A klebsiella pneumoniae vaccine

Abstract

A Klebsiella pneumoniae vaccine. A composition comprising one or more immunogens is provided. The composition is for use in vaccine therapy to treat or prevent K. pneumoniae infection in subject.

Description

[0001] Title of the invention

[0002] A Klebsiella pneumoniae vaccine.

[0003] Field of the Invention

[0004] This invention relates to a vaccine for the prevention or treatment of Klebsiella pneumoniae infection in a subject.

[0005] Background of the Invention

[0006] Klebsiella pneumoniae is a Gram-negative encapsulated bacterium that causes septicaemia, respiratory tract infections, urinary tract infections, and soft tissue infections, by colonising the mucosa.

[0007] K. pneumoniae is a major opportunistic pathogen and a leading cause of hospital acquired infections. It has been identified as one of the ESKAPE pathogens, i.e. a group associated with some of the deadliest hospital acquired infections across the world. It is responsible for outbreaks in closed hospital units, especially in paediatric (neonatal) units and a common cause of infections in spinal injury centres. It also causes outbreaks in long-term acute care hospitals and nursing homes.

[0008] K. pneumoniae is prevalent among nosocomial infections and is described as the fastest growing antibiotic resistant threat in Europe. It possesses several mechanisms of antibiotic resistance including beta lactamases, extended spectrum beta lactamases (EBSL), carbapenemases (KPC). It is resistant to third-generation cephalosporins, carbapenems, and colistins, with 44% of European countries reporting that 50% of isolates are resistantto these antibiotics. Recently, K. pneumoniae strains with resistance to colistin, one of the antibiotics of last resort were identified.

[0009] K. pneumoniae is associated with high mortality rates globally with 33%, 50% and 44% reported for North America, Europe and Asia, respectively. It is more prevalent in Latin America and Asia-Pacific regions than in North America and Europe It was the third most commonly reported bacterial species in 2021 , accounting for 11 .9 % of all reported cases in the EU / EEA (ECDC Annual epidemiology report 2021 ).

[0010] There are two pathotypes of K. pneumoniae, classical K. pneumoniae and hypervirulent K pneumoniae. Classical K. pneumoniae is one of the most commonly acquired opportunistic pathogens and is more likely to cause healthcare-acquired infections in hosts who are immunocompromised or with co-morbidities. In contrast to classical K. pneumoniae, hypervirulent infections are not limited to immune compromised individuals but can cause severe infections among a wider population, especially healthy people. It has the ability to infect healthy individuals of all ages, and infected patients have a tendency to present with multiple sites of infection and / or develop subsequent metastaticspread, which is very unusual in other Enterobacteriaceae. Hypervirulent . pneumoniae has caused a significant number of community acquired infections worldwide.

[0011] The first line treatment for K. pneumoniae infections are -lactams and other antibiotics. However, the increasing level of antibiotic-resistant K. pneumoniae and hypervirulent K. pneumoniae strains (Choby, JE et al., J Internal Medicine (2020) 287, 283-300) with antimicrobial resistance associated with extended -spectrum p-lactamases have emerged separately across the world and are causing an increasingly serious problem. K. pneumoniae has a high fatality rate in patients with chronicobstructive pulmonary disease (COPD) patients: up to 50% patients with antimicrobial resistant strains die. The emergence of hypervirulent carbapenem-resistant strains present an additional threat with a potential for global dissemination.

[0012] Unfortunately, there are no approved vaccines available to prevent K. pneumoniae infection. Various vaccine technologies have been tested in preclinical trials, including outer membrane vesicles (Lee, W.H., et al., Vaccination with Klebsiella pneumoniae-derived extracellular vesicles protects against bacteria-induced lethality via both humoral and cellular immunity. Experimental & Molecular Medicine, 2015. 47(9): p. e183-e183), lipopolysaccharide (LPS) (Yadav V et al. ,(2005) Folia microbiologica 50, 83-86), capsular polysaccharides (Feldman, M.F., et al., A promising bioconjugate vaccine against hypervirulent Klebsiella pneumoniae. Proc Natl Acad Sci U S A, 2019. 116(37): p. 18655-18663) and subunit proteins (Assoni et al., Infectious diseases and Therapy 10 (4)2157 -21752021), but to date, each approach presents limitations in terms of toxicity, variability and / or cross-reactivity with host microbiota. In addition, many candidates have shown limited T cell or other cellular responses, which are important for protection. Examples of tested subunit vaccines include OmpA, Ompk36 and OmpK17, but none have progressed to clinical trial.

[0013] Visweswaran GR, et al., (Exploiting the peptidoglycan-binding motif, LysM, for medical and industrial applications. Appl Microbiol Biotechnol.2014 May;98(10):4331 -45) is a review article that discusses the lysin motif (LysM) and the characteristics and possible use of LysM in various applications. This review does not mention Klebsiella pneumoniae.

[0014] There is a real clinical need to identify new vaccine antigens, particularly due to the dramatic increase in microbial resistance and incidence of this infection.

[0015] The current invention serves to address the problems of the prior art. Summary of the Invention

[0016] The inventors have identified a series of K. pneumoniae proteins for use as antigens in a vaccine that can prevent or treat serious infection of subjects with K. pneumoniae (“proteins of the invention”). The use of multiple antigens widens the protective response against bacterial infection amongst a greater range of strains of the bacterium.

[0017] The inventors are the first to discover that these proteins were involved in the attachment of K. pneumoniae to host cells, in particular to human lung epithelial cells. For example, Figure 7 confirms that LysM, 0mpP1 and DPS are all involved in host cell attachment. In addition, the inventors discovered that LysM, 0mpP1 and DPS are protective in a sepsis mouse model (Figure 9), reducing both bacterial colonisation and dissemination to the spleen (indicative of invasive disease).

[0018] The proteins of the invention, or vaccine antigens, are listed below in Table 1.

[0019] Table 1 : Proteins of the Invention. Accordingly, in an aspect of the current invention, there is provided a composition comprising one or more immunogens of the invention (herein “composition of the invention”).

[0020] The immunogens of the invention may be one or more proteins of the invention, or an immunogenic portion of the protein of the invention.

[0021] In an embodiment, the composition comprises a nucleotide or mRNA encoding the immunogen of the invention.

[0022] In an embodiment, the immunogenic portion is a polypeptide or a peptide.

[0023] The immunogen may be a conjugate comprising one or more immunogens of the invention conjugated, linked or otherwise fused.

[0024] In an embodiment, the composition of the invention is an immunogenic composition.

[0025] In an embodiment, the composition of the invention is a vaccine.

[0026] In an embodiment, the vaccine of the invention is an adenoviral vector vaccine.

[0027] In an embodiment, the vaccine is a mRNA vaccine.

[0028] In an embodiment, the vaccine is a liposome-based vaccine.

[0029] The composition may optionally include one or more adjuvants.

[0030] The composition may optionally include one or more additional antigens.

[0031] The composition of the invention may be a pharmaceutical composition and optionally comprising one or more pharmaceutically acceptable excipients.

[0032] Preferably, the immunogen is present in the composition of the invention in a therapeutically effective amount.

[0033] It will be appreciated that the one or more immunogens in the composition of the invention may be any combination of the proteins of the invention, or immunogenic portions thereof.

[0034] In an embodiment, the composition comprises all the proteins of the invention.

[0035] In an embodiment, the one or more immunogen comprises LysM / BON. In an embodiment, the only immunogen present in the composition is LysM / BON.

[0036] In an embodiment, the one or more immunogen comprisesOmpPI . In an embodiment, the only immunogen in the composition is OmpP1.

[0037] In an embodiment, the one or more immunogen comprises DPS. In an embodiment, the only immunogen in the composition is DPS. In an embodiment, the one or more immunogen comprises LysM / BON and OmpP1 . In an embodiment, the only immunogens in the composition are LysM / BON and OmpP1.

[0038] In an embodiment, the one or more immunogen comprises LysM / BON and DPS. In an embodiment, the only immunogens in the composition are LysM / BON and DPS.

[0039] In an embodiment, the one or more immunogen comprises OmpP1 and DPS. In an embodiment, the only immunogens in the composition are OmpP1 and DPS.

[0040] In an embodiment, the one or more immunogen comprise LysM / BON, OmpP1 and DPS. In an embodiment, the composition comprises these three immunogens only.

[0041] In any embodiment, the composition comprises all of the immunogens of the invention.

[0042] An aspect of the invention provides an immunogen of the invention (ora nucleotide or mRNA encoding the same) , for use in a vaccine therapy to preventortreat infection by K. pneumoniae in a subject.

[0043] An aspect of the invention provides the composition of the invention for use in a vaccine therapy to prevent or treat infection by K. pneumoniae in a subject.

[0044] An aspect of the invention provides an immunogen of the invention (ora nucleotide or mRNA encoding the same), for use as a therapy to induce an immune response against K. pneumoniae in a subject.

[0045] The invention also provides a method of treating or preventing infection by K. pneumoniae in a subject, the method comprising a step of administering a therapeutically effective amount of one or more immunogens of the invention (or a nucleotide or mRNA encoding the same) to the subject.

[0046] The invention also provides a method of treating or preventing infection by K. pneumoniae in a subject, comprising administering the composition of the invention to the subject.

[0047] An embodiment of the current invention includes the use of an antibody directed or raised against, to one or more immunogens of the invention, which are suitable for immunotherapy for use to treat infection by K. pneumoniae. The antibody is typically a monoclonal antibody specific to the immunogen of the invention

[0048] The invention also provides a method of treating K. pneumoniae infection in a subject, the method comprising a step of administering an antibody against one or more immunogens of the invention to the subject.

[0049] In an embodiment of the above aspects, the antibody is a monoclonal antibody. In an embodiment, the monoclonal antibody is one or more of anti-LysM, anti-DPS or anti- OmpP1.

[0050] In an embodiment of any aspect of the invention, the subject is human.

[0051] The K. pneumoniae may be hypervirulent K. pneumoniae (hvKp). This may occur in any subject, including an otherwise healthy subject, .e.g. a subject who does not have an underlying health condition, or has not critical ill or admitted to hospital.

[0052] The K. pneumoniae may be antibiotic-resistant K. pneumoniae.

[0053] In an aspect of the invention concerned with prevention, the subject may be one at risk of an infection with K. pneumoniae.

[0054] In an embodiment, the subject is one admitted in a hospital unit, including a paediatric (neonatal) unit.

[0055] In an embodiment, the subject is a subject that is critically ill, such as a subject in an intensive care unit (ICU). Subjects on ventilators, or subjects with prolonged use of invasive medical devices are also a target.

[0056] In an embodiment, the subject is one admitted in a spinal injury centre.

[0057] In an embodiment, the subject may be one with community acquired pneumonia, or a urinary tract infection.

[0058] In an embodiment, the subject is one with any nosocomial infection.

[0059] In an embodiment, the subject may be one with an underlying condition such as but not limited to diabetes, chronic alcohol use and prolonged use of antibiotics.

[0060] It will be appreciated that prevention or treatment may take place in subjects listed.

[0061] It will also be appreciated that the subject may be any subject, e.g. any otherwise healthy subject.

[0062] Definitions

[0063] Where used herein and unless specifically indicated otherwise, the following terms are intended to have the following meanings in addition to any broader (or narrower) meanings the terms might enjoy in the art:

[0064] Unless otherwise required by context, the use herein of the singular is to be read to include the plural and vice versa. The term "a" or "an" used in relation to an entity is to be read to refer to one or more of that entity. As such, the terms "a" (or "an"), "one or more," and "at least one" are used interchangeably herein. As used herein, the term "comprise," orvariationsthereof such as "com prises" or "com prising," are to be read to indicate the inclusion of any recited integer (e.g. a feature, element, characteristic, property, method / process step or limitation) or group of integers (e.g. features, element, characteristics, properties, method / process stepsorlimitations) but not the exclusion of any other integer or group of integers. Thus, as used herein the term "comprising" is inclusive or open-ended and does not exclude additional, unrecited integers or method / process steps.

[0065] As used herein, the term “disease” is used to define any abnormal condition that impairs physiological function and is associated with specific symptoms. The term is used broadly to encompass any disorder, illness, abnormality, pathology, sickness, condition, or syndrome in which physiological function is impaired irrespective of the nature of the aetiology (or indeed whether the aetiological basis for the disease is established). It therefore encompasses conditions arising from infection, trauma, injury, surgery, radiologica l ablation, poisoning, or nutritional deficiencies.

[0066] As used herein, the term "treatment" or "treating" refer to an intervention (e.g. the administration of an agentto a subject) which cures, ameliorates or lessens the symptoms of a disease or removes (or lessens the impact of) its cause(s). In this case, the term is used synonymously with the term “therapy”. It can be manifested by a perman ent or temporary improvement in the subject's condition. In this context, it includes limiting and / or reversing disease progression.

[0067] As used herein the terms "prevention" or "preventing" refers to an intervention (e.g., the administration of an agent or vaccine to a subject), which prevents or delays the onset or progression of a disease or infection, e.g., K. pneumoniae infection, or the severity of a disease in a subject, or reduces (or eradicates) its incidence within a treated population.

[0068] When used herein, the term “composition” should be understood to mean something made by the hand of man, and not including naturally occurring compositions. Compositions may be formulated in unit dosage form, i.e. , in the form of discrete portions containing a unit dose, or a multiple or sub-unit of a unit dose.

[0069] In this specification, the term “sequence identity” should be understood to mean the amount of nucleic acids or amino acids which match between different sequences. For example, a sequence, such as a polypeptide or a polynucleotide sequence, that shares 90% sequence identity with a sequence is one in which any 90% of aligned residues are identical to the corresponding nucleic acids or amino acids in the reference sequence across the entire length of the sequence. Sequence homology can be calculated by a BLAST program (www.ncbi.nlm.nih.gov / cgi-bin Blast). Hereby, gaps are not counted. The term subject (which is to be read to include "individual", "animal", "patient" or "mammal" where the context permits) defines any subject, particularly a mammalian subject, for whom treatment or prevention is indicated. Mammal is a human or an animal, typically a human. In many cases, the subject will be a mammal which has, or which is suspected of having, a Klebsiella pneumoniae infection, or at risk of having a Klebsiella pneumoniae infection.

[0070] The term “immunogen” is a substance that is capable of generating a B-cell and / or T-cell immune response in a host organism. In this context, the immunogen may be a protein or an immunogenic portion of the protein such as an isolated polypeptide, or peptid e. In an embodiment, the immunogens is purified, recombinant or synthetic.

[0071] In this specification, the term “immunogenic portion thereof” as applied to any of the proteins of the invention should be understood to mean a portion of the protein that when administered to a mammal using the methods described below elicits an immune response that is specific to the portion that is administered. In this context immune response includes one or more of IFN-y response, TNF-alpha, IL-12, IL17, II-22, IL-23 in, CD4+ and CD8+ T cells, natural killer cells and humoral responses, i.e. (B-cells and antibodies). In an embodiment, the immunogenic portion is a peptide. In an embodiment, the peptide is a short fragment of any one of the amino acid sequences of the invention. The peptide may have a length of up to 50 amino acids, or up to 20 amino acids. It may have from 10 to 25 amino acids in length from any portion of any one of the amino acid sequences of the invention. Functional variants of the immunogenic portion are also part of the current invention.

