Enterococcus faecalis vaccine and uses thereof

Abstract

The present disclosure provides an immunogenic composition comprising at least one recombinant polypeptide antigen derived from Enterococcus bacteria (e.g., E. faecalis, E. faecium, E. durans). The present disclosure further provides a method and a use of the immunogenic composition for protecting or treating a subject against infection with Enterococcus bacteria. Such infections may cause or exacerbate conditions such as root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection or wound infection.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 332,110, filed April 18, 2022, the disclosure of which is incorporated herein by reference in its entirety.

[0002] Description of electronically submitted text files The contents of the electronic sequence listing (STRO_023_01WO_SeqList_ST26.xml; size: 28,215 bytes; and creation date: March 15, 2023) are incorporated herein by reference in their entirety.

[0003] The present invention relates generally to the fields of microbiology and vaccine development. [Background technology]

[0004] Approximately 15 million root canal procedures are performed annually in the United States, with approximately 20% (3 million) failing due to infection. The infection is primarily caused by Enterococcus faecalis, a type of bacteria in the large gram-positive Enterococcus genus. The cost of follow-up treatment or implants for failed root canals currently averages $2,000. Assuming a 50% reimbursement, the cost to insurance companies would be approximately $3 billion per year. In addition, E. faecalis causes other serious and sometimes fatal opportunistic infections in humans, such as endocarditis, urinary tract infections, prostatitis, intraperitoneal infections, cellulitis, and wound infections.

[0005] Enterococcus species also have a high intrinsic antibiotic resistance, with some strains acquiring resistance to potent second-generation antibiotics such as vancomycin (see, e.g., Arias, CA, et al. Nat. Rev. Microbiol. 2012, 266-278; Puchter, L., et al. Antimicrob. Resist. Infect. Control 2018). A vaccine that would reduce root canal failure rates, or any of the other conditions described herein, would have a favorable impact on pharmaceutical economics and human health implications. However, there are currently no approved or promising vaccine candidates. Thus, there is a need for first-in-class prophylactics and treatments against Enterococcus infections. Summary of the Invention [Problem to be solved by the invention]

[0006] Generally, an immunogenic composition for Enterococcus infection comprises at least one recombinant Enterococcus polypeptide antigen or a fragment thereof. The immunogenic composition may comprise at least one recombinant Enterococcus faecalis polypeptide antigen or a fragment thereof. The polypeptide antigen may be, for example, an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, an Esp polypeptide antigen, or any suitable recombinant polypeptide antigen derived from an Enterococcus protein. [Means for solving the problem]

[0007] The immunogenic composition may, for example, comprise a first and a second recombinant Enterococcus polypeptide antigen. The immunogenic composition may also comprise a first, a second, and a third recombinant Enterococcus polypeptide antigen. Some immunogenic compositions may comprise a first, a second, a third, and a fourth recombinant Enterococcus polypeptide antigen. Some immunogenic compositions may comprise a first, a second, a third, a fourth, and a fifth recombinant Enterococcus polypeptide antigen. The immunogenic composition may also comprise six or more recombinant Enterococcus polypeptide antigens.

[0008] In general, the method of inducing a protective immune response against Enterococcus bacteria may comprise administering an immunogenic composition comprising at least one recombinant Enterococcus polypeptide antigen.In general, the use of the immunogenic composition can induce a protective immune response against Enterococcus bacteria.In general, the immunogenic composition or the method or use thereof may be useful for patients suffering from or at risk of suffering from root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal flora dysbiosis, prosthetic joint infection, or wound infection. [Brief description of the drawings]

[0009] [Figure 1] FIG. 1 shows the average total and average soluble protein (μg / mL) of expressed polypeptide antigens and the accompanying SDS-PAGE gel. [Diagram 2] FIG. 2 shows the relative expression of two different EbpA constructs: full-length and EbpA-IpaH domain. [Diagram 3] FIG. 3 shows a gel of E. faecalis EfaA polypeptide antigen expression / purification and following TEV cleavage. [Figure 4] FIG. 4 shows a gel of E. faecalis Ace polypeptide antigen expression / purification and following TEV cleavage. [Diagram 5] FIG. 5 shows a gel of E. faecalis Esp polypeptide antigen expression / purification and following TEV cleavage. [Figure 6]FIG. 6 shows a gel of E. faecalis AdcA polypeptide antigen expression / purification and following TEV cleavage. [Figure 7] FIG. 7 shows a gel of E. faecalis AdcAII polypeptide antigen expression / purification and following TEV cleavage. [Figure 8] FIG. 8 shows a gel of E. faecalis EpbA polypeptide antigen expression / purification and following TEV cleavage. [Figure 9] FIG. 9 shows a chromatograph of SEC-MALS molecular weight analysis of the polypeptide antigens of SEQ ID NOs:12-18. [Figure 10] Figures 10A and 10B show SDS-PAGE and SEC-MALS data, respectively, of the batches of expressed polypeptide antigens used in animal studies. [Figure 11] FIG. 11A shows that AdcA is relatively stable to freeze-thaw cycles, with only a slight decrease in the intensity and molecular weight of the main peak. FIG. 11B shows that AdcAII is also relatively stable to freeze-thaw cycles, with only a slight decrease in the intensity of the main peak and a slight increase in molecular weight. FIG. 11C shows that Ace is stable to freeze-thaw cycles. FIG. 11D shows that the stability of two different Esp samples to freeze-thaw cycles. FIG. 11D shows that Esp is stable when 10% glycerol is added to the storage buffer. FIG. 11E and 11F show that EfaA and EpbA, respectively, are relatively stable to freeze-thaw cycles. [Figure 12]Figure 12A shows immunoblots resulting from probing cell lysates generated from wild-type E. faecalis OG1RF, E. faecalis OG1RF ΔAdcACB / ΔAdcA double null mutant, and 0.25 μg each of the AdcA and AdcAII antigens used to generate the mouse antisera, along with AdcA mouse antiserum. Figure 12B shows immunoblots resulting from probing cell lysates generated from wild-type E. faecalis OG1RF, E. faecalis OG1RF ΔAdcACB / ΔAdcA double null mutant, and 0.25 μg each of the AdcA and AdcAII antigens used to generate the mouse antisera, along with AdcAII mouse antiserum. [Figure 13] FIG. 13 shows survival data from a G. mellonella immunization model utilizing rabbit antisera from an exemplary E. faecalis polypeptide antigen-treated arm. [Figure 14] Figure 14A shows the dosing schedule for an immunization study in rabbits utilizing exemplary E. faecalis immunogenic compositions, and Figure 14B shows each immunogenic composition tested in the rabbit cohort. [Figure 15] FIG. 15 shows that antisera obtained from animals administered individual E. faecalis polypeptide antigens elevate antigen-specific IgG titers compared to pre-immune animals and alum controls. [Figure 16] FIG. 16 shows a schematic of an opsonophagocytosis (OPA) assay used to assess the efficacy of antibody-dependent killing by neutrophils following immunization with selected E. faecalis polypeptide antigens. [Figure 17] FIG. 17 shows the influence of the opsonizing / neutralizing effect of antiserum antibodies on neutrophil-mediated killing of E. faecalis in vitro. [Figure 18] FIG. 18 shows a schematic diagram of the passive immunization mouse peritonitis model. [Figure 19]FIG. 19 shows a summary of colony forming unit (CFU) data generated from a mouse peritoneal (peritonitis) infection model in two body sites (peritoneal cavity and spleen) tested for the presence of active E. faecalis after passive immunization. [Figure 20] FIG. 20 shows a summary of colony forming unit data for kidneys in a mouse peritoneal (peritonitis) infection model examined for the presence of active E. faecalis after passive immunization. [Figure 21] Figures 21A, 21B, and 21C show end-bleed IgG titers for 10 treatment groups in a passive immunization study to determine the protective efficacy of antisera generated against combinations of E. faecalis antigens. [Figure 22] FIG. 22 shows a schematic diagram of the passive immunization mouse peritonitis model without a second booster injection of serum. [Figure 23] Figures 23A, 23B, 23C, 23D, 23E, 23F, 23G, and 23H show survival data generated from a G. merulonella immunization model utilizing rabbit antisera obtained from an exemplary E. faecalis polypeptide antigen treatment arm. [Figure 24] Figures 24A and 24B show the efficacy of E. faecalis OG1RF antigen-specific antisera against clinically relevant E. faecalis and E. faecium strains, respectively. Figure 24C shows a summary of colony forming unit data generated from a mouse peritoneal (peritonitis) infection model for the presence of active E. faecalis and E. faecium following passive immunization. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0010] Although vaccines against enterococcal infections have been previously investigated (see, for example, Kalfopoulou, E., et al. Cells, 2020, 2397), no prophylactic or therapeutic vaccine has been successfully developed, despite the potentially serious health impacts caused by pathogenic Enterococcus species (e.g., E. faecalis, E. faecium, E. durans).

[0011] immunogenic composition In some embodiments, the disclosure provides immunogenic compositions comprising at least one recombinant Enterococcus (e.g., E. faecalis, E. faecium, E. durans) polypeptide antigen or fragment thereof.

[0012] As used herein, the term "immunogenic" refers to the ability of an antigen (e.g., a polypeptide) to elicit an immune response, including either a humoral or cellular immune response, preferably both. In some embodiments, a subject exhibits either a therapeutic or protective immunological response to administration of an "effective amount" or an "immunologically effective amount" of an immunogenic composition herein, thereby enhancing resistance to new infections and / or reducing the clinical severity of disease. An immunological response is usually indicated by the alleviation or elimination of at least one symptom associated with the infection.

[0013] An "immunologically effective amount" or "effective amount" of an immunogenic composition is an amount that is effective, either in a single dose or as part of a series of two or more doses, to treat or prevent an enterococcal infection or an enterococcal-related disease or condition. The amount administered will vary depending on several factors, including the subject's overall health and physical condition, the subject's age, the capacity of the subject's immune system to synthesize the relevant antibodies, the form of the composition (e.g., injectable liquid, nasal spray, etc.), and other factors known to the medical practitioner overseeing administration.

[0014] The term "treatment" refers to therapeutic treatment by administration of an immunogenic composition for the purpose of reducing or eliminating an infection. For example, "treatment" can include directly affecting, suppressing, inhibiting, and eliminating an infection, as well as reducing the severity of an infection, delaying its onset, and / or reducing symptoms associated with an infection. Unless otherwise expressly indicated or implied by context, the term "treatment" encompasses "prevention" (or preventative measures or prophylactic treatment), where "prevention" can refer to reducing the risk of a subject developing an infection, delaying the onset of symptoms, preventing the recurrence of an infection, or preventing the occurrence of an infection.

