Isolation Of Bacteriophages Research Articles

Isolation and characterisation of novel lytic bacteriophages for therapeutic applications in biofilm-associated prosthetic joint infections

Abstract Prosthetic joint infections are devastating complications of joint arthroplasties. Without effective management, they can lead to limb amputation and even death. A significant proportion of these infections is caused by the primarily commensal Coagulase-negative Staphylococci pathogens, which form thick, antibiotic-resistant biofilms at the site of infection. Combinatorial therapy involving antibiotics and bacteriophages may represent a strategy to overcome resistance. Previous research indicates that as bacteria develop resistance to antibiotics, they often become more susceptible to bacteriophages. In this study, we produced a cocktail of novel bacteriophages and assessed their viability to eradicate nosocomial staphylococcal biofilms. Here, we used clinical isolates from prosthetic joint infections to isolate and identify four new bacteriophages from sewage effluent. These novel phages were characterized through electron microscopy and full genome sequencing. Subsequently, we combined them into a phage cocktail, which effectively re-sensitized biofilms to vancomycin and flucloxacillin. Notably, this phage cocktail demonstrated low cytotoxicity in vitro to human epithelial cells, even when used alongside antibiotic treatments. These findings highlight the potential of the phage cocktail as a tool to increase antibiotic treatment success in prosthetic joint infections.

Novel Bacteriophage KG853 Exhibits Potent Lytic Activity and Biofilm Inhibition Against Pseudomonas aeruginosa

This study reports the isolation and characterization of a novel bacteriophage, KG853, specifically targeting Pseudomonas aeruginosa ATCC 27853. Morphological analysis using transmission electron microscopy revealed that bacteriophage KG853 belongs to the Bruynoghevirus genus. The phage demonstrated favorable characteristics for potential therapeutic applications, including a short latent period of 30 minutes and a large burst size of 136 plaque-forming units (PFU) per cell. KG853 exhibited stability across various temperatures and pH values, indicating its robustness under various environmental conditions. Genomic analysis showed that KG853 possesses a circular DNA genome of 45,390 base pairs with a GC content of 52.2%. No lysogenic or virulence genes were detected among the 84 open reading frames annotated in the genome, suggesting its safety for potential therapeutic use. Phylogenetic analysis revealed that phage KG853 is closely related to phage PaP3. Notably, KG853 demonstrated the ability to inhibit the formation of 4-hour biofilms by P. aeruginosa, a critical virulence factor in many infections. Host range analysis showed that KG853 is specific to P. aeruginosa, an important characteristic for targeted therapy. These findings suggest that bacteriophage KG853 represents a promising candidate for combating drug-resistant P. aeruginosa infections. Its specific host range, robust physical characteristics, lack of harmful genes, and anti-biofilm activity make it a potential alternative to conventional antibiotics. Further research is warranted to explore its efficacy in in vivo models and potential clinical applications.

Isolation and Antibiofilm Activity of Bacteriophages against Cutibacterium acnes from Patients with Periprosthetic Joint Infection

Isolation and Antibiofilm Activity of Bacteriophages against Cutibacterium acnes from Patients with Periprosthetic Joint Infection

A Vibrio cholerae anti-phage system depletes nicotinamide adenine dinucleotide to restrict virulent bacteriophages.

Bacteria and their predatory viruses (bacteriophages or phages) are in a perpetual molecular arms race. This has led to the evolution of numerous phage defensive systems in bacteria that are still being discovered, as well as numerous ways of interference or circumvention on the part of phages. Here, we identify a unique molecular battle between the classical biotype of Vibrio cholerae and virulent phages ICP1, ICP2, and ICP3. We show that classical biotype strains resist almost all isolates of these phages due to a 25-kb genomic island harboring several putative anti-phage systems. We observed that one of these systems, Nezha, encoding SIR2-like and helicase proteins, inhibited the replication of all three phages. Bacterial SIR2-like enzymes degrade the essential metabolic coenzyme nicotinamide adenine dinucleotide (NAD+), thereby preventing replication of the invading phage. In support of this mechanism, we identified one phage isolate, ICP1_2001, which circumvents Nezha by encoding two putative NAD+ regeneration enzymes. By restoring the NAD+ pool, we hypothesize that this system antagonizes Nezha without directly interacting with its proteins and should be able to antagonize other anti-phage systems that deplete NAD+.IMPORTANCEBacteria and phages are in a perpetual molecular arms race, with bacteria evolving an extensive arsenal of anti-phage systems and phages evolving mechanisms to overcome these systems. This study identifies a previously uncharacterized facet of the arms race between Vibrio cholerae and its phages. We identify an NAD+-depleting anti-phage defensive system called Nezha, potent against three virulent phages. Remarkably, one phage encodes proteins that regenerate NAD+ to counter the effects of Nezha. Without Nezha, the NAD+ regeneration genes are detrimental to the phage. Our study provides new insight into the co-evolutionary dynamics between bacteria and phages and informs the microbial ecology and phage therapy fields.

