Hidden diversity and potential ecological function of phosphorus acquisition genes in widespread terrestrial bacteriophages.

Phosphorus (P) limitation of ecosystem processes is widespread in terrestrial habitats. While a few auxiliary metabolic genes (AMGs) in bacteriophages from aquatic habitats are reported to have the potential to enhance P-acquisition ability of their hosts, little is known about the diversity and potential ecological function of P-acquisition genes encoded by terrestrial bacteriophages. Here, we analyze 333 soil metagenomes from five terrestrial habitat types across China and identify 75 viral operational taxonomic units (vOTUs) that encode 105 P-acquisition AMGs. These AMGs span 17 distinct functional genes involved in four primary processes of microbial P-acquisition. Among them, over 60% (11/17) have not been reported previously. We experimentally verify in-vitro enzymatic activities of two pyrophosphatases and one alkaline phosphatase encoded by P-acquisition vOTUs. Thirty-six percent of the 75 P-acquisition vOTUs are detectable in a published global topsoil metagenome dataset. Further analyses reveal that, under certain circumstances, the identified P-acquisition AMGs have a greater influence on soil P availability and are more dominant in soil metatranscriptomes than their corresponding bacterial genes. Overall, our results reinforce the necessity of incorporating viral contributions into biogeochemical P cycling.

Isolation, characterization and application of noble bacteriophages targeting potato common scab pathogen Streptomyces stelliscabiei.

Bacteriophages have emerged as promising alternatives to pesticides for controlling bacterial pathogens in crops. Among these pathogens, Streptomyces stelliscabiei (syn. S. stelliscabiei) is a primary causative agent of potato common scab (PCS), resulting in substantial global economic losses. The traditional management methods for PCS face numerous challenges, highlighting the need for effective and environmentally friendly control strategies. In this study, we successfully isolated three novel bacteriophages, namely Psst1, Psst2, and Psst4, which exhibited a broad host range encompassing seven S. stelliscabiei strains. Morphological analysis revealed their distinct features, including an icosahedral head and a non-contractile tail. These phages demonstrated stability across a broad range of temperatures (20-50°C), pH (pH 3-11), and UV exposure time (80 min). Genome sequencing revealed double-stranded DNA phage with open reading frames encoding genes for phage structure, DNA packaging and replication, host lysis and other essential functions. These phages lacked genes for antibiotic resistance, virulence, and toxicity. Average nucleotide identity, phylogenetic, and comparative genomic analyses classified the three phages as members of the Rimavirus genus, with Psst1 and Psst2 representing novel species. All three phages efficiently lysed S. stelliscabiei in the liquid medium and alleviated scab symptom development and reduced pathogen abundance on potato slices. Furthermore, phage treatments of radish seedlings alleviated the growth inhibition caused by S. stelliscabiei with no disease symptoms. In soil potted experiments, phages significantly reduced disease incidence by 40%. This decrease is attributed to a reduction in pathogen density and the selection of S. stelliscabiei strains with reduced virulence and slower growth rates in natural environments. Our study is the first to report the isolation of three novel phages that infect S. stelliscabiei as a host bacterium. These phages exhibit a broad host range, and demonstrate stability under a variety of environmental conditions. Additionally, they demonstrate biocontrol efficacy against bacterial infections in potato slices, radish seedlings, and potted experiments, underscoring their significant potential as biocontrol agents for the effective management of PCS.

Molecular engineering of a spheroid-penetrating phage nanovector for photodynamic treatment of colon cancer cells.

Photodynamic therapy (PDT) represents an emerging strategy to treat various malignancies, including colorectal cancer (CC), the third most common cancer type. This work presents an engineered M13 phage retargeted towards CC cells through pentavalent display of a disulfide-constrained peptide nonamer. The M13CC nanovector was conjugated with the photosensitizer Rose Bengal (RB), and the photodynamic anticancer effects of the resulting M13CC-RB bioconjugate were investigated on CC cells. We show that upon irradiation M13CC-RB is able to impair CC cell viability, and that this effect depends on i) photosensitizer concentration and ii) targeting efficiency towards CC cell lines, proving the specificity of the vector compared to unmodified M13 phage. We also demonstrate that M13CC-RB enhances generation and intracellular accumulation of reactive oxygen species (ROS) triggering CC cell death. To further investigate the anticancer potential of M13CC-RB, we performed PDT experiments on 3D CC spheroids, proving, for the first time, the ability of engineered M13 phage conjugates to deeply penetrate multicellular spheroids. Moreover, significant photodynamic effects, including spheroid disruption and cytotoxicity, were readily triggered at picomolar concentrations of the phage vector. Taken together, our results promote engineered M13 phages as promising nanovector platform for targeted photosensitization, paving the way to novel adjuvant approaches to fight CC malignancies.

