Search Results (111)

Antimicrobial resistance (AMR) and virulence have traditionally been viewed as competing traits in bacterial evolution due to fitness costs. However, Streptococcus pneumoniae has emerged as a paradigm of successful coevolution, with multidrug-resistant clones simultaneously maintaining or enhancing pathogenic potential. This review examines the molecular mechanisms, epidemiological patterns, and clinical consequences of the convergence between AMR and virulence in Streptococcus pneumoniae. Resistance to β-lactams is driven by mosaic penicillin-binding protein genes (pbp1a, pbp2b, pbp2x), while macrolide resistance is mediated primarily by the erm(B) gene (MLS phenotype) and mef(A/E)–msr(D) genes encoding an efflux system. These determinants are frequently co-localized on integrative and conjugative elements, ICEs, (e.g., Tn916 family) within successful clonal complexes such as CC271/320 and lineages including ST320 and GPSC10. Contrary to the classical fitness cost hypothesis, compensatory epistasis, capsular recombination, metabolic adaptations, and intra-serotype phenotypic variation enable certain clones to combine high-level resistance to β-lactams, macrolides, and tetracyclines with enhanced colonization, biofilm formation, immune evasion, and invasive capacity. Post-pneumococcal conjugate vaccine (PCV) surveillance reveals the persistence and expansion of these high-risk lineages, contributing to treatment-refractory invasive pneumococcal disease (IPD), increased morbidity, and mortality. Although PCVs have reduced vaccine-type resistant strains in some settings, serotype replacement and emerging metabolic genotypes continue to drive adaptation. This review highlights the need for integrated genomic surveillance, novel therapeutics (e.g., omadacycline, lefamulin, endolysins), monoclonal antibodies, and next-generation vaccines targeting both resistance and conserved virulence determinants. A multifaceted strategy combining antimicrobial stewardship, strengthened surveillance, and innovative interventions is essential to curb the evolving threat of resistant and virulent S. pneumoniae. Full article

Staphylococcus aureus remains to be one of the leading causes of global mortality. The most common class of antibiotics used to treat S. aureus infections are next-generation β-lactams (NGBs), as they are highly efficacious and have low adverse effects. NGB resistance in S. aureus is classically attributed to penicillin-binding protein-2a (PBP2a), but previous studies from our group have also implicated an altered expression of penicillin-binding protein-4 (PBP4) with NGB resistance. PBP4 is the sole low-molecular-mass (LMM) PBP present in S. aureus; it is also the only known LMM PBP with transpeptidase activity, giving it the unique ability to bring about peptidoglycan cross-linking. In this article, we review some of the recent findings from our group, which reveal that mutations associated with PBP4 lead to altered protein expression and NGB resistance in both methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) backgrounds. We discuss the clinical relevance of PBP4-associated mutations, particularly in methicillin-resistant lacking mec (MRLM) isolates, as well as the synergistic effect of altered PBP4 and GdpP functions. Finally, this review summarizes the potential role played by PBP4 in S. aureus virulence. Together, we highlight the increasing relevance of PBP4 as a mediator of NGB resistance and discuss its potential as an important factor during infection diagnosis and therapy. Full article

Tebipenem is a new carbapenem antibiotic that binds to penicillin-binding proteins (PBPs). Given the need for effective antibiotics against multidrug-resistant (MDR) bacteria, this review evaluated the in vitro antimicrobial activity of tebipenem against Gram-negative and Gram-positive bacteria, focusing on uropathogens. Five resources (Google Scholar, Web of Science, Embase, Scopus, and PubMed) were used to identify relevant articles. Of the 1322 articles identified, 9 relevant studies were included, which evaluated 12,501 Gram-negative and 122 Gram-positive pathogens. All nine studies (100%) assessed the activity of tebipenem against Escherichia coli, with an MIC₉₀ value range of 0.015–>4 mg/L. Seven studies (77.8%) included Klebsiella pneumoniae, with an MIC₉₀ value range of 0.015–0.5 mg/L. Six studies (66.7%) reported data on Proteus mirabilis, with an MIC₉₀ value range of ≤0.125–0.5 mg/L. Two studies (22.2%) evaluated the activity of tebipenem against Enterococcus faecalis, with MIC₉₀ of 1 mg/L among vancomycin-susceptible isolates and 32 mg/L in isolates with not-reported mechanisms of resistance. Two studies (22.2%) evaluated the activity of tebipenem against Enterococcus faecium, with MIC₉₀ of >4 mg/L among both vancomycin-susceptible and vancomycin-resistant isolates and MIC₉₀ of 128 mg/L among isolates with no resistance mechanism reported. Tebipenem demonstrated good activity against Enterobacterales, such as E. coli and K. pneumoniae. The antimicrobial agent exhibited higher MICs and a higher proportion of resistance among P. mirabilis isolates. Tebipenem could be effective for outpatient treatment of infections caused by MDR Gram-negative pathogens. However, given its potential to exert selective pressure for the development of antimicrobial resistance, it should be considered for patients with cUTIs when none of the first-line treatment options demonstrate in vitro antimicrobial activity. Full article