[0072] The term “vaccine therapy” should be understood to mean the administration of an immunogen, e.g., a protein or immunogenic portion thereof, (or a nucleotide or mRNA encoding the immunogen, to a mammal with a view to eliciting a response by the host immune system. Typically, it results in the immunogen being destroyed and subsequently recognised by the host immune system.

[0073] The term “immune response” should be understood to mean induced humoral or cellular response in a host subject. In this context immune response includes one or more of IFN-y response, TNF-alpha, IL-12, IL17, II-22, IL-23 in, CD4+ and CD8+ T cells, natural killer (NK) cells, natural killer T cells (NKT cells) and humoral responses, i.e. (B-cells and antibodies).

[0074] As used herein, the term “active pharmaceutical ingredient” or “API” refers to an immunogen that may be used in vaccine therapy such as a protein, protein subunit, polypeptide, botanical (e.g., a plant extract) a fraction of a cell such as a bacterial cell, viral cell, or yeast cell (e.g. a cell wall fraction), or a nucleic acid encoding an immunogen. The nucleic acid may be an RNA (e.g. mRNA) or DNA molecule. In this specification, the term “vaccine” should be understood to mean a composition comprising at least one immunogen, e.g., a protein of the invention, and optionally a suitable adjuvant and / or carrier. The preparation of vaccines comprising a protein or peptide as an active agent is well known in the art, for example US4599230 and US4601903 as examples. It is a subunit vaccine comprising purified proteins or synthetic peptides. In an embodiment, it includes nucleotides or mRNA. A “subunit” vaccine contains one or more specific antigens from a pathogen. It is acellular, i.e. it does not contain whole bacteria.

[0075] In this specification, the term “infection by Klebsiella pneumoniae” should be understood to mean infections of mammals that are caused by Klebsiella pneumoniae.

[0076] In this specification, the term “effective variant’ or “functional variant” as applied to any of the amino acid sequences discussed herein means an amino acid sequence that shares sequence identity, e.g. at least 90% or more, with the sequence, and that is capable of eliciting a protective immune response in a mammal immunised with the variant against subsequent challenge by Klebsiella pneumoniae as described below. The term can include peptides or proteins having an amino acid sequence that is substantially identical to the reference, and which is therapeutically effective as defined herein. Thus, for example, the term should be taken to include variants that are altered in respect of one or more amino acid residues. Preferably, such alterations involve the insertion, addition, deletion and / orsubstitution of 6 or fewer amino acids, preferably 5 or fewer, 4 or fewer, even more preferably of 3 or fewer, most preferably of 1 or 2 amino acids only. Insertion , addition and substitution with natural and modified amino acids is envisaged. It should be noted thatany variant will have principally the same therapeutic effect, or may have an enhanced effect when tested in in vitro or in vivo models of the disease. This term includes “immunogenic portions” of the protein.

[0077] The term “effective variant” or “functional variant” may also be applied to the immunogenic portions, mRNA and nucleotide sequences of the invention. For instance, the “effective variant”, of the nucleotide is one that encodes a protein or immunogenic portion thereof that is capable of eliciting a protective immune response in a mammal immunised with the protein or immunogenic portion thereof against subsequent challenge by Klebsiella pneumoniae species as described below. The “effective variant” or “functional variant” as applied to any of the nucleic acid sequences may be a sequence with at least 90% or more sequence identity with one or more of the nucleic acid sequences disclosed herein.

[0078] The term “DNA vaccine” should be understood to mean a composition that comprises a nucleic acid, typically on a nucleic acid construct that is capable of being delivered to a subject and encodes one or more proteins or immunogenic portions of the invention . The nucleic acid construct expresses said one or more protein or immunogenic portions in said subject. The protein or immunogenic portion thereof may be expressed as separate entities or in the form of a fusion protein. The vaccine may include one or more suitable adjuvants and / or one or more suitable pharmaceutically acceptable carriers. The construct may be a n expression vector. Expression vectors suitable forthis purpose are known in the art, forexample a plasmid or a virus. In an embodiment, the vector is an adenovirus vector.

[0079] The term “adjuvant” should be understood to mean an agent that enhances the subject, i.e., the recipient’s, immune response to an immunogenic protein or portion thereof. Details of a suitable adjuvant composition are well known to a person skilled in the a rt.

[0080] The term “immunogenic composition” refers to a composition comprising one or more proteins of the invention, or an immunogenieportion thereof, (or nucleotide or mRNA encoding same) that is capable of inducing an immune response in an individual.

[0081] Proteins, peptides, and polypeptides (and their variants, including fragments) of and for use in the invention may be generated wholly or partly by chemical synthesis or by expression from nucleic acid. The proteins and peptides of and for use in the present invention can be readily prepared according to well-established, standard liquid or, preferably, solid-phase peptide synthesis methods known in the art (see, for example, J. M. Stewart and J. D. Young, Solid Phase Peptide Synthesis, 2nd edition, Pierce Chemical Company, Rockford, Illinois (1984), in M. Bodanzsky and A. Bodanzsky, The Practice of Peptide Synthesis, Springer Verlag, New York (1984).

[0082] In this specification, the term “amount effective” or “therapeutically effective amount” should be taken to mean an amount which results in a clinically significant reduction or prevention of Klebsiella pneumoniae infection.

[0083] Suitably, the immunogenic component of the vaccine is administered at a dose of from 1 pg to 1 OOpg. In one embodiment, it is administered at a dose of from 1 Opg to 70pg, orfrom 10pg to 50pg, or from 20pg to 40pg, or from 25pg to 30pg.

[0084] Typically, it is given as a bolus dose.

[0085] In the context of the therapeutic aspects of the present invention, the term “individual in need thereof” shall be taken to mean a mammal, preferably a human, who has an infection caused by Klebsiella pneumoniae.

[0086] The term "symptom" is defined as an indication of disease, illness, injury, or that something is not right in the body.

[0087] The term "excipient" refers to a diluent, adjuvant, carrier or vehicle with which the therapeutic or active is administered. Such pharmaceutical carriers or excipients can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water oraqueous buffered solution, are the preferred carriers when the pharmaceutical composition is administered intramuscularly. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers, particularly for injectable solutions, oil in wateremulsionscan also be used in some formulations of the invention. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, driedskim milk, glycerol, propyleneglycol, water, ethanol and the like.

[0088] The vaccine or composition of the invention can be formulated in any suitable means for the desired route of administration. Formulation should suit the mode of administration. Various delivery systems are known and can be used to administer a therapeuticof the invention, e.g., intra-nasally. Methodsof introduction include but are not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, intracerebral, and oral routes. Typically, the mode is intramuscular, subcutaneous, oral or transdermal, such as by microneedle patches. In a typical embodiment, the mode is intramuscular. The vaccine or compositions may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.). The vaccine or composition may be administered togetherwith adjuvants and or other biologically active agents. Administration can be systemic or local. Pulmonary administration can also be employed, e.g., by use of an inhaler or nebulizer, and formulation with an aerosolizing agent.

[0089] An “mRNA vaccine” is a vaccine that uses a copy of messenger RNA (mRNA) to produce an immune response. The vaccine delivers the immunogen encoding mRNA into the subjects immune cells. This uses the mRNA to provide the protein, in this case one or more proteins of the invention or immunogenic portions thereof. It is translated in the cell cytosol. This protein then stimulates the immune response which trains the body to identify, retain a memory for and destroy the pathogen. Typically, the mRNA is delivered by a co-formulation of the RNA encapsulated in a lipid nanoparticle that protects the RNA strand and aids absorption into the cells. Methods to make mRNA and mRNA vaccines are known in the art. mRNA vaccines are described in Kowalski PS, Rudra A, Miao L, Anderson DG (April 2019). "Delivering the Messenger: Advances in Technologies forTherapeuticmRNADelivery". Mol Then 27 (4): 710- 28; and Verbeke R, Lentacker l, De Smedt SC, Dewitte H (October 2019). "Three decades of messenger RNA vaccine development". Nano Today. 28: 100766. In this specification, the term “sample” or “fluid” is a biological sample. The sample is one obtained from a subject, such as, for example, urine or urine cell-sediment, blood, saliva, or a tissue sample.

[0090] In this specification, the term “one or more” should be understood to mean and encompass that one or more, two or more, three or more, four or more, five or more, and so on. In an embodiment, it includes all 19 proteins of the invention.

[0091] The term “peptide” used herein refers to a polymer composed of up to 50 amino acids, for example 5 to 50 amino acid monomers typically linked via peptide bond linkage. The peptide may have up to 20 amino acids. Peptides may be generated wholly or partly by chemical synthesis or by expression from nucleic acid. For example, the peptides can be readily prepared according to well-established, standard liquid or, preferably, solid-phase peptide synthesis methods known in the art (see, for example, J. M. Stewart and J. D. Young, Solid Phase Peptide Synthesis, 2nd edition, Pierce Chemical Company, Rockford, Illinois (1984), in M. Bodanzsky and A. Bodanzsky, The Practice of Peptide Synthesis, Springer Verlag, New York (1984).

[0092] Brief Description of the Figures

[0093] The current invention will now be described with reference to the following Figures in which;

[0094] Figure 1 is a bar graph showing a comparison of the attachment of K. pneumoniae strain CIP52.145 and CIP52.145 eps strain to lung epithelial cell lines A549 and 16HBE14O-. Each bar represents mean±SEM of three independent experiments. Statistically significant differences in bacterial strain attachment to lung epithelial cells were analysed by a two-way ANOVA test.

[0095] Figure 2 shows the identification of K. pneumoniae Cl P52.145 proteins bound toA549 cells. (A) shows PageBlue™ stained gel containing membrane protein preparations of K. pneumoniae CIP52.145 and (B) shows a PVDF membrane probed with 1 x 106cell / ml A549 cells. 2-D gel and blots are representative of three independent experiments.

[0096] Figure 3 shows the identification of K. pneumoniae Cl P52.145 proteins bound to 16HBE14o- cells. (A) PageBlue stained gel containing membrane protein preparations of K. pneumoniae CIP52.145. (B) PVDF membrane probed with 1 x 106cell / ml 16HBE14o~ cells. 2-D gel and blots are representative of three independent experiments.

[0097] Figure 4 illustrates identification of K. pneumoniae Acps proteins bound to A549 cells. (A) PageBlue stained gel containing membrane protein preparations of K. pneumoniae Acps. (B) PVDF membrane probed with 1 x 106 cell / ml A549 cells. 2-D gel and blots are representative of three independent experiments. Figure 5 shows identification of K. pneumoniae Acps proteins bound to 16HBE14o- cells. (A) PageBlue™ stained gel containing membrane protein preparations of K. pneumoniae Acps. (B) PVDF membrane probed with 1 x 106cell / ml 16HBE14o- cells. 2-D gel and blots are representative of three independent experiments.

[0098] Figure 6 illustrates negative control of attachment K. pneumoniae CIP52.145 or K. pneumoniae Acps proteins to lung epithelia cells. (A) PageBlue stained gel containing membrane protein preparations of K. pneumoniae Cl P.52.145. (B) PVDF membrane without probing any lung epithelia cells. (C) PageBlue stained gel containing membrane protein preparations of . pneumoniae Acps. (D) PVDF membrane without probing any lung epithelia cells.

[0099] Figure 7 shows that the expression of K. pneumoniae vaccine candidates increases bacterial attachment to human lung epithelial cells. Induced BL21 cells expressing recombinant protein increased the level of bacterial attachment to lung epithelial cells compared to induced BL21 cells transformed with empty pET100 / D-Topo (control) in vitro. (A) Induced BL21 cells expressing LysM / BON protein increased the level of bacterial attachment to lung epithelial cells (p = 0.012). (B) Induced BL21 cells expressing OMPP1 protein increased the level of bacterial attachment to lung epithelial cells (p = 0.0199). (C) Induced BL21 cells expressing Dps protein increased the level of bacterial attachment to lung epithelial cells ( p = 0.0011). Each error bar represents mean±SEM from three independent experiments. Significant differences in adhesion were analysed by two-way ANOVA test (A, C) or t - test (B).

[0100] Figure 8 shows serological analysis of immune response to LysM / BON immunisation (top panel) and OmpP1 (lower panel) in mice: Total IgG, I gG 1 and lgG2a antibody responses after LysM / BON:SAS or OmpP1 :SAS immunisation, three doses, two weeks apart.

[0101] Figure 9 demonstrates the protective efficacy of K. pneumoniae vaccine candidates. K. pneumoniae colonisation in peritoneal cavity (A) and dissemination to spleen (B) after LysM / BON, OMPP1 or Dps immunisation. Afteri.p. challenge with 1 x 105CFU . pneumoniae CIP52.145. (A) IP lavage fluid from BALB / c mice (n=8 or n=6) were collected at 24 hours post infection. Bacterial burden was determined as CFU / ml and is shown on a log scale. All colonisation data are displayed as means + standard deviation (SD). Significant differences in adhesion were analysed by Mann-Whitney test, p = 0.0080, p = 0.0007, p = 0.0007 respectively. (B) The spleens from BALB / c mice (n=8 or n=6) were collected at 24 hours post infection. Bacterial burden was determined as CFU / ml and is shown on a log scale. All colonisation data are displayed as means + standard deviation (SD). Significant differences in adhesion were analysed by Mann-Whitney test, p = 0.0263, p = 0.0003, p = 0.0043, respectively. Figure 10 illustrates LysM / BON or OMPP1 -activated CD4+T cells, CD8+T cells, y6 T cells, NK cells and NKT cells, a): DP cells, b): NK cells, c): NKT cells, d-e): total or activated y6 T cells, f): CD4+regulatory T cells, g-l): total, naive, memory, effector activated or regulatory helper T cells, m-r: total, naive, memory, effector activated or regulatory cytotoxic T cells. Asterisks denote statistically significant differences according to a t-test. The levels of significance are represented as follows: ****p < 0.0001 , ***p < 0.001 , **p < 0.01 , *p < 0.05, ns > 0.05. Each spot represents a single immunised mouse.

[0102] Figure 11 shows LysM / BON or OMPPI immunisation stimulated IL-4, IL-17, IL-22, TNF and I FN-y recall responses in: a-e) CD4+T cells, f-i ) CD8+T cells, j-n) y6 T cells. Asterisks denote statistically significant differences according to a t-test. The levels of significance are represented as follows****p < 0.0001 ; ***p < 0.001 ; **p < 0.01 ; *p < 0.05; ns > 0.05. Each spot represents a single immunised mouse.

[0103] Figure 12 shows LysM / BON or OMPPI immunisation stimulated IL-4, IL-17, IL-22, TNF and I FN-y recall responses in: a-d) NK cells, e-i ) NKT cells, j-n) CD3- cells. Asterisks denote statistically significant differences according to a t-test. The levels of significance are represented as follows****p < 0.0001 ; ***p < 0.001 ; **p < 0.01 ; *p < 0.05; ns > 0.05. Each spot represents a single immunised mouse.

