Biology subjects

Dahl, J.-U.

Publications and source records attributed to Dahl, J.-U..

9 recordsLinked to original sources

bioRxiv2026

A newly identified detoxification system protects uropathogenic Escherichia coli from reactive chlorine species

Neutrophils eliminate invading pathogens through the production of reactive oxygen and chlorine species (ROS/RCS), with hypochlorous acid (HOCl) representing the most abundant and bactericidal oxidant produced in this process. Compared to bacterial defenses against ROS, which are well studied, little is known about how pathogens respond to and counter RCS, including HOCl. Here, we identify and mechanistically characterize RcrB, a protein of the uncharacterized DUF417 protein family, for which no role in oxidative stress defense has been described yet. We report a previously unrecognized role as an RCS detoxification system that confers high-level resistance to uropathogenic Escherichia coli (UPEC). We show that RcrB is an inner membrane protein and strongly induced during RCS exposure and phagocytosis. Loss of RcrB results in profound HOCl hypersensitivity, accompanied by elevated macromolecular damage and severe metabolic perturbations, establishing RcrB as a central determinant of UPECs RCS stress resistance. Heterologous expression of RcrB in HOCl-sensitive intestinal E. coli strains is sufficient to restore resistance but requires functional glutathione biosynthesis. Quantitative HOCl trapping assays demonstrate that RcrB expression protects the bacterial population by significantly reducing extracellular HOCl, indicating active chemical quenching rather than passive membrane protection. Structure-function analysis of RcrB confirms this conclusion and demonstrate that conserved, redox-active amino acids facing the periplasm are essential for its detoxification activity. In summary, our study reveals a hitherto unknown bacterial strategy for mitigating RCS and reveal a distinct mechanism by which UPEC may survive the mammalian host defense. Significance StatementDuring infection, human immune cells such as neutrophils kill bacteria by releasing powerful oxidants, including hypochlorous acid (HOCl), the active ingredient of household bleach. How pathogenic bacteria survive this chemical attack is still poorly understood. This study identifies a membrane protein in uropathogenic Escherichia coli as a key defense factor that neutralizes HOCl before it can damage the cell. RcrB acts at the bacterial cell envelope, where it detoxifies HOCl in a glutathione-dependent manner to maintain a balanced redox homeostasis and cellular integrity, thereby even protect neighboring bacteria. These findings reveal a previously unrecognized frontline defense strategy that helps pathogens survive immune attack and may represent a new target for antimicrobial therapies.

microbiology↗

bioRxiv2026

Enhancement of low-temperature growth of Staphylococcus aureus by low concentrations of antimicrobial unsaturated fatty acids

The incorporation of oleic acid, a physiologically relevant and relatively non-toxic fatty acid, into membrane phospholipids and glycolipids has major impacts on the membrane biophysical properties of Staphylococcus aureus, including the promotion of low-temperature growth. Inclusion of oleic acid in growth media decreased the minimum growth temperature of S. aureus. It has not been studied yet whether other physiologically relevant, yet more antimicrobial straight-chain unsaturated fatty acids (SCUFAs), elicit similar responses. Here, we report that exogenous addition of these antimicrobial SCUFAs at 12{degrees}C led to their incorporation into the membrane lipids in only relatively small amounts, which, however, was sufficient to strongly promote growth at low temperatures. Intriguingly, these SCUFAs showed a preferential incorporation into diglucosyldiacylglyceride (DGDG) over phosphatidyl glycerol, and increased DGDG levels [~] 4-fold in SCUFA-grown S. aureus. Two glycolipid-deficient mutants lost the growth-enhancing effects of SCUFAs at low temperatures and showed increased susceptibility to SCUFAs at 37{degrees}C, suggesting that glycolipids are important membrane components and are required for low-temperature adaptation. The presence of SCUFAs at low temperatures also enhanced the production of the carotenoid staphyloxanthin. Overall, our results suggest that multiple strategies are at play in the membrane lipid remodeling of S. aureus to adapt to the stress of low-temperature growth and likely to other environmental stresses. Incorporation of antimicrobial SCUFAs into membrane lipids may facilitate the insertion of free forms of these SCUFAs into the membrane and be a part of their mode of action.

