Search Results (206)

Background/Objectives: Pseudomonas aeruginosa is one of the leading causes of ventilator-associated pneumonia (VAP) and ventilator-associated tracheobronchitis (VAT), with a worldwide spread of difficult-to-treat high-risk clones. This study aimed to investigate the virulence potential and to characterize phenotypic and genotypic antimicrobial resistance (AMR) in P. aeruginosa causing VAP/VAT in the Intensive Care Unit (ICU), University Hospital of Split, Croatia. Methods: The study included P. aeruginosa isolates obtained from ICU patients who met the criteria for VAP or VAT, between January 2023 and January 2024. Isolates were identified using MALDI-TOF MS and tested for antimicrobial susceptibility (AST). A subset of phenotypically multidrug-resistant (MDR) isolates was further analyzed using whole-genome sequencing (WGS) and multilocus sequence typing. Results: A high rate of resistance was detected to ceftazidime (23.4%), imipenem (39.6%), and meropenem (43.8%). WGS confirmed the presence of multiple AMR genes, including the bla_VIM-2 gene, whose genetic environment highlights a complex MDR locus integrating multiple AMR determinants and mobile genetic elements. All tested isolates possessed genes for class C (bla_PDC34, bla_PDC374 or bla_PDC16) and class D (bla_OXA-2, bla_OXA-10 or bla_OXA-50) β-lactamases, fosA, aph(3′)-IIb and crpP genes. Additionally, WGS analysis revealed the presence of numerous virulence genes including those for adherence (Type IV pili and Fap protein production), motility (such as flgF), biofilm formation (e.g., algE and mucE), quorum sensing (lasI, lasR, rhlI and rhlR), exotoxin (toxA and plcH) and exoenzyme activity (exoU, exoT, exoS, exoY, pcrV, hcp1 and lasA). The isolates belonged to four different sequence types: ST235, ST446, the high-risk ST253 and the widely distributed ST395. Phylogenomic comparison demonstrated that the isolates from this study do not originate from a single clonal source, but instead represent multiple globally distributed high-risk P. aeruginosa lineages introduced into the clinical setting. Conclusions: Due to the emergence of high-risk clones with broad AMR and strong virulence potential, ineffectiveness of standard empirical therapy may be anticipated, highlighting the urgent need for new therapeutic approaches (including those targeting major virulence factors). Full article

Background: Aggregatibacter actinomycetemcomitans is a Gram-negative, facultative anaerobic, immobile oral bacterium responsible for the secretion of virulence factors, namely leukotoxin (LtxA), a large exotoxin of the RTX family that enables the bacterium to evade the immune system by destroying leukocytes, resulting in aggressive periodontitis (AP) leading to tooth loss. Methods: This study aimed to screen 106 molecules derived from Moroccan propolis in order to identify potential inhibitors of the active sites of LtxA based on molecular docking, ADMET property evaluation, and molecular dynamics (MD) simulation. Results: Epigallocatechin gallate (EGCg), used as a reference compound, showed binding energies of −6.9 kcal/mol, −6.1 kcal/mol, −6.5 kcal/mol, and −5.9 kcal/mol with the four active sites P1, P2, P3, and P4, respectively. By establishing conventional hydrogen bonds, pi-alkyl bonds, and non-covalent pi–pi bonds. Chrysin and luteolin showed favorable binding affinities with the four active sites, named as follows: P1–P4 (P1–chrysin = −7.5 kcal/mol; P2–chrysin = −7.9 kcal/mol; P3–chrysin = −8.1 kcal/mol; P4–chrysin = −6.9 kcal/mol; P1–luteolin = −7.3 kcal/mol; P2–luteolin = −7.6 kcal/mol; P3–luteolin = −8.1 kcal/mol; P4–luteolin = −7.3 kcal/mol). The binding affinity of these two propolis derivatives was stabilized by pi−sigma bonds, pi−alkyl bonds, conventional hydrogen bonds, pi-cation interactions, non-covalent pi–pi bonds, and carbon–hydrogen bonds. According to free energy calculations performed with Prime MM-GBSA, the complexes formed by chrysin demonstrated the most stable interactions due to Van der Waals and lipophilic forces. Luteolin formed significant interactions, but slightly weaker than those of chrysin. These results reveal the inhibitory potential of chrysin and luteolin with protein active sites. MD simulations corroborated the excellent stability of complexes formed by chrysin, as indicated by low RMSD values, suggesting favorable dynamic behavior. Conclusions: These results highlight the potential of chrysin as a versatile inhibitor capable of interacting with the four active sites. These findings are a strong foundation for further experimental confirmations. Full article

