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Comparative analysis of Salmonella susceptibility and tolerance to the biocide chlorhexidine identifies a complex cellular defense network.

Condell O, Power KA, Händler K, Finn S, Sheridan A, Sergeant K, Renaut J, Burgess CM, Hinton JC, Nally JE, Fanning S - Front Microbiol (2014)

Bottom Line: Typhimurium compared with its isogenic sensitive progenitor.The defense network involved multiple cell targets including those associated with the synthesis and modification of the cell wall, the SOS response, virulence, and a shift in cellular metabolism toward anoxic pathways, some of which were regulated by CreB and Fur.In addition, results indicated that chlorhexidine tolerance was associated with more extensive modifications of the same cellular processes involved in this proposed network, as well as a divergent defense response involving the up-regulation of additional targets such as the flagellar apparatus and an altered cellular phosphate metabolism.

View Article: PubMed Central - PubMed

Affiliation: UCD Centre for Food Safety, School of Public Health, Physiotherapy and Population Science, University College Dublin Belfield, Dublin, Ireland ; European Program for Public Health Microbiology Training, European Centre for Disease Prevention and Control Stockholm, Sweden.

ABSTRACT
Chlorhexidine is one of the most widely used biocides in health and agricultural settings as well as in the modern food industry. It is a cationic biocide of the biguanide class. Details of its mechanism of action are largely unknown. The frequent use of chlorhexidine has been questioned recently, amidst concerns that an overuse of this compound may select for bacteria displaying an altered susceptibility to antimicrobials, including clinically important anti-bacterial agents. We generated a Salmonella enterica serovar Typhimurium isolate (ST24(CHX)) that exhibited a high-level tolerant phenotype to chlorhexidine, following several rounds of in vitro selection, using sub-lethal concentrations of the biocide. This mutant showed altered suceptibility to a panel of clinically important antimicrobial compounds. Here we describe a genomic, transcriptomic, proteomic, and phenotypic analysis of the chlorhexidine tolerant S. Typhimurium compared with its isogenic sensitive progenitor. Results from this study describe a chlorhexidine defense network that functions in both the reference chlorhexidine sensitive isolate and the tolerant mutant. The defense network involved multiple cell targets including those associated with the synthesis and modification of the cell wall, the SOS response, virulence, and a shift in cellular metabolism toward anoxic pathways, some of which were regulated by CreB and Fur. In addition, results indicated that chlorhexidine tolerance was associated with more extensive modifications of the same cellular processes involved in this proposed network, as well as a divergent defense response involving the up-regulation of additional targets such as the flagellar apparatus and an altered cellular phosphate metabolism. These data show that sub-lethal concentrations of chlorhexidine induce distinct changes in exposed Salmonella, and our findings provide insights into the mechanisms of action and tolerance to this biocidal agent.

No MeSH data available.


Related in: MedlinePlus

qRT-PCR confirmation of microarray data from (A) the reference ST24WT chlorhexidine treated, relative to the same isolate in the absence of chlorhexidine exposure and (B) the mutant isolate, ST24CHX, relative to the reference ST24WT. Details of protein results are provided in Table S4, with qRT-PCR and transcriptomic results shown in Table S3.
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Figure 4: qRT-PCR confirmation of microarray data from (A) the reference ST24WT chlorhexidine treated, relative to the same isolate in the absence of chlorhexidine exposure and (B) the mutant isolate, ST24CHX, relative to the reference ST24WT. Details of protein results are provided in Table S4, with qRT-PCR and transcriptomic results shown in Table S3.

Mentions: A total of 247 genes were significantly differentially expressed (2-fold-change, p < 0.05), following sub-lethal exposure to chlorhexidine (Table S1); 81 of these genes were up-regulated whilst 166 were down-regulated (Figure 2). Differentially expressed genes of known function were assigned to functional categories (Figure 3 and Table S1). Transcriptomic data were subsequently validated using qRT-PCR for 5 selected genes. Similar fold-change values were obtained from both independent analyses, for all five genes (Figure 4A, Table S3). Results from transcriptomic profiling revealed that the functional categories containing the largest number of up-regulated genes were involved in general cell metabolism; amino acid transport/metabolism, cofactor metabolism and carbohydrate metabolism. Conversely, the functional groups containing the largest number of down-regulated genes included virulence, transcription, translation and ribosomal structure, along with phage-associated genes. For each functional category, the numbers of differentially expressed genes are shown as a percentage of the total number of genes in that category (as determined by KEGG, Figure 3). Corresponding gene lists are provided in Table S1.


