Vectors optimized for delivery of microbial populations

Abstract

The present invention relates to vectors, preferably contained in a delivery vehicle, that contain no more than (100), preferably no more than (10), restriction sites recognized by restriction enzymes encoded by each bacterium of a bacterial group of interest. The present invention also relates to the use of the above vector, preferably contained in a delivery vehicle, as a medicament, in the treatment of a disease, especially in a patient in need thereof.

Description

[Technical field]

[0001] The present invention relates to the field of medicine, in particular to the field of bacterial infections. The present invention provides a novel phage therapy strategy. [Background technology]

[0002] Today, the treatment of bacterial infections relies primarily on the use of antibiotics. However, excessive and inappropriate use of antibiotics has encouraged the emergence and spread of antibiotic-resistant microorganisms. In fact, infections caused by antibiotic-resistant microorganisms, also known as "superbugs," often no longer respond to conventional treatments, thereby prolonging the duration of illness associated with the infection and even causing death to the patient. Due to the emergence of this antibiotic resistance phenomenon and the lack of discovery of new antibiotic classes, humanity is currently facing the possibility of a future without effective treatments for bacterial infections.

[0003] Moreover, with recent developments in the field of microbiome, an increasing number of studies have highlighted the detrimental effects of antibiotic treatment: in fact, due to their low specificity, antibiotics reduce bacterial diversity, which is essential for the development and preservation of a healthy microbiome.

[0004] Bacteriophages are viruses that are able to infect and reproduce in certain bacterial species, most often causing their death. Since their discovery in 1917, they have been used as a treatment against pathogenic bacteria in Eastern Europe. There, they continue to be an alternative to antibiotics or complementary therapies. This method is called "phage therapy".

[0005] Unlike classical chemical-based antibiotics, which are active against a broad spectrum of bacterial species, each bacteriophage can only infect and kill a small number of different closely related bacteria. This narrow spectrum of bactericidal activity is one of the main bottlenecks in the use of phage therapy. To date, two methods have been used to circumvent the specificity problem.

[0006] The first method consists of using a mixture of different naturally occurring phages, also called phage cocktails, where each phage targets a different subset of bacterial strains. The main drawback of such cocktails is that they are difficult to meet regulatory approval criteria.

[0007] The second method is to broaden the host range of a particular phage by mutation / selection cycles or engineering.

[0008] In the mutation / selection cycle strategy, the phage undergoes cycles of mutation and selection in the presence of targeted bacteria. However, this method has several drawbacks. Indeed, these mutation / selection cycles must be repeated for each targeted strain that is immune to the phage, and the selection of mutations that allow replication in one targeted strain may reduce or abolish the possibility of entering other strains. Furthermore, the mutation spectrum (space of available mutations) of the phage is limited.

[0009] In contrast, phage engineering allows for a more rational approach in which specific features of phages known to be important for infection are added or modified. Most of the efforts in phage engineering have so far focused on the ability of phages to recognize targeted strains via tail fiber proteins. Swapping tail fibers between two phages could, in principle, allow for the exchange of their recognition properties. As an example, by exchanging the gp17 tail fiber genes of phages T3 and T7, T7 was able to expand its host range and infect E. coli strains that were previously immune to T7 but not to T3 (Pires DP et al., 2016, Microbiology and Molecular Biology Reviews, 80(3): 523-543).

[0010] However, tail fiber protein modifications have shown limited success in expanding bacteriophage host range.

[0011] Moreover, phage engineering can be a tedious task for several reasons, including the phage genome size and the difficulty of engineering modifications in the host (especially when dealing with lytic phages). Thus, researchers have recently used the phage machinery as a nanocarrier to inject DNA of interest in the form of plasmids into bacteria. For example, these plasmid-phage hybrids, also called phagemids, have been used to inject genetic circuits encoding CRISPR Cas9 nucleases that allow sequence-specific killing of bacteria (Bikard D et al., 2014, Nature Biotechnology, 32(11): 1146-1150; Citorik RJ et al., 2014, Nature Biotechnology, 32(11): 1141-1145; WO 2015 / 034872). This strategy has been efficient in specifically killing subpopulations of bacteria that contain resistance to a given antibiotic.

[0012] However, there is currently no real broad-spectrum alternative treatment to conventional antibiotics on the market.Therefore, there is a persistent and urgent medical need to develop novel broad-spectrum phage or phagemid therapies that can effectively overcome global pathogen resistance, especially broad-spectrum therapies that target bacterial pathogen strains without compromising the diversity of the microbiome.The present invention aims to meet these and other needs. [Prior art documents] [Patent documents]

[0013] [Patent Document 1] International Publication No. 2015 / 034872 [Patent Document 2] International Publication No. 2014 / 124226 [Patent Document 3] U.S. Patent Application Publication No. 2015 / 0064138

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[0015] The inventors surprisingly discovered that the main obstacle to DNA entry from a phage / phagemid into a target bacterial strain is not related to the specificity of the tail fiber, but rather to the number of restriction sites present in its genome. Thus, the same bacteriophage scaffold with the same tail fiber protein can deliver its DNA cargo with varying efficiencies (ranging from 0 to 100%) according to the number of restriction sites present in the cargo. These results address the problem of phage / phagemid host range in an entirely different way, providing an unexplored avenue for the design of novel engineered, highly efficient phage or phagemid-based particles. Indeed, here, instead of engineering different attachment and entry points for viral particles as has been done previously, the solution is to rationally design a phagemid or bacteriophage genome with a reduced number of restriction sites.

[0016] Thus, the present invention relates to a bacteriophage or packaged phagemid, which does not contain a restriction site for any of the restriction enzymes frequently encoded in the bacterial population of interest.

[0017] In particular, a restriction enzyme is frequently encoded in a bacterial population of interest if at least 10% of the bacteria in the bacterial population encode the restriction enzyme.

[0018] Preferably, the group of bacteria of interest consists of a group of n bacterial strains, where n comprises a positive integer of 2 to about 100,000, preferably 10 to about 10,000. Preferably, the bacterial strains are selected from a single species.

[0019] Optionally, the bacterium of interest is selected from the group consisting of Yersinia spp., Escherichia spp., Klebsiella spp., Acinetobacter spp., Bordetella spp., Neisseria spp., Aeromonas spp., Franciella spp., Corynebacterium spp., Citrobacter spp., Chlamydia spp., Haemophilus spp., Brucella spp., spp.), Mycobacterium spp., Legionella spp., Rhodococcus spp., Pseudomonas spp., Helicobacter spp., Vibrio spp., Bacillus spp., Erysipelothrix spp., Salmonella spp., Streptomyces spp., Streptococcus spp., Staphylococcus spp., Bacteroides spp. spp., Prevotella spp., Clostridium spp., Bifidobacterium spp., Clostridium spp., Brevibacterium spp., Lactococcus spp., Leuconostoc spp., Actinobacillus spp., Selnomonas spp.), Shigella spp., Zymonas spp., Mycoplasma spp., Treponema spp., Leuconostoc spp., Corynebacterium spp., Enterococcus spp., Enterobacter spp., Pyrococcus spp., Serratia spp., Morganella spp., Parvimonas spp., Fusobacterium spp., Actinomyces spp. spp., Porphyromonas spp., Micrococcus spp., Bartonella spp., Borrelia spp., Brucelia spp., Campylobacter spp., Chlamydophilia spp., Cutibacterium spp. (formerly Propionibacterium spp.), Ehrlichia spp., Haemophilus spp., Leptospira spp., Listeria spp. spp., Mycoplasma spp., Nocardia spp., Rickettsia spp., Ureaplasma spp., and Lactobacillus spp., and mixtures thereof, and preferably, the bacterium of interest is selected from the group consisting of Escherichia spp..

[0020] Optionally, the bacterium of interest is selected from the group consisting of Bacteroides thetaiotaomicron, Bacteroides fragilis, Bacteroides distasonis, Bacteroides vulgatus, Clostridium leptum, Clostridium coccoides, Staphylococcus aureus, Bacillus subtilis, Clostridium butyricum, Brevibacterium lactofermentum, Streptococcus pyogenes, and the like. agalactiae, Lactococcus lactis, Leuconostoc lactis, Actinobacillus actinobycetemcomitans, cyanobacteria, Escherichia coli, Helicobacter pylori, Selnomonas ruminatium, Shigella sonnei, Zymomonas mobilis, Mycoplasma mycoides, Treponema denticola, Bacillus thuringiensis, Staphylococcus lugdunensis, lugdunensis, Leuconostoc oenos, Corynebacterium xerosis, Lactobacillus plantarumplantarum, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus acidophilus, Enterococcus faecalis, Bacillus coagulans, Bacillus cereus, Bacillus popillae, Synechocystis strain PCC6803, Bacillus liquefaciens, Pyrococcus abyssi, Selenomonas nominantium, Lactobacillus hilgardii, Streptococcus ferus, Lactobacillus pentosus, Bacteroides fragilis, Staphylococcus epidermidis, Streptomyces phaechromogenes, Streptomyces ghanaenis, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Serratia marcescens, Morganella morganii, morganii, Citrobacter freundii, Pseudomonas aerigunosa, Parvimonas micra, Prevotella intermediaintermedia, Fusobacterium nucleatum, Prevotella nigrescens, Actinomyces israelii, Porphyromonas endodontalis, Porphyromonas gingivalis, Micrococcus luteus, Bacillus megaterium, Aeromonas hydrophila, Aeromonas caviae, Bacillus anthracis, Bartonella henselae, Bartonella quintana Quintana, Borrelia pertussis, Borrelia burgdorferi, Borrelia garinii, Borrelia afzelii, Borrelia recurrentis, Brucella abortus, Brucella canis, Brucella melitensis, Brucella suis, Campylobacter jejuni, Campylobacter coli, Campylobacter fetus, Chlamydia pneumoniae, Chlamydia trachomatis trachomatis, Chlamydophila psittaci, Clostridium botulinum, Clostridium difficiledifficile, Clostridium perfringens, Clostridium tetani, Corynebacterium diphtheria, Cutibacterium acnes (formerly Propionibacterium acnes), Ehrlichia canis, Ehrlichia chaffeensis, Enterococcus faecium, Francisella tularensis, Haemophilus influenza, Legionella pneumophila, Leptospira interrogans interrogans, Leptospira santarosai, Leptospira weilii, Leptospira noguchii, Listeria monocytogenes, Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium ulcerans, Mycoplasma pneumonia, Neisseria gonorrhoeae, Neisseria meningitides, Nocardia asteroids, Rickettsia rickettsia, Salmonella enteritidis enteritidis, Salmonella typhi, Salmonella paratyphiparatyphi, Salmonella typhimurium, Shigella flexnerii, Shigella dysenteriae, Staphylococcus saprophyticus, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus viridans, Treponema pallidum, Ureaplasma urealyticum, Vibrio cholera, Vibrio parahaemolyticus, Yersinia pestis pestis, Yersinia enterocolitica, Yersinia pseudotuberculosis, Actinobacter baumanii, Pseudomonas aerigunosa, and mixtures thereof, preferably, the bacterium of interest is selected from the group consisting of Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanii, Pseudomonas aeruginosa, Enterobacter cloacae, and Enterobacter aerogenes, and mixtures thereof, more preferably, the bacterium of interest is selected from the group consisting of Escherichia coli strains.

[0021] In one embodiment, the bacterium of interest is selected from the group in Table 1 or Table 2, and the bacteriophage or phagemid does not contain at least one restriction site selected from the list of restriction sites in Table 1 or Table 2 corresponding to the bacterial group of interest. Preferably, the bacteriophage or phagemid does not contain a restriction site from the list of restriction sites in Table 1 or Table 2 corresponding to the bacterial group.

[0022] In one embodiment, the bacterium of interest is selected from an E. coli strain and the bacteriophage or phagemid does not contain at least one restriction site selected from the group consisting of CACNNNNNNNCTGG (SEQ ID NO: 1); AACNNNNNNGTGC (SEQ ID NO: 2); AACNNNNCTTT (SEQ ID NO: 4); CACNNNNGTAY (SEQ ID NO: 3); GGTCTC (SEQ ID NO: 36); CTGCAG (SEQ ID NO: 37); and GAAABCC (SEQ ID NO: 34). Preferably, the bacteriophage or phagemid does not contain the restriction sites CACNNNNNNNCTGG (SEQ ID NO: 1), CTGCAG (SEQ ID NO: 37), and GAAABCC (SEQ ID NO: 34). Optionally, the bacteriophage or phagemid does not contain the restriction sites CACNNNNNNNCTGG (SEQ ID NO:1), CTGCAG (SEQ ID NO:37), and GAAAABCC (SEQ ID NO:34), preferably the restriction sites CACNNNNNNNCTGG (SEQ ID NO:1); AACNNNNNNGTGC (SEQ ID NO:2); AACNNNNCTTT (SEQ ID NO:4); CACNNNNGTAY (SEQ ID NO:3); GGTCTC (SEQ ID NO:36); CTGCAG (SEQ ID NO:37); and GAAAABCC (SEQ ID NO:34).

[0023] Optionally, the bacteriophage is selected from the group consisting of IKe, CTX-φ, Pf1, Pf2, Pf3, Myoviridae (e.g., P1-like, P2-like, Mu-like, SPO1-like, and phiH-like bacteriophages); Siphoviridae (e.g., λ-like, γ-like, T1-like, c2-like, L5-like, psiM1-like, phiC31-like, and N15-like bacteriophages); Podoviridae (e.g., phi29-like, P22-like, and N4-like bacteriophages); Tectiviridae (e.g., Tectivirus); Corticoviridae (e.g., Corticovirus); Liposthrixviridae (e.g., Alphaliposthrixvirus, Betaliposthrixvirus, Gammaliposthrixvirus, and Desalisvirus). Rutaliposthrixvirus); Plasmaviridae (e.g., Plasmavirus); Rudiviridae (e.g., Rudiviridae); Fuselloviridae (e.g., Fusellovirus); Inoviridae (e.g., Inovirus, Pretovirus, M13-like and fd-like bacteriophages); Microviridae (e.g., Microvirus, Spiromicrovirus, Bdelomicrovirus, and Chlamydiamicrovirus); Leviviridae (e.g., Levivirus, Allolevivirus); Cystoviridae (e.g., Cystovirus), E. coli phage (e.g., infecting E. coli), B1 (e.g., infecting Bacteroides thetaiotamicron), ATCC 51477-B1, B40-8, or Bf-1 (e.g., infecting B. fragilis), phiHSC01 (e.g., infecting B. cacae), phiHSC02 (e.g., infecting B.ovatus), phiC2, phiC5, phiC6, phiC8, phiCD119, or phiCD27 (e.g., Clostridium difficile infection), KP01K2, K11, Kpn5, KP34, or JD001 (e.g., Klebsiella pneumoniae infection), phiNM1 or 80alpha (e.g., Staphylococcus aureus infection), IME-EF1 (e.g., Enterococcus faecalis infection), The bacteriophage is selected from the group consisting of ENB6 or C33 (e.g., infecting Enterococcus faecium), and phiKMV, PAK-P1, LKD16, LKA1, delta, sigma-1, J1 (e.g., infecting Pseudomonas aerignosa), T2, T4, T5, T7, RB49, phiX174, R17, PRD1, or the phagemid is packaged into the capsid of one of these bacteriophages.

[0024] Optionally, the phagemid is a lambda-derived phagemid, a P4-derived phagemid, an M13-derived phagemid, such as one that contains an f1 origin for filamentous phage packaging, e.g., pBluescript II SK(+ / -) and KS(+ / -) phagemids, pBC The phagemid according to the present invention is selected from the group consisting of SK and KS phagemids, pADL and P1 derived phagemids, preferably the phagemid according to the present invention is selected from lambda derived phagemids and P4 derived phagemids, more preferably the phagemid according to the present invention is selected from lambda derived phagemids, preferably HK022 derived phagemid, mEP237 derived phagemid, HK97 derived phagemid, HK629 derived phagemid, HK630 derived phagemid, mEP043 derived phagemid, mEP213 derived phagemid, mEP234 derived phagemid, mEP390 derived phagemid, mEP460 derived phagemid, mEPx1 derived phagemid, mEPx2 derived phagemid, phi80 derived phagemid, mEP234 derived phagemid.

[0025] In a second aspect, the present invention relates to the use of a bacteriophage or a packaged phagemid according to the invention for infecting a bacterium, preferably a bacterium selected from the above group of bacteria of interest.

[0026] In a third aspect, the present invention also relates to a pharmaceutical or veterinary composition comprising or essentially consisting of a bacteriophage or packaged phagemid according to the invention.

[0027] Furthermore, in a fourth aspect, the present invention relates to a bacteriophage or a packaged phagemid according to the invention, or a pharmaceutical or veterinary composition according to the invention, for use as a drug, in particular for improving the overall health of a subject, for eradicating pathogenic or virulent bacteria, for improving the efficacy of a drug and / or for modifying the composition of the microbiota, in particular for treating infectious diseases, inflammatory diseases, autoimmune diseases, cancer and brain disorders.

[0028] Preferably, the infection is a bacterial infection, preferably a bacterial infection caused by a bacterium selected from the bacterial group of interest. [Brief description of the drawings]

[0029] [Figure 1AB] Injection efficiency of wild-type lambda phage and lambda-based phagemids. A) A set of 89 E. coli strains was treated with wild-type lambda phage particles (48.5 kb genome) and injection efficiency was assessed by plaque formation. White, plaques present; grey, no plaques; cross, strain did not grow (not analyzed). B) The same strains shown in (A) were treated with packaged lambda phagemid carrying chloramphenicol resistance and a 3.3 kb DNA cargo encoding GFP. White, fluorescent colonies present; grey, no colonies; cross, strain was naturally resistant to chloramphenicol (not analyzed). [Figure 1C]C) Examples of strains from (A) and (B) showing that a 3.3 kb packaged lambda-based phagemid can be injected into a target strain (bottom row), whereas a lambda wild-type bacteriophage cannot be injected (top row). [Diagram 2] Transduction efficiency of lambda phagemids: 8kb type I and 8kb RF (A), 3kb type I and 3kb RF (B) for test strain 1 and permissive control strain (control) assessed by CFU. Lambda phagemids 8kb and 3kb RF have removed the restriction site recognized by the RM nuclease of test strain 1, while lambda phagemids 8kb type I and 3kb type II contain a single restriction site. The control strain is permissive for both RF and non-restriction free lambda phagemid. The dilution [10^(-number given)] of the phagemid used for transduction is indicated above each streak. The streak with the phagemid dilution that gave the highest number of individual countable CFU was used for calculation of the phagemid titer. [Diagram 3] Growth of 87 different E. coli strains after transduction with lambda phagemids: 8kb WT and 8kb RF (A), 3kb WT and 3kb RF (B). Individual strains are indicated by their position on the 96-well plate. Strains that showed growth (more than 10 colonies visible per plated spot) are shown in grey. 96-well plate positions not included in the analysis are shown in black. Position B7 contains E. coli MG1655, a permissive control strain that showed growth after transduction with all phagemids tested. A large number of strains were transduced with lambda phagemids 8kb and 3kb RF, in which most of the restriction sites recognized by E. coli RM nuclease have been removed, compared to the corresponding WT phagemid mutants in which no restriction sites have been depleted. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0030] The inventors have surprisingly discovered that the main obstacle for DNA delivery from phages / phagemids to target bacterial strains is not related to the properties of the bacteriophage, particularly the specificity of its tail fiber, but rather to the number of restriction sites present in its genome. Therefore, the same bacteriophage scaffold with the same tail fiber protein can deliver its DNA cargo with various efficiencies (ranging from 0 to 100%) according to the number of restriction sites present in its genome and recognized by the restriction enzymes of the targeted bacterial group. Thus, the inventors have discovered a new strategy for the design of engineered, highly efficient phagemid-based particles that rely on a reduced number of restriction sites, thereby improving the spectrum in the targeted bacterial group.

[0031] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used to practice or test the present invention, preferred methods and materials are described. All patents and scientific literature cited in this application attest to the level of knowledge in this field and are incorporated herein by reference. For clarity of explanation, the following terms are defined below.

[0032] As used herein, the term "nucleic acid" refers to at least two nucleotides covalently linked together and, in some cases, may include phosphodiester bonds (e.g., a phosphodiester "backbone"). In some embodiments, the nucleic acids of the present disclosure may be considered to be nucleic acid analogs that may contain other backbones, including, for example, phosphoramide, phosphorothioate, phosphorodithioate, O-methyl phosphoramidite linkages, and / or peptide nucleic acids. The nucleic acids of the present disclosure (e.g., components or portions of nucleic acids) may be naturally occurring or engineered. Engineered nucleic acids include recombinant and synthetic nucleic acids. "Recombinant nucleic acid" may refer to a molecule constructed by joining nucleic acid molecules and, in some embodiments, capable of replicating in a living cell. "Synthetic nucleic acid" may refer to a molecule synthesized or amplified by chemical or other means, including those that are chemically or otherwise modified but capable of base pairing with naturally occurring nucleic acid molecules. Recombinant and synthetic nucleic acids also include those molecules resulting from replication of any of the foregoing. Nucleic acids may be single-stranded (ss) or double-stranded (ds) or may contain portions of both single-stranded and double-stranded sequences, as specified. Nucleic acids may be DNA, both genomic and cDNA, RNA or hybrids / chimeras, where the nucleic acid contains any combination of deoxyribonucleotides and ribonucleotides, and any combination of bases including uracil, adenine, thymine, cytosine, guanine, inosine, xanthine, hypoxanthine, isocytosine, and isoguanine. Preferably, recombinant and synthetic nucleic acids are not naturally occurring, in particular they contain two sequences that are not found in the same nucleic acid molecule in nature.

[0033] As used herein, the terms "vector" and "DNA cargo" are synonymous and refer to a nucleic acid molecule, typically DNA or RNA, that serves to transfer a passenger nucleic acid sequence, i.e., DNA or RNA, to a host cell. A vector may contain an origin of replication, a selection marker, and optionally a site suitable for inserting genes, such as a multiple cloning site. There are several common types of vectors, including plasmids, bacteriophage genomes, phagemids, viral genomes, cosmids, and artificial chromosomes. A "vector" can refer to a phagemid or a bacteriophage genome.

[0034] As used herein, the term "expression vector" refers to a vector designed for gene expression in a cell. An expression vector allows the introduction of a particular gene into a target cell and can direct the cell's mechanism for protein synthesis to produce the protein encoded by that gene. An expression vector contains expression elements including, for example, a promoter, a ribosome binding site, and the correct translation initiation sequence, such as a start codon, a stop codon, and a transcription termination sequence. An expression vector may also contain other regulatory regions, such as enhancers, silencers, and boundary elements / insulators, to direct the transcription level of a given gene. An expression vector may be a vector for stable or transient expression of a gene.

[0035] As used herein, the term "delivery vehicle" refers to a structure or composition that allows the transfer of a vector into bacteria. There are several common types of delivery vehicles, including bacteriophage scaffolds, virus scaffolds, chemical-based delivery vehicles (e.g., cyclodextrin, calcium phosphate, cationic polymers, cationic liposomes), nanoparticle-based delivery vehicles (or platforms), non-chemical-based delivery vehicles (e.g., electroporation, sonoporation, optical transfection), particle-based delivery vehicles (e.g., gene guns, magnetofection, impalefection, particle bombardment, cell-penetrating peptides), or donor bacteria.

[0036] As used herein, the term "conjugative plasmid" refers to a plasmid that is transferred from one bacterial cell to another during conjugation, and a "donor bacterium" as used herein is a bacterium that can transfer a conjugative plasmid to another bacterium.

[0037] As used herein, the terms "bacteriophage" or "phage" are synonymous and refer to a virus that infects and replicates within bacteria. A bacteriophage is composed of a bacteriophage scaffold and a bacteriophage genome. The bacteriophage genome can be wild-type or engineered. In particular, the bacteriophage is an engineered bacteriophage. By "engineered" it is intended that the bacteriophage genome has been genetically engineered, in particular by removing some restriction sites.

[0038] As used herein, the terms "bacteriophage scaffold," "capsid," or "coat protein" are synonymous and refer to proteins that encapsulate a bacteriophage genome, phagemid, or plasmid. Preferably, they refer to a bacteriophage capsid or coat protein.