Coordinated transcriptional response to environmental stress by a Synechococcus virus.

Viruses are a major control on populations of microbes. Often, their virulence is examined in controlled laboratory conditions. Yet, in nature, environmental conditions lead to changes in host physiology and fitness that may impart both costs and benefits on viral success. Phosphorus (P) is a major abiotic control on the marine cyanobacterium Synechococcus. Some viruses infecting Synechococcus have acquired, from their host, a gene encoding a P substrate binding protein (PstS), thought to improve virus replication under phosphate starvation. Yet, pstS is uncommon among cyanobacterial viruses. Thus, we asked how infections with viruses lacking PstS are affected by P scarcity. We show that the production of infectious virus particles of such viruses is reduced in low P conditions. However, this reduction in progeny is not caused by impaired phage genome replication, thought to be a major sink for cellular phosphate. Instead, transcriptomic analysis showed that under low P conditions, a PstS-lacking cyanophage increased the expression of a specific gene set that included mazG, hli2, and gp43 encoding a pyrophosphatase, a high-light inducible protein and DNA polymerase, respectively. Moreover, several of the upregulated genes were controlled by the host's phoBR two-component system. We hypothesize that recycling and polymerization of nucleotides liberates free phosphate and thus allows viral morphogenesis, albeit at lower rates than when phosphate is replete or when phages encode pstS. Altogether, our data show how phage genomes, lacking obvious P-stress-related genes, have evolved to exploit their host's environmental sensing mechanisms to coordinate their own gene expression in response to resource limitation.

Bacteriophage and antibiotic combination therapy for recurrent Enterococcus faecium bacteremia.

Enterococcus faecium is a member of the human gastrointestinal (GI) microbiota but can also cause invasive infections, especially in immunocompromised hosts. Enterococci display intrinsic resistance to many antibiotics, and most clinical E. faecium isolates have acquired vancomycin resistance, leaving clinicians with a limited repertoire of effective antibiotics. As such, vancomycin-resistant E. faecium (VREfm) has become an increasingly difficult to treat nosocomial pathogen that is often associated with treatment failure and recurrent infections. We followed a patient with recurrent E. faecium bloodstream infections (BSIs) of increasing severity, which ultimately became unresponsive to antibiotic combination therapy over the course of 7 years. Whole-genome sequencing (WGS) showed that the patient was colonized with closely related E. faecium strains for at least 2 years and that invasive isolates likely emerged from a large E. faecium population in the patient's gastrointestinal (GI) tract. The addition of bacteriophage (phage) therapy to the patient's antimicrobial regimen was associated with several months of clinical improvement and reduced intestinal burden of VRE and E. faecium. In vitro analysis showed that antibiotic and phage combination therapy improved bacterial growth suppression compared to therapy with either alone. Eventual E. faecium BSI recurrence was not associated with the development of antibiotic or phage resistance in post-treatment isolates. However, an anti-phage-neutralizing antibody response occurred that coincided with an increased relative abundance of VRE in the GI tract, both of which may have contributed to clinical failure. Taken together, these findings highlight the potential utility and limitations of phage therapy to treat antibiotic-resistant enterococcal infections. IMPORTANCE: Phage therapy is an emerging therapeutic approach for treating bacterial infections that do not respond to traditional antibiotics. The addition of phage therapy to systemic antibiotics to treat a patient with recurrent E. faecium infections that were non-responsive to antibiotics alone resulted in fewer hospitalizations and improved the patient's quality of life. Combination phage and antibiotic therapy reduced E. faecium and VRE abundance in the patient's stool. Eventually, an anti-phage antibody response emerged that was able to neutralize phage activity, which may have limited clinical efficacy. This study demonstrates the potential of phages as an additional option in the antimicrobial toolbox for treating invasive enterococcal infections and highlights the need for further investigation to ensure phage therapy can be deployed for maximum clinical benefit.