[0015] As used herein, a "recombinant" polypeptide refers to a polypeptide produced by recombinant DNA technology (e.g., generated from a cell transformed with an exogenous DNA construct encoding a desired polypeptide). In some embodiments, one or more recombinant polypeptides are synthesized using scalable cell-free protein synthesis (CFPS) as described in U.S. Pat. No. 9,040,253, U.S. Pat. No. 9,650,621, and Murray, et al. 2013 Current Opin. Chem. Biol. 17(3): 420-26, all of which are incorporated herein by reference. The expressed polypeptide may include a leader sequence to aid in isolation and / or purification. For example, the peptide may have a terminal His (e.g., 6xHis) tag. The peptide may have a cleavable (e.g., protease-cleavable) linker between the terminal tag and the primary polypeptide sequence. It should be understood that the polypeptides of the immunogenic compositions described herein are recombinant polypeptides.

[0016] In some embodiments, the present disclosure provides purified recombinant Enterococcus polypeptide antigens. As used herein, when the term "purified" is used in reference to a molecule, it means that the concentration of the purified molecule is increased compared to the concentration of the molecule in the environment in which it is produced. The term may also refer to the purification of a chemically synthesized molecule from a reaction mixture in which it is produced as a reaction product. As used herein, when the term "isolated" is used in reference to a molecule, it means that the molecule has been removed from its natural environment. For example, a polynucleotide or polypeptide that is naturally present in a living organism is "not isolated," but the same polynucleotide or polypeptide that is separated from coexisting materials in its natural state is in an "isolated" state. An isolated portion, whether separated from a natural or non-native environment (e.g., recombinant expression, cell-free expression, chemical synthesis, etc.), is preferably at least about 1% pure, 5% pure, 10% pure, 20% pure, 30% pure, 40% pure, 50% pure, 60% pure, 70% pure, 80% pure, 90% pure, 95% pure, 99% pure, or 100% pure. As used herein, the term "% purity" refers to the percent by weight of a composition that is composed of the molecule of interest.

[0017] As used herein, the term "polypeptide" is intended to include any structure composed of one or more amino acids, and thus includes dipeptides, oligopeptides, polypeptides, and / or polypeptide fragments thereof. The amino acids forming all or part of a polypeptide may be any of the 20 conventional naturally occurring amino acids, i.e., alanine (A), cysteine ​​(C), aspartic acid (D), glutamic acid (E), phenylalanine (F), glycine (G), histidine (H), isoleucine (I), lysine (K), leucine (L), methionine (M), asparagine (N), proline (P), glutamine (Q), arginine (R), serine (S), threonine (T), valine (V), tryptophan (W), tyrosine (Y), as well as non-conventional amino acids, e.g., They may be isomers and variants of conventional amino acids, (e.g., D-amino acids, non-protein amino acids, post-translationally modified amino acids, enzymatically modified amino acids, β-amino acids, etc.), constructs or structures designed to mimic amino acids (e.g., α,α-substituted amino acids, N-alkyl amino acids, lactate, β-alanine, naphthylalanine, 3-pyridylalanine, 4-hydroxyproline, O-phosphoserine, N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine, and nor-leucine), and other unconventional amino acids.

[0018] In some embodiments, the polypeptide antigen described herein is an Enterococcus membrane-associated polypeptide. For example, in some embodiments, the polypeptide antigen is an Enterococcus collagen adhesin (e.g., "Ace" polypeptide antigen from E. faecalis, E. faecium, E. durans). In certain embodiments, the polypeptide antigen is a zinc ABC transporter substrate-binding lipoprotein (e.g., "AdcA" polypeptide antigen from E. faecalis, E. faecium, E. durans, "AdcAII" polypeptide antigen from E. faecalis, E. faecium, E. durans). In certain embodiments, the polypeptide antigen is an endocarditis and biofilm-associated pili protein (e.g., "EbpA" polypeptide antigen from E. faecalis, E. faecium, E. durans). In certain embodiments, the polypeptide antigen is an ABC superfamily ATP-binding cassette transporter (e.g., "EfaA" polypeptide antigen from E. faecalis, E. faecium, E. durans). In some embodiments, the polypeptide antigen is an Enterococcal surface protein (eg, the "Esp" polypeptide antigen from E. faecalis, E. faecium, E. durans).

[0019] The amino acid sequences of exemplary Enterococcus polypeptides are shown in Table 1 below.

[0020] [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5]

[0021] The recombinant polypeptide antigen can be a full-length wild-type (WT) polypeptide sequence from an Enterococcus species (e.g., E. faecalis, E. faecium, E. durans), such as SEQ ID NOs: 1-12. The polypeptide antigen can additionally or alternatively be a fragment of a full-length wild-type protein from an Enterococcus species. For example, in some embodiments, the polypeptide antigen of the compositions and methods described herein can be a full-length wild-type E. faecalis Ace polypeptide antigen (e.g., SEQ ID NO: 1) or a full-length wild-type E. faecium Ace polypeptide antigen (e.g., SEQ ID NO: 7). In some embodiments, the polypeptide antigen can be a fragment of a wild-type full-length polypeptide antigen (e.g., SEQ ID NO: 13, residues 30-366 of the E. faecalis Ace polypeptide). In some embodiments, the polypeptide antigen can be a fragment of any of the polypeptide antigens described herein. For example, a fragment can be at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of a polypeptide antigen (e.g., a wild-type polypeptide antigen or an antigen sequence specified herein). In other words, a fragment can have at least 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the number of residues of the parent polypeptide antigen. In some embodiments, a polypeptide antigen fragment is at least 10-15%, 10-20%, 10-25%, 10-30%, 10-35%, 10-40%, 10-45%, 10-50%, 10-55%, 10-65%, 10-70%, 10-75%, 10-80%, 10-85%, 10-90%, 10-95%, ...30%, 10-40%, 10-45%, 10-50%, 10-55%, 10-65%, 10-70%, 10-75%, 10-80%, 10-85%, 10-90%, 10-95%, 10-15%, 10-20%, 10-20% or 10-30% of the length of a polypeptide antigen or antigen sequence designated herein. ~20%, 15~25%, 15~30%, 15~35%, 15~40%, 15~45%, 15~50%, 15~55%, 15~65%, 15~70%, 15~75%, 15~80%, 15~85%, 15~90%, 15~95%, 20~25%, 20~30%, 20~35%, 20~40%, 20~45%, 20~50%, 20~55%, 20~65%, 20~70%, 20~75%,20~80%, 20~85%, 20~90%, 20~95%, 25~30%, 25~35%, 25~40%, 25~45%, 25~50%, 25~55%, 25~65%, 25~70%, 25~75%, 25~80%, 25~85%, 25~90%, 25~95%, 30~35%, 30~40%, 30~45%, 30~50%, 30~55%, 30~65%, 30~70%, 30~75%, 30~ 80%, 30~85%, 30~90%, 30~95%, 35~40%, 35~45%, 35~50%, 35~55%, 35~65%, 35~70%, 35~75%, 35~80%, 35~85%, 35~90%, 35~95%, 40~45%, 40~50%, 40~55%, 40~65%, 40~70%, 40~75%, 40~80%, 40~85%, 40~90%, 40~95%, 45~50%, 45~55%, 45~65%, 45~70%, 45~75%, 45~80%, 45~85%, 45~90%, 45~95%, 50~55%, 50~65%, 50~70%, 50~75%, 50~80%, 50~85%, 50~90%, 50~95%, 55~65%, 55~70%, 55~75%, 55~80%, 55~85%, 55~90%, 55~95%, 65~70%, 65~75%, 65~8 0%, 65-85%, 65-90%, 65-95%, 65-75%, 65-80%, 65-85%, 65-90%, 65-95%, 70-75%, 70-80%, 70-85%, 70-90%, 70-95%, 75-75%, 75-80%, 75-85%, 75-90%, 75-95%, 80-85%, 80-90%, 80-95%, 85-90%, 85-95%, or 85-95%. Generally, useful fragments are antigenic, e.g., the fragments are of sufficient length to elicit a protective immune response (e.g., as measured by methods including animal models described herein, including in the Examples).

[0022] The immunogenic compositions described herein include at least one recombinant Enterococcus (e.g., E. faecalis, E. faecium, E. durans) polypeptide antigen or fragment thereof. In some embodiments of such immunogenic compositions, the at least one recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen. In some embodiments of such immunogenic compositions, the at least one recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen. In some embodiments of such immunogenic compositions, the at least one recombinant E. durans polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0023] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an Ace polypeptide antigen or a fragment thereof. The Ace polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO: 1, 7, or 13. In some embodiments, the Ace polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NO: 1, 7, or 13. In some embodiments, the Ace polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO: 1, 7, or 13.

[0024] "Percent identity" (e.g., "90% identical") in the context of the polypeptide sequences described herein refers to two or more sequences that are identical or have a certain percentage of identical amino acid residues when compared and aligned for maximum correspondence over a given length (comparison window) as measured using a sequence comparison algorithm (e.g., BLASTP or Smith-Waterman homology search algorithm). In the context of the present specification, percent sequence identity can be determined for the entire length of a polypeptide or only a portion of it. One method for calculating percent sequence identity is the BLASTP program with default settings of word length (W) of 3, expectation (E) of 10, and BLOSUM62 scoring matrix; see, e.g., Henikoff et al. (1989) Proc. Natl. Acad. Sci. USA 89:15915. Exemplary determinations of sequence alignment and percent sequence identity use the BESTFIT or GAP programs of the GCG Wisconsin Software package (Accelrys, Madison Wis.) using default parameters. If these preferred methods of calculating sequence identity yield different amounts, preference is given to the method showing the higher sequence identity.

[0025] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an AdcA polypeptide antigen or a fragment thereof. The AdcA polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO: 2, 8, or 14. In some embodiments, the AdcA polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NO: 2, 8, or 14. In some embodiments, the AdcA polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO: 2, 8, or 14.

[0026] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an AdcAII polypeptide antigen or a fragment thereof. The AdcAII polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO: 3, 9, or 15. In some embodiments, the AdcAII polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NO: 3, 9, or 15. In some embodiments, the AdcAII polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO: 3, 9, or 15.

[0027] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an EbpA polypeptide antigen or a fragment thereof. The EbpA polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO: 4, 15, or 16. In some embodiments, the EbpA polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NO: 4, 15, or 16. In some embodiments, the EbpA polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO: 4, 15, or 16.

[0028] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an EfaA polypeptide antigen or a fragment thereof. The EfaA polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO:5, 11, or 17. In some embodiments, the EfaA polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any of SEQ ID NO:5, 11, or 17. In some embodiments, the EfaA polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO:5, 11, or 17.

