Search Results (37)

Listeria monocytogenes is a life-threatening bacterial foodborne pathogen that can persist in food-processing facilities for years. Although phages can control L. monocytogenes during food production, phage-resistant bacterial subpopulations can regrow in phage-treated environments. In this study, an L. monocytogenes hly defective strain, NJ05-Δhly, was produced, which considerably regulated the interactions between L. monocytogenes and phages. Specifically, we observed a 76.92-fold decrease in the efficiency of plating of the defective strain following infection with the Listeria phage vB-LmoM-NJ05. The lytic effect was notably diminished at multiplicities of infection of 1 and 10. Furthermore, the inactivation of LLO impaired biofilm formation, which was completely suppressed and eliminated following treatment with 10⁸ PFU/mL of phage. Additionally, phages protected cells from mitochondrial membrane damage and the accumulation of mitochondrial reactive oxygen species induced by L. monocytogenes invasion. Transcriptomic analysis confirmed these findings, revealing the significant downregulation of genes associated with phage sensitivity, pathogenicity, biofilm formation, and motility in L. monocytogenes. These results underscore the vital role of LLO in regulating the pathogenicity, phage susceptibility, and biofilm formation of L. monocytogenes. These observations highlight the important role of virulence factors in phage applications and provide insights into the potential use of phages for developing biosanitizers. Full article

Background: Intracellular bacteria frequently result in chronic and recurrent infections. MRSA is one of the most prevalent facultative intracellular bacteria in clinical infections. The drug resistance of MRSA and the difficulty of most antibiotics in entering cells result in a suboptimal clinical efficacy of antibiotics in the treatment of intracellular MRSA. Bacteriophages represent a promising alternative therapy in the context of the current antimicrobial resistance crisis. Nevertheless, the low efficiency of phage entry into cells and their rapid inactivation remain challenges in the treatment of intracellular MRSA using phages. The utilization of functionalized carriers for the delivery of phages into cells and their protection represents a feasible strategy. Methods: In this study, a new MRSA bacteriophage (vB_SauS_PHM) was isolated from hospital sewage, exhibiting the characteristics of short incubation period, large lytic amount, and good environmental tolerance. Subsequently, vB_SauS_PHM was encapsulated by TAT peptide-functionalized liposomes through microfluidic technology and size-exclusion chromatography (SEC), forming a phage delivery system, designated TAT-Lip@PHM. Results: The encapsulation rate of the phage by TAT-Lip@PHM was 20.3%, and the cell entry efficiency was ≥90% after 8 h. The 24 h eradication rate of 300 μg/mL TAT-Lip@PHM against intracellular MRSA was 94.05% (superior to the 21.24% and 44.90% of vB_SauS_PHM and Lip@PHM, respectively), while the mammalian cell activity was >85% after 24 h incubation. Conclusions: The TAT-Lip@PHM effectively delivered the phage into the cell and showed an excellent killing effect on intracellular MRSA with low cytotoxicity. This work provides a technical reference for the application of phages in the treatment of intracellular bacterial infection. Full article

The emergence of antimicrobial resistance among foodborne pathogens has intensified the search for alternative biocontrol strategies. Among these, essential oils (EOs) and bacteriophages have gained increasing attention, due to their natural origin and antimicrobial potential. This narrative review investigates their individual and combined use as innovative tools for improving food safety. We discuss the mechanisms of action, current food applications, and regulatory or technical limitations associated with both EOs and phages. Particular emphasis is placed on their complementary characteristics, which may enhance efficacy when used together. An in-depth analysis of five key studies investigating synergistic EO–phage combinations against Staphylococcus aureus, Escherichia coli, and Salmonella Typhimurium is presented. These studies, conducted in both in vitro and food-based systems, reveal that antimicrobial synergy is often dose- and temperature-dependent. Optimized combinations lead to enhanced bacterial reduction and reduced resistance development. However, several challenges remain, including sensory alterations in food products, phage inactivation by EO compounds, and host cell destruction at high EO doses. The review concludes that while EOs and phages face limitations when applied independently, their strategic combination shows substantial promise. Future research should focus on formulation development, delivery systems, and regulatory alignment to unlock their full synergistic potential. Full article