The venom of the Asian hornet (Vespa affinis) comprises a complex mixture of biologically active substances, including various enzymes such as phospholipase A and hyaluronidase; amines such as histamine, serotonin, and catecholamines; peptides such as mastoparan and vespakinin; and other components including acetylcholine and antigen 5. This complexity reflects the highly evolved nature of V. affinis as a venomous insect. The composition of animal venoms often exhibits a certain degree of variability, making the study of biogenic amines particularly intriguing. The objective of this research was to confirm and identify the presence of tetradecylamine in the venom of Vespa affinis using the scientific computational analysis software MetFrag. In addition, the study aimed to construct the biosynthetic pathway of this compound and to predict its potential biological roles. The predicted biosynthetic route of tetradecylamine suggested its possible involvement in antibacterial activity. Antibacterial assays were performed against four bacterial strains Escherichia coli, Staphylococcus aureus, Bacillus cereus, and Klebsiella pneumoniae. The results revealed that tetradecylamine exhibited notable inhibitory effects, with minimum inhibitory concentration (MIC) values of 2, 4, 8, and 4 µg/mL, and minimum bactericidal concentration (MBC) values of 2, 4, 8, and 4 µg/mL, respectively. Furthermore, molecular docking studies were conducted using penicillin-binding protein 2x (PBP2x, PDB ID: 5OIZ) as the target protein. Among eight tested ligands, streptomycin exhibited the highest binding affinity with a docking score of 64.76. In contrast, biogenic amines such as 2-phenylethylamine and tetradecylamine showed docking scores of 33.74 and 48.2, respectively. In the MurA protein (PDB ID: 3VCY), the biogenic amine ligand tetradecylamine exhibited a binding affinity comparable to that of certain reference drugs. Specifically, tetradecylamine achieved a GOLD score of 52.58, whereas ampicillin showed a higher score of 61.53. Notably, tetradecylamine demonstrated a higher binding affinity to the target protein compared with certain conventional antibiotics such as doxycycline and gentamycin. Full article

Background/Objectives: Granulicatella adiacens infective endocarditis is conventionally managed with penicillin, ampicillin, or ceftriaxone in combination with gentamicin, although double beta-lactam regiments have been proposed a safer alternative to reduce aminoglycoside-associated nephrotoxicity. To date, the High-Molecular-Mass Penicillin-Binding Proteins (HMM-PBPs) of G. adiacens and their affinities for beta-lactam antibiotics have not been previously characterized. This study investigated the HMM-PBP profile of G. adiacens, with particular interest on sequence alterations and beta-lactam binding properties, both as single agents and in combination. Methods: Beta-lactam activity, synergistic interactions and PBP binding affinities were evaluated in a clinical isolate (IS 48) and compared with those in the reference strain ATCC 49175. Binding of PBPs to ampicillin, ceftriaxone, and ceftobiprole, alone or in combination, was investigated by Bocillin-FL labeling. PBP homology and conserved active-sites motifs were assessed by sequence alignment, and pbp gene mutations were identified by whole-genome sequencing. Results: The clinical isolate was non-susceptible to ampicillin, resistant to ceftriaxone and exhibited higher minimum inhibitory concentrations (MICs) for ceftobiprole relative to the fully susceptible ATCC reference strain. Five HMM PBPs with high enterococcal homology, were identified. In the IS 48 isolate, the class A PBP showed distinct amino acid substitutions in proximity to the catalytic centers. Despite these alterations, PBP1A and PBP2A were strongly inhibited by the tested beta-lactams, whereas PBP2 and PBP2B demonstrated low acylation rates. Combination of ampicillin with either ceftobiprole or ceftriaxone resulted in enhanced acylation of the three bifunctional HMM PBPs compared with monotreatment. IC₅₀ values were consistently higher for the IS 48 clinical isolate, suggesting decreased target availability and/or reduced beta-lactam affinity under clinical conditions. Conclusions: The resistance phenotype of G. adiacens clinical isolate appears to be primarily associated with altered PBP beta-lactam interactions. Nonetheless, beta-lactam combination regimes remain effective by achieving substantial inhibition of key HMM-PBPs involved in peptidoglycan synthesis, thereby supporting the rationale for dual beta-lactam therapy in this setting. Full article

