Calcium transcriptionally regulates the biofilm machinery of Xylella fastidiosa to promote continued biofilm development in batch cultures

Abstract

The functions of calcium (Ca) in bacteria are less characterized than in eukaryotes, where its role has been studied extensively. The plant-pathogenic bacterium Xylella fastidiosa has several virulence features that are enhanced by increased Ca concentrations, including biofilm formation. However, the specific mechanisms driving modulation of this feature are unclear. Characterization of biofilm formation over time showed that 4 mM Ca supplementation produced denser biofilms that were still developing at 96 h, while biofilm in non-supplemented media had reached the dispersal stage by 72 h. To identify changes in global gene expression in X. fastidiosa grown in supplemental Ca, RNA-Seq of batch culture biofilm cells was conducted at three 24-h time intervals. Results indicate that a variety of genes are differentially expressed in response to Ca, including genes related to attachment, motility, exopolysaccharide synthesis, biofilm formation, peptidoglycan synthesis, regulatory functions, iron homeostasis, and phages. Collectively, results demonstrate that Ca supplementation induces a transcriptional response that promotes continued biofilm development, while biofilm cells in nonsupplemented media are driven towards dispersion of cells from the biofilm structure. These results have important implications for disease progression in planta, where xylem sap is the source of Ca and other nutrients for X. fastidiosa.

Calcium transcriptionally regulates movement, recombination and other functions of Xylella fastidiosa under constant flow inside microfluidic chambers

“… –, No changes; Down, downregulated gene expression; Flask, Flask‐RNA‐Seq (Parker et al , ); MC, MC‐RNA‐Seq (this study); and Up, upregulated gene expression. …”

“… Comparison of the effect of Ca on gene expression profiles of X. fastidiosa grown in microfluidic chamber (MC‐RNA‐Seq) and batch cultures (Flask‐RNA‐Seq, Parker et al, ). A. Pie charts for distribution of differentially expressed genes (DEGs) in different functional categories between MC‐RNA‐Seq and Flask‐RNA‐Seq.…”

“…These genes are located in a putative genomic island comprising PD0906‐PD0943 (Parker et al , ). 21 out of the 38 genes in this island do not have homologues in the closely related avirulent strain X. fastidiosa EB92‐1 (Zhang et al , ; Parker et al , ). These findings suggest that the role of these phage‐related genes may be associated with twitching motility, biofilm formation and virulence of X. fastidiosa , but to confirm this, further investigation is needed.…”

“…Calcium‐enhanced twitching motility is associated with the interaction between Ca and the Ca‐binding protein PilY1 of the type IV pili (TFP) (Cruz et al , ). Whole transcriptome analysis of X. fastidiosa biofilm cells in batch culture at different Ca concentration showed that genes involved in attachment and biofilm formation maintained a higher expression level in Ca‐supplemented media; in contrast, these gene expression levels were gradually decreased in non‐supplemented media, which indicates Ca promotes continued biofilm development in X. fastidiosa (Parker et al , ). Unravelling the molecular basis of the effect of Ca on virulence‐related traits of X. fastidiosa is essential for understanding the biology of X. fastidiosa in plant xylem.…”

In vitro Determination of Extracellular Proteins from Xylella fastidiosa

“…Similarly, Parker et al (2016) showed that calcium supplementation changed the global expression of genes that promote continued biofilm development. These genes are related to attachment, motility, exopolysaccharide synthesis, biofilm formation, peptidoglycan synthesis, regulatory functions, iron homeostasis, and phages.…”