[0029] In some embodiments, at least one recombinant Enterococcus polypeptide antigen is an Esp polypeptide antigen or a fragment thereof. The Esp polypeptide antigen may, for example, comprise or consist of the amino acid sequence of SEQ ID NO: 6, 12, or 18. In some embodiments, the Esp polypeptide antigen comprises or consists of an amino acid sequence that is at least 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to any one of SEQ ID NO: 6, 12, or 18. In some embodiments, the Esp polypeptide antigen comprises or consists of an amino acid sequence that is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of any one of SEQ ID NO: 6, 12, or 18.

[0030] The immunogenic compositions described herein may comprise or consist of any number of recombinant Enterococcus polypeptide antigens. For example, the immunogenic compositions may comprise or consist of one, two, three, four, five, six, or more Enterococcus polypeptide antigens. The immunogenic compositions may comprise or consist of, for example, 1-6, 1-5, 1-4, 1-3, 1-2, 2-6, 2-5, 2-4, 2-3, 3-6, 3-5, 3-4, 4-6, 4-5, or 5-6 polypeptide antigens. In some embodiments, the immunogenic compositions may comprise polypeptide antigens of different Enterococcus proteins (e.g., a wild-type Ace polypeptide antigen and a wild-type AdcA polypeptide antigen). Additionally or alternatively, the immunogenic compositions may comprise multiple polypeptide antigens of the same Enterococcus protein (e.g., two different fragments of an AdcA polypeptide antigen). In some embodiments, 1, 2, 3, 4, 5, 6, or more of the Enterococcus polypeptide antigens are fragments of the polypeptide antigen.

[0031] In some embodiments, the immunogenic composition may comprise more than one recombinant polypeptide antigen, or fragment thereof, from the same Enterococcus species (e.g., two or more polypeptide antigens from E. faecalis). In certain embodiments, the immunogenic composition may comprise more than one polypeptide antigen, or fragment thereof, from different Enterococcus species (e.g., at least one polypeptide antigen from E. faecalis and at least one polypeptide antigen from E. faecium; at least one polypeptide antigen from E. faecalis and at least one polypeptide antigen from E. faecium; at least one polypeptide antigen from E. faecalis and at least one polypeptide antigen from E. durans; at least one polypeptide antigen from E. faecium and at least one polypeptide antigen from E. durans; at least one polypeptide antigen from each of E. faecium, E. faecalis, and E. durans).

[0032] In some embodiments, the immunogenic compositions described herein comprise at least one recombinant E. faecalis polypeptide antigen or a fragment thereof. For example, the immunogenic compositions may comprise a first recombinant E. faecalis polypeptide antigen and a second recombinant E. faecalis polypeptide. The immunogenic compositions may comprise a third, fourth, fifth, or sixth E. faecalis polypeptide. Thus, in certain embodiments, the immunogenic compositions may comprise a first recombinant E. faecalis polypeptide antigen, a second recombinant E. faecalis polypeptide, and a third recombinant E. faecalis polypeptide. In certain embodiments, the immunogenic compositions may comprise a first recombinant E. faecalis polypeptide antigen, a second recombinant E. faecalis polypeptide, a third recombinant E. faecalis polypeptide, and a fourth recombinant E. faecalis polypeptide. In certain embodiments, the immunogenic composition may include a first recombinant E. faecalis polypeptide antigen, a second recombinant E. faecalis polypeptide, a third recombinant E. faecalis polypeptide, a fourth recombinant E. faecalis polypeptide, and a fifth recombinant E. faecalis polypeptide. In some embodiments, the immunogenic composition may include a first recombinant E. faecalis polypeptide antigen, a second recombinant E. faecalis polypeptide, a third recombinant E. faecalis polypeptide, a fourth recombinant E. faecalis polypeptide, a fifth recombinant E. faecalis polypeptide, and a sixth recombinant E. faecalis polypeptide.

[0033] In certain embodiments, the immunogenic compositions described herein comprise at least one recombinant E. faecium polypeptide antigen or a fragment thereof. For example, the immunogenic compositions may comprise a first recombinant E. faecium polypeptide antigen and a second recombinant E. faecium polypeptide. The immunogenic compositions may comprise a third, fourth, fifth, or sixth E. faecium polypeptide. Thus, in certain embodiments, the immunogenic compositions may comprise a first recombinant E. faecium polypeptide antigen, a second recombinant E. faecium polypeptide, and a third recombinant E. faecium polypeptide. In certain embodiments, the immunogenic compositions may comprise a first recombinant E. faecium polypeptide antigen, a second recombinant E. faecium polypeptide, a third recombinant E. faecium polypeptide, and a fourth recombinant E. faecium polypeptide. In certain embodiments, the immunogenic composition may comprise a first recombinant E. faecium polypeptide antigen, a second recombinant E. faecium polypeptide, a third recombinant E. faecium polypeptide, a fourth recombinant E. faecium polypeptide, and a fifth recombinant E. faecium polypeptide. In some embodiments, the immunogenic composition may comprise a first recombinant E. faecium polypeptide antigen, a second recombinant E. faecium polypeptide, a third recombinant E. faecium polypeptide, a fourth recombinant E. faecium polypeptide, a fifth recombinant E. faecium polypeptide, and a sixth recombinant E. faecium polypeptide.

[0034] In certain embodiments, the immunogenic compositions described herein comprise at least one recombinant E. durans polypeptide antigen or a fragment thereof. For example, the immunogenic compositions may comprise a first recombinant E. durans polypeptide antigen and a second recombinant E. durans polypeptide. The immunogenic compositions may comprise a third, fourth, fifth, or sixth E. durans polypeptide. Thus, in certain embodiments, the immunogenic compositions may comprise a first recombinant E. durans polypeptide antigen, a second recombinant E. durans polypeptide, and a third recombinant E. durans polypeptide. In certain embodiments, the immunogenic compositions may comprise a first recombinant E. durans polypeptide antigen, a second recombinant E. durans polypeptide, a third recombinant E. durans polypeptide, and a fourth recombinant E. durans polypeptide. In certain embodiments, the immunogenic composition may comprise a first recombinant E. durans polypeptide antigen, a second recombinant E. durans polypeptide, a third recombinant E. durans polypeptide, a fourth recombinant E. durans polypeptide, and a fifth recombinant E. durans polypeptide. In some embodiments, the immunogenic composition may comprise a first recombinant E. durans polypeptide antigen, a second recombinant E. durans polypeptide, a third recombinant E. durans polypeptide, a fourth recombinant E. durans polypeptide, a fifth recombinant E. durans polypeptide, and a sixth recombinant E. durans polypeptide.

[0035] For clarity, it should be noted that the numbering designations used herein (e.g., first, second, third, etc. recombinant polypeptide antigens) are interchangeable within a given composition. For example, in an immunogenic composition comprising a first, second, and third recombinant polypeptide antigen, where the three recombinant polypeptide antigens are AdcAII, EfaA, and EbpA, each of these three recombinant polypeptide antigens may be referred to as a first polypeptide antigen, where the remaining two are (interchangeably) the second and third.

[0036] Additionally or alternatively, the immunogenic compositions described herein may not include a second recombinant Enterococcus polypeptide antigen. For example, in some embodiments, the immunogenic compositions may not include a second recombinant E. faecalis polypeptide antigen (e.g., the immunogenic composition may include only one recombinant E. faecalis polypeptide antigen). In certain embodiments, the immunogenic composition does not include a third recombinant polypeptide antigen (e.g., the immunogenic composition may include no more than one or two polypeptide antigens). In certain embodiments, the immunogenic composition does not include a fourth recombinant polypeptide antigen (e.g., the immunogenic composition may include no more than one, two, or three polypeptide antigens). In certain embodiments, the immunogenic composition does not include a fifth recombinant polypeptide antigen (e.g., the immunogenic composition may include no more than one, two, three, or four polypeptide antigens). In some embodiments, the immunogenic composition does not include a sixth recombinant polypeptide antigen (e.g., the immunogenic composition may include no more than one, two, three, four, or five polypeptide antigens). In some embodiments, one or more of the at least one polypeptide antigen is a fragment of a polypeptide antigen.

[0037] Different combinations of recombinant polypeptide antigens have different immunogenic properties, which may allow fine-tuning the activity of the final immunogenic composition. In some embodiments, the immunogenic composition comprises a first and a second recombinant E. faecalis polypeptide antigen. In some embodiments, the first polypeptide antigen is an AdcAII polypeptide antigen and the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15, and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:15, and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:17. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0038] In some embodiments, the immunogenic composition comprises a first and a second recombinant E. faecium polypeptide antigen or fragment thereof. In some embodiments, the first polypeptide antigen is an AdcAII polypeptide antigen and the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9 and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9 and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0039] In some cases, the immunogenic composition comprises a first, a second, and a third recombinant E. faecalis polypeptide antigen. In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; the Esp polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:6 or SEQ ID NO:18, and the Ace polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:1 or SEQ ID NO:13. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:15, the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:18, and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:13. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0040] In some cases, the immunogenic composition comprises a first, a second, and a third recombinant E. faecium polypeptide antigen. In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:9; the Esp polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:12; and the Ace polypeptide antigen comprises an amino acid sequence at least 95% identical to SEQ ID NO:7. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9, the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12, and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0041] In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15, the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:15, the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:17, and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:16. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0042] In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigens.

[0043] In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:15; the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:13; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:16. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigen.

[0044] In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigens.

[0045] In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16. In certain embodiments, the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:14, the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:17, and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:16. In some embodiments, one or more of the multimeric antigens are fragments of polypeptide antigens.

[0046] In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:15. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigens.

[0047] In some cases, the immunogenic composition comprises a first, a second, a third, and a fourth recombinant E. faecalis polypeptide antigen. In some cases, the immunogenic composition comprises a first, a second, a third, and a fourth recombinant E. faecium polypeptide antigen. In some embodiments, one or more of the polypeptide antigens is a fragment of the polypeptide antigen.

[0048] In some cases, the immunogenic composition comprises a first, a second, a third, a fourth, and a fifth recombinant E. faecalis polypeptide antigen. In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:14; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13. In certain embodiments, the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 14, the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 17, the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 16, the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 18, and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 13. In some embodiments, one or more of the polypeptide antigens is a fragment of a polypeptide antigen.

[0049] In some cases, the immunogenic composition comprises a first, a second, a third, a fourth, and a fifth recombinant E. faecium polypeptide antigen. In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:8; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigens.

[0050] In some embodiments, the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13. In certain embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 15, the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 17, the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 16, the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 18, and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 13. In some embodiments, one or more of the polypeptide antigens is a fragment of a polypeptide antigen.

[0051] In some embodiments, the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen. In some embodiments, the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7. In some embodiments, one or more of the polypeptide antigens are fragments of the polypeptide antigens.

