Search Results (34)

Persister cells are a subset of bacterial cells that exhibit transient antibiotic tolerance without genetic resistance, contributing to the persistence of chronic infections. This study investigates the ability of diosgenin, a naturally occurring steroidal saponin, to inhibit persister cell formation in Staphylococcus aureus through metabolic suppression and membrane modulation. Diosgenin treatments at 80 µM and 160 µM significantly reduced persister cell survival under oxacillin, ciprofloxacin, and gentamicin stress, with reductions ranging from 82% to 94% after 3 h diosgenin pre-exposure. Gene expression analysis revealed that diosgenin downregulated relP and relQ, key genes involved in (p)ppGpp synthesis, by up to 60%, accompanied by 36–38% decreases in intracellular ATP levels. Diosgenin did not significantly alter membrane permeability or membrane potential but reduced membrane fluidity by 35% and 41% at 80 µM and 160 µM, respectively. Taken together, our findings suggest that diosgenin exerts a dual-action regulatory effect on persister cell formation by disrupting metabolic pathways essential for dormancy and altering membrane dynamics, potentially affecting membrane-associated signaling. This study provides a framework for the further exploration of diosgenin as a potential anti-persister agent with particular promise for use in combination with conventional antibiotics to enhance therapeutic efficacy against chronic bacterial infections. Full article

Tuberculosis (TB) remains a threat to human health worldwide. Mycobacterium tuberculosis (Mtb) and other nontuberculous mycobacteria (NTM) can form biofilms, and in vitro and animal experiments have shown that biofilms cause serious drug resistance and mycobacterial persistence. Deeper investigations into the mechanisms of mycobacterial biofilm formation and, consequently, the exploration of appropriate antibiofilm treatments to improve the efficiency of current anti-TB drugs will be useful for curing TB. In this review, the genes and molecules that have been recently reported to be involved in mycobacterial biofilm development, such as ABC transporter, Pks1, PpiB, GroEL1, MprB, (p)ppGpp, poly(P), and c-di-GMP, are summarized. Biofilm-induced clinical problems, including biofilm-related infections and enhanced virulence, as well as their possible mechanisms, are also discussed in detail. Moreover, we also illustrate newly synthesized anti-TB agents that target mycobacterial biofilm, as well as some assistant methods with high efficiency in reducing biofilms in hosts, such as the use of nanoparticles. Full article

Resistance evolution during exposure to non-lethal levels of antibiotics is influenced by various stress responses of bacteria which are known to affect growth rate. Here, we aim to disentangle how the interplay between resistance development and associated fitness costs is affected by stress responses. We performed de novo resistance evolution of wild-type strains and single-gene knockout strains in stress response pathways using four different antibiotics. Throughout resistance development, the increase in minimum inhibitory concentration (MIC) is accompanied by a gradual decrease in growth rate, most pronounced in amoxicillin or kanamycin. By measuring biomass yield on glucose and whole-genome sequences at intermediate and final time points, we identified two patterns of how the stress responses affect the correlation between MIC and growth rate. First, single-gene knockout E. coli strains associated with reactive oxygen species (ROS) acquire resistance faster, and mutations related to antibiotic permeability and pumping out occur earlier. This increases the metabolic burden of resistant bacteria. Second, the ΔrelA knockout strain, which has reduced (p)ppGpp synthesis, is restricted in its stringent response, leading to diminished growth rates. The ROS-related mutagenesis and the stringent response increase metabolic burdens during resistance development, causing lower growth rates and higher fitness costs. Full article

In addition to comprising monomers of nucleic acids, nucleotides have signaling functions and act as second messengers in both prokaryotic and eukaryotic cells. The most common example is cyclic AMP (cAMP). Nucleotide signaling is a focus of great interest in bacteria. Cyclic di-AMP (c-di-AMP), cAMP, and cyclic di-GMP (c-di-GMP) participate in biological events such as bacterial growth, biofilm formation, sporulation, cell differentiation, motility, and virulence. Moreover, the cyclic-di-nucleotides (c-di-nucleotides) produced in pathogenic intracellular bacteria can affect eukaryotic host cells to allow for infection. On the other hand, non-cyclic nucleotide molecules pppGpp and ppGpp are alarmones involved in regulating the bacterial response to nutritional stress; they are also considered second messengers. These second messengers can potentially be used as therapeutic agents because of their immunological functions on eukaryotic cells. In this review, the role of c-di-nucleotides and cAMP as second messengers in different bacterial processes is addressed. Full article

