with H2O2 to B11 roGFP, although less growth were observed with increasing nickel concentrations without H2O2. In addition, the compound effect between nickel and H2O2 introduced greater oxidative stress than H2O2 alone. This combination was slightly less lethal than the combination of zinc and hydrogen peroxide. The B11 strain was able to recover from the oxidated state induced by hydrogen peroxide in the presence of nickel, but it could not recover in the presence of zinc. An increase in nickel concentration caused a decrease in growth rate and final population density, and B11 was more tolerant to nickel than Psm. Confocal imaging suggested younger plants are more susceptible to infection, but more replications are needed. Luminescence imaging showed inconclusive results in bacterial colony distributions between nickel plants grown in different nickel concentrations. The first replicate experiment indicated the expected trend that N. caerulescens grown with less nickel are more susceptible to infections. However, the second replicate showed no clear trends, and more replicates are needed.
I am grateful for this invaluable opportunity from BSPP. It gave me more confidence in going into postgraduate research. I am also very thankful for my supervisors, Professor Gail Preston and Rose Bourdon, who taught me practical techniques and key qualities to be a good researcher, such as curiosity and attention to detail.
Anna (Yuyi) Li
This is the report from a BSPP Undergraduate ‘Vacation’ Bursary.
Click here to read more/apply for one yourself.