[0052] The present disclosure provides immunogenic compositions that include additional adjuvants and excipients. In some embodiments, the immunogenic compositions include one or more adjuvants to enhance the immune response to one or more of the polypeptide antigens described herein. Suitable vaccine adjuvants include, by way of example only, aluminum salts, saponins, bacteria-associated adjuvants, TLR agonists, and / or combinations thereof. Examples of aluminum salts include aluminum phosphate, aluminum hydroxide, and aluminum sulfate (e.g., alum). In some embodiments, saponins include saponin Quil A and saponin QS-21 derived from Quil A, and immune stimulating complexes (ISCOMs) formed by mixing cholesterol, phospholipids, and saponins. Bacterial-related adjuvants include, but are not limited to, cell wall peptidoglycans and lipopolysaccharides from Gram-negative bacteria such as Mycobacterium spp., Corynebacterium parvum, C. granulosum, Bordetella pertussis, Neisseria meningitis, etc., such as lipid A, monophosphoryl lipid A (MPLA), other lipid A derivatives and mimetics (such as RC529), enterobacterial lipopolysaccharide ("LPS"), TLR4 ligands, and trehalose dimycolate ("TDM"). In some embodiments, the TLR agonist adjuvant may include BCG, PAM3CSK4, MALP-2, zymosan, poly I:C, LPS, flagellin, imiquimod, resiquimod, and CpG ODN.

[0053] Excipients may be added, for example, to optimize the stability or delivery of the immunogenic compositions described herein. An excipient is an immunologically and pharmacologically inactive ingredient that is "pharmacologically acceptable." As used herein, a "pharmacologically acceptable" ingredient is one that (1) may be included in an immunogenic composition administered to a subject without causing significant undesirable biological effects or adversely interacting with any other ingredients of the formulation, and (2) meets the criteria set forth in the "Inactive Ingredients" established by the U.S. Food and Drug Administration, and is preferably also designated as "Generally Regarded as Safe" ("GRAS"). The type of excipient incorporated into the immunogenic compositions described herein depends, in part, on the selected route of administration and the particular formulation type or form, e.g., liquid formulation for injection, nasal spray formulation, etc.; routes of administration and corresponding formulations are discussed below. In general, however, inactive ingredients that can be conveniently incorporated into the immunogenic compositions described herein include, but are not limited to, vehicles, solubilizers, emulsifiers, stabilizers, preservatives, isotonicity agents, buffer systems, dispersants, diluents, viscosity modifiers, absorption enhancers, and combinations thereof. A detailed description of pharma- ceutically acceptable inactive additives is available in Gennaro (2000) Remington: The Science and Practice of Pharmacy, 20th Ed., ISBN: 0683306472.

[0054] In some embodiments, the immunogenic compositions described herein are provided in a sterile formulation for administration to a subject, e.g., a suspension, solution, or lyophilized form that is rehydrated prior to use.

[0055] Treatment method As discussed herein, the immunogenic compositions of the present disclosure may be suitable for both therapeutic and prophylactic use against Enterococcus infection. In some embodiments, the present disclosure provides a method for inducing a protective immune response against Enterococcus bacteria (e.g., E. faecalis, E. faecium, E. durans), comprising administering to a subject an immunogenic composition as described herein. In certain embodiments, the use of the immunogenic composition as described herein for inducing a protective immune response against Enterococcus bacteria in a subject is provided. In some embodiments, the use of the immunogenic composition as described herein in the manufacture of a medicament for inducing a protective immune response against Enterococcus bacteria in a subject is provided herein.

[0056] As used herein, the term "protective immune response" includes eliciting an antibody response (e.g., anti-Enterococcus) in a subject. The titer of antibodies generated following administration of an immunogenic composition described herein can be determined by means known in the art, such as an ELISA assay of serum samples from immunized subjects (see Example 4 herein). In some embodiments, the immunogenic compositions described herein elicit an antibody response in a treated subject, where the antibodies generated bind to one or more (i.e., two or more) Enterococcus peptides. For example, an immunogenic composition described herein that includes a polypeptide from one Enterococcus species (e.g., E. faecium, E. durans, or any combination of E. faecium, E. faecium, and E. durans) may generate antibodies in a subject that cross-react with corresponding peptides present in a different Enterococcus species (e.g., E. faecium, E. durans, or any combination of E. faecium, E. faecium, and E. durans). This allows the vaccines described herein to protect against or treat infections with multiple pathogenic Enterococcus species. In some embodiments, the immunogenic compositions described herein do not elicit an antibody response against human proteins or tissues.

[0057] As used herein, the term "subject" refers to a mammal. In some embodiments, the subject is a mouse, rat, dog, cat, guinea pig, rabbit, sheep, horse, cow, non-human primate, or human. In some embodiments, the subject is a human.

[0058] As mentioned above, the present specification provides methods and uses of immunogenic compositions for inducing a protective immune response against Enterococcus bacteria in a subject.These methods and uses may involve administering the immunogenic compositions therapeutically, i.e., to treat a subject suffering from Enterococcus-associated root canal defect, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal flora dysbiosis, prosthetic joint infection, or wound infection.The method includes administering the immunogenic composition prophylactically, which means, for example, that the method reduces the risk of Enterococcus-associated root canal defect, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal flora dysbiosis, prosthetic joint infection, or wound infection occurring in the subject. When the immunogenic composition is used prophylactically, the subject may be predisposed to an Enterococcus infection as a result of a number of risk factors, including but not limited to, location (e.g., hospitalization, veterinary hospital admission) or predisposing factors (e.g., previous Enterococcus infection, weakened immune system). In some embodiments, the Enterococcus bacterium is E. faecalis, E. faecium, or E. durans. In some embodiments, the Enterococcus bacterium is E. faecalis.

[0059] Also provided herein is a method and use of the immunogenic composition for treating disorders in a subject in need thereof.As described above with respect to prophylactic treatment, the subject in need thereof may suffer from, for example, root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection, or wound infection.In some embodiments, the Enterococcus bacterium is E. faecalis or E. faecium.In some embodiments, the Enterococcus bacterium is E. faecalis.In some embodiments, the Enterococcus bacterium is E. faecium.In some embodiments, the Enterococcus bacterium is E. durans.

[0060] In any of the methods, immunogenic compositions for use, or uses of the immunogenic compositions described herein, the subject may in some embodiments be at risk of or experience one or more of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, gut dysbiosis, prosthetic joint infection, or wound infection. In some of the embodiments of the methods, immunogenic compositions for use, or uses of the immunogenic compositions described herein, the risk of one or more of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, gut dysbiosis, prosthetic joint infection, or wound infection in the subject is reduced compared to before administration of the immunogenic composition.

[0061] The mode of administration largely determines the type of formulation or dosage form that contains the immunogenic composition. Compositions formulated for parenteral administration include sterile aqueous and non-aqueous solutions, suspensions, and emulsions. Aqueous solutions for injection contain the active agent in water-soluble form. Examples of non-aqueous solvents or vehicles include fatty oils such as olive oil and corn oil, synthetic fatty acid esters such as ethyl oleate or triglycerides, low molecular weight alcohols such as propylene glycol, synthetic hydrophilic polymers such as polyethylene glycol, liposomes, and the like. Parenteral formulations may include excipients such as solubilizers, emulsifiers, stabilizers, preservatives, isotonicity agents, buffer systems, dispersants, diluents, viscosity adjusters, absorption enhancers, and combinations thereof. Injectable preparations are sterilized by incorporation of bactericides, filtration through a bacteria-retaining filter, irradiation, or heat. They may also be manufactured using a sterile injectable medium. The immunogenic composition or its individual components may also be in a dried, e.g., lyophilized, form and rehydrated with a suitable vehicle immediately prior to administration by injection.

[0062] The immunogenic compositions described herein are administered to a subject according to a suitable dosing regimen. The composition may be administered once or two or more times spaced apart over an extended period of time. For example, an initial "prime" dose may be followed by at least one "boost" dose. The time interval between the prime dose and the subsequent boost dose, and between the boost doses, typically ranges from about 2 to about 24 weeks, more typically from about 2 to about 12 weeks, e.g., 2 to 8 weeks, 3 to 6 weeks, etc. Regardless of the mode of administration, e.g., intramuscular injection, intranasal administration, etc., the volume of a single dose of the vaccine generally ranges from about 1 μL to about 500 μL, typically from about 1 μL to about 250 μL, about 2.5 μL to about 200 μL, or about 5 μL to about 150 μL. It will be understood that the concentration of the total antigen in the immunogenic composition corresponds to an immunologically effective amount of the composition per unit volume, based on the dose volume guidelines set forth above.

[0063] For ease of use, the immunogenic compositions of the present invention can be incorporated into a packaged product, or "kit," that includes instructions for self-administration or administration by a medical professional. The kit includes a sealed container that contains a dose of the vaccine formulation, usually a "unit dose" suitable for an immunologically effective single-administration event. The vaccine may be in liquid form, ready to be administered, such as as an injection, or in other forms that require the user to carry out a preparation process before administration, such as hydration of a lyophilized formulation, activation of inactive ingredients, etc. The kit may include two or more sealed containers that contain a prime dose in a first container and a boost dose in one or more additional containers, or an Enterococcus vaccine formulation in a first container and a vaccine against another infectious disease that may or may not be related to Enterococcus infection in another container.

[0064] Although the present invention has been described in conjunction with a number of specific embodiments, it should be understood that the foregoing description and the following experimental section are intended to illustrate, but not to limit, the scope of the present invention. In this regard, no attempt has been made to show details of the structure of the present invention beyond the scope necessary for a fundamental understanding of the present invention, and the description in view of the drawings and / or examples will enable those skilled in the art to understand how the invention may be embodied in practice. This disclosure includes all modifications and equivalents of the subject matter described in the appended claims to the extent permitted by applicable law. Furthermore, any combination of the elements of the present invention described herein is encompassed in this disclosure unless otherwise stated herein or clearly contradicted by context.

[0065] Enumerated Embodiments Embodiment I-1. An immunogenic composition comprising at least one recombinant Enterococcus faecalis (E. faecalis) polypeptide antigen or a fragment thereof.

[0066] Embodiment I-2. The immunogenic composition of embodiment I-1, wherein at least one recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0067] Embodiment I-3. The immunogenic composition of embodiment I-1 or I-2, wherein the at least one recombinant E. faecalis polypeptide antigen comprises a first recombinant E. faecalis polypeptide antigen and a second recombinant E. faecalis polypeptide antigen.

[0068] Embodiment I-4. The immunogenic composition of embodiment I-3, wherein the first recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0069] Embodiment I-5. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen.

[0070] Embodiment I-6. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen.

[0071] Embodiment I-7. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen.

[0072] Embodiment I-8. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Ebp polypeptide antigen.

[0073] Embodiment I-9. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen.

[0074] Embodiment I-10. The immunogenic composition of any one of embodiments I-1 to I-4, wherein at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen.

