Bacteriophage therapy is considered one of the most promising therapeutic approaches against multi-drug resistant bacterial infections. Infections caused by Staphylococcus aureus are very efficiently controlled with therapeutic bacteriophage cocktails, containing a number of individual phages infecting a majority of known pathogenic S. aureus strains. We assessed the contribution of individual bacteriophages comprising a therapeutic bacteriophage cocktail against S. aureus in order to optimize its composition. Two lytic bacteriophages vB_SauM-515A1 (Myoviridae) and vB_SauP-436A (Podoviridae) were isolated from the commercial therapeutic cocktail produced by Microgen (Russia). Host ranges of the phages were established on the panel of 75 S. aureus strains. Phage vB_SauM-515A1 lysed 85.3% and vB_SauP-436A lysed 68.0% of the strains, however, vB_SauP-436A was active against four strains resistant to vB_SauM-515A1, as well as to the therapeutic cocktail per se. Suboptimal results of the therapeutic cocktail application were due to extremely low vB_SauP-436A1 content in this composition. Optimization of the phage titers led to an increase in overall cocktail efficiency. Thus, one of the effective ways to optimize the phage cocktails design was demonstrated and realized by using bacteriophages of different families and lytic spectra.

Antibiotic resistance is a major public health concern in many countries worldwide. The rapid spread of multidrug-resistant (MDR) bacteria is the main driving force for the development of novel non-antibiotic antimicrobials as a therapeutic alternative. Here, we isolated and characterized three virulent bacteriophages that specifically infect and lyse MDR Klebsiella pneumoniae with K23 capsule type. The phages belonged to the Autographiviridae (vB_KpnP_Dlv622) and Myoviridae (vB_KpnM_Seu621, KpS8) families and contained highly similar receptor-binding proteins (RBPs) with polysaccharide depolymerase enzymatic activity. Based on phylogenetic analysis, a similar pattern was also noted for five other groups of depolymerases, specific against capsule types K1, K30/K69, K57, K63, and KN2. The resulting recombinant depolymerases Dep622 (phage vB_KpnP_Dlv622) and DepS8 (phage KpS8) demonstrated narrow specificity against K. pneumoniae with capsule type K23 and were able to protect Galleria mellonella larvae in a model infection with a K. pneumoniae multidrug-resistant strain. These findings expand our knowledge of the diversity of phage depolymerases and provide further evidence that bacteriophages and phage polysaccharide depolymerases represent a promising tool for antimicrobial therapy.

Escherichia coli (E. coli) has been increasingly implicated in the pathogenesis of Crohn’s disease (CD). The phylogeny of E. coli isolated from Crohn’s disease patients (CDEC) was controversial, and while genotyping results suggested heterogeneity, the sequenced strains of E. coli from CD patients were closely related. We performed the shotgun genome sequencing of 28 E. coli isolates from ten CD patients and compared genomes from these isolates with already published genomes of CD strains and other pathogenic and non-pathogenic strains. CDEC was shown to belong to A, B1, B2 and D phylogenetic groups. The plasmid and several operons from the reference CD-associated E. coli strain LF82 were demonstrated to be more often present in CDEC genomes belonging to different phylogenetic groups than in genomes of commensal strains. The operons include carbon-source induced invasion GimA island, prophage I, iron uptake operons I and II, capsular assembly pathogenetic island IV and propanediol and galactitol utilization operons. Our findings suggest that CDEC are phylogenetically diverse. However, some strains isolated from independent sources possess highly similar chromosome or plasmids. Though no CD-specific genes or functional domains were present in all CD-associated strains, some genes and operons are more often found in the genomes of CDEC than in commensal E. coli. They are principally linked to gut colonization and utilization of propanediol and other sugar alcohols.

Background: The combined use of bacteriophages and antibiotics represents a promising strategy for combating multidrug-resistant bacterial pathogens. However, the lack of uniformity in methods for assessing combination effects and experimental protocols has resulted in inconsistent findings across studies. This study aimed to evaluate the effects of interactions between phages and antibiotics on Klebsiella pneumoniae strains using various statistical approaches to formalize combination effects.Methods: Effects were assessed for four antibiotics from distinct classes (gentamicin, levofloxacin, meropenem, chloramphenicol), three phages from different genera (Dlv622, Seu621, FRZ284), and a depolymerase (Dep622) on three K. pneumoniae strains of the KL23 capsule type. Antibiotics were used at C_max concentrations, and phages at sublethal levels. A modified t-test, Bliss independence model, two-way ANOVA, and checkerboard assay were employed to evaluate the results.Results: Among 48 combinations, 33 effects were statistically significant, including 26 cases of synergy and 7 of antagonism. All statistical methods showed consistency in identifying effects; however, the t-test and Bliss method detected a greater number of effects. The strongest synergy was observed with levofloxacin in combination with Seu621 or Dep622 across all bacterial strains. Checkerboard assays confirmed synergy in selected cases but indicated that combined effects could vary with antimicrobial concentrations.Conclusion: The choice of analytical method substantially impacts the detection of phage-antibiotic effects. The t-test and Bliss method, due to their simplicity and sensitivity, may be optimal for clinical application, while two-way ANOVA for confirming strong interactions. These results emphasize the need to consider interaction characteristics when designing therapeutic strategies.

The issue of antibiotic resistance in healthcare worldwide has led to a pressing need to explore and develop alternative approaches to combat infectious diseases. Among these methods, phage therapy has emerged as a potential solution to tackle this growing challenge. Virulent phages of the Herelleviridae family, known for their ability to cause lysis of Staphylococcus aureus, a clinically significant pathogen frequently associated with multidrug resistance, have proven to be one of the most effective viruses utilized in phage therapy. In order to utilize phages for therapeutic purposes effectively, a thorough investigation into their physiology and mechanisms of action on infected cells is essential. The use of omics technologies, particularly total RNA sequencing, is a promising approach for analyzing the interaction between phages and their hosts, allowing for the assessment of both the behavior of the phage during infection and the cell’s response. This review aims to provide a comprehensive overview of the physiology of the Herelleviridae family, utilizing existing analyses of their total phage transcriptomes. Additionally, it sheds light on the changes that occur in the metabolism of S. aureus when infected with virulent bacteriophages, contributing to a deeper understanding of the phage–host interaction.

In order to address the upcoming crisis in the treatment of Klebsiella pneumoniae infections, caused by an increasing proportion of resistant isolates, new approaches to antimicrobial therapy must be developed. One approach would be to use (bacterio)phages and/or phage derivatives for therapy. In this study, we present a description of the first K. pneumoniae phage from the Zobellviridae family. The vB_KpnP_Klyazma podovirus, which forms translucent halos around the plaques, was isolated from river water. The phage genome is composed of 82 open reading frames, which are divided into two clusters located on opposite strands. Phylogenetic analysis revealed that the phage belongs to the Zobellviridae family, although its identity with the closest member of this family was not higher than 5%. The bacteriophage demonstrated lytic activity against all (n = 11) K. pneumoniae strains with the KL20 capsule type, but only the host strain was lysed effectively. The receptor-binding protein of the phage was identified as a polysaccharide depolymerase with a pectate lyase domain. The recombinant depolymerase protein showed concentration-dependent activity against all strains with the KL20 capsule type. The ability of a recombinant depolymerase to cleave bacterial capsular polysaccharides regardless of a phage’s ability to successfully infect a particular strain holds promise for the possibility of using depolymerases in antimicrobial therapy, even though they only make bacteria sensitive to environmental factors, rather than killing them directly.