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BioRxiv : the Preprint Server For... Jun 2024The multidrug-resistant, nosocomial pathogen is a major threat to human health. A sensor kinase-response regulator system, BfmS-BfmR, is a potential antimicrobial...
The multidrug-resistant, nosocomial pathogen is a major threat to human health. A sensor kinase-response regulator system, BfmS-BfmR, is a potential antimicrobial target in the bacterium due to its essential role in promoting drug resistance and virulence. Important questions remain, however, about how the system controls resistance and pathogenesis. Although knockout of BfmR is known to alter expression of >1000 genes, its direct regulon is undefined. Moreover, how phosphorylation controls BfmR is unclear. Here, we address these problems by combining mutagenesis, ChIP-seq, and reactions using a small phosphodonor to study how phosphorylation affects BfmR function. We show that phosphorylation requires the BfmR active site Asp58 and is essential to gene regulation, antibiotic resistance, and virulence in a sepsis model. Consistent with activation of the protein, phosphorylation induces dimerization and increases its affinity for target DNA. Integrated analysis of the genome-wide binding and transcriptional profiles of BfmR led to several key findings: (1) Phosphorylation dramatically expands the number of genomic sites bound by BfmR, from 4 to >250; (2) BfmR recognizes a direct repeat motif widespread across promoters; (3) BfmR directly regulates >300 genes as activator (eg, capsule, peptidoglycan, and outer membrane biogenesis) or repressor (eg, type IV pilus machinery); (4) The regulator also directly controls a set of non-coding sRNAs. These studies reveal the centrality of a phosphorylation signal in driving resistance and pathogenicity and unravel the extensive gene regulatory network under its control.
PubMed: 38948834
DOI: 10.1101/2024.06.16.599214 -
BioRxiv : the Preprint Server For... Jun 2024Emerging antibiotic resistance requires continual improvement in the arsenal of antimicrobial drugs, especially the critical macrolide antibiotics. Formation of the...
Emerging antibiotic resistance requires continual improvement in the arsenal of antimicrobial drugs, especially the critical macrolide antibiotics. Formation of the macrolactone scaffold of these polyketide natural products is catalyzed by a modular polyketide synthase (PKS) thioesterase (TE). The TE accepts a linear polyketide substrate from the termina PKS acyl carrier protein to generate an acyl-enzyme adduct that is resolved by attack of a substrate hydroxyl group to form the macrolactone. Our limited mechanistic understanding of TE selectivity for a substrate nucleophile and/or water has hampered development of TEs as biocatalysts that accommodate a variety of natural and non-natural substrates. To understand how TEs direct the substrate nucleophile for macrolactone formation, acyl-enzyme intermediates were trapped as stable amides by substituting the natural serine OH with an amino group. Incorporation of the unnatural amino acid, 1,3-diaminopropionic acid (DAP), was tested with five PKS TEs. DAP-modified TEs (TE ) from the pikromycin and erythromycin pathways were purified and tested with six full-length polyketide intermediates from three pathways. The erythromycin TE had permissive substrate selectivity, whereas the pikromycin TE was selective for its native hexaketide and heptaketide substrates. In a crystal structure of a native substrate trapped in pikromycin TE , the linear heptaketide was curled in the active site with the nucleophilic hydroxyl group positioned 4 Å from the amide-enzyme linkage. The curled heptaketide displayed remarkable shape complementarity with the TE acyl cavity. The strikingly different shapes of acyl cavities in TEs of known structure, including those reported here for juvenimicin, tylosin and fluvirucin biosynthesis, provide new insights to facilitate TE engineering and optimization.
PubMed: 38948807
DOI: 10.1101/2024.06.20.599880 -
Biofilm Jun 2024platforms capable of mimicking the hydrodynamic conditions prevailing in natural aquatic environments have been previously validated and used to predict the fouling...
platforms capable of mimicking the hydrodynamic conditions prevailing in natural aquatic environments have been previously validated and used to predict the fouling behavior on different surfaces. Computational Fluid Dynamics (CFD) has been used to predict the shear forces occurring in these platforms. In general, these predictions are made for the initial stages of biofilm formation, where the amount of biofilm does not affect the flow behavior, enabling the estimation of the shear forces that initial adhering organisms have to withstand. In this work, we go a step further in understanding the flow behavior when a mature biofilm is present in such platforms to better understand the shear rate distribution affecting marine biofilms. Using 3D images obtained by Optical Coherence Tomography, a mesh was produced and used in CFD simulations. Biofilms of two different marine cyanobacteria were developed in agitated microtiter plates incubated at two different shaking frequencies for 7 weeks. The biofilm-flow interactions were characterized in terms of the velocity field and shear rate distribution. Results show that global hydrodynamics imposed by the different shaking frequencies affect biofilm architecture and also that this architecture affects local hydrodynamics, causing a large heterogeneity in the shear rate field. Biofilm cells located in the streamers of the biofilm are subjected to much higher shear values than those located on the bottom of the streamers and this dispersion in shear rate values increases at lower bulk fluid velocities. This heterogeneity in the shear force field may be a contributing factor for the heterogeneous behavior in metabolic activity, growth status, gene expression pattern, and antibiotic resistance often associated with nutrient availability within the biofilm.
