Urinary tract infections (UTIs) are among the most common bacterial infections worldwide and are associated with high recurrence rates and substantial morbidity. New therapeutic approaches are urgently needed, especially for recurrent UTIs (rUTIs), among which phage therapy represents a promising option. Here, we systematically evaluated the performance of six E. coli phages under urinary tract-relevant conditions and established a quantitative area-under-the-curve (AUC)-based framework for the rational selection of therapeutic phages. Phage lysis behavior was tested under oxic and anoxic conditions, revealing modulation by oxygen availability. Phages were also screened for biofilm-degrading activity, and in silico depolymerase predictions were compared with in vitro results, showing strong correlation. Phage-antibiotic interactions and refined treatment strategies were further investigated. Phages G9062, G10400, and MM02 emerged as the most promising candidates, remaining stable and active in urine under physiological pH fluctuations and exhibiting synergistic effects with gentamicin and fosfomycin, but not with the bacteriostatic nitrofurantoin. Importantly, we demonstrate that treatment sequence is a critical determinant of efficacy, with phage administration prior to antibiotic exposure consistently producing the strongest synergistic effects. These findings highlight the potential of phages as an integral component of a holistic therapeutic strategy for rUTIs.Importance Recurrent urinary tract infections (rUTIs) are a major clinical challenge characterized by repeated episodes, increasing antimicrobial resistance, and a substantial socioeconomic burden. Although bacteriophages represent a promising alternative or adjunct to antibiotics, their clinical translation has been hindered by the lack of standardized criteria for phage selection, dosing, and combination with antibiotics. This study addresses these gaps by defining key determinants of phage efficacy under conditions relevant to the urinary tract. Our findings highlight that phage performance is strongly shaped by the treatment environment and by interactions with antibiotics, and that inappropriate combinations or treatment sequences may compromise efficacy. By providing quantitative and transferable frameworks for evaluating phage effectivity, this work advances the rational development of phage-based therapies and helps bridge the gap between experimental phage research and clinically actionable treatment strategies for rUTI.

The release of petroleum and its constituents, such as polycyclic aromatic hydrocarbons (PAHs), into the environment poses a significant global threat due to their toxic, genotoxic, mutagenic, and/or carcinogenic properties. Using indigenous PAH-degrading bacteria to decontaminate the PAHs is an eco-friendly, convenient, and non-expensive way in comparison to other traditional physical and chemical methods. In the present study, several bacteria were isolated from petroleum-contaminated soils using a culture-enrichment technique. Phenanthrene and naphthalene were used as representative examples of the PAHs and as sole carbon and energy sources for the isolated bacteria. Three isolates (A1, A3, and B8-1) were selected as the most potent phenanthrene and naphthalene degraders. These three most potent isolates were used to build up four different consortiums, and both the individual isolates and developed consortiums were induced to set up biofilms. The results showed that the highest growth and degradation rate of phenanthrene and naphthalene were achieved at 37 °C and pH 7. Individual strains had higher phenanthrene degrading capacities compared to microbial consortiums in both planktonic and biofilm forms. Additionally, the developed-biofilm displays an increase in the degradation rate of individual isolates A1, A3, and consortiums. Finally, the selected biofilm degraders were used to investigate the PAHs degradation ability in an artificially contaminated soil. The finding of this experiment demonstrate that the biofilm mode of growth could be more effective in PAHs degradation compared to planktonic cultures, thus it can be applied effectively to remediate PAHs-contaminated soils. The most potent PAHs degraders were identified using 16 S rRNA, and the phylogenetic sequence analysis of the three isolates was affiliated as B8-1(Bacillus paramycoides), A1(Bacillus paranthracis), and A3(Bacillus licheniformis).