The development of vaccines against RNA viruses has undergone a rapid evolution in recent years, particularly driven by the COVID-19 pandemic. This review examines the key roles that RNA viruses, with their high mutation rates and zoonotic potential, play in fostering vaccine innovation. We also discuss both traditional and modern vaccine platforms and the impact of new technologies, such as artificial intelligence, on optimizing immunization strategies. This review evaluates various vaccine platforms, ranging from traditional approaches (inactivated and live-attenuated vaccines) to modern technologies (subunit vaccines, viral and bacterial vectors, nucleic acid vaccines such as mRNA and DNA, and phage-like particle vaccines). To illustrate these platforms’ practical applications, we present case studies of vaccines developed for RNA viruses such as SARS-CoV-2, influenza, Zika, and dengue. Additionally, we assess the role of artificial intelligence in predicting viral mutations and enhancing vaccine design. The case studies underscore the successful application of RNA-based vaccines, particularly in the fight against COVID-19, which has saved millions of lives. Current clinical trials for influenza, Zika, and dengue vaccines continue to show promise, highlighting the growing efficacy and adaptability of these platforms. Furthermore, artificial intelligence is driving improvements in vaccine candidate optimization and providing predictive models for viral evolution, enhancing our ability to respond to future outbreaks. Advances in vaccine technology, such as the success of mRNA vaccines against SARS-CoV-2, highlight the potential of nucleic acid platforms in combating RNA viruses. Ongoing trials for influenza, Zika, and dengue demonstrate platform adaptability, while artificial intelligence enhances vaccine design by predicting viral mutations. Integrating these innovations with the One Health approach, which unites human, animal, and environmental health, is essential for strengthening global preparedness against future RNA virus threats. Full article

Bacteria from genus Vibrio continue to be one of the most common threats to aquaculture sustainability. Vibrio spp. have been associated with infectious outbreaks in fish, shrimp, bivalves and even algae farms worldwide. Moreover, several Vibrio spp. are also pathogens that impact human health and are a threat to public health when transferred to consumers through contaminated seafood products. The use of bacteriophages is an evolving technology that could be applied in the treatment of Vibrio spp. either to protect aquaculture farms or to decontaminate seafood, namely bivalves during their depuration. In the present study, bacteriophages vB_VpS_LMAVpS1 (S1) vB_VpS_LMAVpVPP (VPP), vB_VpS_LMAVpSH (SH) and vB_VpS_LMAVpH (H) infecting V. parahaemolyticus were isolated and characterized. All phages presented fast adsorption rates and were able to control V. parahaemolyticus at all multiplicity of infections (MOIs) tested (MOI of 1, 10 and 100), with reductions of more than 4 log CFU/mL being recorded, but only in the presence of divalent cation calcium. The rate of emergence of phage-resistant mutants was very low (1.8 × 10⁻⁶ to 3.1 × 10⁻⁶). Bacterial phage resistance was not permanent and led to a loss of bacterial fitness. All four phages presented with lysins encoded in their genomes. The results presented provide valuable insights for future studies in the application of these bacteriophages in different scenarios to control, decontaminate or treat bacterial infections or contaminations of V. parahaemolyticus. Full article

Pseudomonas aeruginosa is a common cause of hospital-acquired infections and exhibits a strong resistance to antibiotics. An alternative treatment option for bacterial infections is the use of bacteriophages (or phages). In this study, two distinct phages, VB_PaD_phPA-G (phPA-G) and VB_PaN_phPA-Intesti (phPA-Intesti), were used as single suspensions or in a phage cocktail to inactivate the planktonic cells and biofilms of P. aeruginosa. Preliminary experiments in culture medium showed that phage phPA-Intesti (reductions of 4.5–4.9 log CFU/mL) outperformed phPA-G (reductions of 0.6–2.6 log CFU/mL) and the phage cocktail (reduction of 4.2 log CFU/mL). Phage phPA-Intesti caused a maximum reduction of 5.5 log CFU/cm² in the P. aeruginosa biofilm in urine after 4 h of incubation. The combination of phage phPA-Intesti and ciprofloxacin did not improve the efficacy of bacterial inactivation nor reduce the development of resistant mutants. However, the development of resistant bacteria was lower in the combined treatment with the phage and the antibiotic compared to treatment with the antibiotic alone. This phage lacks known toxins, virulence, antibiotic resistance, and integrase genes. Overall, the results suggest that the use of phage phPA-Intesti could be a potential approach to control urinary tract infections (UTIs), namely those caused by biofilm-producing and multidrug-resistant strains of P. aeruginosa. Full article