microbiology↗

bioRxiv2025

The Synergy between a Silver-Ruthenium Antimicrobial and aminoglycosides is based on severe macromolecular damage

The rise of multidrug-resistant (MDR) bacterial pathogens, including uropathogenic Escherichia coli (UPEC), highlights the urgent need for alternative treatment strategies to restore antibiotic efficacy. The silver-ruthenium antimicrobial AGXX(R) exerts potent bactericidal effects through the production of reactive oxygen species (ROS); however, its potential synergy with antibiotics has not been thoroughly investigated. Here, we show that sublethal concentrations of AGXX(R) strongly enhance aminoglycoside-mediated killing across a diverse panel of Gram-negative and Gram-positive MDR clinical isolates, including highly aminoglycoside-resistant strains. Combinational treatments significantly reduced the effective concentrations of gentamicin, tobramycin, kanamycin, and amikacin required for bacterial killing. Mechanistic analyses revealed that AGXX(R)/aminoglycoside co-treatment induces pronounced intracellular ROS accumulation, resulting in severe proteotoxic stress, extensive protein aggregation, and DNA damage. Scavenging ROS abolished synergistic killing, establishing oxidative imbalance as the primary driver of the synergy between both antimicrobials. We further identify polyphosphate as a key bacterial defense mechanism that mitigates ROS accumulation, proteotoxicity, and genotoxic stress during combinational treatment. Moreover, AGXX(R)-aminoglycoside synergy was preserved in an artificial urine medium and across clinical UPEC isolates, underscoring its relevance to urinary tract infections. Together, these findings position AGXX(R) as a potent aminoglycoside adjuvant that restores antibiotic efficacy through ROS-driven macromolecular damage, supporting its development for combination therapies against MDR bacterial infections.

microbiology↗

bioRxiv2025

A Novel Silver-Ruthenium-Based Antimicrobial Kills Gram-Negative Bacteria Through Oxidative Stress-Induced Macromolecular Damage

Amplified by the decline in antibiotic discovery, the rise of antibiotic resistance has become a significant global challenge in infectious disease control. Extraintestinal Escherichia coli (ExPEC), known to be the most common instigators of urinary tract infections (UTIs), represent such global threat. Novel strategies for more efficient treatments are therefore desperately needed. These include silver nanoparticles, which have been used as antimicrobial surface-coatings on catheters to eliminate biofilm-forming uropathogens and reduce the risk of nosocomial infections. AGXX(R) is a promising silver coating that presumably kills bacteria through the generation of reactive oxygen species (ROS) but is more potent than silver. However, neither is AGXX(R)s mode of action fully understood, nor have its effects on Gram-negative bacteria or bacterial response and defense mechanisms towards AGXX(R) been studied in detail. Here, we report that the bactericidal effects of AGXX(R) are primarily based on ROS formation, as supplementation of the media with a ROS scavenger completely abolished AGXX(R)-induced killing. We further show that AGXX(R) impairs the integrity of the bacterial cell envelope and causes substantial protein aggregation and DNA damage already at sublethal concentrations. ExPEC strains appear to be more resistant to the proteotoxic effects of AGXX(R) compared to non-pathogenic E. coli, indicating improved defense capabilities of the uropathogen. Global transcriptomic studies of AGXX(R)-stressed ExPEC revealed a strong oxidative stress response, perturbations in metal homeostasis, as well as the activation of heat shock and DNA damage responses. Finally, we present evidence that ExPEC counter AGXX(R) damage through the production of the chaperone polyphosphate.

microbiology↗

bioRxiv2024

Lipidomics of homeoviscous adaptation to low temperatures in Staphylococcus aureus utilizing exogenous straight-chain unsaturated fatty acids over biosynthesized endogenous branched-chain fatty acids

