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A Purification and In Vitro Activity Assay for a pppGpp Synthetase from Clostridium difficile
Published on: November 3, 2018
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(p)ppGpp synthetases regulate the pathogenesis of zoonotic Streptococcus suis
Jiawen Zhu1, Tengfei Zhang2, Zhipeng Su1
1State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, China.
Microbiological Research
|August 16, 2016
Summary
The stringent response, regulated by (p)ppGpp synthetases (RelA and RelQ), is crucial for bacterial adaptation. Disrupting these enzymes in Streptococcus suis significantly reduces its virulence and ability to cause infection.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- The stringent response, mediated by guanosine tetraphosphate (ppGpp) and guanosine pentaphosphate (pppGpp), is a key bacterial adaptation mechanism linked to virulence.
- Streptococcus suis, a zoonotic pathogen, possesses two enzymes responsible for (p)ppGpp synthesis: RelA and RelQ.
Purpose of the Study:
- To investigate the role of (p)ppGpp synthetases (RelA and RelQ) in regulating the pathogenesis of Streptococcus suis.
- To compare the phenotypic characteristics of a double mutant lacking both synthetases with the wild-type strain.
Main Methods:
- Construction and phenotypic analysis of a ΔrelAΔrelQ double mutant in Streptococcus suis.
- Microscopy (light and electron) to assess bacterial morphology.
- In vitro assays for adhesion, invasion, resistance to blood killing, and phagocytosis.
- In vivo mouse infection models to evaluate virulence and clearance.
- Quantitative reverse transcription PCR (qRT-PCR) to analyze virulence gene expression.
Main Results:
- The ΔrelAΔrelQ mutant exhibited increased chain length compared to the parental strain.
- Disruption of RelA and RelQ significantly decreased bacterial adhesion and invasion of HEp-2 cells.
- The mutant strain showed increased sensitivity to host immune defenses, including blood killing and phagocytosis.
- In vivo, the mutant strain was attenuated, demonstrating reduced virulence and easier clearance from infected mice.
- Expression of virulence-associated genes was downregulated in the mutant strain.
Conclusions:
- The (p)ppGpp synthetases, RelA and RelQ, play a critical role in the pathogenesis of Streptococcus suis.
- (p)ppGpp signaling is essential for regulating bacterial morphology, host cell interaction, and resistance to host immunity.
- Targeting (p)ppGpp synthesis represents a potential strategy for controlling Streptococcus suis infections.
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