The HD-GYP Domain Protein RpfG of Xanthomonas oryzae pv. oryzicola Regulates Synthesis of Extracellular Polysaccharides that Contribute to Biofilm Formation and Virulence on Rice

Yuanbao Zhang,Yunyue Wang,John Maxwell Dow,Wendi Jiang,Lei Wang,Churui Li,Chao Wei,Wenxian Sun

doi:10.1371/journal.pone.0059428

Abstract

Bacterial leaf streak caused by Xanthomonas oryzae pv. oryzicola (Xoc) is one of the most important diseases in rice. However, little is known about the pathogenicity mechanisms of Xoc. Here we have investigated the function of three HD-GYP domain regulatory proteins in biofilm formation, the synthesis of virulence factors and virulence of Xoc. Deletion of rpfG resulted in altered production of extracellular polysaccharides (EPS), abolished virulence on rice and enhanced biofilm formation, but had little effect on the secretion of proteases and motility. In contrast, mutational analysis showed that the other two HD-GYP domain proteins had no effect on virulence factor synthesis and tested phenotypes. Mutation of rpfG led to up-regulation of the type III secretion system and altered expression of three putative glycosyltransferase genes gumD, pgaC and xagB, which are part of operons directing the synthesis of different extracellular polysaccharides. The pgaABCD and xagABCD operons were greatly up-regulated in the Xoc ΔrpfG mutant, whereas the expression of the gum genes was unaltered or slightly enhanced. The elevated biofilm formation of the Xoc ΔrpfG mutant was dramatically reduced upon deletion of gumD, xagA and xagB, but not when pgaA and pgaC were deleted. Interestingly, only the ΔgumD mutant, among these single gene mutants, exhibits multiple phenotype alterations including reduced biofilm and EPS production and attenuated virulence on rice. These data indicate that RpfG is a global regulator that controls biofilm formation, EPS production and bacterial virulence in Xoc and that both gumD- and xagB-dependent EPS contribute to biofilm formation under different conditions.

Highlights

Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak (BLS) in rice, one of the most important bacterial diseases in tropical and subtropical Asia, some parts of Africa, as well as ricegrowing regions of northern Australia
BLAST searches revealed that three genes encode HD-GYP domain proteins in Xoc BLS256; these are XOC2264, XOC1984, and XOC4564
To determine the effect of these HD-GYP domain proteins on bacterial behaviors and in vivo virulence of Xoc, the single, double and triple mutants involving rpfG, hgdA and hgdC genes were constructed as described in Materials and Methods and confirmed by Southern blot analyses (Figure S2)

Summary

Introduction

Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak (BLS) in rice, one of the most important bacterial diseases in tropical and subtropical Asia, some parts of Africa, as well as ricegrowing regions of northern Australia. Oryzicola (Xoc) causes bacterial leaf streak (BLS) in rice, one of the most important bacterial diseases in tropical and subtropical Asia, some parts of Africa, as well as ricegrowing regions of northern Australia. The BLS disease can cause yield loss up to 30% in epidemic years [1,2,3]. Xoc does not invade the xylem, which is in contrast to another rice bacterial pathogen Xanthomonas oryzae pv. Oryzae (Xoo) that causes bacterial blight by invading vascular tissues [2]. Interactions between Xoo/Xoc and rice have become models for understanding fundamental aspects of bacterial pathogenesis in host plants and plant disease resistance, as well as functional and comparative genomics in microbial biology [4]

Methods

Results

Conclusion