Bacteriophage therapy for drug-resistant Staphylococcus aureus infections.

Drug-resistant Staphylococcus aureus stands as a prominent pathogen in nosocomial and community-acquired infections, capable of inciting various infections at different sites in patients. This includes Staphylococcus aureus bacteremia (SaB), which exhibits a severe infection frequently associated with significant mortality rate of approximately 25%. In the absence of better alternative therapies, antibiotics is still the main approach for treating infections. However, excessive use of antibiotics has, in turn, led to an increase in antimicrobial resistance. Hence, it is imperative that new strategies are developed to control drug-resistant S. aureus infections. Bacteriophages are viruses with the ability to infect bacteria. Bacteriophages, were used to treat bacterial infections before the advent of antibiotics, but were subsequently replaced by antibiotics due to limited theoretical understanding and inefficient preparation processes at the time. Recently, phages have attracted the attention of many researchers again because of the serious problem of antibiotic resistance. This article provides a comprehensive overview of phage biology, animal models, diverse clinical case treatments, and clinical trials in the context of drug-resistant S. aureus phage therapy. It also assesses the strengths and limitations of phage therapy and outlines the future prospects and research directions. This review is expected to offer valuable insights for researchers engaged in phage-based treatments for drug-resistant S. aureus infections.

Bacteriophage T5 dUTPase: Combination of Common Enzymatic and Novel Functions.

The main function of dUTPases is to regulate the cellular levels of dUTP and dTTP, thereby playing a crucial role in DNA repair mechanisms. Despite the fact that mutant organisms with obliterated dUTPase enzymatic activity remain viable, it is not possible to completely knock out the dut gene due to the lethal consequences of such a mutation for the organism. As a result, it is considered that this class of enzymes performs an additional function that is essential for the organism's survival. In this study, we provide evidence that the dUTPase of bacteriophage T5 fulfills a supplemental function, in addition to its canonical role. We determined the crystal structure of bacteriophage T5 dUTPase with a resolution of 2.0 Å, and we discovered a distinct short loop consisting of six amino acid residues, representing a unique structural feature specific to the T5-like phages dUTPases. The removal of this element did not affect the overall structure of the homotrimer, but it had significant effects on the development of the phage. Furthermore, it was shown that the enzymatic function and the novel function of the bacteriophage T5 dUTPase are unrelated and independent from each other.

Optimizing in vitro phage-ciprofloxacin combination formulation for respiratory therapy of multi-drug resistant Pseudomonas aeruginosa infections.

Respiratory infection caused by multi-drug resistant (MDR) Pseudomonas aeruginosa is challenging to treat. In this study, we investigate the optimal dose of anti-pseudomonas phage PEV31 (103, 105, and 108 PFU/mL) combined with ciprofloxacin (ranging from 1/8× MIC to 8× MIC) to treat the MDR P. aeruginosa strain FADD1-PA001 using time-kill studies. We determined the impact of phage growth kinetics in the presence of ciprofloxacin through one-step growth analysis. Single treatments with either phage PEV31 or ciprofloxacin (except at 8× MIC) showed limited bactericidal efficiency, with bacterial regrowth observed at 48 h. The most effective treatments were PEV31 at multiplicity of infection (MOI) of 0.1 and 100 combined with ciprofloxacin at concentrations above 1× MIC, resulting in a >4 log10 reduction in bacterial counts. While the burst size of phage PEV31 was decreased with increasing ciprofloxacin concentration, robust antimicrobial effects were still maintained in the combination treatment. Aerosol samples collected from vibrating mesh nebulization of the combination formulation at phage MOI of 100 with 2× MIC effectively inhibited bacterial density. In summary, our combination treatments eradicated in vitro bacterial growth and sustained antimicrobial effects for 48 h. These results indicated the potential application of nebulization-based strategies for the combination treatment against MDR lung infections.