Slaughterhouses serve as critical surveillance hubs for identifying subclinical and economically important diseases in food-producing animals. Trueperella (Arcanobacterium) pyogenes, an opportunistic pathogen commonly found on the mucous membranes of livestock, is associated with mastitis, abortion, and suppurative infections such as abscesses. In this study, we investigated 30 pig carcasses fully condemned due to vertebral osteomyelitis (VO) at two slaughterhouses in Gwangju, Republic of Korea, between November 2023 and May 2024. From abscess lesions, 11 T. pyogenes strains were isolated and characterized morphologically, biochemically, and genetically. The hemolytic exotoxin pyolysin (plo gene), a major virulence factor, was detected in five isolates (45.46%). Phylogenetic analysis of partial 16S rDNA sequences confirmed close clustering with known T. pyogenes reference strains. All 11 isolates exhibited multidrug resistance, showing resistance to 8–14 antimicrobial agents per strain. Complete resistance (11/11, 100%) was observed against amikacin (AMI), nalidixic acid (NAL), chloramphenicol (CHL), florfenicol (FFN), and trimethoprim/sulfamethoxazole (SXT). High resistance rates were also detected for erythromycin (ERY) and clindamycin (CLI) (10/11, 90.9%), ceftazidime (TAZ), ceftriaxone (AXO), ciprofloxacin (CIP) (7/11, 63.6%), and tetracycline (TET) and streptomycin (STR) (5/11, 45.5%), while gentamicin (GEN) resistance was found in three isolates (27.3%). In contrast, none of the isolates showed resistance to ampicillin, cefoxitin, or cefotaxime. These findings underscore the epidemiological value of abattoir-based monitoring in detecting emerging pathogens and tracking antimicrobial resistance. The results provide important baseline data to inform disease control strategies, guide antimicrobial stewardship, and support One Health approaches, including the development of preventive measures such as vaccines. Full article

Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen frequently associated with delayed wound healing, particularly in chronic skin injuries. Its capability to form biofilms, secrete virulence factors, and the faculty to compete with other microorganisms makes it a major challenge in clinical wound management. Recent literature reveals different molecular and cellular mechanisms through which P. aeruginosa disrupts the wound healing process. Findings highlight that it interferes with key phases of healing by modulating host immune responses, degrading extracellular matrix components, and inhibiting keratinocyte migration. Its quorum-sensing systems regulate the expression of critical virulence factors such as exotoxin A, elastases, pyocyanin, and rhamnolipids. Additionally, the production of the biofilm matrix components alginate, and polysaccharides provide protection against host defenses and antibiotics. Interactions with other microorganisms, including antagonistic effects on Staphylococcus epidermidis and synergistic relationships with Staphylococcus aureus, modify the wound microbiota. Promising therapeutic alternatives have shown efficacy in disrupting biofilms and reducing virulence. These insights remark the importance of targeting both P. aeruginosa and its ecological interactions to enhance wound healing outcomes and develop more effective treatments. This review aimed to highlight the pathogenic role of P. aeruginosa and its interactions with other microbial species in the context of skin wound healing. Full article

Furunculosis caused by Aeromonas salmonicida is one of the most common diseases in aquaculture, leading to significant economic losses. This study comprehensively investigated the dynamics of pathophysiological and histopathological disorders in rainbow trout (Oncorhynchus mykiss) infected with the moderately virulent strain A. salmonicida SL0n. Whole-genome analysis showed that strain SL0n belongs to the A. salmonicida species complex, possessing a single circular chromosome. The genome encodes a wide range of virulence factors, including adhesion systems (type IV pili, fimbriae), toxins (aerolysin, hemolysins), and a type II secretion system (T2SS), but notably lacks plasmids and a type III secretion system (T3SS). This genomic profile likely dictates a pathogenic mechanism reliant on secreted exotoxins (via T2SS), explaining the observed systemic cytotoxic damage. In an acute experiment, the 4-day LD₅₀ was determined to be 1.63 × 10⁶ CFU/fish. In a prolonged experiment, fish were injected with a sublethal dose (1.22 × 10⁶ CFU/fish—75% of LD50). The disease progressed through three consecutive stages. The early stage (1–2 DPI) was characterized by maximal bacterial load and activation of nonspecific immunity. The acute stage (4 DPI) manifested as severe septicemia and anemia, associated with systemic organ damage, which correlated with peak AST and ALT enzyme activity. The recovery stage (6 DPI) was marked by partial regression of inflammation, key biochemical and histological parameters indicated persistent liver and kidney dysfunction, signifying an incomplete recovery. These results demonstrate the pathogenesis of acute furunculosis and reveal that the genomic profile of the SL0n strain causes a sequential, systemic infection characterized by severe organ dysfunction. Full article

