The effect of Rhopalosiphum padi and barley yellow dwarf virus on the wheat and aphid metabolome under increasing atmospheric CO2

Submission note: A thesis submitted in total fulfilment of the requirements for the degree of Doctor of Philosophy to the School of Applied Systems Biology, College of Science, Health and Engineering, La Trobe University, Bundoora.

The effects of CO2 fertilisation due to rising atmospheric CO2 in crop systems have been well studied, but little is known about how a changing climate will affect the specific interactions between plants, pests and pathogens. The work presented in this thesis investigates the effects of an Australian isolate of Barley yellow dwarf virus (BYDV-PAV) and its aphid vector, Rhopalosiphum padi, on wheat (Triticum aestivum L. cv. Yitpi) biochemistry under current ambient (aCO2; 400 micromol mol-1 ) and future predicted elevated CO2 concentrations (eCO2; 650 micromol mol-1 ). Growth chamber studies monitoring plant physiological factors and biochemistry, via metabolomics, were employed to dissect these changes. Untargeted (1H NMR) and targeted (LC-MS) approaches were utilised to explore changes in wheat primary metabolism due to aphid herbivory and virus infection. eCO2 generally increased plant growth, BYDV-PAV disease symptoms, and also enhanced host plant sugars (fructose, mannitol and trehalose), irrespective of disease status. Virus infection increased concentrations of several amino acids essential to the aphid diet (histidine, lysine, phenylalanine and tryptophan), irrespective of CO2 concentration. Further LC-MS/MS methods were employed to elucidate changes in wheat secondary metabolism resulting from aphid and virus interactions under eCO2. Nine metabolites were identified as having a statistically significant difference in concentration between treatments. These were mostly up-regulated with virus infection and are involved in either plant hormone synthesis or plant defence. Virus concentrations (titre) were assessed at different wheat growth stages in order to correlate virus load with an altered wheat metabolome. Compared to aCO2, eCO2 reduced virus titre in wheat; however, this was correlated with enhanced plant growth. An optimal feeding time for aphids under future eCO2 was identified at 20 days post inoculation, when peak virus concentrations are complimented by reduced defence-related metabolites. Finally, 1H NMR analyses was used to explore the effects of virus and eCO2 on the aphid metabolome. Eleven metabolites were identified and elevated alanine was an indicator of aphids harbouring virus, regardless of CO2 concentration. Lactate, aminobutyrate and glucose were associated with eCO2 in viruliferous aphids (compared to aCO2). Alterations in aphid weight and metabolite profiles were assumed to be due to altered host plant quality. In summary, information provided by this thesis indicates that wheat grown under eCO2 will likely influence aphid and BYDV-PAV interactions by altering host plant biochemistry. This is likely to have an impact on the spread of BYDV-PAV disease in the future.