It is well established that Staphylococcus aureus can incorporate exogenous straight-chain unsaturated fatty acids (SCUFAs) into membrane phospho- and glyco-lipids from various sources in supplemented culture media, and when growing in vivo in an infection. Given the enhancement of membrane fluidity when oleic acid (C18:1{Delta}9) is incorporated into lipids, we were prompted to examine the effect of medium supplementation with C18:1{Delta}9 on growth at low temperatures. C18:1{Delta}9 supported the growth of a cold-sensitive, branched-chain fatty acid (BCFA)-deficient mutant at 12{degrees}C. Interestingly, we found similar results in the BCFA-sufficient parental strain. We show that incorporation of C18:1{Delta}9 and its elongation product C20:1{Delta}9 into membrane lipids was required for growth stimulation and relied on a functional FakAB incorporation system. Lipidomics analysis of the phosphatidylglycerol (PG) and diglycosyldiacylglycerol (DGDG) lipid classes revealed major impacts of C18:1{Delta}9 and temperature on lipid species. Growth at 12{degrees}C in the presence of C18:1{Delta}9 also led to increased production of the carotenoid pigment staphyloxanthin; however, this was not an obligatory requirement for cold adaptation. Enhancement of growth by C18:1{Delta}9 is an example of homeoviscous adaptation to low temperatures utilizing an exogenous fatty acid. This may be significant in the growth of S. aureus at low temperatures in foods that commonly contain C18:1{Delta}9 and other SCUFAs in various forms. IMPORTANCEWe show that S. aureus can use its known ability to incorporate exogenous fatty acids to enhance its growth at low temperatures. Individual species of phosphatidylglycerols and diglycosyldiacylglycerol bearing one or two degrees of unsaturation derived from incorporation of C18:1{Delta}9 at 12{degrees}C are described for the first time. In addition, enhanced production of the carotenoid staphyloxanthin occurs at low temperatures. The studies describe a biochemical reality underlying in membrane biophysics. This is an example of homeoviscous adaptation to low temperatures utilizing exogenous fatty acids over the regulation of the biosynthesis of endogenous fatty acids. The studies have likely relevance to food safety in that unsaturated fatty acids may enhance growth of S. aureus in the food environment.

microbiology↗

bioRxiv2023

Pseudomonas aeruginosa kills Staphylococcus aureus in a polyphosphate-dependent manner

Due to their frequent coexistence in many polymicrobial infections, including in patients with burn or chronic wounds or cystic fibrosis, recent studies have started to investigate the mechanistic details of the interaction between the opportunistic pathogens Pseudomonas aeruginosa and Staphylococcus aureus. P. aeruginosa rapidly outcompetes S. aureus under in vitro co-cultivation conditions, which is mediated by several of P. aeruginosas virulence factors. Here, we report that polyphosphate (polyP), an efficient stress defense system and virulence factor in P. aeruginosa, plays a role for the pathogens ability to inhibit and kill S. aureus in a contact-independent manner. We show that P. aeruginosa cells characterized by low polyP level are less detrimental to S. aureus growth and survival while the gram-positive pathogen is significantly more compromised by the presence of P. aeruginosa cells that produce high level of polyP. We show that the polyP-dependent phenotype could be a direct effect by the biopolymer, as polyP is present in the spent media and causes significant damage to the S. aureus cell envelope. However, more likely is that polyPs effects are indirect through the regulation of one of P. aeruginosas virulence factors, pyocyanin. We show that pyocyanin production in P. aeruginosa occurs polyP-dependent and harms S. aureus through membrane damage and the generation of reactive oxygen species, resulting in increased expression of antioxidant enzymes. In summary, our study adds a new component to the list of biomolecules that the gram-negative pathogen P. aeruginosa generates to compete with S. aureus for resources. IMPORTANCEHow do interactions between microorganisms shape the course of polymicrobial infections? Previous studies have provided evidence that the two opportunistic pathogens P. aeruginosa and S. aureus generate molecules that modulate their interaction with potentially significant impact on disease outcomes. Our study identified the biopolymer polyP as a new effector molecule that impacts P. aeruginosas interaction with S. aureus. We show that P. aeruginosa kills S. aureus in a polyP-dependent manner, which occurs primarily through the polyP-dependent production of the P. aeruginosa virulence factor pyocyanin. Our findings add a new role for polyP to an already extensive list of functions. A more in-depth understanding of how polyP influences interspecies interactions is critical, as targeting polyP synthesis in bacteria such as P. aeruginosa may have a significant impact on other microorganisms and potentially result in dynamic changes in the microbial composition.