Protein toxins are biologically active polypeptides produced by a variety of organisms, including bacteria, plants, fungi, and animals. These molecules exert potent and specific toxic effects on target cells and are primarily associated with pathogenicity and defense mechanisms of the organisms. In the past few decades, significant progress has been made in understanding their structure, mechanisms of action, and regulation. Among these, bacterial protein toxins have emerged as valuable tools particularly in the development of targeted therapies. A notable example is Botulinum toxin, originally known for its neurotoxic effects, which was approved as a therapeutic agent in 1989 for strabismus treatment, paving way for repurposing bacterial toxins for clinical use. This review provides an overview of the different classes of bacterial toxin-based therapeutics, with a particular focus on Pseudomonas exotoxin A (PE) from Pseudomonas aeruginosa and anthrax toxin from Bacillus anthracis. The modular architecture and potent cytotoxicity of these A-B type toxins have enabled their successful adaptation into targeted cancer therapies. The clinical approval of the PE-based immunotoxin, moxetumomab pasudotox, for the treatment of hairy cell leukemia, underscores the potential of this strategy. This review also discusses current challenges and outlines future directions for the advancement of bacterial toxin-based therapeutics. Full article

Urinary tract infections (UTIs) remain a major global health concern, with rising antimicrobial resistance prompting the search for alternative therapies. Selenium nanoparticles (Se NPs) are promising antimicrobial agents due to their unique physicochemical properties and ability to disrupt bacterial physiology. This study evaluated the antibacterial efficacy of Se NPs against four uropathogens and conducted comparative proteomic analyses to elucidate stress responses. Enumeration assays showed that Se NPs effectively inhibited bacterial growth, with Pseudomonas aeruginosa being the most susceptible and Proteus mirabilis the most resistant. Microscopy revealed Se NP-induced membrane rupture and cellular deformation across all species. Proteomic and bioinformatic analyses showed more pronounced protein regulation in P. mirabilis than in P. aeruginosa. Cluster of Orthologous Groups (COG) analysis revealed both shared and species-specific responses, while Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated activation of key stress pathways. Virulence-associated proteins were modulated in both species, with P. mirabilis uniquely upregulating stress survival and exotoxin-related proteins. Both regulated efflux pumps, suggesting active transport mitigates Se NP toxicity. P. aeruginosa showed mercury resistance, while P. mirabilis expressed tellurite resistance proteins. These findings highlight distinct yet overlapping strategies and support the potential of Se NPs in novel antimicrobial development. Full article

Background. We are developing cytotoxic anti-HIV immunoconjugates to attack the reservoir of infected cells that persist after years of fully suppressive anti-retroviral therapy. Methods. We have produced a chimeric fusion protein, J3ExoA, consisting of J3VHH, a broadly reactive anti-gp120 camelid nanobody, joined to the de-immunized PE38 fragment of Pseudomonas exotoxin A. The efficacy of J3ExoA was compared to that of a well-studied anti-gp41 immunotoxin (IT), 7B2-dgA, in cytotoxicity assays and for inhibition of infectivity. Immunogenicity of the ITs was tested in mice. Results. J3ExoA killed cells expressing the HIV envelope with specificity in concentrations in the ng/mL range. Of all anti-HIV ITs we have tested, only J3ExoA compared to 7B2-dgA in cytotoxic efficacy, although there were differences between the two ITs on different target cells. J3ExoA suppressed the spread of HIV infection in tissue culture. J3ExoA was less immunogenic than 7B2-dgA, but mice made antibodies to both portions of the fusion protein. Conclusions. J3ExoA represents a novel IT that may be used to eliminate infected cells in the persistent HIV reservoir of infection, the barrier to an HIV “cure.” Additional approaches for addressing IT immunogenicity are discussed. Full article