[0075] Embodiment I-11. The immunogenic composition of any one of embodiments I-3 to I-10, wherein at least one recombinant E. faecalis polypeptide antigen comprises a second recombinant E. faecalis polypeptide antigen, and the second recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0076] Embodiment I-12. The immunogenic composition of any one of embodiments I-3 to I-11, wherein at least one recombinant E. faecalis polypeptide antigen comprises a third recombinant E. faecalis polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0077] Embodiment I-13. The immunogenic composition of any one of embodiments I-3 to I-12, wherein at least one recombinant E. faecalis polypeptide antigen comprises a fourth E. faecalis polypeptide antigen, and the fourth E. faecalis polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0078] Embodiment I-14. The immunogenic composition of any one of embodiments I-3 to I-13, wherein at least one recombinant E. faecalis polypeptide antigen comprises a fifth recombinant E. faecalis polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0079] Embodiment I-15. The immunogenic composition of any one of embodiments I-3 to I-14, wherein at least one recombinant E. faecalis polypeptide antigen comprises a sixth recombinant E. faecalis polypeptide antigen, and the sixth recombinant E. faecalis polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0080] Embodiment I-16. The Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13; the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:14; The AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; The EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; The immunogenic composition of any one of embodiments I-2 to I-15, wherein the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18.

[0081] Embodiment I-17. The immunogenic composition of any one of embodiments I-1 to I-16, wherein at least one recombinant E. faecalis polypeptide antigen comprises or consists of 3 to 5 recombinant E. faecalis polypeptide antigens.

[0082] Embodiment I-18. The immunogenic composition of any one of embodiments I-3 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen and the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen.

[0083] Embodiment I-19. The immunogenic composition of any one of embodiments I-13 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen.

[0084] Embodiment I-20. The immunogenic composition of any one of embodiments I-13 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen.

[0085] Embodiment I-21. The immunogenic composition of any one of embodiments I-13 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen.

[0086] Embodiment I-22. The immunogenic composition of any one of embodiments I-13 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen.

[0087] Embodiment I-23. The immunogenic composition of any one of embodiments I-15 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen.

[0088] Embodiment I-24. The immunogenic composition of any one of embodiments I-14 to I-17, wherein the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen.

[0089] Embodiment I-25. The immunogenic composition of any one of embodiments I-1, I-2, I-4 through I-15, or I-16, which does not include a second recombinant E. faecalis polypeptide antigen.

[0090] Embodiment I-26. The immunogenic composition of any one of embodiments I-3 to I-11 or I-16, which does not include a third recombinant E. faecalis polypeptide antigen.

[0091] Embodiment I-27. The immunogenic composition of any one of embodiments I-3 to I-12 or I-16 to I-22, which does not include a fourth recombinant E. faecalis polypeptide antigen.

[0092] Embodiment I-28. The immunogenic composition of any one of embodiments I-3 to I-13, or I-16 to I-22, which does not include a fifth recombinant E. faecalis polypeptide antigen.

[0093] Embodiment I-29. The immunogenic composition of any one of embodiments I-3 to I-14 or I-16 to I-24, which does not include a sixth recombinant E. faecalis polypeptide antigen.

[0094] Embodiment I-30. The immunogenic composition of any one of embodiments I-2 to I-24 or I-26 to I-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; and the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:5 or SEQ ID NO:17.

[0095] Embodiment I-31. The immunogenic composition of any one of embodiments I-2 to I-24 or I-26 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17.

[0096] Embodiment I-32. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:1 or SEQ ID NO:13.

[0097] Embodiment I-33. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13.

[0098] Embodiment I-34. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:5 or SEQ ID NO:17; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4 or SEQ ID NO:16.

[0099] Embodiment I-35. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16.

[0100] Embodiment I-36. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:1 or SEQ ID NO:13; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4 or SEQ ID NO:16.

[0101] Embodiment I-37. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16.

[0102] Embodiment I-38. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16.

[0103] Embodiment I-39. The immunogenic composition of any one of embodiments I-2 to I-24 or I-27 to I-29, wherein the AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:2 or SEQ ID NO:14; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:5 or SEQ ID NO:17; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4 or SEQ ID NO:16.

[0104] Embodiment I-40. The immunogenic composition of any one of embodiments I-2 to I-24, or I-27 to I-29, wherein the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:14; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16.

[0105] Embodiment I-41. The immunogenic composition of any one of embodiments I-2 to I-24 or I-28 to I-29, wherein the AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:2 or SEQ ID NO:14; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:1 or SEQ ID NO:13.

[0106] Embodiment I-42. The immunogenic composition of any one of embodiments I-2 to I-24 or I-28 to I-29, wherein the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:2 or SEQ ID NO:14; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13.

[0107] Embodiment I-43. The immunogenic composition of any one of embodiments I-2 to I-24 or I-28 to I-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:1 or SEQ ID NO:13.

[0108] Embodiment I-44. The immunogenic composition of any one of embodiments I-2 to I-24 or I-28 to I-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:3 or SEQ ID NO:15; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:5 or SEQ ID NO:17; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:4 or SEQ ID NO:16; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:18; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:1 or SEQ ID NO:13.

[0109] Embodiment I-45. The immunogenic composition of any one of embodiments I-1 to I-44, further comprising one or more adjuvants selected from alum, saponin, monophosphoryl lipid A (MPLA), a TLR agonist, or combinations thereof.

[0110] Embodiment I-46. The immunogenic composition of any one of embodiments I-1 to I-45, further comprising at least one excipient.

[0111] Embodiment I-47. The immunogenic composition of embodiment I-46, wherein at least one excipient is selected from vehicles, solubilizers, emulsifiers, stabilizers, preservatives, isotonicity agents, buffer systems, dispersants, diluents, viscosity modifiers, and absorption enhancers.

[0112] Embodiment I-48. The immunogenic composition of any one of embodiments I-45 to I-47, formulated as a sterile injectable solution.

[0113] Embodiment I-49. A method of inducing a protective immune response against Enterococcus bacteria in a subject comprising administering to the subject an immunogenic composition of any one of embodiments I-1 to I-48.

[0114] Embodiment I-50. The method of embodiment I-49, wherein the immunogenic composition induces an antibody response in the subject against Enterococcus bacteria and does not induce an antibody response in the subject against human tissue.

[0115] Embodiment I-51. The immunogenic composition of any one of embodiments I-1 to I-48 for use in treating a disorder in a subject in need thereof.

[0116] Embodiment I-52. The immunogenic composition of any one of embodiments I-1 to I-48, for use in treating a disorder in a subject in need thereof, wherein the disorder is associated with infection with Enterococcus bacteria.

[0117] Embodiment I-53. Use of the immunogenic composition of any one of embodiments I-1 to I-48 for inducing a protective immune response against Enterococcus bacteria in a subject.

[0118] Embodiment I-54. Use of the immunogenic composition of any one of embodiments I-1 to I-44 in the manufacture of a medicament for inducing a protective immune response against Enterococcus bacteria in a subject.

[0119] Embodiment I-55. The immunogenic composition for the method of embodiment I-49 or I-50, the use of embodiment I-51 or I-52, or the use of embodiment I-53 or I-54, wherein the Enterococcus bacterium is E. faecalis, E. faecium, or E. durans.

[0120] Embodiment I-56. The immunogenic composition for use in any one of the methods of embodiments I-49 to I-50, the use of embodiment I-51 or I-52, or the use of embodiment I-53 or I-54, wherein the Enterococcus bacterium is E. faecalis.

[0121] Embodiment I-57. The immunogenic composition for use in the method of embodiment I-49 to I-50, the use of embodiment I-51 or I-52, or the use of embodiment I-53 or I-54, wherein the subject is at risk for or experiencing root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection, or wound infection.

[0122] Embodiment I-58. The method of any one of embodiments I-49 or I-50, the immunogenic composition for use of embodiment I-51 or I-52, or the use of embodiment I-53 or I-54, in which the risk of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection, or wound infection in a subject is reduced compared to before administration of the immunogenic composition.

[0123] Embodiment II-1. An immunogenic composition comprising at least one recombinant Enterococcus faecium (E. faecium) polypeptide antigen or fragment thereof.

[0124] Embodiment II-2. The immunogenic composition of embodiment II-1, wherein at least one recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0125] Embodiment II-3. The immunogenic composition of embodiment II-1 or II-2, wherein the at least one recombinant E. faecium polypeptide antigen comprises a first recombinant E. faecium polypeptide antigen and a second recombinant E. faecium polypeptide antigen.

[0126] Embodiment II-4. The immunogenic composition of embodiment II-3, wherein the first recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0127] Embodiment II-5. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen.

[0128] Embodiment II-6. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an AdcA polypeptide antigen.

[0129] Embodiment II-7. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen.

[0130] Embodiment II-8. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an Ebp polypeptide antigen.

[0131] Embodiment II-9. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen.

[0132] Embodiment II-10. The immunogenic composition of any one of embodiments II-1 to II-4, wherein at least one recombinant E. faecium polypeptide antigen or the first recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen.

[0133] Embodiment II-11. The immunogenic composition of any one of embodiments II-3 to II-10, wherein the at least one recombinant E. faecium polypeptide antigen comprises a second recombinant E. faecium polypeptide antigen, and the second recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0134] Embodiment II-12. The immunogenic composition of any one of embodiments II-3 to II-11, wherein the at least one recombinant E. faecium polypeptide antigen comprises a third recombinant E. faecium polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0135] Embodiment II-13. The immunogenic composition of any one of embodiments II-3 to II-12, wherein the at least one recombinant E. faecium polypeptide antigen comprises a fourth E. faecium polypeptide antigen, and the fourth E. faecium polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0136] Embodiment II-14. The immunogenic composition of any one of embodiments II-3 to II-13, wherein the at least one recombinant E. faecium polypeptide antigen comprises a fifth recombinant E. faecium polypeptide antigen, and the fifth recombinant E. faecium polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0137] Embodiment II-15. The immunogenic composition of any one of embodiments II-3 to II-14, wherein the at least one recombinant E. faecium polypeptide antigen comprises a sixth recombinant E. faecium polypeptide antigen, and the sixth recombinant E. faecium polypeptide antigen is: an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen.

[0138] Embodiment II-16. The Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7; the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:8; the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; The immunogenic composition of any one of embodiments II-2 to II-15, wherein the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12.

[0139] Embodiment II-17. The immunogenic composition of any one of embodiments II-1 to II-16, wherein the at least one recombinant E. faecium polypeptide antigen comprises or consists of 3 to 5 recombinant E. faecium polypeptide antigens.

[0140] Embodiment II-18. The immunogenic composition of any one of embodiments II-3 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen and the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen.

[0141] Embodiment II-19. The immunogenic composition of any one of embodiments II-13 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen.

