Analysis of the Streptococcus mutans proteome during acid and oxidative stress reveals modules of co-expression and an expanded role for the TreR transcriptional regulator

Streptococcus mutans promotes a tooth-damaging dysbiosis in the oral microbiota because it can form biofilms and survive acid stress better than most of its health-associated, ecological competitors. Many of these commensals produce hydrogen peroxide, therefore S. mutans must manage both oxidative stress and acid stress with coordinated and complex physiological responses. In this study, the proteome of S. mutans was examined during regulated growth in acid and oxidative stresses, as well as in deletion mutants with impaired oxidative stress phenotypes, Δnox and ΔtreR. 607 proteins exhibited significantly different abundance levels across the conditions tested, and correlation network analysis identified several modules of co-expressed proteins that were responsive to the deletion of nox and/or treR, as well as acid and oxidative stress. The data provided evidence explaining the ROS-sensitive and mutacin-deficient phenotypes exhibited by the ΔtreR strain. SMU.1069-1070, a poorly understood LytTR system, had elevated abundance in the ΔtreR strain. S. mutans LytTR systems regulate mutacin production and competence, which may explain how TreR affects mutacin production. Furthermore, the gene cluster that produces mutanobactin, important in ROS tolerance, displayed reduced abundance in the ΔtreR strain. The role of Nox as a keystone in the oxidative stress response was also emphasized. This dataset illustrates, at high resolution, how the paradigm cariogenic species activates the stress responses which enable it to cause disease, and it provides oral microbiota researchers with a resource to both reexamine previous hypotheses regarding proteins of interest and explore newly identified co-expression modules and pathways.

The critical role of MetR/MetB/MetC/MetX in cysteine and methionine metabolism, fungal development and virulence of Alternaria alternata

Methionine is a unique sulfur-containing amino acid, which plays an important role in biological protein synthesis and various cellular processes. Here, we characterized the biological functions of AaMetB, AaMetC, and AaMetX in the tangerine pathotype of Alternaria alternata. Morphological analysis showed that the mutants lacking AaMetB, AaMetC, or AaMetX resulted in less aerial hypha and fewer conidia in artificial media. Pathogenicity analysis showed that AaMetB, AaMetC, and AaMetX are required for full virulence. The defects in vegetative growth, conidiation and virulence of ΔMetB, ΔMetC, and ΔMetX can be restored by exogenous methionine and homocysteine, indicating that AaMetB, AaMetC, and AaMetX are required for methionine biosynthesis. However, exogenous cysteine only restored the growth and virulence defects of ΔMetR but not ΔMetB/C/X, suggesting that AaMetR is essential for cysteine biosynthesis. Oxidant sensitivity assay showed that only ΔMetR is sensitive to H2O2 and many ROS-generating compounds, indicating that AaMetR is essential for oxidative tolerance. Interestingly, fungicides indoor bioassays showed that only the ΔMetR mutants are susceptive to chlorothalonil, a fungicide that could bind to the cysteine of glyceraldehyde-3-phosphate dehydrogenase. Comparative transcriptome analysis showed that the inactivation of MetB, MetC, MetX, or MetR significantly affected the expression of methionine metabolism-related genes. Moreover, the inactivation of AaMetR significantly affected the expression of many genes related to glutathione metabolism, which is essential for ROS tolerance. Taken together, our study provides genetic evidence to define the critical roles of AaMetB, AaMetC, AaMetX, and AaMetR in cysteine and methionine metabolism, fungal development and virulence of Alternaria alternata. IMPORTANCE The transcription factor METR regulating methionine metabolism is essential for reactive oxygen species (ROS) tolerance and virulence in many phytopathogenic fungi. However, the underlying regulatory mechanism of METR involved in this process is still unclear. In the present study, we generated AaMetB, AaMetC and AaMetX deletion mutants and compared these mutants with AaMetR disrupted mutants. Interestingly, we found that AaMetB, AaMetC and AaMetX are required for vegetative growth, conidiation, and pathogenicity in Alternaria alternata, but not for ROS tolerance and cysteine metabolism. Furthermore, we found that METR is involved in the biosynthesis of cysteine, which is an essential substrate for the biosynthesis of methionine and glutathione. This study emphasizes the critical roles of MetR, MetB, MetC, MetX in the regulation of cysteine and methionine metabolism, as well as the cross-link with glutathione-mediated ROS tolerance in phytopathogenic fungi, which provides a foundation for future investigations.