Exploration of Phage-Agrochemical Interaction Based on a Novel Potent Phage LPRS20-Targeting Ralstonia solanacearum.

Phage therapy has the potential to alleviate plant bacterial wilt. However, the knowledge gap concerning the phage-agrochemical interaction impedes the broader application of phages in agriculture. This study characterized a phage isolate and investigated its interactions with agrochemicals. A novel species within the Ampunavirus genus was proposed, serving phage LPRS20 as a type phage with a broad lytic range and significant antibacterial activity against Ralstonia solanacearum strains infecting tobacco, chili, or tomato. Sensory evaluation of the morphology of tobacco leaves suggested that phage application resulted in negligible harm to plants. Investigations into phage-agrochemical interactions revealed synergisms when LPRS20 was delivered 4 h before thiodiazole-copper as well as LPRS20 in combination with low-concentration berberine. Overall, our findings reveal that phage LPRS20 represents a novel, effective, and eco-friendly biocontrol agent against tobacco bacterial wilt in vivo and in vitro and contributes to the potential integration of phages and agrochemicals for controlling soil-borne pathogens.

Mycobacteriophages and Their Applications.

Mycobacterial infections caused by tuberculous and non-tuberculous strains pose significant treatment challenges, especially among immunocompromised patients. Conventional antibiotic therapies often fail due to bacterial resistance, highlighting the need for alternative therapeutic strategies. Mycobacteriophages are emerging as promising candidates for the treatment of mycobacteria. This review comprehensively explores phage isolation, characterization, and clinical applications. Despite the need for more extensive in vitro and in vivo studies, existing evidence shows their efficacy against both sensitive and antibiotic-resistant mycobacterial strains, even under disease-mimicking conditions, particularly when used in cocktails to minimize resistance development. Mycobacteriophages can be engineered and evolved to overcome limitations associated with lysogeny and narrow host range. Furthermore, they exhibit activity in ex vivo and in vivo infection models, successfully targeting mycobacteria residing within macrophages. Delivery methods such as bacterial and liposomal vectors facilitate their entry into human cells. Considering the potential for phage-treatment-induced bacterial resistance, as described in this review, the combination of mycobacteriophages with antibiotics shows efficacy in countering mycobacterial growth, both in the laboratory setting and in animal models. Interestingly, phage-encoded products can potentiate the activity of relevant antibiotics. Finally, the application of phages in different compassionate cases is reported. The positive outcomes indicate that phage therapy represents a promising solution for the treatment of antibiotic-resistant mycobacteria.

Isolation of bacteriophages as therapeutic agents for combating trauma-related infections in Ukraine

Isolation of bacteriophages as therapeutic agents for combating trauma-related infections in Ukraine

Isolation, characterization, and potential application of Acinetobacter baumannii phages against extensively drug-resistant strains.

One of the significant issues in treating bacterial infections is the increasing prevalence of extensively drug-resistant (XDR) strains of Acinetobacter baumannii. In the face of limited or no viable treatment options for extensively drug-resistant (XDR) bacteria, there is a renewed interest in utilizing bacteriophages as a treatment option. Three Acinetobacter phages (vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln) were identified from hospital sewage and analyzed for their morphology, host ranges, and their genome sequences were determined and annotated. These phages and vB_AbaS_SA1 were combined to form a phage cocktail. The antibacterial effects of this cocktail and its combinations with selected antimicrobial agents were evaluated against the XDR A. baumannii strains. The phages exhibited siphovirus morphology. Out of a total of 30 XDR A. baumannii isolates, 33% were sensitive to vB_AbaS_Ftm, 30% to vB_AbaS_Gln, and 16.66% to vB_AbaS_Eva. When these phages were combined with antibiotics, they demonstrated a synergistic effect. The genome sizes of vB_AbaS_Ftm, vB_AbaS_Eva, and vB_AbaS_Gln were 48487, 50174, and 50043 base pairs (bp), respectively, and showed high similarity. Phage cocktail, when combined with antibiotics, showed synergistic effects on extensively drug-resistant (XDR) strains of A. baumannii. However, the need for further study to fully understand the mechanisms of action and potential limitations of using these phages is highlighted.

