Search Results (62)

Serratia marcescens is a nosocomial pathogen that has acquired resistance to multiple antibiotics, necessitating alternative antimicrobial strategies. The aim of this study was to isolate and characterize a novel phage (ØAS2) against Serratia marcescens and evaluate its biocontrol potential in dairy matrices. In this study, a lytic bacteriophage specific to S. marcescens, designated ØAS2, was isolated from sewage samples collected in Assiut, Egypt. Phage ØAS2 was characterized using plaque assays, transmission electron microscopy (TEM), host range determination, pH and thermal stability tests, and one-step growth curve analysis. Its ability to inhibit bacterial growth and disrupt biofilms was also evaluated in vitro. TEM revealed that ØAS2 possesses an icosahedral head approximately 47.2 nm in diameter and a very short tail, consistent with the morphology of a member of the class Caudoviricetes that exhibits podovirus-like morphology. The phage exhibited a broad host range, infecting various Serratia strains as well as other Gram-negative bacteria, including Klebsiella spp., Escherichia coli, Salmonella typhi, and Shigella spp. ØAS2 was thermostable up to 60 °C and showed maximum activity at pH 8. One-step growth curve analysis revealed a short latent period of 10 min and a burst size of 115 PFU per infected cell. ØAS2 effectively inhibited the growth of S. marcescens SM02 in vitro and significantly reduced preformed biofilms at different multiplicities of infection (MOIs). When applied to skim milk and fresh soft cheese at various MOIs (Multiplicities of Infection), the phage successfully controlled bacterial contamination under refrigerated storage (7 °C for 7–10 days). At MOI 5.0, phage ØAS2 reduced biofilm biomass by 25.6%, planktonic growth by 85.7%, and achieved a reduction of 2.1 log₁₀ CFU/mL in skim milk. These findings indicate that ØAS2 is a promising biocontrol candidate for managing S. marcescens spoilage in dairy products. Full article

Background: The global rise in multidrug-resistant (MDR) Gram-negative bacteria (GNB) poses an urgent challenge for infection control and antibiotic stewardship. Among these, Klebsiella pneumoniae is a major cause of hospital-acquired infections and is listed as a critical priority pathogen by the World Health Organization. Peru reports an exceptionally high prevalence of MDR K. pneumoniae, underscoring the need for innovative antimicrobial approaches. Methods: Here, we describe the isolation and characterization of lytic Klebsiella bacteriophage from sewage samples collected from the Huatanay River (Cusco, Peru) in 2023. Phages were isolated using the reference strain MDR K. pneumoniae ATCC BAA-2814. Then, they were screened against 50 clinical MDR K. pneumoniae strains. Results: The phage H33IIK demonstrated effective antibacterial capability, showing strict host specificity for K. pneumoniae, thermal stability, moderate pH tolerance, and high burst size. Whole-genome sequencing analysis classified it within the class Caudoviricetes, family Ackermannviridae, and genus Taipeivirus. The genomic analysis confirmed the absence of lysogeny-related, antimicrobial resistance, and virulence genes, supporting its suitability and safety for potential biotechnological applications. Conclusions: These findings highlight phage H33IIK as a lytic agent effective against MDR K. pneumoniae. It could contribute to the development of phage-based approaches to combat MDR GNB. Full article

The global surge in multidrug-resistant (MDR) bacterial pathogens has created an urgent imperative for innovative antimicrobial strategies. Enterococcus faecalis, Staphylococcus aureus, and Acinetobacter baumannii demonstrate remarkable antibiotic resistance and dominate hospital-acquired infections. These bacteria often form biofilms, a complex community structure that shields them from immune system phagocytosis, resists antibiotic penetration, and enhances their survival in hostile environments. In clinical cases, these bacteria often form mixed biofilms and lead to treatment failures. Phages and their derivatives have emerged as promising candidates in the fight against drug-resistant bacteria. Lys22, an endolysin derived from an enterococcus phage, has been cloned and demonstrated to possess a broad host range, effectively targeting E. faecalis, various Staphylococcus species, and A. baumannii. When applied to the biofilms formed by these bacteria, Lys22 was found to significantly inhibit both simple and complex biofilms in vitro. Virulent genes, including agrA, sarA, and icaA in S. aureus; asa1, cylA, and gelE in E. faecalis; and OmpA and lpsB in A. baumannii were also downregulated by Lys22. Notably, Lys22 also exhibited a robust protective effect against dual or triple infections involving E. faecalis, S. aureus, and A. baumannii in a zebrafish embryos model, highlighting its potential as a therapeutic agent in combatting multi-bacterial infections. Full article