Cutaneous adverse reactions (CARs) are common complications of epidermal growth factor receptor (EGFR) inhibitor therapy, with papulopustular eruptions and paronychia being the most frequent. Growing scientific evidence implies that Staphylococcus aureus is involved in the pathogenesis of these reactions. This observational prospective study characterized 42 S. aureus strains isolated from CARs, analyzing antibiotic resistance, biofilm formation, soluble virulence factors, and virulence/resistance genes using multiplex polymerase chain reaction (PCR). S. aureus was identified in 90% of lesions; in 33% of cases, nasal and skin isolates were genetically identical. High resistance rates were noted for penicillins (85%) and tetracyclines (57%), while all strains remained susceptible to fluoroquinolones, vancomycin, and rifampicin. All isolates formed biofilms, and DNase/esculinase production significantly correlated with CAR severity. An enzymatic score based on these markers was associated with an 18-fold increased risk of severe reactions. Genotypically, clfA and clfB were prevalent (85.7%), while exotoxin genes were less common. These findings support a key role for S. aureus in exacerbating CARs via antibiotic resistance, biofilm production, and the expression of virulence factor. Additionally, we emphasize the role of routine microbial screening—including nasal swabs—and therapy guided by antibiograms. Furthermore, the enzymatic score may further be validated as a predictive biomarker. Full article

We present a case of fatal necrotizing Staphylococcus aureus pneumonia with underlying influenza A (H3) infection. Next-generation-sequencing-based analysis revealed that the S. aureus isolate harbored the newly recognized exfoliative toxin etE2 gene. Molecular epidemiologic analysis showed that the isolate belonged to the MSSA ST152 lineage, harboring PVL genes and edinB co-located to etE2 as distinctive virulence factors. The etE2 gene is present in all isolates of this lineage co-located to the exotoxin gene edinB, both implicated in the destruction of tissue integrity. We alert as to the global emergence of this lineage causing serious infections in patients. Full article

Cystic fibrosis (CF) is a life-limiting autosomal recessive disorder affecting a large number of individuals in Europe. The disease arises from mutations in the CFTR gene encoding the cystic fibrosis transmembrane conductance regulator, a chloride ion channel crucial for maintaining epithelial ion and fluid homeostasis. Dysfunctional CFTR disrupts mucociliary clearance, particularly in the respiratory tract, resulting in persistent bacterial colonization, chronic inflammation, and progressive pulmonary damage—ultimately leading to respiratory failure, the principal cause of mortality in CF patients. Early diagnosis and advances in therapy have substantially improved both survival and quality of life. A hallmark of CF pathology is the establishment of polymicrobial infections within the thickened airway mucus. Pseudomonas aeruginosa is the dominant pathogen in chronic CF lung infections and demonstrates a remarkable capacity for adaptation via biofilm formation, metabolic reprogramming, and immune evasion. Biofilms confer increased tolerance to antimicrobial agents and facilitate long-term persistence in hypoxic, nutrient-limited microenvironments. P. aeruginosa exhibits a wide range of virulence factors, including exotoxins (e.g., ExoU, ExoS), pigments (pyoverdine, pyochelin), and motility structures (flagella and pili), which contribute to tissue invasion, immune modulation, and host damage. During chronic colonization, P. aeruginosa undergoes significant genotypic and phenotypic changes, such as mucoid conversion, downregulation of acute virulence pathways, and emergence of hypermutator phenotypes that facilitate rapid adaptation. Persistent cells, a specialized subpopulation characterized by metabolic dormancy and antibiotic tolerance, further complicate eradication efforts. The dynamic interplay between host environment and microbial evolution underlies the heterogeneity of CF lung infections and presents significant challenges for treatment. Elucidating the molecular mechanisms driving persistence, hypermutability, and biofilm resilience is critical for the development of effective therapeutic strategies targeting chronic P. aeruginosa infections in CF. Full article

Clostridioides difficile (C. difficile) is a major pathogen responsible for antibiotic-associated diarrhea, frequently observed in hospital settings. Due to the widespread use of antibiotics, the incidence and severity of C. difficile infection (CDI) are rising across the world. CDI is primarily driven by two homologous protein exotoxins, toxin A (TcdA) and toxin B (TcdB). Other putative virulence factors include binary toxin CDT, surface layer proteins, phosphorylated polysaccharides, and spore coat proteins. These C. difficile virulence factors are potential targets for vaccine development. Although several C. difficile vaccines have entered clinical trials, there is currently no approved vaccine on the market. This review outlines the intoxication mechanism during CDI, emphasizing the potential antigens that can be used for vaccine development. We aim to provide a comprehensive overview of the current status of research and development of C. difficile vaccines. Full article

