Summary
One key to the success of Pseudomonas spp. is their ability to reside in hostile environments. Pseudomonas spp. possess a cis–trans isomerase (Cti) an enzyme that converts the cis‐unsaturated fatty acids (FAs) of the membrane lipids to their trans‐isomers to rigidify the membrane and thereby resist stresses. Whereas the posttranslational Cti regulation has been previously reported, transcriptional cti regulation remains to be studied in more details. Here, we have studied cti transcriptional regulation in the solvent‐tolerant strain Pseudomonas putida F1. Two cti transcriptional start sites (cti‐279 and cti‐77) were identified with cti‐279 transcript being dominant. Expression of cti was found to increase with temperature increase, addition of the organic solvent, octanol and in the stationary growth phase. We found that cti expression was repressed by the cyclic‐AMP receptor protein (Crp) and repression required the cyclic‐AMP ligand of Crp. Production of trans‐unsaturated FAs was found to decrease after 24 h of growth. Although this decrease was accompanied by an increase in cyclopropane FA content, this was not at the expense of trans‐unsaturated FAs demonstrating the absence of competition between Cti and Cfa in FA modification.
Human exposure to nitrogen dioxide (NO2), an air pollutant of increasing interest in biology, results in several toxic effects to human health and also to the air microbiota. The aim of this study was to investigate the bacterial response to gaseous NO2. Two Pseudomonas fluorescens strains, namely the airborne strain MFAF76a and the clinical strain MFN1032 were exposed to 0.1, 5, or 45 ppm concentrations of NO2, and their effects on bacteria were evaluated in terms of motility, biofilm formation, antibiotic resistance, as well as expression of several chosen target genes. While 0.1 and 5 ppm of NO2did not lead to any detectable modification in the studied phenotypes of the two bacteria, several alterations were observed when the bacteria were exposed to 45 ppm of gaseous NO2. We thus chose to focus on this high concentration. NO2-exposed P. fluorescens strains showed reduced swimming motility, and decreased swarming in case of the strain MFN1032. Biofilm formed by NO2-treated airborne strain MFAF76a showed increased maximum thickness compared to non-treated cells, while NO2 had no apparent effect on the clinical MFN1032 biofilm structure. It is well known that biofilm and motility are inversely regulated by intracellular c-di-GMP level. The c-di-GMP level was however not affected in response to NO2 treatment. Finally, NO2-exposed P. fluorescens strains were found to be more resistant to ciprofloxacin and chloramphenicol. Accordingly, the resistance nodulation cell division (RND) MexEF-OprN efflux pump encoding genes were highly upregulated in the two P. fluorescens strains. Noticeably, similar phenotypes had been previously observed following a NO treatment. Interestingly, an hmp-homolog gene in P. fluorescens strains MFAF76a and MFN1032 encodes a NO dioxygenase that is involved in NO detoxification into nitrites. Its expression was upregulated in response to NO2, suggesting a possible common pathway between NO and NO2 detoxification. Taken together, our study provides evidences for the bacterial response to NO2 toxicity.
Nowadays air pollution is increasing due to anthropogenic activity. Among all air pollutants, nitrogen oxides (NOx) such as NO 2 are predominant. It is well known that those compounds exhibit direct toxic effects on human health. However, microorganisms are also exposed to them, but the effect of NOx on the virulence of air microbiota is still poorly understood. In this study, we evaluated the impact of NO 2 on the adaptability and virulence of an airborne strain of P. fluorescens, MFA76a, by exposition of this strain to 45 ppm of NO 2. The growth kinetics and cultivability were analysed. A decrease of cultivability coupled with an increase of the lag phase was observed suggesting a potential toxicity of NO 2. Since NOx particularly target lipids, the membrane permeability was assessed thanks to Live Dead tests and confocal microscopy. A significant alteration of membrane permeability was observed. Furthermore, more abundant bacterial aggregates were detected compared to the control. Thus, a lipidomic study was performed using MALDI-TOF MS Imaging coupled to HPTLC. Interestingly, bacteria exposed to NO 2 were lacking one putative glycerophospholipid molecule. In agreement with a previous study from Kondakova et al., these data demonstrate the adaptation potential of P. fluorescens MFAF76a to an air pollutant such as NO 2 .
Anthropogenic atmospheric pollution and immune response regularly expose bacteria to toxic nitrogen oxides such as NO• and NO2. These reactive molecules can damage a wide variety of biomolecules such as DNA, proteins and lipids. Several components of the bacterial envelope are susceptible to be damaged by reactive nitrogen species. Furthermore, the hydrophobic core of the membranes favors the reactivity of nitrogen oxides with other molecules, making membranes an important factor in the chemistry of nitrosative stress. Since bacteria are often exposed to endogenous or exogenous nitrogen oxides, they have acquired protection mechanisms against the deleterious effects of these molecules. By exposing bacteria to gaseous NO2, this work aims to analyze the physiological effects of NO2 on the cell envelope of the airborne bacterium Pseudomonas fluorescens MFAF76a and its potential adaptive responses. Electron microscopy showed that exposure to NO2 leads to morphological alterations of the cell envelope. Furthermore, the proteomic profiling data revealed that these cell envelope alterations might be partly explained by modifications of the synthesis pathways of multiple cell envelope components, such as peptidoglycan, lipid A, and phospholipids. Together these results provide important insights into the potential adaptive responses to NO2 exposure in P. fluorescens MFAF76a needing further investigations.
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