A universal dual-mode hydrogel array based on phage-DNA probe for simultaneous rapid screening and precisely quantitative detection of Escherichia coli O157:H7 in foods by the fluorescent/microfluidic chip electrophoresis methods.

Rapid and specific detection of virulent bacterial strains is a great challenge for food safety regarding large amounts of contaminated samples. Herein, a dual-mode hydrogel array biosensor was constructed to simultaneously rapidly screen and precisely quantitatively detect virulent Escherichia coli O157:H7 (E. coli O157:H7) based on a novel DNA-modified phage probe. First, E. coli O157:H7 was incubated with alginate to form the E. coli O157:H7/hydrogel premix complex. Subsequently, hydrogel formation by cross-linking upon the addition of calcium ions and phages for E. coli O157:H7 modified with a DNA primer (phage-DNA) was added to the alginate hydrogel. The DNA on the complex could trigger rolling circle amplification (RCA) to form a phage probe containing a long-chain DNA skeleton (phage@RCA-DNA). The RCA-DNA was then hybridized with the complementary DNA (cDNA) to form double-stranded DNA fragments (phage@RCA-dsDNA), which could be stained by the SYBR Green dye to emit visual green fluorescence (FL) and determined by a smartphone for rapid screening. Meanwhile, the unreacted cDNA in the supernatant could be quantitatively detected by microfluidic chip electrophoresis (MCE). The signal decrement was also proportional to the bacterial concentration. The detection limit values of E. coli O157:H7 were 50 CFU mL-1 by the FL signal and 6 CFU mL-1 by the MCE signal. The two results could be mutually corrected to decrease the false-positive results. This assay was also employed to detect virulent Salmonella Typhimurium (S. Typhimurium) using the corresponding S. Typhimurium phage@RCA-DNA probe. All these results demonstrated that the universal bioassay was suitable for simultaneous rapid screening and precisely quantitative detection of virulent bacterial strains.

Alternatives to conventional antibiotics for the prevention and treatment of commonly occurring diseases in feedlot cattle.

Antibiotic-resistant bacteria are a problem in human medicine. The development of antibiotic resistance in bacteria in feedlot cattle could have negative effects on their health and welfare and there is a theoretical possibility of transmission of antibiotic-resistant bacteria from food animals to humans. Alternatives to conventional antibiotics in feedlot health management could reduce the selective pressure for the development of antibiotic resistance. This review assesses the evidence supporting potential alternatives to conventional antibiotics in the prevention and treatment of diseases in feedlot cattle, including nitric oxide, plant extracts, supplemental yeast or yeast products, bacterial probiotics, organic acids, bacteriophages and non-specific immunostimulants. Further research is warranted with lactate utilising bacteria, the organic acid malate, bacteriophages and the non-specific immunostimulants ß-1,3 glucan and those based on pox viruses. However, none of the alternatives to conventional antibiotics investigated in this review have sufficient supporting evidence to date to justify their use with feedlot cattle. Frequently, statistically weak results and studies without negative controls are cited as support for similar studies. The health and welfare of feedlot cattle are dependent on the use of products that have robust supporting data to ensure efficacy and to avoid adverse outcomes.

Enhancement of bactericidal effects of bacteriophage and gentamicin combination regimen against Staphylococcus aureus and Pseudomonas aeruginosa strains in a mice diabetic wound model.

Diabetic patients are more susceptible to developing wound infections resulting in poor and delayed wound healing. Bacteriophages, the viruses that target-specific bacteria, can be used as an alternative to antibiotics to eliminate drug-resistant bacterial infections. Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) are among the most frequently identified pathogens in diabetic foot ulcers (DFUs). The aim of this study was assessment of bacteriophage and gentamicin combination effects on bacterial isolates from DFU infections. Specific bacteriophages were collected from sewage and animal feces samples and the phages were enriched using S. aureus and P. aeruginosa cultures. The lytic potential of phage isolates was assessed by the clarity of plaques. We isolated and characterized four lytic phages: Stp2, Psp1, Stp1, and Psp2. The phage cocktail was optimized and investigated in vitro. We also assessed the effects of topical bacteriophage cocktail gel on animal models of DFU. Results revealed that the phage cocktail significantly reduced the mortality rate in diabetic infected mice. We determined that treatment with bacteriophage cocktail effectively decreased bacterial colony counts and improved wound healing in S. aureus and P. aeruginosa infections, especially when administrated concomitantly with gentamicin. The application of complementary therapy using a phage cocktail and gentamicin, could offer an attractive approach for the treatment of wound diabetic bacterial infections.