The increasing incidence of hospital-acquired infections and antimicrobial-resistant pathogens poses a major clinical challenge. Nearly all medical devices are vulnerable to bacterial biofilm formation, which acts as a protective coating against the host defense systems and antibiotics. The persistence of biofilm infections, accounting for around 65% of all microbial infections, and poor conventional treatment outcomes has driven interest in alternative approaches like bacteriophage therapy. This review encompasses key aspects of biofilm biology, taking into account the clinically significant ESKAPE pathogens, and provides an in-depth analysis of the role of phage agents in biofilm control as a new biofilm control strategy. Diving deeper into the mechanisms of phage-mediated processes, the review examines how bacteriophages penetrate and disrupt biofilm architecture and evaluates current therapeutic strategies that exploit these actions, acknowledging their limitations and considering possible future directions. Full article

Antibiotic-resistant Pseudomonas aeruginosa presents a critical global health challenge, particularly in hospital-acquired infections. Bacteriophages offer a promising therapeutic avenue due to their ability to target and lyse resistant strains. This study characterizes Pseudomonas phage Banzai, a newly isolated Pbunavirus (family Lindbergviridae) with lytic activity against multiple P. aeruginosa isolates, including multidrug-resistant strains. Genomic analysis revealed a 66,189 bp genome, lacking antibiotic resistance or virulence factors, and suggested a headful packaging mechanism and the presence of a bidirectional component in the replication. In vivo experiments using Galleria mellonella showed therapeutic potential, significantly improving larval survival (87% at 24 h). Host range analysis revealed activity against 13 of 30 P. aeruginosa isolates, including members of O1, O3, O5 and O6 in silico predicted serogroups. Phylogenomic analyses place phage Banzai within the genus Pbunavirus, sharing 94.8% intergenomic similarity with its closest relatives, supporting its classification as a novel species. These findings highlight phage Banzai as a potential candidate for phage therapy, demonstrating genomic stability, a strictly lytic lifestyle, and in vivo efficacy. Full article

Multidrug-resistant (MDR) bacterial infections present a major challenge in cancer therapy, particularly for immunocompromised patients undergoing chemotherapy, radiation, or surgery. These infections often arise from prolonged antibiotic use, hospital-acquired pathogens, and weakened immune defenses, leading to increased morbidity and mortality. As conventional antibiotics become less effective against MDR strains, there is an urgent need for alternative treatment options. This review highlights phage therapy as a promising approach to managing MDR bacterial infections in cancer patients. Once widely used, phage therapy has recently regained attention as a targeted antimicrobial strategy that can specifically eliminate harmful bacteria while preserving the beneficial microbiota. Phages work by directly lysing bacteria, disrupting biofilms, and synergizing with antibiotics to restore bacterial susceptibility. These mechanisms make phage therapy especially appealing for treating infections that complicate cancer treatments. However, the clinical application of phage therapy faces challenges such as variability in phage–host interactions, regulatory hurdles, and immune responses in patients. This review identifies gaps in current research regarding the use of phage therapy for MDR infections in cancer patients. By examining recent innovations, therapeutic mechanisms, and associated limitations, we provide valuable insights into the potential of phage therapy for improving infection management in oncology. Future research should focus on refining phage delivery methods, assessing long-term safety, and exploring combination therapies to maximize clinical efficacy. Overcoming these challenges could position phage therapy as a valuable complement to existing antimicrobial strategies in cancer care. Full article