Pseudomonas aeruginosa is a major cause of ventilator-associated pneumonia (VAP) and is increasingly associated with extensive antimicrobial resistance, limiting therapeutic options and contributing to poor clinical outcomes. The emergence of high-risk clones that combine multidrug resistance and virulence-associated traits represents a growing public health concern.In this study, 100 clinical P. aeruginosa isolates recovered from VAP patients were screened using culture-based identification and antimicrobial susceptibility testing. Two isolates exhibiting the most extensive resistance profiles, consistent with pandrug-resistant (PDR) phenotypes, and associated with fatal clinical outcomes were selected for whole-genome sequencing using Oxford Nanopore technology.Comprehensive genomic analyses were performed to characterize antimicrobial resistance determinants, chromosomal resistance mutations, virulence-associated genes, mobile genetic elements, genomic islands, and phylogenomic relationships. Both isolates were assigned to sequence type ST308 and serogroup O11, a globally disseminated high-risk lineage. Core-genome phylogenomic analysis of 52 genomes identified 144,617 informative SNPs across 5,085 shared core genes and showed that the two Egyptian isolates differed by only 74 core-genome SNPs while clustering within the international ST308 lineage.The isolates harbored an extensive resistome including blaNDM-1, blaOXA-10, blaOXA-488, qnrVC1, rmtF, sul1, sul2, floR, and msrE. High-confidence chromosomal resistance mutations were detected in gyrA (T83I), parC (S87L), oprD (Q402HfsTer99), nalC, nalD, and pmrB, supporting resistance to fluoroquinolones, carbapenems, and polymyxins. Two complete class 1 integrons and one In0 structure, all carrying the class 1 integrase IntI1, were identified, comprising integron-associated multidrug resistance modules containing qnrVC1, aadA11, aac(6')-II, dfrB5, sul1, qacEΔ1, rmtF, blaOXA-10, and blaPAC-1. In contrast, blaNDM-1 was located within a distinct ISVsa3-associated resistance island together with ble and msrE. Genomic island analysis further demonstrated the co-localization of key resistance determinants and the exoU virulence factor within predicted genomic islands.Both isolates carried a highly conserved virulome, including exoU, exoT, type VI secretion system components, iron-acquisition systems, and biofilm-associated genes. Collectively, these findings indicate the convergence of resistance islands, integron-associated multidrug resistance modules, chromosomal resistance mechanisms, and virulence-associated determinants within a globally disseminated ST308/O11 high-risk clone associated with fatal VAP cases in Egypt.To our knowledge, this study provides the first comprehensive genomic characterization of ST308/O11 P. aeruginosa exhibiting a pandrug-resistant phenotype based on the antimicrobial panel tested and associated with fatal ventilator-associated pneumonia in Egypt. Furthermore, by integrating cohort-level screening of 100 clinical isolates with comparative whole-genome analysis, this work provides additional insight into the genomic diversity of globally disseminated high-risk lineages through the accumulation of resistance and virulence determinants.

The clinical implementation of mycobacteriophages is hampered by the lack of standardized in vitro methods and inconsistent host range results, limiting reliable assessment of their therapeutic potential. Nontuberculous mycobacterial (NTM) infections are frequently difficult to treat, emphasizing the urgent need for alternative or adjunctive strategies to antibiotics. To facilitate reproducible translational research, a detailed and systematic understanding of in vitro assay conditions is essential. Here, we comprehensively evaluated commonly used liquid and solid media for their suitability in supporting growth of multiple NTM strains and subsequent phage application. We compared bacterial growth kinetics, aggregation tendencies, and the efficiency of phage-induced lysis, including plaque formation, across three liquid media and corresponding solid agars. Our results demonstrate that Middlebrook-based media and agar consistently support robust NTM growth, reduce bacterial clumping, and enable reproducible detection of phage activity, outperforming TSB-based alternatives despite higher cost and labor requirements. These findings provide a standardized framework for in vitro NTM cultivation and phage testing, which is critical for reliable evaluation of host range, infection dynamics, and the development of mycobacteriophage-based therapies.

