Search Results (2,401)

Bacterial extracellular vesicles (BEVs) are nanoscale spherical lipid bilayer structures secreted by bacteria, including outer membrane vesicles (OMVs) released by Gram-negative bacteria and membrane vesicles (MVs) produced by Gram-positive bacteria. Although the biogenesis of BEVs requires substantial energy expenditure, these vesicles provide bacteria with strategic advantages in the evolutionary interplay between pathogens and hosts. BEVs contribute to bacterial adaptation to environmental stress by remodeling membrane components, eliminating toxic substances, promoting biofilm formation, and mediating the interbacterial transfer of antibiotic resistance determinants. They can also function as decoys to protect bacteria from bacteriophage or antibiotic attack, deliver virulence factors, modulate host immune responses to facilitate bacterial colonization, and mediate interspecies competition. This review summarizes the central roles of BEVs as bacterial mediators of environmental responses, with particular emphasis on their involvement in immune regulation, environmental adaptation, and interspecies competition, thereby providing new insights into pathogen evolutionary strategies. Full article

Wound infections due to antibiotic resistance pose a global public health problem. Phage therapy is a promising approach to address this issue. To improve localization, phage stability, delivery, and antibacterial performance, we propose polymer mix gel microbeads encapsulated with phages as a model for the delivery of phiKZ bacteriophage to combat Pseudomonas aeruginosa. Phages were loaded into the alginate pre-gel under magnetic stirring, with further cross-linking by chitosan and/or Ca²⁺ ions. The obtained gel microbeads were characterized using FTIR and Raman spectroscopy, and their cytotoxicity and antimicrobial properties were evaluated. This study demonstrated the efficient loading of high-titer phage lysate, achieving up to 99% encapsulation efficiency for alginate–chitosan microbeads. The key characteristics of the microbeads include stable physicochemical properties, slow but continuous phage release over 48 h in physiological saline, and low cytotoxicity. The phage-loaded microbeads demonstrated strong in vitro antimicrobial activity against P. aeruginosa PAO1, resulting in mean reductions of 6.9 log₁₀ and 4.8 log₁₀ CFU/mL for alginate and alginate–chitosan formulations, respectively. This corresponded to a decrease in bacterial concentration from approximately 1.1 × 10¹¹ CFU/mL in untreated controls to 1.1 × 10⁵ CFU/mL and 7.7 × 10⁶ CFU/mL for alginate and alginate–chitosan formulations after 3 h of incubation. Full article

Lactic acid bacteria (LAB), as important probiotics, face challenges in applications from bacteriophage infection and the instability of foreign genetic elements. Although Gabija systems and their GajA nuclease components have been characterized in model bacteria, their distribution, biochemical properties, and defensive functions in LAB remain largely unexplored. Here, we provide the first systematic characterization of a naturally occurring Gabija system from Ligilactobacillus salivarius Ren and clarify its distribution among LAB. Approximately 9.3% of LAB strains encode the Gabija system, which exists as a gajA-gajB gene cluster. We found that the Gabija system originated independently in different bacterial lineages. The GajA of L. salivarius Ren (LsGajA) was purified and exhibited non-specific nuclease activity that could efficiently cleave various nucleic acid substrates, including plasmids and linear double-stranded DNA (dsDNA). This activity displayed a temperature-dependent profile, with high activity observed from 45 to 60 °C and at pH 8.0. Mg²⁺ markedly enhanced its degradative nuclease activity, whereas high concentrations of dNTPs inhibited DNA cleavage. LsGajA exhibited substrate-dependent differences in cleavage efficiency, indicating that substrate origin and associated physicochemical features may influence its activity. Additionally, we demonstrate that LsGajA exhibits exceptional stability as a nuclease, retaining activity under a wide range of conditions. The LsGabija system significantly enhanced the ability to reject foreign plasmids and provided strong resistance to the bacteriophage T5 in Escherichia coli. This study provides the first systematic biochemical and functional characterization of the Gabija system in LAB, advancing our understanding of this prokaryotic defense system and highlighting its potential for industrial applications. Full article

