MXPA98004261A - Therapeutic compositions and diagnosti - Google Patents

Abstract

The present invention relates generally to therapeutic compositions for the treatment and / or prophylaxis of intestinal disease conditions in animals and birds caused or exacerbated by Lawsonia intracellularis or the like or microorganism reacted in another way. The present invention also contemplates method for the treatment and / or prophylaxis of such intestinal disease conditions and diagnostic agents and methods for detecting Lawsonia intracellularis or the like or microorganism related in another way.

Description

THERAPEUTIC AND DIAGNOSTIC COMPOSITIONS The present invention relates generally to therapeutic compositions for the treatment and / or prophylaxis of intestinal disease conditions in animals and birds caused or exacerbated by Lawsonia intracellularis or the like or microorganism related in another way. The present invention also contemplates methods for the treatment and / or prophylaxis of such intestinal disease conditions and for diagnostic agents and methods for detecting Lawsonia intracellularis or the like or microorganism related in another manner. The bibliographic details of the publications numerically referenced in this specification are gathered at the end of the description. The Sequence Identity Numbers (SEQ ID Nos.) For the nucleotide and amino acid sequences referred to in the specification are defined after the literature. Throughout this specification, unless the context requires otherwise, the word "comprises", or variations such as "comprises" or "comprising", shall be understood to imply the inclusion of a declared or whole or group element. of elements or integers, but not the exclusion of any other element or integer or group of elements or integers. The meat industry in Australia and, indeed, in most countries of the world, is an important aspect of the global livestock industry. However, the meat industry is subject to rapid economic losses in response to disease conditions that affect animals as well as human diseases putatively borne by animals. Therefore, it is important to have well-defined treatment, prophylactic and diagnostic procedures available to treat infections or potential infections in animals and humans. Pigs form a major component in the meat industry. However, pigs are sensitive to a broad spectrum of intestinal diseases collectively referred to as porcine proliferative enteropathy (PPE). This disease has been previously known as a complex of intestinal adentomatosis (1), porcine intestinal adenomatosis (PIA), necrotic enteritis (2), proliferative hemorrhagic enteropathy (3), regional ileitis (4), hemorrhagic bowel syndrome (5), enteritis proliferative swine and enteritis induced by Campylobacter spp (6). There are two main forms of PPE: a non-hemorrhagic form represented by intestinal adenomatosis, which frequently causes growth retardation and mild diarrhea; and a hemorrhagic form, which is frequently fatal, represented by proliferative hemorrhagic enteropathy (PHE) where the lumen of the distal small intestine becomes congested with blood. PPE has been reported in a number of animal species including pigs (14), hamsters (7), ferrets (15), guinea pigs (16), rabbits (17) as well as poultry species (18). The PPE-provoking organism is an organism similar to Campylobacter referred to herein as "Lawsonia intracellularis" (26). The organism has also previously been referred to as lleal symbiont intracellularis (7). PPE-like diseases in pigs can also be caused by other pathogens such as several species of Campylobacter (8). Lawsonia intracellularis is an intracellular bacterium, possibly linked intracellular. It can only be cultured in vitro with tissue culture cells (9, 26). Pigs suffering from PPE are characterized by multiple abnormal immature crypts and L. intracellularis is located in the cytoplasm of these crypt cells. PPE is an important cost component associated with the pig industry, especially in terms of feed costs, medication costs, reduced growth rates of pigs and increased feed costs. PPE also contributes to indirect costs downstream in, for example, additional labor costs and environmental costs to deal with contamination of antibiotic residues and for control measures to prevent the organism from being passed or carried to other animals or humans. Current control strategies for PEP depend on the use of antibiotics. Nevertheless, such a strategy is considered to be of short to medium term, especially as government regulation pressures tend to target animal management practices, which are only supported by prophylactic antibiotics. Consequently, there is a need to develop effective, safe and low-cost alternatives for the use of antibiotics. There is also a need to extend this alternative to antibiotics for similar organisms, which infect other animals such as humans.