Bacteriophage infection and killing of intracellular Mycobacterium abscessus.

IMPORTANCE: As we rapidly approach a post-antibiotic era, bacteriophage (phage) therapy may offer a solution for treating drug-resistant bacteria. Mycobacterium abscessus is an emerging, multidrug-resistant pathogen that causes disease in people with cystic fibrosis, chronic obstructive pulmonary disease, and other underlying lung diseases. M. abscessus can survive inside host cells, a niche that can limit access to antibiotics. As current treatment options for M. abscessus infections often fail, there is an urgent need for alternative therapies. Phage therapy is being used to treat M. abscessus infections as an option of last resort. However, little is known about the ability of phages to kill bacteria in the host environment and specifically in an intracellular environment. Here, we demonstrate the ability of phages to enter mammalian cells and to infect and kill intracellular M. abscessus. These findings support the use of phages to treat intracellular bacterial pathogens.

LysZX4-NCA, a new endolysin with broad-spectrum antibacterial activity for topical treatment.

The prevalence of multidrug-resistant highly virulent Klebsiella pneumoniae (MDR-hvKP) requires the development of new therapeutic agents. Herein, a novel lytic phage vB_KpnS_ZX4 against MDR-hvKP was discovered in hospital sewage. Phage vB_KpnS_ZX4 had a short latent period (5 min) and a large burst size (230 PFU/cell). It can rapidly reduce the number of bacteria in vitro and improve survival rates of bacteremic mice in vivo from 0 to 80 % with a single injection of 108 PFU. LysZX4, an endolysin derived from vB_KpnS_ZX4, exhibits potent antimicrobial activity in vitro in combination with ethylenediaminetetraacetic acid (EDTA). The antimicrobial activity of LysZX4 was further enhanced by the fusion of KWKLFKI residues from cecropin A (LysZX4-NCA). In vitro antibacterial experiments showed that LysZX4-NCA exerts broad-spectrum antibacterial activity against clinical Gram-negative bacteria, including MDR-hvKP. Moreover, in the mouse model of MDR-hvKP skin infection, treatment with LysZX4-NCA resulted in a three-log reduction in bacterial burden on the skin compared to the control group. Therefore, the novel phages vB_KpnS_ZX4 and LysZX4-NCA are effective reagents for the treatment of systemic and local MDR-hvKP infections.

Bacteriophage treatment as an alternative therapy for multidrug-resistant bacteria.

Multidrug-resistant (MDR) bacteria constitute one of the most serious global health threats. The increasing incidence rate of bacterial infections caused by MDR strains and the decrease in the number of newly developed antibiotics have prompted the scientific community to search for alternatives. One such alternative is the use of bacteriophages. In this review, we discuss the most critical MDR organisms, including Acinetobacter baumanni, Pseudomonas aeruginosa, and methicillin-resistant Staphylococcus aureus The efficacy of phage therapy against MDR bacteria is also discussed. We included studies from the last 10 years that examined the efficacy of phage therapy against MDR pathogens. In addition, this review highlights the effect of bacteriophages against bacterial biofilms. The existing knowledge indicates that phage therapy is a potential therapeutic strategy against MDR bacteria. However, the adverse effects of phage therapy, such as toxicity, and the emergence of phage resistance have not yet been resolved.

Functional mimicry of sea urchin biomineralization proteins with CaCO3-binding peptides selected by phage display.