[0104] Detailed Description of the Invention

[0105] All publications, patents, patent applications and other references mentioned herein are hereby incorporated by reference in their entireties for all purposes as if each individual publication, patent or patent application were specifically and individually indicated to be incorporated by reference and the content thereof recited in full.

[0106] There is a need to provide a vaccine to prevent or treat infections of K. pneumoniae in a subject, preferably a human. There are currently no vaccines available.

[0107] The invention provides compositions, such as a subunit vaccine, comprising one or more immunogens of the invention. The one or more immunogens are generally one or more proteins of the invention or immunogenieportions thereof. The proteinsof the invention are selected from the group comprising LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC, Ribokinase, Wzi family protein, LptD LPS assembly protein, Outer membrane channel protein, Polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, Outer membrane protein assembly factor BamA, Malate dehydrogenase, alcohol dehydrogenase, Putative outer membrane lipoprotein Sly B, long-chain fatty acid outer membrane transporter, Putative porin, and Maltoporin. The proteins of the invention or immunogenic portions thereof are for use as vaccine antigens.

[0108] The vaccine of the invention can be a multivalent vaccine comprising a mixture of antigens. In an embodiment, the composition of the invention comprises two or more immunogens, three or more immunogens, four or more immunogens, five or more immunogens, six or more immunogens, seven or more immunogens, eight or more immunogens, nine or more immunogens, ten or more immunogens, eleven or more immunogens, twelve or more immunogens, thirteen or more immunogens, fourteen or more immunogens, fifteen or more immunogens, sixteen or more immunogens, seventeen or more immunogens, eighteen or more immunogens, or nineteen immunogens.

[0109] In an embodiment, the numberof immunogens consist of two immunogens, three, four, five, six, seven, eight, nine, ten, eleven, twelve, thirteen, fourteen, fifteen, sixteen, seventeen, eighteen, or nineteen immunogens.

[0110] In an embodiment, the composition comprises all 19 proteins of the invention or immunogenic portions thereof.

[0111] The composition of the invention may comprise a combination of immunogenic portions. This may be any combination of the immunogenic portions of the invention.

[0112] It will be appreciated that in the embodiments disclosed above, the composition may comprise a nucleotide encoding the one or more immunogen, or an mRNA encoding the one or more immunogen instead of the immunogen itself. In such an embodiment, the vaccine may be a DNA vaccine. It may be an adenoviral vector vaccine which comprise one or more nucleotide sequences encoding one or more immunogen of the invention.

[0113] In such an embodiment, the vaccine may be an mRNA vaccine.

[0114] In an embodimentofthe invention the composition is a vaccine composition. The vaccine may have one or more adjuvants. Typically, the adjuvant is one that induces a T cell and B cell response, e.g., the CAF series, such asCAF-01 orCAF-09 orCpG ODNS (ODN 2006). It may be manno-heptose capsular polysaccharide (CPS) (Scott et al., Infect Immun. 2014 Aug;82(8):3206-13). It will be appreciated that such adjuvants are known in the art.

[0115] The invention further provides a DNA vaccine comprising an active agent, optionally a pharmaceutically acceptable carrierand optionally an adjuvant. In an embodiment, the active agent is a nucleic acid sequence encoding one or more immunogens of the invention. In an embodiment, the nucleotide acid sequence comprises (or consists of) SEQUENCE ID NO. 1 to 19 or a therapeutically effective variant thereof, typically having at least 90% sequence identity with SEQUENCE ID NO. 1 to 19. In an embodiment, the vaccine of the invention is an mRNA vaccine. Thus, in an aspect, a vaccine is provided that comprises an mRNA that encodes one or more immunogens of the invention.

[0116] The vaccine or composition of the invention may comprise a combination of immunogenic portions.

[0117] Notably, the composition is for use as a vaccine to prevent ortreat infection by K. pneumoniae in a subject. A vaccine for use to treat an infection can be referred to as “post exposure vaccines”.

[0118] The composition can be used to induce an immune response against K. pneumoniae in a subject.

[0119] An aspect of the invention includesone or more immunogens of the invention ora composition comprising said one or more immunogens, for use in immunotherapy.

[0120] Thus, also provided by the invention is an antibody (or a composition comprising the same) directed to one or more immunogens of the invention and its use in immunotherapy. The composition may contain multiple antibodies directedto different immunogens of the invention. Antibodies to said immunogens or antigens can be obtained with routine methods known in the art, for example hybridoma technology. The use of monoclonal antibodies and generation of such antibodies, to provide long term protection has been investigated for K. pneumoniae infection (Diago-Navarro E, et al., 2017. Antibody-Based Immunotherapy To Treat and Prevent Infection with Hypervirulent Klebsiella pneumoniae. Clin Vaccine Immunol 24:e00456- 16)

[0121] An embodimentof the treatment aspect of the current invention includes the use of an antibody directed or raised against, to one ormore immunogens of the invention, which are suitable for immunotherapy and administered to the subject. The antibody is typ ically a monoclonal antibody specific to the immunogen of the invention .

[0122] The nucleotide sequence and amino acid sequence of the proteins of the invention are as follows:

[0123] 1. LysM Peptidoglycan associated protein (also known as LysM / BON)

[0124] The term LysM encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.20.

[0125] Accession Code: KPHS_26690 / A0A0H3GSY1

[0126] Gene Sequence:

[0127] SEQUENCE ID NO.1 atgggattattaaattttgtgaaagaagcgggcgagaaaatatgggatgccgtctccggcgacagtaaagaagatcgggctga aaagctgaaaaaacatatcgatggacttaatctgcctggcgccgaaaaggtcaatattgatgttgcggaagatggcaccgcaa cggtcactggtgatgtcgcttcgcaggaggataaggaaaaaattctggttgcggtcgggaatgtcactggcgtaggccaggtca gtgacggcgttaaggtgacgcaaagcggggcggaaagtcgtttttacacggtcaaatccggtgacactctgagcgcaatctcaa aagcgatgtatggctcggccaatgagtatcaacgcatttttgaagccaataaaccgatgctgacccatcccgacaaaatctatcc gggtcaggtgctgattatcccggcgaagtag

[0128] Protein Sequence:

[0129] SEQUENCE ID NQ.20

[0130] MGLLNFVKEAGEKIWDAVSGDSKEDRAEKLKKHIDGLNLPGAEKVNIDVAEDGTATVTGDVA SQEDKEKILVAVGNVTGVGQVSDGVKVTQSGAESRFYTVKSGDTLSAISKAMYGSANEYQR IFEANKPMLTHPDKIYPGQVLIIPAK

[0131] 2. DNA protection during starvation protein

[0132] The term DNA protection during starvation protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.21 .

[0133] Accession Code: KPHS_16730 / A0A0H3GQL3

[0134] SEQUENCE ID NO.2 atgagtaccgcaaaactggttaaatcaaaagcgtctaatctggtctatacccgcaacgatgtcgctgacagcgaaaagaaagc gaccattgagctgttgaatcgccaggtgatccagttcattgacctgtcacttatcaccaagcaggcacactggaatatgcgcggtg cgaactttatcgccgtgcacgaaatgctggatggcttccgtaccgcgctgaccgaacatctggacaccatggccgagcgcgcgg tgcagctgggcggcgtcgccctgggtaccactcaggtcattaacagcaaaaccccgctgcagagctacccactggacatccat cacgttcaggatcacctgaaagccctggctgaccgctacgcggtagtggcgaacgatgtgcgtaaggcgatcgacgaagcaa aagacgaagacactgcagatatcttcaccgcggcttcccgcgatctggataaattcctgtggttcattgaagccaacatcgagta a

[0135] Protein Sequence: SEQUENCE ID NO. 21 MSTAKLVKSKASNLVYTRNDVADSEKKATIELLNRQVIQFIDLSLITKQAHWNMRGANFIAVH EMLDGFRTALTEHLDTMAERAVQLGGVALGTTQVINSKTPLQSYPLDIHHVQDHLKALADRY AWANDVRKAI DEAKDEDTADI FTAASRDLDKFLWFI EANI E

[0136] 3. OMPP1 / Fadl_ / TodX family outer membrane transporter

[0137] The term 0mpP1 encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.22.

[0138] Accession Code: KPHS_37960 / A0A0H3H155

[0139] SEQUENCE ID NO.3 atgtgggagctgtccggctataaccgcgtggcgccgcagtgggcggtccactatagcctgacctacaccagctggagtcagttc caggagctgaaagccaccggcagcaacggccagacgctgttctataaagaggaaggcttcaaggacgcctatcgtcttgcgct g g g g acca cctattattatg a eg a taa etgg accttccgta ccg g tat eg ccttcga tg aca gcccg gtg ccgg ctaataaccgct eta teteta tteeg g a cca g ga tcgcctgtg g etg a geg egg g ta ccaccta eg eg ttca ata a ag atg cctcg g ttg aegteg gc gtttcatacatgcacgggcagcatgttgaaattaaggaaggcccgtataccttccgttccgaaggaaccgcctggctgtacggcg eta a ettea a eta ccg ettet g a

[0140] Protein Sequence: SEQUENCE ID NO. 22

[0141] MWELSGYNRVAPQWAVHYSLTYTSWSQFQELKATGSNGQTLFYKEEGFKDAYRLALGTTY YYDDNWTFRTGIAFDDSPVPANNRSISIPDQDRLWLSAGTTYAFNKDASVDVGVSYMHGQH VEIKEGPYTFRSEGTAWLYGANFNYRF

[0142] 4. Outer membrane protein assembly factor BamC

[0143] The term BamC encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.23.

[0144] Accession Code: KPHS_38830 / A0A0H3GWJ5

[0145] Gene Sequence:

[0146] SEQUENCE ID NO.4 atggcttactcagtacagaagtcgcgcctggcgaaggttgcgggtgtttcgctggttctactcctcgcggcctgtagttctgactcgc gctataagcgtcaggtgagcggtgatgaggcctatttacaggcatcaccgttgagcgaacttcatgcgccggcggggatgatcct gccgattcaggtgggtgattacaatattccggtggctaacagcaccggcgcggtcggtaaagcgctggacattcgtccgccggc gcagccgctggcgctggtgagcggtgcgcggacccagtttaacggcgataccgcgacgctgatggtcgagaacggccgcag cggttcgctgtgggcacaggtcaccagtattcttcagtcgaagaactacgttatcgccaagcgcgatgatgcgagccagacgctg aacaccgactgggttgagtggaaccgtcttgatgaagatcagcagtatcgcggtcgttatcaaatctccgtcaaaccgcagggtt atcagcaagcggtggtcgttaagctggtgaatctggagcaggcgggtaaaccggttgccgatccggcgtctctgcagcgctaca gcaccgctatgctgaacgtcatttcggaaggccttgacatgaatgccaccagcgcccagaacgccgcccagcgcagcgcggg g g cca ccttcg a eg tgcaga g egeeg ccg aegataeeg geetg ccg atg ctcgta gtteg eg eg ccg ttta a cctg g tg tg g ca g eg tetg cctg g eg ccctcg ag a aa g tg g gg atg aaag tg accga cag ca eg eg ctctcag g gca g ca tg geg etg a cctat a a g ccg etg tetg aca g cag etgg cag g ag etgg g eg ca eg tg acccg ca g etg g teteeg geg acta caa a etg cag g teg gcgacctcgataaccgcagtagcctgcagtttatcgacccgaaaggccacaccctgacgcagtcgcaaaatgacgccctggta g ccg tattcca g g ccg cattta a caa g ta a

[0147] Protein Sequence:

[0148] SEQUENCE ID NO.23

[0149] MAYSVQKSRLAKVAGVSLVLLLAACSSDSRYKRQVSGDEAYLQASPLSELHAPAGMILPIQV GDYNIPVANSTGAVGKALDIRPPAQPLALVSGARTQFNGDTATLMVENGRSGSLWAQVTSIL

[0150] QSKNYVIAKRDDASQTLNTDWVEWNRLDEDQQYRGRYQISVKPQGYQQAWVKLVNLEQA GKPVADPASLQRYSTAM LNVISEGLDM NATSAQNAAQRSAGATFDVQSAADDTGLPM LWR APFNLVWQRLPGALEKVGMKVTDSTRSQGSMALTYKPLSDSSWQELGARDPQLVSGDYKL QVGDLDNRSSLQFIDPKGHTLTQSQNDALVAVFQAAFNK

[0151] 5. Ribokinase

[0152] The term ribokinase encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.24.

[0153] Accession Code: KPHS_26410 / A0A0H3GXH5

[0154] Gene Sequence:

[0155] SEQUENCE ID NO.5 atgtcaggaaaagtctgtgtttttggttcattcaatttcgacatggtcgccagggtggatcgctttccggtaccaggtgagtcgttagtg gcctgcggaagtatgaccagcgccggaggtaaaggggcgaaccaggccaccgccgcgctaaaagcgggagcaaatgtcc actatatcggcaagattggcaatgacaccttcggccattttgcccgccgtcatctgaaaggcgtgggcttcaatgctgtgacgctgc tggtcgccgaagagatccccaccggcaatgcgctgatttatgtcgccggtaacgatgccgaaaacatgattgccgtcgatcccg gcgcgaatatgacggtgaccgatgatgaaatcgccggctgcattcccgctatcggctgcgcggatgtggttctggttcagctgga gaacaatctttctgctattgaacaggtcatagacgccggcaaacaagccggggccctggtaatacttaatcccgcgccctggca gcccgttgagcacgcccttctacgcaaagtggatttacttacaccgaacgccacggaggccgggttaatgaccggacgccggg ttgacagtctgacggccgcggctgaggccgccgacgtgttacatgctcagggcgcgcgcaacgtcattatcactctcggcgcca gcggcgccctgctgagcgagcatggggtgaaatcccctattccctgctttccttctcacccccgagacactaccggtgcgggcga tg ccttca acg g eg egetg geg geg egg etgg cctgcg gg g aa ccgcta cag g ccg ca gcccga ttcgcg geg g cctatgcc gccgtcagtgtcgaaaagcaaggcgcctcatctttacctgagtacctggaggcacaggaacggcttctccgtgccgccgctgatt atgaaatggcataa

[0156] Protein Sequence:

[0157] SEQUENCE ID NO.24

[0158] MSGKVCVFGSFNFDMVARVDRFPVPGESLVACGSMTSAGGKGANQATAALKAGANVHYIG KIGNDTFGHFARRHLKGVGFNAVTLLVAEEIPTGNALIYVAGNDAENMIAVDPGANMTVTDD EIAGCIPAIGCADWLVQLENNLSAI EQVI DAGKQAGALVI LNPAPWQPVEHALLRKVDLLTPN ATEAGLMTGRRVDSLTAAAEAADVLHAQGARNVIITLGASGALLSEHGVKSPIPCFPSHPRD TTGAGDAFNGALAARLACGEPLQAAARFAAAYAAVSVEKQGASSLPEYLEAQERLLRAAAD YEMA 6. Wzi family protein

[0159] The term Wzi family protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.25.

