Antimicrobial Resistance Crisis Research Articles

Biofilm targeting with chitosan-based nanohydrogel containing Quercus infectoria G. Olivier extract against Streptococcus mutans: new formulations of a traditional natural product.

Biofilm formation has a crucial role in the cariogenic virulence of Streptococcus mutans, which leads to resistance to common antibacterials. The antimicrobial resistance crisis has led to increased research about traditional natural products. Quercus infectoria extract (QI extract) and nano hydrogels containing QI extract (QI-NH) and tannic acid (TA-NH) were evaluated against this pathogen. QI extract was analyzed by HPLC and the physiological characteristics of nanohydrogels were assessed by SEM, FTIR, zeta potential, DLS and determination of release kinetics and encapsulation efficiency. Determination of MIC and MBC of the material and their anti-biofilm effect was done by the microtiter method and on the extracted tooth surface. The properties of extracts and nano hydrogels in the expression of genes codifying glucosyltransferases (gtfB, gtfC and gtfD) and glucan binding protein B (gbpB) were quantified. Their toxicity was tested by the MTT method against the KB cell line. According to HPLC, 55.18% of QI extract contained TA. The encapsulation efficiency of QI-NH and TA-NH was equal to 60% and 80%, respectively. SEM and FTIR exhibited that QI extract and TA were successfully entrapped in the networks resulting from the chemical bonding of chitosan and TPP. The average size of QI-NH and TA-NH was 70.45 and 58.43nm, and their zeta potential was 6.17 ± 2.58 and 0.25 ± 0.03 mv, respectively. PDI < 0.3 of nano hydrogels indicated the favorable polydispersity of nanohydrogels. MIC of QI extract, QI-NH and TA-NH were 937.5, 30 and 10µg/ml, respectively. Also their MBIC50 was 35.1, 2.1 and 0.95µg/ml, respectively, and the extracts and nano hydrogels restrained the biofilm maturation on enamel. The pivotal genes of S. mutans in biofilm formation were significantly less expressed by treatment with QI-NH and TA-NH than others. Based on the MTT test, QI-NH had less acute toxicity than QI extract and TA-NH. IC50 of QI-NH was calculated as 775.4µg/ml, while it was equal to 3.12µg/ml for chlorhexidine as a common antibacterial agent. QI-NH, a new formulation derived from traditional anti-caries, can be a safe and efficient option to combat dental biofilm.

ACTINOBACTERIAL DIVERSITY DETERMINATION IN MALAYSIAN BRIS (BEACH RIDGES INTERSPERSED WITH SWALES) SOIL BY CULTURE-DEPENDENT APPROACH

Globally, the rapid emergence of antibiotic-resistant bacteria poses a major threat to public health, threatening the effectiveness of antibiotics. Addressing the crisis of antimicrobial resistance requires urgency in new drug discovery and the effort in finding novel compounds is dramatically evolving. However, one of the main obstacles is the rediscovery of known compounds from natural resources. To overcome this challenge, exploring underexploited location for new resource of actinomycetes, such as Beach Ridges Interspersed with Swales (BRIS) soil in Malaysia, was selected. Soil samples from Jambu Series and Rudua Series in Setiu, Terengganu were collected and analysed. A total of 87 actinomycetes strains were isolated on yeast malt agar (ISP2), Actinomyces agar and Zobell marine agar, with the highest number from the Jambu series using serial dilution and spread plate method. Antimicrobial screening revealed that 31 isolates exhibited activity against ESKAPE pathogens, with 28 and 12 isolates showing activity against Gram-positive and Gram-negative bacteria, respectively. These isolates showed strong inhibiting response towards one or more ESKAPE pathogens. This study highlights the potential of BRIS soil as an underexploited habitat for discovering actinobacterial communities which could lead to the discovery of a new potential antimicrobial producer to serve as novel drug leads.