microbiology↗

bioRxiv2023

Expression of RcrB confers resistance to hypochlorous acid in uropathogenic Escherichia coli

To eradicate bacterial pathogens, neutrophils are recruited to the sites of infection, where they engulf and kill microbes through the production of reactive oxygen and chlorine species (ROS/RCS). The most prominent RCS is antimicrobial oxidant hypochlorous acid (HOCl), which rapidly reacts with various amino acids side chains, including those containing sulfur and primary/tertiary amines, causing significant macromolecular damage. Pathogens like uropathogenic Escherichia coli (UPEC), the primary causative agent of urinary tract infections (UTIs), have developed sophisticated defense systems to protect themselves from HOCl. We recently identified the RcrR regulon as a novel HOCl defense strategy in UPEC. The regulon is controlled by the HOCl-sensing transcriptional repressor RcrR, which is oxidatively inactivated by HOCl resulting in the expression of its target genes, including rcrB. rcrB encodes the putative membrane protein RcrB, deletion of which substantially increases UPECs susceptibility to HOCl. However, many questions regarding RcrBs role remain open including whether (i) the proteins mode of action requires additional help, (ii) rcrARB expression is induced by physiologically relevant oxidants other than HOCl, and (iii) expression of this defense system is limited to specific media and/or cultivation conditions. Here, we provide evidence that RcrB expression is sufficient to E. colis protection from HOCl and induced by and protects from several RCS but not from ROS. RcrB plays a protective role for RCS-stressed planktonic cells under various growth and cultivation conditions but appears to be irrelevant for UPECs biofilm formation. IMPORTANCEBacterial infections pose an increasing threat to human health exacerbating the demand for alternative treatment options. UPEC, the most common etiological agent of urinary tract infections (UTIs), are confronted by neutrophilic attacks in the bladder, and must therefore be well equipped with powerful defense systems to fend off the toxic effects of RCS. How UPEC deal with the negative consequences of the oxidative burst in the neutrophil phagosome remains unclear. Our study sheds light on the requirements for the expression and protective effects of RcrB, which we recently identified as UPECs most potent defense system towards HOCl-stress and phagocytosis. Thus, this novel HOCl-stress defense system could potentially serve as an attractive drug target to increase the bodys own capacity to fight UTIs.

microbiology↗

bioRxiv2023

The Novel Silver-Containing Antimicrobial Potentiates Aminoglycoside Activity Against Pseudomonas aeruginosa.