[0039] As used herein, the term "bacteriophage genome" refers to a DNA or RNA genome that is packaged in a bacteriophage scaffold. The bacteriophage genome can be wild-type or engineered.

[0040] The term "lethal bacteriophage" as used herein refers to a bacteriophage that causes the death of a bacterium after injecting its genome into the bacterial cytoplasm. Lethal bacteriophages according to the present invention include, but are not limited to, bacteriophages that have a lytic replication cycle that results in bacterial cell lysis.

[0041] The term "non-lethal bacteriophage" as used herein refers to a bacteriophage that does not cause the death of the bacterium after injecting its genome into the bacterial cytoplasm. In particular, the non-lethal bacteriophage according to the present invention exhibits a non-lytic replication cycle, which leaves the bacterial cell intact. The bacteriophage of the present invention may be, in some embodiments, a non-lethal bacteriophage.

[0042] In a preferred embodiment, the bacteriophage according to the invention is a lethal bacteriophage.

[0043] As used herein, the terms "phagemid" or "phasmid" are synonymous and refer to vectors derived from both plasmid and bacteriophage genomes. The phagemids of the present invention comprise a phage packaging site and, optionally, an origin of replication (ori), in particular a bacterial and / or phage origin of replication. In one embodiment, the phagemids according to the present invention do not comprise a bacterial origin of replication and therefore cannot replicate by themselves when injected into a bacterium. Alternatively, the phagemids according to the present invention comprise a plasmid origin of replication, in particular a bacterial and / or phage origin of replication. According to the present invention, the term phagemid is not limited to vectors having an f1 origin of replication.

[0044] As used herein, the term "packaged phagemid" or "phagemid-derived engineered particle" refers to a phagemid that is encapsidated in a bacteriophage scaffold or capsid. In particular, it refers to a bacteriophage scaffold or capsid that lacks a bacteriophage genome.

[0045] Packaged phagemids according to the invention can be produced using helper phage strategies well known to those skilled in the art. The helper phage contains all the genes encoding the structural and functional proteins essential for the phagemids according to the invention to be encapsidated.

[0046] The packaged phagemids according to the invention may be produced using satellite virus strategies, also known to those skilled in the art. Satellite viruses are subviral agents, composed of nucleic acids that depend on the co-infection of the host cell with a helper virus for all morphogenetic functions, whereas for all their episomal functions (integration and immunity, multicopy plasmid replication), the satellite is completely independent of the helper. In one embodiment, the satellite gene may code for a protein that promotes capsid size reduction of the helper phage, as described for the P4 Sid protein, which regulates the P2 capsid size to fit its smaller genome.

[0047] As used herein, the term "lethal packaged phagemid" refers to a packaged phagemid that results in the death of the bacterium after injection of its genome into the bacterial cytoplasm.

[0048] As used herein, the term "non-lethal packaged phagemid" refers to a phagemid that does not lead to the death of the bacteria after injection of its genome into the bacterial cytoplasm.

[0049] In a preferred embodiment, the packaged phagemid according to the invention is a lethal packaged phagemid.

[0050] Alternatively, the packaged phagemid according to the invention is a non-lethal packaged phagemid.

[0051] As used herein, the terms "promoter" and "transcription promoter" are synonymous and refer to a regulatory region of a nucleic acid sequence where the initiation and rate of transcription of the remainder of the nucleic acid sequence are regulated. A promoter may also contain subregions to which regulatory proteins and molecules, such as RNA polymerase and other transcription factors, may bind. A promoter drives the transcription of the nucleic acid sequence it controls. As used herein, a promoter is considered to be "operably linked" when it is in the correct functional location and orientation with respect to the nucleic acid sequence it controls so as to regulate ("drive") the initiation of transcription of that sequence.

[0052] Promoters can be classified as strong or weak according to their affinity for RNA polymerase (and / or sigma factors); this relates to how closely the promoter sequence resembles the ideal consensus sequence for polymerase or sigma factors. The strength of a promoter can depend on whether transcription initiation occurs at that promoter frequently or infrequently. Different promoters with different strengths can be used in the present invention to lead to different levels of gene / protein expression (e.g., the expression level initiated from an mRNA derived from a weak promoter is lower than that initiated from a strong promoter).

[0053] A promoter may be one that is naturally associated with a gene or sequence, as can be obtained by isolating the 5' non-coding sequences located upstream of the coding segment of a given gene or sequence. Such a promoter may be referred to as "endogenous." Similarly, an activator / enhancer may be one that is naturally associated with a nucleic acid sequence located either within or downstream or upstream of that sequence.

[0054] In some embodiments, the encoding nucleic acid segment may be placed under the control of a recombinant or heterologous promoter, which refers to a promoter that is not normally associated with an encoded nucleic acid sequence in its natural environment. A recombinant or heterologous enhancer refers to an enhancer that is not normally associated with a nucleic acid sequence in its natural environment.

[0055] As used herein, an "inducible promoter" is characterized by initiating or enhancing transcriptional activity in the presence of, affected by, or contacted by an inducer or inducer agent. An "inducer" or "inducer agent" can be an endogenous or usually exogenous condition, compound, or protein that induces transcriptional activity from an inducible promoter. Examples of inducible promoters for use herein include, but are not limited to, bacteriophage promoters (e.g., Plslcon, T3, T7, SP6, PL) and bacterial promoters (e.g., Pbad, PmgrB, Ptrc2, Plac / ara, Ptac, Pm), or hybrids thereof (e.g., PLlacO, PLtetO).

[0056] Examples of bacterial promoters for use in accordance with the present invention include, but are not limited to, positively regulated E. coli promoters, such as the positively regulated σ70 promoter (e.g., inducible pBad / araC promoter, Lux cassette lite promoter, modified lambda Prm promoter, plac Or2-62(positive), pBad / AraC containing additional REN sites, pBad, P(Las)TetO, P(Las)CIO, P(Rhl), Pu, FecA, pRE, cadC, hns, pLas, pLux), the "s" promoter (e.g., Pdps), the σ32 promoter (e.g., heat shock), and the σ54 promoter (e.g., glnAp2); negatively regulated E. coli promoters, such as the negatively regulated σ70 promoter (e.g., promoter(PRM+), modified lambda Prm promoter, TetR-TetR-4C P(Las)TetO, P(Las)CIO, P(Lac)IQ, RecA_DlexO_DLac01, dapAp, FecA, Pspac-hy, pel, plux-cl, plux-lac, CinR, CinL, glucose-regulated, modified Pr, modified Prm+, FecA, Pcya, rec A(SOS), RecA(SOS), EmrR_regulated, Betl_regulated, pLac_lux, pTet_Lac, pLac / Mnt, pTet / Mnt, LsrA / cI, pLux / cI, Lacl, LacIQ, pLacIQl, pLas / cI, pLas / Lux, pLux / Las, pRecA with LexA binding site, reverse BBa_R0011, pLacI / ara-1, pLacIq, rrnB PI, cadC, hns, PfhuA, pBad / araC, nhaA, OmpF, RcnR), σS promoter (e.g., Lutz-Bujard LacO with alternative sigma factor σ38), σ32 promoter (e.g., Lutz-Bujard LacO with alternative sigma factor σ32), σ54 promoter (e.g., glnAp 2); negatively regulated Bacillus subtilis (B.subtilis promoters, such as the repressible B. subtilis σA promoter (e.g., gram-positive IPTG-inducible, Xyl, hyperspunk), σ promoters, and BioFAB promoters disclosed in Mutalik VK et al. (Nature Methods, 2013, 10: 354-360, see especially the Supplementary Data) and the BioFAB website (http: / / biofab.synberc.org / data). Other inducible microbial and / or bacterial promoters can be used in accordance with the present invention. Inducible promoters for use in accordance with the present disclosure can be induced (or repressed) by one or more physiological conditions, such as changes in pH, temperature, radiation, osmolality, saline gradients, cell surface binding, and the concentration of one or more exogenous or endogenous inducers. Exogenous inducers or agents may include, but are not limited to, amino acids and amino acid analogs, sugars and polysaccharides, nucleic acids, protein transcription activators and inhibitors, cytokines, toxins, petroleum-based compounds, metal-containing compounds, salts, ions, enzyme substrate analogs, hormones, or combinations thereof.

[0057] Particularly preferred bacterial promoters for use in accordance with the present invention are the promoters from the Anderson collection (http: / / parts.igem.org / Promoters / Catalog / Anderson): The promoter may be selected from constitutive promoters controlled by σ70, such as BBa_J23100, BBa_J23101, BBa_J23102, BBa_J23103, BBa_J23104, BBa_J23105, BBa_J23106, BBa_J23107, BBa_J23108, BBa_J23109, BBa_J23110, BBa_J23111, BBa_J23112, BBa_J23113, BBa_J23114, BBa_J23115, BBa_J23116, BBa_J23117, BBa_J23118, and BBa_J23119.

[0058] In some embodiments of the invention, a promoter may or may not be used in conjunction with an "enhancer," which refers to a ds-acting regulatory sequence involved in the transcriptional activation of a nucleic acid sequence downstream of the promoter. The enhancer may be placed in any functional location before or after the promoter.

[0059] In some embodiments, the vector of the present invention may include a terminator sequence or terminator. A "terminator" as used herein is a nucleic acid sequence that stops transcription. A terminator may be unidirectional or bidirectional. It is composed of a DNA sequence that is involved in the specific termination of an RNA transcript by an RNA polymerase. A terminator sequence prevents the transcriptional activation of a downstream nucleic acid sequence by an upstream promoter. Thus, in certain embodiments, a terminator that terminates the production of an RNA transcript is contemplated. A terminator may be necessary in vivo to achieve a desired gene / protein expression level.

[0060] The most commonly used type of terminator is the forward terminator. When placed downstream of a nucleic acid sequence that is normally transcribed, a forward transcription terminator will stop transcription. In some embodiments, a bidirectional transcription terminator is provided, which will normally stop transcription on both the forward and reverse strands. In some embodiments, a reverse transcription terminator is provided, which will normally terminate transcription only on the reverse strand. In prokaryotic systems, terminators are generally classified into two categories: (1) rho-independent terminators, and (2) rho-dependent terminators. Rho-independent terminators are generally composed of palindromic sequences that form a stem-loop rich in GC base pairs, followed by a series of uracil bases.

[0061] Terminators for use according to the present invention include any transcription terminator described herein or known to those skilled in the art. Examples of terminators include, but are not limited to, gene termination sequences, such as the bovine growth hormone terminator, and viral terminator sequences, such as the TO terminator, TE terminator, lambda T1 and T1T2 terminators found in bacterial cells. In some embodiments, the termination signal may be a sequence that cannot be transcribed or translated, such as those resulting from sequence truncation.

[0062] Terminators for use according to the present invention also include those disclosed in Chen YJ et al. (2013, Nature Methods, 10: 659-664) and the BioFAB terminator disclosed in Cambray G et al. (Nucl Acids Res, 2013, 41(9): 5139-5148).

[0063] Other genetic elements are known in the art and can be used in accordance with the present disclosure.

[0064] As used herein, the term "codon usage table" refers to a database that provides codons, the amino acids coded for by each codon, and the frequency with which these codons are found for a defined type of amino acid.

[0065] As used herein, the term "codon usage satisfying" refers to optimizing the codon frequencies in a nucleic acid to be as close as possible to the codon frequencies in a codon usage table for the host or host group under consideration.

[0066] As used herein, the terms "modification," "alteration," and "mutation" are used interchangeably and refer to changes in an amino acid or nucleic acid sequence, such as substitutions, insertions, and / or deletions.

[0067] As used herein, a "substitution" refers to the replacement of a nucleotide or amino acid at a particular position in a parent nucleic acid or amino acid sequence with a different nucleotide or amino acid.

[0068] By "insertion" is meant the addition of a nucleic acid or amino acid at a particular position in a parent nucleic acid or amino acid sequence.

[0069] A "deletion" refers to the removal of a nucleotide or amino acid at a particular position in a parent nucleic acid or amino acid sequence.

[0070] Nucleotide substitutions can be neutral. A neutral nucleotide substitution is the replacement of a given nucleotide with another nucleotide, resulting in a codon that codes for the same amino acid. Codons that code for the same amino acid are provided by a codon usage table.

[0071] As used herein, the term "parent sequence" refers to a nucleic acid or amino acid sequence that is subsequently modified to produce a variant. The term also refers to a reference sequence, which may be a naturally occurring sequence or an engineered sequence, that is mutated or modified to generate a variant. A "parent sequence" may refer to a vector sequence, such as a phagemid sequence.

[0072] As used herein, the term "mutant sequence" or "mutant" is synonymous and refers to a nucleic acid or amino acid sequence that differs from that of a parent sequence by at least one modification. A mutant may contain one or several substitutions, and / or one or several insertions, and / or one or several deletions. In some embodiments, a nucleic acid mutant may contain one or several neutral substitutions. In some other embodiments, an amino acid mutant may contain one or several conservative substitutions.

[0073] As used herein, the terms "restriction site" and "restriction enzyme site" are synonymous and refer to a position on a nucleic acid that contains a specific sequence of nucleotides that is recognized by a restriction enzyme. In particular, the nucleic acid contains a specific sequence that is bound and cleaved by a restriction enzyme. A restriction site is generally a palindromic sequence that is 4-8 base pairs in length. More precisely, a restriction site refers to a specific sequence and modification state so that it is bound and cleaved by a restriction enzyme. In particular, it refers to a specific unmodified sequence so that it is bound and cleaved by a restriction enzyme. In particular, the sequence is not methylated, hydroxymethylated and glucosyl-hydroxymethylated. In this context, the restriction enzyme is of type I, II or III. Alternatively, it may refer to a specific modified sequence so that it is bound and cleaved by a restriction enzyme, for example, methylated, hydroxymethylated and glucosyl-hydroxymethylated DNA. In this context, the restriction enzyme is of type IV.

[0074] As used herein, "recognized by," in reference to a restriction site and a restriction enzyme, means that the restriction site is cleaved by the restriction enzyme.

[0075] In the restriction site sequences, N means that the nucleotide may be A, C, G, or T; B means that the nucleotide may be C, G, or T; Y means that the nucleotide may be C or T; W means that the nucleotide may be A or T; R means that the nucleotide may be A or G; D means A, G, or T.

[0076] As used herein, the terms "restriction enzyme" and "restriction endonuclease" are synonymous and refer to enzymes that cut nucleic acids at or near a restriction site. Restriction enzymes are generally classified into four types (type I to type IV). The REBASE database makes it possible to list the restriction sites that a given bacterium can recognize according to the restriction enzymes it expresses.

[0077] As used herein, the term "bacterium" or "bacteria" refers to any prokaryotic microorganism that exists as a single cell or as a cluster or aggregate of single cells. The term "bacteria" encompasses all variants of bacteria (e.g., endogenous bacteria naturally occurring in closed systems, environmental bacteria, or bacteria released for bioremediation or other activities). Bacteria of the present disclosure include bacterial subdivisions of Eubacteria and Archaea. Eubacteria can be further subdivided into Gram-positive and Gram-negative eubacteria, which depend on differences in cell wall structure. Also included herein are those classified solely on the basis of macroscopic morphology (e.g., cocci, bacilli). In some embodiments, the bacteria are Gram-negative cells, and in some embodiments, the bacteria are Gram-positive cells.

[0078] As used herein, the terms "microbiome" or "microbiota" are synonymous and refer literally to the ecological community of commensal, symbiotic, and pathogenic microorganisms that share the bodily space of a subject, preferably a human subject.

[0079] The microbiota can be specific to a given species, body area, body site, organ or tissue. Thus, as used herein, the term human microbiota refers to the ecological community of commensal, symbiotic, and pathogenic microorganisms that make up the human microbiota.

[0080] As used herein, the term "infection" refers to the invasion of an organism's body tissues by disease-causing bacteria, their proliferation, and the response of host tissues to these bacteria and ultimately the toxins they produce.

[0081] As used herein, the terms "infectious agent," "microbial agent," "pathogen," "disease-causing microorganism," and "virulent bacteria" are synonymous and refer to bacteria that cause infection.

[0082] Virulent bacteria according to the present invention also include antibacterial resistant bacteria.

[0083] As used herein, the term "selection marker" refers to a gene used to confirm the cloning of a gene or to confirm or ensure the presence of a plasmid in a bacterium. A selection marker can be a marker gene that provides a selectable phenotype, such as drug resistance, auxotrophy, resistance to cytotoxic agents, or surface protein expression. For example, antibiotic resistance genes, genes that allow overcoming auxotrophy, chromogenic enzyme genes, or luminescent / fluorescent genes can be used. This confers a "selective advantage" to bacteria carrying such a selection marker, allowing them to grow on media supplemented with antibiotics, heavy metals, or on media that lack essential components such as amino acids.

[0084] As used herein, the terms "antibiotic" and "antibacterial" are synonymous and refer to a type of antibacterial active ingredient used to treat and prevent bacterial infections.

[0085] As used herein, the term "antimicrobial resistance" refers to the ability of bacteria to resist the effects of drugs previously used to treat the bacteria.

[0086] As used herein, the terms "treatment," "treat," or "treating" refer to any action intended to improve the health status of a patient, for example, the treatment, prevention, prophylaxis, and delay of an infectious disease. In certain embodiments, such terms refer to the amelioration or eradication of an infectious disease or symptoms associated therewith. In other embodiments, the terms refer to minimizing the spread or worsening of an infectious disease resulting from the administration of one or more therapeutic agents to a subject with such a disease.

[0087] As used herein, the terms "subject", "individual" or "patient" are interchangeable and refer to animals, preferably mammals, and even more preferably humans, such as adults, children, newborns and prenatal stages of humans. However, the term "subject" can also refer to non-human animals, particularly mammals, such as dogs, cats, horses, cows, pigs, sheep, donkeys, rabbits, ferrets, gerbils, hamsters, chinchillas, rats, mice, guinea pigs and non-human primates, among others, in need of treatment.

[0088] The terms "quantity," "amount," and "dose" are used interchangeably herein and may refer to the absolute quantification of a molecule.

[0089] As used herein, the terms "active ingredient," "active ingredient," and "active pharmaceutical ingredient" are synonymous and refer to the component of a pharmaceutical composition that has a therapeutic effect.

[0090] As used herein, the term "therapeutic effect" refers to an effect induced by an active ingredient, pharmaceutical or veterinary composition, kit, product or combined preparation according to the invention that can prevent or delay the onset of an infectious disease, or cure or reduce the effects of an infectious disease.

[0091] As used herein, the term "effective amount" refers to an amount of active ingredient or pharmaceutical or veterinary composition that prevents, eliminates, or reduces the harmful effects of an infection. It will be apparent that the amount administered can be adapted by those skilled in the art depending on the subject to be treated, on the nature of the infection, etc. In particular, the dosage and regimen of administration will be a function of the nature, stage and severity of the infection to be treated, as well as the weight, age and general health of the subject to be treated, and the judgment of the physician.

[0092] As used herein, the term "consisting essentially of" is intended to refer to a pharmaceutical or veterinary composition that contains no other active ingredients.

[0093] As used herein, the term "about" refers to a range of values ​​of ±10% of a particular value. For example, "about 50" includes values ​​of 50 ±10%, i.e., values ​​in the range of 45 to 55. Preferably, the term "about" refers to a range of values ​​of ±5% of a particular value.

[0094] The present invention relates to a vector with a reduced number of restriction sites for a bacterial group of interest, a vector packaged in a bacteriophage scaffold, and the use of the vector packaged in a bacteriophage scaffold and a method of using the same. In other words, it has a small number of restriction sites corresponding to the restriction enzymes encoded by the bacterial group of interest. By small number, it is preferably understood that the vector contains 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 or less restriction sites. In a preferred embodiment, the vector does not have a restriction site for the restriction enzymes encoded by the bacterial group of interest or for the restriction enzymes that are highly present in the bacterial group of interest. However, the selection of the restriction sites to be removed from the vector depends on the number of bacteria from the bacteria of interest that encode the restriction enzymes.

[0095] Number of restriction sites The vectors according to the invention are such that they contain no or very few restriction sites for the restriction enzymes encoded by the bacterial group of interest.

[0096] More specifically, some restriction enzymes encoded by the bacterial group of interest are frequent, in particular very frequent, in the bacterial group of interest, and their restriction sites are rare or absent, preferably absent, in the vector according to the invention. By "rare" it is meant that there are one or two restriction sites in the vector. In a preferred embodiment, the restriction sites are absent in the vector according to the invention.

[0097] "High frequency" or "high frequency" in a bacterial population of interest means that at least 10, 20, 30, 40, 50, 60, 70, 75, 80, 85, 90, 95 or 99% of the bacterial population encodes a restriction enzyme. "Very high frequency" in a bacterial population of interest means that at least 50, 60, 70, 75, 80, 85, 90, 95 or 99% of the bacterial population encodes a restriction enzyme. When several restriction enzymes have the same restriction site, the number of these restriction enzymes is considered to determine the frequency. More specifically, the sum of the occurrences of these restriction enzymes is considered to determine the frequency. The frequency can be determined based on the coding sequences present in the bacteria, the genome of the bacteria, or the coding sequences of the extrachromosomal or mobile genetic elements (also called accessory genomes) of the bacteria, including but not limited to plasmid sequences, transposon sequences, chromosomal cassette sequences and genomic island (islet) sequences. A bacterium is considered to encode a restriction enzyme if its genome or its accessory genome as described above contains a sequence encoding the restriction enzyme.

[0098] In one embodiment, the vector according to the invention does not contain any restriction sites for restriction enzymes that are present at high or very high frequency in the bacterial population of interest.

[0099] If some restriction enzymes are encoded by the bacterial group of interest at low frequency, then those restriction sites are rare in the vector according to the invention. By "rare" it is meant that there are one or two restriction sites in the vector. In one embodiment, the restriction sites are absent in the vector according to the invention.

[0100] In certain embodiments, "high frequency" in a bacterial group of interest means that at least 10% of the bacteria in that group encode a restriction enzyme.

[0101] In certain embodiments, a vector according to the invention does not contain any restriction sites for restriction enzymes encoded by more than 10, 15, 20, or 25% of the bacteria of the bacterial group of interest.

[0102] Optionally, if less than 10% of the bacteria from the group encode a restriction enzyme, that restriction site can be present in the vector.

[0103] The vector according to the invention, preferably contained in the delivery vehicle, preferably in a bacteriophage capsid, contains 100 or less restriction sites. Preferably, the vector according to the invention, preferably contained in the delivery vehicle, contains 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 or less restriction sites. In a preferred embodiment, the vector according to the invention, preferably contained in the delivery vehicle, contains 10 or less restriction sites. In a most preferred embodiment, the vector according to the invention, preferably contained in the delivery vehicle, does not contain any restriction sites.

[0104] In a particular embodiment, the present invention relates to a vector, preferably a vector contained in a delivery vehicle, in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% or 100% of the restriction sites originally present in the vector have been removed. The term "originally present" as used herein refers to the number of restriction enzymes that are naturally present in the vector or that are present after construction or modification of the vector but before any attempt to reduce the number of restriction enzymes. A person skilled in the art can use a restriction site database, such as the REBASE database, to identify all known restriction sites present in a given vector. Preferably, the restriction sites to be removed are selected from the restriction sites of restriction enzymes that are present at high or very high frequency in the bacterial group of interest. Preferably, the vector, preferably the vector contained in the delivery vehicle, has at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% or 100% of the restriction sites originally present in the vector removed, preferably such that the restriction sites are for a restriction enzyme that is highly or very frequently present in the bacterial population of interest.

[0105] In a preferred embodiment, the present invention relates to a bacteriophage genome or phagemid comprising 100 or less recognized restriction sites. Preferably, the bacteriophage genome or phagemid according to the invention comprises 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 or less recognized restriction sites. In a preferred embodiment, the bacteriophage genome or phagemid according to the invention comprises 10 or less restriction sites recognized by restriction enzymes encoded by each bacterium of the bacterial group of interest. In a most preferred embodiment, the bacteriophage genome or phagemid according to the present invention does not comprise any restriction sites.