[0160] Accession Code: KPHS_35700 / A0A0H3GSK2

[0161] Gene Sequence:

[0162] SEQUENCE ID NO.6 atgataaaaattgcgcgcattgccgtgacgttgggtttgctttcctcactgggagcccaggcttacgcggccgggttagtggtaaat gacaacgacttacgtaacgacctggcctggctttccgatcgcggggtcatccatctgagcctgtcgacctggccgctgagccagg aagagattgctcgggcgctaaagaaggccaagccgtcttattcttctgagcaagtggtgctggcccgtatcaaccagcgactgtc tgctttaaaagccgatttcagggtcaccggctacacctcaaccgaccagcctggcaccccgcaggggtttggccagacgcagc cggcggataattctctgggcctggcattcaacaacagcggtgagtggtgggatgtccacctgcagggcaacgtcgaaggaggg gagcggattagcaacggctcgcgcttcaacgccaacggcgcatacggtgcggtgaagttctggaaccagtggctctcttttggtc aggttccgcagtggtggggacctggctatgaaggtagcctgatccgcggggatgccatgcggccgatgaccggtttcctgatgc agcgcgcagagcaggcagcgccagagacctggtggttgcgctgggtcggcccatggcagtaccagatctccgccagccaga tgaatcaatataccgcggtaccccatgccaaaattatcggcggtcgttttaccttttcaccattccagtctttagaattaggcgcgtca cgcattatgcagtggggtggggaagggcggcctgagtcatttagcagcttctgggatggcttcactggccatgataataccggga cggataacgaaccgggtaaccagctggccggatttgactttaaattcaaactcgagccgactctagggtggccggtgagtttctat gggcaaatggtcggcgaggatgagtctggctacctgccttccgcgaatatgttccttggtggagtggaagggcaccacggttggg gtaaagatgcggttaactggtatgtggaagcgcatgacacgcgtaccaacatgagccgaaccaattacagctatacccaccac atctataaagatggttattaccaacaagggtatccactgggggatgcgatgggtggggatggtcaactcattgccgggaaggttg agctgattaccgaagataaccagcgttggagtacacgcctcgtttacgccaaagttaaccctgagaaccagtcgatcaataaag cattcccccatgccgatactttgaaaggtgtacagctaggctggagcggcgatgtgtatcagtctgttcgtctgaatacctctttgtgg tacacaaacgcgaacaacagcgacagcgatgacgttggggcaagcgcagggatagaaataccgtttagtttataa

[0163] Protein Sequence:

[0164] SEQUENCE ID NO.25

[0165] MIKIARIAVTLGLLSSLGAQAYAAGLWNDNDLRNDLAWLSDRGVIHLSLSTWPLSQEEIARAL KKAKPSYSSEQWLARINQRLSALKADFRVTGYTSTDQPGTPQGFGQTQPADNSLGLAFNN SGEWWDVHLQGNVEGGERISNGSRFNANGAYGAVKFWNQWLSFGQVPQWWGPGYEGS LI RGDAM RPMTGFLMQRAEQAAPETWWLRWVGPWQYQISASQM NQYTAVPHAKI IGGRF TFSPFQSLELGASRIMQWGGEGRPESFSSFWDGFTGHDNTGTDNEPGNQLAGFDFKFKL EPTLGWPVSFYGQMVGEDESGYLPSANMFLGGVEGHHGWGKDAVNWYVEAHDTRTNMS RTNYSYTHHIYKDGYYQQGYPLGDAMGGDGQLIAGKVELITEDNQRWSTRLVYAKVNPEN QSINKAFPHADTLKGVQLGWSGDVYQSVRLNTSLWYTNANNSDSDDVGASAGIEIPFSL

[0166] 7. LptD LPS assembly protein

[0167] The term LptD encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.26.

[0168] Accession Code: KPHS_07630 / A0A0H3GMF8

[0169] Gene Sequence: SEQUENCE ID N0.7 atgattgccagcgccttgtatagccaacaaggcctcgctgccgatctcgcaacgcaatgtatgcttggcgtgccaagctatgatcg tccg ct eg tg g a agg tegteetg geg atetgeeg g tg a egatta a egeeg atcatg eg aa g gg caactacccg ga caacg ccg tctttaccggcaacgtcgatattaaccaggggaatagtcgcctccgcgccgacgaagttcagctgcaccagcagcaggccgcg ggccaggcgcagccggtgcgcacggtggacgcgctgggcaacgtgcattacgacgataaccaggtgatcctcaaagggcca aaagcctggtcgaatctgaataccaaagataccaacgtctggcagggcgattatcaaatggtcggacgccagggacgcggca ccgccgacctgatgaaacagcgcggtgaaaaccgctataccattctcgaaaacggcagctttacctcctgtctgccgggctccg acacctggagcgtcgtcggcagcgaagttatccacgatcgcgaagagcaggttgccgagatctggaacgcccgcttcaagctt g g ctctg tg ccg a tttt ctata g ccccta cct g cag etgeeg gtg gg eg ata ag eg teg ttca g gcttcctg a tcccg aa eg egaaa tacagcaccaaaaacggcgtggaattctccctgccgtactactggaacatcgcgccaaacttcgatgccaccattactccgcact atatgaacaaacgcggcggcgtgatgtgggaaaacgagttccgctatctgacccagctcggcagcggcttaactgaattcgact a cctg ccg teg g ataaa gtctacg aag a eg a cca ctcga g ega cag ca acagccg ccg etgg ctgttctactg ga a ccactca ggggttatcgatcaggtatggcgtctgaacgctgactacaccaaggtcagcgatcctgactacttcaacgacttcagctcgaaat atggttccagtaccgatggctatgcgacgcagaaattcagcgccggttacgtcaaccagaactttgacgccacggtatcgacca aacagttccaggtctttgaccgcgaatcgagcaactcctattcggctgagccgcagctcgacgtcaactactaccagaatgatgt cggtccgttcgatacccatctctatggacaggttgcccattttgttaactcgaataacaacatgccggaagcgacccgcgttcacttc gaaccgacgatcaacctgccgctgtccaacggttggggcagtctgaataccgaagccaagctgctggcgactcactaccagca gagcaacctcgataagtacaatgccgccaacggcactgactataaagagtccgtcagccgcgtaatgccgcagtttaaagtcg acggcaaaatggtctttgaacgcgacctgcaggagggattcacccaaacgctggaaccgcgcgtgcagtatctgtacgtgccgt accgcgatcagagtgaaatcggcaactacgactccacgctgttgcagtcggattacaccggtctgttccgcgaccgtacctatag eg g tetg g a ccg ca teg egteg getaatea gg tcacca ccgg gctcacctcg egeg tgtatgatgeeg ccg eg gtg gaa eg tttt aatatttccgttggtcaaatctactatttcaccgagtcacgcaccggtgatgacaacatcaactgggagaacaacgataccacgg gttcactggtctgggccggcgatacctactggcgcattgccgatgaatggggtctgcgcggagggatccagtacgatacgcgtct ggataacgttgccactggtaacggcaccattgaataccgtcgcgatgagaaccgcttagttcagcttaactatcgttacgccagcc cggaatatattcaggccacgctgccgtcatattccaccgcggcacaatataaacagggtatttcgcaggtggggatgaccgcca gttggccgattgtcgatcgctggtccgtggtcggggcctactactacgatactaatacccggaaagcagcaaaccagatgttggg tgtgcagtataactcctgctgctacgccatccgtcttggctacgaacgtaaagtcaa eg gctggaacagcaacga caacg gegg cgagagcaaatacgataacaccttcggaatcaatatcgaattgcgcggtctgagctctaactacggcctcggcacccagcagat gctgcgctcgaacattttaccgtaccagagctccctgtga

[0170] Protein Sequence:

[0171] SEQUENCE ID NO.26

[0172] MIASALYSQQGLAADLATQCMLGVPSYDRPLVEGRPGDLPVTINADHAKGNYPDNAVFTGN VDINQGNSRLRADEVQLHQQQAAGQAQPVRTVDALGNVHYDDNQVILKGPKAWSNLNTK DTNVWQGDYQMVGRQGRGTADLMKQRGENRYTILENGSFTSCLPGSDTWSWGSEVIHD REEQVAEIWNARFKLGSVPIFYSPYLQLPVGDKRRSGFLIPNAKYSTKNGVEFSLPYYWNIA PNFDATITPHYMNKRGGVMWENEFRYLTQLGSGLTEFDYLPSDKVYEDDHSSDSNSRRWL FYWNHSGVIDQVWRLNADYTKVSDPDYFNDFSSKYGSSTDGYATQKFSAGYVNQNFDATV STKQFQVFDRESSNSYSAEPQLDVNYYQNDVGPFDTHLYGQVAHFVNSNNNMPEATRVHF EPTINLPLSNGWGSLNTEAKLLATHYQQSNLDKYNAANGTDYKESVSRVMPQFKVDGKMV FERDLQEGFTQTLEPRVQYLYVPYRDQSEIGNYDSTLLQSDYTGLFRDRTYSGLDRIASAN QX / TTGLTSRVYDAAAVERFNISVGQIYYFTESRTGDDNINWENNDTTGSLVWAGDTYWRIA DEWGLRGGIQYDTRLDNVATGNGTIEYRRDENRLVQLNYRYASPEYIQATLPSYSTAAQYK QGISQVGMTASWPIVDRWSWGAYYYDTNTRKAANQMLGVQYNSCCYAIRLGYERKVNG WNSNDNGGESKYDNTFGINIELRGLSSNYGLGTQQMLRSNILPYQSSL 8. Outer membrane channel protein

[0173] The term outer membrane channel protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or

[0174] 99% sequence identity with SEQUENCE ID NO.27.

[0175] Accession Code: KPHS_45760 / A0A0H3H370

[0176] Gene Sequence:

[0177] SEQUENCE ID NO.8 atgaagaaattgctccccattcttatcggcctgagcctgaccgggttcagcgccatgagccaggcggaaaacctgcttcaggttta ccagcaggcacgcatcagcaaccccgatctgcgtaaatcggcagccgatcgtgacgccgcgttcgaaaagatcaacgaagc gcgcagtccattactgcctcagcttgggctgggagcggattatacctataccaatggctatcgcgacagcaacggcgtcaactct aacgtcaccagcggttcgctgcagttaacgcaggttctgtttgatatgtcgaaatggcgcgccctgacgctgcaggaaaaaacg gcagggattcaggatgtcacgtatcagaccgatcagcaaacactgattctgaataccgcgacggcctattttaaagtgctggccg ccatcgacacgctctcctataccgaagcgcagaaacaggctatttaccgccagttggatcaaaccacgcagcgctttaacgtag gcctggtggcgatcaccgacgtacagaacgcccgttcacaatacgatgccgtgctggcgaacgaagtcaccgcgcgtaacga tctcgacaacgccgtcgaaggactgcgtcaagtcaccggtaattactatccggagctggcctccctgaacgtgaacggcttcaa aaccaacaagccgcaggcggtcaacgccctgctgaaggaagcggagaaccgcaacctgtcactgctgcaggcgcgtctga gccaggacctggcccgcgagcagattcgtcaggcgcaggacggccatttgccgaccctgaacctttccgcctctaccggggtct cgaatacgcgttacaacggttcgaaaaccaataccccgctcgcctacaacgacagtgataacgggcagaaccaaatcggcct gaacttctctctgccactgtatcagggcggcgcggtgacctcgcaggtcaagcaggcacaatacaacttcgttggcgccagcga g ca g ctg g a a agcg cccaccg ca g cgtcg tg ca ga ctgtg cgttcatcg ttta aca a cgtg aatgcctccatca gcag catcaa cgcctacaaacaggcggtggtctctgcgcaaagctccctggatgccatggaagctggctactcggtgggtacgcgtactatcgtt gacgtcctcgacgccaccactacgctgtataacgctaagcagcagctctcgaatgcgcgctacaactacctgatcaacgagctg aacattaagtcggcgttaggtaccctgaacgagcaggatctggtcgccctgaacaacacgctgggtaaacccatctccacctcc gcagatagcgtcgcgccggaaaatccgcaacaggatgccaccgctgatggctacggcaacactaccgcggcggtgaagcc ggcgtccgcacggaccacccagagcagcggcagcaatccgttccgtcagtaa

[0178] Protein Sequence:

[0179] SEQUENCE ID NO.27

[0180] MKKLLPILIGLSLTGFSAMSQAENLLQVYQQARISNPDLRKSAADRDAAFEKINEARSPLLPQ LGLGADYTYTNGYRDSNGVNSNVTSGSLQLTQVLFDMSKWRALTLQEKTAGIQDVTYQTD QQTLI LNTATAYFKVLAAI DTLSYTEAQKQAIYRQLDQTTQRFNVGLVAITDVQNARSQYDAVL ANEVTARNDLDNAVEGLRQVTGNYYPELASLNVNGFKTNKPQAVNALLKEAENRNLSLLQA RLSQDLAREQIRQAQDGHLPTLNLSASTGVSNTRYNGSKTNTPLAYNDSDNGQNQIGLNFS LPLYQGGAVTSQVKQAQYNFVGASEQLESAHRSWQTVRSSFNNVNASISSINAYKQAWS AQSSLDAMEAGYSVGTRTIVDVLDATTTLYNAKQQLSNARYNYLINELNIKSALGTLNEQDLV ALNNTLGKPISTSADSVAPENPQQDATADGYGNTTAAVKPASARTTQSSGSNPFRQ

[0181] 9. Polysaccharide export protein

[0182] The term polysaccharide export protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.28.

[0183] Accession Code: KPHS_35690 / A0A0H3GV97 Gene Sequence:

[0184] SEQUENCE ID N0.9 atgaagaaaaaacttgttagattttcggcattagcgttggctattgggtttttatcgggttgtaccattatccctggtcagggattaaata gtctgcggaagaacgtagtggagcttccggacagcgattacgatttggataaactggttaacgtttatccgatgacgccgggcctg atcgagcaactccgtccggagactgtgctcgctcgtccaaacccacagttggataatttactacgtagttacgaatatcgcattggg gtaggtgatgtactgatggttacggtatgggatcacccggaattgacaacgcctgcaggtcaataccgtagcgccagcgacacc ggcaactgggttaactccgacggtaccattttctatccttatatcggtaaagtacaggtggcggggaaaacgcttagccaggtacg ccaggatatagccagccgtttgacaacatatatcgaaagtccacaggtggacgtaagtattgctgcattccgctcgcaaaaagct tatgtgactggggaagttgctaaatcaggacaacaaccgattacaaacattcctcttactgtcatggatgctattaatgcagcaggt ggtttagctgaaaatgcagattggcgtaacgttgttctaacccataatggtaaagatactaaaatctcattatatgcattaatgcaaa agggcgacttaacgcaaaaccatatgctctatccaggagacatcctttttgttcctcgtaatgatgatcttaaagtttttgtgatgggtg aagttggtaaacaaaccacaatgaagatggatcgaagtggtatgaccctagccgaggctttagggaatgctgaaggggtatctc aggaaatgagcgatgcaacagggatatttgttatacgtcaattgaaatccaataagcagggaaaaattgctgatatatatcaatta aatgcacaagacgcatcagcaatggttctaggaacagaattccaattacaaccatatgacatcgtttatgtcactacagcaccttt ggtacgctggaatcgagtaatctctcaattagtaccaactataacaggtgtacatgatatgacagaaacagttcgttatattaagag atggcctaattaa

[0185] Protein Sequence:

[0186] SEQUENCE ID NO.28

[0187] MKKKLVRFSALALAIGFLSGCTIIPGQGLNSLRKNWELPDSDYDLDKLVNVYPMTPGLIEQL RPETVLARPNPQLDNLLRSYEYRIGVGDVLMVTVWDHPELTTPAGQYRSASDTGNWVNSD GTIFYPYIGKVQVAGKTLSQVRQDIASRLTTYIESPQVDVSIAAFRSQKAYVTGEVAKSGQQP ITNIPLTVMDAINAAGGLAENADWRNWLTHNGKDTKISLYALMQKGDLTQNHMLYPGDILFV PRNDDLKVFVMGEVGKQTTMKMDRSGMTLAEALGNAEGVSQEMSDATGIFVIRQLKSNKQ GKIADIYQLNAQDASAMVLGTEFQLQPYDIVYVTTAPLVRWNRVISQLVPTITGVHDMTETVR YIKRWPN

[0188] 10. DNA-binding transcriptional activator OsmE

[0189] The term OsmE encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.29.