Pseudomonas aeruginosa is a major pathogen associated with hospital-acquired infections, and the spread of carbapenem-resistant isolates highlights the urgency of developing non-conventional therapies, such as phage therapy. For this alternative to be effective, understanding phage–host interactions is crucial for the selection of candidate phages and offers new insights into these dynamics. Background/Objectives: This study aimed to characterize prophage diversity in clinical P. aeruginosa genomes, assess the relationship between phages and the CRISPR/Cas system, and investigate the potential role of prophages in disseminating resistance genes. Methods: A total of 141 genomes from Brazilian hospitals were analyzed. Prophage detection was performed using VIBRANT, and in silico analyses were conducted to evaluate taxonomic diversity, the presence of resistance genes, phage life cycle, genomic distribution, and the presence of the CRISPR/Cas system. Results: A total of 841 viral sequences were identified by the VIBRANT tool, of which 498 were confirmed by CheckV, with a predominance of the class Caudoviricetes and high overall phage diversity. No statistically significant difference was observed in the number of prophages between isolates with and without CRISPR/Cas systems. Prophages carrying resistance genes such as rsmA, OXA-56, SPM-1, and others were detected in isolates harboring the type I-C CRISPR/Cas system. Additionally, prophages showed no preference for specific insertion sites along the bacterial genome. Conclusions: These findings provide evidence of a well-established phage–host relationship. The dual role of prophages—as vectors of antimicrobial resistance and as potential therapeutic agents—reflects their dynamic impact on bacterial communities and reinforces their importance in developing new strategies to combat antimicrobial resistance. Full article

Pseudomonas aeruginosa is associated with both community and hospital-acquired infections. It colonizes the lungs of cystic fibrosis (CF) patients, establishing an ecological niche where it adapts and evolves from early to chronic stages, resulting in deteriorating lung function and frequent exacerbations. With antibiotics resistance on the rise, there is a pressing need for alternative personalized treatments (such as bacteriophage therapy) to combat P. aeruginosa infections. In this study, we aimed to isolate and characterize phages targeting both early and chronic P. aeruginosa isolates and evaluate their potential for phage therapy. Four highly virulent phages belonging to myoviral, podviral, and siphoviral morphotypes were isolated from sewage samples. These phages have a broad host range and effectively target 62.5% of the P. aeruginosa isolates with a positive correlation to the early isolates. All the phages have a virulence index of ≥0.90 (0.90–0.98), and one has a large burst size of 331 PFU/cell and a latency period of 30 min. All phages are stable under a wide range of temperature and pH conditions. Genomic analysis suggests the four phages are strictly lytic and devoid of identifiable temperate phage repressors and genes associated with antibiotic resistance and virulence. More significantly, two of the phages significantly delayed the onset of larval death when evaluated in a lethal Galleria mellonella infection model, suggesting their promise as phage therapy candidates for P. aeruginosa infections. Full article

Background/Objectives: Antibiotic resistance in pathogenic bacteria poses a critical global health threat, with multidrug-resistant (MDR) strains increasingly undermining conventional treatments. Among these, Pseudomonas aeruginosa is a high-priority pathogen due to its resistance to carbapenems and frequent presence in hospital settings, contributing to severe healthcare-associated infections. This study aimed to isolate and characterize novel bacteriophages from environmental wastewater samples that could specifically target MDR P. aeruginosa. Methods: Two bacteriophages, M8-2 and M8-3, were isolated from wastewater in Monterrey, Mexico. A genomic analysis classified M8-2 and M8-3 within the Caudoviridae family, and next-generation sequencing (NGS) was used to confirm the absence of undesirable antibiotic resistance or virulence genes. Optimization of viral amplification was performed to achieve high titers, with structural proteins characterized by SDS-PAGE. Results: Phages M8-2 and M8-3 exhibited specific lytic activity against MDR strains of P. aeruginosa, offering a targeted approach to combat antibiotic-resistant infections. High genetic similarity (>95%) to known Gram-negative bacterial phages was observed. Optimized viral amplification yielded titers of 4.2 × 10⁷ and 1.03 × 10⁹ PFUs/mL for M8-2 and M8-3, respectively. The specificity of these phages minimized disruption to the host microbiome, and their significant efficacy in suppressing bacterial growth positions bacteriophages as promising candidates for localized and personalized phage therapy, especially in chronic and hospital-acquired infection settings. Conclusions: These findings highlight the therapeutic potential of M8-2 and M8-3 in addressing antibiotic-resistant P. aeruginosa infections. Their safety profile, high target specificity, and robust lytic activity underscore the feasibility of incorporating phage-based strategies into current antimicrobial protocols. This study contributes to the broader goal of developing sustainable and effective phage therapies for diverse clinical and environmental contexts. Full article