The emergence of multidrug-resistant (MDR) bacterial pathogens has heightened the urgency for novel antibacterial agents. The bacterial cell wall usually comprises peptidoglycan, which presents a prime target for antibacterial drug development due to its indispensable role in maintaining cellular integrity. Conventional antibiotics such as β-lactams and glycopeptides hinder peptidoglycan synthesis through competitive binding of penicillin-binding proteins (PBPs) and sequestration of lipid-linked precursor molecules. Nevertheless, prevalent resistance mechanisms including target modification, β-lactamase hydrolysis, and multi-drug efflux pumps have limited their clinical utility. This comprehensive analysis explicates the molecular machinery underlying bacterial cell wall assembly, evaluates both explored and unexplored enzymatic nodes within this pathway, and highlights the transformative impact of high-resolution structural elucidation in accelerating structure-guided drug discovery. Novel targets such as GlmS, GlmM, GlmU, Mur ligases, D,L-transpeptidases are assessed for their inclusiveness for the discovery of next-generation antibiotics. Additionally, cell wall inhibitors are also examined for their mechanisms of action and evolutionary constraints on MDR development. High-resolution crystallographic data provide valuable insights into molecular blueprints for structure-guided optimization of pharmacophores, enhancing binding affinity and circumventing resistance determinants. This review proposes a roadmap for future innovation, advocating for the convergence of computational biology platforms, machine learning-driven compound screening, and nanoscale delivery systems to improve therapeutic efficacy and pharmacokinetics. The synergy of structural insights and cutting-edge technologies offers a multidisciplinary framework for revitalizing the antibacterial arsenal and combating MDR infections efficiently. Full article

Streptococcus agalactiae are bacteria that can cause a range of infections, some of them life-threatening. Currently, antimicrobial resistance has become a global problem that puts public health at risk. Despite the widespread use of β-lactams, penicillin remains the first-line antimicrobial for the treatment of invasive S. agalactiae infections. However, reduced susceptibility and resistance to penicillin have been identified in several countries. Penicillin-binding proteins, mainly PBP2X, have been associated with reduced susceptibility to β-lactams in streptococci. The aim of this review is to summarize currently published data on penicillin-binding proteins in S. agalactiae and penicillin susceptibility, highlighting the increasing number of strains with reduced susceptibility and resistance to penicillin commonly used in the prophylaxis and treatment of invasive infections by this pathogen. Data on invasive S. agalactiae strains with high levels of penicillin resistance have been found in Japan, the United States, Canada, and Africa. The data on antibiotic resistance are alarming and require increased monitoring of strains with reduced penicillin susceptibility, as well as preventive control measures to avoid the spread of resistant mutant strains. Full article