[0106] In a particular embodiment, the present invention relates to a bacteriophage genome or phagemid in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% of the restriction sites originally present in the bacteriophage genome or phagemid have been removed. Preferably, the restriction sites to be removed are selected from restriction sites of restriction enzymes that are present at high or very high frequency in the bacterial population of interest. Preferably, the vector, preferably the vector contained in the delivery vehicle, is such that at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% or 100% of the restriction sites originally present in the vector have been removed, the restriction sites being restriction sites of restriction enzymes that are present at high or very high frequency in the bacterial population of interest.

[0107] In a particular embodiment, the invention relates to a bacteriophage or packaged phagemid in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% of the restriction sites originally present in the bacteriophage or packaged phagemid have been removed, preferably the restriction sites are restriction sites for restriction enzymes that are present at high or very high frequency in the bacterial population of interest.

[0108] In a first embodiment, the restriction site is a restriction site recognised by the bacterial strain of interest, i.e. by a restriction enzyme expressed by the bacterial strain of interest.

[0109] In a second embodiment, the restriction site is for a restriction enzyme that is frequently encoded in the bacterial population of interest.

[0110] In a third embodiment, the restriction site is a restriction site recognized by each bacterial strain of the group of bacterial strains of interest, i.e. by a restriction enzyme expressed by each bacterial strain of the group of bacterial strains of interest. Preferably, the bacterial strains of the group of bacterial strains are derived from the same bacterial species.

[0111] In a fourth embodiment, the restriction site is a restriction site recognised by the bacterial species of interest, i.e. by a restriction enzyme expressed by the bacterial species of interest.

[0112] In a fifth embodiment, the restriction site is a restriction site recognized by each bacterial species of the group of bacterial species of interest, i.e. by a restriction enzyme expressed by each bacterial species of the group of bacterial species of interest. Preferably, the bacterial species of the group of bacterial species are from the same bacterial genus.

[0113] In a sixth embodiment, the restriction site is a restriction site recognised by the bacterial genus of interest, i.e. by a restriction enzyme expressed by the bacterial genus of interest.

[0114] In a seventh embodiment, the restriction sites are all restriction sites that can be recognised by a restriction enzyme expressed by bacteria, ie by all bacteria.

[0115] In an eighth embodiment, the restriction site is a restriction site recognised by a restriction enzyme expressed by a population of bacteria, such as, for example, bacteria from the human microbiome, i.e. by all bacterial populations.

[0116] A list of restriction sites recognized by the restriction enzymes of a bacterial strain, a group of bacterial strains, a bacterial species, a group of bacterial species, or a bacterial genus of interest can be established by a person skilled in the art based on a restriction site database such as REBASE. Alternatively, other databases well known to a person skilled in the art can be used, such as NCBI's RefSeq (www.ncbi.nlm.nih.gov / refseq / , in particular Release 86); Enterobase (enterobase.warwick.ac.uk, in particular accessed January 30, 2018); Genome Online Database (gold.jgi.doe.gov / , in particular Gold Release v-6); and Sanger (ftp.sanger.ac.uk / pub / project / pathogens / ).

[0117] To determine the frequency of each restriction enzyme in a bacterial group of interest, two complementary approaches can be taken, both of which rely on REBASE, a comprehensive database of restriction-modification (RM) systems (Roberts, R. et al., 2003, Nucleic Acids Research 31, 1805-1812). REBASE identifies RM genes and their cognate DNA recognition sites in a given bacterium by comparing the bacterial genome to a library of RM systems whose recognition sites have been experimentally confirmed. REBASE associates new recognition sites only if the two protein sequences are highly similar or identical (e.g., have 100% identity). More recently, single molecule real-time sequencing (SMRT) allows the identification of methylation patterns across the genome. Combined with the identification of RM systems, this allows the association of RM systems with their recognition sites. With the increasing number of available genomes and SMRT sequencing, the number of characterized RM systems is growing exponentially.

[0118] The first approach may consist of extracting from REBASE all type IS (including PacBIO data), type II R, type III R, RM gene names and their associated sites. Thanks to the gene nomenclature (EcoRI stands for Escherichia coli strain RY13 enzyme I) (Roberts, R. et al., 2015, Nucleic Acids Res 43, D298-D299), most species and strains of these RMs are identified and it is possible to estimate the most frequent sites within each species. In particular, RMs present in at least 10% of the strains of the bacterial species of interest are selected.

[0119] The second approach relies on the sequencing of a specific collection of strains of interest of a given bacterial species. The raw sequencing data can be assembled into contigs thanks to a genome assembler (Spages) (Bankevich, A. et al., 2012, J. Comput. Biol., 19, 455-77). The contigs are then annotated thanks to the RASTK (Brettin, T. et al., 2015, Sci Rep 5, 8365) or prokka pipelines (Seemann, T., 2014, Bioinformatics, 30, 2068-9). For each strain, all annotated proteins were blasted against each REBASE extracted list of RMs, using an e-value of 1e-120 as threshold. Only RM systems with a percentage of identity equal to or exceeding 80% and present in at least 10% of the strains of the bacterial species of interest are selected. The 10% threshold can be adapted to the chosen frequency.

[0120] In a particular embodiment, the present invention relates to a vector, preferably a vector contained in a delivery vehicle, that comprises 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 or less restriction sites from the list of restriction sites.Preferably, the vector, preferably the vector contained in a delivery vehicle, does not comprise any restriction site from the list of restriction sites.The above list of restriction sites comprises at least one restriction site sequence.Preferably, the above list of restriction sites comprises at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000 restriction site sequences. Preferably, said list of restriction sites comprises at least 1%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 99%, 100% of known restriction sites. More preferably, said list of restriction sites comprises at least 1%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 99%, 100% of the restriction sites known to be recognized by the restriction enzymes of a given bacterial genus, group of species, species, group of strains or strain of interest. In one embodiment, said list of restriction sites comprises restriction sites of restriction enzymes that are frequently encoded in the bacterial group of interest. In a very specific embodiment, said list of restriction sites comprises restriction sites of restriction enzymes that are present in at least 10% of bacteria from the bacterial group of interest. In a specific embodiment, the list is one of the lists provided in Table 1 or Table 2 for each bacterial group of interest.

[0121] The present invention then relates to a vector, preferably a vector contained in a delivery vehicle, that contains 0, 1 or 2 restriction sites from a list of restriction sites, preferably a list of restriction sites that contains a restriction site for a restriction enzyme that is frequently present in the bacterial group of interest, preferably present in at least 10% of the bacteria from the bacterial group of interest. In a particular embodiment, the list is one of the lists provided in Table 1 or Table 2 for each bacterial group of interest. Preferably, the vector does not contain any of the restriction sites from the list provided in Table 1 or Table 2 for the particular bacterial group of interest.

[0122] In a particular embodiment, the present invention relates to a vector, preferably a vector contained in a delivery vehicle, that does not contain one restriction site selected from the group consisting of a list of restriction sites that includes a restriction site of a restriction enzyme that is frequently present in the bacterial group of interest, preferably present in at least 10% of the bacteria from the bacterial group of interest. For example, the vector, preferably the vector contained in the delivery vehicle, does not contain 1, 2, 3, 4, 5, 6 or 7 restriction sites selected from the group consisting of a list of restriction enzymes, preferably a list of restriction sites that includes a restriction site of a restriction enzyme that is frequently present in the bacterial group of interest, preferably present in at least 10% of the bacteria from the bacterial group of interest. In a particular embodiment, the vector, preferably the vector contained in the delivery vehicle, does not contain a restriction site of a list of restriction enzymes, preferably a list of restriction sites that includes a restriction site of a restriction enzyme that is frequently present in the bacterial group of interest, preferably present in at least 10% of the bacteria from the bacterial group of interest. In a particular embodiment, the list is one of the lists provided in Table 1 or Table 2 for each bacterial group of interest. Preferably, the vector does not contain any of the restriction sites listed in Table 1 or Table 2 for the particular bacterial group of interest.

[0123] In certain embodiments, the vector does not contain any restriction sites for any restriction enzymes encoded in the bacterial population of interest.

[0124] Preferably, the vector according to the invention is a bacteriophage genome or a phagemid.

[0125] More preferably, the vector according to the invention is a bacteriophage or a packaged phagemid.

[0126] A person skilled in the art can establish such a list based on a database such as REBASE. In a particular embodiment, said list of restriction sites comprises all restriction enzyme sequences disclosed in the REBASE database. Alternatively, said list of restriction sites comprises all restriction enzyme sequences disclosed in the REBASE database that are recognized by the restriction enzymes of a given bacterial genus, species group, species, strain group or strain of interest.

[0127] The technique for modifying the number of restriction sites in vector is well known to those skilled in the art.Specific databases such as REBASE can be used to identify the restriction sites in vector and the corresponding restriction enzymes.Then, the restriction sites can be modified, for example, one by one, until the desired number or percentage of restriction sites is reached.

[0128] Modification of a restriction site means that the modified sequence of a restriction site exhibits at least one nucleotide difference from its unmodified sequence and that the restriction enzyme can no longer recognize the restriction site.

[0129] Modification of a restriction site can be accomplished by deletion of part or all of the restriction site or by nucleotide substitution of at least one nucleotide of the restriction site.

[0130] Preferably, restriction sites in non-coding regions of the vector are modified first, and only if the restriction sites in the coding regions of the vector do not reach the desired number or percentage of restriction sites, and then all restriction sites in the non-coding regions are modified.

[0131] Optionally, deletions are made only in non-coding regions.

[0132] Substitutions at restriction sites are preferably neutral substitutions, especially in coding regions. Codon usage tables can be used to determine potential neutral substitutions.

[0133] If neutral substitutions are not available at a restriction site and other restriction sites cannot be modified, nucleotide mutations that result in conservative amino acid mutations can be made.

[0134] If the restriction site contains an "N" type nucleotide, the substitution is made to a non-N nucleotide.

[0135] Preferably, the substitutions are selected to meet bacterial codon usage.

[0136] Vectors can be modified by mutation, for example by direct mutagenesis, or by any suitable technique known to those of skill in the art, or by nucleic acid synthesis if the sequence is designed (see, e.g., Huges RA et al., 2017, Cold Spring Harb Perspect Biol, 9:a023812).

[0137] In particular, restriction sites can be modified using classical cloning techniques well known to those of skill in the art, including but not limited to restriction enzyme cloning, Gibson assembly, and inverse PCR.

[0138] For example, bacteriophage can be mutated by yeast-mediated cloning (see, for example, Joska TM et al., 2014, Journal of Microbiological Methods, 100: 46-51). Other mutation methods include, but are not limited to, lambda red-mediated insertion / deletion (see, for example, Murphy KC, 1998, J Bacteriol., 180(8): 2063-71), multiplex automated genome engineering (MAGE) (see, for example, Wang HH et al., 2009, Nature, 460(7257): 894-898), and Cas9-mediated mutation (see, for example, Jiang Yu et al., 2015, Appl. Environ. Microbiol, 81(7): 2506-2514).

[0139] Bacteria of interest Preferably, the bacterial group of interest consists of a group of n bacterial species, n being a positive integer comprised between 1 and about 100, preferably between 1 and about 50, more preferably between 1 and about 10, even more preferably between 1 and about 5, and even more preferably the bacterial group of interest consists of a single bacterial species. Preferably, the bacterial species according to the present invention is selected from a single genus.

[0140] More preferably, the bacterial group of interest is a group of n bacterial strains, n being a positive integer between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 and about 1000, 10,000, or 100,000, preferably between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 and about 500, more preferably between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 and about 250, even more preferably between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 and about 100, even more preferably between 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 and about 50. In the most preferred embodiment, the bacterial group of interest is a group of n bacterial strains, n being a positive integer between 1 and about 30, preferably between 1 and about 20, more preferably between 1 and about 10, even more preferably between 1 and about 5. In a particular embodiment, the bacterial population of interest consists of a single bacterial strain. Preferably, the bacterial strains according to the invention are selected from a single species.

[0141] Bacterial groups of interest may include bacterial strains that have been reported to cause a defined pathology.

[0142] Bacteria of interest according to the invention may be eubacteria or archaebacteria, preferably eubacteria. Bacteria of interest according to the invention may be gram-positive or gram-negative eubacteria.

[0143] In a preferred embodiment, the bacteria of interest according to the present invention are toxin bacteria.

[0144] In another preferred embodiment, the bacteria of interest according to the present invention are antibacterial resistant bacteria.

[0145] In yet another preferred embodiment, the bacteria of interest are different strains of the same species.

[0146] In another preferred embodiment, the bacteria of interest are different strains of different species.

[0147] In another preferred embodiment, the bacteria of interest are different strains of different species of members of the human microbiota, in particular the gut microbiota.

[0148] Examples of bacteria of interest according to the present invention include, but are not limited to: Yersinia spp., Escherichia spp., Klebsiella spp., Acinetobacter spp., Bordetella spp., Neisseria spp., Aeromonas spp., Francisella spp., Corynebacterium spp., Citrobacter spp., Chlamydia spp., Haemophilus spp., Brucella spp., Mycobacterium spp., Legionella spp., Rhodococcus spp., Pseudomonas spp., Helicobacter spp., Vibrio spp., Bacillus spp., Erysipelothrix spp., Salmonella spp., Streptomyces spp., Streptococcus spp., Staphylococcus spp., Bacteroides spp., Prevotella spp., Clostridium spp., Bifidobacterium spp., Clostridium spp., Brevibacterium spp., Lactococcus spp., Leuconostoc spp., Actinobacillus spp., and S. cerevisiae. The bacteria include bacteria selected from Lunomonas spp., Shigella spp., Zymonas spp., Mycoplasma spp., Treponema spp., Leuconostoc spp., Corynebacterium spp., Enterococcus spp., Enterobacter spp., Pyrococcus spp., Serlazia spp., Morganella spp., Parvimonas spp., Fusobacterium spp., Actinomyces spp., Porphyromonas spp., Micrococcus spp., Bartonella spp., Borrelia spp., Campylobacter spp., Chlamydophilia spp., Ehrlichia spp., Haemophilus spp., Leptospira spp., Listeria spp., Mycoplasma spp., Nocardia spp., Rickettsia spp., Ureaplasma spp., Cutibacterium spp. (formerly Propionibacterium spp.), Lactobacillus spp., or mixtures thereof.

[0149] Preferably, the bacterium of interest according to the present invention is selected from the group consisting of Yersinia spp., Escherichia spp., Klebsiella spp., Acinetobacter spp., Pseudomonas spp., Helicobacter spp., Vibrio spp., Salmonella spp., Streptococcus spp., Staphylococcus spp., Bacteroides spp., Clostridium spp., Shigella spp., Enterococcus spp., Enterobacter spp., and Listeria spp.

[0150] More preferably, the bacterium of interest according to the present invention is selected from the Escherichia species.

[0151] In a preferred embodiment, the bacterium of interest according to the present invention is selected from the group consisting of Bacteroides thetaiotaomicron, Bacteroides fragilis, Bacteroides distasonis, Bacteroides vulgatus, Clostridium leptum, Clostridium coccoides, Staphylococcus aureus, Bacillus subtilis, Clostridium butyricum, Brevibacterium lactofermentum, Group B hemolytic streptococcus, Lactococcus lactis, Leuconostoc lactis, Actinobacillus actinobisetemcomitans, Cyanobacteria, Escherichia coli, Helicobacterium erythropoeis, Bacillus subtilis ... Helicobacter pylori, Obligate anaerobes, Shigella dysenteriae, Zymomonas mobilis, Mycoplasma mycoides, Treponema denticola, Bacillus thuringiensis, Staphylococcus lugdunensis, Leuconostoc oenos, Corynebacterium xerosis, Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus acidophilus, Enterococcus faecalis, Bacillus coagulans, Bacillus cereus, Bacillus popirae, Synechocystis strain PCC6803, Bacillus S. liqueffaciens, Pyrococcus abysii, Selenomonas nominantium, Lactobacillus hilgardii, Streptococcus ferus, Lactobacillus pentosus, Bacteroides fragilis, Staphylococcus epidermidis, Streptomyces faechromogenes, Streptomyces ganaenis, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Serrachia marcescens, Morganella morganii, Citrobacter freundii, Pseudomonas aerignon, sa, Parvimonas micra, Prevotella intermedia, Fusobacterium nucleatum, Prevotella nigrescens, Actinomyces israelii, Porphyromonas endodontalis, Porphyromonas gingivalis, Micrococcus luteus, Bacillus megaterium, Aeromonas hydrophila, Aeromonas caviae, Bacillus anthracis, Bartonella henselae, Bartonella quintana, Borrelia pertussis, Borrelia burgdorferi, Borrelia garinii, Borrelia afzelii, Borrelia leucorrhizias,Brucella abortus, Brucella canis, Brucella melitensis, Brucella suis, Campylobacter jejuni, Campylobacter coli, Campylobacter fetus, Chlamydia pneumoniae, Chlamydia trachomatis, Chlamydophila psittaci, Clostridium botulinum, Clostridium difficile, Clostridium felfringens, Clostridium tetani, Corynebacterium diphtheriae, Cutibacterium acnes (formerly Propionibacterium acnes), Ehrlichia canis, Ehrlichia chaffeensis, Enterococcus faecium, Francisella tularensis, Haemophilus influenzae, Legionella pneumophila, Leptospira interrogans, Leptospira santalosii, Leptospira weilii, Leptospira noguchii, Listeria monocytogenes, monocytogenes, Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium ulcerans, Mycoplasma pneumoniae, Neisseria gonorrhoeae, Neisseria meningitides, Nocardia asteroides, Rickettsia rickettsii, Salmonella enteritidis, Salmonella typhi, Salmonella paratyphi, Salmonella typhimurium, Shigella flexneri, Shigella dysenteriae, Staphylococcus suppurativa, lophiticus, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus viridans, Treponema pallidum, Ureaplasma urealyticum, Vibrio cholerae, Vibrio parahaemolyticus, Yersinia pestis, Yersinia enterocolitica, Yersinia pseudotuberculosis, Actinobacter baumannii, Pseudomonas aeruginosa, and mixtures thereof. Preferably, the bacterium of interest according to the invention is selected from the group consisting of Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter cloacae, and Enterobacter aerogenes, and mixtures thereof. More preferably, the bacterium of interest according to the invention is selected from Escherichia coli strains. Other bacteria may also be selected.

[0152] In another preferred embodiment, the bacterium of interest according to the present invention is an antibacterial resistant bacterium, preferably selected from the group consisting of extended-spectrum beta-lactamase producing (ESBL) Escherichia coli, ESBL Klebsiella pneumoniae, vancomycin-resistant Enterococcus (VRE), methicillin-resistant Staphylococcus aureus (MRSA), multidrug-resistant (MDR) Acinetobacter baumannii, MDR Enterobacter spp., carpavenem-resistant Enterobacteriaceae (CRE), colicitin-resistant E. coli, and combinations thereof. Preferably, the bacterium of interest according to the present invention is an antibacterial resistant bacterium selected from the group consisting of extended-spectrum beta-lactamase producing (ESBL) E. coli strains.

[0153] The bacterium of interest can also be selected from Shiga toxin-producing E. coli (STEC) and Verotoxigenic E. coli (VTEC).

[0154] The bacterium of interest may also be selected from enteropathogenic E. coli (EPEC), enterohemorrhagic E. coli (EHEC), enteroaggregative E. coli (EAEC), enterotoxigenic E. coli (ETEC), adherent and invasive E. coli (AIEC), uropathogenic E. coli (UPEC).

[0155] In yet another embodiment, the bacterium of interest according to the invention is a bacterium of the microbiota of a given species, preferably a bacterium of the human microbiota.

[0156] In a particular embodiment, the bacterium of interest is selected from an E. coli strain, and the restriction sites recognized by the restriction enzymes of the E. coli strain are: CACNNNNNNNCTGG (SEQ ID NO: 1), AACNNNNNNGTGC (SEQ ID NO: 2), CACNNNNGTAY (SEQ ID NO: 3), AACNNNNCTTT (SEQ ID NO: 4), CCANNNNNNNCTTC (SEQ ID NO: 5), TACNNNNNNNRTRTC (SEQ ID NO: 6), GAGNNNNNNNGTCA (SEQ ID NO: 7), TGANNNNNNNNTGCT (SEQ ID NO: 8), AGCANNNNNNTGA (SEQ ID NO: 9), TGA NNNNNNCTTC (SEQ ID NO: 10), GAGNNNNNGTTY (SEQ ID NO: 11), GATGNNNNNNTAC (SEQ ID NO: 12), GAANNNNNNRTCG (SEQ ID NO: 13), RTCANNNNNNCTC (SEQ ID NO: 14), GNAGNNNNRTDCA (SEQ ID NO: 15), GAANNNNNNNRTCG (SEQ ID NO: 16), GGANNNNNNNNATGC (SEQ ID NO: 17), GAGNNNNNTCC (SEQ ID NO: 18), CACNNNNNNNGTTG (SEQ ID NO: 19), YTCANNNNNNGTTY (SEQ ID NO: 20), GATGNNNN NCTG (SEQ ID NO: 21), CCAYNNNNNGTTY (SEQ ID NO: 22), RTCANNNNNNNNGTGG (SEQ ID NO: 23), GAANNNNNNNTAAA (SEQ ID NO: 24), TCANNNNNNNRTTC (SEQ ID NO: 25), GACNNNNNNGTC (SEQ ID NO: 26), TTCANNNNNNNNCTGG (SEQ ID NO: 27), TTANNNNNNNGTCY (SEQ ID NO: 28), CCANNNNNNNRTGC (SEQ ID NO: 29), CCANNNNNNNNTGAA (SEQ ID NO: 30), GAGNNNNNNNATGC (SEQ ID NO: 31), CAGN NNNNNCGT (SEQ ID NO: 32), GATGNNNNNGGC (SEQ ID NO: 33), GAAABCC (SEQ ID NO: 34), CCWGG (SEQ ID NO: 35), GGTCTC (SEQ ID NO: 36), CTGCAG (SEQ ID NO: 37), GCCGGC (SEQ ID NO: 38), RGGNCCY (SEQ ID NO: 39), GTCGAC (SEQ ID NO: 40), GCGCGC (SEQ ID NO: 41), RCCGGY (SEQ ID NO: 42), CCNGG (SEQ ID NO: 43), AAGCTT (SEQ ID NO: 44), CANCATC (SEQ ID NO: 45), GRCGYC (SEQ ID NO: 46), CYCGRG (SEQ ID NO: 47),GCNGC (SEQ ID NO: 48), YGGCCR (SEQ ID NO: 49), CCGCGG (SEQ ID NO: 50), GRGCYC (SEQ ID NO: 51), CTGAAG (SEQ ID NO: 52), GGWCC (SEQ ID NO: 53), TGGCCA (SEQ ID NO: 54), CCWWGG (SEQ ID NO: 55), GGNCC (SEQ ID NO: 56), GAGCTC (SEQ ID NO: 57), GGTACC (SEQ ID NO: 58), GGCGCC (SEQ ID NO: 59), ACCYAC (SEQ ID NO: 60), GAATTC (SEQ ID NO: 61), GATATC (SEQ ID NO: 62), CCTNAGG (SEQ ID NO: 63), GGTNACC (SEQ ID NO: 64), ATGCAT (SEQ ID NO: 65), GGYRCC (SEQ ID NO: 66), AGGCCT (SEQ ID NO: 67), CTCAAT (SEQ ID NO: 68), GCWGC (SEQ ID NO: 69), TCGCGA (SEQ ID NO: 70), CCTNNNNNAGG (SEQ ID NO: 71), ACCACC (SEQ ID NO: 72), CACAG (SEQ ID NO: 73), GAACC (SEQ ID NO: 74), GAGAC (SEQ ID NO: 75), CAGCAG (SEQ ID NO: 76), AGACC (SEQ ID NO: 77), and CCGAG (SEQ ID NO: 78).

