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Response to Gaseous NO2 Air Pollutant of P. fluorescens Airborne Strain MFAF76a and Clinical Strain MFN1032.

Kondakova T, Catovic C, Barreau M, Nusser M, Brenner-Weiss G, Chevalier S, Dionnet F, Orange N, Poc CD - Front Microbiol (2016)

Bottom Line: 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.Finally, NO2-exposed P. fluorescens strains were found to be more resistant to ciprofloxacin and chloramphenicol.Taken together, our study provides evidences for the bacterial response to NO2 toxicity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology Signals and Microenvironment EA 4312, Normandy University, University of Rouen, SéSa, IRIBEvreux, France; Aerothermic and Internal Combustion Engine Technological Research CentreSaint Etienne du Rouvray, France.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus

Transcription of hmp is increased in response to NO2 exposure. The nucleotide sequences of the hmp-homolog gene in P. fluorescens strains were obtained using the non-annotated genome drafts of airborne P. fluorescens MFAF76a () and clinical MFN1032 (). The GenBank accession numbers of hmp nucleotide sequences are listed in Table S2. Quantification of mRNA level was assayed using qRT-PCR on RNAs extracted from NO and synthetic air-exposed P. fluorescens. The PCR reactions were performed in triplicate and the standard deviations were lower than 0.15 Ct. Statistical analysis used pairwise strain comparisons (t-test) p < 0.01 (**). Dotted line shows the gene expression in air-exposed control.
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Figure 8: Transcription of hmp is increased in response to NO2 exposure. The nucleotide sequences of the hmp-homolog gene in P. fluorescens strains were obtained using the non-annotated genome drafts of airborne P. fluorescens MFAF76a () and clinical MFN1032 (). The GenBank accession numbers of hmp nucleotide sequences are listed in Table S2. Quantification of mRNA level was assayed using qRT-PCR on RNAs extracted from NO and synthetic air-exposed P. fluorescens. The PCR reactions were performed in triplicate and the standard deviations were lower than 0.15 Ct. Statistical analysis used pairwise strain comparisons (t-test) p < 0.01 (**). Dotted line shows the gene expression in air-exposed control.

Mentions: Remarkably, we have shown herein a link between gaseous NO2 and soluble NO treatment. Indeed, NO is found to induce the expression of mexEF-oprN genes (Fetar et al., 2011) and modulates bacterial resistance to several antibiotics (Gusarov et al., 2009; McCollister et al., 2011; van Sorge et al., 2013). Since NO2 and NO are related chemical toxic compounds, and since NO detoxification pathways have been deeply investigated, the NO2 effects on several chosen target genes were tested. The most well-studied pathway for NO detoxification is based on flavohemoglobin (FlavoHb) (Hmp for E. coli and Fhp for P. aeruginosa), which acts as an NO dioxygenase to transform NO to NO (Figure 7A) (Corker and Poole, 2003; Arai et al., 2005). After exposure to 45 ppm of NO2, the hmp mRNA levels were increased almost 25- and 23-fold in MFAF76a and in MFN1032 (respectively KR818822 and KR818823 in Table S2 and Figure 8), indicating that NO2 induces hmp expression in both P. fluorescens and suggesting a possible involvement of Hmp in NO2 detoxification. The NO2 effect on the Hmp synthesis was observed in other studies, where, to activate the Hmp-dependent detoxification pathway, NO2 was proposed to be reduced to NO (Poole et al., 1996). In Pseudomonas spp., NO2 reduction can be performed by nitrite reductase (NIR) enzymes (Figures 7B,C), including the well-studied respiratory cytochrome cd1 nitrite reductase (Figure 7B) of the denitrification pathway (Arai et al., 2005; Shiro, 2012). According to the genome draft analysis


Response to Gaseous NO2 Air Pollutant of P. fluorescens Airborne Strain MFAF76a and Clinical Strain MFN1032.

Kondakova T, Catovic C, Barreau M, Nusser M, Brenner-Weiss G, Chevalier S, Dionnet F, Orange N, Poc CD - Front Microbiol (2016)

Transcription of hmp is increased in response to NO2 exposure. The nucleotide sequences of the hmp-homolog gene in P. fluorescens strains were obtained using the non-annotated genome drafts of airborne P. fluorescens MFAF76a () and clinical MFN1032 (). The GenBank accession numbers of hmp nucleotide sequences are listed in Table S2. Quantification of mRNA level was assayed using qRT-PCR on RNAs extracted from NO and synthetic air-exposed P. fluorescens. The PCR reactions were performed in triplicate and the standard deviations were lower than 0.15 Ct. Statistical analysis used pairwise strain comparisons (t-test) p < 0.01 (**). Dotted line shows the gene expression in air-exposed control.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4814523&req=5

Figure 8: Transcription of hmp is increased in response to NO2 exposure. The nucleotide sequences of the hmp-homolog gene in P. fluorescens strains were obtained using the non-annotated genome drafts of airborne P. fluorescens MFAF76a () and clinical MFN1032 (). The GenBank accession numbers of hmp nucleotide sequences are listed in Table S2. Quantification of mRNA level was assayed using qRT-PCR on RNAs extracted from NO and synthetic air-exposed P. fluorescens. The PCR reactions were performed in triplicate and the standard deviations were lower than 0.15 Ct. Statistical analysis used pairwise strain comparisons (t-test) p < 0.01 (**). Dotted line shows the gene expression in air-exposed control.
Mentions: Remarkably, we have shown herein a link between gaseous NO2 and soluble NO treatment. Indeed, NO is found to induce the expression of mexEF-oprN genes (Fetar et al., 2011) and modulates bacterial resistance to several antibiotics (Gusarov et al., 2009; McCollister et al., 2011; van Sorge et al., 2013). Since NO2 and NO are related chemical toxic compounds, and since NO detoxification pathways have been deeply investigated, the NO2 effects on several chosen target genes were tested. The most well-studied pathway for NO detoxification is based on flavohemoglobin (FlavoHb) (Hmp for E. coli and Fhp for P. aeruginosa), which acts as an NO dioxygenase to transform NO to NO (Figure 7A) (Corker and Poole, 2003; Arai et al., 2005). After exposure to 45 ppm of NO2, the hmp mRNA levels were increased almost 25- and 23-fold in MFAF76a and in MFN1032 (respectively KR818822 and KR818823 in Table S2 and Figure 8), indicating that NO2 induces hmp expression in both P. fluorescens and suggesting a possible involvement of Hmp in NO2 detoxification. The NO2 effect on the Hmp synthesis was observed in other studies, where, to activate the Hmp-dependent detoxification pathway, NO2 was proposed to be reduced to NO (Poole et al., 1996). In Pseudomonas spp., NO2 reduction can be performed by nitrite reductase (NIR) enzymes (Figures 7B,C), including the well-studied respiratory cytochrome cd1 nitrite reductase (Figure 7B) of the denitrification pathway (Arai et al., 2005; Shiro, 2012). According to the genome draft analysis

Bottom Line: 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.Finally, NO2-exposed P. fluorescens strains were found to be more resistant to ciprofloxacin and chloramphenicol.Taken together, our study provides evidences for the bacterial response to NO2 toxicity.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Microbiology Signals and Microenvironment EA 4312, Normandy University, University of Rouen, SéSa, IRIBEvreux, France; Aerothermic and Internal Combustion Engine Technological Research CentreSaint Etienne du Rouvray, France.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus