Targeting a bacterial stress response to enhance antibiotic action

Lee, Samuel; Hinz, Aaron; Bauerle, Elizabeth; Angermeyer, Angus; Juhaszova, Katy; Kaneko, Yukihiro; Singh, Pradeep Kumar; Manoil, Colin

doi:10.1073/pnas.0903619106

Cited by 123 publications

(151 citation statements)

References 55 publications

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“…In Vibrio cholerae, natural transformation is induced by chitin, which is recognized by a membrane sensor that activates tfoX (76). We hypothesize that in A. actinomycetemcomitans, a similar sensor recognizes membrane stress (77,78) caused by calcium, erythromycin, and potentially other antibiotics. Experiments are under way in our laboratory to test this hypothesis.…”

Section: Discussionmentioning

confidence: 99%

Defining Genetic Fitness Determinants and Creating Genomic Resources for an Oral Pathogen

Narayanan

Ramsey

Stacy

et al. 2017

Appl Environ Microbiol

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Periodontitis is a microbial infection that destroys the structures that support the teeth. Although it is typically a chronic condition, rapidly progressing, aggressive forms are associated with the oral pathogen Aggregatibacter actinomycetemcomitans. One of this bacterium's key virulence traits is its ability to attach to surfaces and form robust biofilms that resist killing by the host and antibiotics. Though much has been learned about A. actinomycetemcomitans since its initial discovery, we lack insight into a fundamental aspect of its basic biology, as we do not know the full set of genes that it requires for viability (the essential genome). Furthermore, research on A. actinomycetemcomitans is hampered by the field's lack of a mutant collection. To address these gaps, we used rapid transposon mutant sequencing (Tn-seq) to define the essential genomes of two strains of A. actinomycetemcomitans, revealing a core set of 319 genes. We then generated an arrayed mutant library comprising Ͼ1,500 unique insertions and used a sequencing-based approach to define each mutant's position (well and plate) in the library. To demonstrate its utility, we screened the library for mutants with weakened resistance to subinhibitory erythromycin, revealing the multidrug efflux pump AcrAB as a critical resistance factor. During the screen, we discovered that erythromycin induces A. actinomycetemcomitans to form biofilms. We therefore devised a novel Tnseq-based screen to identify specific factors that mediate this phenotype and in follow-up experiments confirmed 4 mutants. Together, these studies present new insights and resources for investigating the basic biology and disease mechanisms of a human pathogen. IMPORTANCE Millions suffer from gum disease, which often is caused by Aggregatibacter actinomycetemcomitans, a bacterium that forms antibiotic-resistant biofilms. To fully understand any organism, we should be able to answer: what genes does it require for life? Here, we address this question for A. actinomycetemcomitans by determining the genes in its genome that cannot be mutated. As for the genes that can be mutated, we archived these mutants into a library, which we used to find genes that contribute to antibiotic resistance, leading us to discover that antibiotics cause A. actinomycetemcomitans to form biofilms. We then devised an approach to find genes that mediate this process and confirmed 4 genes. These results illuminate new fundamental traits of a human pathogen.KEYWORDS Aggregatibacter, Tn-seq, antibiotics, biofilms, essential genome O ne of the longest-studied human microbiomes is the "animalcules" (bacteria), first observed by van Leeuwenhoek (1), that inhabit the oral cavity. Oral bacteria readily attach to tooth surfaces and develop into spatially organized, multispecies biofilms (2). These communities also persist beneath the gum line in the subgingival pocket, where they are kept in check by host immune cells (3). Though normally benign, subgingival communities can accumulate opportunistic oral p...

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Section: Discussionmentioning

confidence: 99%

Defining Genetic Fitness Determinants and Creating Genomic Resources for an Oral Pathogen

Narayanan

Ramsey

Stacy

et al. 2017

Appl Environ Microbiol

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“…Previous studies on the bacterial sensitivity affected by electromagnetic fields were carried out and different mechanisms described this phenomenon: a) Surface charges and membrane potential. Changes in membrane potential and surface charge, make disruption in the electron transport system and energy generation by proton motive force of bacteria would be impaired (40)(41)(42) and can increase sensitivity to antibiotics (43). Since membrane potential was crucial for bacterial binary fusion, it can be interpreted that exposure to EMFs influences the behavior of bacteria in the environment (44).…”

Section: Discussionmentioning

confidence: 99%

Evaluation of the 900 MHz Radiofrequency Radiation Effects on the Antimicrobial Susceptibility and Growth Rate of Klebsiella pneumoniae

et al. 2017

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Introduction: Drug resistance to one or more antibiotics is mainly believed to be a serious threat to global public health which occurs by various mechanisms. The electromagnetic field as a nonchemical tool is capable of altering the antimicrobial susceptibility of bacteria. The aim of this study is the evaluation of the antimicrobial susceptibility of the Klebsiella pneumoniae to antibiotics before and after exposure to 900 MHz radiofrequency (RF) radiation produced by a mobile phone.Methods: Antimicrobial susceptibility of the bacteria to antibiotics disks were performed after exposure to 900 MHz radiofrequency radiation. Antimicrobial susceptibility was carried out by Kirby-Bauer and the inhibition zone of each antibiotic disk was measured (as mean). Additionally, growth variations under exposure were recorded and an outgrowth curve was drawn.Results: Based on the results, after 12 hours of exposure to 900MHz radiofrequency, there was observed a maximum increase in the sensitivity of K. pneumoniae and shown that radiofrequency (RF) radiation can change antimicrobial sensitivity significantly (P < 0.05). Also, the radiofrequency radiation effect on the bacterial proliferation was evaluated and showed that exposure enables the bacteria to grow faster in the exponential phase than the non-exposed bacteria. Conclusions:The findings of the study indicated that the sensitivity of K. pneumoniae was significantly increased to the various antibiotics after 12 hours of exposure to 900 MHz radiofrequency radiation. These observations could be interpreted by the concept of non-linearity in the responses of K. pneumoniae to different antibiotics after exposure to electromagnetic radiofrequency radiation and can be useful in the management of serious infectious diseases.

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“…The Keio collection has previously been used to identify mutants hypersensitive to at least 1 of 22 different antibiotics (16,17). Similarly, screening of transposon libraries in Acinetobacter baylyi and P. aeruginosa has identified genes whose disruption hypersensitizes these organisms to a variety of antibiotics (18)(19)(20). These studies yielded sets of proteins that could be targeted with codrugs to potentiate the activities of extant antibiotics.…”

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confidence: 99%

Recruitment of genes and enzymes conferring resistance to the nonnatural toxin bromoacetate

Desai

Miller

2010

Proc. Natl. Acad. Sci. U.S.A.

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Microbial niches contain toxic chemicals capable of forcing organisms into periods of intense natural selection to afford survival. Elucidating the mechanisms by which microbes evade environmental threats has direct relevance for understanding and combating the rise of antibiotic resistance. In this study we used a toxic small-molecule, bromoacetate, to model the selective pressures imposed by antibiotics and anthropogenic toxins. We report the results of genetic selection experiments that identify nine genes from Escherichia coli whose overexpression affords survival in the presence of a normally lethal concentration of bromoacetate. Eight of these genes encode putative transporters or transmembrane proteins, while one encodes the essential peptidoglycan biosynthetic enzyme, UDP- N -acetylglucosamine enolpyruvoyl transferase (MurA). Biochemical studies demonstrate that the primary physiological target of bromoacetate is MurA, which becomes irreversibly inactivated via alkylation of a critical active-site cysteine. We also screened a comprehensive library of E. coli single-gene deletion mutants and identified 63 strains displaying increased susceptibility to bromoacetate. One hypersensitive bacterium lacks yliJ , a gene encoding a predicted glutathione transferase. Herein, YliJ is shown to catalyze the glutathione-dependent dehalogenation of bromoacetate with a k cat / K m value of 5.4 × 10 3 M -1 s -1 . YliJ displays exceptional substrate specificity and produces a rate enhancement exceeding 5 orders of magnitude, remarkable characteristics for reactivity with a nonnatural molecule. This study illustrates the wealth of intrinsic survival mechanisms that can be exploited by bacteria when they are challenged with toxins.

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Targeting a bacterial stress response to enhance antibiotic action

Cited by 123 publications

References 55 publications

Defining Genetic Fitness Determinants and Creating Genomic Resources for an Oral Pathogen

Defining Genetic Fitness Determinants and Creating Genomic Resources for an Oral Pathogen

Evaluation of the 900 MHz Radiofrequency Radiation Effects on the Antimicrobial Susceptibility and Growth Rate of Klebsiella pneumoniae

Recruitment of genes and enzymes conferring resistance to the nonnatural toxin bromoacetate

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