The biocide triclosan triggers multiple regulatory systems in Staphylococcus aureus to induce antibiotic tolerance

ABSTRACTAn experimental model that mimicked the structure and characteristics ofin vivobiofilm infections, such as those occurring in the lung or in dermal wounds where no biomaterial surface is present, was developed. In these infections, microbial biofilm forms as cell aggregates interspersed in a layer of mucus or host matrix material. This structure was modeled by filling glass capillary tubes with an agarose gel that had been seeded withStaphylococcus aureusbacteria and then incubating the gel biofilm in medium for up to 30 h. Confocal microscopy showed that the bacteria formed in discrete pockets distributed throughout the gel matrix. These aggregates enlarged over time and also developed a size gradient, with the clusters being larger near the nutrient- and oxygen-supplied interface and smaller at greater depths. Bacteria entrapped in gels for 24 h grew slowly (specific growth rate, 0.06 h−1) and were much less susceptible to oxacillin, minocycline, or ciprofloxacin than planktonic cells. Microelectrode measurements showed that the oxygen concentration decreased with depth into the gel biofilm, falling to values less than 3% of air saturation at depths of 500 μm. An anaerobiosis-responsive green fluorescent protein reporter gene for lactate dehydrogenase was induced in the region of the gel where the measured oxygen concentrations were low, confirming biologically relevant hypoxia. These results show that the gel biofilm model captures key features of biofilm infection in mucus or compromised tissue: formation of dense, distinct aggregates, reduced specific growth rates, local hypoxia, and antibiotic tolerance.

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Characterizing the Effect of the Staphylococcus aureus Virulence Factor Regulator, SarA, on Log-Phase mRNA Half-Lives

Journal of Bacteriology ◽

10.1128/jb.188.7.2593-2603.2006 ◽

2006 ◽

Vol 188 (7) ◽

pp. 2593-2603 ◽

Cited By ~ 73

Author(s):

Corbette Roberts ◽

Kelsi L. Anderson ◽

Ellen Murphy ◽

Steven J. Projan ◽

William Mounts ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Virulence Factor ◽

Mrna Stability ◽

Transcription Initiation ◽

Target Gene ◽

Dependent Manner ◽

Reading Frame ◽

Rna Molecules ◽

Regulatory Systems ◽

Small Stable Rna

ABSTRACT Bacterial pathogens regulate virulence factor expression at both the level of transcription initiation and mRNA processing/turnover. Within Staphylococcus aureus, virulence factor transcript synthesis is regulated by a number of two-component regulatory systems, the DNA binding protein SarA, and the SarA family of homologues. However, little is known about the factors that modulate mRNA stability or influence transcript degradation within the organism. As our entree to characterizing these processes, S. aureus GeneChips were used to simultaneously determine the mRNA half-lives of all transcripts produced during log-phase growth. It was found that the majority of log-phase transcripts (90%) have a short half-life (<5 min), whereas others are more stable, suggesting that cis- and/or trans-acting factors influence S. aureus mRNA stability. In support of this, it was found that two virulence factor transcripts, cna and spa, were stabilized in a sarA-dependent manner. These results were validated by complementation and real-time PCR and suggest that SarA may regulate target gene expression in a previously unrecognized manner by posttranscriptionally modulating mRNA turnover. Additionally, it was found that S. aureus produces a set of stable RNA molecules with no predicted open reading frame. Based on the importance of the S. aureus agr RNA molecule, RNAIII, and small stable RNA molecules within other pathogens, it is possible that these RNA molecules influence biological processes within the organism.

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Interruption of the tricarboxylic acid cycle in Staphylococcus aureus leads to increased tolerance to innate immunity

AIMS Microbiology ◽

10.3934/microbiol.2021031 ◽

2021 ◽

Vol 7 (4) ◽

pp. 513-527

Author(s):

Alexis M. Hobbs ◽

◽

Kennedy E. Kluthe ◽

Kimberly A. Carlson ◽

Austin S. Nuxoll

Keyword(s):

Staphylococcus Aureus ◽

Immune System ◽

Innate Immune System ◽

Tricarboxylic Acid Cycle ◽

Fold Increase ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Innate Immune ◽

Antibiotic Tolerance ◽

Males And Females

<abstract> <p><italic>Staphylococcus aureus</italic> is widely known for its resistance and virulence causing public health concerns. However, antibiotic tolerance is also a contributor to chronic and relapsing infections. Previously, it has been demonstrated that persister formation is dependent on reduced tricarboxylic acid (TCA) cycle activity. Persisters have been extensively examined in terms of antibiotic tolerance but tolerance to antimicrobial peptides (AMPs) remains largely unexplored. AMPs are a key component of both the human and <italic>Drosophila</italic> innate immune response. TCA cycle mutants were tested to determine both antibiotic and AMP tolerance. Challenging with multiple classes of antibiotics led to increased persister formation (100- to 1,000-fold). Similarly, TCA mutants exhibited AMP tolerance with a 100- to 1,000-fold increase in persister formation when challenged with LL-37 or human β-defensin 3 (hβD3). The ability of TCA cycle mutants to tolerate the innate immune system was further examined with a <italic>D. melanogaster</italic> model. Both males and females infected with TCA cycle mutants exhibited increased mortality and had higher bacterial burdens (1.5 log) during the course of the infection. These results suggest increasing the percentage of persister cells leads to increased tolerance to components of the innate immune system.</p> </abstract>