[0142] Embodiment II-20. The immunogenic composition of any one of embodiments II-13 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen.

[0143] Embodiment II-21. The immunogenic composition of any one of embodiments II-13 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen.

[0144] Embodiment II-22. The immunogenic composition of any one of embodiments II-13 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen.

[0145] Embodiment II-23. The immunogenic composition of any one of embodiments II-15 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen.

[0146] Embodiment II-24. The immunogenic composition of any one of embodiments II-14 to II-17, wherein the first recombinant E. faecium polypeptide antigen is an AdcAII polypeptide antigen, the second recombinant E. faecium polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecium polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecium polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecium polypeptide antigen is an Ace polypeptide antigen.

[0147] Embodiment II-25. The immunogenic composition of any one of embodiments II-1, II-2, II-4 through II-15, or II-16, which does not include a second recombinant E. faecium polypeptide antigen.

[0148] Embodiment II-26. The immunogenic composition of any one of embodiments II-3 to II-11 or II-16, which does not include a third recombinant E. faecium polypeptide antigen.

[0149] Embodiment II-27. The immunogenic composition of any one of embodiments II-3 to II-12, or II-16 to II-22, not comprising a fourth recombinant E. faecium polypeptide antigen.

[0150] Embodiment II-28. The immunogenic composition of any one of embodiments II-3 to II-13 or II-16 to II-22, not comprising a fifth recombinant E. faecium polypeptide antigen.

[0151] Embodiment II-29. The immunogenic composition of any one of embodiments II-3 to II-14 or II-16 to II-24, not comprising a sixth recombinant E. faecium polypeptide antigen.

[0152] Embodiment II-30. The immunogenic composition of any one of embodiments II-2 to II-24 or II-26 to II-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:9; and the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:11.

[0153] Embodiment II-31. An immunogenic composition of any one of embodiments II-2 to II-24 or II-26 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; and the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17.

[0154] Embodiment II-32. The immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:9; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:12; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:7.

[0155] Embodiment II-33. An immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7.

[0156] Embodiment II-34. The immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:9; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:11; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:10.

[0157] Embodiment II-35. An immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10.

[0158] Embodiment II-36. The immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:9; the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:7; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:10.

[0159] Embodiment II-37. An immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10.

[0160] Embodiment II-38. An immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10.

[0161] Embodiment II-39. The immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:8; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:11; and the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:10.

[0162] Embodiment II-40. An immunogenic composition of any one of embodiments II-2 to II-24, or II-27 to II-29, wherein the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:8; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; and the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10.

[0163] Embodiment II-41. The immunogenic composition of any one of embodiments II-2 to II-24, or II-28 to II-29, wherein the AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:8; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:11; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:10; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:12; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:7.

[0164] Embodiment II-42. An immunogenic composition of any one of embodiments II-2 to II-24 or II-28 to II-29, wherein the AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:8; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7.

[0165] Embodiment II-43. The immunogenic composition of any one of embodiments II-2 to II-24 or II-28 to II-29, wherein the AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO:9; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:11; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:10; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:12; and the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO:7.

[0166] Embodiment II-44. An immunogenic composition of any one of embodiments II-2 to II-24, or II-28 to II-29, wherein the AdcAII polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:9; the EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:11; the EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:10; the Esp polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:12; and the Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO:7.

[0167] Embodiment II-45. The immunogenic composition of any one of embodiments II-1 to II-44, further comprising one or more adjuvants selected from alum, saponin, monophosphoryl lipid A (MPLA), a TLR agonist, or combinations thereof.

[0168] Embodiment II-46. The immunogenic composition of any one of embodiments II-1 to II-45, further comprising at least one excipient.

[0169] Embodiment II-47. The immunogenic composition of embodiment II-46, wherein at least one excipient is selected from a vehicle, a solubilizer, an emulsifier, a stabilizer, a preservative, a tonicity agent, a buffer system, a dispersant, a diluent, a viscosity modifier, and an absorption enhancer.

[0170] Embodiment II-48. The immunogenic composition of any one of embodiments II-45 to II-47, formulated as a sterile injectable solution.

[0171] Embodiment II-49. A method for inducing a protective immune response against Enterococcus bacteria in a subject, comprising administering to the subject an immunogenic composition of any one of embodiments II-1 to II-48.

[0172] Embodiment II-50. The method of embodiment II-49, wherein the immunogenic composition induces an antibody response in the subject against Enterococcus bacteria and does not induce an antibody response in the subject against human tissue.

[0173] Embodiment II-51. The immunogenic composition of any one of embodiments II-1 to II-48 for use in treating a disorder in a subject in need thereof.

[0174] Embodiment II-52. The immunogenic composition of any one of embodiments II-1 to II-48 for use in treating a disorder in a subject in need thereof, wherein the disorder is associated with infection with Enterococcus bacteria.

[0175] Embodiment II-53. Use of an immunogenic composition according to any one of embodiments II-1 to II-48 for inducing a protective immune response against Enterococcus bacteria in a subject.

[0176] Embodiment II-54. Use of an immunogenic composition of any one of embodiments II-1 to II-44 in the manufacture of a medicament for inducing a protective immune response against Enterococcus bacteria in a subject.

[0177] Embodiment II-55. The immunogenic composition for the method of embodiment II-49 or II-50, the use of embodiment II-51 or II-52, or the use of embodiment II-53 or II-54, wherein the Enterococcus bacterium is E. faecalis, E. faecium, or E. durans.

[0178] Embodiment II-56. The immunogenic composition for use in any one of the methods of embodiment II-49 or II-50, embodiment II-51 or II-52, or embodiment II-53 or II-54, wherein the Enterococcus bacterium is E. faecium.

[0179] Embodiment II-57. The immunogenic composition for use in the method of embodiment II-49 to II-50, in embodiment II-51 or II-52, or in embodiment II-53 or II-54, wherein the subject is at risk for or experiencing root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection, or wound infection.

[0180] Embodiment II-58. The immunogenic composition for use of any one of the methods of embodiment II-49 or II-50, embodiment II-51 or II-52, or embodiment II-53 or II-54, wherein the risk of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal dysbiosis, prosthetic joint infection, or wound infection in a subject is reduced compared to before administration of the immunogenic composition.

[0181] Embodiment II-59. The immunogenic composition of any one of embodiments I-1 to I-48, or II-1 to II-48, the method of any one of embodiments I-49 to I-50, I-55 to I-58, II-49 to II-50, or II-55 to II-58, the immunogenic composition for use of embodiment I-51, I-52, II-51, or II-52, or the use of embodiment I-53, I-54, II-53, or II-54, wherein one or more of the at least one polypeptide antigen is a fragment of a polypeptide antigen.

[0182] Embodiment II-60. The immunogenic composition, method, immunogenic composition for use, or use of embodiment I-59 or II-59, wherein the fragment of the at least one polypeptide antigen is at least 15%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of the at least one polypeptide antigen.

[0183] Embodiment II-61. A product, process, system, kit, or use invention characterized by one or more elements disclosed in this application. EXAMPLES

[0184] Unless otherwise defined, all technical and scientific terms used herein have the meanings that are generally understood.Practitioners are particularly referred to Green & Sambrook (eds.) Molecular Cloning: A Laboratory Manual, 4th ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (2012); Ausubel et al., Current Protocols in Molecular Biology (Supplement 99) (New York: John Wiley & Sons, 2012), and Plotkin et al., Vaccines, Sixth Ed. (London: Elsevier, 2013). Examples of molecular techniques suitable for recombinant nucleic acid generation, cloning, activation and derivatization of biomolecules, purification and identification of proteins and peptides, and other related techniques are also described and / or cited in US Patent Publication No. 2018 / 0333484 [Fairman et al. (Vaxcyte, Inc.)] and PCT Publication No. WO 2014 / 172631, each of which is incorporated herein by reference. For examples of techniques and components required for parenteral administration of biomolecules as described herein, practitioners are referred to Remington, Essentials of Pharmaceutics, Pharmaceutical Press, London (2012). Methods of cell-free protein synthesis are also described in Spirin & Swartz (2008) Cell-free Protein Synthesis, Wiley-VCH, Weinheim, Germany.

[0185] Example 1: Enterococcal antigen expression screening To determine how different Enterococcal polypeptide antigens are expressed in a cell-free platform, microscale cell-free protein synthesis reactions were performed in the presence of radiolabeled leucine, the incorporation of which allows assessment of total and soluble protein expression in the reaction mixture.

[0186] Microscale expression test reactions were performed using 2% 14 Test reactions were performed with 150 μL of supermix containing C-labeled leucine, plasmid DNA of each E. faecalis polypeptide construct, cell-free E. coli extract, T7 polymerase, and water. Test reactions were performed with constructs corresponding to SEQ ID NOs: 13-18. Figure 1 shows the total average protein and average soluble protein (μg / mL) of the expressed polypeptides and the accompanying SDS-PAGE gel. Most E. faecalis polypeptide antigens expressed high amounts of soluble protein. Because the initial EbpA construct (vWA domain, 28 kD) did not express well, additional constructs were expressed (Figure 2). The EbpA construct (aa193-1532) gave better expression than the vWA domain fragment.

[0187] Example 2: Scale-up of expression and purification of Enterococcal antigens Cell-free extracts were thawed to room temperature and incubated with 50 μM iodoacetamide for 30 min as previously described (Knapp et al. 2007). OCFS reactions were carried out for up to 15 h at 25°C in 30% (v / v) iodoacetamide-treated extract containing 8 mM magnesium glutamate, 15 mM ammonium glutamate, 130 mM potassium glutamate, 35 mM sodium pyruvate, 1.2 mM AMP, 0.86 mM each of GMP, UMP, and CMP, 2 mM amino acids (1 mM tyrosine), 4 mM sodium oxalate, 1 mM putrescine, 1.5 mM spermidine, 15 mM potassium phosphate, 150 nM T7 RNAP, 2–15 μg / mL plasmid DNA template, and 1–15 μM E. coli DsbC. Reduced (GSH) and oxidized (GSSG) glutathione were added to a total concentration of 5 mM. The initial redox potential was calculated using the Nernst equation with E0 = 205 mV as the standard potential for the GSH / GSSG couple at 30 °C and pH 7.

[0188] Proteins produced at scales of 250 μL or less were monitored by incorporation of l-[U-14C]-leucine by measuring TCA-precipitable soluble and total protein in 24-well plates or Petri dishes without shaking (Voloshin and Swartz, 2005) or in 96-well plates at a 30 μL scale (with shaking). Proteins were analyzed by Sypro staining of reducing or nonreducing 12% SDS-PAGE gels and autoradiography using a PhosphoImager according to the manufacturer's recommendations.