Background: Chromobacterium violaceum is an emerging multidrug-resistant (MDR) Gram-negative bacterium associated with severe septicemia, abscess formation, and high mortality, particularly in immunocompromised individuals. Increasing antimicrobial resistance and the absence of approved vaccines underscore the urgent need for alternative preventive strategies. Traditional vaccine approaches are often inadequate against genetically diverse MDR pathogens, prompting the use of computational immunology and reverse vaccinology for vaccine design. Objectives: This study aimed to design and characterize a novel multi-epitope subunit vaccine (MEV) candidate against C. violaceum using a comprehensive pangenome-guided subtractive proteomics and immunoinformatics pipeline to identify conserved antigenic targets capable of eliciting strong immune responses. Methods: Comparative genomic analysis across eight C. violaceum strains identified 3144 core genes. Subtractive proteomics filtering yielded two essential, non-homologous, surface-accessible, and antigenic proteins—penicillin-binding protein 1A (Pbp1A) and organic solvent tolerance protein (LptD)—as vaccine targets. Cytotoxic T-lymphocyte (CTL), helper T-lymphocyte (HTL), and B-cell epitopes were predicted and integrated into a 272-amino-acid MEV construct adjuvanted with human β-defensin-4A using optimal linkers. The construct was evaluated through structural modeling, molecular docking with TLR4, molecular dynamics simulation, immune simulation, and in silico cloning into the pET-28a(+) vector. Results: The MEV construct exhibited strong antigenicity, non-allergenicity, and non-toxicity, with stable tertiary structure and favorable physicochemical properties. Docking and dynamics simulations demonstrated high binding affinity and stability with TLR4 (ΔG = −16.2 kcal/mol), while immune simulations predicted durable humoral and cellular immune responses with broad population coverage (≈89%). Codon optimization confirmed high expression potential in E. coli K12. Conclusions: The pangenome-guided immunoinformatics approach enabled the identification of conserved antigenic proteins and rational design of a promising multi-epitope vaccine candidate against MDR C. violaceum. The construct exhibits favorable immunogenic and structural features, supporting its potential for experimental validation and future development as a preventive immunotherapeutic against emerging MDR pathogens. Full article

LD-Transpeptidases (LDTs) are a widely conserved class of peptidoglycan (PG) crosslinking enzymes in bacteria. They are sometimes overlooked as they often act secondary to penicillin binding proteins (PBPs) under standard conditions. However, LDTs are essential in key pathogens such as Clostridioides difficile and are responsible for β-lactam resistance in Mycobacterium tuberculosis and Enterococcus faecium due their low affinity for penicillins and cephalosporins, allowing them to form LD-crosslinks when DD-crosslinking PBPs are inactivated. This role makes LDTs a promising target when developing new treatments for these pathogens. LDTs can perform different enzymatic reactions. Most commonly they reinforce the PG with 3,3-LD-crosslinks or, in a few cases, 1,3-LD-crosslinks, during stationary phase or stress responses. Some LDTs also incorporate endogenous and exogenous non-canonical D-amino acids into the PG. In many Gram-negative bacteria, specialised LDTs tether lipoproteins or outer membrane proteins (OMPs) to the PG to maintain cell envelope integrity; in some cases this regulates virulence factors. Specialised LDTs have also been implied to have roles in polar growth, toxin secretion, and symbiotic colonisation. Recent discoveries include novel subgroups of the major YkuD family and the identification of the VanW family; this has opened new research directions surrounding LDTs. We aim to understand LDTs and their roles to expand our knowledge of PG synthesis and modification and how these enzymes can be targeted for antibiotic treatment. Full article

The synthesis of novel pyrazole derivatives (SPDs) and their evaluation for antibacterial potential against Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa), Staphylococcus aureus (S. aureus), and Streptococcus pneumoniae (S. pneumoniae) was developed herein. These compounds were obtained via a microwave-assisted eco-friendly multicomponent reaction (MCR) and were characterized for structural confirmation using ¹H NMR, ¹³C NMR, 2D experiments, TOF-MS, and FTIR spectrometry. Antibacterial activity, as measured by minimum inhibitory concentrations (MICs) of SPDs, ranged between 0.212 and 2.50 mg/mL against S. aureus, S. pneumoniae, P. aeruginosa, and E. coli. Compound 4e was the most potent against S. pneumoniae, with an MIC value of 0.0156 mg/mL compared with Amoxicillin’s MIC value of 0.0306 mg/mL. Thus, compound 4e was observed as a potential lead candidate against S. pneumoniae. Further corroboration by molecular docking at the active site of the key penicillin-binding protein (PBP) revealed that the most potent compounds against each organism showed comparable docking scores to those of amoxicillin. In addition, a pharmacokinetics study showed that synthesized SPDs were predicted to be orally bioavailable and non-inhibitors of cytochrome 3A4 and belong to drug classes 4 and 6. Hence, they were suitable for drug development and warrant further studies such as in vitro assays, in silico modeling, DFT studies, and machine learning for drug design. Full article