Astroviruses are becoming a growing concern in public and veterinary health. In humans, astrovirus infections can cause severe diarrhea and may lead to neuropathological encephalitis, whereas in wildlife, these enteropathogenic viral infections often lack overt symptoms and thus remain unnoticed. Yet their close interaction with the host's gastrointestinal microbiome might drive cascading effects with disadvantages for host health. Bats harbor many zoonotic viruses without showing signs of disease, and many species move freely along the gradient from pristine to agricultural landscapes. To better understand the impact of astrovirus (AstV) infection under a One Health framework, we investigated the gut microbiome of naturally AstV-infected Seba's short-tailed bats (Carollia perspicillata, n = 234) inhabiting old-growth lowland forests or forest fragments embedded in an agricultural matrix in Panama. AstV prevalence was higher in forest fragments. We observed that AstV infection is associated with a shift in microbial beta but not alpha diversity, which points towards the replacement of common gut microbial taxa when infected. Indeed, potentially beneficial bacteria, such as Lactococcus, decreased in abundance, whereas potentially pathogenic bacteria from the Helicobacter genus increased in AstV-positive bats. Two Helicobacter haplotypes closely related to avian Helicobacter species were identified. We conclude that even though the impact of infection on the microbiome was not amplified in forest fragments, the higher infection likelihood in landscapes altered by humans implies more frequent or prolonged health repercussions for bats.

Avian pathogenic Escherichia coli (APEC) poses a substantial economic and health challenges for poultry and meat industries. One of the promising alternative approaches to overcome bacterial contamination in food products is bacteriophage, due to its high specificity, safety, and effectiveness. We aimed to isolate a bacteriophage against E. coli causing poultry diseases and investigate its potential as biocontrol/therapeutic agent. A novel E. coli phage vB_EcoP_ZCEC16 was isolated from poultry samples. vB_EcoP_ZCEC16 is a podophage as visualized by transmission electron microscope, and it exhibited a high thermal and pH stability, and high lytic activity at MOIs 0.1 and 1 demonstrated by rapid reduction in bacterial optical density. Whole-genome analysis clustered the phage within Kuravirus genus and confirmed its virulence and the absence of antimicrobial resistance genes. The phage was also safe at high concentration (10[9] PFU/mL) on human fibroblast cells (HSF) and human colon carcinoma (Caco-2) cell lines as no cytotoxic effect was observed after 24 h. In addition, the lowest tested phage MOI (0.1) was the most protective to HSF cells against bacterial infection. Finally, phage vB_EcoP_ZCEC16 was evaluated for its biocontrol efficacy, where it significantly reduced the count of E. coli contaminating fresh poultry meat after 2 h of treatment. Our study introduced a novel virulent phage with the potential of biocontrol agent for enhancing food safety.

The global rise of antibiotic resistance has rekindled interest in bacteriophages (phages) as alternative therapeutics. Here, we isolated and characterized a novel Escherichia coli phage, Ec_Ou_ZC1, from the family Drexlerviridae, genus Tunavirus, sharing 94.72% genomic identity with Escherichia phage phiD2-2. Ec_Ou_ZC1 encodes an orphan methyltransferase and a Cor superinfection exclusion (SIE) protein. Heterologous expression of Cor blocked all tested FhuA-dependent Drexlerviridae phages (Ec_Ou_ZC1, T1, Bas03, Bas12, Bas13) but not T4 or T7, confirming family-specific SIE through FhuA receptor inactivation. Plaque assays against four antiviral STAND (Avs) systems revealed differential susceptibility within Drexlerviridae: Bas03 (Braunvirinae) was sensitive to SeAvs3, while Ec_Ou_ZC1, Bas05, Bas12, Bas13, and T1 (Tempevirinae and Tunavirinae) resisted all four systems, suggesting that Avs resistance depends on specific genomic features. Species-level phylogenetic analyses using concatenated single-copy orthologs robustly resolved evolutionary relationships within Tunavirus, separating Escherichia- and Shigella-infecting phages and providing a framework for predicting host range of new isolates.