[0157] In certain embodiments, the restriction enzyme of E. coli is the following type I restriction enzyme: CACNNNNNNNCTGG (SEQ ID NO: 1), AACNNNNNNGTGC (SEQ ID NO: 2), CACNNNNGTAY (SEQ ID NO: 3), AACNNNNCTTT (SEQ ID NO: 4), CCANNNNNNNCTTC (SEQ ID NO: 5), TACNNNNNNNRTRTC (SEQ ID NO: 6), GAGNNNNNNNGTCA (SEQ ID NO: 7), TGANNNNNNNNTGCT (SEQ ID NO: 8), AGCANNNNNNTGA (SEQ ID NO: 9), TGANNNNNNCTTC (SEQ ID NO: 10), GAGNNNNNGTTY (SEQ ID NO: 11), GATGNNNNNNTAC (SEQ ID NO: 12), GAANNNNNNRTCG (SEQ ID NO: 13), RTCANNNNNNCTC (SEQ ID NO: 14), GNAGNNNNRTDCA (SEQ ID NO: 15), GAANNNNNNNRTCG (SEQ ID NO: 16), GGANNNNNNNNATGC (SEQ ID NO: 17), 7), GAGNNNNNTCC (SEQ ID NO: 18), CACNNNNNNNGTTG (SEQ ID NO: 19), YTCANNNNNNGTTY (SEQ ID NO: 20), GATGNNNNNCTG (SEQ ID NO: 21), CCAYNNNNNGTTY (SEQ ID NO: 22), RTCANNNNNNNNGTGG (SEQ ID NO: 23), GAANNNNNNNTAAA (SEQ ID NO: 24), TCANNNNNNNRTTC (SEQ ID NO: 25), GACNNNNNNGTC (SEQ ID NO: 26), TTCANNNNNNNNCTGG (SEQ ID NO: 27), TTANNNNNNNGTCY (SEQ ID NO: 28), CCANNNNNNNRTGC (SEQ ID NO: 29), CCANNNNNNNNTGAA (SEQ ID NO: 30), GAGNNNNNNNATGC (SEQ ID NO: 31), CAGNNNNNNCGT (SEQ ID NO: 32), GATGNNNNNGGC (SEQ ID NO: 33).

[0158] In another particular embodiment, the restriction enzyme of E. coli is one of the following type II restriction enzymes: GAAABCC (SEQ ID NO: 34), CCWGG (SEQ ID NO: 35), GGTCTC (SEQ ID NO: 36), CTGCAG (SEQ ID NO: 37), GCCGGC (SEQ ID NO: 38), RGGNCCY (SEQ ID NO: 39), GTCGAC (SEQ ID NO: 40), GCGCGC (SEQ ID NO: 41), RCCGGY (SEQ ID NO: 42), CCNGG (SEQ ID NO: 43), AAGCTT (SEQ ID NO: 44), CANCATC (SEQ ID NO: 45), GRCGYC (SEQ ID NO: 46), CYCGRG (SEQ ID NO: 47), GCNGC (SEQ ID NO: 48), YGGCCR (SEQ ID NO: 49), CCGCGG (SEQ ID NO: 50), GRGCYC (SEQ ID NO: 51), CTGAAG (SEQ ID NO: 52), GGWCC (SEQ ID NO: 53), 3), TGGCCA (SEQ ID NO: 54), CCWWGG (SEQ ID NO: 55), GGNCC (SEQ ID NO: 56), GAGCTC (SEQ ID NO: 57), GGTACC (SEQ ID NO: 58), GGCGCC (SEQ ID NO: 59), ACCYAC (SEQ ID NO: 60), GAATTC (SEQ ID NO: 61), GATATC (SEQ ID NO: 62), CCTNAGG (SEQ ID NO: 63), GGTNACC (SEQ ID NO: 64), ATGCAT (SEQ ID NO: 65), GGYRCC (SEQ ID NO: 66), AGGCCT (SEQ ID NO: 67), CTCAAT (SEQ ID NO: 68), GCWGC (SEQ ID NO: 69), TCGCGA (SEQ ID NO: 70), CCTNNNNNAGG (SEQ ID NO: 71), ACCACC (SEQ ID NO: 72).

[0159] In yet another specific embodiment, the E. coli restriction enzymes are the following type III restriction enzymes: CACAG (SEQ ID NO: 73), GAACC (SEQ ID NO: 74), GAGAC (SEQ ID NO: 75), CAGCAG (SEQ ID NO: 76), AGACC (SEQ ID NO: 77), and CCGAG (SEQ ID NO: 78).

[0160] In another particular embodiment, the bacterial restriction enzyme of interest is selected from type I restriction enzymes and / or type II restriction enzymes and / or type III restriction enzymes and / or type IV restriction enzymes. Preferably, the bacterial restriction enzyme of interest is selected from type I restriction enzymes, type II restriction enzymes, type III restriction enzymes, and type IV restriction enzymes.

[0161] In a particular embodiment, list A of restriction sites for restriction enzymes frequently encoded by the bacterial group of interest derived from E. coli comprises: - CACNNNNNNNCTGG (SEQ ID NO: 1) (present in 24.5% of E. coli reference strains in REBASE); - AACNNNNNNGTGC (SEQ ID NO: 2) (present in 18% of E. coli reference strains in REBASE); - AACNNNNCTTT ((SEQ ID NO: 4) (present in 12% of E. coli reference strains in REBASE); - CACNNNNGTAY (SEQ ID NO: 3) (present in 11% of E. coli reference strains in REBASE). - GGTCTC (SEQ ID NO: 36) (present in 18% of E. coli reference strains in REBASE); - CTGCAG (SEQ ID NO: 37) (present in 18% of E. coli reference strains in REBASE); and - GAAABCC (SEQ ID NO: 34) (present in 26% of E. coli reference strains in REBASE).

[0162] In one embodiment, the invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the species E. coli, that does not contain 1, 2, 3, 4, 5, 6 or 7 restriction sites selected from the group consisting of this list A of restriction sites. In a particular embodiment, the invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the species E. coli, that does not contain any of the restriction sites CACNNNNNNNCTGG (SEQ ID NO: 1), CTGCAG (SEQ ID NO: 37) and GAAABCC (SEQ ID NO: 34). Preferably, it relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the species E. coli, that does not contain any of the restriction sites of this list A of restriction sites.

[0163] In another particular embodiment, the list B of restriction sites for restriction enzymes frequently encoded by the group of bacteria of interest derived from E. coli STEC or a subgroup of this species comprises: - TACNNNNNNNRTRTC (SEQ ID NO: 6) (present in 24% of STEC strain collections); - CACNNNNNNNCTGG (SEQ ID NO: 1) (present in 24% of the STEC strain collection); - CTGCAG (SEQ ID NO: 37) (present in 27% of the STEC strain collection); - GAAABCC (SEQ ID NO: 34) (present in 20% of the STEC strain collection); and - GATCAG (SEQ ID NO: 79) (present in 15% of the STEC strain collection).

[0164] In one embodiment, the invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the E. coli STEC species, that does not contain one, two, three, four or five restriction sites selected from the group consisting of this list B of restriction sites. In a particular embodiment, the invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the E. coli STEC species, that does not contain any of the restriction sites CACNNNNNNNCTGG (SEQ ID NO: 1), CTGCAG (SEQ ID NO: 37) and GAAABCC (SEQ ID NO: 34). Preferably, it relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest belonging to the E. coli STEC species, that does not contain any of the restriction sites of this list B of restriction sites.

[0165] [Table 1A]

[0166] [Table 1B]

[0167] [Table 1C]

[0168]

Table 1D

[0169]

Table 1E

[0170]

Table 2A

[0171]

Table 2B

[0172]

Table 2C

[0173]

Table 2D

[0174]

Table 2E

[0175]

Table 2F

[0176]

Table 2G

[0177]

Table 2H

[0178]

Table 2I

[0179]

Table 2J

[0180]

Table 2K

[0181]

Table 2L

[0182]

Table 2M

[0183]

Table 2N

[0184]

Table 2O

[0185]

Table 2P

[0186]

Table 2Q

[0187] In one embodiment, the present invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group or subgroup of bacteria shown in Table 1 or Table 2 above, and does not contain at least one restriction site selected from the group consisting of the list of restriction sites in Table 1 or Table 2 corresponding to the group of bacteria, for example 1, 2, 3, 4, 5, 6 or 7 restriction sites selected from the group consisting of the list of restriction sites in Table 1 or Table 2. Preferably, the present invention relates to a vector, preferably comprised in a delivery vehicle, designed for a group of bacteria of interest shown in Table 1 or Table 2 or a subgroup thereof, and does not contain any of the restriction sites in this list of restriction sites in Table 1 or Table 2 corresponding to the group of bacteria.

[0188] In yet another embodiment, the present invention relates to a bacteriophage genome or phagemid comprising no more than 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 restriction sites recognized by the restriction enzymes encoded by each bacterium of the bacterial group of interest. In a preferred embodiment, the bacteriophage genome or phagemid according to the present invention comprises no more than 10 restriction sites recognized by the restriction enzymes encoded by each bacterium of the bacterial group of interest. In a most preferred embodiment, the bacteriophage genome or phagemid according to the present invention does not comprise any restriction sites recognized by the restriction enzymes encoded by each bacterium of the bacterial group of interest.

[0189] In an alternative embodiment, the present invention relates to a bacteriophage genome or phagemid in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% of the restriction sites recognized by the restriction enzymes encoded by each bacterium of the bacterial group of interest and originally present in the vector have been removed.

[0190] In another alternative embodiment, the present invention relates to a bacteriophage genome or phagemid in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% of the restriction sites originally present in the vector and listed in the table above for the bacterial group of interest have been removed.

[0191] In an alternative embodiment, the invention relates to a bacteriophage or packaged phagemid in which at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 99% of the restriction sites recognized by restriction enzymes encoded by each bacterium of the bacterial group of interest and originally present in the vector have been removed.

[0192] The present invention further relates to a vector as defined above packaged in a bacteriophage capsid, the bacteriophage capsid being suitable for targeting a bacterial population of interest. The present invention also relates to a pharmaceutical composition comprising the vector as defined above packaged in a bacteriophage capsid, the use of the vector as defined above packaged in a bacteriophage capsid as a drug, and a method for treating a disease in a subject, comprising administering to the subject the vector as defined above packaged in a bacteriophage capsid. More specifically, the present invention relates to the use of the vector as defined above packaged in a bacteriophage capsid for delivering the vector to a bacterial population of interest.

[0193] Bacteria of the genus Actinomyces can be infected by the following phages: Av-1, Av-2, Av-3, BF307, CT1, CT2, CT3, CT4, CT6, CT7, CT8 and 1281, or by vectors packaged in the capsids of the above phages.

[0194] Bacteria of the genus Aeromonas can be infected by the following phages: AA-1, Aeh2, N, PM1, TP446, 3, 4, 11, 13, 29, 31, 32, 37, 43, 43-10T, 51, 54, 55R.1, 56, 56RR2, 57, 58, 59.1, 60, 63, Aehl, F, PM2, 1, 25, 31, 40RR2.8t, (syn=44R), (syn=44RR2.8t), 65, PM3, PM4, PM5 and PM6, or by vectors packaged in the capsids of the above phages.

[0195] Bacillus bacteria are the following phages: A, aizl, Al-K-I, B, BCJAl, BCl, BC2, BLLl, BLl, BP142, BSLl, BSL2, BSl, BS3, BS8, BS15, BS18, BS22, BS26, BS28, BS31, BS104, BS105, BS106, BTB, B1715V1, C, CK-I, Coll, Corl, CP-53, CS-I, CSi, D, D, D, D5, entl, FP8, FP9, FSi, FS2, FS3, FS5, FS8, FS9, G, GH8, GT8, GV-I, GV-2, GT-4, g3, gl2, gl3, gl4, gl6, gl7, g21, g23, g24, g29, H2, kenl, KK-88, Kuml, Kyul, J7W-1, LP52, (syn=LP-52), L7, Mexl, MJ-I, mor2, MP-7, MPlO, MP12, MP14, MP15, Neol, N°2, N5, N6P, PBCl, PBLA, PBPl, P2, S-a, SF2, SF6, Shal, Sill, SP02, (syn=ΦSPP1), SPβ, STI, STi, SU-Il, t, TbI, Tb2, Tb5, TbIO, Tb26, Tb51, Tb53, Tb55, Tb77, Tb97, Tb99, Tb560, Tb595, Td8, Td6, Tdl5, TgI, Tg4, Tg6, Tg7, Tg9, TgIO, TgIl, Tgl3, Tgl5, Tg21, Tinl, Tin7, Tin8, Tinl3, Tm3, Tocl, Togl, toll, TP-I, TP-10vir, TP-15c, TP-16c, TP-17c, TP-19, TP35, TP51, TP-84, Tt4, Tt6, A type, B type, C type, D type, E type, Tφ3, VA-9, W, wx23, wx26, Yunl, α, γ, pll, φmed-2, φT, φμ-4, φ3T, φ75, φlO5, (syn=φlO5), IA, IB, 1-97A, 1-97B, 2, 2, 3, 3, 3, 5, 12, 14, 20, 30, 35, 36, 37, 38, 41C, 51, 63, 64, 138D, I, II, IV, NN-Bacillus(13), alel, ARl, AR2, AR3, AR7, AR9, Bace-11, (syn=11), Bastille, BLl, BL2, BL3, BL4, BL5, BL6, BL8, BL9, BP124, BS28, BS80, Ch, CP-51, CP-54, D-5,darl, denl, DP-7, entl, FoSi, FoS2, FS4, FS6, FS7, G, gall, GEl, GF-2, G Si, GT-I, GT-2, GT-3, GT-4, GT-5, GT-6, GT-7, GV-6, gl5, 19, 110, ISi, K, MP9 MP13, MP21, MP23, MP24, MP28, MP29, MP30, MP32, MP34, MP36, MP37, MP39, MP4 MP41, MP43, MP44, MP45, MP47, MP50, NLP-I, No.l, N17, N19, PBSl, PKl, PMBl. PMB12, PMJl, S, SPOl, SP3, SP5, SP6, SP7, SP8, SP9, SPlO, SP-15, and SP50 (syn=S P-50)、SP82、SST、subl、SW、Tg8、Tgl2、Tgl3、Tgl4、thul、thuΛ、thuS、Tin4、Ti n23, TP-13, TP33, TP50, TSP-I, V, VI, V, Vx, β22, φe, φNR2, φ25, φ63, 1, 1, 2 2C, 3NT, 4, 5, 6, 7, 8, 9, 10, 12, 12, 17, 18, 19, 21, 138, III, 4(B. megateriwn))、4(バル·スァァ・(B. sphaericus)) AR13 BPP-IO BS32 BS107 B1 B2 GA-I GP -IO, GV-3, GV-5, g8, MP20, MP27, MP49, Nf, PP5, PP6, SF5, Tg l8, TP-I, Versailles, φ15, φ29, 1-97, 837 / IV, mi-バチルス(1) BatlO, BSLlO, BSLI1, BS6, BSI1, BS16, BS23, BS101, BS102 gl8, morl, PBLl, SN45, thu2, thu3, TmI, Tm2, ​​TP-20, TP21, T P52, BacF, BacG, Bac IV, HN-BacMus(3), BLE (syn=θc), BS2 BS4, BS5, BS7, B10, B12, BS20, BS21, F, MJ-4, PBA12, AP50, A P50-04, AP50-11, AP50-23, AP50-26, AP50-27, and Bam35.or by vectors packaged in the capsids of the above phages. The following Bacillus-specific phages have been deleted: DLP10716, DLP-11946, DPB5, DPB12, DPB21, DPB22, DPB23, GA-2, M, No. 1M, PBLB, PBSH, PBSV, PBSW, PBSX, PBSY, PBSZ, phi, SPa, Type 1 and μ.

[0196] Bacteroides bacteria can be infected by the following phages: ad I2, Baf-44, Baf-48B, Baf-64, Bf-1, Bf-52, B40-8, F1, β1, φA1, φBr01, φBrO2, 11, 67.1, 67.3, 68.1, mt-Bacteroides (3), Bf42, Bf71, HN-Bdellovibrio (1), and BF-41, or by vectors packaged into the capsids of the above phages.

[0197] Bacteria of the genus Bordetella can be infected by the following phages: 134 and NN-Bordetella (3) or by vectors packaged in the capsids of the above phages.

[0198] Bacteria of the genus Borrelia can be infected by the following phages: NN-Borrelia (1) and NN-Borrelia (2) or by vectors packaged in the capsids of the above phages.

[0199] Brucella bacteria are resistant to the following phages: A422, Bk, (syn=Berkeley), BM29, FOi, (syn=FOl), (syn=FQl), D, FP2, (syn=FP2), (syn=FD2), Fz, (syn=Fz75 / 13), (syn=Firenze75 / 13), (syn=Fi), Fi, (syn=Fl), Fim, (syn=FIm), (syn= Fim), FiU, (syn=FlU), (syn=FiU), F2, (syn=F2), F3, (syn=F3), F4, (syn=F4), F5, (syn=F5), F6, F7, (sy n=F7), F25, (syn=F25), (syn=￡25), F25U, (syn=F25u), (syn=F25U), (syn=F25V), F44, (syn-F44), F45, (syn=F45), F48, (syn=F48), I, Im, M, MC / 75, M51, (syn=M85), P, (syn=D), S708, R, Tb, (syn=TB), (syn=T bilisi), W, (syn=Wb), (syn=Weybridge), X, 3, 6, 7, 10 / 1, (syn=10), (syn=F8), (syn=F8), 12m, 24 / 11, ( It can be infected by (syn=24), (syn=F9), (syn=F9), 45 / 111, (syn=45), 75, 84, 212 / XV, (syn=212), (syn=Fi0), (syn=FlO), 371 / XXIX, (syn=371), (syn=Fn), (syn=Fll) and 513, or by vectors packaged in the capsids of the above-mentioned phages.

[0200] Bacteria of the genus Burkholderia can be infected by the following phages: CP75, NN-Burkholderia (1) and 42, or by vectors packaged in the capsids of the above phages.

[0201] Campylobacter bacteria are identified by the following phages: C type, NTCC12669, NTCC12670, NTCC12671, NTCC12672, NTCC12673, NTCC12674, NTCC12675, NTCC12676, NTCC12677, NTCC12678, NTCC12679, NTCC12680, NTCC12681, NTCC12682, NTCC12683, NTCC12684, NTCC12685, NTCC12686, NTCC12687, NTCC12688, NTCC12689, NTCC12690, NTCC12691, NTCC12692, NTCC12693, NTCC12694, NTCC12695, NTCC12696, NTCC12697, NTCC12698, NTCC12699, NTCC12680, NTCC12681, NTCC12698, NTCC12699, NTCC12690, NTCC12691, NTCC12692, NTCC12693, NTCC12694, NTCC12695, NTCC12696, NTCC12697, NTCC12698, NTCC12699, NTCC12610, NTCC12611, NTCC12612, NTCC12613, NTCC12614, NTCC12615, NTCC12616, NTCC12617, NTCC12618, NTCC12619, NTCC12620, NTCC12621, NTCC12622, NTCC12623, NTCC12624, NTCC12625, NTCC126 3, NTCC12684, 32f, 111c, 191, NN-Campylobacter (2), Vfi-6, (syn=V19), VfV-3, V2, V3, V8, V16, (syn=Vfi-1), V19, V20(V45), V45, (syn=V-45) and NN-Campylobacter (1), or by vectors packaged in the capsids of the above phages.

[0202] Bacteria of the genus Chlamydia can be infected by the following phages: Chpl, or by vectors packaged in the phage capsids mentioned above.

[0203] Clostridium bacteria are classified as follows: CAKl, CA5, Ca7, CEβ, (syn=1C), CEγ, Cldl, c-n71, c-203 Tox-, DEβ, (syn=ID), (syn=1Dt0X+), HM3, KMl, KT, Ms, NAl, (syn=Naltox+), PA135Oe, Pfo, PL73, PL78, PL81, Pl, P50, P5771, P19402, 1Ct0X+, 2Ct0X\2D3 (syn=2Dt0X+), 3C, (syn=3Ctox+), 4C, (syn=4Ct0X+), 56, III-1, NN-Clostridium (61), NBlt0X+, αl, CAl, HMT, HM2, PFl5 P-23, P-46, Q-05, Q-oe, Q-16, Q-21, Q-26, Q-40, Q-46, S111, SA02, WA01, WA03, Wm, W523, 80, C, CA2, CA3, CPTl, CPT4, cl, c4, c5, HM7, H11 / A1, H18 / Ax, FWS23, Hi58ZA1, K2ZA1, K21ZS23, ML, NA2t0X;Pf2, Pf3, Pf4, It can be infected by S9ZS3, S41ZA1, S44ZS23, α2, 41, 112ZS23, 214 / S23, 233 / Ai, 234 / S23, 235 / S23, II-1, II-2, II-3, NH-Clostridium (12), CA1, F1, K, S2, 1, 5 and NH-Clostridium (8), or by vectors packaged in the capsids of the above-mentioned phages.

[0204] Corynebacterium spp. was identified by the following phages: CGKl (deletion), A, A2, A3, AlOl, A128, A133, A137, A139, A155, A182, B, BF, B17, B18, B51, B271, B275, B276, B277, B279, B282, C, capi, CCl, CGl, CG2, CG33, CL31, Cog, (syn=CG5), D, E, F, H, HI, hqi, hq2, 11ZH33, Ii / 31, J, K, K, (syn=Ktox"), L, L, (syn=Ltox+), M, MC-I, MC-2, MC-3, MC-4, MLMa, N, O, ovi, o It can be infected by v2, ov3, P, P, R, RP6, RS29, S, T, U, UB1, ub2, UH1, UH3, uh3, uh5, uh6, β, (syn=βtox+), βhv64, βvir, γ, (syn=γtoχ-), γl9, δ, (syn=δ'ox+), p, (syn=ptoχ-), Φ9, φ984, ω, IA, 1 / 1180, 2, 2 / 1180, 5 / 1180, 5ad / 9717, 7 / 4465, 8 / 4465, 8ad / 10269, 10 / 9253, 13Z9253, 15 / 3148, 21 / 9253, 28, 29, 55, 2747, 2893, 4498 and 5848.

[0205] Bacteria of the genus Enterococcus are infected with the following phages: DF78, Fl, F2, 1, 2, 4, 14, 41, 867, Dl, SB24, 2BV, 182, 225, C2, C2F, E3, E62, DS96, H24, M35, P3, P9, SBlOl, S2, 2BII, 5, 182a, 705, 873, 881, 940, 1051, 1057, 21096C, NN-Enterococcus (1), PEl, Fl, F3, F4, VD13, 1, 200, 235 and 341.

[0206] Bacteria of the genus Erysipelothrix can be infected by the following phages: NN-Erysipelothrix (1).