The treatment of many bacterial and fungal infections remains a problem due to increasing antibiotic resistance and biofilm formation by pathogens. In the present article, a methodology for the chemoselective synthesis of 2-(1H-indol-3-yl)-1H-benzo[d]imidazole derivatives is presented. We report on the antimicrobial activity of synthesized 2-(1H-indol-3-yl)-1H-benzo[d]imidazoles with significant activity against Staphylococcus aureus ATCC 25923, Staphylococcus aureus ATCC 43300 (MRSA), Mycobacterium smegmatis (mc(2)155/ATCC 700084), and Candida albicans ATCC 10231. High activity against staphylococci was shown by indolylbenzo[d]imidazoles 3ao and 3aq (minimum inhibitory concentration (MIC) < 1 µg/mL) and 3aa and 3ad (MIC 3.9–7.8 µg/mL). A low MIC was demonstrated by 2-(1H-indol-3-yl)-1-methyl-1H-benzo[d]imidazole (3ag) against M. smegmatis and against C. albicans (3.9 µg/mL and 3.9 µg/mL, respectively). 2-(5-Bromo-1H-indol-3-yl)-6,7-dimethyl-1H-benzo[d]imidazole (3aq) showed a low MIC of 3.9 µg/mL against C. albicans. Compounds 3aa, 3ad, 3ao, and 3aq exhibited excellent antibiofilm activity, inhibiting biofilm formation and killing cells in mature biofilms. Molecular docking analysis identified three potential interaction models for the investigated compounds, implicating (p)ppGpp synthetases/hydrolases, FtsZ proteins, or pyruvate kinases in their antibacterial action mechanism. Full article

DksA is a proteobacterial regulator that binds directly to the secondary channel of RNA polymerase with (p)ppGpp and is responsible for various bacterial physiological activities. While (p)ppGpp is known to be involved in the regulation and response of fatty acid metabolism pathways in many foodborne pathogens, the role of DksA in this process has yet to be clarified. This study aimed to characterize the function of DksA on fatty acid metabolism and cell membrane structure in Yersinia enterocolitica. Therefore, comparison analysis of gene expression, growth conditions, and membrane permeabilization among the wide-type (WT), DksA-deficient mutant (YEND), and the complemented strain was carried out. It confirmed that deletion of DksA led to a more than four-fold decrease in the expression of fatty acid degradation genes, including fadADEIJ. Additionally, YEND exhibited a smaller growth gap compared to the WT strain at low temperatures, indicating that DksA is not required for the growth of Y. enterocolitica in cold environments. Given that polymyxin B is a cationic antimicrobial peptide that targets the cell membrane, the roles of DksA under polymyxin B exposure were also characterized. It was found that DksA positively regulates the integrity of the inner and outer membranes of Y. enterocolitica under polymyxin B, preventing the leakage of intracellular nucleic acids and proteins and ultimately reducing the sensitivity of Y. enterocolitica to polymyxin B. Taken together, this study provides insights into the functions of DksA and paves the way for novel fungicide development. Full article

The treatment of many bacterial diseases remains a significant problem due to the increasing antibiotic resistance of their infectious agents. Among others, this is related to Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA) and Mycobacterium tuberculosis. In the present article, we report on antibacterial compounds with activity against both S. aureus and MRSA. A straightforward approach to 2-(1H-indol-3-yl)quinazolin-4(3H)-one and their analogues was developed. Their structural and functional relationships were also considered. The antimicrobial activity of the synthesized compounds against Mycobacterium tuberculosis H₃₇Rv, S. aureus ATCC 25923, MRSA ATCC 43300, Candida albicans ATCC 10231, and their role in the inhibition of the biofilm formation of S. aureus were reported. 2-(5-Iodo-1H-indol-3-yl)quinazolin-4(3H)-one (3k) showed a low minimum inhibitory concentration (MIC) of 0.98 μg/mL against MRSA. The synthesized compounds were assessed via molecular docking for their ability to bind long RSH (RelA/SpoT homolog) proteins using mycobacterial and streptococcal (p)ppGpp synthetase structures as models. The cytotoxic activity of some synthesized compounds was studied. Compounds 3c, f, g, k, r, and 3z displayed significant antiproliferative activities against all the cancer cell lines tested. Indolylquinazolinones 3b, 3e, and 3g showed a preferential suppression of the growth of rapidly dividing A549 cells compared to slower growing fibroblasts of non-tumor etiology. Full article