Discovery of a family of menaquinone-targeting cyclic lipodepsipeptides for multidrug-resistant Gram-positive pathogens

Menaquinone (MK) in bacterial membrane is an attractive target for the development of novel therapeutic agents. Mining the untapped chemical diversity encoded by Gram-negative bacteria presents an opportunity to identify additional MK-binding antibiotics (MBAs). By MK-binding motif searching of bioinformatically predicted linear non-ribosomal peptides from 14,298 sequenced genomes of 45 underexplored Gram-negative bacterial genera, here we identify a novel MBA structural family, including silvmeb and pseudomeb, using structure prediction-guided chemical synthesis. Both MBAs show rapid bacteriolysis by MK-dependent membrane depolarization to achieve their potent activities against a panel of Gram-positive pathogens. Furthermore, both MBAs are proven to be effective against methicillin-resistant Staphylococcus aureus in a murine peritonitis-sepsis model. Our findings suggest that MBAs are a kind of structurally diverse and still underexplored antibacterial lipodepsipeptide class. The interrogation of underexplored bacterial taxa using synthetic bioinformatic natural product methods is an appealing strategy for discovering novel biomedically relevant agents to confront the crisis of antimicrobial resistance.

Characterization of two Friunavirus phages and their inhibitory effects on biofilms of extremely drug resistant Acinetobacter baumannii in Dakar, Senegal

BackgroundAcinetobacter baumannii is a gram-negative, opportunistic pathogen, that is responsible for a wide variety of infections and is a significant cause of hospital-acquired infections. A. baumannii is listed by the World Health Organization (WHO) as a critical priority pathogen because of its high level of antibiotic resistance and the urgent need for alternative treatment solutions. To address this challenge, bacteriophages have been used to combat bacterial infections for more than a century, and phage research has regained interest in recent years due to antimicrobial resistance (AMR). However, although the vast majority of deaths from the AMR crisis will occur in developing countries in Africa and Asia, few phages’ studies have been conducted in these regions. In this study, we present a comprehensive characterization of the bacteriophages vAbBal23 and vAbAbd25, actives against extremely drug-resistant (XDR) A. baumannii.MethodsPhages were isolated from environmental wastewaters in Dakar, Senegal. The host-range, thermal and pH stabilities, infection kinetics, one step growth assay, antibiofilm activity assay, sequencing, and genomic analysis, were performed to characterize the isolated phages.ResultsComparative genomic and phylogenetic analyses revealed that vAbBal23 and vAbAbd25 belong to the Caudoviricetes class, Autographiviridae family and Friunavirus genus. Both phages demonstrated activity against strains with capsular type KL230. They were stable over a wide pH range (pH 3 to 9) and at temperatures ranging from 25 °C to 40 °C. Additionally, the phages exhibited notable activity against both planktonic and biofilm cells of targeted extremely drug resistant A. baumannii. The results presented here indicate the lytic nature of vAbBal23 and vAbAbd25. This is further supported by the absence of genes encoding toxins, resistance genes and bacterial virulence factors, highlighting their potential for future phage applications.ConclusionPhages vAbBal23 and vAbAbd25 are promising biological agents that can infect A. baumannii, making them suitable candidates for use in phage therapies.

Combating Antimicrobial Resistance by Resensitising Bacteria to Antibiotics Using CRISPR: A Narrative Review

Background: Antimicrobial resistance (AMR) remains a formidable global health threat. Conventional strategy of developing new antibiotics is costly and unsustainable. Thus, innovative approaches for resensitising bacteria using clustered regularly inter-spaced short palindromic repeats (CRISPR) technology are sought. Aims and Objectives: The aim of this study was to review existing technologies and approaches with the aim to identify the most feasible and practical approach to solve AMR globally. Methods: It is a narrative review of CRISPR research to identify a practical medical solution for combating AMR. By considering biological, social and pharmaceutical factors, feasible solutions and their limitations were identified. Next, CRISPR solutions were examined, then filtered based on strict medical expectations. A suitable cargo type and delivery mechanism will be established. Finally, the feasibility from a practical and economic perspective is considered. Results: Among the Cas effector modules examined, Cas12k was identified as the most promising candidate. Notably, Cas12k possesses a small protein size and lacks trans-cleavage activity, enhancing its specificity for targeted bacterial strains. Additionally, its naturally inactivated endonuclease domain further contributes to its precision. In line with this investigation, DNA plasmid was identified as the optimal cargo storage, while lipid nanoparticles were identified as the optimal delivery method. Conclusion: This is the first time an end-to-end consideration to AMR was considered for CRISPR-based strategies. This review offers valuable insights into a promising solution to addressing the AMR crisis. Such knowledge is essential to guide the development of novel AMR solutions.