Biofilms are structured communities of microorganisms embedded in a self-produced extracellular polymeric substance (EPS) matrix, whose development significantly enhances microbial resistance to antibiotics, disinfectants, and host immune defenses, posing major challenges in clinical, industrial, and environmental settings. Compared with planktonic cells, biofilm-associated microorganisms can exhibit up to 10- to 1000-fold increased tolerance to antimicrobial agents, contributing to the persistence of biofilm-associated infections (BAIs). These infections remain difficult to eradicate due to reduced penetration, altered metabolic states, and the presence of dormant or persister cells. Anti-biofilm strategies can be broadly classified into physical approaches (e.g., ultrasound, mechanical stress, and light-based approaches) that target biofilm structure; chemical and enzymatic methods (e.g., EPS-degrading enzymes) that destabilize the matrix; and biological and molecular strategies (e.g., quorum-sensing (QS) inhibitors, anti-virulence agents, bacteriophages, phage-derived antimicrobial molecules, antimicrobial peptides, and natural bioactive compounds) that modulate biofilm development and integrity by targeting regulatory pathways and matrix stability through distinct mechanisms of action. Natural compounds, including lactoferrin, lactoferrin-derived peptides, and probiotic and postbiotic fractions of lactic acid bacteria (LAB), as well as plant-derived metabolites, have shown promising anti-biofilm effects, with efficacy often enhanced through complementary or potentially synergistic interactions. However, despite these advancements, clinical translation remains limited. For example, BAIs account for approximately 80% of chronic infections, with high recurrence rates and therapeutic failure reported in device-associated infections and chronic wounds. These limitations highlight the need for clinically translatable, multimodal approaches that integrate structural biofilm disruption, antimicrobial targeting, and host response modulation to design more effective and sustainable anti-biofilm strategies. Full article

Effective hand washing takes time and hand sanitizers that contain alcohol have a number of drawbacks, and frequent use of alcohol may cause skin damage. The objective of this study is to formulate nanostructured lipid carrier systems containing chlorhexidine digluconate to be applied topically for hand hygiene, especially for people sensitive to alcohol. A cytotoxicity experiment was conducted to ascertain the safe dosage for each of the three nano-cream formulas (F1, F2 and F3). Following each treatment, the viral titer was assessed using tissue culture infectious dose₅₀ and standard plaque assays. The selected formulation was characterized rheologically. Furthermore, fifteen volunteers of various ages and genders participated in the vivo antimicrobial test of the selected formulation as a hand sanitizer. All of the formulas were found to be safe. Using the disc diffusion method, the three formulations exhibited in vitro antimicrobial effects against different microbes. F1 showed biphasic release, reasonable skin deposition and spherical droplets under a microscope. F1 exhibited a non-Newtonian shear thinning flow behavior. After 30 min, the reduction values for rotavirus and Phix-174 were 21 and 4%, respectively. Additionally, the impact of F1 was assessed on the infectivity of simian rotavirus sa-11 (ds RNA) and Phix-174 (ss DNA) bacteriophage. According to the findings of the in vivo study, the percentage of total bacterial counts that were removed varied from 91 to 100%. Moreover, the range of the removal percentage of total fungi was 95.38 to 100%. In summary, F1 can be used as an economic, safe, and effective hand antiseptic. It can also completely replace alcohol in the market. Full article