The rapid dissemination of antibiotic resistance combined with the decline in the discovery of novel antibiotics represents a major challenge for infectious disease control that can only be mitigated by investments into novel treatment strategies. Alternative antimicrobials, including silver, have regained interest due to their diverse mechanisms of inhibiting microbial growth. One such example is AGXX(R), a broad-spectrum silver containing antimicrobial that produces highly cytotoxic reactive oxygen species (ROS) to inflict extensive macromolecular damage. Due to connections identified between ROS production and antibiotic lethality, we hypothesized that AGXX(R) could potentially increase the activity of conventional antibiotics. Using the gram-negative pathogen Pseudomonas aeruginosa, we screened possible synergistic effects of AGXX(R) on several antibiotic classes. We found that the combination of AGXX(R) and aminoglycosides tested at sublethal concentrations led to a rapid exponential decrease in bacterial survival and restored sensitivity of a kanamycin-resistant strain. ROS production contributes significantly to the bactericidal effects of AGXX(R)/aminoglycoside treatments, which is dependent on oxygen availability and can be reduced by the addition of ROS scavengers. Additionally, P. aeruginosa strains deficient in ROS detoxifying/repair genes were more susceptible to AGXX(R)/aminoglycoside treatment. We further demonstrate that this synergistic interaction was associated with significant increase in outer and inner membrane permeability, resulting in increased antibiotic influx. Our study also revealed that AGXX(R)/aminoglycoside-mediated killing requires an active proton motive force across the bacterial membrane. Overall, our findings provide an understanding of cellular targets that could be inhibited to increase the activity of conventional antimicrobials. IMPORTANCEThe emergence of drug-resistant bacteria coupled with the decline in antibiotic development highlights the need for novel alternatives. Thus, new strategies aimed at repurposing conventional antibiotics have gained significant interest. The necessity of these interventions is evident especially in gram-negative pathogens as they are particularly difficult to treat due to their outer membrane. This study highlights the effectiveness of the silver containing antimicrobial AGXX(R) in potentiating aminoglycoside activities against P. aeruginosa. The combination of AGXX(R) and aminoglycosides not only reduces bacterial survival rapidly but also significantly re-sensitizes aminoglycoside-resistant P. aeruginosa strains. In combination with gentamicin, AGXX(R) induces increased endogenous oxidative stress, membrane damage and iron sulfur cluster disruption. These findings emphasize AGXX(R)s potential as a route of antibiotic adjuvant development and shed light into potential targets to enhance aminoglycoside activity.

microbiology↗

bioRxiv2021

Redox-mediated inactivation of the transcriptional repressor C3600 makes uropathogenic Escherichia coli exquisitely resistant to reactive chlorine species

The ability to overcome stressful environments is critical for pathogen survival in the host. One challenge for bacteria is the exposure to reactive chlorine species (RCS), which are generated by innate immune cells as critical part of the oxidative burst. Hypochlorous acid (HOCl) is the most potent antimicrobial RCS and associated with extensive macromolecular damage in the phagocytized pathogen. However, bacteria have evolved defense strategies to alleviate the effects of HOCl-mediated damage. Among these are RCS-sensing transcriptional regulators that control the expression of HOCl-protective genes under non- and HOCl stress. Uropathogenic Escherichia coli (UPEC), the major causative agent of urinary tract infections (UTIs), is particularly exposed to infiltrating neutrophils during pathogenesis, however, their responses to and defenses of HOCl are still completely unexplored. Here, we present evidence that UPEC strains tolerate higher levels of HOCl and are better protected from neutrophil-mediated killing compared to other E. coli. Transcriptomic analysis of HOCl-stressed UPEC revealed the upregulation of an operon consisting of three genes, one of which encodes the transcriptional regulator C3600. We identified C3600 as a HOCl-sensing transcriptional repressor, which, under non-stress conditions, is bound to the operator and represses the expression of its target genes. During HOCl exposure, however, the repressor forms reversible intermolecular disulfide bonds and dissociates from the DNA resulting in the de-repression of the operon. Deletion of one of the target genes renders UPEC significantly more susceptible to HOCl indicating that the HOCl-mediated induction of the regulon plays a major role for UPECs HOCl resistance. IMPORTANCEHow do pathogens deal with antimicrobial oxidants produced by the innate immune system during infection? Uropathogenic Escherichia coli (UPEC), the most common etiological agent of urinary tract infections (UTIs), is particularly exposed to infiltrating neutrophils and, therefore, must counter elevated levels of the antimicrobial oxidant HOCl to establish infection. Our study provides fundamentally new insights into a defense mechanism that enables UPEC to fend off the toxic effects of HOCl stress. Intriguingly, the defense system is predominantly found in UPEC and absent in non-invasive enteropathogenic E. coli. Our data suggest that expression of the target gene c3601 is exclusively responsible for UPECs increased HOCl tolerance in culture and therefore potentially contributes to UPECs survival during phagocytosis. Thus, this novel HOCl stress defense system could potentially serve as an attractive drug target to increase the bodys own capacity to fight UTIs.

microbiology↗