RovM and CsrA Negatively Regulate Urease Expression in Yersinia pseudotuberculosis

Stewart

Front. Cell. Infect. Microbiol.

et al. 2018

The formation of biofilms by Yersinia pseudotuberculosis (Yptb) and Y. pestis requires the hmsHFRS genes, which direct production of a polysaccharide extracellular matrix (Hms-ECM). Despite possessing identical hmsHFRS sequences, Yptb produces much less Hms-ECM than Y. pestis. The regulatory influences that control Yptb Hms-ECM production and biofilm formation are not fully understood. In this study, negative regulators of biofilm production in Yptb were identified. Inactivation of the BarA/UvrY two-component system or the CsrB regulatory RNA increased binding of Congo Red dye, which correlates with extracellular polysaccharide production. These mutants also produced biofilms that were substantially more cohesive than the wild type strain. Disruption of uvrY was not sufficient for Yptb to cause proventricular blockage during infection of Xenopsylla cheopis fleas. However, this strain was less acutely toxic toward fleas than wild type Yptb. Flow cytometry measurements of lectin binding indicated that Yptb BarA/UvrY/CsrB mutants may produce higher levels of other carbohydrates in addition to poly-GlcNAc Hms-ECM. In an effort to characterize the relevant downstream targets of the BarA/UvrY system, we conducted a proteomic analysis to identify proteins with lower abundance in the csrB::Tn5 mutant strain. Urease subunit proteins were less abundant and urease enzymatic activity was lower, which likely reduced toxicity toward fleas. Loss of CsrB impacted expression of several potential regulatory proteins that may influence biofilms, including the RcsB regulator. Overexpression of CsrB did not alter the Congo-red binding phenotype of an rcsB::Tn5 mutant, suggesting that the effect of CsrB on biofilms may require RcsB. These results underscore the regulatory and compositional differences between Yptb and Y. pestis biofilms. By activating CsrB expression, the Yptb BarA/UvrY two-component system has pleiotropic effects that impact biofilm production and stability.

Section: Resultssupporting

confidence: 93%

Yersinia pseudotuberculosis BarA-UvrY Two-Component Regulatory System Represses Biofilms via CsrB

Stewart

Front. Cell. Infect. Microbiol.

et al. 2018

“…Secondly, Gao et al ’s comparative transcriptomic analysis found that that rovM expression was not affected in an ompR ‐negative mutant (Gao et al , ). Thirdly, the urease locus in Yersinia pseudotuberculosis (the recent ancestor of Y. pestis ) is regulated by OmpR and the carbon storage regulator (Csr)‐RovM cascade because both OmpR and RovM bind to the urease locus promoter and not because OmpR regulates rovM (Achtman et al , ; Hu et al , ; Dai et al , ). Even though it has been suggested that RovM is involved in Y. pestis ’s metabolic adaptation to the flea gut (because it provides a fitness advantage), it has been clearly stated that RovM does not contribute to biofilm blockage or Y. pestis ’s growth or survival in the flea gut (Vadyvaloo et al , ).We therefore hypothesize that during flea infection, nutrient cues activate the Csr‐RovM regulatory cascade and the OmpR‐EnvZ regulatory system independently.…”

Section: Discussionmentioning

confidence: 99%

Nutrient depletion may trigger the Yersinia pestis OmpR‐EnvZ regulatory system to promote flea‐borne plague transmission

Bontemps-Gallo

Fernandez

Dewitte

et al. 2019

Molecular Microbiology

The flea's lumen gut is a poorly documented environment where the agent of flea-borne plague, Yersinia pestis, must replicate to produce a transmissible infection. Here, we report that both the acidic pH and osmolarity of the lumen's contents display simple harmonic oscillations with different periods. Since an acidic pH and osmolarity are two of three known stimuli of the OmpR-EnvZ two-component system in bacteria, we investigated the role and function of this Y. pestis system in fleas. By monitoring the in vivo expression pattern of three OmpR-EnvZregulated genes, we concluded that the flea gut environment triggers OmpR-EnvZ. This activation was not, however, correlated with changes in pH and osmolarity but matched the pattern of nutrient depletion (the third known stimulus for OmpR-EnvZ). Lastly, we found that the OmpR-EnvZ and the OmpF porin are needed to produce the biofilm that ultimately obstructs the flea's gut and thus hastens the flea-borne transmission of plague. Taken as a whole, our data suggest that the flea gut is a complex, fluctuating environment in which Y. pestis senses nutrient depletion via OmpR-EnvZ. Once activated, the latter triggers a molecular program (including at least OmpF) that produces the biofilm required for efficient plague transmission. ; Tel 33 320 871 193; Fax 33 320 871 178. § Bontemps-Gallo and Fernandez contributed equally to this work.

“…The chromosomal deletion mutants were constructed by sacB -based allelic exchange [ 24 ]. Two DNA fragments were amplified by PCR with C. sakazakii BAA-894 chromosomal DNA as a template with primer pairs UP-F/UP-R and DOWN-F/DOWN-R, respectively ( Table 2 ).…”

Section: Methodsmentioning

confidence: 99%

The Role of ptsH in Stress Adaptation and Virulence in Cronobacter sakazakii BAA-894

Sun

Yang

et al. 2022

Foods

Cronobacter sakazakii, an emerging foodborne pathogen that was isolated primarily from powdered infant formula, poses an important issue in food safety due to its high stress tolerance and pathogenicity. The Hpr (encoded by ptsH gene) has been shown to regulate carbon metabolism as well as stress response and virulence. However, the functional properties of ptsH in C. sakzakii have not been investigated. In this study, we clarified the role of ptsH in the C. sakzakii stress response and virulence, and explored its possible regulatory mechanism by RNA-seq. Compared with wild-type, the ΔptsH mutant showed a slower growth rate in the log phase but no difference in the stationary phase. Moreover, the resistance to heat stress (65 °C, 55 °C), simulated gastric fluid (pH = 2.5), biofilm formation and adhesion to HT-29 cells of ΔptsH mutant were significantly decreased, whereas the oxidative resistance (1, 5, 10 mM H2O2), osmotic resistance (10%, 15%, 20% NaCl), and superoxide dismutase activity were enhanced. Finally, RNA-seq analysis revealed the sulfur metabolism pathway is significantly upregulated in the ΔptsH mutant, but the bacterial secretion system pathway is dramatically downregulated. The qRT-PCR assay further demonstrated that the ΔptsH mutant has elevated levels of genes that are related to oxidative and osmotic stress (sodA, rpoS, cpxA/R, osmY). This study provides a great understanding of the role of ptsH in diverse stress responses and virulence in C. sakazakii, and it contributes to our understanding of the genetic determinant of stress resistance and pathogenicity of this important foodborne pathogen.