Global emergence of Escherichia coli with PBP3 insertions.

Escherichia coli producing metallo-β-lactamases with penicillin-binding protein 3 (PBP3) insertions have reduced susceptibility to aztreonam-avibactam and cefiderocol. Here, we analysed high-quality E. coli genomes for PBP3 insertions. E. coli genomes (n = 167 518) were retrieved from EnteroBase with CheckM2 for quality control, fastANI for species confirmation, multi-locus sequencing typing for sequence type (ST) determination and AMRFinderPlus for resistance gene identification. For PBP3 insertion analysis, we used Prokka for predicting coding sequences, BLAST+ for comparing resulted protein sequences and SnpEff for annotating variants. Among the included 159 341 genomes, PBP3 insertions with 11 variants were found in 2.01% (n = 3198). The predominant variant is a duplication of 334-337 amino acids (aa) (94.75%, n = 3030), comprising YRIN (65.92%, n = 2108) and its single-aa-variant YRIK (28.83%, n = 922), followed by a similar PYRI duplication of 333-336 (4.16%, n = 133). The less common ones are a TIPY duplication of 331-334 (n = 24) and its single-aa-variant TVPY's duplication: TVVPY (n = 1), TVPYTVPY (n = 1) and TVPYPVPY (n = 1). Rare duplications include VGDR of 106-109 (n = 3), ANALNIPL of 114-121 (n = 3), AL of 567-568 (n = 1) and TG of 584-585 (n = 1). Insertion variants were detected across 62 countries on six continents, primarily in human samples, and associated with 85 STs, concentrated in high-risk clones ST410 (29.18%, n = 1923), ST167 (23.40%, n = 1740) and ST405 (10.56%, n = 1334), with 83.32% (n = 2218) encoding metallo-β-lactamase NDM. Global spread of E. coli harbouring PBP3 insertion, often with NDM β-lactamase, high-risk ST410, ST167 and ST405 clones and various hosts, underscores the escalating antimicrobial resistance crisis and the urgency for a 'One Health' strategy.

Recent Developments in Infectious Disease Research

The field of infectious disease research is rapidly evolving and advancing, driven by the demand to address both current and emerging threats to global health. The recent COVID-19 pandemic underscored the criticality of infectious disease outbreak preparedness and response, while factors such as climate change, migration, and globalization continue to influence disease transmission dynamics. Innovative approaches to infectious disease research are more essential than ever to prevent future pandemics and the spread of vectorborne diseases. This comprehensive review article investigates some recent pertinent developments in infectious disease research, including the development of a novel serosurveillance platform, modeled after the one designed during the COVID-19 pandemic, aimed at tackling the growing global threat of arboviruses. Also investigated in this article is the repurposing and application of mRNA vaccine technology, initially developed during the pandemic, to address the complex challenge of tick-borne Lyme disease. Finally, we outline several novel and cutting-edge strategies aimed at tackling the ever-mounting crisis of antimicrobial resistance (AMR). Each of these innovations marks a significant step forward in global health security and preparedness, offering promising solutions to the challenges of a rapidly evolving world, while drawing on the lessons learned from past outbreaks and epidemics.

