Identification of bacteria associated with the acute oak decline syndrome in England using Next Generation Sequencing. Bacteria associated with acute oak decline

Outbreaks of acute oak decline (AOD) have been documented in England from 2006 and this condition is spreading through the Midlands and South East England. Both species of native oaks (Quercus robur and Quercus petraea) are affected. At Forest Research, certain bacteria were consistently isolated from symptomatic trees which are thought to cause necrotic patches in the bark and sapwood. To complement isolation efforts for identification of putative causative biotic agents and increase our understanding of bacterial communities associated with oak tissue in general, 5 different sites in England namely Attingham Wood, Runs Wood, Great Monks Wood, Stratfield Brake and Bisham Wood were chosen for this study. Samples of outer bark, inner bark, sapwood and heartwood were taken from healthy oak and trees with symptoms at varying stages of the syndrome. Furthermore, larval galleries were included in the study. All samples were subjected to molecular analysis using Next Generation Sequencing (NGS). After DNA extraction and purification the V3-V5 fragment of the 16S rRNA gene was amplified followed by pyrosequencing. The dataset was analysed by QIIME followed by multivariate statistical methods to identify patterns in bacterial communities between the different sites and syndrome stages. Overall, the composition of bacterial communities differed greatly according to the geographical location of their hosts. Within each site the composition of the bacteria associated with symptomatic tissue varied between advanced stages of the syndrome and healthy tissue. Key players in healthy and symptomatic tissue were identified and included members of the Gammaproteobacteria and Firmicutes.