<i>Xanthomonas</i> biofilm rupture by DNAse I.

<p>A, DNAse treatment of 1 h duration. B, DNAse treatment for an overnight period. The absorbance values were normalized to the control XVM2 plus buffer in order to compare the response for different strains. Error bars represents the standard deviation. Graphs are a representative assay of at least three assays with three replicates per assay</p

Presence of eDNA at the early stages of biofilm formation in <i>Xcc</i>.

<p>Representative light (crystal violet, CV staining) and fluorescence (SYTO-9 staining) images of 72 h static cultures on LB or XVM2 media. Fibers were observed after both staining for strains <i>Xcc</i> 306 and <i>Xcc</i> 12879 A^w at the early stages of biofilm formation. Fibers interconnected cells at different stages of aggregation, from one to several cells are shown. In XVM2 medium, fibers were thicker and more uniform after staining with CV and SYTO-9. eDNA in XVM2 medium appeared to cover the surface like a sheet in contrast to individual fibers produced in LB medium.</p

Strains of <i>Xanthomonas</i> spp. used in the study.

<p>Strains of <i>Xanthomonas</i> spp. used in the study.</p

Presence of eDNA in preformed biofilms of <i>Xcc</i>.

<p>Representative light (CV staining) and fluorescence (SYTO-9 staining) images of mature biofilms on LB or XVM2 media. Both <i>Xcc</i> 306 and <i>Xcc</i> 12879 A^w strains were more aggregated in XVM2 than LB. A high level of aggregation for strain <i>Xcc</i> 12879 A^w in XVM2 made it difficult to observe eDNA fibers in aggregates. In XVM2, strain <i>Xcc</i> 306 was less aggregated and CV and SYTO-9 staining revealed eDNA surrounding the cells. In LB both staining revealed long fibers interconnecting aggregates.</p

Presence of Extracellular DNA during Biofilm Formation by <i>Xanthomonas citri</i> subsp. <i>citri</i> Strains with Different Host Range

<div><p><i>Xanthomonas citri</i> subsp. <i>citri</i> (<i>Xcc</i>) A strain causes citrus bacterial canker, a serious leaf, fruit and stem spotting disease of several <i>Citrus</i> species. <i>X</i>. <i>alfalfae</i> subsp. <i>citrumelonis</i> (<i>Xac</i>) is the cause of citrus bacterial spot, a minor disease of citrus nursery plants and <i>X</i>. <i>campestris</i> pv. <i>campestris</i> (<i>Xc</i>) is a systemic pathogen that causes black rot of cabbage. <i>Xanthomonas</i> spp. form biofilms <i>in planta</i> that facilitate the host infection process. Herein, the role of extracellular DNA (eDNA) was evaluated in the formation and stabilization of the biofilm matrix at different stages of biofilm development. Fluorescence and light microscopy, as well as DNAse treatments, were used to determine the presence of eDNA in biofilms and bacterial cultures. DNAse treatments of <i>Xcc</i> strains and <i>Xac</i> reduced biofilm formation at the initial stage of development, as well as disrupted preformed biofilm. By comparison, no significant effect of the DNAse was detected for biofilm formation by <i>Xc</i>. DNAse effects on biofilm formation or disruption varied among <i>Xcc</i> strains and <i>Xanthomonas</i> species which suggest different roles for eDNA. Variation in the structure of fibers containing eDNA in biofilms, bacterial cultures, and in twitching motility was also visualized by microscopy. The proposed roles for eDNA are as an adhesin in the early stages of biofilm formation, as an structural component of mature bacterial aggregates, and twitching motility structures.</p></div

Effect of DNAse I during biofilm formation in <i>Xanthomonas</i>.

<p>Biofilm formation after treatment with DNAse I at different times after bacterial seeded with different <i>Xanthomonas</i> species and strains (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0156695#pone.0156695.t001" target="_blank">Table 1</a>). A, DNAse added at 0 h post seeding (hps); B, DNAse added at 24 hps; C, DNAse added at 48 hps; and D, DNAse added at 72 hps. The absorbance values were normalized to the control XVM2 plus buffer in order to compare the response for different strains. Error bars represent the standard deviation. Graphs are a representative assay of at least three assays with three replicates per assay. Statistical analyses were performed using STATGRAPHICS Plus, version 5.1 (Copyright Manugistics Inc.).</p

Effect of DNAse in the extracellular matrix of <i>Xcc</i>.

<p>Transmission Electron Microscopy of <i>Xcc</i> 306 and 12879 A^w strains in exponential growth phase, after DNAse and no-DNAse (Buffer) treatments. DNAse treated bacteria showed less extracellular structures than de buffer control. The remaining structures observed after treatment could be associated to extracellular proteins. Bacterial cell division is shown under DNAse treatment.</p