Peptic ulcer disease, affecting almost 20% of the worldwide population, depicts an urgent need for effective treatment due to the limited therapeutic options available and the side effects associated with current drugs. The disease is often linked with Helicobacter pylori infection and NSAID usage, both of which compromise the mucosal lining of the stomach. There is growing evidence that dietary polyphenols can contribute to the prevention and management of various chronic diseases, including cancer and gastrointestinal disorders. Among these, green tea has garnered significant attention due to its rich polyphenolic content and associated health benefits. The abundance of green tea polyphenols (GTPs) exhibits chemoprotective, antimicrobial, and antioxidant properties. This study explores a set of 65 GTPs against penicillin-binding proteins (PBPs) as a molecular target to prevent peptic ulceration. Our molecular docking analysis revealed that the polyphenol ‘Epigallocatechin gallate’ (EGCG) exhibited effective binding affinity towards PBPs (PDB code: 1QMF), with a docking score of (−17.23 kcal/mol), followed then by Theaflavin-3-gallate (−16.57 kcal/mol) and Epigallocatechin (−15.91 kcal/mol). In silico ADME profiling indicated favorable pharmacokinetics for EGCG, including no AMES toxicity, low hERG inhibition, and good intestinal absorption. Our study highlights EGCG as a potential inhibitor of H. pylori, providing a promising natural therapeutic candidate for the management of peptic ulcer disease. Full article

Background: Helicobacter pylori is associated with a wide range of gastroduodenal diseases, including chronic gastritis, peptic ulcer disease, and gastric cancer. Eradication efforts are challenged by increasing antimicrobial resistance rates, particularly in Southeast Asia. We sequenced the whole genomes of clinical H. pylori isolates from Southern Thailand to elucidate their resistance profiles, virulence determinants, and evolutionary relationships. Methods: Three clinical H. pylori isolates (004, 117, and 189) were subjected to whole-genome sequencing, phenotypic antimicrobial susceptibility testing, and comparative genomic analyses. Results: All strains exhibited high-level resistance to metronidazole. Additionally, H. pylori 117 was resistant to both amoxicillin and levofloxacin, classifying it as multidrug-resistant. Genomic analysis revealed mutations in rdxA, frxA, and rpoB, as well as in penicillin-binding protein genes (pbp2 and pbp3), supporting the phenotypic findings. While all isolates harboured clarithromycin resistance mutations (A2142G and A2143G in the 23S rRNA gene), they were phenotypically susceptible, highlighting a potential discordance that requires further investigation. Virulence gene profiling identified 115–118 conserved genes per strain, including cagA, vacA, oipA, babA, and flagellar, urease, and lipopolysaccharide biosynthesis genes. Phylogenetic analysis using core-genome single-nucleotide polymorphisms demonstrated that these strains formed a distinct Southern Thai monophyletic clade, suggesting localised clonal expansion driven by regional selective pressures. Conclusions: Region-specific surveillance strategies and treatment guidelines are urgently needed in Thailand. The combination of high-risk virulence genes and rising antimicrobial resistance in H. pylori strains necessitates tailored therapeutic approaches, the integration of genomic surveillance into clinical diagnostics, and expanded studies linking genotype to clinical outcomes in diverse populations. Full article

