The Two-Component System DcuS-DcuR Is Involved in Virulence and Stress Tolerance in the Poplar Canker Bacterium Lonsdalea populi.

The gram-negative bacterium Lonsdalea populi causes an emerging poplar (Populus × euramericana) canker resulting in severe losses to poplar production in China and Europe. Two-component signal transduction systems play important roles in the regulation of virulence and stress responses in phytopathogenic bacteria. We identified a two-component pair (Lqp2625-Lqp2624) in L. populi, highly homologous to DcuS-DcuR of Escherichia coli. Mutants lacking DcuS or DcuR displayed normal growth while their virulence on poplar twigs was impaired. An inability to produce flagella indicated that DcuS and DcuR are involved in biofilm formation and swimming motility. Moreover, the loss of DcuS or DcuR led to increased sensitivity to oxidative stress and chloramphenicol through downregulation of genes associated with catalases and the multidrug efflux pump, suggesting that the two-component pair contributes to cellular adaptation to oxidative and antibiotic stresses. We identified key domains and putative phosphorylation sites important for virulence and stress responses. Our findings reveal the functions of DcuS-DcuR in virulence and stress responses in L. populi and provide increasing evidence that two-component systems are crucial during the infection process and stress adaptation in bacteria.

miR159a modulates poplar resistance against different fungi and bacteria.

Trees are inevitably attacked by different kinds of pathogens in their life. However, little is known about the regulatory factors in poplar response to different pathogen infections. MicroRNA159 (miR159) is a highly conserved microRNA (miRNA) in plants and regulates plant development and stress responses. Here, transgenic poplar overexpressing pto-miR159a (OX-159) showed antagonistic regulation mode to poplar stem disease caused by fungi Cytospora chrysosperma and bacteria Lonsdalea populi. OX-159 lines exhibited a higher susceptibility after inoculation with bacterium L. populi, whereas enhanced disease resistance to necrotrophic fungi C. chrysosperma compared with wild-type (WT) poplars. Intriguingly, further disease assay found that OX159 line rendered the poplar susceptible to hemi-biotrophic fungi Colletotrichum gloeosporioide, exhibiting larger necrosis and lower ROS accumulation than WT lines. Transcriptome analyses revealed that more down-regulated differentially expressed genes with disease-resistant domains in OX-159 line compared with WT line. Moreover, the central mediator NPR1 of salicylic acid (SA) pathway showed a decrease in expression level, while jasmonic acid/ethylene (JA/ET) signal pathway marker genes ERF, as well as PR3, MPK3, and MPK6 genes showed an increase level in OX159-2 and OX159-5 compared with WT lines. Further spatio-temporal expression analysis revealed JA/ET signaling was involved in the dynamic response process to C. gloeosporioides in WT and OX159 lines. These results demonstrate that overexpression of pto-miR159a resulted in the crosstalk changes of the downstream hub genes, thereby controlling the disease resistance of poplars, which provides clues for understanding pto-miR159a role in coordinating poplar-pathogen interactions.

Crosstalk of DNA Methylation Triggered by Pathogen in Poplars With Different Resistances.

DNA methylation plays crucial roles in responses to environmental stimuli. Modification of DNA methylation during development and abiotic stress responses has been confirmed in increasing numbers of plants, mainly annual plants. However, the epigenetic regulation mechanism underlying the immune response to pathogens remains largely unknown in plants, especially trees. To investigate whether DNA methylation is involved in the response to infection process or is related to the resistance differences among poplars, we performed comprehensive whole-genome bisulfite sequencing of the infected stem of the susceptible type Populus × euramerican '74/76' and resistant type Populus tomentosa 'henan' upon Lonsdalea populi infection. The results revealed that DNA methylation changed dynamically in poplars during the infection process with a remarkable decrease seen in the DNA methylation ratio. Intriguingly, the resistant P. tomentosa 'henan' had a much lower basal DNA methylation ratio than the susceptible P. × euramerican '74/76'. Compared to mock-inoculation, both poplar types underwent post-inoculation CHH hypomethylation; however, significant decreases in mC and mCHH proportions were found in resistant poplar. In addition, most differentially CHH-hypomethylated regions were distributed in repeat and promoter regions. Based on comparison of DNA methylation modification with the expression profiles of genes, DNA methylation occurred in resistance genes, pathogenesis-related genes, and phytohormone genes in poplars during pathogen infection. Additionally, transcript levels of genes encoding methylation-related enzymes changed during pathogen infection. Interestingly, small-regulator miRNAs were subject to DNA methylation in poplars experiencing pathogen infection. This investigation highlights the critical role of DNA methylation in the poplar immune response to pathogen infection and provides new insights into epigenetic regulation in perennial plants in response to biotic stress.

Genomic Characterization Provides an Insight into the Pathogenicity of the Poplar Canker Bacterium Lonsdalea populi.

An emerging poplar canker caused by the gram-negative bacterium, Lonsdalea populi, has led to high mortality of hybrid poplars Populus × euramericana in China and Europe. The molecular bases of pathogenicity and bark adaptation of L. populi have become a focus of recent research. This study revealed the whole genome sequence and identified putative virulence factors of L. populi. A high-quality L. populi genome sequence was assembled de novo, with a genome size of 3,859,707 bp, containing approximately 3434 genes and 107 RNAs (75 tRNA, 22 rRNA, and 10 ncRNA). The L. populi genome contained 380 virulence-associated genes, mainly encoding for adhesion, extracellular enzymes, secretory systems, and two-component transduction systems. The genome had 110 carbohydrate-active enzyme (CAZy)-coding genes and putative secreted proteins. The antibiotic-resistance database annotation listed that L. populi was resistant to penicillin, fluoroquinolone, and kasugamycin. Analysis of comparative genomics found that L. populi exhibited the highest homology with the L. britannica genome and L. populi encompassed 1905 specific genes, 1769 dispensable genes, and 1381 conserved genes, suggesting high evolutionary diversity and genomic plasticity. Moreover, the pan genome analysis revealed that the N-5-1 genome is an open genome. These findings provide important resources for understanding the molecular basis of the pathogenicity and biology of L. populi and the poplar-bacterium interaction.

Molecular Aspects of an Emerging Poplar Canker Caused by Lonsdalea populi.

The Gram-negative bacterium Lonsdalea populi causes a lethal disease known as bark canker on Populus × euramericana in China and Europe. Typical symptoms of bark canker include an abundant white-colored fluid, which oozes from the infected tissues. The availability of the genomic sequence of the bacterium provided the necessary resource to launch genome-scale investigations into the mechanisms fundamental to pathogenesis. Functional analyses of a diverse group of genes encoding virulence factors and components of signaling pathways indicate that successful bark infection depends on specific responses by the pathogen to various stresses, including oxidative stress. Although physiology of resistance is well studied, the molecular processes underlying the defense responses and the genetic basis of resistance to L. populi and in other poplar species remain largely unknown. Control of the disease has relied on chemical measures. Due to the genetic amenability of Lonsdalea and poplar, this pathosystem will become an important model system to unravel molecular mechanisms of bacterial pathogenicity on woody plants. Increased understanding of pathogenesis and signaling in the interaction will facilitate the management of this kind of poplar canker.