Pseudomonas aeruginosa is a bacteria responsible for many hospital-acquired infections. Phages are promising alternatives for treating P. aeruginosa infections, which are often intrinsically resistant. The combination of phage and antibiotics in clearing bacterial infection holds promise due to increasing reports of enhanced effectiveness when both are used together. The aim of the study is to isolate and characterize a novel P. aeruginosa phage and determine its effectiveness in in vitro combination with antibiotics in controlling P. aeruginosa. In this study, a novel jumbo myophage HPP-Temi infecting P. aeruginosa Pa9 (PP334386) was isolated from household sewage. Electron micrographs of the phage were obtained to determine the morphological features of HPP-Temi virions. Complete genome analysis and a combination of Pseudomonas phage HPP-Temi with antibiotics were examined. The phage HPP-Temi was able to productively infect P. aeruginosa ATCC 9027 but was unable to infect a closely related genus. The phage was stable at 4–37 °C, 0.5% NaCl, and pH 8 for at least one hour. The HPP-Temi genome is a 302,719-bp-long dsDNA molecule with a GC content of 46.46%. The genome was predicted to have 436 ORFs and 7 tRNA genes. No virulence factor-related genes, antimicrobial resistance, or temperate lifestyle-associated genes were found in the phage HPP-Temi genome. Phage HPP-Temi is most closely related to the known or tentative representatives of the Pawinskivirus genus and can be proposed as a representative for the creation of a novel phage species in that genus. The phage and antibiotics (Ciprofloxacin) combination at varying phage titers (10³, 10⁶, 10⁹) were used against P. aeruginosa Pa9 (PP334386) at 3.0 × 10⁸ CFU/mL, which was carried out in triplicate. The result showed that combining antibiotics with phage significantly reduced the bacteria count at 10³ and 10⁶ titers, while no growth was observed at 10⁹ PFU/mL. This suggests that the effect of phage HPP-Temi in combination with antibiotics is a potential and promising agent for the control of P. aeruginosa infections. 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

Klebsiella pneumoniae, a member of the ESKAPE pathogen group, is a prominent cause of hospital-acquired infections. The WHO has recognized carbapenem-resistant K. pneumoniae as a critical-one priority pathogen. These resilient superbugs have the ability to form biofilms and present a significant global threat. In the present study, we isolated and characterized a bacteriophage SAKp02, from hospital sewage, infectious to carbapenem-resistant K. pneumoniae patient isolates. SAKp02 could infect 43 of 72 clinical isolates, indicating a broad host spectrum. Whole genome analysis classified SAKp02 within the family Casjensviridae, with a 59,343 bp genome encoding 82 ORFs. Comparative genomic analysis revealed significant differences between SAKp02 and its closest viruses, indicating a distinct genetic makeup positioning it as a novel phage strain within the lineage. The SAKp02 genome comprises bacteriolytic enzymes, including holin, endolysin, and phage depolymerase, crucial for bacterial lysis and biofilm disruption. It reduced biofilm biomass by over threefold compared to the control and eradicated 99% of viable cells within a 4 h treatment period. Scanning electron microscopy corroborated the ability of the phage to dismantle biofilm matrices and lyse bacterial cells. Safe and effective treatments are warranted, and hence, the fully characterized lytic phages with therapeutic potential against drug-resistant clinical isolates of bacteria are needed. Our study is the first to report the antibacterial and antibiofilm activity of Casjensviridae phages, and our discovery of a novel K. pneumoniae phage broadens the arsenal against the bacteria. Full article