In the work leading to the present invention, the inventors sought to develop vaccines for the prophylaxis and treatment of PPE in animals and birds. The vaccines of the present invention provide an effective alternative for the use of antibiotics with a range of consequential management and medical benefits. Accordingly, one aspect of the present invention provides a vaccine composition for the prophylaxis or treatment of infection in an animal or bird by L. intracellularis or the like or a microorganism otherwise related, said vaccine composition comprising an immunogenic form, non-pathogenic L. intracellularis or related microorganism or an immunogenic component thereof and one or more carriers, diluents and / or auxiliaries suitable for veterinary or pharmaceutical use. The present invention is particularly useful and is exemplified hereinafter in relation to the protection and / or treatment of swine of infection with L. intracellularis. However, this is done on the understanding that the present invention extends to the prophylaxis and treatment of all animals including humans and birds of infection with L. intracellularis and / or related microorganisms. The animals contemplated by the present invention include, but are not limited to, humans, primates, companion animals (e.g., cats, dogs), supply animals (e.g., pigs, sheep, cattle, horses, donkeys, goats) , laboratory test animals (eg, mice, rats, guinea pigs, rabbits) and captive wild animals (eg, kangaroos, foxes, deer). The present invention also extends to birds such as poultry, game birds and caged birds. Additionally, the present invention extends to all isolates and sub-types of L. intracellularis as well as other species of Lawsonia genus or other microorganisms related thereto at the nucleotide, biochemical, structural, physiological and / or immunointeractive level. The reference below to "Lawsonia intracellularis" or its abbreviation "L. intracellularis", includes all microorganisms similar to or otherwise related to this microorganism. For example, a related microorganism may have a nucleotide sequence similarity at the chromosome or extrachromosomal level of at least about 60%, more preferably at least about 70% and even more preferably greater than at least 80% with respect to all or part of the nucleotide sequence within the chromosome or extrachromosomal elements of L. intracellularis. For example, these percentage similarities may relate to the sequence set forth in SEQ ID NO: 5. This sequence is a portion of the chromosome of L. intracellularis. Accordingly, this aspect of the present invention is directed to a vaccine composition for the prophylaxis and / or treatment of infection in a pig by L. intracellularis, said vaccine composition comprising an immunogenic, non-pathogenic form of L. intracellularis or a related microorganism or an immunogenic component thereof and one or more carriers, diluents and / or auxiliaries suitable for veterinary or pharmaceutical use.

The term "immunogenic component" refers to L. intracellularis (in an attenuated or killed non-pathogenic form) or a component of L. intracellularis including a peptide, polypeptide or protein encoded by DNA of or derived from L. intracellularis, which is able to induce a protective immune response in a pig. A protective immune response can be at the humoral and / or cellular level and generally results in a substantial reduction in the PPE symptoms in pigs. The vaccine compositions will comprise an effective amount of immunogenic component in a manner that allows the induction of a protective immune response. According to this aspect of the present invention, a vaccine composition for the prophylaxis and treatment of a pig by L. intracellularis is provided., said vaccine composition comprising an amount of at least one immunogenic component of L. intracellularis or related microorganism effective to induce a protective immune response in said pig against L. intracellularis or related microorganism, said vaccine composition further comprising one or more carriers , auxiliaries and / or diluents suitable for veterinary or pharmaceutical use. The immunogenic component may be a naturally occurring peptide, polypeptide or protein, a carbohydrate, lipid or nucleic acid (e.g., DNA) or any combination thereof isolated from L. intracellularis or a cell culture thereof, or a recombinant form of a peptide, polypeptide or protein encoded by DNA of or derived from L. intracellularis, or is a derivative of said peptide, polypeptide or protein. An isolated component of L. intracellularis is a component which has undergone at least one purification step, or which has experienced at least partial concentration of a cell culture comprising L. intracellularls or a Used preparation of L cells. intracellularis The purity of such component of L. intracellularis, which has the required immunogenic properties is preferably at least about 40%, more preferably at least about 50%, even more preferably at least about 60%, still more preferably at least about 70% and even more preferably at least about 80-90% or greater relative to other components in a preparation as determined by molecular weight, immunogenic activity or other suitable means. A particularly useful form of the vaccine is a whole cell vaccine, which comprises L. intracellularis in an attenuated or otherwise non-pathogenic form or dead cells or various fractions thereof. Attenuated or nonpathogenic cells include killed L. intracellularis cells prepared, for example, by heating, formalin or other chemical treatment, electrical shock or pressure and such cells are particularly useful in the practice of the present invention.