Presence of a resident species aids invader evolution

Phytoplankton populations are intrinsically large and genetically variable, and interactions between species in these populations shape their physiological and evolutionary responses. Yet, evolutionary responses of microbial organisms in novel environments are investigated almost exclusively through the lens of species colonising new environments on their own, and invasion studies are often of short duration. Although exceptions exist, neither type of study usually measures ecologically relevant traits beyond growth rates. Here, we experimentally evolved populations of fresh- and seawater phytoplankton as monocultures (the green algae Chlamydomonas moewusii and Ostreococcus tauri, each colonising a novel, unoccupied salinity) and co-cultures (invading a novel salinity occupied by a resident species) for 200 generations. Colonisers and invaders differed in extinction risks, phenotypes (e.g. size, primary production rates) and strength of local adaptation: invaders had systematically lower extinction rates and broader salinity and temperature preferences than colonisers – regardless of the environment that the invader originated from. We emphasise that the presence of a locally adapted species has the potential to alter the invading species’ eco-evolutionary trajectories in a replicable way across environments of differing quality, and that the evolution of small cell size and high ROS tolerance may explain high invader fitness. To predict phytoplankton responses in a changing world, such interspecific relationships need to be accounted for.

Methyl viologen can affect mitochondrial function in Arabidopsis

Reactive oxygen species (ROS) are key signalling intermediates in plant metabolism, defence, and stress adaptation. The chloroplast and mitochondria are centres of metabolic control and ROS production, which coordinate stress responses in other cell compartments. The herbicide and experimental tool, methyl viologen (MV) induces ROS generation in the chloroplast under illumination, but is also toxic in non-photosynthetic organisms. We used MV to probe plant ROS signalling in compartments other than the chloroplast. Taking a genetic approach in Arabidopsis thaliana, we used natural variation, QTL mapping, and mutant studies with MV in the light, but also under dark conditions, when the chloroplast electron transport is inactive. These studies revealed a light-independent MV-induced ROS-signalling pathway, suggesting mitochondrial involvement. Mitochondrial Mn SUPEROXIDE DISMUTASE was required for ROS-tolerance and the effect of MV was enhanced by exogenous sugar, providing further evidence for the role of mitochondria. Mutant and hormone feeding assays revealed roles for stress hormones in organellar ROS-responses. The radical-induced cell death1 mutant, which is tolerant to MV-induced ROS and exhibits altered mitochondrial signalling, was used to probe interactions between organelles. Our studies implicate mitochondria in the response to ROS induced by MV.

Mechanisms of Bacterial Tolerance and Persistence in the Gastrointestinal and Respiratory Environments

SUMMARYPathogens that infect the gastrointestinal and respiratory tracts are subjected to intense pressure due to the environmental conditions of the surroundings. This pressure has led to the development of mechanisms of bacterial tolerance or persistence which enable microorganisms to survive in these locations. In this review, we analyze the general stress response (RpoS mediated), reactive oxygen species (ROS) tolerance, energy metabolism, drug efflux pumps, SOS response, quorum sensing (QS) bacterial communication, (p)ppGpp signaling, and toxin-antitoxin (TA) systems of pathogens, such asEscherichia coli,Salmonellaspp.,Vibriospp.,Helicobacterspp.,Campylobacter jejuni,Enterococcusspp.,Shigellaspp.,Yersiniaspp., andClostridium difficile, all of which inhabit the gastrointestinal tract. The following respiratory tract pathogens are also considered:Staphylococcus aureus,Pseudomonas aeruginosa,Acinetobacter baumannii,Burkholderia cenocepacia, andMycobacterium tuberculosis. Knowledge of the molecular mechanisms regulating the bacterial tolerance and persistence phenotypes is essential in the fight against multiresistant pathogens, as it will enable the identification of new targets for developing innovative anti-infective treatments.