Maintaining high system performance is critical for a multi-hop flying mesh network (FlyMesh) to perform missions in different environments. Although the Poisson point process (PPP) has been widely used for the performance analysis of FlyMesh, it still has flaws in describing the spatial distribution of the UAVs since it does not restrict the minimum distance between them. The spatial deployment of FlyMesh varies depending on the environment. Considering the relevance and practicality, we modeled the multi-hop FlyMesh using the $\beta$-Ginibre point process ($\beta$-GPP) and equipped each UAV with a directional antenna. Under the condition of the decode-and-forward protocol, we derived the connection probability and ergodic capacity of a multi-hop FlyMesh utilizing the Laplace transform of interference. Then, we calculated an approximate expression for the interference Laplace transform based on the diagonal approximation and further obtained the coverage probability. Finally, the numerical simulation results verified the correctness of the theoretical derivation, indicating that it is possible to optimize the system’s performance based on the expressions derived in this paper. Full article

The stringent response is a rapid response system that is ubiquitous in bacteria, allowing them to sense changes in the external environment and undergo extensive physiological transformations. However, the regulators (p)ppGpp and DksA have extensive and complex regulatory patterns. Our previous studies demonstrated that (p)ppGpp and DksA in Yersinia enterocolitica positively co-regulated motility, antibiotic resistance, and environmental tolerance but had opposite roles in biofilm formation. To reveal the cellular functions regulated by (p)ppGpp and DksA comprehensively, the gene expression profiles of wild-type, ΔrelA, ΔrelAΔspoT, and ΔdksAΔrelAΔspoT strains were compared using RNA-Seq. Results showed that (p)ppGpp and DksA repressed the expression of ribosomal synthesis genes and enhanced the expression of genes involved in intracellular energy and material metabolism, amino acid transport and synthesis, flagella formation, and the phosphate transfer system. Additionally, (p)ppGpp and DksA inhibited amino acid utilization (such as arginine and cystine) and chemotaxis in Y. enterocolitica. Overall, the results of this study unraveled the link between (p)ppGpp and DksA in the metabolic networks, amino acid utilization, and chemotaxis in Y. enterocolitica and enhanced the understanding of stringent responses in Enterobacteriaceae. Full article

The stringent response, originally identified in Escherichia coli as a signal that leads to reprogramming of gene expression under starvation or nutrient deprivation, is now recognized as ubiquitous in all bacteria, and also as part of a broader survival strategy in diverse, other stress conditions. Much of our insight into this phenomenon derives from the role of hyperphosphorylated guanosine derivatives (pppGpp, ppGpp, pGpp; guanosine penta-, tetra- and tri-phosphate, respectively) that are synthesized on starvation cues and act as messengers or alarmones. These molecules, collectively referred to here as (p)ppGpp, orchestrate a complex network of biochemical steps that eventually lead to the repression of stable RNA synthesis, growth, and cell division, while promoting amino acid biosynthesis, survival, persistence, and virulence. In this analytical review, we summarize the mechanism of the major signaling pathways in the stringent response, consisting of the synthesis of the (p)ppGpp, their interaction with RNA polymerase, and diverse factors of macromolecular biosynthesis, leading to differential inhibition and activation of specific promoters. We also briefly touch upon the recently reported stringent-like response in a few eukaryotes, which is a very disparate mechanism involving MESH1 (Metazoan SpoT Homolog 1), a cytosolic NADPH phosphatase. Lastly, using ppGpp as an example, we speculate on possible pathways of simultaneous evolution of alarmones and their multiple targets. Full article

Urinary tract infections are one of the most frequent bacterial diseases worldwide. UPECs are the most prominent group of bacterial strains among pathogens responsible for prompting such infections. As a group, these extra-intestinal infection-causing bacteria have developed specific features that allow them to sustain and develop in their inhabited niche of the urinary tract. In this study, we examined 118 UPEC isolates to determine their genetic background and antibiotic resistance. Moreover, we investigated correlations of these characteristics with the ability to form biofilm and to induce a general stress response. We showed that this strain collection expressed unique UPEC attributes, with the highest representation of FimH, SitA, Aer, and Sfa factors (100%, 92.5%, 75%, and 70%, respectively). According to CRA (Congo red agar) analysis, the strains particularly predisposed to biofilm formation represented 32.5% of the isolates. Those biofilm forming strains presented a significant ability to accumulate multi-resistance traits. Most notably, these strains presented a puzzling metabolic phenotype—they showed elevated basal levels of (p)ppGpp in the planktonic phase and simultaneously exhibited a shorter generation time when compared to non-biofilm-forming strains. Moreover, our virulence analysis showed these phenotypes to be crucial for the development of severe infections in the Galleria mellonella model. Full article