Cold plasma is a promising alternative for water treatment owing to pathogen control and a plethora of issues in the agriculture and food sectors. Shellfish pose a serious risk to public health and are linked to large viral and bacterial outbreaks. Hence, current European regulations mandate a depuration step for shellfish on the basis of their geographical growth area. This study investigated the inactivation of relevant viral and bacterial pathogens of three plasma-activated seawaters (PASWs), and their reactive oxygen and nitrogen species (RONS) composition, as being primarily responsible for microbial inactivation. Specifically, F-specific (MS2) and somatic (φ174) bacteriophage, cultivable surrogate (murine norovirus, MNV, and Tulane virus, TV), and human norovirus (HuNoV GII.4) inactivation was determined using plaque counts and infectivity assays, including the novel human intestinal enteroid (HIE) model for HuNoV. Moreover, the kinetic decay of Escherichia coli, Salmonella spp., and Vibrio parahaemolyticus was characterized. The results showed the complete inactivation of phages (6–8 log), surrogates (5–6 log), HuNoV (6 log), and bacterial (6–7 log) pathogens within 24 h while preventing cytotoxicity effects and preserving mussel viability. Nitrites (NO₂⁻) were found to be mostly correlated with microbial decay. This research shows that PASWs are a suitable option to depurate bivalve mollusks and control the biohazard risk linked to their microbiological contamination, either viral or bacterial. Full article

Numerous treatment modalities have been employed over the years to eradicate bacterial communities in industrial wastewater. Oxidizing agents and chemical additives, such as ozone, permanganate, glutaraldehyde, and chlorine, are effective in treating microbial contaminants that are typically found in domestic wastewater. However, the chemical complexity of water produced from fracking requires novel approaches, because the microbes have developed mechanisms to overcome typical disinfectants. In this work, we test the effectiveness of bacteriophages for the eradication of two model bacteria from produced water: Pseudomonas aeruginosa and Bacillus megaterium. These bacteria were grown in low salinity produced water and exposed to their corresponding phage. Overall, the total inactivation of the P. aeruginosa population was achieved, as well as the inactivation of B. megaterium. These promising results provide a potentially useful tool for bacterial elimination in overall PW treatment, at an industrial scale. Particularly, since phage treatment is a rapid and cost-effective alternative. Moreover, these results fall within the objectives proposed as part of the sustainable development goals adopted worldwide. Full article

We analyzed metagenome data of feces from sows at different physiological periods reared on large-scale farms in Anhui Province, China, to provide a better understanding of the microbial diversity of the sow intestinal microbiome and the structure of antibiotic-resistance genes (ARGs) and virulence genes it carries. Species annotation of the metagenome showed that in the porcine intestinal microbiome, bacteria were dominant, representing >97% of the microorganisms at each physiological period. Firmicutes and Proteobacteria dominated the bacterial community. In the porcine gut microbiome, the viral component accounted for an average of 0.65%, and the species annotation results indicated that most viruses were phages. In addition, we analyzed the microbiome for ARGs and virulence genes. Multidrug-like, MLS-like, and tetracycline-like ARGs were most abundant in all samples. Evaluation of the resistance mechanisms indicated that antibiotic inactivation was the main mechanism of action in the samples. It is noteworthy that there was a significant positive correlation between ARGs and the total microbiome. Moreover, comparative analysis with the Virulence Factor Database showed that adhesion virulence factors were most abundant. Full article