Rumicidins are a family of mammalian host-defense peptides plugging the 70S ribosome exit tunnel

The antimicrobial resistance crisis along with challenges of antimicrobial discovery revealed the vital necessity to develop new antibiotics. Many of the animal proline-rich antimicrobial peptides (PrAMPs) inhibit the process of bacterial translation. Genome projects allowed to identify immune-related genes encoding animal host defense peptides. Here, using genome mining approach, we discovered a family of proline-rich cathelicidins, named rumicidins. The genes encoding these peptides are widespread among ruminant mammals. Biochemical studies indicated that rumicidins effectively inhibited the elongation stage of bacterial translation. The cryo-EM structure of the Escherichia coli 70S ribosome in complex with one of the representatives of the family revealed that the binding site of rumicidins span the ribosomal A-site cleft and the nascent peptide exit tunnel interacting with its constriction point by the conservative Trp23-Phe24 dyad. Bacterial resistance to rumicidins is mediated by knockout of the SbmA transporter or modification of the MacAB-TolC efflux pump. A wide spectrum of antibacterial activity, a high efficacy in the animal infection model, and lack of adverse effects towards human cells in vitro make rumicidins promising molecular scaffolds for development of ribosome-targeting antibiotics.

Natural compound-induced downregulation of antimicrobial resistance and biofilm-linked genes in wastewater Aeromonas species.

Addressing the global antimicrobial resistance (AMR) crisis requires a multifaceted innovative approach to mitigate impacts on public health, healthcare and economic systems. In the complex evolution of AMR, biofilms and the acquisition of antimicrobial resistance genes (ARGs) play a pivotal role. Aeromonas is a major AMR player that often forms biofilm, harbors ARGs and is frequently detected in wastewater. Existing wastewater treatment plants (WWTPs) do not have the capacity to totally eliminate antimicrobial-resistant bacteria favoring the evolution of ARGs in wastewater. Besides facilitating the emergence of AMR, biofilms contribute significantly to biofouling process within the activated sludge of WWTP bioreactors. This paper presents the inhibition of biofilm formation, the expression of biofilm-linked genes and ARGs by phytochemicals andrographolide, docosanol, lanosterol, quercetin, rutin and thymohydroquinone. Aeromonas species were isolated and purified from activated sludge samples. The ARGs were detected in the isolated Aeromonas species through PCR. Aeromonas biofilms were quantified following the application of biocompounds through the microtiter plate assay. qPCR analyses of related genes were done for confirmation. Findings showed that the natural compounds inhibited the formation of biofilms and reduced the expression of genes linked to biofilm production as well as ARGs in wastewater Aeromonas. This indicates the efficacy of these compounds in targeting and controlling both ARGs and biofilm formation, highlighting their potential as innovative solutions for combating antimicrobial resistance and biofouling.

Insights Into Antimicrobial Resistance From Dental Students in the Asia–Pacific Region

BackgroundDentists, as prominent prescribers, are key stakeholders in addressing the antimicrobial resistance (AMR) crisis. Dental students’ perceptions about the topic have been underexplored in the Asia–Pacific region, a key location for the development and spread of AMR. Thus, the aim of this study was to evaluate the awareness and confidence to prescribe antimicrobials amongst dental students studying in the region. MethodsStudents from 15 dental schools in 4 countries were invited to participate in a cross-sectional online survey during 2022–2023. A previously validated and standardised 14-item instrument was utilised. ResultsIn all, 1413 responses were collected from Australia (n = 165), Sri Lanka (n = 112), Japan (n = 173), and Vietnam (n = 963). Of those, 201 were from final-year students (14.2%). On a scale from 1 to 10, awareness on AMR was placed at a mean (SEM) priority of 8.09 (0.05). With regards to target areas to address for mitigation of the AMR crisis, participants placed general public awareness at the top (mean [SEM] 8.53 [0.05]). Final-year students presented a mean (SEM) level of confidence to prescribe antibiotics of 6.01 (0.14) on a scale from 1 to 10, whilst 59.7% and 56.8% indicated feeling pressured to prescribe by patients or when lacking time, respectively. Final-year students participating in research activities assigned a higher priority to AMR compared to their peers not involved in research (mean [SEM] 8.6 [0.19] vs 7.81 [0.16]; P = .01). ConclusionsThis study highlights a need for increased awareness and confidence to prescribe amongst dental students in the Asia–Pacific region, an understudied population thus far. To mitigate this issue, the implementation (followed by assessment) of local educational and antibiotic stewardship initiatives is warranted.

