Acute Oak Decline (AOD) is a complex Decline-disease affecting both native species of oak within the United Kingdom. Two novel species of bacteria, Gibbsiella quercinecans and Brenneria goodwinii, have been consistently isolated from necrotic lesions of AOD affected trees and are apparently absent from healthy oak trees. The aim of this thesis was to determine the role of Gibbsiella quercinecans and Brenneria goodwinii in AOD via the application of molecular methods. The first step in this investigation was to identify an appropriate molecular marker to differentiate between closely related oak lesion isolates belonging to the family Enterobacteriaceae. Using ITS1 profiling, a set of Enterobacteriaceae strains, primarily G. quercinecans and B. goodwinii were resolved, and eight strains were selected for sequencing to draft level. This enabled characterisation of strain level variation between sequenced isolates and functional annotation of genome encoded virulence factors, revealing an assortment of phytopathogenic virulence genes As the isolates were sequenced to draft level and no other whole genome sequences of G. quercinecans or B. goodwinii existed, the type strains of both species, i.e. G. quercinecans FRB97 and B. goodwinii FRB141 were sequenced to completion using the Pacific Biosciences RSII sequencing platform. The second component of this investigation characterised the putative phytopathogenic behaviour of G. quercinecans and B. goodwinii. Data collected from whole genome sequencing was used to align metagenomic and metatranscriptomic data recovered from the necrotic lesions of AOD affected trees, and to inform transcriptomic analysis of virulence gene expression within in vitro cultures containing oak tissue. The analysis demonstrated that G. quercinecans and B. goodwinii alter their behaviour in the presence of oak tissue, and active functionality of G. quercinecans and B. goodwinii within necrotic lesions AOD-affected trees. These experiments describe the first whole genome and transcriptome characterisation of G. quercinecans and B. goodwinii.