According to this aspect of the present invention, there is provided a vaccine composition for the prophylaxis and / or treatment of infection in a pig by L. intracellularis or related microorganism, said vaccine composition comprising a killed preparation of L. intracellularis or microorganism related or an immunogenic fraction thereof and one or more carriers, diluents and / or auxiliaries suitable for veterinary or pharmaceutical use. In an alternative embodiment, a recombinant vaccine can be used. The recombinant vaccine may comprise one or more peptides, polypeptides or recombinant proteins derived from L. intracellularis or is a recombinant molecular immunologically related to a peptide, polypeptide or protein derived from L. intracellularis, or may be a fusion molecule having a first a portion comprising a peptide, polypeptide or protein derived from L. intracellularis and a second peptide, polypeptide or heterelogic protein which may be useful, for example, as a carrier molecule or an auxiliary or an immune stimulatory molecule such as a cytokine. A particularly useful recombinant protein of L. intracellularis comprises a peptide, polypeptide or protein derived from the surface or cell membrane of L. intracellularis, is an enzyme in a metabolic pathway within L. intracellularis or is a redoubled and / or shock protein of heat. In a preferred embodiment, the protein is a redoubled protein / heat shock such as, but not limited to, GroEL and GroES. Other putative vaccine candidates include flagellar basal body rod protein, S-adenosylmethionine: tRNA ribosyltransferase-isomerase, enoyl- (acyl-carrier-protein) reductase, N-acetyl muramoyl-L-alanine amidase (autolysin), UOP-3 -0- [3-hydroxymyristoyl] glucosamine N-acetyltransferase and a glucarate transporter. According to a preferred embodiment, the present invention relates to a vaccine composition for the prophylaxis and / or treatment of infection in a pig by L. intracellularis or related microorganism, said vaccine composition comprising at least one peptide, polypeptide or recombinant protein of L. intracellularis and wherein said peptide, polypeptide or recombinant protein is capable of inducing a protective immune response against L. intracellularis in pigs, the vaccine composition further comprising one or more carriers, diluents and / or auxiliaries suitable for use veterinary or pharmaceutical. In a particularly preferred embodiment, the recombinant protein is GroEL having an amino acid sequence as set forth in SEQ ID NO: 2 or is a protein having a predicted amino acid sequence of at least about 40%, at least about 60% , or more preferably at least about 70% and even more preferably at least about 80-90% or greater in similarity to all or part of the amino acid sequence set forth in SEQ ID NO: 2. In another embodiment, the recombinant molecule is GroES having an amino acid sequence as set forth in SEQ ID NO: 4 or is a molecular having an amino acid sequence of at least about 40%, at least about 60%, more preferably at least about 70% and even more preferably at least about 80-90% or greater in similarity to all or part of the amino acid sequence set forth in SEQ ID NO: 4. Another embodiment of the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 1 or having at least 40% similarity thereto or capable of hybridizing thereto. under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 3, or having at least 40% similarity thereto or capable of hybridizing thereto under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 5, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 6, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 8, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 11, or having at least 40% similarity thereto or capable of hybridize to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 13, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 15, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 17, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 18, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 19, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related mode, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 20, or having at least 40% similarity thereto or capable of hybridizing thereto under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 21, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 22, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. In a related embodiment, the present invention includes and comprises a peptide, polypeptide or protein encoded by a nucleotide sequence as set forth in SEQ ID NO: 23, or having at least 40% similarity thereto or capable of hybridizing to it under conditions of low severity and whose nucleotide sequence encodes an immunogenic component of L. intracellularis or related microorganism. The percentages of preferred similarities include at least about 50% or at least about 60% or at least about 70-90%. Herein, the reference to a low severity at 42 ° C includes and ranges from at least about 1% v / v to at least about 15% v / v of formamide and from at least about 1 M to at least about 2M salt for hybridization, and at least 1M to at least about 2M salt for washing conditions. Alternative severity conditions may be applied where necessary, such as average severity, which includes and ranges from at least about 16% v / v to at least about 30% v / v of formamide and from at least about 0.5M to at least about 0.9M salt for hydridation, and at least about 0.5M to at least about 0.9M salt for washing conditions, or high severity, which includes and spans from at least about 31% v / is at least about 50% v / v of formamide and from at least about 0.01 to at least about 0.15M salt for hybridization, and at least about 0.01M to at least about 0.15M salt for conditions of washed. The present invention also contemplates peptides, polypeptides or proteins having an amino acid sequence substantially as set forth in one of SEQ ID NO: 7 or 9 or 10 or 12 or 14 or 16 or having at least 40% similarity of the same or all or part thereof. The percentages of preferred similarities include at least about 50%, or at least about 60% or at least about 70-90%. The present invention further extends to a vaccine comprising a recombinant vaccine vector encoding a peptide, polypeptide or protein derived from L. intracellularis or related microorganism as described above. The vaccine vector may be of viral, yeast or bacterial origin and would be capable of expression of a genetic sequence encoding a peptide, polypeptide or protein of L. intracellularis in a manner effective to induce a protective immune response. For example, a non-pathogenic bacterium could be prepared containing a recombinant sequence capable of encoding a peptide, polypeptide or protein of L. intracellularis. The recombinant sequence would be in the form of an expression vector under the control of a constitutive or inducible promoter. The bacteria would then be allowed to colonize suitable locations in a pig gut and be allowed to grow and produce the recombinant peptide, polypeptide or protein in an amount sufficient to induce a protective immune response against L. intracellularis. In an alternative embodiment, the vaccine can be a DNA vaccine comprising a DNA molecule encoding a peptide, polypeptide or protein of L. intracelfularis and which is injected into a muscle tissue or other suitable tissue in a pig under sufficient conditions to allow transient expression of said DNA to produce an amount of peptide, polypeptide or protein effective to induce a protective immune response. The vaccines of the present invention may contain a single peptide, polypeptide or protein or a range of peptides, polypeptides or proteins covering different or similar epitopes. In addition, or alternatively, a single polypeptide can be provided with multiple epitopes. The last type of vaccine is referred to as a polyvalent vaccine. A multiple epitope includes two or more repetitive epitopes. Vaccine formation is generally known in the art and conveniently reference may be made to Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Co., Easton, Pennsylvania, USA. The present invention, therefore, contemplates a pharmaceutical composition or vaccine composition comprising an effective amount that develops immunity of one or more of: (i) an immunogenic component of L. intracellularis; (ii) a peptide, polypeptide or recombinant protein of L. intracellularis having immunogenic properties; and / or (iii) whole cells or a component or fraction thereof of L. intracellularis.