Plasma activated water pre-treatment substantially enhances phage activity against Proteus mirabilis biofilms

The ongoing antimicrobial resistance crisis has incentivised research into alternative antibacterial and antibiofilm agents. One of them is plasma-activated water (PAW), which is produced by exposing water to a cold plasma discharge. This process generates a diverse array of reactive oxygen and nitrogen species (ROS/RNS) with antimicrobial properties. Another intensively studied class of alternative antimicrobials are bacteriophages, attracting attention due to their specificity and strong antibacterial activity. As combinations of different types of antimicrobials are known to often exhibit synergistic interactions, in this study we investigated the combined use of cold atmospheric-pressure plasma-activated water and the bacteriophage vB_PmiS_PM-CJR against Proteus mirabilis biofilms as a potential option for treatment of catheter-associated urinary tract infections (CAUTIs).We compared the effect of two cold plasma discharge setups for PAW production on its antimicrobial efficacy against P. mirabilis planktonic and biofilm cultures. Next, we assessed the stability of the phage vB_PmiS_PM-CJR in PAW. Finally, we tested the antimicrobial activity of the phages and PAW against biofilms, both individually and in combinations.Our findings demonstrate that the combination of PAW with phage is more effective against biofilms compared to individual treatments, being able to reduce the number of biofilm-embedded cells by approximately 4 log. We were also able to show that the order of treatment plays an important role in the anti-biofilm activity of the phage-PAW combination, as the exposure of the biofilm to PAW prior to phage administration results in a stronger effect than the reverse order.This research underlines PAW's ability to potentiate phage activity, showcasing a considerable reduction in biofilm viability and biomass. Additionally, it contributes to the growing body of evidence supporting the use of phage-based combinatorial treatments. Overall, this sequential treatment strategy demonstrates the potential of leveraging multiple approaches to address the mounting challenge of antibiotic resistance and offers a promising avenue for enhancing the efficacy of CAUTI management.

The treatment of Enterococcus faecalis related root canal biofilms with phage therapy

As one of the global concerns, antimicrobial resistance crisis increases the clinical importance of Enterococcus species. Enterococcus faecalis (E. faecalis) specifically penetrates the dentinal tubules and remains prevalent even after endodontic treatment. It has also biofilm forming character as well as the development of resistance to antibiotics. Sodium hypochlorite (NaOCl) is considered the gold standard among antibacterial washing solutions. However, due to its toxic effects, its usage limitations have led to the search for natural, non-toxic alternatives. Phages can be considered an important alternative because of their effects on specific bacteria. The objective of this study was to compare the effect of isolated active vB_Ef1 phage on the removal of E. faecalis biofilm in dentin, together and separately with the chemical irrigation solution NaOCl. As a result of study, the optimal NaOCl solution concentration to be applied with vB_Ef1 phage is 0.5 %, and the use of solution at this value reduces the biofilm mass by 84 %, reaching the highest biofilm mass reduction value. It was found that the combination of phage and NaOCl at appropriate concentrations had the strongest biofilm disruption effect.

ENOVAT: the European Network for Optimization of Veterinary Antimicrobial Treatment

The global antimicrobial resistance crisis has been the driver of several international strategies on antimicrobial stewardship. For their implementation at the field level, the veterinary sector encounters several specific challenges and in particular: (i) a shortage of experts in key disciplines related to antimicrobial stewardship, (ii) a lack of evidence-based antimicrobial treatment guidelines, and (iii) inferior diagnostic tests available compared to human medicine. The present white paper describes how the COST Action ENOVAT (the European Network for Optimization of Veterinary Antimicrobial Treatment, CA18217), comprising 332 persons from 51 countries, worked towards solutions to these challenges. Initially, surveys were conducted to explore the present state in Europe in terms of existing antimicrobial use guidelines and microbiology practices performed. Concurrently, various research activities were launched to optimize diagnostics, including development of epidemiological cut-offs, clinical breakpoints and matrix-assisted laser desorption ionization time of flight mass spectrometry interpretive criteria. Also, guidelines drafting groups working towards evidence-based antimicrobial treatment guidelines for six conditions in food-producing and companion animals were established. The processes and outcomes, also in terms of capacity building, are summarized in this white paper where emphasis is placed on sustainability of the activities. Although several ENOVAT initiatives and spin-off projects will continue beyond the Action, we recommend that a new European veterinary research agenda is launched focusing on research and funding leading to long-term impacts on veterinary antimicrobial use.