[0190] Accession Code: KPHS_21270 / A0A0H3GLT9

[0191] Gene Sequence:

[0192] SEQUENCE ID NQ.10 atgaacaaaagcttagcaggaatactgggcgtcaccgtcgcgttaaccttactggcgggctgtaccgcttacgatcgtaccaaa gaccagtttacccagccggtggtgaaagatgtgaagaaagggatgtcgcgtcagcaggtgatgcagatcgccggcaaaccttc aacggaagtcaccatggtccatgcccgcggcacctgccagacctatatcctcggtcaacgagatggtaaagttgaaacctacttt gtggccctcgacgaaacgggtcacgtcatgaactccggctaccagacctgcgccgagtatgataccgacccgcgcaacgcca ggtaa

[0193] Protein Sequence:

[0194] SEQUENCE ID NO.29 MNKSLAGILGVTVALTLLAGCTAYDRTKDQFTQPWKDVKKGMSRQQVMQIAGKPSTEX / TM VHARGTCQTYILGQRDGKVETYFVALDETGHVMNSGYQTCAEYDTDPRNAR

[0195] 11. Deoxyribose-phosphate aldolase / deoC

[0196] The term DeoC encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.30.

[0197] Accession Code: KPHS_26440 / A0A0H3GSW1

[0198] Gene Sequence:

[0199] SEQUENCE ID NO.11 atgaaagcaattaaaaacaaaggacaggataagaccatgaatgaaacaacccgtcgttttgcacgtttagttgatctgagcgca gttcaggccaccagtaccgaagcggatgttcgcgcctgcgccgagctggctgccagatacaacattatttccgtccacgtactgc cttg ctg g acg eg ctttctcag ta eg ttgctg ccg ca gca gg g tacg gg agag gtg atg attg geg g teeg gteg gctttcccg g c g g tg g g ca ta ccaccg ata ccaa g gttcag g aa g tgcg gcagctg attg ccga egg eg ctcg gg a ag tgg a tatg gteg tgaa tatcggcaaagtgctctccggcgattatgactatgtgcgcgaagatttgcgccgcgtggtggaagccgccgccccggtaccggc caaagtgatccttgaaacgcactatcttaatgaggagcagattcgtcgcgtgtgcgaaattgccgtagaggtggggatgaagtgg gtcaaaacctctaccggctgggcgcccaccggcgccacggtggaaaaagtcagcatcatcgccgaccagcttaaaggccga atcgatatcaaaggcgccgggggcattcgcgatctggccacggtgcgtgcgctgtaccagcttggcgtgcgccgctttggcatga gccatggcgcggtaaccaaggtgctggctgaactggagcagcacccggaacgttttcccgagctgaacgccgattaa

[0200] Protein Sequence:

[0201] SEQUENCE ID NO. 30

[0202] M KAI KNKGQDKTM NETTRRFARLVDLSAVQATSTEADVRACAELAARYNI ISVHVLPCWTRF LSTLLPQQGTGEVMIGGPVGFPGGGHTTDTKVQEVRQLIADGAREVDMWNIGKVLSGDY DYVREDLRRWEAAAPVPAKVI LETHYLNEEQI RRVCEI AVEVGM KWVKTSTGWAPTGATVE KVSIIADQLKGRIDIKGAGGIRDLATVRALYQLGVRRFGMSHGAVTKVLAELEQHPERFPELN AD

[0203] 12. Putative lipoprotein

[0204] The term putative lipoprotein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.31 .

[0205] Accession Code: KPHS.17750 / A0A0H3GM61

[0206] Gene Sequence:

[0207] SEQUENCE ID NO.12 atgcgctacaaactttttgctgtacttctcccttgcgcgctggcgctcagcgcctgtacgaccgtcaccccggcctataaagataatg gcacccgcagcggcccctgcgtcgaaggcggccctgacgacgtagcgcaaaaattctacgacacccagatccagaatcgta ctca g g a teeg g eg geg ctg eg ccccta tetg ag eg acg geetg geg cag ttactcaatg acg cccg ccag g atcccgccaac agcaaattactgcaggctaatccgttctccagcagcagcaccccggcagacagcgccgtggtcgccagcgcctccaccatccc gaaccgcgatgcgcgcaatattccgctgcgcgtcgatctgaaacagggcacccagagctggaaagatgaagtgctgatgatc caggaaggccagtgctgggccgtcgatgacgttcgctatctcggcaacaacagccatgcgccagccgggacgctgcgacag agtctggagaaacgctga

[0208] Protein Sequence:

[0209] SEQUENCE ID NO.31

[0210] MRYKLFAVLLPCALALSACTTVTPAYKDNGTRSGPCVEGGPDDVAQKFYDTQIQNRTQDPA ALRPYLSDGLAQLLNDARQDPANSKLLQANPFSSSSTPADSAWASASTIPNRDARNIPLRV DLKQGTQSWKDEVLMIQEGQCWAVDDVRYLGNNSHAPAGTLRQSLEKR

[0211] 13. Outer membrane protein assembly factor BamA

[0212] The term BamA encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.32.

[0213] Accession Code: KPHS_09240 / A0A0H3GNJ8

[0214] Gene Sequence:

[0215] SEQUENCE ID NO.13 atggcgatgaaaaagttgctcatagcgtcgctgctgtttagcagcgcgactgtatacggtgctgaagggttcgtggtgaaggacat tcatttcgaaggcttgcagcgtgtcgctgttggtgcggccctcctcagtatgccagtgcgtcctggcgatacggtgaccgacgatga tatcagtaacactattcgcgcgctgtttgccactggcaacttcgaggacgtccgcgtcctgcgcgatggtgataccctgctggtgca ggtgaaagagcgtccgacgatcgccagcatcaccttctccggcaacaagtcggtgaaagatgacatgctgaagcagaacctt gaggcctcaggcgttcgggtgggcgagtcgcttgaccgcacgaccatcgcggatatcgagaagggtcttgaagacttctactac agcgtcggtaaatacagcgccagcgtcaaagcagtcgttacgccgctgccgcgtaaccgtgtcgatttgaagctggtcttccagg a a g g eg teteeg ca aa aa ttcaa cag atcaa catcg teg g ca a ccatg cgttttcg a ccg atg agetg atctcccacttccagctg cgcgatgaggtgccgtggtggaacgtggtcggcgaccgtaaataccagaagcagaagctagcgggcgaccttgaaaccctg cgcagctactacctggatcgcggctatgcccgtttcaacatcgattctacccaggtcagcctgacgccggataagaaagggatct acatcaccgtcaacatcaccgaaggcgatcagtacaagttttccggagtgcaggtgacgggcaacctcgctggccattccgcg gaaatcgaagcgctgactaaagttgagccaggcgaactgtacaacggcgcgaaagtgaccaggatggaaaacgacatcaa gaaactgttgggtcgttatggttacgcctatccgcgcgtgcagtcgcagccggagatcaacgacagcgataaaaccgttaagct gcacgttaacgtcgacgcaggcaaccgttattacgtgcgtaaaattcgcttcgaaggcaacgacacctctaaagatgccgtactg cgccgcgaaatgcgccagatggaaggcgcatggctgggcagcgacctcgtcgatcagggtaaagaccgtctcaatcgtttag gtttctttgaaacggtggatactgatacccagcgcgtgccgggcaacccggaccaggtcgacgttgtctacaaggtgaaagagc gtaacaccggtagcttcaacttcggtatcggctacggcaccgagagcggcgtcagcttccaggcgggcgttcagcaggataact ggttaggtactggctatgctgtcgggatcaacggtaccaaaaacgactaccagacctataccgagctgtcggtgaccaacccgt acttcaccgtagacggtgtaagcctcggcggtcgtgtcttctataatgactttgatgcaaacgatgcggatctgtctgactataccaa caaaagctatggtacagacattacgctgggcttcccggtcaatgaatacaacacgctgcgcgccggcgtcggttatgtgcataac tccctgtccaatatgcagccgcaggtggcaatgtggcgttaccttaactcgatgggccagtatccggacaacaccaacgaccgg aactcgttcagtgcgaatgacttcaccttcaactacggttggacctataacaagcttgaccgcggcttcttcccaacggaaggttcg cgcgtcaacctgaacggtaaggtgaccattccgggctcagacaacgagtactacaaagcgacgctggataccgcgacctacg tgccgatcgacaacgatcatcagtgggtagtactgggtcgtacgcgctttggttatggcgatggtatcggcggcaaagagatgcc gttctatgagaacttctatgccggtggttccagcaccgtgcgtggcttccagtcgaacaccattggtccgaaggcggtgtacttccc gtcgagcagccgtcatgatggcgatagcggttataccaatgactgtaagagcaccgaatccgcaccgtgtaaatccgatgatgc ggttggcggtaacgcgatggcggtggccagccttgagctgattaccccgacgccgtttattagtgacaaatatgcgaactcggtc cggacttccgtcttctgggatatgggtaccgtatgggatactcactgggattcgagcgcgtacgctggttatccggattacagcgat ccgagcaacatccgtatgtctgcgggtattgccgtgcagtggatgtcgccgttggggccgttggtcttctcctacgcccaaccgttca aaaagtacgatggagacaaagccgaacagttccagtttaacattggtaaaacctggtaa Protein Sequence:

[0216] SEQUENCE ID NO.32

[0217] MAMKKLLIASLLFSSATVYGAEGFWKDIHFEGLQRVAVGAALLSMPVRPGDTVTDDDISNTI

[0218] RALFATGNFEDVRVLRDGDTLLVQVKERPTIASITFSGNKSVKDDMLKQNLEASGVRVGESL

[0219] DRTTIADIEKGLEDFYYSVGKYSASVKAWTPLPRNRVDLKLVFQEGVSAKIQQINIVGNHAF

[0220] STDELISHFQLRDEVPWWNWGDRKYQKQKLAGDLETLRSYYLDRGYARFNIDSTQVSLTP

[0221] DKKGIYITVNITEGDQYKFSGVQVTGNLAGHSAEIEALTKVEPGELYNGAKVTRMENDIKKLL

[0222] GRYGYAYPRVQSQPEINDSDKTVKLHVNVDAGNRYYVRKIRFEGNDTSKDAVLRREMRQM

[0223] EGAWLGSDLVDQGKDRLNRLGFFETVDTDTQRVPGNPDQVDWYKVKERNTGSFNFGIGY

[0224] GTESGVSFQAGVQQDNWLGTGYAVGINGTKNDYQTYTELSVTNPYFTVDGVSLGGRVFYN

[0225] DFDANDADLSDYTNKSYGTDITLGFPVNEYNTLRAGVGYVHNSLSNMQPQVAMWRYLNSM

[0226] GQYPDNTNDRNSFSANDFTFNYGWTYNKLDRGFFPTEGSRVNLNGKVTIPGSDNEYYKAT

[0227] LDTATYVPIDNDHQWWLGRTRFGYGDGIGGKEMPFYENFYAGGSSTVRGFQSNTIGPKAV

[0228] YFPSSSRHDGDSGYTNDCKSTESAPCKSDDAVGGNAMAVASLELITPTPFISDKYANSVRT

[0229] SVFWDMGTVWDTHWDSSAYAGYPDYSDPSNIRMSAGIAVQWMSPLGPLVFSYAQPFKKY

[0230] DGDKAEQFQFNIGKTW

[0231] 14. Malate dehydrogenase

[0232] The term malate dehydrogenase encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.33.

[0233] Accession Code: KPHS_47750 / A0A0H3GWA1

[0234] Gene Sequence:

[0235] SEQUENCE ID NO.14 atgaaagttgcagtccttggcgctgccggtggtatcggccaggcgcttgccctactacttaagacccaactgccttcaggttcaga gctctcgttgtacgacatcgctccggttacgccgggcgtggcggtagatctaagtcatatccccacagatgtaaaaattaaaggat tttccg g tg a ag acg ctactccg gcg ctg g aa g gcg egg atg ta gtg etg a teteeg eg g geg tgg eg eg ta ag cccg g catgg atcgttccgacctgtttaatgtgaatgcgggtatcgtgaagaacctcgtgcagcagattgccaaaacctgcccgcaggcctgcatc ggcattatcaccaacccggtgaataccaccgtggctatcgccgccgaagtactgaaaaaagccggcgtgtacgataaaaaca a a ctg tteg g eg ttacca egetg ga catca teeg ttcca ata cctttg tgg eg ga getg aaa gg ta aa tegg caaccga g gtg ga agtcccggtcattggtggtcactccggggtcaccattctgcctttactgtcgcagatccccggcgtcagctttagcgatcaggaaatt gccgacctgactaaacgtattcagaacgccggtaccgaagtcgtggaagcgaaagcgggcggcgggtcggcgaccttgtcg a tg g g ccag g eg getg cccg ttttg gtctctctctg gtteg eg ccatg cag g gg g aa a aa g gcg tgg tgg a gtg cgcctacg tgg aaggcgacggccactatgcgcgtttcttctcccagccgctgctgctggggaaaaacggcgtggaagagcgacagtctatcggc aaactcagcgcctttgaacagcaggcgctggagggcatgctggacaccctgaagaaagatatcgcgctcggcgaagacttcg tcaacaagtaa

[0236] Protein Sequence:

[0237] SEQUENCE ID NO.33

[0238] MKVAVLGAAGGIGQALALLLKTQLPSGSELSLYDIAPVTPGVAVDLSHIPTDVKIKGFSGEDAT PALEGADWLISAGVARKPGMDRSDLFNVNAGIVKNLVQQIAKTCPQACIGIITNPVNTTVAIA AEVLKKAGVYDKNKLFGVTTLDIIRSNTFVAELKGKSATEVEVPVIGGHSGVTILPLLSQIPGV SFSDQEIADLTKRIQNAGTEWEAKAGGGSATLSMGQAAARFGLSLVRAMQGEKGWECAY VEGDGHYARFFSQPLLLGKNGVEERQSIGKLSAFEQQALEGMLDTLKKDIALGEDFVNK

[0239] 15. Alcohol dehydrogenase

[0240] The term alcohol dehydrogenase encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.34.