The above components are referred to hereafter as "active ingredients". The active ingredients of a vaccine composition as contemplated herein exhibit excellent therapeutic activity, for example, in the treatment and / or prophylaxis of PPE when administered in an amount which depends on the particular case. For example, for recombinant molecules, from about 0.5 μg to about 20 mg can be administered. Other useful effective amounts include 1 μg to about 10 mg, 10 μg to about 5 mg and 50 μg to about 1 mg. The important feature is to administer enough to induce an effective protective immune response. The above amounts can be administered as stated or can be calculated per kilogram of body weight. The dosage regimen can be adjusted to provide the optimal therapeutic response. For example, several divided doses may be administered daily or the dose may be proportionally reduced as indicated by the exigencies of the therapeutic situation. Reinforcement administration may also be required. The active ingredients may be administered in a convenient manner such as by routes, oral, intravenous (where it is soluble in water), intramuscular, subcutaneous, intranasal, intradermal or suppository or implant (e.g., using slow release technology). Depending on the route of administration, the active ingredients which comprise, for example, peptides, polypeptides or proteins may be required to be covered in a material to protect said ingredients from the action of enzymes, acids and other natural conditions, which may inactivate said ingredients. The term "auxiliary" is used in its broadest sense and includes any immune stimulating compound such as interferon. Auxiliaries contemplated herein include resorcinols, non-ionic surfactants such as polyoxyethylene oleyl ether and n-hexadecyl polyethylene ether and complete and incomplete Freund's aids. The active compounds can also be administered parenterally or intraperitoneally. The dispersions can also be prepared in glycerol, liquid polyethylene glycols, and mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms. Pharmaceutical forms suitable for injectable use include sterile aqueous solutions (where it is soluble in water) or sterile dispersions and powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the form must be fluid to the extent that it is easily manageable in the syringe unless the dosage form is a solid or semi-solid, such as when slow release technology is employed. In any case, it must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms.