BWC0977, a broad-spectrum antibacterial clinical candidate to treat multidrug resistant infections

The global crisis of antimicrobial resistance (AMR) necessitates the development of broad-spectrum antibacterial drugs effective against multi-drug resistant (MDR) pathogens. BWC0977, a Novel Bacterial Topoisomerase Inhibitor (NBTI) selectively inhibits bacterial DNA replication via inhibition of DNA gyrase and topoisomerase IV. BWC0977 exhibited a minimum inhibitory concentration (MIC90) of 0.03–2 µg/mL against a global panel of MDR Gram-negative bacteria including Enterobacterales and non-fermenters, Gram-positive bacteria, anaerobes and biothreat pathogens. BWC0977 retains activity against isolates resistant to fluoroquinolones (FQs), carbapenems and colistin and demonstrates efficacy against multiple pathogens in two rodent species with significantly higher drug levels in the epithelial lining fluid of infected lungs. In healthy volunteers, single-ascending doses of BWC0977 administered intravenously (https://clinicaltrials.gov/study/NCT05088421) was found to be safe, well tolerated (primary endpoint) and achieved dose-proportional exposures (secondary endpoint) consistent with modelled data from preclinical studies. Here, we show that BWC0977 has the potential to treat a range of critical-care infections including MDR bacterial pneumonias.

Phenotypic bacterial epidemiology and antimicrobial resistance profiles in neonatal sepsis at Jimma medical center, Ethiopia: Insights from prospective study.

Epidemiological profiles and the rundown crisis of antimicrobial resistance from bacterial isolates in neonatal sepsis compel regular surveillance to enhance data-driven decision-making. Accordingly, this study aimed to assess the phenotypic epidemiology and antimicrobial resistance profiles of bacteria isolated from clinically suspected neonatal sepsis in Ethiopia. A total of 342 neonates suspected of clinical sepsis were randomly included in a prospective observational study conducted at the neonatal intensive care unit (NICU) of Jimma medical center (JMC) from May 2022 to July 2023. Blood samples were collected from each neonate and subjected to a culture test for identification of bacterial isolates and their antibiotic resistance profiles following the standardized guidelines. The laboratory results, along with relevant clinical data, were recorded using WHONET and analyzed using STATA software. Out of the 342 blood samples that were analyzed, 138 samples (40.4%, 95% CI: 35.1-45.6, P<0.01) exhibited proven bacterial infection. The infection rates were notably higher in males with 85/138 (61.6%, 95% CI: 53.4-69.8, P<0.01) and neonates aged 0-3 days with 81/138 (58.7%, 95% CI: 50.5-66.9, P<0.01). The majority of the infections were attributed to Gram-negative bacteria, accounting for 101/138(73.2%, 95% CI: 65.6-80.7) cases, with 69/101(68.3%, 95% CI: 63.8-72.8) cases involving ESBL-producing strains, while Gram-positive bacteria were responsible for 26.8% (95% CI: 19.3-34.4) of the infections. The predominant isolates included Klebsiella pneumoniae (37.7%, 95% CI: 29.6-45.8), Coagulase-negative Staphylococci (CoNs) (20.3%, 95% CI: 13.6-27.0), and Acinetobacter species (11.6%, 95% CI: 6.0-17.1). Of the total cases, 43/72 (59.7%, 95% CI: 48.4-71.1, P<0.01) resulted in mortality, with 28/72 (38.9%, 95% CI: 27.70-50.1, P<0.03) deaths linked to Extended-Spectrum Beta-Lactamase (ESBL)-producing strains. Klebsiella pneumoniae displayed high resistance rates to trimethoprim-sulfamethoxazole (100%), ceftriaxone (100%), cefotaxime (98.1%), ceftazidime (90.4%), and gentamicin (84.6%). Acinetobacter species showed resistance to ampicillin (100%), cefotaxime (100%), trimethoprim-sulfamethoxazole (75%), ceftazidime (68.8%), chloramphenicol (68.8%), and ceftriaxone (68.8%). Likewise, CoNs displayed resistance to ampicillin (100%), penicillin (100%), cefotaxime (86.0%), gentamicin (57.2%), and oxacillin (32.2%). Multidrug resistance was observed in 88.4% (95% CI: 81.8-93.0) of isolates, with ESBL-producers significantly contributing (49.3%, 95% CI: 45.1-53.5). Furthermore, 23.0% (95% CI: 15.8-31.6) exhibited a prevalent resistance pattern to seven distinct antibiotic classes. The prevalence and mortality rates of neonatal sepsis were significantly high at JMC, with a notable surge in antibiotic and multidrug resistance among bacterial strains isolated from infected neonates, specifically ESBL-producers. These resistant strains have a significant impact on infection rates and resistance profiles, highlighting the requisite for enhanced diagnostic and antimicrobial stewardship, stringent infection control, and further molecular characterization of isolates to enhance neonatal survival.