[0189] Proteins produced at large scale were seeded evenly into 20 cm Petri dishes to form a thin layer at the bottom, and plates were incubated for 14-16 h at 25 °C. The next day, the pool was harvested at 7,500 × g for 35 min, and the supernatant was collected and adjusted to 50 mM Tris (pH 8), 150 mM NaCl, and 15 mM imidazole before being subjected to 0.22 μM filtration.

[0190] To purify the polypeptide antigens, NiCapture was used to bind His-tagged proteins. A 5 mL HisTrap Excel column was equilibrated with Buffer A (50 mM Tris pH 8, 150 mM NaCl, and 15 mM imidazole) and washed to baseline with 15 CV of Buffer A, 15 CV of Buffer A2 (+0.05% Triton X-150 added for endotoxin removal), followed by 12 CV of Buffer A to remove detergent. The polypeptide antigens were eluted using a 150% Buffer B gradient (50 mM Tris pH 8, 150 mM NaCl, and 300 mM imidazole). Fractions corresponding to the elution peak were pooled and the TeV sequence was cleaved using 1:15 TEV protease:protein. Dialysis was performed overnight in 1x TBS. Samples were collected and passed over 1 mL of HisTrap with 1x TBS as buffer A and 300 mM imidazole as B1. The flow-through was collected and concentrated using an Amicon with a 15 kD cut-off. SDS-PAGE gels were run with the washed and eluted flow-through from the first Akta purification and again after TeV cleavage, and also stained with Coomassie blue to confirm size and purity as well as confirm TeV cleavage. Endotoxin levels were tested using the Charles River Endosafe Nexgen-PTS150 Endotoxin Test Detection System. Aliquots of purified polypeptide antigen were prepared, labeled, and stored at -80°C.

[0191] Table 2 shows the results of expression / purification and TEV cleavage at 150 mL scale for selected E. faecalis polypeptide antigens.

[0192] [Table 2]

[0193] Figures 3-8 show gels of an exemplary E. faecalis polypeptide antigen after expression / purification and TEV cleavage, respectively. The Esp polypeptide antigen of SEQ ID NO: 18 was found to be unstable during multiple freeze / thaw cycles. Adjusting the storage buffer to 1x TBS with 15% glycerol increased the stability of the polypeptide.

[0194] Multi-angle light scattering coupled with size exclusion chromatography (SEC-MALS) was used to confirm the molecular weight of the isolated exemplary polypeptide antigens. Figure 9 shows a chromatograph for the SEC-MALS molecular weight analysis of the polypeptide antigens of SEQ ID NOs: 12-18.

[0195] Final confirmation of molecular weight for the polypeptide antigens used in the animal studies was obtained by SEC-MALS. Figures 15A and 15B show the SDS-PAGE and SEC-MALS data, respectively, of this polypeptide antigen expression batch. Samples of each polypeptide antigen were also subjected to a stability test involving three freeze-thaw cycles. Figures 11A-11F show the SEC-MALS data after freeze-thaw cycles for E. faecalis AdcA, AdcAII, Ace, Esp, EfaA, and EbpA samples, respectively. Figure 11A shows that AdcA is relatively stable, with only a slight decrease in the intensity and molecular weight of the main peak. Figure 11B shows that AdcAII is also relatively stable, with only a slight decrease in the intensity of the main peak and a slight increase in molecular weight. Ace showed a consistent SEC-MALS peak (Figure 11C). Figure 11D shows the results of a stability test with two different Esp samples. As previously described, and shown here in Figure 11 D, Esp was not stable until 15% glycerol was added to the storage buffer. Figures 11 E and 11 F show that EfaA and EpbA, respectively, are relatively stable to freeze-thaw cycles.

[0196] Example 3: Immunization studies in CD-1 mice with individual exemplary E. faecalis immunogenic polypeptide antigens Seven groups of 15 female CD-1 mice each were administered 5 μg of each antigen (EfaA, Ace, Esp, AdcA, AdcAII, and alum control) on days 0, 7, and 14. A terminal bleed was performed on day 21, and serum was analyzed.

[0197] Immunoblotting was performed to analyze antigen specificity. Polypeptides are loaded and run on SDS gels and transferred to PVDF membranes. They are then probed with recovered mouse serum containing antisera (primary antibodies against each antigen), which then bind to the proteins on the PVDF membrane if present. Protein bands can be visualized with goat anti-mouse serum conjugated with horseradish peroxidase (secondary antibody). Figure 12A shows that mouse antiserum raised against AdcA antigen detects the presence of AdcA protein in cell lysates generated from wild-type E. faecalis OG1RF grown in Blood Heart Infusion broth, although in lane 1: "Wt lysate (BHI)", the band elutes slightly higher than the expected Mw of AdcA of 35 kDa. Lane 2: "ΔAdcACB / ΔAdcA lysate" probes cell lysates generated from the ΔAdcACB / ΔAdcA null mutant strain of E. faecalis OG1RF and shows the absence of AdcA protein as expected. The last two lanes are 0.25 μg of each antigen used to immunize mice. Lane 3: "AdcA 0.25 μg" shows that the AdcA antigen is detected by the antiserum and that several fragments are present. Lane 4: "AdcAII 0.25 μg" shows little or no cross-reaction of the AdcA antiserum with the AdcAII antigen. FIG. 12B shows that in lane 1: "Wt lysate (BHI)", mouse antisera raised against the AdcAII antigen detects the presence of AdcAII protein in cell lysates generated from wild-type E. faecalis OG1RF grown in Blood Heart Infusion broth, although the band elutes slightly higher than the expected Mw of AdcAII of 57 kDa. Lane 2: "ΔAdcACB ΔAdcA lysate" probes cell lysates generated from a ΔAdcACB / ΔAdcA null mutant strain of E. faecalis OG1RF, and interestingly detects some presence of AdcA protein, suggesting that the AdcAII antisera cross-reacts with AdcA. The last two lanes are 0.25 μg of each antigen used to immunize mice.Lane 3: "AdcA 0.25 μg" shows that some of the AdcA antigen bands are detected by the AdcAII antiserum, indicating cross-reactivity. Lane 4: "AdcAII 0.25 μg" shows that the AdcAII antigen is detected by the AdcAII antiserum, as expected. Overall, the Western immunoblots show that the expression of antigens by E. faecalis OGRF1 is conditional, that antisera raised against the AdcA antigen are not cross-reactive to the AdcAII antigen, whereas antisera raised against the AdcAII antigen are cross-reactive to the AdcA antigen.

[0198] The G. mellonella immunization model was used to analyze the neutralization and opsonization ability of mouse antisera to modulate the virulence of E. faecalis. Details of this model are described, for example, in Lam, et al. Bacterial Virulence, Methods in Mol. Bio. 2021. Figure 13 shows the survival rate of G. mellonella after treatment with rabbit antisera from each treatment arm. After 72 hours, only about 15% were alive in the WT group. However, in each antiserum arm, the survival rate increased to more than about 90% after 72 hours.

[0199] Example 4: Immunization studies in rabbits with E. faecalis immunogenic compositions New Zealand White rabbits were administered either alum control, heat-killed E. faecalis, or one of the eight immunogenic compositions described herein (according to the schedule in FIG. 14A). The immunogenic compositions tested are shown in FIG. 14B. For each immunogenic composition, rabbits were administered 15 μg per constituent polypeptide antigen.

[0200] Antigen-specific IgG ELISA was used to assess changes in IgG levels following immunization. Figure 15 shows that antisera from animals administered individual E. faecalis polypeptide antigens elevate antigen-specific IgG titers compared to pre-immunized animals and alum controls.

[0201] Example 5: Immunization studies in rabbits with exemplary E. faecalis immunogenic compositions An opsonophagocytic assay (see, e.g., Nahm, et. al., 2014, Protocol for multiplexed opsonophagocytic killing assay (UAB-MOPA) for antibodies against Streptococcus pneumoniae) was used to evaluate the antibody-dependent killing effect by neutrophils after immunization. Figure 16 shows an overview of the opsonophagocytic (OPA) assay. Results at the start of incubation and after 90 minutes of incubation show that opsonophagocytic activity was observed when antisera were added to the bacteria, suggesting that these antigens may be useful as vaccine antigens. Figure 17 shows that antiserum antibodies have an opsonizing / neutralizing effect on neutrophil-mediated killing of E. faecalis in vitro.

[0202] Example 6: Passive and active immunization studies using a mouse model of peritonitis infection To determine the protective efficiency of antisera generated against E. faecalis antigens, a passive / active immunization study was performed in the laboratory of Dr. Jose Lemos at the University of Florida. Figure 18 shows an overview of the mouse peritonitis model used to determine the protective efficiency of different antisera generated against E. faecalis. Briefly, on day 1, groups (N=3 mice per group) of healthy mice (one group for each antigen investigated) are primary vaccinated with the different mouse antisera generated (3 PBS-only negative controls, prebleed serum, and alum-only final bleed serum) as well as 7 final bleeds from the antigens under test (EfaA, AdcA, AdcAII, EbpA, Asa, Ace, Esp). On day 2, individual mouse groups are boosted with the same antisera as on day 1. On day 3, mice are injected intraperitoneally with E. faecalis OG1RF (158 cells / mL). On day 4, the mice groups received a second booster immunization, and on day 5, the mice were sacrificed, various tissues (spleen and peritoneal cavity) were washed, and the wash solutions were collected and transferred to BHI plates to examine colony forming units. The data in Figure 19 show that, compared to the three controls, passive immunization with anti-EfaA or anti-AdcAII antisera significantly reduced (approximately 5-fold) the E. faecalis CFUs recovered from the spleen, whereas mice immunized with EfaA or AdcA specific antisera had reduced bacteria recovered from the peritoneal cavity (1 log and 5-fold, respectively).

[0203] Example 7: Passive immunization study using a mouse model of peritonitis infection A passive immunization study was performed to determine the protective efficiency of antisera generated against E. faecalis antigen combinations according to the procedure of Example 6. Kidneys of test subjects were analyzed for colony forming units. Figure 20 shows the results of this study, showing that antisera from both (AdcAII+AdcA+EfA) and (AdcAII+EfaA+EbpA+Esp+Ace) combinations significantly reduced CFUs of E. faecalis recovered from kidneys. Furthermore, final bleed antigen-specific IgG titers were obtained according to the above protocol. Figures 21A, 21B, and 21C show the final bleed IgG titers of the 10 treatment groups (groups 1-15). All enterococcal peptide combinations show significant IgG titers compared to the alum control (group 1) and heat-killed E. faecalis (group 2).