PubMed: 38948680
DOI: 10.1016/j.bioflm.2024.100204 -
Frontiers in Veterinary Science 2024(), also known as group B (GBS), is a highly infectious pathogen. Prolonged antibiotic usage leads to significant issues of antibiotic residue and resistance....
(), also known as group B (GBS), is a highly infectious pathogen. Prolonged antibiotic usage leads to significant issues of antibiotic residue and resistance. Chelerythrine (CHE) is a naturally occurring benzophenidine alkaloid and chelerythrine chloride (CHEC) is its hydrochloride form with diverse biological and pharmacological activities. However, the antibacterial mechanism of CHEC against GBS remains unclear. Thus, this study aims to investigate the antibacterial activity of CHEC on GBS and elucidate its underlying mechanism. The antibacterial effect of CHEC on GBS was assessed using inhibitory zone, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) assays, as well as by constructing a time-kill curve. The antibacterial mechanism of CHEC was investigated through techniques such as scanning electron microscopy (SEM) and transmission electron microscopy (TEM), measurement of alkaline phosphatase (AKP) activity, determination of Na K, Ca Mg-adenosine triphosphate (ATP) activity, observation of membrane permeability, and analysis of intracellular reactive oxygen species (ROS) and mRNA expression levels of key virulence genes. The results demonstrated that the inhibition zone diameters of CHEC against GBS were 14.32 mm, 12.67 mm, and 10.76 mm at concentrations of 2 mg/mL, 1 mg/mL, and 0.5 mg/mL, respectively. The MIC and MBC values were determined as 256 μg/mL and 512 μg/mL correspondingly. In the time-kill curve, 8 × MIC, 4 × MIC and 2 × MIC CHEC could completely kill GBS within 24 h. SEM and TEM analyses revealed significant morphological alterations in GBS cells treated with CHEC including shrinkage, collapse, and leakage of cellular fluids. Furthermore, the antibacterial mechanism underlying CHEC's efficacy against GBS was attributed to its disruption of cell wall integrity as well as membrane permeability resulting in extracellular release of intracellular ATP, AKP, Na K, Ca Mg. Additionally CHEC could increase the ROS production leading to oxidative damage and downregulating mRNA expression levels of key virulence genes in GBS cells. In conclusion, CHEC holds potential as an antimicrobial agent against GBS and further investigations are necessary to elucidate additional molecular mechanisms.
PubMed: 38948675
DOI: 10.3389/fvets.2024.1408376 -
Journal of Family Medicine and Primary... May 2024Gastrointestinal infections affect many people annually. The most common bacterial agents involved in these infections are enteropathogenic bacteria and in the...
INTRODUCTION
Gastrointestinal infections affect many people annually. The most common bacterial agents involved in these infections are enteropathogenic bacteria and in the continuation of using broad-spectrum antibiotics, -associated diarrhea is involved, especially in hospitalized patients. The aim of the present study was to investigate the pattern of antibiotic resistance among enteropathogenic bacteria.
MATERIALS AND METHODS
In this cross-sectional study, 163 samples of patients with diarrhea in Dezful Ganjavian Hospital were examined. The samples were cultured in MacConkey, Hektoen enteric agar and GN broth, and cycloserine cefoxitin fructose agar media and incubated under standard conditions. In order to identify enteropathogenic bacteria, biochemical tests and serological confirmatory tests were used. Antibiotic resistance pattern of the isolates was investigated by Kirby-Bauer disk diffusion susceptibility test.
RESULTS
The frequency of pathogenic bacteria includes 41.1% of , followed by 41.1% of , 6.7% of Enteropathogenic , 5.5% of Serogroup B, and 5.5% of . The results revealed a total of 46 patients with orders regarding culture, no was isolated from the samples. The studied isolates showed the highest resistance to trimethoprim-sulfamethoxazole, and ceftriaxone (88.3%), and the most effective antibiotic in the treatment of patients was ciprofloxacin with 86% sensitivity.