[0241] Accession Code: KPHS_28230 / A0A0H3GTY9

[0242] Gene Sequence:

[0243] SEQUENCE ID NO.15 atgaaggcagctgttgttacccacgaccatcaggttaacgtcacggaaaaaacgctgcgcccgctggaatacggcgaagcgct gttgaaaatggaatgctgcggcgtgtgtcatactgacctgcacgtgaaaaacggcgattttggcgataaaaccggcgtcattctcg gccatgaagggatcggggtggtacaaaaagtcggcccgggcgtcacctccctgaagccgggcgaccgcgccagcgtggcgt ggttcttcgaaggctgcggccactgcgattactgtaacagcggcaacgagacgctctgccgctcggtgaaaaacgccggctata ccgtcgatggcggcatggcggaagagtgcatcgtcaccgccaactacgcggtaaaagttccggacggcctcgactccgccgc cgccagcagcatcacctgcgcgggcgtcaccacctacaaagcggtcaaggtctcccacatcaaaccgggccagtggatcgcc atctacggcctcggcgggttgggtaacctcgcgctgcagtatgcgaagaatgtctttaacgccaaagtgatcgctatcgacgtca acgacggacagctggagctggcggcctcgatgggcgccgacctgaccatcaactcccgcaatgaagatgcggcgaaagtga ttcaggaaaaaaccggcggcgcccacgctgcggtagtaaccgcggtggctaaagcggcctttaactcggcggtggatgccgtt cgcgccggtggccgcgtggttgcggtgggcctgccgccggaggcgatgagcctcgatattccgcgtctggtgctggacggcatc gaggtggtcggttcgctggtcggcacccgtcaggatctggtggaagccttccagtttgccgccgaaggcaaagtggtgccgaaa gtcaccctgcgtccgctggaagatatcaatgctatcttcaaagagatggagcaaggtcagatccgcggccgtatggttatcgatct gcgtagctaa

[0244] Protein Sequence:

[0245] SEQUENCE ID NO.34

[0246] MKAAWTHDHQVNVTEKTLRPLEYGEALLKMECCGVCHTDLHVKNGDFGDKTGVILGHEGI GWQKVGPGVTSLKPGDRASVAWFFEGCGHCDYCNSGNETLCRSVKNAGYTVDGGMAE ECIVTANYAVKVPDGLDSAAASSITCAGVTTYKAVKVSHIKPGQWIAIYGLGGLGNLALQYAK NVFNAKVIAIDVNDGQLELAASMGADLTINSRNEDAAKVIQEKTGGAHAAWTAVAKAAFNSA VDAVRAGGRWAVGLPPEAMSLDIPRLVLDGIEWGSLVGTRQDLVEAFQFAAEGKWPKVT LRPLEDINAIFKEMEQGQIRGRMVIDLRS

[0247] 16. Outer Membrane Lipoprotein SlyB

[0248] The term outer membrane lipoprotein SlyB encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or

[0249] 99% sequence identity with SEQUENCE ID NO.35.

[0250] Accession Code: KPHS_29580 / A0A0H3GUH2

[0251] Gene Sequence: SEQUENCE ID N0.16 atgattttacgtgttttggctgtatcgatgattggttttacgctcgcaggctgcgtgagcagcagcggcctgtccggcgacgtttattcc gcatctgaagccaaacaggtgcagagcgtgacttacggtactatcgtgcatacgcgcgccgtacagatccagagcggcgacg atagcaatgccatcggggccattggcggtgcggtactgggcgggttccttggaaacaccatcggcggcggtaccggacgctctc tggctacggcggcaggtgcggtcgcgggcggcgtagccggtcagggcgttcagggcgcgatgaataaaacgcagggcgttg aactggaaatccgtaaggacgatggcaataccattatggtagtgcagaaacagggcagcactccgttctccgttggtcagcgcg tg g ccattg ccg g cag eg g cag cca g g tea ccg tctctccg eg ata a

[0252] Protein Sequence:

[0253] SEQUENCE ID NO.35

[0254] M I LRVLAVSM I G FT LAGCVSSSGLSG DVYSASEAKQVQSX / TYGT I VHT RAVQI QSG DDSNAI GAIGGAVLGGFLGNTIGGGTGRSLATAAGAVAGGVAGQGVQGAMNKTQGVELEIRKDDGN Tl M WQKQGSTPFSVGQRVAI AGSGSQVTVSPR

[0255] 17. long-chain fatty acid outer membrane transporter

[0256] The term long chain fatty acid outer membrane transporter protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.36.

[0257] Accession Code: KPHS_37950 / A0A0H3GT78

[0258] Gene Sequence:

[0259] SEQUENCE ID NO.17 atgggcatgagccagaaaacccgctttacccaatctgcccttgcagtggctgtcgcacttgtttccacccaggcctggtcagcagg cttcca g ctca acg a attetetg catcg gg g etg g gteg tgcctattccgg tg aa gg g geg ateg etga ega tg etg g ta a tg cca gtcgtaacccggcattaattatgatgttcgaccgcccgaccatgtcggcgggcgccgtgtttgtcgacccgggcgttaatgtttcgg gcacctctcccaccggcaagagcctgaaagcagacaatatcgccccgacggcgtgggtgcccaacttccactttgtcgcgccg attaacgaccaatttggttggggggcgtctatcacctctaactatggtctggcaacagagtataacgatgattatgcggccggcag catgggcggtaaaaccga cctg a ccaccg eg a aett caa cctga g eg g cgcctaccg cctcg a ca g caa etgg a gettegg c ttaggctttgacgcggtctaa

[0260] Protein Sequence:

[0261] SEQUENCE ID NO.36

[0262] MGMSQKTRFTQSALAVAVALVSTQAWSAGFQLNEFSASGLGRAYSGEGAIADDAGNASRN PALI M M FDRPTMSAGAVFVDPGVNVSGTSPTGKSLKADNIAPTAWVPNFHFVAPI NDQFGW GASITSNYGLATEYNDDYAAGSMGGKTDLTTANFNLSGAYRLDSNWSFGLGFDAV

[0263] 18. putative porin

[0264] The term putative porin protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.37.

[0265] Accession Code: KPHS_06190 / A0A0H3GMR2 Gene Sequence:

[0266] SEQUENCE ID N0.18 atgagaaagcttaacatccttattcttgcagccctgaccgcggtctctggttccgccatggcggtggggtttaccgttgaacagggta aaaacttcactaatctgaatatggaaatgggtaaatcctcttctggcctgtacgccgaaagccactggctgaagaataccgatgat ggtagccagaccggcggcgtgggcgcgggttacaacctggaagtgggcccggtgatgctcaacgccggcgcgaaagccatc tacctcgggccgaaaaaaggcgataacggcgtcgcgtttccggtcggcggcggggtgaacgtgaccctgaccgacagcatcc acgtgtttggcgaagggtatgtcgcgccggatggtctgaacaacagcgtgaaaaactatgttgaagcgaacggcggcgtgagc tggtctccgattggtccagtgacgttgaaagtgggttaccgccatgtgagcgttgacggtaaagagggtcgcccgaaccacacc ctg a ttg a tg g eg ccta tg t g g g eg g eg g g g tea ccttctg a

[0267] Protein Sequence:

[0268] SEQUENCE ID NO.37

[0269] MRKLNILILAALTAVSGSAMAVGFTVEQGKNFTNLNMEMGKSSSGLYAESHWLKNTDDGSQ TGGVGAGYNLEVGPVMLNAGAKAIYLGPKKGDNGVAFPVGGGVNVTLTDSIHVFGEGYVA PDGLNNSVKNYVEANGGVSWSPIGPVTLKVGYRHVSVDGKEGRPNHTLIDGAYVGGGVTF

[0270] 19. Maltoporin

[0271] The term long maltoporin protein encompasses this protein and therapeutically effective variants that share at least 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% sequence identity with SEQUENCE ID NO.38.

[0272] Accession Code: KPHS_02600 / A0A0H3GHG7

[0273] Gene Sequence:

[0274] SEQUENCE ID NO.19 atgatgattactctgcgcaaacttcctctggcggtcgccgtcgcagcaggcgtgatgtctgctcaggcgctggctgtcgatttccatg gctacgcgcgttccggcattggctggaccggcagcggcggcgagcaacagtgcttcaaagcaaccggcgctcaaagtaaata ccgtcttggtaacgaatgtgaaacctatgcggaactgaagctgggccaggagctgtggaaggaaggggataagagtttttatttc gatactaacgttgcctattccgtgaatcaggaagatgactgggaaagcacctctccggcgttccgtgaagccaacatccagggt aaaaacctgatcgactggctgccgggctccacgctgtgggcgggtaaacgcttctatcagcgtcatgacgttcacatgatcgactt ctactactgggatatctccggcccgggtgcaggtctggaaaacgttgaccttggcttcggtaagctctctctggccgctacccgtaa ctcagaaagcggcggctcttatactttctccagcgatgacaccaaaaaatatgctgcgaaaactgccaacgacgtctttgatatcc gtctggcgggtctggaaaccaacccgggcggcgtgctggagttaggggtcgattacgaccgtgctaacccgcaggatgactat cgcctggaagacggcgcgtcgaaagacggctggatgtggaccggtgaacatactcagtctatctggggcggcttcaacaagttt gtggttcagtacgccactgacgcaatgacctcctggaacagcggccactctcaggggaccagcatcgataacaacggcagca tgatccgcgttctggatcacggcgcgatggacttcaacgatgactggggcctgatgtacgtggcaatgtaccaggacgtggatct ggacagcaagaacggttctacctggtacaccgtgggtgtccgtccgatgtacaaatggacgccgatcatgagcacccagctgg aaatcggttacgacaacgtgaaatcccagcgtaccagcgaaaacaacaaccagtacaaaattactctggctcaacagtggca ggcaggcaacagcgtctggtctcgtccggctatccgtatcttcgcaacctacgcgaagtgggatgaaaactggggctacagcaa cacctccggtctgcagacgaaagacagcagcggaagcggcgctttcacctccagccgcggtgacgacagcgaagttaccttc ggtgcccagatggaagtgtggtggtaa

[0275] Protein Sequence:

[0276] SEQUENCE ID NO.38

[0277] MMITLRKLPLAVAVAAGVMSAQALAVDFHGYARSGIGWTGSGGEQQCFKATGAQSKYRLG NECETYAELKLGQELWKEGDKSFYFDTNVAYSVNQEDDWESTSPAFREANIQGKNLIDWLP GSTLWAGKRFYQRHDVHMIDFYYWDISGPGAGLENVDLGFGKLSLAATRNSESGGSYTFS SDDTKKYAAKTANDVFDIRLAGLETNPGGVLELGVDYDRANPQDDYRLEDGASKDGWMWT GEHTQSIWGGFNKFWQYATDAMTSWNSGHSQGTSIDNNGSMIRVLDHGAMDFNDDWGL MYVAMYQDVDLDSKNGSTWYTVGVRPMYKWTPIMSTQLEIGYDNVKSQRTSENNNQYKIT LAQQWQAGNSVWSRPAIRIFATYAKWDENWGYSNTSGLQTKDSSGSGAFTSSRGDDSEV TFGAQMEVWW

[0278] The invention will now be described with reference to specific Examples. These are merely exemplary and for illustrative purposes only: they are not intended to be limiting in any way to the scope of the monopoly claimed or to the invention described. These examples constitute the best mode currently contemplated for practicing the invention.

[0279] EXAMPLES

[0280] MATERIALS AND METHODS

[0281] Determination of adhesion of K. pneumoniae strains to human lung epithelial cells by colony counting

[0282] Human A549 cells or 16HBE14o- cells were seeded in uncoated or coated 24-well plates respectively at a density of 4 x 105cells / well overnight. Bacteria were cultured in 10 m L LB broth overnight and then transferred into a 250 m L conical flask containing 100 m L of fresh LB broth and incubated at 37 °C with agitation at 200 rpm to ODeooO.6-0.8. Mid-logarithmic phase cultures were diluted to 2 x 107(MOI 50: 1 ), 4 x 106(MO1 10:1 ) or 2 x 106(MOI 5:1 ) cells / well in 500 pL of prewarmed fresh media (RPMI or MEM) without antibiotics. The seeded lung epithelial cells were washed with one m L of prewarmed empty media, the bacteria were added into each well in duplicate. The 24-well plates were centrifuged at 300 x g for five minutes before incubating for 30 minutes at 37°C in 5% CO2 to allow the bacteria to attach to the cells. The wells were washed four times by gently adding 500 pL prewarmed sterile PBS to the cells followed by aspiration to remove the unattached bacteria. To remove the attached cells, 500 pL lysis buffer (0.5 % Triton X-100 in PBS) was added to each well and incubated for 20 minutes at RT. The lysates were serially diluted in PBS and plated onto LB agar and were incubated overnight at 37°C and CFU determined.

[0283] Outer membrane protein (OMP) preparation

[0284] Stationary phase bacterial pellets were harvested by centrifuging at 10,000 x g for 1 h at 4 °C (Cl P52.145 strain) or at 4000 x g for 10 minutes at 4 °C (Acps strain). After centrifugation, the pellets were resuspended in 10 m L bacteria lysis buffer containing 50 mM Tris (pH 8.0) with 400 pL protease EDTA-free inhibitorcocktail. Then the resuspended bacteria were sonicated for 10 minutes for 30 second intervals with 30 second rests before being centrifuged at 12,000 x g for 10 mins at 4 °C. The supernatants were retained, transferred to fresh tubes, and treated with 1 pL DNase for 20 mins at RT. The supernatants were centrifuged at 12,000 x g for lO mins at 4 °C again, and then transferred to a fresh tube. After solubilising the supernatants in 2 % sarkosyl solution for 30 mins at 25 °C, the whole solution was centrifuged at 20,500 x g for 50 mins at 4 °C. The supernatants were discarded, and the pellets were resuspended in 1 mL2-D lysis buffer containing 8 M Urea, 2 M Thiourea, 4 % CHAPS, 1 % Triton X, 10 mM Tris base, 65 mM DTT, 0.8 % immobilised pH gradient (IPG) buffer (pH 3-11 NL) and stored at - 20 °C until required.

[0285] Protein separation by 2-D gel electrophoresis

[0286] The OMP fractions of each strain were solubilised for isoelectricfocusing (IEF) in rehydration buffer containing 8 M urea, 2 M thiourea, 4 % CHAPS, 1 % triton X, 10 mM Tris base, 65 mM DTT, 0.8 % immobilised pH gradient (IPG) buffer (pH 3-11 NL) and a trace of bromophenol blue. IPG dry strips, 7 cm long (pH 3-11 NL or pH 4-7NL) were rehydrated overnight at RT with 125 pL of the rehydration solution containing 200 pg of OMPs, and IEF performed as follows: three hours at 300 V for 30 mins, 1 ,000 V for 30 mins and 5,000 V for 4 hours. The IPG strips were then equilibrated in reducing buffer containing 30 % glycerol, 2 % SDS, 6 M urea, 50 mM Tris and 2 % DTT for 20 minutes at RT. The IPG strips were then alkylated for 20 minutes in a similar buffer, containing 2.5 % iodoacetamide instead of DTT and a trace of bromophenol blue and placed on 10% or 12 % SDS-PAGE gels. Separation was performed at 110 V, 100 mA. Two gels were performed in parallel; one was stained, and another was transferred led to PVDF membrane as described below. Proteins were visualised using PageBlue protein stain (Thermo Fisher Scientific) by staining overnight at RT with agitation, followed by destaining overnight in sterile dH2O.