Streptococcus pneumoniae remains a major global health threat and is listed by the World Health Organization as a pathogen in urgent need of new antimicrobial strategies. While primarily considered an extracellular pathogen, S. pneumoniae can persist within splenic macrophages in severe disease, creating a protected intracellular niche that may contribute to fulminant sepsis. We recently demonstrated the concept of an mRNA-based therapeutic approach in which host cells produce the pneumococcal bacteriophage endolysin Cpl-1. Here, we investigated whether expression of Cpl-1 in macrophages can target S. pneumoniae residing within host cells. Using the human THP-1 macrophage line, we demonstrated successful translation and intracellular accumulation of bioactive Cpl-1 following IVT-mRNA transfection. Lysates from Cpl-1 mRNA-transfected cells exhibited bacteriolytic activity, and Western blotting as well as immunofluorescent staining confirmed cytosolic endolysin production. Phagocytosis assays using an encapsulated and unencapsulated pneumococcal strain showed a reduction in intracellular bacterial burden in Cpl-1 mRNA-transfected macrophages compared with control and inactive-mutant Cpl-1 mRNA groups, and a flow cytometry-based assay further corroborated a decreased intracellular bacterial signal. Together, these findings suggest that mRNA-encoded Cpl-1 enhances intracellular killing of S. pneumoniae and supports the feasibility of mRNA-based endolysin therapies to target intracellular pneumococcal reservoirs. Full article

Considering the therapeutic potential of the Proteus mirabilis PM16 podophage, the interaction between PM16, its host strain, and the mouse immune system was investigated. We evaluated how pre-existing humoral immunity to PM16 influences the immune response against P. mirabilis and the neutralization of the phage itself. Balb/c mice were divided into three groups and immunized two times with (1) 0.9% NaCl, (2) adjuvants, or (3) a mixture of PM16 and an adjuvant. Then, each group was subdivided into three subgroups: mock infection, infection with P. mirabilis, and infection with P. mirabilis followed by model phage therapy with PM16. The obtained results demonstrated that pre-immunization with PM16 enhanced the anti-P. mirabilis IgG antibody response upon bacterial challenge, indicating that the phage potentiates antibacterial immunity. In addition, pre-immunization elicited a significant anti-PM16 antibody response that resulted in in vitro neutralization of phage lytic activity. However, phage-neutralizing antibodies neither decreased the efficacy of phage therapy nor influenced bacteria-specific immune response. Thus, while PM16 can boost the host’s immune response against its bacterial host, the resulting humoral immunity also drives phage clearance through both direct and bacteria-mediated neutralization pathways, revealing a complex immunopharmacological relationship central to phage therapy. Full article

Newborns may suffer from dangerous bacterial infections caused by life-threatening multi-drug resistant pathogens. Thus, despite bactericidal capabilities of antibiotics, microorganisms are known to circumvent this therapy, and a new, more effective type of remedy is needed. An increasingly recognized strategy for addressing these challenges is the use of bacteriophages—viruses infecting bacteria—collectively referred to as phage therapy. Nonetheless, the research considering phage therapy amongst newborns is still at a pioneering stage, owing to the scarcity of systematic investigations and the prevalence of case-study data, leaving room for further discovery and analysis. This review summarizes the information needed to understand this complex issue, considering the description of pathogens causing infections affecting newborns, the formation of the early microbiota and phageome (defining its composition followed by an influence on immune system development), and the possible use of bacteriophages in the treatment, which may be complicated by ethical concerns. Full article

Staphylococcus aureus, including methicillin- and vancomycin-resistant variants (MRSA, VRSA), causes infections that are increasingly difficult to treat with conventional antibiotics. One of the approaches to developing new therapeutics to treat staphylococcal infections is the use of bacteriophages specific to these bacteria or the lytic enzymes of such bacteriophages, which are capable of hydrolyzing the cell walls of these bacteria. Phage-encoded endolysins offer an alternative promising class of antimicrobial agents. In this study, LysSA120, a 250-amino-acid endolysin encoded by the S. aureus podophage vB_SauP_120, was cloned, expressed in Escherichia coli, and characterized. The domain organization and tertiary structure of LysSA120 were predicted. Recombinant LysSA120 hydrolyzed cell walls were obtained from S. aureus, S. epidermidis, S. haemolyticus, S. warneri, S. auricularis and S. saprophyticus. It was shown that treatment of S. aureus planktonic cells with endolysin LysSA120 led to reduced viability. Furthermore, LysSA120 could hydrolyze mature biofilm formed by VRSA. The lytic spectrum and antibiofilm activity of LysSA120 warrant its further evaluation as an enzybiotic against drug-resistant staphylococcal infections. Full article