[0207] Advertisement Continue reading the main story BW73、B278、D6、D10 8、E、E、E24、E41、FI-2、FI-4、FI-5、HI8A、F fl8B、i、MM、Mu、(syn=mu)、(syn=MuI)、(syn=Mu-I)、(syn=MU-I)、(syn=MuI)、(syn=μ)、0 25、PhI-5, Pk, PSP3, Pl, PlD, P2, P4 (defective), Sl, Wφ, K13, ϕR73, φ1, ϕ72, 7A, 8φ, 9φ, 15, 18, 28-1, 186,2 99、HH-エエェオキア(2)、AB48、CM、C4、C16、DD-VI、 (syn=Dd-Vi)、(syn=DDVI)、(syn=DDVi)、E4、E7、E28、FIl、FI3、H、Hl、H3、H8、K3、M、N、ND -2、ND-3、ND4、ND-5、ND6、ND-7、Ox-I(syn=OXl)、(syn=HF)、Ox-2(syn=0x2)、(syn=0X2)、 Ox-3、Ox-4、Ox-5、(syn=0X5)、Ox-6、(syn=66F)、(syn=ϕ66t)、(syn=φ66t-)50111、PhI-I 、RB42、RB43、RB49、RB69、S、SaI-I、Sal-2、Sal-3、Sal-4、Sal-5、Sal-6、TC23、TC45、TuII *-6, (syn=TuII*), TuIP-24, TuII*46, TuIP-60, T2, (syn=ganuTia), (syn=γ), (syn=PC), (syn=P.C.), (syn=T-2), (syn=T2), (syn=P4), T4, (syn=T-4), (syn=T4), T6, T35, αl, 1, IA, 3, (syn=Ac3), 3A, 3T+, (syn=3), (syn=Ml), 5φ, (syn=φ5), 9266Q, CFO103, HK620, J, K, KlF, m59, no.A, no.E, no.3, no.9, N4, sd, (syn=Sd), (syn=SD), (syn=Sa)3(syn=sd), (syn=SD), (syn=CD), T3, (syn=T-3), (syn=T3), T7, (syn=T-7), (syn=T7), WPK, W31, ΔH, φC3888, φK3, φK7, φK12, φV-1, Φ04-CF, Φ05, Φ06, Φ07, φ1, φ1.2, φ20, φ95, φ263, φ1092, φ1, φ11, (syn=φW), Ω8, 1, 3, 7, 8, 26, 27, 28-2, 29, 30, 31, 32, 38, 39, 42, 933W, NN-Escherichia (1), Esc-7-11, AC30, CVX-5, Cl, DDUP, ECl, EC2, E21, E29, Fl, F26S, F27S, Hi, HK022, HK97, (syn=ΦHK97), HK139, HK253, HK256, K7, ND-I, no.D, PA-2, q, S2, Tl, (syn=α), (syn=P28), (syn=T-I), (syn=Tx), T3C, T5, (syn=T-5), (syn=T5), UC-I, w, β4, γ2, λ(syn=lambda), (syn=Φλ), ΦD326, φγ, Φ06, Φ7, Φ10, φ80, χ, (syn=χi), (syn=φχ), (syn=φχi), 2, 4, 4A, 6, 8A, 102, 150, 168, 174, 3000, AC6, AC7, AC28, AC43, AC50, AC57, AC81, AC95, HK243, KlO, ZG / 3A, 5, 5A, 21EL, H19-J and 933H may be infected.

[0208] Bacteria of the genus Fusobacterium are infected by the following phages: NN-Fusobacterium (2), fv83-554 / 3, fv88-531 / 2, 227, fv2377, fv2527 and fv8501, or by vectors packaged into the capsids of the above phages.

[0209] Bacteria of the genus Haemophilus are infected by the following phages: HP1, S2 and N3, or by vectors packaged in the capsids of the above phages.

[0210] Bacteria of the genus Helicobacter are infected by the following phages: HP1 and ^^-Helicobacter (1) or by vectors packaged in the capsids of the above phages.

[0211] The following phages were identified in bacteria of the genus Klebsiella: AIO-2, KI4B, Kl6B, Kl9, (syn=K19), Kl14, Kl15, Kl21, Kl28, Kl29, KI32, Kl33, Kl35, Kl106B, Kl171B, Kl181B, Kl832B, AIO-I, AO-I, AO-2, AO-3, FC3-10, K, Kl1, (syn=KIl), Kl2, (syn=K12), Kl3, (syn=K13), (syn= Kl70 / 11), Kl4, (syn=K14), Kl5, (syn=K15), Kl6, (syn=K16), Kl7, (syn=K17), Kl8, (syn=K18), Kl19, (sy n=K19), Kl27, (syn=K127), Kl31, (syn=K131), Kl35, Kl171B, II, VI, IX, CI-I, Kl4B, Kl8, Kl11, Kl12, Kl13 , Kl16, Kl17, Kl18, Kl20, Kl22, Kl23, Kl24, Kl26, Kl30, Kl34, Kl106B, KIi65B, Kl328B, KLXI, K328, P5046 , 11, 380, III, IV, VII, VIII, FC3-11, Kl2B, (syn=K12B), Kl25, (syn=K125), Kl42B, (syn=K142), (syn=K14 2B), Kl181B, (syn=KIl81), (syn=K1181B), Kl765 / !, (syn=K1765 / 1), Kl842B, (syn=K1832B), Kl937B, (syn=K1937B), Ll, φ28, 7, 231, 483, 490, 632 and 864 / 100, or by vectors packaged in the capsids of the above phages.

[0212] Bacteria of the genus Leptospira are infected by the following phages: LE1, LE3, LE4 and .about.NN-leptospira (1) or by vectors packaged in the capsids of the above phages.

[0213] Listeria bacteria are identified by the following phages: A511, 01761, 4211, 4286, (syn=BO54), A005, A006, A020, A500, A502, A511, Al 18, A620, A640, B012, B021, B024, B025, B035, B051, B053, B054, B055, B056, BlOl, BI lO, B545, B604, B653, C707, D441, HSO47, HlOG, H8 / 73, H19, H21, H43, H46, H107, H108, HI lO, H163 / 84, H312, H340, H387, H391 / 73, H684 / 74, H924A, PSA, U153, φMLUP5 , (syn=P35), 00241, 00611, 02971A, 02971C, 5 / 476, 5 / 911, 5 / 939, 5 / 11302, 5 / 11605, 5 / 11704, 184, 575, 633, 699 / 694, 744, 900, 1090, 1317, 1444, 1652, 1806, 1807, 1921 / 959, 1921 / 11367, 1921 / 11500, 1921 / 11566, 1921 / 12460, Listeria can be infected with Listeria 1921 / 12582, 1967, 2389, 2425, 2671, 2685, 3274, 3550, 3551, 3552, 4276, 4277, 4292, 4477, 5337, 5348 / 11363, 5348 / 11646, 5348 / 12430, 5348 / 12434, 10072, 11355C, 11711A, 12029, 12981, 13441, 90666, 90816, 93253, 907515, 910716, and NN-Listeria (15), or by vectors packaged in the capsids of the above-mentioned phages.

[0214] Bacteria of the genus Morganella are infected by the following phages: 47, or vectors packaged in the capsids of the above phages.

[0215] Mycobacterium spp. bacteria are identified by the following phages: 13, AGl, ALi, ATCC11759, A2, B.C3, BG2, BKl, BK5, butyricum, BI, B5, B7, B30, B35, Clark, Cl, C2, DNAIII, DSP1, D4, D29, GS4E, (syn=GS4E), GS7, (syn=GS-7), (syn=GS7), IPa, Lacticola, Legendre, Leo, L5, (syn=ΦL-5), MC-I, MC-3, MC-4, Minetzi, MTPHIl, Mx4, MyF3P / 59a, Frey, (syn=Frey1). , Frey 4, Polonus II, Rabinovich, Smegmatis, TM4, TM9, TMlO, TM20, Y7, YlO, φ630, IB, IF, IH, 1 / 1, 67, 106, 1430, Bl, (syn=Bol), B24, D, D29, FK, FS, HP, Polonus I, Roy, Rl, (syn=Rl-Myb), (syn=Ri), 11, 31, 40, 50, 103a, 103b, 128, 3111-D, 3215-D and NN-Mycobacterium (1) or by vectors packaged in the capsids of the above phages.

[0216] Bacteria of the genus Neisseria are infected by the following phages: group I, group II and NP1, or by vectors packaged in the capsids of the above phages.

[0217] Bacteria of the genus Nocardia are infected by the following phages: MNP8, NJ-L, NS-8, N5 and TtiN-Nocardia or by vectors packaged in the capsids of the above phages.

[0218] Proteus bacteria are the target of the following phages: Pm5, 13vir, 2 / 44, 4 / 545, 6 / 1004, 13 / 807, 20 / 826, 57, 67b, 78, 107 / 69, 121, 9 / 0, 22 / 608, 30 / 680, PmI, Pm3, Pm4, Pm6, Pm7, Pm9, PmIO, PmIl, Pv2, πl, φm, 7 / 549, 9B / 2, 10A / 31, 12 / 55, 14, 15, 16 / 789, 17 / 971, 19A / 653, 23 / 532, 25 / 909, 26 / 219, 27 / 953, 32A / 909, 33 / 971, 34 / 13, 65, 5006M, 7480b, VI, 13 / 3a, Clichy12, π2600, φχ7, 1 / 1004, 5 / 742, 9, 12, 14, 22, 24 / 860, 2600 / D52, Pm8 and 24 / 2514, or by vectors packaged in the capsids of the above phages.

[0219] Bacteria of the genus Providencia are infected by the following phages: PL25, PL26, PL37, 9211 / 9295, 9213 / 921 Ib, 9248, 7 / R49, 7476 / 322, 7478 / 325, 7479, 7480, 9000 / 9402 and 9213 / 921 1a, or vectors packaged into the capsids of the above phages.

[0220] PfI (syn=Pf-I) and Pf2. Pf3、PP7、PRRl、7s、im-シュードモナス(1)、AI-I、AI-2、B 17、B89、CB3、Col 2、Col 11、Col 18、Col 21、C154、C163、C167、C2121、E79、F8、ga、gb、H22、K1、M4、N2、Nu、PB-I、(syn=PBl)、pfl6、 PMN17, PPl, PP8, Psal, PsPl, PsP2, PsP3, PsP4, PsP5, PS3, PS17, PTB80, PX4, PX7, PYOl, PY O2, PYO5, PYO6, PYO9, PYOlO, PYO13, PYO14, PYO16, PYO18, PYO19, PYO20, PYO29, PYO32, PY O33, PYO35, PYO36, PYO37, PYO38, PYO39, PYO41, PYO42, PYO45, PYO47, PYO48, PYO64, PYO6 9、PYO103、PlK、SLPl、SL2、S2、UNL-I、wy、Yai、Ya4、Yan、φBE、φCTX、φC17、ϕKZ、(syn=ΦKZ) φ-LT、Φmu78、φNZ、φPLS-1、φST-1、φW-14、φ-2、1 / 72、2 / 79、3、3 / DO、4 / 237、5 / 406、C6 / 6 / 6 660、7、7v、7 / 184、8 / 280、9 / 95、10 / 502、11 / DE、12 / 100、12S、16、21、24、25F、27、2134、68 71、95、109、188、337、352、1214、ΗN-シュードモナ、(23)、A856、B26、CI-I、CI-2、C5、D、gh-1、Fl 16、HF、H90、K5、K6、Cl 04、K109、K166、K267、N4、N5、O6N-25P、PE69、Pf、PPN25、PPN3 5、PPN89、PPN91、PP2、PP3、PP4、PP6、PP7、PP8、PP56、PP87、PPl14. PP206, PP207, PP306, PP651, Psp231a, Pssy401, Pssy 9220 psi PTB2 PTB20 PTB42 PXl PX3 PXlO PX12 PX14 PYO70, PYO71, R, SH6, SH133, tf, Ya5, Ya7, φBS, ΦKf77, φ- MC, ΦmnF82, φPLS27, φPLS743, φS-1, 1, 2, 2, 3, 4, 5, 6, 7, 7 8, 9, 10, 11, 12, 12B, 13, 14, 15, 14, 15, 16, 17, 18, 19, 20 20, 21, 21, 22, 23, 23, 24, 25, 31, 53, 73, 119x, 145, 0, 267, 284, 308, 525, NN-Ch BI-I, C22, D3, D37, D40, D62, D3112, F7, FlO, gd, ge, gξ Hwl2, Jb 19. KFl, L°, OXN-32P, O6N-52P, PCH-I, PC13-1, PC35-1, PH2, PH51 PH93, PH132, PMW, PM13, PM57, PM61, PM62, PM63, PM69, PM105, PMl 13 PM681, PM682, PO4, PPl, PP4, PP5, PP64, PP65, PP66, PP71, PP86, PP88, PP92, PP401, PP711, PP891 Pssy41, Pssy42, Pssy403, Pssy404, Pssy420, Pssy923, PS4, PS-IO, Pz, SDl, SLl, SL3, SL5, SM, φC5, φCl l, φCl l-1, φC13, φC15, φMO, φX, φO4, φll, φ240, 2, 2F, 5, 7m, 11, 13, 13 / 441, 14, 20, 24, 40, 45, 49, 61, 73, 148, 160, 198, 218, 222, 236, 242, 246, 249, 258, 269, 295, 297, 309, 318, 342, 350, 351, 357-1, 400-1, ΗN-Pseudomonas(6), GlOl, M6, M6a , Ll, PB2, Pssyl5, Pssy4210, Pssy4220, PYO12, PYO34, PYO49, PYO50, PYO51, PYO52, PYO53, PYO57, PYO59, PYO200, PX2, PX5, SL4, φO3, φO6 and 1214, or by vectors packaged in the capsids of the above-mentioned phages.

[0221] Bacteria of the genus Rickettsia are infected by the following phages: NN-Rickettsiae or vectors packaged in the capsid of the above phages.

[0222] References: b, Beccles, CT, d, Dundee, f, FeIs2, GI. GUI、GVI、GVIII、k、K、i、j、L、01、(syn=0-1)、(syn=O1)、(syn=OI )、(syn=7)、02、03、P3、P9a、PlO、Sab3、Sab5、SanlS、Sanl7、SI、 Taunton、ViI、(syn=ViI)、9、im、ルモネラ(1)、NI、N-5、N-IO、N-17、N -22、11、12、16-19、20.2、36、449C / C178、966A / C259、a、BAOR、e 、G4、GUI、L、LP7、M、MG40、N-18、PSA68、P4、P9c、P22、(syn=P22)、 (syn=PLT22)、(syn=PLT22)、P22al、P22-4、P22-7、P22-11、SNT- I、SNT-2、SP6、Villi、ViIV、ViV、ViVI、ViVII、Worksop、Sj5、ε34 、1、37、1(40)、(syn=φ1

[40] )、1、422、2、2.5、3b、4、5、6、14(18) 8、14(6,7), 10、27、28B、30、31、32、33、34、36、37、39、1412、SNT -3、7-11、40.3、c、C236、C557、C625、C966N、g、GV、G5、Gl73、h、I RA、Jersey、MB78、P22-1、P22-3、P22-12、Sat、Sat2、Sat2、Sat4 、Sanl、San2、San3、San4、San6、San7、San8、San9、Sanl3、Sanl4 、Sanl6、Sunl8、Sunl9、Sun20、Sun21、Sun22、Sun23、Sun24、Sun2 5、San26、SasLl、SasL2、SasL3、SasL4、SasL5、SlBL、SII、ViII、φ 1、1、2、3a、3al、1010、Ym-、ルモネラ(1)、N-4、SasL6

[0223] Bacteria from the genus Serrulacea are identified by the following phages: A2P, PS20, SMB3, SMP, SMP5, SM2, V40, V56, ic, ΦCP-3, ΦCP-6, 3M, 10 / la, 20A, 34CC, 34H, 38T, 345G, 345P, 501B, SMB2, SMP2, BC, BT, CW2, CW3, CW4, CW5, Lt232, L2232, L34, L.228, SLP, SMPA, V.43, σ, ΦCWl, ΦCP6-1, ΦCP6-2, ΦCP6-5, 3T, 5 / la, 3 / CWl, ΦCP6-2, ΦCP6-5, ΦCP6-6, ΦCP6-7, ΦCP6-8, ΦCP6-9, ΦCP6-10, ΦCP6-11, ΦCP6-12, ΦCP6-13, ΦCP6-14, ΦCP6-15, ΦCP6-16, ΦCP6-17, ΦCP6-18, ΦCP6-19, ΦCP6-20, ΦCP6-21, ΦCP6-22, ΦCP6-23, ΦCP6-24, ΦCP6-25, ΦCP6-26, ΦCP6-27, ΦCP6-28, ΦCP6-29, ΦCP6-30, ΦCP6-31, ΦCP6-32, ΦCP6-33, ΦCP6-34, ΦCP6-35, ΦCP6-36, ΦCP6-37, ΦCP6-38, ΦCP6-39, ΦCP , 8, 9F, 10 / 1, 2OE, 32 / 6, 34B, 34CT, 34P, 37, 41, 56, 56D, 56P, 6OP, 61 / 6, 74 / 6, 76 / 4, 101 / 8900, 226, 227, 228, 229F, 286, 289, 290F, 512, 764a, 2847 / 10, 2847 / 1Oa, L.359 and SMBl, or by vectors packaged in the capsids of the above phages.

[0224] Bacteria of the genus Shigella are the following phages: Fsa, (syn=a), FSD2d, (syn=D2d), (syn=W2d), FSD2E, (syn=W2e), fv, F6, f7.8, H-Sh, PE5, P90, SfII, Sh, SHm, SHrv, (syn=HIV), SHvi, (syn=HVI), SHVvm, (syn=HVIII), SKγ66, (syn=gamma66), (syn=yββ), (syn=γ66b), SKm, (syn=SIIIb)5(syn=UI), SKw, (syn=Siva), (syn=IV), SIC(trademark), (syn=SIVA.), (syn=IVA), SKvi, (syn=KVI), (syn=Svi), (syn=VI), SKvm, (syn=Svm), (syn=VIII), SKVΠIA, (syn=SvmA), (syn=VIIIA), STvi, STK, STx1, STxn, S66, W2, (syn=D2c), (syn=D20), φl, φIVb3-SO-R, 8368-SO-R, F7, (syn=FS7), (syn=K29), FlO, (syn=FSlO), (syn=K31), I1, (syn=alpha), (syn=FSa), (syn=Kl8), (syn=α), I2, (syn=a), (syn=K19), SG33, (syn=G35), (syn=SO-35 / G), SG35, (syn=SO-55 / G), SG3201, (syn=SO-3201 / G), SHn, (syn=HII), SHv, (syn=SHV), SHx, SHX, SKn, (syn=K2), (syn=KII), (syn=Sn), (syn=SsII), (syn=II), SKrv, (syn=Sm), (syn=SsIV), (syn=IV), SK1Va, (syn=Swab), (syn=SsIVa), (syn=IVa), SKV, (syn=K4), (syn=KV), (syn=SV), (syn=SsV), (syn=V), SKx, (syn=K9), (syn=KX), (syn=SX), (syn=SsX), (syn=X), STV, (syn=T35), (syn=35-50-R), STvm, (syn=T8345), (syn=8345-SO-S-R), W1, (syn=D8), (syn=FSD8), W2a, (syn=D2A), (syn=FS2a), DD-2, Sf6, FSi,(syn=Fl), SF6, (syn=F6), SG42, (syn=SO-42 / G), SG3203, (syn=SO-3203 / G), SKF12, (syn=SsF12), (syn=F12), (syn=F1 2), STn, (syn=1881-SO-R), γ66, (syn=gamma66a), (syn=Ssγ66), φ2, BIl, DDVII, (syn=DD7), FSD2b, (syn=W2B), FS2, (syn =F2), (syn=F2), FS4, (syn=F4), (syn=F4), FS5, (syn=F5), (syn=F5), FS9, (syn=F9), (syn=F9), FIl, P2-S0-S, SG36, (s yn=SO-36 / G), (syn=G36), SG3204, (syn=SO-3204 / G), SG3244, (syn=SO-3244 / G), SHi, (syn=HI), SHvπ, (syn=HVII), SHK , (syn=HIX), SHx1, SHxπ, (syn=HXn), SKI, KI, (syn=S1), (syn=SsI), SKVII, (syn=KVII), (syn=Svπ), (syn=SsVII), SKI X, (syn=KIX), (syn=S1x), (syn=SsIX), SKXII, (syn=KXII), (syn=Sxn), (syn=SsXII), STi, STffl, STrv, STVi, STvπ, S7 0, S206, U2-S0-S, 3210-SO-S, 3859-SO-S, 4020-SO-S, φ3, φ5, φ7, φ8, φ9, φlO, φll, φl3, φl4, φl8, SHm, (syn=Hπi), SHχi, (syn=HXt), and SKxI, (syn=KXI), (syn=Sχi), (syn=SsXI), (syn=XI), or by vectors packaged in the capsids of the above phages.

[0225] Bacteria of the genus Staphylococcus are susceptible to the following phages: A, EW, K, Ph5, Ph9, PhIO, Phl3, Pl, P2, P3, P4, P8, P9, PlO, RG, SB-i, (syn=Sb-I), S3K, Twort, ΦSK311, φ812, 06, 40, 58, 119, 130, 131, 200, 1623, STCl, (syn=stcl), STC2, (syn=stc2), 44AHJD, 68, ACl, AC2, A6"C", A9"C", b581, CA-I, CA-2, CA-3, CA-4, CA-5, DI l, L39x35, L54a, M42, Nl, N2, N3, N4, N5, N7, N8, NlO, Nil, N12, N13, N14, N16, Ph6, Phl2, Phl4, UC-18, U4, U15, Sl, S2, S3, S4, S5, X2, Z1, φB5-2, φD, ω, 11, (syn=φl l), (syn=P11-M15), 15, 28, 28A, 29, 31, 31B, 37, 42D, (syn=P42D), 44A, 48, 51, 52, 52A, (syn=P52A), 52B, 53, 55, 69, 71, (syn=P71), 71A, 72, 75, 76, 77, 79, 80, 80α, 82, 82A, 83A, 84, 85, 86, 88, 88A, 89, 90, 92, 95, 96, 102, 107, 108, 111, 129-26, 130, 130A, 155, 157, 157A, 165, 187, 275, 275A, 275B, 356, 456, 459, 471, 471A, 489, 581, 676, 898, 1139, 1154A, 1259, 1314, 1380, 1405, 1563, 2148, 2638A, 2638B, 2638C, 2731, 2792A, 2792B, 2818, 2835, 2848A, 3619, 5841, 12100, AC3, A8, AlO, A13, b594n, D, HK2, N9, N15, P52, P87, Sl, S6, Z4, φRE, 3A, 3B, 3C, 6, 7, 16, 21, 42B, 42C, 42E, 44, 47, 47A5Staphylococcus sp. (1), or by vectors packaged in the capsids of the above-mentioned phages.

[0226] Bacteria of the genus Streptococcus are identified by the following phages: EJ-I, NN-Streptococcus (1), a, Cl, FL0Ths, H39, Cp-I, Cp-5, Cp-7, Cp-9, Cp-IO, AT298, A5, alO / Jl, alO / J2, alO / J5, alO / J9, A25, BTI l, b6, CAl, c20-l, c20-2, DP-I, Dp-4, DTl, ET42, elO, FA101, FEThs, Fκ, FKKIOI, FKLIO, FKP74, FKH, FLOThs, FyIOl, fl, F10, F20140 / 76, g, GT-234, HB3, (syn=HB-3), HB-623, HB-746, M102, O 1205, φO1205, PST, PO, Pl, P2, P3, P5, P6, P8, P9, P9, P12, P13, P14, P49, P50, P51, P52, P53, P54 , P55, P56, P57, P58, P59, P64, P67, P69, P71, P73, P75, P76, P77, P82, P83, P88, sc, sch, sf, SfIl 1, (syn=SFiI l), (syn=φSFill), (syn=ΦSfil l), (syn=φSfill), sfil9, (syn=SFil9), (syn=φSFil9), (syn=φSfil9), Sfi21, (syn=SFi21), (syn=φSFi21), (syn=φSfi21), ST0, STX, st2, ST2, ST4, S3, (syn=φS3), s265, φ17, φ42, φ57, φ80, φ81, φ82, φ83, φ84, φ85, φ86, φ87, φ88, φ89, φ90, φ91, φ92, φ93, φ94 , φ95, φ96, φ97, φ98, φ99, φlOO, φlOl, φlO2, φ227, φ7201, ωl, ω2, ω3, ω4, ω5, ω6, ω8 , ωlO, 1, 6, 9, 1OF, 12 / 12, 14, 17SR, 19S, 24, 50 / 33, 50 / 34, 55 / 14, 55 / 15, 70 / 35, 70 / 36, 71 / ST15, 71 / 45, 71 / 46, 74F, 79 / 37, 79 / 38, 80 / J4, 80 / J9, 80 / ST16, 80 / 15, 80 / 47, 80 / 48, 101, 103 / 39, 103 / 40, 121 / 41, 121 / 42, 123 / 43, 123 / 44, 124 / 44, 337 / ST17, and mStreptococcus (34) or by vectors packaged in the capsids of the above phages.

[0227] Bacteria of the genus Treponema are infected by the following phages: NN-Treponema (1) or by vectors packaged in the capsids of the above phages.