The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol and liquid polyethylene glycol, and the like), suitable mixtures thereof and vegetable oils. Proper fluidity can be maintained, for example, by the use of a coating such as licithine, by the maintenance of the required particle size in the case of dispersion and by the use of superfactants. Preventions of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars or sodium chloride. Prolonged absorption of the injectable compositions can be achieved by the use in the compositions of absorption retarding agents, for example, aluminum monostearate and gelatin. Injectable solutions can be prepared by incorporating the active compounds in the required amount in the appropriate solvent with several of the other ingredients listed above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle, which contains the basic dispersion medium and the required other ingredients from those listed above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation with vacuum drying and the freeze-drying technique, which produces a powder of the active ingredient plus any additional desired ingredient from the sterile solution previously filtered from them. The carriers and diluents include any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic agents and absorption retardants and the like. The use of such media and agents in vaccines is well known in the art. Except where conventional means or agents are incompatible with an active ingredient, the use thereof in the therapeutic compositions is contemplated. The complementary active ingredients can also be incorporated into the compositions.

Yet another aspect of the present invention is directed to antibodies to peptides, polypeptides or proteins of L. intracellularis or recombinant forms thereof or non-protein molecules such as carbohydrates. Such antibodies can be monoclonal or polyclonal and can be selected from antibodies that occur naturally for L. intracellularis or can be specifically cultured for specific molecules or whole cells or components or fractions thereof. The antibodies of the present invention are particularly useful for immunotherapy and vaccination, and can also be used as a diagnostic tool for infection or to review the progress of a vaccination or therapeutic regimen. For example, recombinant L. intracellularis peptides, polypeptides or proteins can also be used to classify screening for antibodies that occur naturally for L. intracellularis. Alternatively, specific antibodies can be used to screen for L. intracellularis. Techniques for such assays are well known in the art and include, for example, sandwich and ELISA assays. Further, an immunogenic component is considered to encompass an immunogenic component of L. intracellularis and includes recombinant molecules, whole cells and cell extracts. In accordance with this aspect of the present invention, the immunogenic components are particularly useful in the examination for antibodies to L. intracellularis and, hence, provide a diagnostic protocol for detecting L. intracellularis infection. Alternatively, biological samples can be directly examined for L. intracellularis using cultured antibodies for immunogenic components. Accordingly, a method is provided for the diagnosis of L. intracellularis infection in a pig, comprising contacting a biological sample of said pig with an immunogenic component that binds an effective amount of an antibody for a time and under conditions sufficient to form a complex immunogenic component-antibody, and then detect said complex. The presence of immunogenic components (or antibodies thereto) in blood, serum or other pig body fluid can be detected using a wide range of immunoassay techniques such as those described in US Pat. Nos. 4,016,043 4,424,279 and 4,018,653. This includes both single-site and two-site trials, such as "sandwich", of non-competitive types, as well as traditional competitive ligation trials. Sandwich tests are among the most useful and commonly used and are favored for use in the present invention. A number of variations of the sandwich test technique exist, and all are intended to be encompassed by the present invention. Briefly, in a typical forward assay, a specific immunogenic component antibody is immobilized on a solid substrate to form a first complex and the sample to be tested for the immunogenic component is contacted with the ligated molecule. After a suitable period of incubation, for a period of time sufficient to allow the formation of a secondary antibody-immunogenic component complex, a second antibody of immunogenic component, labeled with a reporter molecule capable of producing a detectable signal, is then added to incubated, allowing sufficient time for the formation of a tertiary complex. Any unreacted material is separated by washing and the presence of bound labeled antibody is determined by observation of a signal produced by the reporter molecule. The results can be either qualitative, by simple observation of the visible signal, or they can be quantitative when compared with a control sample. The present invention contemplates a range of variations for the assay in question including an assay for L. intracellularis antibodies using, for example, peptides, polypeptides or recombinant proteins of that organism.