Potential of training of anti-Staphylococcus aureus therapeutic phages against Staphylococcus epidermidis multidrug-resistant isolates is restricted by inter- and intra-sequence type specificity.

Phage therapy appears to be a promising approach to tackle multidrug-resistant bacteria, including staphylococci. However, most anti-staphylococcal phages have been characterized in Staphylococcus aureus, while a limited number of studies investigated phage activity against S. epidermidis. We studied the potential of phage training to extend the host range of two types of anti-S. aureus phages against S. epidermidis isolates. The Appelmans protocol was applied to a mixture of Kayvirus and a mixture of Silviavirus phages repeatedly exposed to seven S. epidermidis strains representative of nosocomial-associated sequence types (ST), including the world-wide disseminated ST2. We observed increased activity only for the Kayvirus mixture against two of these strains (ST2 or ST35). Phage subpopulations isolated from the training mixture using these two strains (five/strain) exhibited different evolved phenotypes, active only against their isolation strain or strains of the same ST. Of note, 16/47 ST2 strains were susceptible to one of the groups of trained phages. A comparative genomic analysis of ancestral and trained phage genomes, conducted to identify potential bacterial determinants of such specific activity, found numerous recombination events between two of the three ancestors. However, a small number of trained phage genes had nucleotide sequence modifications impacting the corresponding protein compared to ancestral phages, two to four of them per phage genome being specific of each group of phage subpopulations exhibiting different host range. The results suggest that anti-S. aureus phages can be adapted to S. epidermidis isolates but with inter- and intra-ST specificity.ImportanceS. epidermidis is increasingly recognized as a threat for public health. Its clinical importance is notably related to multidrug resistance. Phage therapy is one of the most promising alternative therapeutic strategies to antibiotics. Nonetheless, only very few phages active against this bacterial species have been described. In the present study, we showed that phage training can be used to extend the host range of polyvalent Kayvirus phages within the Staphylococcus genera to include S. epidermidis species. In the context of rapid development of phage therapy, in vitro forced adaptation of previously characterized phages could be an appealing alternative to fastidious repeated isolation of new phages to improve the therapeutic potential of a phage collection.

Bioconservation of hard rent cheese using bacteriophages

Several technological approaches are used to increase the safety of perishable food products during their production and realization. In particular, it is the use of temperature, high pressure, ultraviolet radiation and added preservatives. One of the generally recognized and common disadvantages of all these methods is that they have an influence on the microorganisms indiscriminately, that is, they inhibit both pathogenic and potentially beneficial autochthonous lactic acid bacteria of the "normal" microflora of fermented products. The purpose of this investigation was to monitor hard rennet cheeses for the content of Staphylococcus aureus and to develop a technology for its preservation using a lytic staphylococcal bacteriophage. Contamination of milk-raw material and Dutch cheese with Staphylococcus aureus was determined by inoculating the selected samples and carrying out tenfold dilutions on a selective hemoagar medium with 5% sodium chloride, followed by thermostating for +37-48 hours. Isolation of bacteriophages from milk raw materials was carried out by filtering through bacterial filters with pores of 0.45 μm. During the improvement of the technology of Dutch cheese with staphylococcal bacteriophage, two samples of cheese were produced - one experimental with bacteriophage, and the second control - without bacteriophage. It was set up that in milk-raw material, which is processed for the production of hard cheeses, Staphylococcus aureus was not detected in 25 to 40% of the samples, on average 45% of the milk samples were contaminated with this pathogen up to 5×102 CFU/ml and from 18 to 30% samples had a staphylococci content of more than 5×102 CFU/ml. In hard cheeses which are realized in the retail network, Staphylococcus aureus was not isolated in an average of 25% of the samples, while approximately 30% of the samples were contaminated with this microorganism in the amount of more than 5×102 CFU/g, which exceeds the requirements of the standard. Two bacteriophages were selected that showed 75.0-80.0% lytic activity against cultures of Staphylococcus aureus from milk and hard cheeses. These bacteriophages were used in the technology of preserving hard Dutch cheese. The technology of Dutch cheese with staphylococcal bacteriophage has been developed to increase its safety during implementation. The use of lytic bacteriophages for the biocontrol of Staphylococcus aureus in rennet cheese makes it possible to almost completely neutralize them in the technology of production. Therefore, the use of lytic bacteriophages to increase the safety of dairy products is an environmentally clean way of restraining the development of pathogenic bacteria, thereby not influencing on the fermentation microflora of the product.