[0228] Bacteria of the genus Vibrio are the following phages: CTXΦ, fs, (syn=si), fs2, Ivpf5, Vfl2, Vf33, VPIΦ, VSK, v6, 493, CP-Tl, ET25, kappa, K139, Labol, )XN-69P, OXN-86, O6N-21P, PB-I, P147, rp-1, SE3, VA-I, (syn=VcA-I), VcA-2, VPl, VP2, VP4, VP7, VP8, VP9, VPlO, VP17, VP18, VP19, X29, (syn=29d'Herelle), t, ΦHAWI-1, ΦHAWI-2, ΦHAWI-3, ΦHAWI-4, ΦHAWI-5, ΦHAWI-6, ΦHAWI-7, XHAWI-8, ΦHAWI-9, ΦHAWI-10, ΦHCl-1, ΦHC1-2, ΦHC1-3, ΦHC1-4, ΦHC2-1, >HC2-2, ΦHC2-3, ΦHC2-4, ΦHC3-1, ΦHC3-2, ΦHC3-3, ΦHD1S-1, ΦHD1S-2, ΦHD2S-1, ΦHD2S-2, ΦHD2S-3, ΦHD2S-4, ΦHD2S-5, ΦHDO-1, ΦHDO-2, ΦHDO-3, ΦHDO-4, ΦHDO-5, ΦHDO-6, ΦKL-33, ΦKL-34, ΦKL-35, ΦKL-36, ΦKWH-2, ΦKWH-3, ΦKWH-4, ΦMARQ-1, ΦMARQ-2, ΦMARQ-3, ΦMOAT-1, ΦO139, ΦPEL1A-1, ΦPEL1A-2, ΦPEL8A-1, ΦPEL8A-2, ΦPEL8A-3, ΦPEL8C-1, ΦPEL8C-2, ΦPEL13A-1, ΦPEL13B-1, ΦPEL13B-2, ΦPEL13B-3, ΦPEL13B-4, ΦPEL13B-5, ΦPEL13B-6, ΦPEL13B-7, ΦPEL13B-8, ΦPEL13B-9, ΦPEL13B-10, φVP143, φVP253, Φ16, φl38, 1-II, 5, 13, 14, 16, 24, 32, 493, 6214, 7050, 7227, II, (syn=Group II), (syn==φ2), V, VIII, ~m-Vibrio (13), KVP20, KVP40, nt-1, O6N-22P, P68, el, e2, e3, e4, e5, FK, G, I, K, nt-6, Nl, N2, N3, N4, N5, O6N-34P, OXN-72P, OXN-85P, OXN-100P, P, Ph-I, PL163 / 10, Q, S, T, φ92, 1-9, 37, 51,57, 70A-8, 72A-4, 72A-10, 110A-4, 333, 4996, I (syn = group I), III (syn = group III), VI, (syn = A-Saratov), ​​VII, IX, X, HN-Vibrio (6), pAl, 7, 7-8, 70A-2, 71A-6, 72A-5, 72A-8, 108A-10, 109A-6, 109A-8, 110A-1, 110A-5, 110A-7, hv-1, OXN-52P, P13, P38, P53, P65, P 108, Pill, TP13VP3, VP6, VP12, VP13, 70A-3, 70A-4, 70A-10, 72A-1, 108A-3, 109-B1, 110A-2, 149, (syn=φ149), IV, (syn=group IV), NN-Vibrio (22), VP5, VP11, VP15, VP16, α1, α2, α3a, α3b, 353B and HN-Vibrio (7), or by vectors packaged in the capsids of the above phages.

[0229] Bacteria of the genus Yersinia are infected by the following phages: H, HI, H-2, H-3, H-4, Lucas110, Lucas303, Lucas404, YerA3, YerA7, YerA20, YerA41, 3 / M64-76, 5 / G394-76, 6 / C753-76, 8 / C239-76, 9 / F18167, 1701, 1710, PST, 1 / F2852-76, D'Herelle, EV, H, Kotljarova, PTB, R, Y, YerA41, φYerO3-12, 3, 4 / C1324-76, 7 / F783-76, 903, 1 / M6176 and Yer2AT, or by vectors packaged in the capsids of the above phages.

[0230] vector Vectors are well known to those skilled in the art. The vector according to the present invention can be a DNA vector or an RNA vector. The vector can include a site suitable for inserting a gene or sequence of interest, such as a replication origin, a selection marker, and a multicloning site. "Sequence of interest" refers to a sequence that produces a therapeutic effect.

[0231] Preferably, the vector is an expression vector, preferably a vector allowing expression in bacteria.

[0232] The expression vector according to the present invention may comprise a promoter, a translation initiation sequence such as a ribosome binding site and a start codon, a stop codon, and a transcription termination sequence. The expression vector according to the present invention may also comprise regulatory regions such as enhancers, silencers, and boundary elements / insulators to direct the transcription level of a given gene.

[0233] The expression vector can be a vector for stable or transient expression of a gene.

[0234] The vector according to the invention may be selected from the group consisting of a plasmid, a bacteriophage genome, a phagemid, a viral genome, a cosmid, and an artificial chromosome. Preferably, the vector according to the invention is a bacteriophage genome or a phagemid.

[0235] In a preferred embodiment, the vector according to the invention is a bacteriophage genome. The bacteriophage genome according to the invention contains all the genes required for its replication in the bacterium, for the formation of new virions and for their release. The genome can be wild-type or genetically engineered.

[0236] The bacteriophage genome according to the invention is the genome of a bacteriophage as defined above, such as IKe, CTX-φ, Pf1, Pf2, Pf3, Myoviridae (e.g. P1-like, P2-like, Mu-like, SPO1-like and phiH-like bacteriophages); Siphoviridae (e.g. λ-like, γ-like, T1-like, c2-like, L5-like, psiM1-like, phiC31-like and N15-like bacteriophages); Podoviridae (e.g. phi29-like, P22-like and N4-like bacteriophages); Tectiviridae (e.g. Tectivirus); Corticoviridae (e.g. Corticovirus); Liposthrixviridae (e.g. Alphaliposthrixvirus, Betaliposthrixvirus, Gamma ... posthrixvirus, and Deltaliposthrixvirus); Plasmaviridae (e.g., Plasmavirus); Rudiviridae (e.g., Rudiviridae); Fuselloviridae (e.g., Fusellovirus); Inoviridae (e.g., Inovirus, Pretovirus, M13-like and fd-like bacteriophages); Microviridae (e.g., Microvirus, Spiromicrovirus, Bdelomicrovirus, and Chlamydiamicrovirus); Leviviridae (e.g., Levivirus, and Allolevivirus); Cystoviridae (e.g., Cystovirus), E. coli phages (e.g., infecting E. coli), B1 (e.g., Bacteroides thetaiotamicron), thetaiotamicron), ATCC 51477-B1, B40-8, or Bf-1 (e.g., B. fragilis infected), phiHSC01 (e.g., B. cacae infected), phiHSC02 (e.g., B.ovatus), phiC2, phiC5, phiC6, phiC8, phiCD119, or phiCD27 (e.g., Clostridium difficile), KP01K2, K11, Kpn5, KP34, or JD001 (e.g., Klebsiella pneumoniae), phiNM1 or 80alpha (e.g., Staphylococcus aureus), IME-EF1 (e.g., Enterococcus faecalis), The genome may be selected from the group consisting of phiKMV, PAK-P1, LKD16, LKA1, delta, sigma-1, J1 (e.g., infecting Pseudomonas aerignosa), T2, T4, T5, T7, RB49, phiX174, R17, PRD1 bacteriophages, and any bacteriophage derived therefrom.

[0237] A bacteriophage genome according to the invention may be the genome of a bacteriophage having a lytic or non-lytic replication cycle.

[0238] Genomes of bacteriophages having a non-lytic replication cycle according to the present invention include IKe, CTX-φ, Pf1, Pf2, Pf3, Myoviridae (e.g., P1-like, P2-like, Mu-like, SPO1-like, and φH-like bacteriophages); Siphoviridae (e.g., λ-like, γ-like, T1-like, c2-like, L5-like, psiM1-like, phiC31-like, and N15-like bacteriophages); Podoviridae (e.g., phi29-like, P22-like, and N4-like bacteriophages); Tectiviridae (e.g., Tectivirus); Corticoviridae (e.g., Corticovirus); Liposthrixviridae (e.g., Alphalipothrixvirus, Betalipothrixvirus, Gammalipothrixvirus, and the like); viruses, and Deltaliposthrixvirus); Plasmaviridae (e.g., Plasmavirus); Rudiviridae (e.g., Rudiviridae); Fuselloviridae (e.g., Fusellovirus); Inoviridae (e.g., Inovirus, Pretovirus, M13-like and fd-like bacteriophages); Microviridae (e.g., Microvirus, Spiromicrovirus, Bdelomicrovirus, and Chlamydiamicrovirus); Leviviridae (e.g., Levivirus, and Allolevivirus); Cystoviridae (e.g., Cystovirus), E. coli phage (e.g., infects E. coli), B1 (e.g., infects Bacteroides thetaeotamicron), ATCC 51477-B1, B40-8, or Bf-1 (e.g., infects B. fragilis), phiHSC01 (e.g., infects B. cacae), phiHSC02 (e.g., B.ovatus), phiC2, phiC5, phiC6, phiC8, phiCD119, or phiCD27 (e.g., infecting Clostridium difficile), KP01K2, K11, Kpn5, KP34, or JD001 (e.g., infecting Klebsiella pneumoniae), phiNM1 or 80alpha (e.g., infecting Staphylococcus aureus), IME-EF1 (e.g., infecting Enterococcus faecalis), ENB6 or C33 (e.g., infecting Enterococcus faecium), and phiKMV, PAK-P1, LKD16, LKA1, delta, sigma-1, J1 (e.g., infecting Pseudomonas aeruginosa) bacteriophages, and any bacteriophage derived therefrom.

[0239] The genome of the bacteriophage with a non-lytic replication cycle (also called lysogenic cycle) according to the invention is derived from a temperate bacteriophage. Preferably, the genome of the bacteriophage with a non-lytic replication cycle according to the invention is selected from the group consisting of P2-like and lambda-like (lambda-like) bacteriophages. More preferably, the genome of the bacteriophage with a non-lytic replication cycle according to the invention is selected from lambda-like bacteriophages, preferably from the group consisting of HK022, mEP237, HK97, HK629, HK630, mEP043, mEP213, mEP234, mEP390, mEP460, mEPx1, mEPx2, phi80, mEP234 bacteriophages and any bacteriophage derived therefrom.

[0240] The genome of a bacteriophage having a lytic replication cycle according to the present invention may be selected from the group consisting of T2, T4, T5, T7, RB49, phiX174, R17, PRD1 bacteriophage, and any bacteriophage genome derived therefrom.

[0241] The genomes of other bacteriophages may be used in accordance with the present invention.

[0242] Preferably, the bacteriophage genome according to the invention is the genome of a lethal bacteriophage, ie one which only has a lytic reproductive cycle.

[0243] Alternatively, the bacteriophage genome according to the invention is the genome of a non-lethal bacteriophage, i.e. a temperate bacteriophage having both a lysogenic and a lytic reproductive cycle.

[0244] In another preferred embodiment, the vector according to the invention is a phagemid. The phagemid according to the invention comprises a phage packaging site and optionally a plasmid origin of replication (ori), in particular a bacterial origin of replication and / or a phage origin of replication. Preferably, the phagemid according to the invention does not comprise a plasmid origin of replication and therefore cannot replicate by itself when injected into a bacterium.

[0245] The phagemid according to the invention may be derived from any of the bacteriophages disclosed herein. In particular, the phagemid according to the invention is suitable to be packaged in any of the bacteriophages disclosed herein, and in particular comprises a packaging site and optionally a bacteriophage origin of replication, which, with the complementation of the complete phage genome, can initiate replication of the plasmid for subsequent encapsidation into a different capsid. The phage origin of replication can be a wild-type or non-wild-type sequence of M13, f1, φX174, P4, lambda, P2, lambda-like bacteriophage, HK022, mEP237, HK97, HK629, HK630, mEP043, mEP213, mEP234, mEP390, mEP460, mEPx1, mEPx2, phi80, mEP234, T2, T4, T5, T7, RB49, phiX174, R17, PRD1 P1-like, P2-like, P22-like N15-like bacteriophage.

[0246] Packaging sites include, but are not limited to, SPP1 (SPP1pac site), P1 (P1pac site), T1 (T1pac site), T7 (T7 concatemer junction), lambda (λcos site), P4 (P4coc site), mu (mupac site), P22 (P22pac site), φ8 (φ8pac site), Sf6 (Sf6pac site), 149 (149pac site), and A1122 (A1122 concatemer junction). For most bacterial viruses, the packaging site is referred to as a pac site. In some cases, the packaging site is referred to as a concatemer junction (e.g., T7 concatemer junction). In all cases, the packaging site is substantially separated from the sequences naturally occurring adjacent to it in the bacterial virus genome.

[0247] The phagemid according to the present invention may be selected from the group consisting of lambda-derived phagemids, P4-derived phagemids, M13-derived phagemids, such as those containing an f1 origin for filamentous phage packaging, e.g., pBluescript II SK(+ / -) and KS(+ / -) phagemids, pBC SK and KS phagemids, pADL and P1-derived phagemids (see, e.g., Westwater CA et al., Microbiology 148, 943-50 (2002); Kittleson JT et al., ACS Synthetoc Biology 1, 583-89 (2012); Mead DA et al., Biotechnology 10, 85-102 (1988)).

[0248] Preferably, the phagemid according to the invention is selected from a lambda-derived phagemid and a P4-derived phagemid.

[0249] More preferably, the phagemid according to the present invention is selected from lambda derived phagemids, preferably selected from the group consisting of HK022 derived phagemid, mEP237 derived phagemid, HK97 derived phagemid, HK629 derived phagemid, HK630 derived phagemid, mEP043 derived phagemid, mEP213 derived phagemid, mEP234 derived phagemid, mEP390 derived phagemid, mEP460 derived phagemid, mEPx1 derived phagemid, mEPx2 derived phagemid, phi80 derived phagemid, mEP234 derived phagemid.

[0250] Other phagemids can be used in accordance with the present invention.

[0251] For example, the vector according to the invention, preferably the bacteriophage genome or phagemid, may also contain a sequence of interest. A sequence of interest is a sequence that has a technical effect on the bacterium, preferably a therapeutic effect. A sequence of interest may be, for example, a sequence that can kill the bacterium, regulate the expression of a gene, in particular suppress or increase the expression of one or several genes, or regulate the production of a metabolite, in particular decrease or increase the expression of one or several metabolites.

[0252] For example, a vector according to the invention, preferably a bacteriophage genome or a phagemid, may also contain additional genes, in particular genes which are intended to be expressed in the bacterium of interest.

[0253] In one embodiment, the vector may include a sequence of interest, e.g., a sequence encoding a protein of interest. The protein of interest may be a bacteriocin, which is a proteinaceous toxin produced by bacteria that can kill or inhibit the growth of other bacteria. Bacteriocins are classified in several ways, including by strain production, general resistance mechanism, and mechanism of death. Such bacteriocins from Gram-negative bacteria (e.g., microcins, colicin-like bacteriocins, and teilosins) and Gram-positive bacteria (e.g., class I, class II, class III, or class IV bacteriocins) have been reported. The protein of interest may then be selected from the group consisting of toxins selected from the group consisting of microcins, colicin-like bacteriocins, teilosins, class I, class II, class III, and class IV bacteriocins.

[0254] In certain embodiments, a programmable nuclease circuit can be added to a vector to be delivered to a bacterium of interest. This programmable nuclease circuit may be able to mediate in vivo sequence-specific removal of bacteria containing a target gene of interest (e.g., a gene harmful to humans). Some embodiments of the present disclosure relate to engineered variants of the type II CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR associated) system of Streptococcus pyogenes, which can be used according to the present disclosure to reverse or specifically disrupt antibiotic resistance in a wide range of microorganisms. Other programmable nucleases that can be used according to the present disclosure include other CRISPR-Cas systems, engineered TALEN (transcription activator-like effector nuclease) variants, and engineered zinc finger nuclease (ZFN) variants. Thus, in some embodiments, the engineered autonomous decentralized circuits provided herein can be used to selectively cleave DNA encoding a gene of interest, such as, for example, a toxin gene, a virulence factor gene, an antibiotic resistance gene, a remodeling gene, or a regulatory gene (see WO 2014 / 124226 and U.S. Patent Application Publication No. 2015 / 0064138).

[0255] The CRISPR system includes two distinct elements: i) an endonuclease, in this case the CRISPR-associated nuclease (Cas or "CRISPR-associated protein"), and ii) a guide RNA. The guide RNA is in the form of a chimeric RNA consisting of a combination of CRISPR (RNAcr) bacterial RNA and RNAtracr (trans-activating RNA CRISPR) (Jinek et al., Science, 2012). The gRNA combines the target specificity of the cRNA, which corresponds to a "spacing sequence" that serves as a guide to the Cas protein, with the conformational properties of Rtracr in a single transcript. When the gRNA and Cas protein are expressed simultaneously in a cell, the target genomic sequence can be permanently modified or interrupted. The modification is advantageously guided by a repair matrix.

[0256] The sequence of interest according to the present invention comprises a nucleic acid sequence encoding a Cas protein. A variety of CRISPR enzymes are available for use as the sequence of interest on a plasmid according to the present invention. In some embodiments, the CRISPR enzyme is a type II CRISPR enzyme. In some embodiments, the CRISPR enzyme catalyzes DNA cleavage. In some other embodiments, the CRISPR enzyme catalyzes RNA cleavage. In one embodiment, the CRISPR enzyme can be coupled to an sgRNA. In certain embodiments, the sgRNA targets a gene selected from the group consisting of antibiotic resistance genes, virulence genes, toxin genes, and any other gene that confers a selective advantage to a pathogen or allows for the differentiation of pathogenic and non-pathogenic strains of the same bacterial species.

[0257] Non-limiting examples of Cas proteins include Casl, CaslB, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csnl and Csxl2), Cas10, Csy1, Csy2, Csy3, Csel, Cse2, Cscl, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmrl, Cmr3, Cmr4, Cmr5, Cmr6, Csbl, Csb2, Csb3, Csxl7, Csxl4, CsxlO, Csxl6, CsaX, Csx3, Csxl, Csxl5, Csfl, Csf2, Csf3, Csf4, homologs thereof, or mutants thereof. In some embodiments, the CRISPR enzyme cleaves both strands of the target nucleic acid at a protospacer adjacent motif (PAM) site.

[0258] In certain embodiments, the CRISPR enzyme is any Cas9 protein, such as any naturally occurring bacterial Cas9, as well as any mutants, homologs, or orthologs thereof.

[0259] "Cas9" refers to the protein Cas9 (also called Csn1 or Csx12) or a functional protein, peptide, or polypeptide fragment thereof, i.e. capable of exerting an enzymatic activity (nuclease) capable of interacting with a guide RNA and effecting a double-stranded break in the DNA of a target genome. Thus, "Cas9" can refer to a modified protein, e.g., a modified protein that has been truncated to remove domains of the protein that are not essential for a given function of the protein, in particular domains that are not required for interaction with the gRNA.

[0260] The sequence encoding Cas9 (whole protein or fragments thereof) used in the context of the present invention can be obtained from any known Cas9 protein (Makarova et al., 2011). Examples of Cas9 proteins useful in the present invention include, but are not limited to, the Cas9 proteins of Streptococcus pyogenes, Streptococcus thermophilus, Streptococcus mutans, Campylobacter jejuni, Francisella novicida, and Neisseria meningitidis. Other Cas9 proteins that can be used in the present invention are also described in the article by Fonfara et al., 2013.

[0261] In certain embodiments, the nucleic acid sequence of interest is a CRISPR / Cas9 system for reducing expression or inactivating a gene selected from the group consisting of antibiotic resistance genes, virulence genes, toxin genes, and any other gene that confers a selective advantage to a pathogen or allows for the differentiation of pathogenic and non-pathogenic strains of the same bacterial species.

[0262] In one embodiment, the CRISPR / Cas9 system is used to target and inactivate virulence factors. Virulence factors can be any substance produced by a pathogen that alters the host-pathogen interaction by increasing the degree of damage inflicted on the host. Virulence factors are used by pathogens in a number of ways, including, for example, cell adhesion or colonization of niches in the host, to evade the host's immune response, to facilitate invasion and exit from the host cell, to obtain nutrients from the host, or to inhibit other physiological processes in the host. Virulence factors can include enzymes, endotoxins, adhesion factors, motility factors, factors involved in complement evasion, and factors that promote biofilm formation. For example, such targeted virulence factor genes can be E. coli virulence gene factors such as eaeA, EHEC-HlyA, Stx1 (VT1), Stx2 (VT2), Stx2c (VT2c), Stx2d (VT2d), Stx2e (VT2e) and Stx2f (VT2f), fimH, neuC, kpsE, sfa, foc, iroN, aer, iha, papC, papGI, papGII, papGIII, hlyC, cnf1, hra, sat, ireA, usp ompT, ibeA, malX, fyuA, irp2, traT, afaD, ipaH, eltB, estA, bfpA, eae, aaiC, aatA, TEM, CTX, SHV.

[0263] In another embodiment, the CRISPR / Cas9 system is used to target and inactivate antibiotic resistance genes (e.g., the ampicillin resistance gene).

[0264] In another embodiment, the CRISPR / Cas9 system is used to target and inactivate bacterial toxin genes. Bacterial toxins can be classified as either exotoxins or endotoxins. Exotoxins are produced and actively secreted; endotoxins remain part of the bacterium. The reaction to bacterial toxins can involve severe inflammation and can lead to sepsis. Such toxins can be, for example, botulinum neurotoxins, tetanus toxins, staphylococcal toxins, diphtheria toxins, anthrax toxins, alpha toxins, pertussis toxins, shiga toxins, heat-stable enterotoxins (E. coli ST), or any of the toxins described in Henkel et al. (Toxins from Bacteria in EXS. 2010; 100: pp. 1-29).

[0265] Other genetic circuits, preferably other programmable genetic circuits, can be added to the vector and delivered to the bacterium of interest. Preferably, the genetic circuit added to the vector results in cell death of the bacterium of interest. For example, the genetic circuit added to the vector may code for a holin or a toxin. Alternatively, the genetic circuit added to the vector does not result in bacterial death. For example, the genetic circuit may code for a reporter gene that results in a luminescent or fluorescent signal. Alternatively, the genetic circuit may include proteins and enzymes that perform a useful function, such as modifying the metabolism of the bacterium or the composition of its environment.

[0266] The vector may further comprise a selection marker, in certain embodiments, which provides a selective advantage to bacterial cells infected with the plasmid, such as antibiotic resistance, heavy metal resistance, complementation of a host auxotrophy, and / or display of a fluorescent or luminescent protein.

[0267] The inclusion of an appropriate selection marker gene in the plasmid allows testing and / or detection for successful delivery of the plasmid according to the invention. The plasmid according to the invention may contain one or more nucleic acid sequences encoding a detectable label such as a selection marker, such as an auxotrophic marker (e.g., LEU2, URA3, TRP1 or HIS3, DapA, ThyA), a fluorescent or luminescent protein (e.g., GFP, eGFP, DsRed, CFP, YFP), or a protein that confers resistance to chemicals / toxic compounds (MGMT gene that confers resistance to temozolomide, kanamycin resistance, chloramphenicol resistance, etc.) or any combination thereof. These markers can be used to select or detect host cells that contain the vector according to the invention and can be easily selected by the skilled artisan according to the host cell.

[0268] For most purposes, antibiotic resistance genes are commonly used selection markers to facilitate molecular biology cloning of plasmids and to allow detection or selection of bacteria transformed by such plasmids. Antibiotic resistance genes are well known in the art and include, but are not limited to, ampicillin resistance (Amp), chloramphenicol resistance (Cm), tetracycline resistance (Tet), kanamycin resistance (Kan), hygromycin resistance (Qiyg or hph gene), and zeomycin resistance (Zeo).

[0269] delivery vehicle The vector according to the invention is preferably included in a delivery vehicle that allows the transfer of the vector into the bacterium of interest.

[0270] Delivery vehicles are well known to those skilled in the art. There are several common types of delivery vehicles, including but not limited to chemical-based delivery vehicles, non-chemical-based delivery vehicles, particle-based delivery vehicles, nanoparticle-based delivery vehicles, donor bacteria, bacteriophage scaffolds, and viral scaffolds.