CONCLUSION
Susceptibility to antibiotics was different among the isolates, which shows that the early identification of the infection agent and the selection of the correct antibiotic treatment are effective in improving the gastrointestinal infection and preventing the spread of the infection.
PubMed: 38948633
DOI: 10.4103/jfmpc.jfmpc_1594_23 -
Asian Journal of Pharmaceutical Sciences Jun 2024The intrinsic resistance of MRSA coupled with biofilm antibiotic tolerance challenges the antibiotic treatment of MRSA biofilm infections. Phytochemical-based...
The intrinsic resistance of MRSA coupled with biofilm antibiotic tolerance challenges the antibiotic treatment of MRSA biofilm infections. Phytochemical-based nanoplatform is a promising emerging approach for treatment of biofilm infection. However, their therapeutic efficacy was restricted by the low drug loading capacity and lack of selectivity. Herein, we constructed a surface charge adaptive phytochemical-based nanoparticle with high isoliquiritigenin (ISL) loading content for effective treatment of MRSA biofilm. A dimeric ISL prodrug (ISL-G2) bearing a lipase responsive ester bond was synthesized, and then encapsulated into the amphiphilic quaternized oligochitosan. The obtained ISL-G2 loaded NPs possessed positively charged surface, which allowed cis-aconityl-d-tyrosine (CA-Tyr) binding via electrostatic interaction to obtain ISL-G2@TMDCOS-Tyr NPs. The NPs maintained their negatively charged surface, thus prolonging the blood circulation time. In response to low pH in the biofilms, the fast removal of CA-Tyr led to a shift in their surface charge from negative to positive, which enhanced the accumulation and penetration of NPs in the biofilms. Sequentially, the pH-triggered release of d-tyrosine dispersed the biofilm and lipase-triggered released of ISL effectively kill biofilm MRSA. An study was performed on a MRSA biofilm infected wound model. This phytochemical-based system led to ∼2 log CFU (>99 %) reduction of biofilm MRSA as compared to untreated wound ( < 0.001) with negligible biotoxicity in mice. This phytochemical dimer nanoplatform shows great potential for long-term treatment of resistant bacterial infections.
PubMed: 38948398
DOI: 10.1016/j.ajps.2024.100923 -
MLife Jun 2024Members of the multiple antibiotic resistance regulator (MarR) protein family are ubiquitous in bacteria and play critical roles in regulating cellular metabolism and... (Review)
Review
Members of the multiple antibiotic resistance regulator (MarR) protein family are ubiquitous in bacteria and play critical roles in regulating cellular metabolism and antibiotic resistance. MarR family proteins function as repressors, and their interactions with modulators induce the expression of controlled genes. The previously characterized modulators are insufficient to explain the activities of certain MarR family proteins. However, recently, several MarR family proteins have been reported to sense sulfane sulfur, including zero-valent sulfur, persulfide (R-SSH), and polysulfide (R-SnH, ≥ 2). Sulfane sulfur is a common cellular component in bacteria whose levels vary during bacterial growth. The changing levels of sulfane sulfur affect the expression of many MarR-controlled genes. Sulfane sulfur reacts with the cysteine thiols of MarR family proteins, causing the formation of protein thiol persulfide, disulfide bonds, and other modifications. Several MarR family proteins that respond to reactive oxygen species (ROS) also sense sulfane sulfur, as both sulfane sulfur and ROS induce the formation of disulfide bonds. This review focused on MarR family proteins that sense sulfane sulfur. However, the sensing mechanisms reviewed here may also apply to other proteins that detect sulfane sulfur, which is emerging as a modulator of gene regulation.
PubMed: 38948149
DOI: 10.1002/mlf2.12109 -
MLife Jun 2024Methicillin-resistant (MRSA) is a current global public health problem due to its increasing resistance to the most recent antibiotic therapies. One critical approach...