Bacteriophage-based products are gaining attention as effective tools to reduce harmful germs in food and combat antimicrobial resistance throughout the food production process. However, in South America, their use is still limited because of complicated regulations and inconsistent evidence requirements. This review aims to (i) explore the current scientific and technological landscape of using bacteriophages in South American food systems, (ii) identify main regulatory challenges that impact their classification, approval, and use, and (iii) highlight the need for consistent international guidelines, especially from Codex Alimentarius, to help safely and effectively incorporate phage-based products in food. Research on phage-based products is growing, but it is not consistent across different regions. There are more patents and advancements in biotechnology, but they are limited to certain areas. Although progress is being made, the regulatory frameworks are still unclear, especially when it comes to how these products are classified, labeled, and monitored for safety. To address these gaps, risk-based guidelines are needed. These should define product categories and claims, set safety standards, and include rules for tracking products and monitoring them after they hit the market. Creating a new Codex Alimentarius project on phage-based products could help establish global guidelines that promote safe use, reduce uncertainty in regulations, and improve trade in food markets around the world. Full article

The effect of administration routes (oral and rectal) on the pharmacokinetics of Salmonella phage SE40 and Escherichia phage V18 was studied. To detect phages in biomaterial (blood, urine, and feces), the Spot-test, Gratia, and double-nested polymerase chain reaction methods were used. Systemic action of the studied phages with both routes of administration, beginning 15–30 min after administration, was demonstrated. The phages persisted for up to 24 h after both oral and rectal administration. However, the concentration in the blood was higher after oral administration, while concentrations in urine and feces were higher after rectal administration. The need to protect phages from the acidic contents of the stomach was confirmed. Full article

Prophages are bacteriophage genomes that are part of bacterial chromosomes. They are not just dormant passengers; they actively shape pathogen biology. For example, in skin-infecting pathogens such as Staphylococcus aureus, Streptococcus pyogenes, and Pseudomonas aeruginosa, prophages carry important virulence factors, cytotoxins, superantigens, immune evasion clusters, and epigenetic regulators that directly affect the course of skin and soft tissue infections. This same prophage biology provides a therapeutic strategy: prophage-derived molecules, including endolysins, holins, spanins, and polysaccharide depolymerases, demonstrate potent antimicrobial and antibiofilm activity against drug-resistant skin pathogens, with several candidates now in clinical development. Engineered chimeric lysins, CRISPR-encoded prophage delivery systems, and the systematic mining of the skin microbiome phageome collectively enhance the translational potential of this biology. This review integrates mechanistic insights into prophage-mediated virulence. It assesses the translational landscape of prophage-derived therapeutics, delineating the conceptual and clinical frontiers that characterize the forthcoming chapter in this domain. Full article

Background/Objectives: Antimicrobial resistance (AMR) in Escherichia coli is a growing public health concern, particularly in regions affected by environmental contamination and poor wastewater management. Data on locally isolated E. coli-targeting phages in Lebanon remain limited. This study aimed to isolate, characterize, and evaluate two lytic bacteriophages against AMR E. coli. Methods: Two phages, EPIMAM01 (gb:PQ493298) and EPIMRB01 (gb:PQ657784), were isolated from untreated sewage in Beirut using E. coli ATCC 25922. Characterization included double-layer agar assays, one-step growth analysis, and stability testing across temperature and pH ranges. Bacteriolytic activity was assessed in planktonic cultures and preformed biofilms. Host range and efficiency of plating (EOP) were evaluated using clinical isolates. Whole-genome sequencing and comparative analyses were performed. Results: Both phages produced clear plaques and showed a latent period of ~40 min. EPIMAM01 had a higher estimated burst size (140 PFU/infected cell) than EPIMRB01 (75 PFU/infected cell). Both phages remained stable between 4–50 °C and within a pH range of 5–10 but showed marked loss of activity at temperatures ≥60 °C and pH ≤3 or ≥12. EPIMAM01 effectively inhibited planktonic growth of E. coli ATCC 25922, whereas EPIMRB01 showed stronger biofilm-disrupting activity against preformed E. coli biofilms. Both phages lysed several of the 17 tested clinical E. coli isolates. Comparative analyses of gene presence/absence patterns, bacterial defense systems, and adsorption phenotypes among the tested E. coli strains identified mlaA, ydcQ, and ompD-2 as candidate adsorption-associated genes and suggested CRISPR systems may reduce susceptibility. Genomic analysis classified both phages as T4-like phages lacking lysogeny, virulence, or AMR genes. Conclusions: EPIMAM01 and EPIMRB01 are lytic phages with complementary antimicrobial properties, supporting their potential for further development as AMR control agents. Full article