[0271] The chemical-based delivery vehicle according to the present invention may be selected from the group consisting of cyclodextrins, calcium phosphates, cationic polymers, cationic liposomes, and mixtures thereof.

[0272] The non-chemical based delivery vehicle according to the present invention may be selected from the group consisting of electroporation, sonoporation, optical transfection, and mixtures thereof.

[0273] The particle-based delivery vehicle according to the present invention may be selected from the group consisting of gene guns, magnetofection, impalefection, particle bombardment, cell penetrating peptides and mixtures thereof.

[0274] The nanoparticle-based delivery vehicle can be, for example, a nucleic acid nanocage.

[0275] The donor bacterium can be a delivery vehicle for the conjugative plasmid. Donor bacteria according to the present invention include, but are not limited to, commensal bacteria and probiotic bacteria.

[0276] In a preferred embodiment, the delivery vehicle according to the present invention is a bacteriophage scaffold or capsid or a bacteriophage virus-like particle. Preferably, the vector contained in the bacteriophage scaffold is a bacteriophage genome or a phagemid.

[0277] Therefore, in a particularly preferred embodiment, the present invention relates to a bacteriophage or a packaged phagemid, the vector being as defined above.

[0278] The bacteriophage genome or phagemid can be engineered to function on a bacteriophage scaffold derived from a natural bacteriophage, an engineered bacteriophage, or an evolved bacteriophage. Preferably, the bacteriophage scaffold is derived from a natural bacteriophage.

[0279] Preferably, a bacteriophage according to the invention comprises a bacteriophage genome and a bacteriophage scaffold of the same origin, i.e. from the same species, preferably from the same strain. Even more preferably, the bacteriophage scaffold according to the invention is assembled from proteins encoded by genes of the bacteriophage genome.

[0280] Preferably, the packaged phagemids according to the invention comprise a bacteriophage scaffold that is of the same origin as the bacteriophage genes of the phagemid, i.e., the bacteriophage scaffold of the packaged phagemid is of the same species, preferably of the same strain, as the bacteriophage DNA from which the phagemid is derived. Preferably, the packaged phagemids according to the invention comprise a bacteriophage scaffold that is of the same origin as the bacteriophage packaging site of the phagemid.

[0281] A bacteriophage or packaged phagemid according to the invention may be lethal or non-lethal. Preferably, a bacteriophage or packaged phagemid according to the invention is a lethal bacteriophage or a lethal packaged phagemid. Alternatively, a bacteriophage or packaged phagemid according to the invention is a non-lethal bacteriophage or a non-lethal packaged phagemid.

[0282] Uses and methods of vectors according to the invention In another aspect, the present invention also relates to the use of a vector according to the present invention, preferably a bacteriophage genome or a phagemid according to the present invention, or a vector comprised in a delivery vehicle according to the present invention, preferably a bacteriophage or a packaged phagemid according to the present invention, for delivering the vector to a bacterium, preferably a bacterium of interest according to the present invention.

[0283] In certain embodiments, the vector according to the invention comprises a genetic circuit, preferably a programmable genetic circuit. The genetic circuit according to the invention can result in cell death of the bacterium, i.e. the bacterium of interest. For example, the genetic circuit added to the vector can code for a holin or a toxin. Alternatively, the genetic circuit added to the vector does not result in bacterial death. For example, the genetic circuit can code for a reporter gene that results in a luminescent or fluorescent signal. Alternatively, the genetic circuit can include proteins and enzymes that perform a useful function, such as modifying the metabolism of the bacterium or the composition of its environment.

[0284] Yet another aspect relates to the use of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, as a research tool, in particular for improving the frequency of delivery and / or for expanding the strains of bacteria to which the vector can be delivered.

[0285] The present invention also relates to a method for preparing a vector according to the invention, comprising the steps of: (i) selecting a bacterial group of interest: (ii) identifying restriction sites based on the vector sequence that are recognized by restriction enzymes encoded by the bacterial group; (iii) modifying the sequence of a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, so as to contain no more than 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 restriction sites recognised by restriction enzymes encoded by each bacterium in the group of bacteria; (iv) optionally preparing a modified vector, in particular by nucleic acid synthesis or by mutation of the vector; Includes.

[0286] Preferably, in step (ii) the frequency of the restriction enzyme in the bacterial group of interest is determined and in step (iii) the sequence of the vector is modified to remove at least one restriction site for a restriction enzyme that is frequently encoded by the bacterial group of interest, preferably to remove 1, 2, 3, 4, 5, 6 or 7 restriction sites for restriction enzymes that are frequently encoded by the bacterial group of interest, more preferably to remove restriction sites for restriction enzymes that are frequently encoded by the bacterial group of interest.

[0287] The present invention further relates to a method for preparing a vector according to the invention, comprising the steps of: (i) selecting a bacterial group of interest; (ii) identifying restriction enzymes encoded by the bacterial group of interest and determining the frequency of bacteria encoding the restriction enzymes of the bacterial group of interest; (iii) optionally selecting a restriction site for a restriction enzyme that is frequently encoded by the bacterial group of interest; (iv) modifying the sequence of a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, to remove a restriction site for a restriction enzyme that is frequently encoded by the bacterial group of interest; (v) optionally preparing a modified vector, in particular by nucleic acid synthesis or by mutation of the vector; Includes.

[0288] In a preferred embodiment, the sequence of the vector is modified so that it does not contain any restriction sites recognized by the restriction enzymes encoded by each bacterium of the bacterial group. The restriction sites can be modified by changing the sequence or modification state (e.g., methylated or not, glycosylated or not) to prevent recognition by the restriction enzyme (i.e., to prevent the restriction enzyme from binding to and cutting the vector). In the case of type III restriction enzymes, it is not necessary to remove all sites because one site will not cut itself. Restriction only occurs when there are two opposite restriction sites. Thus, the vector is modified to remove the presence of two opposite restriction sites.

[0289] Preferably, the bacterial population is as described above.

[0290] Preferably, the bacterial population consists of a single bacterial strain.

[0291] In another aspect, the present invention relates to an in vitro method for delivering a vector to a bacterium of interest, the method further comprising administering a modified vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a modified vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, to a bacterium of said bacterial group.

[0292] Preferably, the modified vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or the modified vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, contains no more than 100, 90, 80, 70, 60, 50, 40, 30, 20, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1 restriction sites recognised by a restriction enzyme encoded by the bacterium to which it is administered, and preferably no restriction sites.

[0293] Used as a drug In another aspect, the present invention also relates to a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or a packaged phagemid according to the invention, for use as a medicament.

[0294] The present invention also relates to the use of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or a packaged phagemid according to the invention, for the manufacture of a medicament.

[0295] In a preferred embodiment, the present invention relates to a vector according to the present invention, preferably a bacteriophage genome or phagemid according to the present invention, or a vector comprised in a delivery vehicle according to the present invention, preferably a bacteriophage or packaged phagemid according to the present invention, for use in the treatment of a disease selected from the group consisting of infectious diseases, preferably bacterial infections, inflammatory diseases, autoimmune diseases, cancer, and brain disorders. Preferably, the disease is caused by a bacterial group of interest. The ... order to improve the overall health of a subject, to eradicate pathogenic or virulent bacteria, to improve the efficacy of drugs, and / or to modify the composition of the microbiota.

[0296] The present invention also relates to a vector according to the present invention, preferably a bacteriophage genome or phagemid according to the present invention, or a vector comprised in a delivery vehicle according to the present invention, preferably a bacteriophage or packaged phagemid according to the present invention, for the manufacture of a medicament for treating a disease selected from the group consisting of an infectious disease, preferably a bacterial infection, an inflammatory disease, an autoimmune disease, cancer, and brain disorders. Preferably, the disease is caused by a bacterial group of interest. The present invention also relates to a vector according to the present invention, preferably a bacteriophage genome or phagemid according to the present invention, or a vector comprised in a delivery vehicle according to the present invention, preferably a bacteriophage or packaged phagemid according to the present invention, for the manufacture of a medicament for improving the overall health of a subject, for eradicating pathogenic or virulent bacteria, for improving the efficacy of a drug, and / or for modifying the composition of the microbiota.

[0297] The present invention further relates to a method for treating a disease in a subject selected from the group consisting of an infectious disease, preferably a bacterial infection, an inflammatory disease, an autoimmune disease, cancer, a metabolic disease or disorder, obesity, diabetes and brain disorders, wherein a therapeutically effective amount of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, is administered to said subject suffering from the disease. Preferably, the disease is caused by a bacterial group of interest. In particular, diseases caused by bacteria include abdominal cramps, acute epiglottitis, arthritis, bacteremia, botulism, brucellosis, brain abscess, soft pituitary disease, chlamydia, conjunctivitis, cholecystitis, Lyme disease, diarrhea, diphtheria, duodenal ulcer, endocarditis, erythroderma, enteric fever, fever, glomerulonephritis, gastroenteritis, gastric ulcer, Guillain-Barre syndrome, tetanus, gonorrhea, leptospirosis, leprosy, listeriosis, tuberculosis, and gynecological inflammation. symptoms, legionnaires' disease, meningitis, mucopurulent conjunctivitis, mucopurulent conjunctivitis, myonecrotizing gas gangrene, Mycobacterium avium complex, neonatal necrotizing enterocolitis, nocardiosis, hospital-acquired infections, otitis media, pharyngitis, pneumonia, peritonitis, purpuric fever, Rocky Mountain spotted fever, dysentery, syphilis, sinusitis, sigmoiditis, septicemia, subcutaneous abscess, tularemia, tracheobronchitis, tonsillitis, typhoid fever, urinary infection, and whooping cough.

[0298] The infection according to the present invention may be selected from the group consisting of skin infections such as acne, intestinal infections such as esophagitis, gastritis, enteritis, colitis, sigmoiditis, proctitis, and peritonitis, urinary tract infections, vaginal infections, female upper genital tract infections such as salpingitis, endometritis, oophoritis, metritis, endometritis, and infections of the pelvic peritoneum, respiratory tract infections such as pneumonia, intra-amniotic infections, odontogenic infections, endodontic infections, fibrosis, meningitis, bloodstream infections, hospital-acquired infections such as catheter-associated infections, hospital-acquired pneumonia, postpartum infections, hospital-acquired gastroenteritis, hospital-acquired urinary tract infections, or combinations thereof.

[0299] Preferably, the infection according to the present invention is an infection caused by a bacterium that exhibits antibiotic resistance.

[0300] In a most preferred embodiment, the infection is an infection caused by a bacterium of interest listed above.

[0301] Preferably, the vector according to the invention, preferably the bacteriophage genome or phagemid according to the invention, or the vector contained in the delivery vehicle according to the invention, preferably the bacteriophage or packaged phagemid according to the invention, targets only the bacterial strain causing the infection, making it possible to treat the subject and preserve a healthy microbiome.

[0302] The metabolic disease or disorder according to the present invention may be selected from the group consisting of metabolic brain diseases, calcium metabolism disorders, DNA repair deficiencies, glucose metabolism disorders, hyperlactatemia, iron metabolism disorders, lipid metabolism disorders, malabsorption syndromes, metabolic syndrome X, inborn errors of metabolism, mitochondrial diseases, phosphorus metabolism disorders, porphyrias and proteostasis deficiencies.

[0303] The metabolic disease or disorder according to the present invention includes, but is not limited to, type 1 diabetes; type 2 diabetes; metabolic syndrome; Bardet-Biedel syndrome; Prader-Willi syndrome; nonalcoholic fatty liver disease; tuberous sclerosis; Albright hereditary osteodystrophy; brain-derived neurotrophic factor (BDNF) deficiency; Single-minded 1 (sevil) (SEVI1) deficiency; leptin deficiency; leptin receptor deficiency; proopiomelanocortin (POMC) deficiency; proprotein convertase subtilisin / kexin type 1 (PCSK1) deficiency; Src homology 2B1 (SH2B 1) deficiency; prohormone convertase 1 / 3 deficiency; melanocortin-4-receptor (MC4R) deficiency; Wilms' tumor, aniridia, genitourinary anomalies, and mental retardation (WAGR) syndrome; pseudohypoparathyroidism type 1A; fragile X syndrome; Borjeson-Forssmann-Lehmann syndrome; Alstrom syndrome; Cohen syndrome; and ulnar mammary syndrome acid-base imbalance.

[0304] Symptoms associated with the aforementioned diseases and conditions include, but are not limited to, one or more of weight gain, obesity, fatigue, hyperlipidemia, overeating, overdrinking, excessive eating, excessive drinking, excessive urination, pain in the extremities, numbness in the extremities, blurred vision, nystagmus, hearing loss, cardiomyopathy, insulin resistance, photosensitivity, pulmonary disease, liver disease, cirrhosis, liver failure, kidney disease, kidney failure, seizures, hypogonadism, and infertility.

[0305] Metabolic disorders are associated with a variety of physiological changes, including, but not limited to, elevated glucose levels, elevated triglyceride levels, elevated cholesterol levels, insulin resistance, hypertension, hypogonadism, subfertility, infertility, abdominal obesity, prothrombotic conditions, and proinflammatory conditions.

[0306] In a particular embodiment, the present invention also relates to a personalized treatment method for an individual in need of treatment for a bacterial infection, comprising the steps of: i) obtaining a biological sample from the individual and determining a group of bacterial DNA sequences from the sample; ii) identifying one or more pathogenic bacterial strains or species present in the sample based on the sequence determination; and iii) administering to the individual a vector according to the present invention, preferably a bacteriophage genome or phagemid according to the present invention, or a vector contained in a delivery vehicle according to the present invention, preferably a bacteriophage or packaged phagemid according to the present invention, which is modified to contain 100 or less restriction sites, preferably 50 or less restriction sites, more preferably 10 or less restriction sites, and even more preferably no restriction sites, recognized by the restriction enzymes encoded by each identified pathogenic bacterial strain or species in the sample, thereby targeting one or more pathogenic bacterial strains or species. Pathogenic bacteria are defined as a group of bacteria of interest. Preferably, the vector is as defined above for targeting a pathogenic bacterial group, and is modified to remove at least one restriction site of a restriction enzyme frequently encoded by the bacterial group of interest, preferably 1, 2, 3, 4, 5, 6 or 7 restriction sites of a restriction enzyme frequently encoded by the bacterial group of interest, more preferably to remove a restriction site of a restriction enzyme frequently encoded by the bacterial group of interest.

[0307] Preferably, the biological sample comprises pathogenic and non-pathogenic bacterial species, and after administration to an individual of a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, the amount of pathogenic bacteria on or in the individual is reduced, but not the amount of non-pathogenic bacteria. In particular, the vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, results in the death of the targeted bacteria (i.e. by lysis or by expression of a toxin or a programmable nuclease circuit).

[0308] In a particular embodiment, the present invention relates to a vector according to the present invention, preferably a bacteriophage genome or phagemid according to the present invention, or a vector contained in a delivery vehicle according to the present invention, preferably a bacteriophage or packaged phagemid according to the present invention, for use in the treatment of pathologies involving bacteria of the human microbiome, such as inflammatory and autoimmune diseases, cancer, infectious diseases, metabolic diseases, metabolic disorders, such as obesity and diabetes, or brain disorders. Indeed, some bacteria of the microbiome may secrete molecules that induce and / or enhance the development of inflammatory or autoimmune diseases or cancer, without causing any infectious disease. Also, some bacteria of the microbiome secrete molecules that affect the brain.

[0309] In another particular embodiment, the present invention relates to a vector according to the present invention, preferably a bacteriophage genome or a phagemid according to the present invention, or a vector, preferably a bacteriophage or a packaged phagemid according to the present invention, contained in a delivery vehicle according to the present invention, for use to improve the efficacy of a drug. Indeed, it is known that some bacteria of the microbiome, without being pathogenic in themselves, are capable of metabolizing drugs and turning them into ineffective or harmful molecules.

[0310] The present invention also relates to non-therapeutic uses of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or a packaged phagemid according to the invention. For example, the non-therapeutic use may be a cosmetic application or a use for improving the health of a subject, in particular a subject not suffering from a disease. Thus, the present invention also relates to a cosmetic or non-therapeutic composition comprising a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or a packaged phagemid according to the invention.

[0311] Pharmaceutical or veterinary compositions In yet another aspect, the present invention also relates to a pharmaceutical or veterinary composition comprising or consisting essentially of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, or a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention.

[0312] A pharmaceutical or veterinary composition comprises at least one excipient or pharma- ceutically acceptable carrier.

[0313] Preferably, the pharmaceutical or veterinary composition further comprises another active ingredient, preferably an antibiotic or another antibacterial agent, even more preferably an antibiotic.

[0314] Preferably, the antibiotic according to the invention is a penicillin, such as penicillin G, penicillin K, penicillin N, penicillin O, penicillin V, methicillin, benzylpenicillin, nafcillin, oxacillin, cloxacillin, dicloxacillin, ampicillin, amoxicillin, pivampicillin, hetacillin, bacampicillin, metampicillin, talampicillin, epicillin, carbenicillin, ticarcillin, temocillin, mezlocillin, and piperacillin; a cephalosporin, such as cefacillin, acetrile, cefadroxil, cephalexin, cephaloglycin, cephalonium, cephaloridine, cephalothin, cephapirin, cefatrizine, cefazaflur, cefazedone, cefazolin, cephradine, cefroxadine, ceftezole, cefaclor, cefonicid, cefprozil, cefuroxime, cefuzonam, cefmetazole, cefotetan, cefoxitin, loracarbef, cefbuperazone, cefmenox, cefotetan, cefoxitin, cefotiam, cefcapene, cef Daloxime, cefdinir, cefditoren, cefetamet, cefixime, cefmenoxime, cefodizime, cefotaxime, cefovecin, cefpimizole, cefpodoxime, cefteram, ceftamele, ceftibuten, ceftiofur, ceftiolene, ceftizoxime, ceftriaxone, cefoperazone, ceftazidime, latamoxef, cefclidin, cefepime, cefluprenam, cefoselis, cefozopran, cefpirome, cefquinome, flomoxef, ceftobiprole, ceftaroline , ceftolozane, cephaloram, cefaparol, cefcanel, cephedrol, cefepidone, cefempidone, cefetrizole, cefibitril, cefmatilen, cefmepidium, cefoxazole, cefrotil, cefsumide, ceftioxide, cefraacetim, and nitrocefin; polymyxins, such as Polysporin, Neosporin, Polymyxin B, and Polymyxin E, rifampicins, such as Rifampicin, Rifapentine, and Rifaximin; fidaxomicin;Quinolones, such as cinoxacin, nalidixic acid, oxolinic acid, piromidic acid, pipemidic acid, losoxacin, ciprofloxacin, enoxacin, fleroxacin, lomefloxacin, nadifloxacin, norfloxacin, ofloxacin, pefloxacin, rufloxacin, balofloxacin, grepafloxacin, levofloxacin, pazufloxacin, temafloxacin, tosufloxacin, clinafloxacin, gatifloxacin, gemifloxacin, moxifloxacin, sitafloxacin, trovafloxacin, prulifloxacin, dela Floxacin, Nemonoxacin, and Zavofloxacin; Sulfonamides such as Sulfafurazole, Sulfacetamide, Sulfadiazine, Sulfadimidine, Sulfafurazole, Sulfisomidine, Sulfadoxine, Sulfamethoxazole, Sulfamoxole, Sulfanitran, Sulfadimethoxine, Sulfamethoxypyridazine, Sulfamethoxydiazine, Sulfadoxine, Sulfamethpyrazine, and Terephthyl; Macrolides such as Azithromycin, Clarithromycin, Erythromycin, Fidaxomicin, Telithromycin, Carbomycin A, Josamycin, Kitasamycin, Midecamycin, Oleandomycin, Solithromycin, Spiramycin, Troleandomycin, Tylosin, and Roxithromycin; Ketolides such as Telithromycin and Cethromycin; Luoloketolides such as Solithromycin; Lincosamides such as Lincomycin, Clindamycin, and Pirlimycin; Tetracyclines such as Demeclocycline, Doxycycline, Minocycline, Oxytetracycline, and Tetracycline ;aminoglycosides such as amikacin, dibekacin, gentamicin, kanamycin, neomycin, netilmicin, sisomicin, tobramycin, paromomycin, and streptomycin; ansamycins such as geldanamycin, herbimycin, and rifaximin; carbacephems such as loracarbef; carbapenems such as ertapenem, doripenem, imipenem (or cilastatin), and meropenem; glycopeptides such as teicoplanin, vancomycin, telavancin, dalbavancin, and oritavancin;Lincosamides, such as clindamycin and lincomycin; lipopeptides, such as daptomycin; monobactams, such as aztreonam; nitrofurans, such as furazolidone and nitrofurantoin; oxazolidinones, such as linezolid, pozizolid, radezolid, and torezolid; teixobactin, clofazimine, dapsone, capreomycin, cycloserine, ethambutol, ethionamide, isoniazid, pyrazinamide, rifabutin, arsphenamine, chloramphenicol, fosfomycin, fusidic acid, metronidazole, mupirocin, platensimycin, quinupristin (or dalfopristin), thiamphenicol, tigecycline, tinidazole, trimethoprim, alatrofloxacin, fidaxomicin, nalidixic acid, rifampin, derivatives and combinations thereof;

[0315] The present invention also relates to a pharmaceutical or veterinary composition of the invention for use in the treatment of an infection, preferably a bacterial infection.

[0316] The present invention also relates to the use of a pharmaceutical or veterinary composition according to the invention for the manufacture of a medicament for treating an infection, preferably a bacterial infection, in a subject.

[0317] The present invention further relates to a method of treating an infectious disease, preferably a bacterial infection, in a subject, wherein a therapeutically effective amount of a pharmaceutical or veterinary composition of the invention is administered to a subject suffering from an infectious disease.

[0318] Preferably, the infection is as described above for the use of the vector.

[0319] In a most preferred embodiment, the infection is an infection caused by a bacterium of interest listed above.

[0320] Subjects, Regimen, and Administration The subject according to the present invention is an animal, preferably a mammal, and even more preferably a human. However, the term "subject" may also refer to non-human animals, particularly mammals such as dogs, cats, horses, cows, pigs, sheep, donkeys, rabbits, ferrets, gerbils, hamsters, chinchillas, rats, mice, guinea pigs, and non-human primates, in particular, in need of treatment.

[0321] A human subject according to the present invention may be a human in the prenatal stage, a newborn, a child, an infant, an adolescent or an adult, in particular an adult at least 40 years of age, preferably an adult at least 50 years of age, even more preferably an adult at least 60 years of age, even more preferably an adult at least 70 years of age.

[0322] In a preferred embodiment, the subject has been diagnosed with an infectious disease, preferably a bacterial infection. Methods for diagnosing an infectious disease are well known to those of skill in the art.

[0323] In certain embodiments, the infection is resistant to treatment, preferably the infection is resistant to antibiotics.

[0324] In certain embodiments, the subject has already received at least one line of treatment, preferably several lines of treatment, prior to administration of a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition of the invention.

[0325] A vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition of the invention may be administered by any conventional route of administration.

[0326] In particular, a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition of the invention may be formulated for topical, enteral, oral, parenteral, intranasal, intravenous, intramuscular, subcutaneous or intraocular administration, etc.

[0327] Preferably, the vector according to the invention, preferably the bacteriophage genome or phagemid according to the invention, the vector comprised in a delivery vehicle according to the invention, preferably the bacteriophage or packaged phagemid according to the invention, or the pharmaceutical or veterinary composition of the invention may be administered by the enteral or parenteral administration route. When administered parenterally, the vector according to the invention, preferably the bacteriophage genome or phagemid according to the invention, the vector comprised in a delivery vehicle according to the invention, preferably the bacteriophage or packaged phagemid according to the invention, or the pharmaceutical or veterinary composition of the invention is preferably administered by the intravenous administration route. When administered enterally, the vector according to the invention, preferably the bacteriophage genome or phagemid according to the invention, the vector comprised in a delivery vehicle according to the invention, preferably the bacteriophage or packaged phagemid according to the invention, or the pharmaceutical or veterinary composition of the invention is preferably administered by the oral administration route.