The solid substrate is typically glass or a polymer, the most commonly used polymers being cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene. The solid supports may be in the form of tubes, beads, disks or microplates, or any other suitable surface for conducting an immunoassay. Ligation processes are well known in the art and generally consist of covalently crosslinking or physically adsorbing the molecule to the insoluble carrier. By "reporter molecule", as used in the present specification, is meant a molecule which, by its nature -. chemistry, produces an analytically identifiable signal which allows the detection of antibody bound to the antigen. The detection can be either qualitative or quantitative. The most commonly used reporter molecule in this type of assay are either enzymes, fluorophores or molecules containing radionuclide (ie, radioisotopes). In the case of an enzyme immunoassay, an enzyme is conjugated to the second antibody, generally by means of glutaraldehyde or periodate. However, as will be readily recognized, there is a wide variety of different conjugation techniques, which are readily available to one skilled in the art. Commonly used enzymes include horseradish peroxidase, glucose oxidase, β-galactosidase and alkaline phosphatase, among others. Substrates to be used with specific enzymes are generally chosen for the production, upon hydrolysis by the corresponding enzyme, of a change of detectable color. It is also possible to employ fluorogenic substrates, which produce a fluorescent product. Alternatively, fluorescent compounds, such as fluorescein and rhodamine, can be chemically coupled to antibodies without altering their binding capacity. When activated by illumination with light at a particular wavelength, the fluorochrome-labeled antibody adsorbs the light energy, inducing a state of excitability in the molecule, followed by the emission of light to a characteristic color visually detectable with a microscope. of light. As in the EIA, the fluorescently labeled antibody is allowed to bind to the first antibody-hapten complex. After washing the unbound reagent, the remaining ternary compound is then exposed to the light of the appropriate wavelength, the fluorescence observed indicates the presence of the hapten of interest. The immunofluorescence and ElA techniques are both well established in the art and are particularly preferred by the present method. However, other reporter molecules, such as radioisotope, chemiluminescent or bioluminescent molecules, may also be employed. It will be readily apparent to the skilled artisan how to vary the procedure to suit the required purpose. A range of genetic diagnostic assays can be employed, such as polymerase chain reaction (PCR) assays, hybridization assays or protein truncation assays. All these tests are contemplated in the present invention. The present invention is further described by the following, non-limiting, Figures and / or Examples.

In the Figures: Figure 1 is a photographic representation showing the "Western" analysis of L. intracellularis antigens recognized by vaccinated pigs. Footprint 1 (395) was tested with pig serum from a pig (395) that had been immunized three times with the complete L. intracellularis vaccine killed in formalin. Traces 2 to 5 (Y10, Y12, Y14, Y16) were tested with serum obtained from pigs Y10, Y12, Y14 and Y16, respectively on day 0. Figure 2 is a photographic representation of the small intestine of pig Y1 in on day 20. Figure 3 is a photographic representation of the small intestine of pig Y2 on day 20. Figure 4 is a photographic representation of the small intestine of pig Y4 on day 20.