RETRACTED: Teklemariam et al. Isolation and Characterization of a Novel Lytic Phage, vB_PseuP-SA22, and Its Efficacy against Carbapenem-Resistant Pseudomonas aeruginosa. Antibiotics 2023, 12, 497.

The journal retracts the article, "Isolation and Characterization of a Novel Lytic Phage, vB_PseuP-SA22, and Its Efficacy against Carbapenem-Resistant Pseudomonas aeruginosa", cited above [...].

Isolation and Characterization of a Novel Phage against Vibrio alginolyticus Belonging to a New Genus.

Vibrio alginolyticus causes substantial economic losses in the aquaculture industry. With the rise of multidrug-resistant Vibrio strains, phages present a promising solution. Here, a novel lytic Vibrio phage, vB_ValC_RH2G (RH2G), that efficiently infects the pathogenic strain V. alginolyticus ATCC 17749T, was isolated from mixed wastewater from an aquatic market in Xiamen, China. Transmission electron microscopy revealed that RH2G has the morphology of Siphoviruses, featuring an icosahedral head (73 ± 2 nm diameter) and long noncontractile tail (142 ± 4 nm). A one-step growth experiment showed that RH2G had a short latent period (10 min) and a burst size of 48 phage particles per infected cell. Additionally, RH2G was highly species-specific and was relatively stable at 4-55 °C and pH 4-10. A genomic analysis showed that RH2G has a 116,749 bp double-stranded DNA genome with 43.76% GC content. The intergenomic similarity between the genome sequence of RH2G and other phages recorded in the GenBank database was below 38.8%, suggesting that RH2G represents a new genus. RH2G did not exhibit any virulence or resistance genes. Its rapid lysis capacity, lytic activity, environmental resilience, and genetic safety suggested that RH2G may be a safe candidate for phage therapy in combatting vibriosis in aquaculture settings.

Characterization and comparative genomic analysis of a marine Bacillus phage reveal a novel viral genus.

Although recent metagenomics research has obtained a wealth of phage genetic information, much of it is considered "dark matter" because of the lack of similarity with known sequences in the database. Therefore, the isolation and characterization of novel phages will help to interpret the vast unknown viral metagenome data and improve our understanding of phage diversity and phage-host interactions. Bacillus pumilus shows high economic relevance due to its wide applications in biotechnology, industry, biopharma, and environmental sectors. Since phages influence the abundance, metabolism, evolution, fitness, and ecological functions of bacteria through complex interactions, the significance of isolation and characterization of novel phages infecting B. pumilus is apparent. In this study, we isolated and characterized a B. pumilus phage belonging to a novel viral genus, which provides essential knowledge for phage biology as well as the industrial application of B. pumilus.

Isolation and Characterization of Novel Lytic Bacteriophages Infecting Carbapenem-Resistant Pathogenic Diarrheagenic and Uropathogenic Escherichia Coli.

The evolution of antimicrobial resistance has dramatically reduced the efficacy of the first-choice and last-resort antibiotics used to treat E. coli infections. Thus, searching for novel therapeutics to treat and control the emergence of antibiotic resistance is urgent. Therefore, this study aimed to illustrate the lytic effect of phages against carbapenem-resistant pathogenic E. coli. Phages were isolated from hospital effluents by the enrichment assay. This was followed by the evaluation of the host range of the phages by the spot assay. The time taken by phages to bind to the host bacterial cells was determined by the adsorption assay. The phage latent period and burst size were determined using a one-step growth experiment. Phage morphology was determined by the Transmission Electron Microscopy. Molecular characterization of phages was done by whole genome sequencing. Two phages named UGKSEcP1 and UGKSEcP2 were isolated from hospital effluents. The phages were professionally lytic with a broad host range. The two phages recorded an average adsorption time of 11.25 minutes, an adsorption rate of 99.3%, a latency period of 20 minutes, and a burst size of approximately 528 phages/infected cell. Phages UGKSEcP1 and UGKSEcP2 had genome lengths of 167433bp, and 167221bp with 277 and 276 predicted genes, respectively, and no undesirable genes were detected. Phylogenetic analysis revealed the two phages belonged genus Tequatrovirus. TEM micrograph showed that the two phages had a similar morphotype with icosahedral heads and contractile tails; thus, classified as members of the Myoviridae phage family. The findings demonstrate that the study isolated two novel professionally lytic phages with a broad host range and thus, are candidates for phage-mediated biocontrol.