The intricate process of biomineralization, e.g. in sea urchins, involves the precise interplay of highly regulated mineralization proteins and the spatiotemporal coordination achieved through compartmentalization. However, the investigation of biomineralization effector molecules, e.g. proteins, is challenging, due to their very low abundance. Therefore, we investigate the functional mimicry in the bioinspired precipitation of calcium carbonate (CaCO3) with artificial peptides selected from a peptide library by phage display based on peptide-binding to calcite and aragonite, respectively. The structure-directing effects of the identified peptides were compared to those of natural protein mixes isolated from skeletal (test) structures of two sea urchin species (Arbacia lixula and Paracentrotus lividus). The calcium carbonate samples deposited in the absence or presence of peptides were analyzed with a set of complementary techniques with regard to morphology, polymorph, and nanostructural motifs. Remarkably, some of the CaCO3-binding peptides induced morphological features in calcite that appeared similar to those obtained in the presence of the natural protein mixes. Many of the peptides identified as most effective in exerting a structure-directing effect on calcium carbonate crystallization were rich in basic amino acid residues. Hence, our in vitro mineralization study further highlights the important, but often neglected, role of positively charged soluble organic matrices associated with biological and bioinspired CaCO3 deposition.

Progress on Z genome biosynthetic pathway of bacteriophage.

There are abundant base modifications in bacteriophages' genomes, mainly for avoiding the digestion of host endonucleases. More than 40 years ago, researchers discovered that 2-amino-adenine (Z) completely replaced adenine (A) and forms a complementary pairing with three hydrogen bonds with thymine (T) in the DNA of cyanophage S-2L, forming a distinct "Z-genome". In recent years, researchers have discovered and validated the biosynthetic pathway of Z-genome in various bacteriophages, constituting a multi-enzyme system. This system includes the phage-encoded enzymes deoxy-2'-aminoadenylosuccinate synthetase (PurZ), deoxyadenosine triphosphate hydrolase (dATPase/DatZ), deoxyadenosine/deoxyguanosine triphosphate pyrophosphatase (DUF550/MazZ) and DNA polymerase (DpoZ). In this review, we provide a concise overview of the historical discovery on diversely modified nucleosides in bacteriophages, then we comprehensively summarize the research progress on multiple enzymes involved in the Z-genome biosynthetic pathway. Finally, the potential applications of the Z-genome and the enzymes in its biosynthetic pathway are discussed in order to provide reference for research in this field.

Effects of bacteriophage on Salmonella Enteritidis infection in broilers.

Bacteriophages (BP) are viruses that can infect bacteria. The present study evaluated the effect of BP on Salmonella infected broilers. A number of 150 day-old broilers were used in a completely randomized design with five treatments that included: (1) basal diet from day 0 to 28; (2) basal diet + 0.3 g/kg of colistin from day 0 to 28; (3) basal diet from day 1 to 13, and basal diet + 0.4 g/kg of colistin from day 14 to 28; (4) basal diet + 1 g/kg of BP from day 0 to 28; (5) basal diet + 1.5 g/kg of BP from day 0 to 28. On day 13, 15 chickens from each treatment were challenged by Salmonella Enteritidis (SE), while fifteen from each treatment were not; instead, they were kept in the same cage with the challenged chickens (exposed chickens). At 7 and 14 days post-challenge, the number of SE and coliform bacteria in the cecum and liver of colistin and BP-fed birds was lower than the control treatment. In exposed and challenged chickens, the height and surface area of villus were greater in the BP and colistin-supplemented groups. Serum concentrations of aspartate aminotransferase and alanine transaminase were greater, while serum albumin and triglycerides concentrations were lower in the control treatment. The liver of the challenged chickens had more pathological lesions than exposed birds. BP significantly decreased PPARγ gene expression in exposed chickens. In the challenged and exposed chickens, TLR4 gene expression was lower in BP and colistin-treated birds as compared to the control. In conclusion, adding BP to the diet from the day of age prevents the spread of Salmonella.

Bacteriophages: The promising therapeutic approach for enhancing ciprofloxacin efficacy against bacterial infection.