[0287] Detection of Klebsiella pneumoniae adhesins using the Cell blot proteomic platform

[0288] The proteins separated on 2-D gels were transferred to PVDF membranes and the membranes blocked overnight at 4°C with 5% BSA and 3% Marvel® in PBS with gentle rotation. Human 16HBE14o- cells were scraped from T75 flasks and resuspended in five mL PBS with a final concentration of 1 xi o6cells / ml and were incubated with the membranes for 4 hours at 4°C with gentle agitation. The membranes were rinsed with 20 m L of 0.7% (v / v) PBS-Tween 20 (PBS-T) five times, and then any bound epithelial cells were fixed with 3% paraformaldehyde in PBS for 8 minutes. After that, the membranes were incubated with a mouse-anti-epithelial antibody (Millipore) (1 :1000 in 5% (w / v) BSA in 0.04% (v / v) PBS-T) (Merck) overnight at 4°C with gentle rotation. The membranes were then washed five times with 0.7% (v / v) PBS-T. This was followed by the addition of a secondary HRP conjugated antibody (1 :36,000) in 5% BSA in 0.04% PBS-T. The secondary HRP rabbit anti-mouse-IgG antibodies were incubated for 1 hour at RT with gentle rotation. The membranes were then subsequently washed five times with 0.7% PBS-T before chemiluminescence detection was carried out with luminol reagent.

[0289] Mass spectrometry for Protein identification.

[0290] The identification of proteins by Q-Exactive™ was carried out by Mass Spectrometry Core Facility, UCD Conway Institute as published before with modifications. The tryptic peptides after were then dried in a vacuum centrifuge and resuspended in 20 pL of loading buffer containing 2.5 % acetonitrile and 0.5 % acetic acid in LC-MS grade water to be analysed by the Mass Spectrometry Core service at the Conway Institute (University College Dublin). Peptides were separated on C18 home-made column (C18-AQ Dr. Maisch Reprosil-Pur 100 x 0.075 mm x 3 pm) over 180 min at a flow rate of 250 nL / min with a linear gradient of increasing ACN from 1 % to 27%. The mass spectrometer was operated in data dependent mode; a high resolution (70,000) MS scan (300-1600 m / z) was performed to select the twelve most intense ions and fragmented using high energy C-trap dissociation forMS / MS analysis. The protein spectra were obtained by Q-Exactive analysis and matched with the K. pneumoniae reference proteome from HS11286 strain (UniProt accession number UP000007841 ).

[0291] Assessment of protection of antigen candidates using an immunisation and K. pneumoniae sepsis challenge model

[0292] The antigens were examined for their potential to protect mice in a sepsis model. Briefly, female C57BL / 6 mice, originally purchased from Charles River Laboratories (UK), were separately into groups (n = 8 or n = 6). All mice were immunised subcutaneously with 100 uL of SAS adjuvant only control or 50 pg recombinant antigen with SAS adjuvant on Day 0 and received boost s.c. on Days 14 and 28. Mice were monitored every day for 3 days following procedure and every 2 days thereafter. A log phase culture of K. pneumoniae Cl P 52.145 was washed and resuspended in sterile endotoxin -free Dulbecco's phosphate-buffered saline (DPBS) and diluted based on the ODeoo to a final concentration of 1 x 105CFU / mL. Prepared inoculums were plated on the LB agar plates with 100 pL / mL of ampicillin (AMP100 LB agar plates) to confirm CFU dose delivered. Mice were challenged intraperitoneally with 100 pLof inoculum using a 30G needle. Mice were observed every 6 hours for symptoms. At 24 h mice were culled by cervical dislocation. The fur and skin were removed, and the abdomen cavity was injected with ice cold, sterile DPBS. An incision was made in the abdomen to release the DPBS into a falcon. The lavage was serially diluted, plated on low salt AMP100 LB agar and incubated at 37 °C overnight. The spleen was removed and put into two m L of cold DPBS and were transferred to sterile homogenizertubes (with beads) and filled with 1 .5 mL PBS. They were homogenized at 5800rpm, 2 x 15 seconds cycle with 30 seconds pause. The homogenates were diluted and plated as described above.

[0293] Determination of antigen-specific IgG antibodies raised in response immunisation using indirect ELISA

[0294] Purified recombinant antigens (LysM or OmpPI ) were diluted to 0.5 pg / mL in coating buffer containing 0.2 M sodium carbonate, pH 9.6 and each well was coated with 100 pL buffer and incubated overnight at 4°C. The following day, the coating solution was removed and the 96- well plates were washed three times with 300 pl per well washing buffer(1 X PBS and 0.05% Tween) on a plate washer. One hundred mL of blocking buffer (1 X PBS and 1 % FBS) was added into each well. The plates were covered and blocked for 1 h at RT. Each serum sample was serially diluted down (1 :10) in the plate in fresh blocking buffer. After incubating 2 h at RT, the plates were washed three times with 300 pL / well washing buffer (1 X PBS and 0.05 % Tween). The wells were incubated with 100 pL of 1 :5000 dilutions of one of the following secondary HRP-conjugated antibodies in blocking buffer: anti-mouse lgG2a-HRP, antimouse lgG1-HRP or anti-mouse Total IgG-HRP. After incubating for 60 minutes at RT, the plates were washed 3 times as before and then blotted. Two additional washes were applied by multichannel pipette with 400 pL per well PBS only, to remove Tween-20 and avoid bubbles. The plates were blotted as much as possible afterwashing to remove excess liquid. The TMB substrate solution (100 pL) was added to each well and the plates were incubated at room temperature in darkforaround 15 mins until the colourdeveloped. The reactions were stopped by adding an equal amount (100 pL) of 2 M sulfuric acid. The absorbances were read at 450 nm immediately.

[0295] Immunoprofiling the antigen-specific recall responses by flow cytometry

[0296] Groups of six- to eight-week-old male BALB / c mice (n=8, Janvier, France) were subcutaneously (s.c.)immunised on DayOwith 50 pg of recombinant antigen with SAS or with SAS adjuvant only in a final volume of 100 pl. Fourteen days afterthe immunisation, all mice were culled by carbon dioxide exposure. Spleens were collected and single-cell suspensions of splenocytes prepared as previously described

[0038] , The cells were re-stimulated with 50 pg I ml purified recombinant protein and incubated at 37 °C in 5 % CO2environment for approximately 60 h. The flow cytometric analysis was carried out by a Beckman Coulter CytoFLEX LX (NUV full configuration). Quality control of the instrument was performed using Beckman Coulter Daily QC beads and IR Daily QC priorto the sample analysis. Data analysis was carried out by Beckman Coulter CytExpert software v.2.4 as described previously (Tomas- Cortazar, et al., BpOmpW Antigen Stimulates the Necessary Protective T-Cell Responses Against Melioidosis. Front. Immunol., 2021. 12: p. 767359).

[0297] RESULTS

[0298] Attachment to host epithelial cells.

[0299] The abilities of two K. pneumoniae strains, wild-type strain, CIP52.145, and capsule mutant strain, CIP52.145 eps to bind to two independent human lung epithelial cell lines, A549 and 16HBE14o-were measured, to examinetheirsuitability as cell lines to identify K. pneumoniae adhesins. The deletion of cps previously demonstrated the decrease of virulence compared to the wild-type strain. The K. pneumoniae CIP52.145 strain showed 7.3-fold higher binding to 16HBE14O- cells relative to the CIP52.145 Acps strain (two-way ANOVA p = 0.0215) (MOI 50:1 ). In contrast, the CIP52.145 eps strain showed 4.7-fold more attachment to A549 cells than the K. pneumoniae CIP52.145 strain ( two-way ANOVA p = 0.0081 ) (MOI 50:1 ). (Figure 1 ). Both cell lines were therefore used as target cell lines in the proteomic identification of K. pneumoniae to lung epithelial cells.

[0300] Identification of proteins involved in Klebsiella pneumoniae CIP52.145 strain and Zlcps strain attachment lung epithelia cells using the cell blot proteomic platform.

[0301] More than 50 protein spots were observed in the 2-D gel separation of K. pneumoniae CIP52.145 afterstaining with PageBlue. Most ofthespots were identified in the pH4 - 7 range and the 20 kDa -150 kDa range. The intensity of protein spots indicated the expression level of each protein. When a blot prepared in parallel was probed with A549 cells, 12 clear spots were identified as being positive for host cell attachment. These were excised, pooled and identified by LC-MS in three independent experiments and the results are summarised in Table 1 . In summary, 12 spots were identified on the gel of K. pneumoniaeCl P52.145adhesins bound toA549 cells. After samples were pooled and MS identification, 21 proteins with more than 20 % unique sequence coverage or more than four unique peptides were selected.

[0302]

[0303] Table 1 : K. pneumoniae Cl P52.145 proteins identified as involved in attachmentto A549 cells.1Unitprot code; theoretical isoelectric points were determined by Expassy ProtParam model;3Molecularmass as determined by Q-Exactive LC-MS using the K. pneumoniae HS11286 database;4Number of unique peptides identified per sample.

[0304] The 2-D gel separation of the K. pneumoniae CIP52.145 strain after staining with PageBlue (Figure 3) showed that most of the adhesins were identified in the pH 4 - 7 range, consequently, IPG with a range of pH 4-7NL was used in later studies to improve the separation of individual proteins. When blots prepared in parallel were probed with 16HBE14o- cells, 15 clear spots were observed as being positive forcellular attachment (Figure 3), which were excised and identified by LC-MS. Three independent experiments were carried out and the results are summarised in Table 2. In summary, 15 spots identified on the gel of K. pneumoniae Cl P52.145 adhesins bound to 16HBE14o- cells were pooled and identified by MS, 16 proteins were selected with the cut-off more than 20 % unique sequence coverage or with more than four unique peptides.

[0305] Table 2: K. pneumoniae CIP52.145 proteins identified as involved in attachment to 16HBE14O“ cells.1Unitprot code; theoretical isoelectric points were determined by Expassy P rot Pa ram model;3Molecular mass as determined by Q-Exactive LC-MS using the K. pneumoniae HS11286 database;4Number of unique peptides identified per sample. More than 20 protein spots were observed in the 2-D gel separation of the Acps membrane proteins after staining with PageBlue (Figure 4). Most of the proteins were identified in the pH 4 - 7 range and between 20 kDa to 150 kDa range as before. When blots prepared in parallel were probed with A549 cells, 15 clear spots were detected as being positive for cellular attachment (Figure 4), which were excised and identified by LC-MS. Three independent experiments were carried out and the results are summarised in Table 3. In summary, 15 spots were identified on the gel of K. pneumoniae CIP Acps adhesins bound to A549 cells, which were pooled and identified by MS, finally, 22 proteins with more than 20 % unique sequence coverage or with more than four unique peptides were selected .

[0306] Table 3: K. pneumoniae Acps proteins identified as involved in attachment to A549 cells.1Unitprot code; theoretical isoelectric points were determined by Expassy ProtParam model;3Molecularmass as determined by Q-Exactive LC-MS using the K. pneumoniae HS11286 database;4Number of unique peptides identified per sample .

[0307] The 2-D gel separation of K. pneumoniae Acps strain after staining with PageBlue (Figure 5) showed that most of the adhesins were identified in the pH 4 - 7 range, consequently, IPG with a range of pH 4-7NL was used in later studies to improve the separation of individual proteins. When blots prepared in parallel were probed with 16HBE14o- cells, 11 clear spots were identified as being positive for cellular attachment (Figure 5), which were excised and identified by LC-MS. Three independent experiments were carried out and the results are summarised in Table 4. In summary, 11 spots were identified on the gel of K. pneumoniae CIPAcps adhesins bound to 16HBE14o- cells. After samples were pooled and MS identification, 17 proteins with more than 20 % unique sequence coverage or with more than four unique peptides were selected. Negative controls were performed for both strains and confirmed there was no non-specific interactions between the antibodies and the bacterial proteins (Figure 6).

[0308] Table 4: K. pneumoniae Acps proteins identified as involved in attachment to 16HBE14o- cells.1Unitprot code; theoretical isoelectric points were determined by Expassy ProtParam model;3Molecularmass as determined by Q-Exactive LC-MS using the K. pneumoniae HS11286 database;4Number of unique peptides identified per sample.

[0309] Confirming the role of three K. pneumoniae antigens in attachment to epithelial cell lines

[0310] The antigens were cloned and individually expressed in E. coli 8 .2 cells to confirm that they are directly involved in host cell attachment. The expression of each antigen was induced by IPTG and the attachment of IPTG-induced BL21 cells expressing the recombinant antigen to human 16HBE14o- cells was compared to the IPTG induced BL21 cells transformed with empty pET100 / D-TOPO. As shown in Figure 7, a significant 14.3-fold, 6.48-fold and 7.22- fold increase in attachment to 16HBE14o_cells was observed with IPTG- induced BL21 cells expressing LysM / BON protein (p = 0.012), OMPP1 protein (p = 0.0199) and Dps protein (p=0.0011 ) respectively, compared to IPTG-induced BL21 cells transformed with empty pET100 / D-Topo, which confirms that the three antigens tested, LysM / BON protein, OMPP1 protein and Dps protein are involved in host cell attachment.

[0311] Serological analysis of immune response LysM / BON and OMPP1 immunisation in mice

[0312] Serological analysis after the immunisation, followed by two boosts 2 weeks apart, indicates that LysM / Bon and OmpP1 were highly immunogenic as strong antigen -specific IgG titers were raised in mice immunised with either antigen compared with the IgG responses raised in mice treated with adjuvant alone (Figure 8). In both cases the mice showed strong seroconversion to the antigens. The serological response for LysM / BON was dose -dependent, with higher titres being raised as immunisation dose increased. The 50 pg LysM / BON immunised group and 100 pg LysM / BON immunised groups showed higher IgG titresthan the lower dose of 20ug (Figure 8A). The antibody titres elicited by three different doses of LysM / BON or a single dose (50 pg) of OMPP1 indicate the potential expression of high levels of antigen-specific antibodies.

[0313] Protection from K. pneumoniae infection in a murine sepsis model.

[0314] A sepsis infection model to test the vaccine potential of LysM / BON, OMPP1 and Dps. Briefly, mice were immunised subcutaneously and received two boosts before being challenged intraperitoneally. The bacterial burden in peritoneal cavity and spleen were counted 24 hours post-infection and proved all three antigens were protective.