Genetically similar Xanthomonas arboricola pv. pruni strains and associated phage display phenotypic and genotypic variation across 35 years

Bacteriophages (phages), viruses that infect bacteria, have key ecological and evolutionary functions in the phytobiome. Despite the importance of phages as primary drivers for bacterial evolution, phage-bacteria interactions across spatiotemporal scales in natural, agricultural settings are underexplored. With increased interest in phage-based therapies to manage bacterial pathogens, an enhanced understanding of phage genetic and functional diversity at the population level, and how this, in turn, impacts bacterial evolution and virulence, is necessary. This study presents data on the genetic similarity among Xanthomonas arboricola pv. pruni (Xap) strains isolated from different geographic locations that display different lytic phenotypes when challenged with a panel of six phage isolates collected in the same region over four decades. The minor yet significant genetic variation among this small population of Xap strains is structured by both geographic location and response to phage infection. Phage genomes are also highly similar, with conserved and diverse genomic loci that correspond to isolation year. The six phages characterized here cluster into the Kantovirinae subfamily and possibly form a new genus. Only future studies will elucidate the role of Xap and Xapφ phage genes identified here in the virulence and lysis of Xap and how these, in turn, impact bacterial spot disease outcomes. The research and tripartite pathosystem presented here provides a unique opportunity to investigate the coevolution of phage-phytobacterial pathogen-plant host in depth in an agricultural setting with the potential to monitor the rate at which phage populations contribute to bacterial genetic diversity across geographic and temporal scales.

Isolation of lytic bacteriophages and their relationships with the adherence genes of Staphylococcus saprophyticus

ObjectiveThis study aimed to introduce a lytic bacteriophage against Staphylococcus saprophyticus from wastewater in Gorgan, northern Iran.ResultsThe vB_SsapS-46 phage was isolated from urban wastewater and formed round and clear plaques on bacterial culture. It was visualized by electron microscopy and had a large head (approximately 106 nm) and a long tail (approximately 150 nm), indicating that it belongs to the Siphoviridae family. The host range of vB_SsapS-46 was determined using a spot test on 35 S. saprophyticus clinical isolates, and it was able to lyse 12 of the 35 clinical isolates (34%). Finally, the relationship between phage sensitivity and adherence genes was assessed, revealing no significant correlation between phage sensitivity and the frequency of adherence genes. The vB_SsapS-46 phage can be used alone or in a mixture in future studies to control urinary tract infections caused by this bacterium, especially in the elimination of drug-resistant pathogens.

Bacteriophages isolated from mouse feces attenuates pneumonia mice caused by Pseudomonas aeruginosa.

Most of the current bacteriophages (phages) are mostly isolated from environments. However, phages isolated from feces might be more specific to the bacteria that are harmful to the host. Meanwhile, some phages from the environment might affect non-pathogenic bacteria for the host. Here, bacteriophages isolated from mouse feces were intratracheally (IT) or intravenously (IV) administered in pneumonia mice caused by Pseudomonas aeruginosa at 2 hours post-intratracheal bacterial administration. As such, the mice with phage treatment, using either IT or IV administration, demonstrated less severe pneumonia as indicated by mortality, serum cytokines, bacteremia, bacterial abundance in bronchoalveolar lavage fluid (BALF), and neutrophil extracellular traps (NETs) in lung tissue (immunofluorescence of neutrophil elastase and myeloperoxidase). Interestingly, the abundance of phages in BALF from the IT and IV injections was similar, supporting a flexible route of phage administration. With the incubation of bacteria with neutrophils, the presence of bacteriophages significantly improved bactericidal activity, but not NETs formation, with the elevated supernatant IL-6 and TNF-α, but not IL-1β. In conclusion, our findings suggest that bacteriophages against Pseudomonas aeruginosa can be discovered from feces of the host. The phages attenuate pneumonia partly through an enhanced neutrophil bactericidal activity, but not via inducing NETs formation. The isolation of phages from the infected hosts themselves might be practically useful for future treatment. More studies are warranted.