Comparative analysis of Salmonella susceptibility and tolerance to the biocide chlorhexidine identifies a complex cellular defense network.

Condell O, Power KA, Händler K, Finn S, Sheridan A, Sergeant K, Renaut J, Burgess CM, Hinton JC, Nally JE, Fanning S - Front Microbiol (2014)

qRT-PCR confirmation of microarray data from (A) the reference ST24WT chlorhexidine treated, relative to the same isolate in the absence of chlorhexidine exposure and (B) the mutant isolate, ST24CHX, relative to the reference ST24WT. Details of protein results are provided in Table S4, with qRT-PCR and transcriptomic results shown in Table S3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4117984&req=5

Figure 4: qRT-PCR confirmation of microarray data from (A) the reference ST24WT chlorhexidine treated, relative to the same isolate in the absence of chlorhexidine exposure and (B) the mutant isolate, ST24CHX, relative to the reference ST24WT. Details of protein results are provided in Table S4, with qRT-PCR and transcriptomic results shown in Table S3.
Mentions: A total of 247 genes were significantly differentially expressed (2-fold-change, p < 0.05), following sub-lethal exposure to chlorhexidine (Table S1); 81 of these genes were up-regulated whilst 166 were down-regulated (Figure 2). Differentially expressed genes of known function were assigned to functional categories (Figure 3 and Table S1). Transcriptomic data were subsequently validated using qRT-PCR for 5 selected genes. Similar fold-change values were obtained from both independent analyses, for all five genes (Figure 4A, Table S3). Results from transcriptomic profiling revealed that the functional categories containing the largest number of up-regulated genes were involved in general cell metabolism; amino acid transport/metabolism, cofactor metabolism and carbohydrate metabolism. Conversely, the functional groups containing the largest number of down-regulated genes included virulence, transcription, translation and ribosomal structure, along with phage-associated genes. For each functional category, the numbers of differentially expressed genes are shown as a percentage of the total number of genes in that category (as determined by KEGG, Figure 3). Corresponding gene lists are provided in Table S1.

Bottom Line: Typhimurium compared with its isogenic sensitive progenitor.The defense network involved multiple cell targets including those associated with the synthesis and modification of the cell wall, the SOS response, virulence, and a shift in cellular metabolism toward anoxic pathways, some of which were regulated by CreB and Fur.In addition, results indicated that chlorhexidine tolerance was associated with more extensive modifications of the same cellular processes involved in this proposed network, as well as a divergent defense response involving the up-regulation of additional targets such as the flagellar apparatus and an altered cellular phosphate metabolism.

View Article: PubMed Central - PubMed

Affiliation: UCD Centre for Food Safety, School of Public Health, Physiotherapy and Population Science, University College Dublin Belfield, Dublin, Ireland ; European Program for Public Health Microbiology Training, European Centre for Disease Prevention and Control Stockholm, Sweden.

ABSTRACT
Chlorhexidine is one of the most widely used biocides in health and agricultural settings as well as in the modern food industry. It is a cationic biocide of the biguanide class. Details of its mechanism of action are largely unknown. The frequent use of chlorhexidine has been questioned recently, amidst concerns that an overuse of this compound may select for bacteria displaying an altered susceptibility to antimicrobials, including clinically important anti-bacterial agents. We generated a Salmonella enterica serovar Typhimurium isolate (ST24(CHX)) that exhibited a high-level tolerant phenotype to chlorhexidine, following several rounds of in vitro selection, using sub-lethal concentrations of the biocide. This mutant showed altered suceptibility to a panel of clinically important antimicrobial compounds. Here we describe a genomic, transcriptomic, proteomic, and phenotypic analysis of the chlorhexidine tolerant S. Typhimurium compared with its isogenic sensitive progenitor. Results from this study describe a chlorhexidine defense network that functions in both the reference chlorhexidine sensitive isolate and the tolerant mutant. The defense network involved multiple cell targets including those associated with the synthesis and modification of the cell wall, the SOS response, virulence, and a shift in cellular metabolism toward anoxic pathways, some of which were regulated by CreB and Fur. In addition, results indicated that chlorhexidine tolerance was associated with more extensive modifications of the same cellular processes involved in this proposed network, as well as a divergent defense response involving the up-regulation of additional targets such as the flagellar apparatus and an altered cellular phosphate metabolism. These data show that sub-lethal concentrations of chlorhexidine induce distinct changes in exposed Salmonella, and our findings provide insights into the mechanisms of action and tolerance to this biocidal agent.

No MeSH data available.


Related in: MedlinePlus