TL;DR
Findings suggest that AhpC1 functions alongside catalase systems in the oxidative stress defense network of V. parahaemolyticus.
Vibrio anguillarum is a major bacterial pathogen in marine aquaculture, which causes significant economic losses. The histone-like protein HU, a conserved nucleoid-associated protein, plays crucial roles in chromosome organization, global gene regulation, and stress adaptation. In many bacteria, HU exists as both a homodimer and a heterodimer composed of two subunits, HUα and HUβ, which are encoded by hupA and hupB, respectively. However, the functional roles and divergence of HU subunits in V. anguillarum remain poorly understood. In this study, we successfully constructed the single-gene deletion mutants (ΔhupA and ΔhupB) and the hupA/hupB double-gene deletion mutant (ΔHU). Compared with wild-type (WT) strain, ΔhupA and ΔHU exhibited impaired growth, whereas ΔhupB showed enhanced growth. Deletion of hupB completely abolished biofilm formation and autoaggregation, whereas the ΔHU mutant partially restored biofilm formation. Interestingly, ΔHU and ΔhupA displayed enhanced autoaggregation. All mutants displayed significantly reduced motility. Notably, extracellular DNase production was markedly decreased in ΔHU. In addition, ΔHU showed the highest susceptibility to DNA-mediated killing, while ΔhupB displayed enhanced resistance to this stress. Moreover, the virulence of ΔhupA and ΔHU was significantly attenuated, with a 10.00-fold and a 12.60-fold increase in LD50, respectively, compared to the WT strain, while ΔhupB showed no significant change in virulence. Collectively, our findings reveal the functional divergence between HUα and HUβ in regulating growth, motility, biofilm formation, quorum-sensing, DNase production, and DNA resistance in V. anguillarum, and provide a theoretical basis and novel insights for the development of live attenuated vaccines against vibriosis in aquaculture.
ABSTRACT LiaS is a histidine kinase receptor in two-component systems that senses environmental stress signals. This study investigated the roles of LiaS in Listeria monocytogenes stress resistance and virulence. Phenotypic assays showed that deletion of liaS (ΔliaS) resulted in significantly impaired growth under acidic, alkaline, and osmotic stresses, as well as reduced survival under strong acidic conditions. These defects were partially restored in the complemented strain (CΔliaS). The growth defects were also observed in the ΔliaS under Cu2+ or H2O2 exposure. Furthermore, the ΔliaS strain exhibited impaired invasion and intracellular migration in host cells, along with attenuated colonization in the liver and spleen at 24 h post-infection. Bacterial loads in the spleen of infected mice remained lower at 48 h. The mortality was also delayed in ΔliaS‑infected mice. Transcriptional analysis revealed that compared to neutral conditions, osmotic stress-related genes (except opucB and gbuC) were significantly upregulated in the ΔliaS strain under acidic conditions, indicating that LiaS modulates acid tolerance through transcriptional regulation. Collectively, LiaS is essential for the adaptation of Listeria monocytogenes to acidic, osmotic, and oxidative stresses. Its absence attenuates host cell invasion, intracellular motility, and organ-specific colonization, highlighting its dual role in environmental resilience and pathogenicity. These findings provide a theoretical basis for understanding host–pathogen interactions and offer new strategies against antimicrobial-resistant pathogens.
ABSTRACT ClpX functions as a component of the ClpXP protease, a conserved intracellular protease that regulates protein turnover, stress responses, and virulence in multiple bacterial species. Our lab has established that clpX is necessary for resistance to cell envelope targeting antibiotics, such as penicillin and daptomycin in Bacillus anthracis Sterne. Previous microarray data identified the msrA/B gene encoding a bifunctional methionine sulfoxide reductase as upregulated in the ΔclpX mutant. Methionine sulfoxide reductases (Msr) repair oxidatively damaged proteins by reducing methionine sulfoxide residues back to methionine. While Msr enzymes are primarily associated with oxidative stress, cell wall antibiotics induce expression of msrA1 and msrB in S. aureus. Here, we investigated the role of MsrA/B in oxidative and cell envelope stress. Our results show that although hydrogen peroxide and paraquat induce msrA/B expression, the ΔmsrA/B strain was not susceptible to either oxidant, whereas the ΔclpX strain was sensitive to both. We also found that loss of msrA/B conferred penicillin‐specific sensitivity, but, unlike ΔclpX, increased sensitivity was not seen with other cell wall or cell membrane targeting antibiotics. Inactivation of the catalytic cysteine of either Msr domain of MsrA/B failed to complement, suggesting that the reducing activity of MsrA/B is required for penicillin resistance. These findings indicate that while MsrA/B contributes to penicillin resistance, other proteins in the ClpXP modulon must also play a role in oxidative and cell envelope stress.
Campylobacter jejuni is the leading cause of bacterial foodborne diarrheal disease worldwide. Despite its microaerophilic nature, C. jejuni is ubiquitous in aerobic environments and must possess specific adaptation mechanisms against oxidative stress. Here, we identified a novel role for FlhF, a GTPase essential for proper flagellar assembly, in promoting resistance to hydrogen peroxide (H2O2). Comparative transcriptomic analysis under H2O2 stress revealed that deletion of flhF leads to significant downregulation of oxidative stress-related genes. FlhF directly interacts with TonB2, an iron transport-associated protein, via its B and N domains. Codeletion of flhF and tonB2 leads to increased sensitivity to H2O2, suggesting a synergistic interaction. Moreover, the FlhF-TonB2 interaction promotes H2O2 detoxification, potentially by modulating intracellular iron homeostasis and influencing redox processes. Together, these findings reveal a novel function of FlhF in the oxidative stress response of C. jejuni, offering new insights into flagella-associated defense mechanisms in this pathogen.
ABSTRACT This study used a functional genomics approach to explore the role of a xenobiotic response element (XRE)-type regulator (SMU.405c) in Streptococcus mutans physiology, including the expression of biofilm regulatory protein BrpA. Results showed that deletional mutation of xre significantly reduced the ability of the deficient mutant to grow in the presence of methyl viologen, a commonly used oxidative stressor (P < 0.001). When challenged in a hydrogen peroxide killing assay, the survival rate of the ∆xre mutant was >2-log less than the parent strain after 60 min (P < 0.001). Luciferase reporter fusion assays showed that xre deficiency had no significant effect on luciferase expression when it was under the control of the intact brpA promoter, but the reporter activity increased by >6-fold (P < 0.001) when the reporter gene was fused to a brpA promoter derivative with deletion of a putative XRE-binding box. Electrophoretic mobility shift assay (EMSA) showed that recombinant XRE interacted with the brpA promoter, resulting in an electrophoretic shift of the promoter probes. In vitro transcription assay also showed that inclusion of XRE caused transcription to fall off, significantly reducing full-length brpA transcripts. RNA-seq analysis revealed that deficiency of XRE led to altered expression of >102 genes by >2-fold (P < 0.05), including 28 with increased expression, and 74 with decreased expression. Among the down-regulated were genes for DNA repair and oxidative stress tolerance response. These results suggest that XRE (SMU.405c) in S. mutans plays an important role in brpA expression and oxidative stress tolerance response. IMPORTANCE Streptococcus mutans, a keystone pathogen in human dental caries, primarily lives in the highly diverse microbiota on tooth surfaces, where the conditions are often harsh and fluctuate frequently. Locus SMU.405c was annotated to encode a xenobiotic response element (XRE)-like transcriptional regulator, but no information is available concerning the role of this protein in S. mutans pathophysiology. This study used a functional genomics approach along with molecular and transcriptomic analysis to characterize a deletional xre mutant, and the results showed that xre deficiency in S. mutans resulted in weakened oxidative stress tolerance response and alterations in transcription of >102 genes, including those known to play an important role in cell envelope biogenesis and stress tolerance response. Reporter fusion assay, electrophoretic mobility shift assay (EMSA), and in vitro transcription further demonstrated that the XRE-like regulator encoded by SMU.405c is a repressor of brpA expression and plays an important role in oxidative stress tolerance response. Streptococcus mutans, a keystone pathogen in human dental caries, primarily lives in the highly diverse microbiota on tooth surfaces, where the conditions are often harsh and fluctuate frequently. Locus SMU.405c was annotated to encode a xenobiotic response element (XRE)-like transcriptional regulator, but no information is available concerning the role of this protein in S. mutans pathophysiology. This study used a functional genomics approach along with molecular and transcriptomic analysis to characterize a deletional xre mutant, and the results showed that xre deficiency in S. mutans resulted in weakened oxidative stress tolerance response and alterations in transcription of >102 genes, including those known to play an important role in cell envelope biogenesis and stress tolerance response. Reporter fusion assay, electrophoretic mobility shift assay (EMSA), and in vitro transcription further demonstrated that the XRE-like regulator encoded by SMU.405c is a repressor of brpA expression and plays an important role in oxidative stress tolerance response.
Results demonstrate that PlsC is essential for maintaining membrane homeostasis and stress resistance in Brucella, which in turn supports its survival within professional phagocytes and full virulence in vivo and suggests a critical link between phospholipid metabolism and Brucella pathogenicity.