Infections due to antimicrobial-resistant bacteria have become a major threat to global health. Some patients may carry resistant bacteria in their gut microbiota. Specific risk factors may trigger the conversion of these carriages into infections in hospitalized patients. Preventively eradicating these carriages has been postulated as a promising preventive intervention. However, previous attempts at such eradication using oral antibiotics or probiotics have led to discouraging results. Phage therapy, the therapeutic use of bacteriophage viruses, might represent a worthy alternative in this context. Taking inspiration from this clinical challenge, we built Gut-On-A-Chip (GOAC) models, which are tridimensional cell culture models mimicking a simplified gut section. These were used to better understand bacterial dynamics under phage pressure using two relevant species: Pseudomonas aeruginosa and Escherichia coli. Model mucus secretion was documented by ELISA assays. Bacterial dynamics assays were performed in GOAC triplicates monitored for 72 h under numerous conditions, such as pre-, per-, or post-bacterial timing of phage introduction, punctual versus continuous phage administration, and phage expression of mucus-binding properties. The potential genomic basis of bacterial phage resistance acquired in the model was investigated by variant sequencing. The bacterial “escape growth” rates under phage pressure were compared to static in vitro conditions. Our results suggest that there is specific bacterial prosperity in this model compared to other in vitro conditions. In E. coli assays, the introduction of a phage harboring unique mucus-binding properties could not shift this balance of power, contradicting previous findings in an in vivo mouse model and highlighting the key differences between these models. Genomic modifications were correlated with bacterial phage resistance acquisition in some but not all instances, suggesting that alternate ways are needed to evade phage predation, which warrants further investigation. Full article

Escherichia coli (E. coli) is one of the most common sources of infection in humans and animals. The emergence of E. coli which acquires resistance to various antibiotics has made treatment difficult. Bacteriophages can be considered promising agents to expand the options for the treatment of antibiotic-resistant bacteria. This study describes the isolation and characterization of Escherichia phage KIT06, which can infect E. coli resistant to the quinolone antibiotic nalidixic acid. Phage virions possess an icosahedral head that is 93 ± 8 nm in diameter and a contractile tail (116 ± 12 nm × 13 ± 5 nm). The phage was found to be stable under various thermal and pH conditions. A one-step growth curve showed that the latent time of the phage was 20 min, with a burst size of 28 particles per infected cell. Phage KIT06 infected 7 of 12 E. coli strains. It inhibited the growth of the host bacterium and nalidixic acid-resistant E. coli. The lipopolysaccharide and outer membrane proteins of E. coli, tsx and btuB, are phage receptors. Phage KIT06 is a new species of the genus Tequatrovirus with a genome of 167,059 bp consisting of 264 open reading frames (ORFs) that encode gene products related to morphogenesis, replication, regulation, and host lysis. The lack of genes encoding integrase or excisionase indicated that this phage was lytic. Thus, KIT06 could potentially be used to treat antibiotic-resistant E. coli using phage therapy. However, further studies are essential to understand its use in combination with other antimicrobial agents and its safe use in such applications. Full article

The Gram-negative ESKAPE bacterium Pseudomonas aeruginosa has become a pathogen of serious concern due its extensive multi-drug resistance (MDR) profile, widespread incidences of hospital-acquired infections throughout the United States, and high occurrence in wound infections suffered by warfighters serving abroad. Bacteriophage (phage) therapy has received renewed attention as an alternative therapeutic option against recalcitrant bacterial infections, both as multi-phage cocktails and in combination with antibiotics as synergistic pairings. Environmental screening and phage enrichment has yielded three lytic viruses capable of infecting the MDR P. aeruginosa strain PAO1. Co-administration of each phage with the carbapenem antibiotics ertapenem, imipenem, and meropenem generated enhanced overall killing of bacteria beyond either phage or drug treatments alone. A combination cocktail of all three phages was completely inhibitory to growth, even without antibiotics. The same 3× phage cocktail also disrupted PAO1 biofilms, reducing biomass by over 75% compared to untreated biofilms. Further, the phage cocktail demonstrated broad efficacy as well, capable of infecting 33 out of 100 diverse clinical isolate strains of P. aeruginosa. Together, these results indicate a promising approach for designing layered medical countermeasures to potentiate antibiotic activity and possibly overcome resistance against recalcitrant, MDR bacteria such as P. aeruginosa. Combination therapy, either by synergistic phage-antibiotic pairings, or by phage cocktails, presents a means of controlling mutations that can allow for bacteria to gain a competitive edge. Full article