Clinical and epidemiologic data on pustular psoriasis are scarce. To investigate the phenotypes of pustular psoriasis and the patients’ characteristics observed in a real-life retrospective observational study. The number of incident cases of pustular psoriasis registered in the period 2005–2021 was retrieved from the electronic medical records of the University Hospital of Verona. One hundred and forty cases of pustular psoriasis were collected. Ninety-one out of 140 patients (65%) were females, with a median (IQR) age of 57 (43–66) years. According to the ERASPEN classification criteria, 116 patients (83%) had palmoplantar pustulosis (PPP), 13 (9%) generalized pustular psoriasis (GPP), and 11 (8%) acrodermatitis continua of Hallopeau (ACH). Gender distribution and median age were consistent among the three groups. The prevalence of psoriatic arthritis in GPP, ACH, and PPP was 8%, 36%, and 28%, respectively. During the same period, a total of 4718 cases of plaque psoriasis were retrieved, with a 1:34 ratio of pustular over plaque psoriasis. Pustular psoriasis is much rarer than plaque psoriasis, with PPP being the more common subtype. Full article

Antimicrobial resistance (AMR) poses a serious threat to our society from both the medical and economic point of view, while the antibiotic discovery pipeline has been dwindling over the last decades. Targeting non-essential bacterial pathways, such as those leading to antibiotic persistence, a bacterial bet-hedging strategy, will lead to new molecular entities displaying low selective pressure, thereby reducing the insurgence of AMR. Here, we describe a way to target (p)ppGpp (guanosine tetra- or penta-phosphate) signaling, a non-essential pathway involved in the formation of persisters, with a structure-based approach. A superfamily of enzymes called RSH (RelA/SpoT Homolog) regulates the intracellular levels of this alarmone. We virtually screened several fragment libraries against the (p)ppGpp synthetase domain of our RSH chosen model Rel_Seq, selected three main chemotypes, and measured their interaction with Rel_Seq by thermal shift assay and STD-NMR. Most of the tested fragments are selective for the synthetase domain, allowing us to select the aminobenzoic acid scaffold as a hit for lead development. Full article

Bacteria convert active 70S ribosomes to inactive 100S ribosomes to survive under various stress conditions. This state, in which the ribosome loses its translational activity, is known as ribosomal hibernation. In gammaproteobacteria such as Escherichia coli, ribosome modulation factor and hibernation-promoting factor are involved in forming 100S ribosomes. The expression of ribosome modulation factor is regulated by (p)ppGpp (which is induced by amino acid starvation), cAMP-CRP (which is stimulated by reduced metabolic energy), and transcription factors involved in biofilm formation. This indicates that the formation of 100S ribosomes is an important strategy for bacterial survival under various stress conditions. In recent years, the structures of 100S ribosomes from various bacteria have been reported, enhancing our understanding of the 100S ribosome. Here, we present previous findings on the 100S ribosome and related proteins and describe the stress-response pathways involved in ribosomal hibernation. Full article

The stringent response regulators, (p)ppGpp and DksA, modulate various genes involved in physiological processes, virulence, and antimicrobial resistance in pathogenic bacteria. This study investigated the role of DksA in the antimicrobial susceptibility of Acinetobacter baumannii. The ∆dksA mutant (KM0248D) of A. baumannii ATCC 17978 and its complemented strain (KM0248C) were used, in addition to the ∆dksA mutant strain (NY0298D) of clinical 1656-2 strain. The microdilution assay was used to determine the minimum inhibitory concentrations (MICs) of antimicrobial agents. Quantitative real-time PCR was performed to analyze the expression of genes associated with efflux pumps. The KM0248D strain exhibited an increase of MICs to quinolones and tetracyclines, whereas KM0248D and NY0298D strains exhibited a decrease of MICs to aminoglycosides. The expression of genes associated with efflux pumps, including adeB, adeI/J, abeM, and/or tetA, was upregulated in both ∆dksA mutant strains. The deletion of dksA altered bacterial morphology in the clinical 1656-2 strain. In conclusion, DksA modulates the antimicrobial susceptibility of A. baumannii. The ∆dksA mutant strains of A. baumannii upregulate efflux pump gene expression, whereas (p)ppGpp-deficient mutants downregulate efflux pump gene expression. (p)ppGpp and DksA conduct opposite roles in the antimicrobial susceptibility of A. baumannii via efflux pump gene regulation. Full article