Free Energy Analysis of Peptide-Induced Pore Formation in Lipid Membranes by Bridging Atomistic and Coarse-Grained Simulations.

Antimicrobial peptides (AMPs) are attractive materials for combating the antimicrobial resistance crisis because they can kill target microbes by directly disrupting cell membranes. Although thousands of AMPs have been discovered, their molecular mechanisms of action are still poorly understood. One broad mechanism for membrane disruption is the formation of membrane-spanning hydrophilic pores which can be stabilized by AMPs. In this study, we use molecular dynamics simulations to investigate the thermodynamics of pore formation in model single-component lipid membranes in the presence of one of three AMPs: aurein 1.2, melittin and magainin 2. To overcome the general challenge of modeling long time scale membrane-related behaviors, including AMP binding, clustering, and pore formation, we develop a generalizable methodology for sampling AMP-induced pore formation. This approach involves the long equilibration of peptides around a pore created with a nucleation collective variable by performing coarse-grained simulations, then backmapping equilibrated AMP-membrane configurations to all-atom resolution. We then perform all-atom simulations to resolve free energy profiles for pore formation while accurately modeling the interplay of lipid-peptide-solvent interactions that dictate pore formation free energies. Using this approach, we quantify free energy barriers for pore formation without direct biases on peptides or whole lipids, allowing us to investigate mechanisms of pore formation for these 3 AMPs that are a consequence of unbiased peptide diffusion and clustering. Further analysis of simulation trajectories then relates variations in pore lining by AMPs, AMP-induced lipid disruptions, and salt bridges between AMPs to the observed pore formation free energies and corresponding mechanisms. This methodology and mechanistic analysis have the potential to generalize beyond the AMPs in this study to improve our understanding of pore formation by AMPs and related antimicrobial materials.

Small molecule produced by Photorhabdus interferes with ubiquinone biosynthesis in Gram-negative bacteria.

The spread of resistant pathogens has led to the antimicrobial resistance crisis, and the need for new compounds acting against Gram-negative pathogens is especially acute. From a screen of Photorhabdus symbionts of nematodes, we identified 3,6-dihydroxy-1,2-benzisoxazole (DHB) that acts against a range of Gram-negative bacteria, including Escherichia coli, Enterobacter cloacae, Klebsiella pneumoniae, and Acinetobacter baumannii. DHB had previously been isolated from other bacterial species, but its mechanism of action remained unknown. We show that DHB is unique among antimicrobials, with dual action as an inhibitor of an important enzyme, UbiA, in the biosynthesis pathway of ubiquinone and as a prodrug. DHB is a mimic of the natural substrate, and UbiA modifies it into a toxic product, contributing to the antimicrobial action of this unusual antibiotic. We also uncover the mechanism of DHB selectivity, which depends on a particular fold of the UbiA enzyme.