[0204] Figure 22 shows an overview of the mouse peritonitis model used to determine the protective efficacy of various antisera generated against E. faecalis, which is the same as the procedure described in Example 6, except that the second boost was not administered on day 4 of the study. Figures 23A and 23B show the protective efficacy of 2.02 x 15 15 CFU / mL of OG1RF strain and 2.2 × 15 9 Survival data for animals inoculated with OG1RF strain in CFU / mL are shown. 9 For animals dosed at CFU / mL, both combinations (AdcAII+AdcA+EfaA) and (AdcAII+EfA+EbpA+Esp+Ace) resulted in a statistically significant increase in survival versus controls at 72 hours post-inoculation.

[0205] Survival studies using additional antigen combinations (Figures 23C, 23D, 23E, 23F, 23G, 23H) were also performed and demonstrated a statistically significant increase in overall survival of subjects in each experimental group.

[0206] Example 8: Efficacy of E. faecalis OG1RF antigen-specific antisera against other clinical enterococcal strains The efficacy of antigen combinations in other clinically relevant Enterococcus strains was determined in a similar manner as described above. Figure 24A (E. faecalis strains) and Figure 24B (E. faecium) show results for E. faecalis HM201 (blood isolate; VanR), E. faecalis HM202 (fecal isolate; KanR), E. faecalis NR31975 (blood isolate; ErmR GenR), E. faecalis NR31979 (urine isolate; VanR), E. faecium HM952 (normal flora), E. faecium NR31903 (fecal isolate; clone of CC17 that is AmpR, carry esp, hyl, acm), E. faecium NR31909 (fecal isolate; clone of CC17), E. faecium NR31912 (fecal isolate; CC17; VanR) strains obtained from the NIH BEI Resources Repository. E. faecium strains, like E. faecalis strains, showed killing efficiency against the antigen combinations tested, indicating that the combinations described herein are capable of treating infections with multiple Enterococcus species and strains.

[0207] FIG. 24C shows a summary of colony forming unit (CFU) data generated from the mouse peritoneal (peritonitis) infection model (described above) for E. faecalis HM201, E. faecalis NR31979, E. faecium NR31903, and E. faecium NR31912.

Claims

1. An immunogenic composition comprising at least one recombinant Enterococcus faecalis (E. faecalis) polypeptide antigen or a fragment thereof, wherein the at least one recombinant E. faecalis polypeptide antigen is (a) Contains the first recombinant E. faecalis polypeptide antigen; (b) containing a first recombinant E. faecalis polypeptide antigen and a second recombinant E. faecalis polypeptide antigen; (c) containing a first recombinant E. faecalis polypeptide antigen, a second recombinant E. faecalis polypeptide antigen, and a third recombinant E. faecalis polypeptide antigen; (d) containing the first recombinant E. faecalis polypeptide antigen, the second recombinant E. faecalis polypeptide antigen, the third recombinant E. faecalis polypeptide antigen, and the fourth recombinant E. faecalis polypeptide antigen; (e) comprising the first recombinant E. faecalis polypeptide antigen, the second recombinant E. faecalis polypeptide antigen, the third recombinant E. faecalis polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen and the fifth recombinant E. faecalis polypeptide antigen; or (f) comprising the first recombinant E. faecalis polypeptide antigen, the second recombinant E. faecalis polypeptide antigen, the third recombinant E. faecalis polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen, the fifth recombinant E. faecalis polypeptide antigen and the sixth recombinant E. faecalis polypeptide antigen, The immunogenic composition.

2. (a) The at least one recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an EbpA polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (b) The first recombinant E. faecalis polypeptide antigen is either an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (c) The at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen; (d) Whether the at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen; (e) The at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an AdcAII polypeptide antigen; (f) Whether the at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Ebp polypeptide antigen; (g) The at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen; (h) The at least one recombinant E. faecalis polypeptide antigen or the first recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen; (i) The at least one recombinant E. faecalis polypeptide antigen comprises a second recombinant E. faecalis polypeptide antigen, wherein the second recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (j) The at least one recombinant E. faecalis polypeptide antigen comprises a third recombinant E. faecalis polypeptide antigen, wherein the third recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (k) The at least one recombinant E. faecalis polypeptide antigen comprises a fourth recombinant E. faecalis polypeptide antigen, wherein the fourth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (l) The at least one recombinant E. faecalis polypeptide antigen comprises a fifth recombinant E. faecalis polypeptide antigen, wherein the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen, an AdcA polypeptide antigen, an AdcAII polypeptide antigen, an Ebp polypeptide antigen, an EfaA polypeptide antigen, or an Esp polypeptide antigen; (m) The at least one recombinant E. faecalis polypeptide antigen comprises a sixth recombinant E. faecalis polypeptide antigen, wherein the sixth recombinant E. faecalis polypeptide antigen is: Ace polypeptide antigen, AdcA polypeptide antigen, AdcAII polypeptide antigen, Ebp polypeptide antigen, EfaA polypeptide antigen, or Esp polypeptide antigen; (n) The first recombinant E. faecalis polypeptide antigen is AdcAII polypeptide antigen and the second recombinant E. faecalis polypeptide antigen is EfaA polypeptide antigen; (o) The first recombinant E. faecalis polypeptide antigen is the AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is the Esp polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is the Ace polypeptide antigen; (p) Whether the first recombinant E. faecalis polypeptide antigen is AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is EbpA polypeptide antigen; (q) Whether the first recombinant E. faecalis polypeptide antigen is AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is Ace polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is EbpA polypeptide antigen; (r) The first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, and the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen; (s) The first recombinant E. faecalis polypeptide antigen is an AdcA polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is an EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is an EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is an Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is an Ace polypeptide antigen; or (t) The first recombinant E. faecalis polypeptide antigen is AdcAII polypeptide antigen, the second recombinant E. faecalis polypeptide antigen is EfaA polypeptide antigen, the third recombinant E. faecalis polypeptide antigen is EbpA polypeptide antigen, the fourth recombinant E. faecalis polypeptide antigen is Esp polypeptide antigen, and the fifth recombinant E. faecalis polypeptide antigen is Ace polypeptide antigen. The immunogenic composition according to claim 1.

3. The Ace polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 13; The AdcA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 14; The AdcAII polypeptide antigen contains or comprises the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; The EbpA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; The EfaA polypeptide antigen comprises or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; The Esp polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 6 or SEQ ID NO:

18. The immunogenic composition according to claim 2.

4. The immunogenic composition according to claim 1, wherein the at least one recombinant E. faecalis polypeptide antigen comprises or consists of three to five recombinant E. faecalis polypeptide antigens.

5. (a) Not containing a second recombinant E. faecalis polypeptide antigen; (b) Does not contain a third recombinant E. faecalis polypeptide antigen; (c) Fourth recombinant E. faecalis polypeptide antigen is not present; (d) Not containing the fifth recombinant E. faecalis polypeptide antigen; or (e) Recombinant E. 6, which does not contain the Enterococcus faecalis polypeptide antigen. The immunogenic composition according to claim 1.

6. (a) The AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 5 or SEQ ID NO: 17; (b) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; (c) The AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 or SEQ ID NO: 15; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 6 or SEQ ID NO: 18; or the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 1 or SEQ ID NO: 13; (d) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the Esp polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 6 or SEQ ID NO: 18; the Ace polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 13; (e) The AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 5 or SEQ ID NO: 17; or the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4 or SEQ ID NO: 16; (f) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; (g) The AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 or SEQ ID NO: 15; the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 1 or SEQ ID NO: 13; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4 or SEQ ID NO: 16; (h) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the Ace polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 13; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; (i) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; (j) The AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 2 or SEQ ID NO: 14; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 5 or SEQ ID NO: 17; or the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4 or SEQ ID NO: 16; (k) The AdcA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 14; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; (l) The AdcA polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 2 or SEQ ID NO: 14; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4 or SEQ ID NO: 16; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 6 or SEQ ID NO: 18; or the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 1 or SEQ ID NO: 13; (m) The AdcA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 14; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; the Esp polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 6 or SEQ ID NO: 18; the Ace polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 13; (n) The AdcAII polypeptide antigen comprises an amino acid sequence that is at least 85%, at least 90%, or at least 95% identical to SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 4 or SEQ ID NO: 16; the Esp polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 6 or SEQ ID NO: 18; the Ace polypeptide antigen comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 1 or SEQ ID NO: 13; or (o) The AdcAII polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 3 or SEQ ID NO: 15; the EfaA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 5 or SEQ ID NO: 17; the EbpA polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 4 or SEQ ID NO: 16; the Esp polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 6 or SEQ ID NO: 18; the Ace polypeptide antigen contains or consists of the amino acid sequence of SEQ ID NO: 1 or SEQ ID NO: 13 The immunogenic composition according to claim 2.

7. (a) further comprising one or more adjuvants selected from alum, saponin, monophosphoryl lipid A (MPLA), TLR agonist, or a combination thereof; and / or (b) further comprising at least one excipient, wherein, if required, the at least one excipient is selected from a medium, a solubilizer, an emulsifier, a stabilizer, a preservative, an isotonic agent, a buffer system, a dispersant, a diluent, a viscosity modifier, and an absorption enhancer. The immunogenic composition according to claim 1, wherein the immunogenic composition is formulated as a sterile injection solution if necessary.

8. The immunogenic composition according to claim 1, wherein one or more of the at least one polypeptide antigens are fragments of a polypeptide antigen, and if required, the fragments of the polypeptide antigen comprise at least 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, or 95% of the length of the at least one polypeptide antigen.

9. A pharmaceutical composition comprising the immunogenic composition according to claim 1 for inducing a protective immune response against Enterococcus bacteria in a subject, In short, the immunogenic composition induces an antibody response against Enterococcus bacteria in the subject, but does not induce an antibody response against human tissue in the subject; In particular, the Enterococcus bacterium is E. faecalis, E. faecium, or E. durans; in particular, the Enterococcus bacterium is E. faecalis; In short, the subject is at risk of or has experienced root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, enterobacteriamal abnormalities, prosthesis infection, or wound infection; and In short, compared to before administration of the immunogenic composition, the risk of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal flora abnormalities, prosthesis infection, or wound infection in the subject is reduced. The aforementioned pharmaceutical composition.

10. (a) for the treatment of a disorder in the subject, (b) A pharmaceutical composition comprising the immunogenic composition according to claim 1 for the treatment of disorders related to infections caused by Enterococcus bacteria in a subject, In particular, the Enterococcus bacterium is E. faecalis, E. faecium, or E. durans; in particular, the Enterococcus bacterium is E. faecalis; In short, the subject is at risk of or has experienced root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, enterobacteriamal abnormalities, prosthesis infection, or wound infection; and In short, compared to before administration of the immunogenic composition, the risk of root canal failure, endocarditis, bacteremia, urinary tract infection, prostatitis, intraperitoneal infection, cellulitis, intestinal flora abnormalities, prosthesis infection, or wound infection in the subject is reduced. The aforementioned pharmaceutical composition.

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