The bacterial adhesion to food processing surfaces is a threat to human health, as these surfaces can serve as reservoirs of pathogenic bacteria. Escherichia coli is an easily biofilm-forming bacterium involved in surface contamination that can lead to the cross-contamination of food. Despite the application of disinfection protocols, contamination through food processing surfaces continues to occur. Hence, new, effective, and sustainable alternative approaches are needed. Bacteriophages (or simply phages), viruses that only infect bacteria, have proven to be effective in reducing biofilms. Here, phage phT4A was applied to prevent and reduce E. coli biofilm on plastic and stainless steel surfaces at 25 °C. The biofilm formation capacity of phage-resistant and sensitive bacteria, after treatment, was also evaluated. The inactivation effectiveness of phage phT4A was surface-dependent, showing higher inactivation on plastic surfaces. Maximum reductions in E. coli biofilm of 5.5 and 4.0 log colony-forming units (CFU)/cm² after 6 h of incubation on plastic and stainless steel, respectively, were observed. In the prevention assays, phage prevented biofilm formation in 3.2 log CFU/cm² after 12 h. Although the emergence of phage-resistant bacteria has been observed during phage treatment, phage-resistant bacteria had a lower biofilm formation capacity compared to phage-sensitive bacteria. Overall, the results suggest that phages may have applicability as surface disinfectants against pathogenic bacteria, but further studies are needed to validate these findings using phT4A under different environmental conditions and on different materials. Full article

Therapeutic application of bacterial viruses (phage therapy) has in recent years been rediscovered by many scientists, as a method which may potentially replace conventional antibacterial strategies. However, one of the main problems related to phage application is the stability of bacterial viruses. Though many techniques have been used to sustain phage activity, novel tools are needed to allow long-term phage storage and application in versatile forms. In this study, we combined two well-known methods for bacteriophage immobilization. First, encapsulated phages were obtained by means of extrusion–ionic gelation, and then alginate microspheres were dried using the lyophilization process (freeze-drying). To overcome the risk of phage instability upon dehydration, the microspheres were prepared with the addition of 0.3 M mannitol. Bacteriophage-loaded microspheres were stored at room temperature for 30 days and subsequently exposed to simulated gastric fluid (SGF). The survival of encapsulated phages after drying was significantly higher in the presence of mannitol. The highest number of viable bacteriophages exceeding 4.8 log₁₀ pfu/mL in SGF were recovered from encapsulated and freeze-dried microspheres, while phages in lyophilized lysate were completely inactivated. Although the method requires optimization, it may be a promising approach for the immobilization of bacteriophages in terms of practical application. Full article

Phage characterization for research and therapy can involve newly isolated phages propagated in pathogenic bacteria. If so, characterization requires safety-managing the bacteria. In the current study, we adapt a common and inexpensive reagent, PrimeStore (Longhorn Vaccines and Diagnostics, San Antonio, TX, USA), to safety-manage bacteria in 20 min by selectively inactivating the bacteria. No bacterial survivors are observed among >10⁹ bacteria per ml for a representative of both Gram-negative bacteria (Escherichia coli) and Gram-positive bacteria (Bacillus thuringiensis). This procedure causes no detected inactivation of podophage T3, myophage T4 and siphophage 0105phi7-2. Margins of safety for PrimeStore concentration exist for bacterial inactivation and phage non-inactivation. Thus, general applicability is expected. Subsequent dialysis is used to block long-term effects on phages. Nonetheless, comparable tests should be performed for each pathogenic bacterial strain/phage. Electron microscopy of thin sections reveals inactivation-altered bacterial cytoplasm and a non-disintegrated bacterial envelope (ghosts). Ghosting of E. coli includes re-arrangement of the cytoplasm and the release of endotoxin. The activity of the released endotoxin is >99% reduced after subsequent dialysis, which also removes PrimeStore components. Ghosting of B. thuringiensis includes apparent phase separation within the cytoplasm. The primary application envisaged is biophysical and other screening of phages for therapy of infectious disease. Full article