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Faculty Opinions recommendation of Reversible antibiotic tolerance induced in Staphylococcus aureus by concurrent drug exposure.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.725312043.793505288 ◽

2015 ◽

Author(s):

Peter Wilson ◽

Carmel Curtis

Keyword(s):

Staphylococcus Aureus ◽

Drug Exposure ◽

Antibiotic Tolerance

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Characterization of NorR Protein, a Multifunctional Regulator of norA Expression in Staphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.185.10.3127-3138.2003 ◽

2003 ◽

Vol 185 (10) ◽

pp. 3127-3138 ◽

Cited By ~ 81

Author(s):

Que Chi Truong-Bolduc ◽

Xiamei Zhang ◽

David C. Hooper

Keyword(s):

Gene Expression ◽

Staphylococcus Aureus ◽

Efflux Pump ◽

Protein A ◽

Protein Overexpression ◽

Wild Type ◽

Liquid Media ◽

Cell Surface Properties ◽

Regulatory Systems

ABSTRACT We characterized a Staphylococcus aureus norA gene expression regulator, NorR, initially identified from its binding to the norA promoter. The norR gene was 444 bp in length, located ∼7 kb upstream from the norA gene, and encoded a predicted 17.6-kDa protein. Overexpression of norR in wild-type S. aureus strain ISP794 led to a fourfold decrease in sensitivity to quinolones and ethidium bromide and an increase in the level of norA transcripts, suggesting that NorR acts as a positive regulator of norA expression. Overexpression of norR in sarA and agr mutants did not alter quinolone sensitivity or levels of norA transcription, indicating that the presence of these two global regulatory systems is necessary for NorR to affect the expression of norA. Insertion and disruption of norR in ISP794 increased resistance to quinolones by 4- to 16-fold but had no effect on norA transcription, suggesting that NorR acts as a repressor for another unidentified efflux pump or pumps. These mutants also exhibited an exaggerated clumping phenotype in liquid media, which was complemented fully by a plasmid-encoded norR gene. Collectively, these results indicate that NorR is a multifunctional regulator, affecting cell surface properties as well as the expression of NorA and likely other multidrug resistance efflux pumps.

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sarZ, a sarA Family Gene, Is Transcriptionally Activated by MgrA and Is Involved in the Regulation of Genes Encoding Exoproteins in Staphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.01555-08 ◽

2008 ◽

Vol 191 (5) ◽

pp. 1656-1665 ◽

Cited By ~ 36

Author(s):

Anand Ballal ◽

Binata Ray ◽

Adhar C. Manna

Keyword(s):

Staphylococcus Aureus ◽

Target Genes ◽

Virulence Gene ◽

Promoter Regions ◽

Regulatory Relationship ◽

Virulence Gene Expression ◽

Regulatory Systems ◽

Expression Of Genes ◽

Genes Encoding ◽

Gel Shift

ABSTRACT The expression of genes involved in the pathogenesis of Staphylococcus aureus is controlled by global regulatory loci, including two-component regulatory systems and transcriptional regulators (e.g., sar family genes). Most members of the SarA family have been partially characterized and shown to regulate a large numbers of target genes. Here, we describe the characterization of sarZ, a sarA paralog from S. aureus, and its regulatory relationship with other members of its family. Expression of sarZ was growth phase dependent with maximal expression in the early exponential phase of growth. Transcription of sarZ was reduced in an mgrA mutant and returned to a normal level in a complemented mgrA mutant strain, which suggests that mgrA acts as an activator of sarZ transcription. Purified MgrA protein bound to the sarZ promoter region, as determined by gel shift assays. Among the sarA family of genes analyzed, inactivation of sarZ increased sarS transcription, while it decreased agr transcription. The expression of potential target genes involved in virulence was evaluated in single and double mutants of sarZ with mgrA, sarX, and agr. Northern and zymogram analyses indicated that the sarZ gene product played a role in regulating several virulence genes, particularly those encoding exoproteins. Gel shift assays demonstrated nonspecific binding of purified SarZ protein to the promoter regions of the sarZ-regulated target genes. These results demonstrate the important role played by SarZ in controlling regulatory and virulence gene expression in S. aureus.

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