Methicillin-resistant (MRSA) is a current global public health problem due to its increasing resistance to the most recent antibiotic therapies. One critical approach is to develop ways to revitalize existing antibiotics. Here, we show that the phytogenic compound cinnamaldehyde (CIN) and β-lactam antibiotic combinations can functionally synergize and resensitize clinical MRSA isolates to β-lactam therapy and inhibit MRSA biofilm formation. Mechanistic studies indicated that the CIN potentiation effect on β-lactams was primarily the result of inhibition of the expression by targeting the staphylococcal accessory regulator . CIN alone or in combination with β-lactams decreased gene expression and increased SarA protein phosphorylation that impaired SarA binding to the promoter element and downregulated virulence genes such as those encoding biofilm, α-hemolysin, and adhesin. Perturbation of SarA- binding thus interfered with PBP2a biosynthesis and this decreased MRSA resistance to β-lactams. Furthermore, CIN fully restored the anti-MRSA activities of β-lactam antibiotics in vivo in murine models of bacteremia and biofilm infections. Together, our results indicated that CIN acts as a β-lactam adjuvant and can be applied as an alternative therapy to combat multidrug-resistant MRSA infections.
PubMed: 38948140
DOI: 10.1002/mlf2.12121 -
Infection and Drug Resistance 2024Contact lenses (CL) have become an immensely popular means of vision correction, offering comfort to millions worldwide. However, the persistent issue of biofilm... (Review)
Review
Contact lenses (CL) have become an immensely popular means of vision correction, offering comfort to millions worldwide. However, the persistent issue of biofilm formation on lenses raises significant problems, leading to various ocular complications and discomfort. The aim of this review is to develop safer and more effective strategies for preventing and managing microbial biofilms on CL, improving the eye health and comfort of wearers. Taking these into consideration, the present study investigates the intricate mechanisms of biofilm formation, by exploring the interplay between microbial adhesion, the production of extracellular polymeric substances, and the properties of the lens material itself. Moreover, it emphasizes the diverse range of microorganisms involved, encompassing bacteria, fungi, and other opportunistic pathogens, elucidating their implications within lenses and other medical device-related infections and inflammatory responses. Going beyond the challenges posed by biofilms on CL, this work explores the advancements in biofilm detection techniques and their clinical relevance. It discusses diagnostic tools like confocal microscopy, genetic assays, and emerging technologies, assessing their capacity to identify and quantify biofilm-related infections. Finally, the paper delves into contemporary strategies and innovative approaches for managing and preventing biofilms development on CL. In Conclusion, this review provides insights for eye care practitioners, lens manufacturers, and microbiology researchers. It highlights the intricate interactions between biofilms and CL, serving as a foundation for the development of effective preventive measures and innovative solutions to enhance CL safety, comfort, and overall ocular health. Research into microbial biofilms on CL is continuously evolving, with several future directions being explored to address challenges and improve eye health outcomes as far as CL wearers are concerned.
PubMed: 38947374
DOI: 10.2147/IDR.S463779 -
Infection and Drug Resistance 2024The emergence of carbapenem-resistant (CRKP) has garnered international concern due to its significant antibiotic resistance. Notably, children exhibit distinct...
BACKGROUND
The emergence of carbapenem-resistant (CRKP) has garnered international concern due to its significant antibiotic resistance. Notably, children exhibit distinct resistance mechanisms compared to adults, necessitating a differential approach to antibiotic selection. A thorough analysis of CRKP's epidemiology and drug resistance mechanisms is essential for establishing a robust foundation for clinical anti-infection strategies and precise prevention and control measures.
METHODS
This study involved the collection of 31 non-repetitive strains from pediatric and adult patients at a tertiary hospital in China, spanning from July 2016 to July 2022, testing for resistance genes, antimicrobial susceptibility, and homology analysis.
RESULTS
Infants (0-1 year) were the largest pediatric CRKP group, with 61.3% of cases. The neonatal intensive care unit (NICU) and pediatrics were the main departments affected. Adults with CRKP had a mean age of 67 years, with the highest prevalence in neurology and emergency ICU. Antimicrobial susceptibility testing revealed that adult CRKP strains exhibited higher resistance to amikacin, ciprofloxacin, cotrimoxazole, and aztreonam compared to pediatric strains. Conversely, pediatric strains showed a higher rate of resistance to ceftazidime/avibactam. The predominant resistance genes identified were in children (58.1%) and in adults (87.1%), with over 93% of both groups testing positive for extended-spectrum beta-lactamase (ESBL) genes. Multilocus Sequence Typing (MLST) indicated ST2735 and ST11 as the predominant types in children and adults, respectively. Pulsed-field gel electrophoresis (PFGE) identified clonal transmission patterns of ST11 and ST15 across both age groups. Notably, this study reports the first instance of ST1114-type CRKP co-producing and in the NICU.
CONCLUSION
This study reveals distinct resistance mechanisms and epidemiology in CRKP from children and adults. The identified clonal transmission patterns emphasize the need for improved infection control to prevent the spread of resistant strains.
PubMed: 38947371
DOI: 10.2147/IDR.S460155