A novel genus of Pectobacterium bacteriophages display broad host range by targeting several species of Danish soft rot isolates

The bacterial diseases black leg and soft rot in potatoes cause heavy losses of potatoes worldwide. Bacteria within the genus Pectobacteriaceae are the causative agents of black leg and soft rot. The use of antibiotics in agriculture is heavily regulated and no other effective treatment currently exists, but bacteriophages (phages) have shown promise as potential biocontrol agents. In this study we isolated soft rot bacteria from potato tubers and plant tissue displaying soft rot or black leg symptoms collected in Danish fields. We then used the isolated bacterial strains as hosts for phage isolation. Using organic waste, we isolated phages targeting different species within Pectobacterium. Here we focus on seven of these phages representing a new genus primarily targeting P. brasiliense; phage Ymer, Amona, Sabo, Abuela, Koroua, Taid and Pappous. TEM image of phage Ymer showed siphovirus morphotype, and the proposed Ymer genus belongs to the class Caudoviricetes, with double-stranded DNA genomes varying from 39 kb to 43 kb. In silico host range prediction using a CRISPR-Cas spacer database suggested both P. brasiliense, P. polaris and P. versatile as natural hosts for phages within the proposed Ymer genus. A following host range experiment, using 47 bacterial isolates from Danish tubers and plants symptomatic with soft rot or black leg disease verified the in silico host range prediction, as the genus as a group were able to infect all three Pectobacterium species. Phages did, however, primarily target P. brasiliense isolates and displayed differences in host range even within the species level. Two of the phages were able to infect two or more Pectobacterium species. Despite no nucleotide similarity with any phages in the NCBI database, the proposed Ymer genus did share some similarity at the protein level, as well as gene synteny, with currently known phages. None of the phages encoded integrases or other genes typically associated with lysogeny. Similarly, no virulence factors nor antimicrobial resistance genes were found, and combined with their ability to infect several soft rot-causing Pectobacterium species from Danish fields, demonstrates their potential as biocontrol agents against soft rot and black leg diseases in potatoes.

Expanding the Phage Galaxy: Isolation and Characterization of Five Novel Streptomyces Siphoviruses Ankus, Byblos, DekoNeimoidia, Mandalore, and Naboo.

Key features of the actinobacterial genus Streptomyces are multicellular, filamentous growth, and production of a broad portfolio of bioactive molecules. These characteristics appear to play an important role in phage-host interactions and are modulated by phages during infection. To accelerate research of such interactions and the investigation of novel immune systems in multicellular bacteria, phage isolation, sequencing, and characterization are needed. This is a prerequisite for establishing systematic collections that appropriately cover phage diversity for comparative analyses. As part of a public outreach program within the priority program SPP 2330, involving local schools, we describe the isolation and characterization of five novel Streptomyces siphoviruses infecting S. griseus, S. venezuelae, and S. olivaceus. All isolates are virulent members of two existing genera and, additionally, establish a new genus in the Stanwilliamsviridae family. In addition to an extensive set of tRNAs and proteins involved in phage replication, about 80% of phage genes encode hypothetical proteins, underlining the yet underexplored phage diversity and genomic dark matter still found in bacteriophages infecting actinobacteria. Taken together, phages Ankus, Byblos, DekoNeimoidia, Mandalore, and Naboo expand the phage diversity and contribute to ongoing research in the field of Streptomyces phage-host interactions.

Isolation and characterization of bacteriophages against E.coli urinary tract infection and evaluating their anti-biofilm activity and antibiotic synergy

Urinary tract infections (UTIs) by Uropathogenic Escherichia coli (UPEC) are a significant health concern, especially due to the increasing prevalence of antibiotic resistance. This study focuses on isolating and characterizing bacteriophages specific to UPEC strains isolated from UTI samples. The isolated phages were assessed for their ability to target and lyse UPEC in vitro, focusing on their efficacy in disrupting biofilms, a key virulence factor contributing to UTI recurrence and antibiotic resistance. The morphological structure observed by TEM belongs to Myoviridae, the phage exhibited icosahedral symmetry with a long non-constricting tail, the approximate measurement of the phage head was 39 nm in diameter, and the phage tail was 105.317 nm in length. One-step growth experiments showed that the latent period was approximately 20 min, followed by a rise period of 40 min, and a growth plateau was reached within 20 min and the burst size observed was 26 phages/infected bacterial cells. These phages were capable of killing cells within the biofilms, leading to a reduction in living cell counts after a single treatment. This study highlights the potential of phages to play a significant role in disrupting, inactivating, and destroying Uropathogenic Escherichia coli (UPEC) biofilms. Such findings could be instrumental in developing treatment strategies that complement antibiotics and disinfectants. The phage-antibiotic synergistic activity was compared to have the possibility to facilitate the advancement of focused and enduring alternatives to traditional antibiotic therapies for UTIs.