The spoilage of juices by Alicyclobacillus spp. remains a serious problem in industry and leads to economic losses. Compounds such as guaiacol and halophenols, which are produced by Alicyclobacillus, create undesirable flavors and odors and, thus, decrease the quality of juices. The inactivation of Alicyclobacillus spp. constitutes a challenge because it is resistant to environmental factors, such as high temperatures, and active acidity. However, the use of bacteriophages seems to be a promising approach. In this study, we aimed to isolate and comprehensively characterize a novel bacteriophage targeting Alicyclobacillus spp. The Alicyclobacillus phage strain KKP 3916 was isolated from orchard soil against the Alicyclobacillus acidoterrestris strain KKP 3133. The bacterial host’s range and the effect of phage addition at different rates of multiplicity of infections (MOIs) on the host’s growth kinetics were determined using a Bioscreen C Pro growth analyzer. The Alicyclobacillus phage strain KKP 3916, retained its activity in a wide range of temperatures (from 4 °C to 30 °C) and active acidity values (pH from 3 to 11). At 70 °C, the activity of the phage decreased by 99.9%. In turn, at 80 °C, no activity against the bacterial host was observed. Thirty minutes of exposure to UV reduced the activity of the phages by almost 99.99%. Based on transmission-electron microscopy (TEM) and whole-genome sequencing (WGS) analyses, the Alicyclobacillus phage strain KKP 3916 was classified as a tailed bacteriophage. The genomic sequencing revealed that the newly isolated phage had linear double-stranded DNA (dsDNA) with sizes of 120 bp and 131 bp and 40.3% G+C content. Of the 204 predicted proteins, 134 were of unknown function, while the remainder were annotated as structural, replication, and lysis proteins. No genes associated with antibiotic resistance were found in the genome of the newly isolated phage. However, several regions, including four associated with integration into the bacterial host genome and excisionase, were identified, which indicates the temperate (lysogenic) life cycle of the bacteriophage. Due to the risk of its potential involvement in horizontal gene transfer, this phage is not an appropriate candidate for further research on its use in food biocontrol. To the best of our knowledge, this is the first article on the isolation and whole-genome analysis of the Alicyclobacillus-specific phage. Full article

Bacteriophages (phages) are viruses that are the natural predators of bacteria and highly abundant in human milk and the infant gut microbiome. However, the effect of pasteurisation on human milk phages is unknown. This study, therefore, assessed the effect of holder pasteurisation (HP) and UV-C irradiation (UV) on exogenous bacteriophages inoculated into human milk. Ten donor human milk samples inoculated with a thermotolerant Escherichia coli phage (T4) and a thermosensitive Staphylococcus aureus phage (BYJ20) were subjected to HP and UV treatments. We found that UV effectively inactivated both phages (8/10 samples; 80%), however, HP was ineffective against the thermotolerant T4 phages (0/10; 0% inactivated). This is the first study to assess the impact of UV and HP methods on the viability of human milk phages. This pilot data suggests that HP methods used by milk banks likely destroy thermosensitive, but not thermotolerant, phages, with implications for early-life virome and bacterial microbiome assembly in donor milk fed infants. Full article

Phage therapy of ventilator-associated pneumonia (VAP) is of great interest due to the rising incidence of multidrug-resistant bacterial pathogens. However, natural or therapy-induced immunity against therapeutic phages remains a potential concern. In this study, we investigated the innate and adaptive immune responses to two different phage cocktails targeting either Pseudomonas aeruginosa or Escherichia coli—two VAP-associated pathogens—in naïve mice without the confounding effects of a bacterial infection. Active or UV-inactivated phage cocktails or buffers were injected intraperitoneally daily for 7 days in C57BL/6J wild-type mice. Blood cell analysis, flow cytometry analysis, assessment of phage distribution and histopathological analysis of spleens were performed at 6 h, 10 days and 21 days after treatment start. Phages reached the lungs and although the phage cocktails were slightly immunogenic, phage injections were well tolerated without obvious adverse effects. No signs of activation of innate or adaptive immune cells were observed; however, both active phage cocktails elicited a minimal humoral response with secretion of phage-specific antibodies. Our findings show that even repetitive injections lead only to a minimal innate and adaptive immune response in naïve mice and suggest that systemic phage treatment is thus potentially suitable for treating bacterial lung infections. Full article