IDENTIFICATION OF TRAITS OF XYLELLA FASTIDIOSA CONFERRING VIRULENCE TO GRAPE AND INSECT TRANSMISSION BY ANALYSIS OF GLOBAL GENE EXPRESSION USING DNA MICROARRAYS

• Author(s): Lindow, Steven; Feil, Bill; Wang, Nian;
• Abstract: Xylella fastidiosa (Xf) regulates virulence factors important in both virulence to grape as well as colonization of sharpshooter vectors via its production of a fatty acid molecule (known as DSF) whose production is encoded by rpfF. The RpfF homologue of Xf strains that cause Pierces disease (PD), synthesizes a fatty acid cell-cell signal (DSF) that is apparently similar to that produced by Xanthomonas campestris pv. campestris (Xcc). Xf rpfF mutants exhibit increased virulence to plants, however, they are unable to be spread from plant to plant by their insect vectors. While we have identified a key regulator of virulence and insect transmission in Xf we lack an understanding of the traits that are regulated by this pathogen in response to the DSF signal molecule. We thus are initiating studies to determine the rpf-regulation in Xf. We are exploiting a DNA microarray developed in another project that addresses host specificity genes in Xf to assess gene expression differences in isogenic RpfF(+) and RpfF(-) strains of Xf strain Temecula. The microarray contains 2555 gene-specific 70 bp oligodeoxynucleotides including negative and positive controls. Microarray analysis was performed to identify genes that are controlled by DSF and/or RpfC. DSF bioassay with reporter strain Xcc 8523 (pKLN55) indicated that DSF production is most abundant 10 days after inoculation when rpfF expression is most active. Preliminary results reveal that at least 124 genes are controlled in response to rpfF in Xf, including those encoding gum production, type IV pili and hemagglutinin. Clearly this regulator has a large effect on the physiological function of Xf. Microarray analysis revealed that more than 300 genes are also controlled by RpfC, including some of the same genes regulated by rpfF as well as genes such as tonB. Microarray-based gene expression results are being verified using quantitative Reverse Transcriptase-PCR. Comparison of the RpfF and RpfC regulons reveals that a complex pattern of expression of potential virulence genes contribute to the virulence of Xf and explains the hypervirulence of rpfF mutants and the reduced virulence of rpfC mutants. Work is also underway to determine the subset of Xf genes that is plant-inducible and the identity of those whose plant-inducible genes whose expression is also dependent on DSF production.
• Publication Date: Nov 2006
• Journal: 2006 Pierce's Disease Research Symposium