BACKGROUND: The emergence of ciprofloxacin-resistant bacteria is a serious challenge worldwide, bringing the need to find new approaches to manage this bacterium. Bacteriophages (phages) have been shown inhibitory effects against ciprofloxacin-resistance bacteria; thus, ciprofloxacin resistance or tolerance may not affect the phage's infection ability. Additionally, researchers used phage-ciprofloxacin combination therapy for the inhibition of multidrug-resistant bacteria. RESULTS: The sublethal concentrations of ciprofloxacin could lead to an increase in progeny production. Antibiotic treatments could enhance the release of progeny phages by shortening the lytic cycle and latent period. Thus, sublethal concentrations of antibiotics combined with phages can be used for the management of bacterial infections with high antibiotic resistance. In addition, combination therapy exerts various selection pressures that can mutually decrease phage and antibiotic resistance. Moreover, phage ciprofloxacin could significantly reduce bacterial counts in the biofilm community. Immediate usage of phages after the attachment of bacteria to the surface of the flow cells, before the development of micro-colonies, could lead to the best effect of phage therapy against bacterial biofilm. Noteworthy, phage should be used before antibiotics usage because this condition may have allowed phage replication to occur first before ciprofloxacin interrupted the bacterial DNA replication process, thereby interfering with the activity of the phages. Furthermore, the phage-ciprofloxacin combination showed a promising result for the management of Pseudomonas aeruginosa infections in mouse models. Nevertheless, low data are existing about the interaction between phages and ciprofloxacin in combination therapies, especially regarding the emergence of phage-resistant mutants. Additionally, there is a challenging and important question of how the combined ciprofloxacin with phages can increase antibacterial functions. Therefore, more examinations are required to support the clinical usage of phage-ciprofloxacin combination therapy.

DePolymerase Predictor (DePP): a machine learning tool for the targeted identification of phage depolymerases.

Biofilm production plays a clinically significant role in the pathogenicity of many bacteria, limiting our ability to apply antimicrobial agents and contributing in particular to the pathogenesis of chronic infections. Bacteriophage depolymerases, leveraged by these viruses to circumvent biofilm mediated resistance, represent a potentially powerful weapon in the fight against antibiotic resistant bacteria. Such enzymes are able to degrade the extracellular matrix that is integral to the formation of all biofilms and as such would allow complementary therapies or disinfection procedures to be successfully applied. In this manuscript, we describe the development and application of a machine learning based approach towards the identification of phage depolymerases. We demonstrate that on the basis of a relatively limited number of experimentally proven enzymes and using an amino acid derived feature vector that the development of a powerful model with an accuracy on the order of 90% is possible, showing the value of such approaches in protein functional annotation and the discovery of novel therapeutic agents.

Isolation and Characterization of a Novel Phage Collection against Avian-Pathogenic Escherichia coli.

The increase in antibiotic-resistant avian-pathogenic Escherichia coli (APEC), the causative agent of colibacillosis in poultry, warrants urgent research and the development of alternative therapies. This study describes the isolation and characterization of 19 genetically diverse, lytic coliphages, 8 of which were tested in combination for their efficacy in controlling in ovo APEC infections. Genome homology analysis revealed that the phages belong to nine different genera, one of them being a novel genus (Nouzillyvirus). One phage, REC, was derived from a recombination event between two Phapecoctavirus phages (ESCO5 and ESCO37) isolated in this study. Twenty-six of the 30 APEC strains tested were lysed by at least one phage. Phages exhibited varying infectious capacities, with narrow to broad host ranges. The broad host range of some phages could be partially explained by the presence of receptor-binding protein carrying a polysaccharidase domain. To demonstrate their therapeutic potential, a phage cocktail consisting of eight phages belonging to eight different genera was tested against BEN4358, an APEC O2 strain. In vitro, this phage cocktail fully inhibited the growth of BEN4358. In a chicken lethality embryo assay, the phage cocktail enabled 90% of phage-treated embryos to survive infection with BEN4358, compared with 0% of nontreated embryos, indicating that these novel phages are good candidates to successfully treat colibacillosis in poultry. IMPORTANCE Colibacillosis, the most common bacterial disease affecting poultry, is mainly treated by antibiotics. Due to the increased prevalence of multidrug-resistant avian-pathogenic Escherichia coli, there is an urgent need to assess the efficacy of alternatives to antibiotherapy, such as phage therapy. Here, we have isolated and characterized 19 coliphages that belong to nine phage genera. We showed that a combination of 8 of these phages was efficacious in vitro to control the growth of a clinical isolate of E. coli. Used in ovo, this phage combination allowed embryos to survive APEC infection. Thus, this phage combination represents a promising treatment for avian colibacillosis.