Penicillin-binding proteins (PBPs) are essential enzymes involved in bacterial cell wall biosynthesis and represent the primary targets of β-lactam antibiotics. However, the efficacy of these agents is threatened by β-lactamase production and PBP alterations, prompting the search for alternative strategies. In this context, boronic acids, long established as potent inhibitors of serine β-lactamases (SBLs), have been proposed as scaffolds for PBP inhibition based on the shared structural and mechanistic features of these enzyme families. This perspective provides a literature-based survey with structural analysis to evaluate emerging evidence on the potential role of boronic acids as PBP-targeting agents, with a particular focus on peptidomimetic boronic acids, repurposed β-lactamase inhibitors, and novel scaffold architectures. While early work showed limited activity against low-molecular-mass PBPs, more recent compounds, particularly certain bicyclic boronates, have demonstrated potent binding and, in some cases, antibacterial activity. Structural analyses reveal diverse binding modes and underscore the role of conformational dynamics in modulating affinity. Despite these advances, significant challenges remain, including target selectivity, membrane permeability, and species-specific differences. Nevertheless, the direct inhibition of PBPs by boronic acids, while still in early development, may offer a viable complement or alternative to β-lactam therapy, warranting further exploration through structure-guided design and comprehensive biological evaluation. Here, we analyze the potential of boronic acid inhibitors (BAIs) to target PBP enzymes, considering their promise as non-β-lactam antimicrobial agents with possible clinical relevance. Full article

Background: Antibacterial resistance (ABR) poses a major challenge to global health, with methicillin-resistant Staphylococcus aureus (MRSA) being one of the prominent multidrug-resistant strains. MRSA has developed resistance through the expression of Penicillin-Binding Protein 2a (PBP2a), a key transpeptidase enzyme involved in bacterial cell wall biosynthesis. Objectives: The objective was to design and characterize a novel small-molecule inhibitor targeting PBP2a as a strategy to combat MRSA. Methods: We synthesized a new indole triazole conjugate (ITC) using eco-friendly and click chemistry approaches. In vitro antibacterial tests were performed against a panel of strains to evaluate the ITC antibacterial potential. Further, a series of in silico evaluations like molecular docking, MD simulations, free energy landscape (FEL), and principal component analysis (PCA) using the crystal structure of PBP2a (PDB ID: 4CJN), in order to predict the mechanism of action, binding mode, structural stability, and energetic profile of the 4CJN-ITC complex. Results: The compound ITC exhibited noteworthy antibacterial activity, which effectively inhibited the selected strains. Binding score and energy calculations demonstrated high affinity of ITC for the allosteric site of PBP2a and significant interactions responsible for complex stability during MD simulations. Further, FEL and PCA provided insights into the conformational behavior of ITC. These results gave the structural clues for the inhibitory action of ITC on the PBP2a. Conclusions: The integrated in vitro and in silico studies corroborate the potential of ITC as a promising developmental lead targeting PBP2a in MRSA. This study demonstrates the potential usage of rational drug design approaches in addressing therapeutic needs related to ABR. Full article

The repurposing potential of Efavirenz (EFV), a clinically established non-nucleoside reverse transcriptase inhibitor, was comprehensively evaluated for its in vitro antibacterial effect either alone or in combination with other antibacterial agents on several Gram-positive clinical strains showing different antibiotic resistance profiles. The binding potential assessed by an in silico study included Penicillin-binding proteins (PBPs) and WalK membrane kinase. Despite the relatively high minimum inhibitory concentration (MIC) limiting the use of EFV as a single antibacterial agent, it exhibits significant synergistic activity at sub-MIC levels when paired with various antibiotics against Enterococcus species and Staphylococcus aureus. EFV showed restored sensitivity of β-lactams against Methicillin-resistant S. aureus (MRSA). It increased the effectiveness of antibiotics tested against Methicillin-sensitive S. aureus (MSSA). It also helped to overcome the intrinsic resistance barrier for several antibiotics in Enterococcus spp. In silico binding studies aligned remarkably with experimental antimicrobial testing results and highlighted the potential of EFV to direct the engagement of PBPs with moderate to strong binding affinities (pK_a 5.2–6.1). The dual-site PBP2 binding mechanism emerged as a novel inhibition strategy, potentially circumventing resistance mutations. Special attention should be paid to WalK binding predictions (pK_a = 4.94), referring to the potential of EFV to interfere with essential regulatory pathways controlling cell wall metabolism and virulence factor expression. These findings, in general, suggest the possibility of EFV as a promising lead for the development of new antibacterial agents. Full article