[0315] The mice group immunised with LysM / BON, OMPP1 or Dps with SAS adjuvant showed significantly reduced levels (1 .54 log, 2.69 log, 2.50 log, respectively) of colonisation in the peritoneal cavity 24 hours post infection, compared to SAS adjuvant alone groups (Mann- Whitney test, p = 0.0080, p = 0.0007, p = 0.0007, respectively) (Figure 9A). Moreover, the mouse groups immunisedwith LysM / BON, OMPP1 or Dps showed significantly reduced levels (0.86 log, 1.44 log, 1.01 log, respectively) of dissemination to the spleen 24 hours post infection, compared to SAS adjuvant alone groups (Mann-Whitney test, p = 0.0263, p = 0043, p = 0.0003, respectively) (Figure 9B). Both bacterial clearance in peritone al cavity and reduced dissemination to the spleen showed that LysM / BON, OMPP1 and Dps were protective in this sepsis model, with OMPP1 and Dps showing greater protection than LysM / BON.

[0316] Antigen-specific recall responses in LysM / BON-immunised mice stimulated IL-4, IL-17, IL-22, TNF, and IFN-y, whereas OMPP1 primarily induced IL-17

[0317] Both LysM / Bon and OMPP1 stimulated cellular immune recall responses in immunised mice, in addition to antibodies responses. Antigen-specific recall responses in LysM / BON- immunised mice stimulated IL-4, IL-17, IL-22, TNF, and IFN-y, whereas OMPP1 primarily induced IL-17 as detailed below:

[0318] Induction of effector T cells and other immune cells that contribute to the removal of pathogen via cell-mediated effector mechanisms is important for an effective K. pneumoniae vaccine. Antigen-specific recall responses in cultured splenocytes of LysM / BON or OMPP1 immunised mice were stimulated with the respective antigen in vitro and examined by flow-cytometry. There was no significant change in total percentage of CD3+cells expressing both CD4+and CD8+(double positive (DP) cells) (PCT Fig 10a). However, y6+cells and Natural killer (NK) cell percentages were significantly decreased after re-stimulation with LysM / BON in the splenocytes relative to controls, while no significant change was found after re -stimulation with 0MPP1 (PCT Fig 10b-c). The total percentage of NKT cells and CD4+regulatory T cells was significantly increased in the LysM / BON immunised group but not in the 0MPP1 immunised group (PCT Fig 10d-e). Moreover, the levels of Helper T (Th) cells or naive Th cells were significantly decreased in response to LysM / BON re-exposure (PCT Fig 10f-g). A greater number of effector Th cells and memory Th cells were observed in re-stimulated splenocytes from LysM / BON-immunised mice compared to controls (PCT Fig 10h-i). In addition, LysM / BON-re-stimulated splenocytes showed activation of CD4+T cells, compared to splenocytes from adjuvant only control mice (PCT Fig 10j). By contrast, increased Th cell percentage was observed in response to re-stimulation with OMPP1 , with no significant changes in naive Th cells, effector Th cells, memory Th cells or activated Th cells (PCT Fig 10f-j). No significant change was found in cytotoxic T (Tc) cells in response to re -stimulation with LysM / BON or OMPP1 (PCT Fig 10k). LysM / BON-re-stimulated splenocytes showed a significant decrease in cytotoxicT (Tc) cells, butnotin OMPPI -re-stimulated splenocytes (PCT Fig 101). A significant increase in effectorTc cells, memory Tc cells and activated Tc cells was observed after re-stimulation with LysM / BON in the splenocytes but not in response to OM PP1 re-exposure (PCT Fig 10m-o).

[0319] Cytokine analyses showed that LysM / BON, but not OMPP1 , induced the expression of IL -4, I L-17, 1 L-22, TNF and IFN-y in Th cells (PCT Fig 11 a-e) and Tccells (PCT Fig 11 f-i), compared with the splenocytes from adjuvant only treated mice, while OMPP1 had no significant effect on any of these parameters. yb+T cells play an important role in preventing dissemination of K. pneumoniae during the initial pulmonary infection by producing IFN-y and TNF. Splenocyte cytokine responses from LysM / BON immunised mice showed upregulated IL-4, IL-22, TNF and IFN-y in yb+T cells compared with the control group splenocytes, while the expression of I L-17 was unaltered (PCT Fig 11 j-n). Previous studies showed that NK cells were the major source of IL-22 and IFN-y during K. pneumoniae infection. Although TNF producing NKcells did not change (PCT Fig 12 a), LysM / BON immunisation elicited high levels of IL-17, IL-22 and IFN-y in NK cells (PCT Fig 12 b-d). In NKT cells, production of TNF, IL-17 and IL-22 remained unchanged following LysM / BON immunisation (PCT Fig 12 e-g), whereas slight increased expression of IFN-y was observed (PCT Fig 12 h). In addition, the expression of IL- 4 in NKT cells induced by LysM / BON immunisation was not significantly changed (PCT Fig 12 i). Together, these data showed that immunisation with LysM / BON resulted in the differentiation of Th cells to Th1 , Th2, and Th17 cells and activation of Tc cells, yb+T cells and NK cells. Although the OMPP1 immunisation showed comparable protection against K. pneumoniae infection in the sepsis model, the T-cell recall responses induced by OMPP1 restimulation in splenocytes were not significantly changed relative to control. Rather, IL-22 expressing NKcells were elevated in the OMPP1 -immunised group relative to the SAS control group, in addition, the IL-4 and TNF producing-NKT cells were increased in the OMPP1 - immunised group (PCT Fig 12 c, e & i). Overall, the stimulation of IL-17 and IL-22, which are important during K. pneumoniae infection were demonstrated in NK cells. Finally, OMPP1 significantly stimulated the expression of TNF (PCT Fig 12 j), while both LysM / BON and OMPP1 significantly induced the IL-17 production in CD3- cells (PCT Fig 12 k). LysM / BON also induced the expression of IL-22 (PCT Fig 12 I), while OMPP1 significantly stimulated the expression of IFN-y (PCT Fig 12 m) in CD3- cells. The increased expression of IL-4 in CD3- cells was only observed in LysM / BON restimulated cells (PCT Fig 12 n).

[0320] DISCUSSION

[0321] There are no clear correlates of protection yet identified for K. pneumoniae vaccines.

[0322] LysM / BON is a novel adhesin in K. pneumoniae, a function which was not previously identified. Previous studies indicated that it was important for carbapenem resistance in K. pneumoniae. Its homolog in Acinetobacter baumannii also played an important role in carbapenem resistance. However, the role of LysM / BON protein in Klebsiella pathogenesis has not been established. K. pneumoniae OMPP1 has never been evaluated as a potential vaccine candidate. DPS plays an important role in forming highly stable complexes with DNA and developing resistance to hydrogen peroxide during starvation -phase. Previous studies demonstrated that DPS was an important virulence factor contributing to the caecal colonisation of C. jejuni.

[0323] This invention demonstrated the function of K. pneumoniae LysM / BON, OMPP1 and DPS in attaching to lung epithelial cells 16HBE14o- / n vitro and showed the potential of those three antigens in the development of a vaccine against K. pneumoniae infection in vivo.

[0324] The inventors found that K. pneumoniae LysM / BON, OMPP1 and DPS antigens showed dramatic increases in attachment to human lung epithelial cells, 16HBE14O-, when recombinantly expressed in E. coli BL21 cells relative to E. coli cells that did not express the recombinant antigens. Immunisation with three different doses of LysM / BON and immunisation with 50 pg rOMPPI adjuvantedwith SAS showed strong seroconversion relative to the adjuvant only group, indicating that LysM / BON and OMPP1 were highly immunogenic proteins.

[0325] A sepsis model was used to determine the potential of LysM / BON, OMPP1 and DPS as prophylactic antigens. Significant reductions in K. pneumoniae colonisation in the peritoneal cavity were observed in all mice groups immunised with LysM / BON, OMPP1 or DPS compared with SAS adjuvant. In addition, concomitant reduction in dissemination to the spleen were observed. The inventors further examined the immune responses elicited by the antigens to identify which parameters contribute to protection . Splenocytesfrom the LysM / BON-im munised group re-exposed to LysM / BON showed higher levels of T-cell activation and differentiation compared to the control group and the substantial proliferation of effector CD4+and effector CD8+, memory CD4+and memory CD8+, and CD4+regulatory T cells in the LysM / BON- immunised mice indicated a robust effector recall response elicited by this antigen. While the overall splenocyte recall responses of the two antigens were quite distinct, it is apparent that restimulation with either LysM / BON or OMPPI led to IL-17 and IL-22 expression, highlighting that these cytokines are important for protection against K. pneumoniae. IL-17 and IL-22 are reported to be essential during the activation of the effective innate immune response in the lungs during K. pneumoniae infection. Stimulation of expression of IL-17 and IL-22 was shown in CD4+and CD8+T cells and NK cells in response to LysM / BON but IL-17 and IL-22 expression and in CD3- cells and NK cells respectively in OMPP1 immunised mice.

[0326] Overall, a mixed Th1 , Th2 and Th17 response was induced by LysM / BON immunisation, while a Th17 immune-skewed response was induced by OMPP1 immunisation, which suggest that IL-17 and IL-22 produced by NK cells and NKT cells are important against K. pneumoniae infection.

[0327] Recombinant LysM / BON, OMPP1 and DPS antigen candidates contain minimal amounts of LPS after purification and have a high potential to work as a bivalent or trivalent vaccine against K. pneumoniae.

[0328] Equivalents

[0329] The foregoing description details presently preferred embodiments of the present invention. Numerous modifications and variations in practice thereof are expected to occur to those skilled in the art upon consideration of these descriptions. Those modifications and variations are intended to be encompassed within the claims appended hereto.

Claims

1. CLAIMS1 . A vaccine composition comprising one or more immunogens (or a nucleotide or mRNA encoding the immunogen) selected from Klebsiella pneumoniae LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC, Ribokinase, Wzi family protein, LptD LPS assembly protein, outer membrane channel protein, polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, outer membrane protein assembly factor BamA, malate dehydrogenase, alcohol dehydrogenase, outer membrane lipoprotein SlyB, long-chain fatty acid outer membrane transporter, putative porin, and maltoporin.

2. The vaccine composition of Claim 1 , wherein the one or more immunogens selected comprise at least LysM / BON, OMPP1 and / or DPS (or a nucleotide ormRNA encoding the immunogen).

3. The vaccine composition of Claim 1 , wherein the immunogens in the composition consist of LysM / BON, OmpP1 and DPS (or a nucleotide or mRNA encoding the immunogen).

4. The vaccine composition of Claim 1 , wherein the immunogens in the composition comprise LysM / BON and 0mp1 (ora nucleotide or mRNA encoding the immunogen).

5. The vaccine composition of any one of the preceding claims, comprising at 5 or more of the one or more immunogens.

6. The vaccine composition of any one of the preceding claims, comprising 10 or more of the one or more immunogens.

7. The vaccine composition of any one of the preceding claims, comprising 15 or more of the one or more immunogens.

8. The vaccine composition of any one of the preceding claims, comprising Klebsiella pneumoniae LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC, Ribokinase, Wzi family protein, LptD LPS assembly protein, outer membrane channel protein, polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, outer membrane protein assembly factor BamA, malate dehydrogenase, alcohol dehydrogenase, outermembrane lipoprotein Sly B, long-chain fatty acid outer membrane transporter, putative porin, and maltoporin.

9. The vaccine composition of any one of the preceding claims, comprising one or more adjuvants.

10. The vaccine composition of any one of the preceding claims, for use in a vaccine therapy to prevent or treat infection by Klebsiella pneumoniae in a subject.1 1 . The vaccine composition for use of Claim 10, wherein vaccine therapy is to prevent infection and the subject is one at risk of an infection with K. pneumoniae.

12. The vaccine composition for use of Claim 10 or 11 , wherein the K. pneumoniae is hypervirulent K. pneumoniae (hvKp).

13. The vaccine composition for use of Claim 10 or 11 , wherein the K. pneumoniae is antibiotic-resistant K. pneumoniae.

14. The vaccine composition for use of any one of Claim 10, 12 or 13, wherein the use is to prevent infection, and the subject is one at risk of infection.

15. The vaccine composition for use of any one of Claims 10 to 14, wherein the subject is one admitted in a hospital unit, a spinal injury centre, a subject that is critically ill, a subject on a ventilators, a subject with prolonged use of invasive medical device, and / or a subject with an underlying condition.

16. A composition comprising an antibody that binds a Klebsiella pneumoniae protein selected from LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC, Ribokinase, Wzi family protein, LptD LPS assembly protein, outer membrane channel protein, polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, outer membrane protein assembly factor BamA, malate dehydrogenase, alcohol dehydrogenase, outer membrane lipoprotein Sly B, long-chain fatty acid outer membrane transporter, putative porin, and maltoporin.

17. The composition of Claim 16, comprising a plurality of antibodies, wherein each antibody of the plurality binds a Klebsiella pneumoniae protein independently selected from LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC,Ribokinase, Wzi family protein, LptD LPS assembly protein, outer membrane channel protein, polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, outer membrane protein assembly factor BamA, malate dehydrogenase, alcohol dehydrogenase, outer membrane lipoprotein SlyB, long-chainfatty acid outermembrane transporter, putative porin, and maltoporin.

18. The composition of Claim 16 or 17, wherein the antibody is a monoclonal antibody.

19. The composition of any one of Claims 16 to 18, wherein the antibody is selected from anti-LysM, anti-DPS or anti-OmpP1 .

20. The composition of Claim 19, wherein the antibodies in the composition consist of anti- LysM, anti-DPS and anti-OmpP1.

21. The composition of any one of Claims 16 to 20, for use in a method of treating K pneumoniae infection in a subject.

22. The composition for use of Claim 21 , wherein the K. pneumoniae is antibiotic-resistant K. pneumoniae.

23. The composition for use of Claim 21 or 22, wherein the use is to prevent infection, and the subject is one at risk of infection.

24. The composition for use of any one of Claims 21 to 23 wherein the subject is one admitted in a hospital unit, a spinal injury centre, a subject that is critically ill, a subject on a ventilator, a subject with prolonged use of invasive medical device, and / or a subject with an underlying condition.

25. The vaccine composition of any one of Claims 1 to 9, or the composition of any one of Claims 16 to 20, for use as a medicament.

26. An immunogen (or a nucleotide or mRNA encoding the immunogen) selected from Klebsiella pneumoniae LysM / BON, OmpP1 , DPS, Outer membrane protein assembly factor BamC, Ribokinase, Wzi family protein, LptD LPS assembly protein, outer membrane channel protein, polysaccharide export protein, DNA-binding transcriptional activator OsmE, Deoxyribose-phosphate aldolase / deoC, Putative lipoprotein, outer membrane protein assembly factor BamA, malate dehydrogenase,46alcohol dehydrogenase, outer membrane lipoprotein Sly B, long-chain fatty acid outer membrane transporter, putative porin, and maltoporin, for use as a therapy to induce an immune response against K. pneumoniae in a subject.

27. A method of treating or preventing infection by K. pneumoniae in a subject, the method comprising a step of administering the vaccine of any one of Claims 1 to 9 to the subject.

28. A method of treating K. pneumoniae infection in a subject, the method comprising a step of administering the composition of any one of Claim 16 to 20 to the subject.

29. The composition of any one of Claims 1 to 9, wherein the immunogen is isolated, recombinant or synthetic.

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