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FadP affects the virulence of Ralstonia pseudosolanacearum by altering nitrogen metabolism processes and reducing motility, cellulase, and extracellular polysaccharide production

  • Jia-chun Wu
  • Min Zhong
  • Xing-wei Li
  • Cheng-peng Yu
  • Qiong-guang LiuEmail author
Original Article
  • 7 Downloads

Abstract

Ralstonia pseudosolanacearum, the causative agent of bacterial wilt disease, is a significant pathogen of economically-important agricultural crops. FadP is a transcriptional regulator of fatty acid degradation; however, its function remains unknown in R. pseudosolanacearum. Herein, a R. pseudosolanacearum fadP deletion mutant (ΔfadP) was constructed to examine the role of FadP. Compared with wild-type strain Tm1401 and fadP complementation strain R-ΔfadP, ΔfadP showed reduced motility and cellulase and extracellular polysaccharide production, along with decreased resistance to tetracycline and virulence on tomato plants. Further, the mutant strain induced only a weak hypersensitive response in tobacco plants. To further understand the underlying virulence mechanisms, RNA-Seq technology was used to compare the transcriptomes of the wild-type and mutant strains. Analysis revealed 848 differentially-expressed genes, amongst which 505 were up-regulated and 343 were down-regulated in ΔfadP. Both flagella and chemotaxis genes were up-regulated, while genes coding for extracellular hydrolytic enzymes, cytochrome c oxidase, and carbon and nitrogen metabolism proteins were down-regulated. Two-step reverse-transcription polymerase chain reaction analysis targeting 13 genes confirmed that the expression of egl, rsl, pehC, pme, cspD3, narH, norB, ccoP, tek, and xpsR was significantly decreased. Our results suggest that FadP affects the virulence of R. pseudosolanacearum by altering nitrogen metabolism processes, reducing bacterial motility and extracellular polysaccharide and cellulase production, and affecting protein-carbohydrate interactions involved in host recognition, attachment, and invasion.

Keywords

Ralstonia pesudosolanacearum fadP Virulence Transcriptomics 

Notes

Acknowledgments

We thank Tamsin Sheen, PhD, from Liwen Bianji, Edanz Editing China (www.liwenbianji.cn/ac), for editing the English text of a draft of this manuscript.

Funding

This study was funded by by Special Fund for Agro-scientific Research in the Public Interest of China(201303015).

Compliance with ethical standards

Conflict of interest

All authors have received research grants from College of Agriculture, South China Agricultural University, Guangzhou, China; and Guangdong Province Key Laboratory of Microbial Signals and Disease Control, Guangzhou, China. All authors declare that they have no conflicts of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

42161_2019_343_Fig8_ESM.png (319 kb)
Fig S1

The construction and verification of the plasmid pK18-fadP-gm and the mutant ΔfadP. (A), PCR amplification of the up and down fragments of fadP gene and the Gm gene. M: DL 2000 marker; 1: the up fragments of fadP gene; 2: the down fragments of fadP gene; 3: gentamicin gene (gm). (B), The verification of pK18-fadP-gm using PCR amplification and double enzymes restriction. M: DL2000 marker; 1: PCR amplification of pK18-fadP-gm using primers pk-18-F/R; 2: double enzymes restriction of pK18-fadP-gm by restriction enzyme EcoR I and Hind III. (C), PCR amplification of the mutant ΔfadP using primers fadP-CX-F/fadP-CX-R. M: DL2000 marker; 1: ddH2O; 2:Tm1401; 3:ΔfadP. (PNG 318 kb)

42161_2019_343_MOESM1_ESM.tif (99 kb)
High Resolution (TIF 98 kb)
42161_2019_343_Fig9_ESM.png (245 kb)
Fig S2

Construct and verify the complement strain of the mutant. (A) Amplification of fadP gene complement fragment. M: DL2000 marker, HB: fadP gene complement fragment. (B) Verify among Tm1401, ΔfadP and R-ΔfadP using primers fadP-HB-F/fadP-HB-R. M: DL2000; 1: Tm1401; 2: ΔfadP; 3: R-ΔfadP; 4: ddH2O. (PNG 245 kb)

42161_2019_343_MOESM2_ESM.tif (66 kb)
High Resolution (TIF 65 kb)

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Copyright information

© Società Italiana di Patologia Vegetale (S.I.Pa.V.) 2019

Authors and Affiliations

  1. 1.College of AgricultureSouth China Agricultural UniversityGuangzhouChina
  2. 2.Guangdong Province Key Laboratory of Microbial Signals and Disease ControlGuangzhouChina

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