ABD-3, the confluence of powerful antibacterial modalities: ABDs delivering and expressing lss, the gene encoding lysostaphin.

In response to the antimicrobial resistance crisis, we have developed a powerful and versatile therapeutic platform, the Antibacterial Drone (ABD) system. The ABD consists of a highly mobile staphylococcal pathogenicity island re-purposed to deliver genes encoding antibacterial proteins. The chromosomally located island is induced by a co-resident helper phage, packaged in phage-like particles, and released in very high numbers upon phage-induced lysis. ABD particles specifically adsorb to bacteria causing an infection and deliver their DNA to these bacteria, where the bactericidal cargo genes are expressed, kill the bacteria, and cure the infection. Here, we report a major advance of the system, incorporation of the gene encoding a secreted, bactericidal, species-specific lytic enzyme, lysostsphin. This ABD not only kills the bacterium that has been attacked by the ABD, but also any surrounding bacteria that are sensitive to the lytic enzyme which is released by secretion and by lysis of the doomed cell. So while the killing field is thus expanded, there are no civilian casualties (bacteria that are insensitive to the ABD and its cargo protein(s) are not inadvertently killed). Without amplifying the number of ABD particles (which are not re-packaged), the expression and release of the cargo gene's product dramatically extend the effective reach of the ABD. A cargo gene that encodes a secreted bactericidal protein also enables the treatment of a mixed bacterial infection in which one of the infecting organisms is insensitive to the ABD delivery system but is sensitive to the ABD's secreted cargo protein.

In treacherous waters: detection of colistin-resistant bacteria in water and plastic litter from a recreational estuary.

Colistin resistance poses a major therapeutic challenge and resistant strains have now been reported worldwide. However, the occurrence of such bacteria in aquatic environments is considerably less understood. This study aimed to isolate and characterize colistin-resistant strains from water and plastic litter collected in an urban recreational estuary. Altogether, 64 strains with acquired colistin resistance were identified, mainly Acinetobacter spp. and Enterobacter spp. From these, 40.6% were positive for at least one mcr variant (1-9), 26.5% harbored, extended-spectrum beta-lactamases, 23.4% harbored, sulfonamide resistance genes, and 9.3% harbored, quinolone resistance genes. merA, encoding mercury resistance, was detected in 10.5% of these strains, most of which were also strong biofilm producers. The minimum inhibitory concentration toward colistin was determined for the mcr-positive strains and ranged from 2 to≥512µg ml-1. Our findings suggest that Gram-negative bacteria highly resistant to a last-resort antimicrobial can be found in recreational waters and plastic litter, thereby evidencing the urgency of the One Health approach to mitigate the antimicrobial resistance crisis.

In Silico Prediction of Antibacterial Activity of Quinolone Derivatives

AbstractThe rising antimicrobial resistance crisis has diminished the effectiveness of traditional antibiotics against pathogenic bacteria. This study addresses this urgent challenge by exploring the antibacterial potential of novel quinolone derivatives (1–33). Using computational in silico modeling to simulate biological interactions, we aimed to identify candidates with potent antibacterial activity. A total of 33 quinolone derivatives were assessed for their physicochemical properties and effectiveness against a range of clinically relevant pathogens, including methicillin‐resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae, Streptococcus pneumoniae, and Enterococcus faecalis. Molecular docking studies identified compounds 28, 29, 32, and 33 as having notable binding affinities, particularly against MRSA. Further molecular dynamics simulations of compound 29 confirmed its favorable stability and potential for disrupting MRSA, reinforcing the docking results and showing strong alignment with in vitro findings. These findings position compound 29 as a promising lead for developing alternative MRSA therapies and underscore the need for further in vivo studies to evaluate its therapeutic potential.