Introduction: Pseudomonas aeruginosa produces many exotoxins which are essential for the bacterial pathogenesis. The aim of this study was to identify Pseudomonas aeruginosa from clinical specimens, detect the sensitivity pattern, biofilm production, and the frequency of exogenes. Methods: Pseudomonas aeruginosa clinical isolates were identified by conventional and genotypic methods. Antibiotic susceptibility patterns and biofilm production were performed. Molecular detection of exotoxin genes exoS, exoT, exoU, and exoY in Pseudomonas aeruginosa isolates was performed by PCR. Results: Seventy-five Pseudomonas aeruginosa were identified in 400 clinical specimens. Sixty-six (88%) isolates were carbapenem-resistant. A total of 25 (33.3%) isolates were extensively drug resistant, 18 (24%) were multidrug resistant, and 11 (14.7%) were pandrug resistant. Sixty-three (84%) isolates were biofilm producers. Biofilm formation was detected in 56 (85%) of carbapenem-resistant isolates. Totally, 70 (93.3%) isolates carried exoS, 68 (90.7%) carried exoY, 65 (86.7%) carried exoT, and 28 (37.3%) carried exoU. Exogenes were highly expressed in carbapenem-resistant isolates. Coexistence of more than one gene was detected in nearly all isolates. Conclusions: Pseudomonas aeruginosa clinical isolates were resistant to many anti-pseudomonal antibiotics. Most of isolates were biofilm-producers. The genes exoT, exoS and exoY were identified in almost all P. aeruginosa strains and are considered an inevitable component of P. aeruginosa virulence. Full article

Introduction: The increase in the rates of multidrug-resistant bacteria in healthcare environments has been recognized as a global public health problem. In view of the scarcity of data on the neonatal population, this study aimed to provide information on the genotypic and epidemiological characteristics of Gram-negative microorganisms isolated from colonization and infection sites in neonates admitted to a tertiary university center of high complexity. Methods: Enterobacterales and non-fermenting Gram-negative bacilli previously collected in a prospective cohort study were submitted to genotypic identification, detection of extended-spectrum β-lactamases (ESBL), carbapenemases and biofilm production, detection of specific virulence markers in Pseudomonas aeruginosa, and typing by pulsed-field gel electrophoresis. Results: The data found here revealed higher rates of infection by Klebsiella spp. and Serratia marcescens that caused bloodstream infection and pneumonia, respectively. In this study, high biofilm production was observed, with 95.0% of Enterobacterales and 100% of non-fermenting Gram-negative bacilli being producers. Most of the P. aeruginosa isolates carried pathogenicity factors such as alginate, hemolytic phospholipase C, exotoxin A, and rhamnolipids. The phenotypic analysis of ESBL revealed that 16 (5.3%) isolates produced these enzymes. Four of these isolates (66.7%) carried the CTX-M-9 gene, three (50%) carried the TEM gene, and one (16.7%) was positive for the SHV and CMY-2 genes. Univariate and multivariate Cox regression analyses were used to identify risk factors for colonization and infection by Gram-negative microorganisms. The results of multivariate analysis revealed that biofilm production by these microorganisms was associated with the persistence of colonization by the same pathogen in the newborn and increased by 75% the daily probability of the newborn developing infection. The production of ESBL also increased the daily probability of infection by 46.8 times. Conclusions: Enterobacterales showed average biofilm production, while the majority of non-fermenting Gram-negative bacilli were strong producers. The present data increase our knowledge of the molecular epidemiology of important Enterobacterales species, with emphasis on ESBL-producing Enterobacter cloacae and Klebsiella pneumoniae with emerging epidemiological potential in the neonatal intensive care unit of a tertiary university hospital. Furthermore, the results highlight the need for the monitoring and implementation of control measures and for restricting the use of broad-spectrum antibiotics. Full article

The bacterium Bacillus anthracis secretes three protein exotoxins: Protective Antigen 83 (PA83), Lethal Factor (LF), and Edema Factor (EF). A cleaved form of PA83 (PA63) aids LF and EF entry into the cytoplasm, which leads to anthrax-induced cell death. The Protein Data Bank (PDB) has more than 25 structures of LF: the monomer alone, bound with inhibitors, or bound to PA63. The structures are all—with only minor shifts of a few Ångströms—nearly congruent. We have measured the structure of LF at equilibrium in D₂O solution by small-angle neutron scattering (SANS). The shape is modeled well by a parallelepiped (all angles 90°) with dimensions of 12 Å × 49 Å × 129 Å. For a protein with a typical density of 1.4, the molecular weight would be between 55 and 94 kDa, which is comparable to that of the 90.2 kDa monomer. However, the LF crystal structure PDB 1pwu (a generally V-shaped molecule with equal arm lengths ≈ 70 Å) with the same model fits the dimensions 30 Å × 48 Å × 104 Å. Given the large changes in the long and short dimensions, straightforward physical modeling of the solution structure from the crystal form is unable to match the SANS results. Full article