The antimicrobial resistance crisis has led to the use of metals and bacteriophages as possible alternatives to antibiotics. Experimental studies have examined interactions between ionic/nano-silver and bacteriophages against multidrug-resistant bacteria. However, these approaches have often failed to examine whether silver affects the stability and infectivity of bacteriophages. Here, we utilized experimental evolution to evolve resistance to ionic silver in bacteriophage T7. High ionic silver concentrations that do not represent physiological exposure conditions were used to impose strong selective pressure. Evolution of ionic silver resistance in phage T7 was rapid, as evidenced by recovery of bacteriophage growth in E. coli following repeated exposures to ionic silver, enhanced infectivity of silver-selected populations relative to parallel control and ancestral populations under increasing ionic silver concentrations, and greater suppression of E. coli growth in standard medium. Furthermore, silver resistance evolved without loss of thermal or pH stability under the conditions tested. The genomic foundation of silver resistance was relatively simple, with positive and negative natural selection differentiating the silver-selected populations from the controls and ancestral populations across serial passages in silver. Support for replication-associated adaptation under ionic silver selection may be reflected in recurrent mutations identified in genes involved in transcription, DNA replication, and genome maintenance, including T7p07 (RNA polymerase), T7p10 (DNA ligase), and T7p29 (DNA polymerase I). These findings highlight the importance of evaluating phage –silver combination strategies within an evolutionary framework that accounts for the adaptive capacity of bacteriophages under silver selection. Full article

Background: Escherichia coli remains a critical multidrug-resistant nosocomial pathogen, driving interest in bacteriophage-based biocontrol. The genus Kayfunavirus (family Autotranscriptaviridae) exhibits obligately lytic replication cycles and favorable biosafety profiles, yet each new phage requires comprehensive genomic characterization to expand therapeutic candidate pools. This study aimed to isolate and genomically characterize a novel Kayfunavirus from an environmental reservoir in Vietnam. Methods: Escherichia phage vB_EcoA-Sparklingdew was isolated from Can Tho River water using host E. coli AgE9. The genome was assembled using SPAdes. The termini were resolved with PhageTerm. The annotation was done via the Pharokka pipeline and HHpred. Taxonomic classification was performed using taxMyPhage, VIRIDIC intergenomic comparisons, and maximum likelihood phylogeny of concatenated structural proteins. Results: The complete genome comprises a 37,944 bp linear dsDNA molecule (49.9% GC), encoding 51 open reading frames in a predominantly unidirectional arrangement. Key features include a virion-encoded T7-like RNA polymerase, a 723-residue T7-like DNA polymerase, a canonical lysis triad, and two putative tailspike proteins. A 212 bp direct terminal repeat and coverage profiles support a headful (pac) packaging mechanism. Comprehensive screening confirmed the absence of lysogeny, virulence, and antibiotic resistance determinants. A single synonymous SNP indicated high clonal purity. Intergenomic identity peaked at 87.7% against ICTV references, confirming placement in a novel species. Conclusions: Phage Sparklingdew represents a strictly lytic Kayfunavirus with a compact genomic architecture. Its favorable safety profile and absence of temperate markers support further evaluation for targeted therapeutic applications against pathogenic E. coli. Full article