[0328] Pharmaceutical or veterinary compositions are formulated according to standard pharmaceutical or veterinary practice known to those skilled in the art (Lippincott Williams & Wilkins, 2000 and Encyclopedia of Pharmaceutical Technology, edited by J. Swarbrick and JC Boylan, pp. 1988-1999, Marcel Dekker, New York).

[0329] For oral administration, the pharmaceutical or veterinary compositions can be formulated into conventional oral dosage forms such as tablets, capsules, powders, granules, and liquid preparations, such as syrups, elixirs, and concentrated drops. Non-toxic solid carriers or diluents can be used, including, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharin, talcum, cellulose, glucose, sucrose, magnesium, carbonates, and the like. For compressed tablets, binders are also required, which are agents that impart cohesiveness to the powdered material. For example, starch, gelatin, sugars such as lactose or dextrose, and natural or synthetic gums can be used as binders. Disintegrants are also required in tablets to facilitate the breakup of the tablet. Disintegrants include starch, clay, cellulose, algins, gums, and cross-linked polymers. In addition, lubricants and glidants are also included in the tablet to prevent adhesion of tablet materials to the surface during the manufacturing process and improve the flow properties of powdered materials during manufacture. Colloidal silicon dioxide is most commonly used as a glidant, and compounds such as talc or stearic acid are most commonly used as lubricants.

[0330] For transdermal administration, the pharmaceutical or veterinary compositions may be formulated in the form of ointments, creams or gels, and penetration may be enhanced using suitable penetrants or surfactants, such as dimethylsulfoxide, dimethylacetamide and dimethylformamide.

[0331] For transmucosal administration, nasal sprays, rectal or vaginal suppositories can be used.The active compound can be incorporated into any of the known suppository bases by methods known in the art.Examples of such bases include cocoa butter, polyethylene glycol (carbowax), polyethylene monostearate sorbitan, and mixtures thereof with other compatible materials that modify the melting point or dissolution rate.

[0332] Pharmaceutical or veterinary compositions according to the invention may be formulated to release the active ingredient substantially immediately after administration, or at any predetermined time or period after administration.

[0333] Preferably, treatment with a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition of the invention is started within one month, preferably within one week, after diagnosis of infection, in the most preferred embodiment treatment is started on the day of diagnosis.

[0334] The vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, the vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or the pharmaceutical or veterinary composition of the invention may be administered in a single dose or in multiple doses.

[0335] Preferably, the treatment is administered periodically, preferably between daily and monthly, more preferably between daily and every two weeks, more preferably between daily and weekly, and even more preferably the treatment is administered daily. In certain embodiments, the treatment is administered several times a day, preferably two or three times a day, and even more preferably three times a day.

[0336] The duration of treatment with a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, a vector comprised in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition of the invention preferably comprises from 1 day to 20 weeks, more preferably from 1 day to 10 weeks, even more preferably from 1 day to 4 weeks, even more preferably from 1 day to 2 weeks. In a particular embodiment, the duration of treatment is about 1 week. Alternatively, treatment may last for as long as the infection persists.

[0337] The amount of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or a packaged phagemid according to the invention, or a pharmaceutical or veterinary composition to be administered must be determined by standard procedures well known to those skilled in the art. To determine the appropriate dosage, it is necessary to take into account the physiological data of the patient (e.g. age, size and weight) and the route of administration, so that a therapeutically effective amount is administered to the patient.

[0338] In a preferred embodiment, the total amount per administration of a vector according to the invention, preferably a bacteriophage genome or phagemid according to the invention, or a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, is greater than or equal to 10 4 ~10 14 active particles, preferably 10 8 ~10 14 active particles, and even more preferably 10 12 ~10 14 active particles are included.

[0339] The form of the pharmaceutical or veterinary composition, the route of administration and the dosage of the vector according to the invention, preferably the bacteriophage genome or phagemid according to the invention, the vector contained in a delivery vehicle according to the invention, preferably the bacteriophage or packaged phagemid according to the invention, or the pharmaceutical or veterinary composition according to the invention can be adjusted by the skilled artisan depending on the type and severity of the infection and on the patient, in particular its age, weight, sex and general physical condition.

[0340] Kits and kit usage The present invention also relates to a kit for treating an infection, preferably a bacterial infection, in a subject, comprising a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention, a vector contained in a delivery vehicle according to the invention, preferably a bacteriophage or packaged phagemid according to the invention, or a pharmaceutical or veterinary composition according to the invention, and optionally a booklet providing guidelines for using such a kit.

[0341] Preferably, the kit further comprises another active ingredient, preferably an antibiotic.

[0342] Optionally, the kit further comprises a helper bacteriophage and / or a satellite bacteriophage.

[0343] In particular, the kit further comprises a helper bacteriophage and / or a satellite bacteriophage which facilitates encapsidation of a vector according to the invention, preferably a bacteriophage genome or a phagemid according to the invention.

[0344] The present invention also relates to the use of the above-mentioned kit in the treatment of a disease selected from the group consisting of infectious diseases, preferably bacterial infections, inflammatory diseases, autoimmune diseases, cancer, and brain disorders in a subject in need thereof.In particular, the above-mentioned kit can be used to improve the overall health of a subject, eradicate pathogenic or toxic bacteria, improve the efficacy of drugs, and / or modify the composition of the microbiota.The above-mentioned kit can also be used for non-therapeutic purposes, such as cosmetic applications, or to improve the health of a subject.

[0345] Preferably, the subject is a human.

[0346] All references cited in this application, including scientific articles and abstracts, published patent applications, granted patents or other references, are incorporated herein in their entirety, including all results, tables, figures and text of these references.

[0347] Although having different meanings, the terms "comprise," "have," "consist," and "containing" may be used interchangeably throughout this application.

[0348] Further aspects and advantages of the present invention are described in the following examples, which should be considered as illustrative and not limiting. EXAMPLES

[0349] Example 1 As a proof of concept, a set of different E. coli strains was tested for infection with bacteriophage lambda. Cells were grown overnight in liquid LB + maltose 0.2% at 37°C. The next day, cells were diluted 1:100 in fresh liquid LB medium + 0.2% maltose and grown for 2 hours at 37°C. Bacterial lawns were prepared by plating 500 μL of bacterial culture on LB-agar plates. Purified lambda bacteriophage particles, containing the wild-type 48.5 kb genome, were spotted onto the bacterial lawn and grown for 18 hours at 37°C. Lambda injection efficiency was assessed by the presence of plaques (Figure 1A and C). Only the K12-MG1655 and REL606 strains were observed to form plaques, indicating lambda phage delivery, while no effect was observed in any of the other 89 tested strains.

[0350] Next, we tested whether injection efficiency could be improved using packaged lambda-based phagemids. Potential restriction sites can be avoided by using a smaller size phagemid. For this, we constructed a 3.3 kb phagemid containing a lambda phage cos site, a constitutively expressed GFP and a chloramphenicol resistance gene (hereafter referred to as lambda phagemid) and transformed it into the CY2120b strain. This strain lacks the wild-type lambda cos site but otherwise possesses all the machinery for induction of the lambda phage lytic cycle as well as the DNA packaging system. CY2120b cells containing lambda phagemids were grown in liquid LB medium at 30°C. At an OD600 of 0.6, the culture was shifted to 42°C for 25 min to induce entry into the lytic cycle. The cells were then returned to 37°C for 3 h to allow virion assembly containing the lambda cosmid. The cells were then centrifuged and washed in lambda buffer (10 mM Tris pH 7.5, 100 mM NaCl, 10 mM MgSO4). Chloroform was added and the samples were centrifuged at 17,000 g for 5 min. Finally, the aqueous phase was collected and filtered through a 0.2 μm pore size filter. This phase contained the pure lambda packaged phagemids and was used to perform the transduction assay. 95 different strains of E. coli were grown overnight at 37°C in liquid LB + 0.2% maltose and the next day diluted 1:100 in fresh LB + maltose. After 2 h incubation at 37°C, 45 μL of cell culture was added to 45 μL of purified, packaged phagemids and further incubated at 37°C for 30 min. Finally, 10 μL of this mixture was plated on LB agar plates containing 25 μg / mL chloramphenicol and incubated at 37° C. for 18 h (FIGS. 1B and C). Of the 95 strains tested, the packaged lambda phagemid was able to inject its cargo into 34 of them, representing approximately 36% of all strains tested.

[0351] Example 2 We used six packed rhamnosidase-based phagemids to test whether removal of restriction sites within the phagemid DNA sequence would improve the efficiency of phagemid delivery and / or expand the number of bacterial strains to which the phagemid can be delivered.

[0352] For this, we constructed three variants of a larger (between 7.2 and 8.7 kb) phagemid (hereafter referred to as lambda phagemid 8 kb), and three variants of a smaller (between 2.9 and 3.3 kb) phagemid (hereafter referred to as lambda phagemid 3 kb). The three variants of each of the two phagemids differ in the presence of restriction sites recognized by the type I and type II restriction-modification (RM) systems of E. coli strains. The wild-type (WT) variants of lambda phagemid 8 kb and lambda phagemid 3 kb are not depleted of E. coli restriction sites and therefore contain multiple sequences recognized by the RM nuclease of E. coli strains. In contrast, the DNA sequences of the restriction-free (RF) variants of lambda phagemid 8 kb and 3 kb have been genetically recoded to remove most of the restriction sites of the E. coli strain. These two RF lambda phagemids are completely depleted of restriction sites known to be recognized by the RM system of a particular E. coli strain, hereafter referred to as tester strain 1. Finally, the lambda phagemid 8 kb variant "Type I" and the lambda phagemid 3 kb variant "Type II" each contain only a single restriction site (CACNNNNNNNCTGG (SEQ ID NO: 1) and GAABCC, respectively) recognized by the Type I or Type II RM system of tester strain 1.

[0353] Both lambda phagemid 8kb and lambda phagemid 3kb contain a p15a origin of replication, a lambda phage cos site, and a constitutively expressed chloramphenicol resistance gene. Lambda phagemid 8kb further contains a small guide RNA scaffold sequence and the cas9 and phlF genes.

[0354] Six lambda phagemids were transformed into strain CYC3, which lacks the wild-type lambda cos site but otherwise retains all the machinery for induction of the lambda phage lytic cycle, as well as the DNA packaging system. CYC3 cells containing individual lambda phagemids were grown in liquid LB medium at 30°C. At an OD600 of 0.7, the CYC3 culture was shifted to 42°C for 30 min to induce entry into the lytic cycle. The cells were then shifted to 37°C for 3 h to allow assembly of virions containing the lambda phagemids. The cells were then centrifuged and washed in lambda buffer (10 mM Tris pH 7.5, 100 mM NaCl, 10 mM MgSO4). Chloroform was added and the samples were centrifuged at 17,000 g for 5 min. Finally, the aqueous phase was collected and filtered through a 0.2 μm pore size filter. This phase contained pure lambda packaged phagemids and was used to perform transduction assays with tester strain I and / or a collection of 96 different E. coli strains.

[0355] E. coli strains were grown overnight at 37°C in liquid LB and diluted 1:60 or 1:100 the next day in fresh LB + 0.2% maltose + 5 mM CaCl2. Transduction assays were performed after 2.5 hours of incubation at 37°C for the recovery of 96 different strains or after the cultures of test strain I reached an optical density of 2.5.

[0356] To determine whether the presence of a single restriction site recognized by either type I or type II RM nucleases affects the frequency of phagemid delivery, test strain I has been transduced with four lambda phagemids: i) 8 kb type I; ii) 8 kb RF; iii) 3 kb type II and 3 kb RF. 50 μL of undiluted culture of test strain I was diluted undiluted or diluted at 10 in LB + 5 mM CaCl2. -1 ~10 -8The phagemids were mixed with 50 μL of purified phagemids diluted between 0.01 and 0.1 μg / mL. The phagemid:bacteria mixture was incubated at 37 °C for 30 min with shaking. Finally, 10 μL of these mixtures were placed on LB agar plates containing 25 μg / mL chloramphenicol. Streaks were formed by tilting the plate so that each 10 μL drop ran down the LB agar surface. The LB agar plates were incubated at 37 °C for 18 h. Colony forming units (CFU) were quantified for the streaks using the phagemid dilution giving the highest number of individual countable colonies (Figure 2).

[0357] CFU corresponds to the number of test strain I cells successfully transduced with the phagemid. To evaluate the delivery efficiency of the four phagemids in test strain I, the obtained CFU numbers were normalized to the total titer of each phagemid (Table 3). The total titer of the phagemids was evaluated in parallel by transducing the E. coli strain MG1655, which is permissive for the tested phagemids. The delivery efficiency of lambda phagemids 8kb and 3kb RF was compared with that of lambda phagemids 8kb type I and lambda phagemids 3kb type II, respectively (Table 3). Lambda phagemids 8kb RF and 3kb RF, deprived of restriction sites recognized by RM nuclease of test strain 1, were delivered 3-5 logs and 1-2 logs more efficiently in test strain 1 than lambda phagemids 8kb type I and 3kb type II, which contain single restriction sites for type I and type II RM systems, respectively. These results indicate that removal of restriction sites recognized by the RM system of a particular strain improves phagemid delivery efficiency. Moreover, the presence of even a single additional restriction site within the phagemid DNA sequence can significantly reduce its delivery into a bacterial strain.

[0358] We tested whether removing multiple restriction sites recognized by various RM nucleases of different strains of E. coli from the phagemid DNA sequence would improve the efficiency of phagemid delivery and / or expand the number of strains to which phagemids can be delivered. For this, 87 different E. coli strains were transduced with four lambda phagemids: i) 8kb WT; ii) 8kb RF; iii) 3kb WT and 3kb RF. 90 μL of undiluted culture of the 87 strains was mixed with 10 μL of purified phagemid. The phagemid:bacteria mixture was incubated at 37° C. for 30 min with shaking. Finally, 10 μL of these mixtures were spotted on LB agar plates containing 25 μg / mL chloramphenicol. Growth of the 87-spot strains of E. coli, indicating successful phagemid delivery, was evaluated after 18 h of incubation of the plates at 37°C (Figure 3, Table 4). The lambda phagemid 8kb RF, depleted of multiple E. coli restriction sites, was transduced into 38 of the 87 strains tested, which represents approximately 44% of all strains tested (Table 2, Figure 2). In contrast, the packaged lambda phagemid 8kb WT, which does not have a reduced number of restriction sites, was transduced into 27 (31%) of the strains tested. Similarly, the lambda phagemid 3kb RF was transduced into a higher number of E. coli strains tested (64), which represents 74% of the strains tested when compared to the lambda phagemid 3kb WT (45 strains; 52%). The high number of strains grown after transduction with the RF variants of the phagemids indicates that removal of multiple restriction sites expands the number of strains to which the two phagemids are delivered (Table 4, FIG. 3).

[0359] [Table 3]

[0360] [Table 4]

Claims

1. 1. A method for preparing a vector containing 10 or fewer restriction sites for restriction enzymes encoded by a bacterial group of interest, the method comprising: (i) selecting a bacterial group of interest: (ii) identifying restriction sites based on the vector sequence that are recognized by restriction enzymes encoded by the bacterial group; (iii) modifying the sequence of the vector to contain a desired number of restriction sites recognized by restriction enzymes encoded by each bacterium in the population; wherein restriction sites in non-coding regions of the vector are modified first and only those restriction sites in coding regions of the vector that do not reach the desired number of restriction sites, thereafter all restriction sites in the non-coding regions are modified until the desired number of restriction sites is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10, and the vector is a bacteriophage genome or a phagemid.

2. The method of claim 1 , wherein the vector is contained in a delivery vehicle.

3. The method of claim 2 , wherein the vector is a bacteriophage or a packaged phagemid.

4. 2. The method of claim 1, wherein in step (ii) the frequency of a restriction enzyme in the bacterial group of interest is determined, and in step (iii) the sequence of the vector is modified to remove at least one restriction site for a restriction enzyme that is frequently encoded by the bacterial group of interest.

5. 5. The method of claim 4, wherein a restriction enzyme is present at a high frequency in a bacterial population of interest if at least 10% of the bacteria in the bacterial population encode the restriction enzyme.

6. The method of claim 1, wherein at least 10% of the restriction sites originally present in the vector have been removed.

7. The method of claim 1, wherein at least 90% of the restriction sites originally present in the vector have been removed.

8. The method of claim 1, wherein 100% of the restriction sites originally present in the vector have been removed.

9. The method described in claim 1, wherein the vector does not contain a restriction site for a restriction enzyme encoded by the target bacterial group.

10. The method of claim 1, wherein the modification is carried out by deletion of part or all of the restriction site or by nucleotide substitution of at least one nucleotide of the restriction site.

11. The method of claim 10, wherein the deletion is only in a non-coding region.

12. 2. The method of claim 1, wherein the bacterial population of interest consists of a population of between 2 and 100,000 bacterial strains.

13. 13. The method of claim 12, wherein the bacterial group of interest is selected from a single species.

14. The target bacteria are Yersinia spp., Escherichia spp., Klebsiella spp., Acinetobacter spp., Bordetella spp., Neisseria spp., Aeromonas spp., Franciella spp., Corynebacterium spp., Citrobacter spp., Chlamydia spp., Hemophilus spp., Brucella spp., Mycobacterium spp., and spp.), Legionella spp., Rhodococcus spp., Pseudomonas spp., Helicobacter spp., Vibrio spp., Bacillus spp., Erysipelothrix spp., Salmonella spp., Streptomyces spp., Streptococcus spp., Staphylococcus spp., Bacteroides spp., Prevotella spp. spp., Clostridium spp., Bifidobacterium spp., Clostridium spp., Brevibacterium spp., Lactococcus spp., Leuconostoc spp., Actinobacillus spp., Selnomonas spp.), Shigella spp., Zymonas spp., Mycoplasma spp., Treponema spp., Leuconostoc spp., Corynebacterium spp., Enterococcus spp., Enterobacter spp., Pyrococcus spp., Serratia spp., Morganella spp., Parvimonas spp., Fusobacterium spp., Actinomyces spp. spp., Porphyromonas spp., Micrococcus spp., Bartonella spp., Borrelia spp., Brucelia spp., Campylobacter spp., Chlamydophilia spp., Ehrlichia spp., Haemophilus spp., Leptospira spp., Listeria spp., Mycoplasma spp., Nocardia spp., Rickettsia spp. spp., Ureaplasma spp., Cutibacterium spp. (formerly Propionibacterium spp.), and Lactobacillus spp., and mixtures thereof.

15. 15. The method of claim 14, wherein the bacterium of interest is selected from the Escherichia species.

16. The target bacteria are Bacteroides thetaiotaomicron, Bacteroides fragilis, Bacteroides distasonis, Bacteroides vulgatus, Clostridium leptum, Clostridium coccoides, Staphylococcus aureus, Bacillus subtilis, Clostridium butyricum, Brevibacterium lactofermentum, Streptococcus pyogenes, and Streptococcus pyogenes. agalactiae, Lactococcus lactis, Leuconostoc lactis, Actinobacillus actinobycetemcomitans, cyanobacteria, Escherichia coli, Helicobacter pylori, Selnomonas ruminatium, Shigella sonnei, Zymomonas mobilis, Mycoplasma mycoides, Treponema denticola, Bacillus thuringiensis, Staphylococcus lugdunensis, lugdunensis, Leuconostoc oenos, Corynebacterium xerosis, Lactobacillus plantarumplantarum, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus acidophilus, Enterococcus faecalis, Bacillus coagulans, Bacillus cereus, Bacillus popillae, Synechocystis strain PCC6803, Bacillus liquefaciens, Pyrococcus abyssi, Selenomonas nominantium, Lactobacillus hilgardii hilgardii, Streptococcus ferus, Lactobacillus pentosus, Bacteroides fragilis, Staphylococcus epidermidis, Streptomyces phaechromogenes, Streptomyces ghanaenis, Klebsiella pneumoniae, Enterobacter cloacae, Enterobacter aerogenes, Serratia marcescens, Morganella morganii morganii, Citrobacter freundii, Pseudomonas aerigunosa, Parvimonas micra, Prevotella intermediaintermedia, Fusobacterium nucleatum, Prevotella nigrescens, Actinomyces israelii, Porphyromonas endodontalis, Porphyromonas gingivalis, Micrococcus luteus, Bacillus megaterium, Aeromonas hydrophila, Aeromonas caviae, Bacillus anthracis, Bartonella henselae, Bartonella quintana Quintana, Bordetella pertussis, Borrelia burgdorferi, Borrelia garinii, Borrelia afzelii, Borrelia recurrentis, Brucella abortus, Brucella canis, Brucella melitensis, Brucella suis, Campylobacter jejuni, Campylobacter coli, Campylobacter fetus, Chlamydia pneumoniae, Chlamydia trachomatis trachomatis, Chlamydophila psittaci, Clostridium botulinum, Clostridium difficiledifficile, Clostridium perfringens, Clostridium tetani, Corynebacterium diphtheria, Cutibacterium acnes (formerly Propionibacterium acnes), Ehrlichia canis, Ehrlichia chaffeensis, Enterococcus faecium, Francisella tularensis, Haemophilus influenza, Legionella pneumophila, Leptospira interrogans interrogans, Leptospira santarosai, Leptospira weilii, Leptospira noguchii, Listeria monocytogenes, Mycobacterium leprae, Mycobacterium tuberculosis, Mycobacterium ulcerans, Mycoplasma pneumonia, Neisseria gonorrhoeae, Neisseria meningitides, Nocardia asteroids, Rickettsia rickettsia, Salmonella enteritidis enteritidis, Salmonella typhi, Salmonella paratyphiparatyphi, Salmonella typhimurium, Shigella flexnerii, Shigella dysenteriae, Staphylococcus saprophyticus, Streptococcus pneumoniae, Streptococcus pyogenes, Streptococcus viridans, Treponema pallidum, Ureaplasma urealyticum, Vibrio cholera, Vibrio parahaemolyticus, Yersinia pestis 2. The method of claim 1, wherein the bacterial strain is selected from the group consisting of Yersinia pestis, Yersinia enterocolitica, Yersinia pseudotuberculosis, Actinobacter baumanii, Pseudomonas aerigunosa, and mixtures thereof.

17. The method of claim 16, wherein the target bacterium is selected from the group consisting of Escherichia coli, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumanii, Pseudomonas aeruginosa, Enterobacter cloacae, Enterobacter aerogenes, Cutibacterium acnes, and Fusobacterium nucleatum, and mixtures thereof.

18. 17. The method of claim 16, wherein the bacterium of interest is selected from an E. coli strain.

19. Bacteriophages include IKe, CTX-φ, Pf1, Pf2, Pf3, Myoviridae, Siphoviridae, Podoviridae, Tectiviridae, Corticoviridae, Liposthrixviridae, Plasmaviridae, Rudiviridae, Fuselloviridae, Inoviridae, Microviridae, Leviviridae, Cystoviridae, E. coli phage, B1, ATCC 5. The method of claim 4, wherein the bacteriophage is selected from the group consisting of 51477-B1, B40-8, Bf-1, phiHSC01, phiHSC02, phiC2, phiC5, phiC6, phiC8, phiCD119, phiCD27, KP01K2, K11, Kpn5, KP34, JD001, phiNM1, 80alpha, IME-EF1, ENB6, C33, phiKMV, PAK-P1, LKD16, LKA1, delta, sigma-1, J1, T2, T4, T5, T7, RB49, phiX174, R17, and PRD1, or the phagemid is packaged into the capsid of one of these bacteriophages.

20. 2. The method of claim 1, wherein the phagemid is selected from the group consisting of a lambda-derived phagemid, a P4-derived phagemid, and an M13-derived phagemid.

21. 21. The method of claim 20, wherein the phagemid is selected from the group consisting of HK022-derived phagemid, mEP237-derived phagemid, HK97-derived phagemid, HK629-derived phagemid, HK630-derived phagemid, mEP043-derived phagemid, mEP213-derived phagemid, mEP234-derived phagemid, mEP390-derived phagemid, mEP460-derived phagemid, mEPx1-derived phagemid, mEPx2-derived phagemid, phi80-derived phagemid, and mEP234-derived phagemid.

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