The following one- and three-letter abbreviations are used for amino acid residues: Amino Acid Three-letter abbreviation Symbol of a letter Alanine Ala A Arginine Arg R Asparagine Asn N Aspartic Acid Aps D Cysteine Cys C Glutamine Gln Q Glutamic Acid Glu E Glycine Gly G Histidine His H Isoleucine He I Leucine Leu L Lysine Lys K Methionine Met M Phenylalaine Phe. F Proline Pro P Serine Ser S Threonine Thr T Tryptophan Trp W Tyrosine Tyr and Valine Val V Any Xaa residue X SUMMARY OF SEQUENCE IDENTITY NUMBERS SEQ ID NO. Description 1 Sequence of GroEL nucleotides 2 GroEL amino acid sequence 3 GroES nucleotide sequence 4 GroES amino acid sequence 5 Sequence of nucleotide component of L. intraceful / aris 6 Sequence of nucleotides of L. intracelullaris component 7 Sequences of amino acids of SEQ ID NO 6 8 Sequence of nucleotides of L. intracellular component 9 Sequences of amino acids of SEQ ID NO: 8 (first coding sequence) 10 Sequences of amino acids of SEQ I D NO: 8 (second coding sequence) 11 Sequencing of nucleotides of L component. intracellular 12 Sequence of amino acids of SEQ ID NO 1 1 13 Sequence of nucleotides of L. intracelullaris component 14 Sequence of amino acids of SEQ ID NO 13 15 Sequence of nucleotides of L. intracelullaris component 16 Sequence of amino acids of SEQ ID NO: 15 17 Sequence of nucleotides of L. intracelullaris component 18 Sequence of nucleotides of L. intracelullaris component 19 Nucleotide sequence of L. intracelullaris component 0 Nucleotide sequence of L. intracelullaris component 1 Nucleotide sequence of L. component intraceluHaris 2 Nucleotide sequence of L. intracellullaris component 3 Nucleotide sequence of component of L. intracelullaris EXAMPLE 1 PORK FABRIC SOURCES Infected pig intestines Sections of excessively thickened lions were taken from pigs affected naturally or experimentally by PPE. The presence of L. intracellularis bacteria in the lions was confirmed using immunofluorescent staining with specific monoclonal antibodies (10). An example of a suitable antibody is the IG4 monoclonal antibody from the University of Edinburgh, UK.

EXAMPLE 2 ISOLATION OF BACTERIA LAWSONIA INTRACELLULARIS OF THE ILEON INFECTED PIG The Lawsonia intracellularis bacteria were extracted directly from PPE lesions in pigs by filtration and further purified on a Percoll gradient (Pharmacia, Uppsala, Sweden). The infected lions were collected from pigs and the presence of L. intracellularis was confirmed histologically before storage at -80CC. The ileum sections were thawed and approximately 8 g of infected mucosa were scraped from the intestinal wall. The mucosa was homogenized with 40 ml of sterile phosphate buffered saline (PBS) at medium speed for 10 s using a Sorvall omnimixer. This suspension was centrifuged at 2000 xg for 4 minutes. The supernatant was discarded and the cell pellet was resuspended in 40 ml of PBS and recentrifuged. This washing step was repeated twice. The cell pellet was resuspended in 20 ml of PBS and homogenized at full speed for one minute to release the L. intracellularis bacteria. This homogenate was centrifuged at 1000 xg for 4 minutes giving a pellet containing a crude mixture of intestinal bacteria and homogenized epithelial cells. The supernatant was filtered using filters with a pore size of 3 μm, 1.2 μm and 0.8 μm (Millipore Corporation, MA, USA). The filtrate was centrifuged at 8000 xg for 30 minutes, resulting in a small pellet of L. intracellularis bacteria. The L. intracellularis bacteria were further purified using a Percoll gradient of 45% self-formation: 2 ml of bacterial preparation were mixed by inversion in 30 ml of a 45% self-training Percoll gradient (Pharmacia LKB, Uppsala, Sweden) (45% v / v Percoll, 150 mM NaCl). The gradients were centrifuged in a Sorval centrifuge using the SS34 rotor at 20,000 rpm for 30 minutes at 4 ° C. Usually a number of bands are formed within the gradient. The band (usually located approximately 10-20 mm from the base of the tube) containing the L. intracellularis bacteria was harvested and capped to 16 ml with PBS. The solution was then centrifuged for 15 minutes at 8000 rpm. The resulting pellet was washed with PBS before being resuspended in a final volume of about one ml.

EXAMPLE 3 PURIFICATION OF GENOMIC DNA OF LAWSONIA INTRACELLULARIS Genomic DNA was extracted from purified Lawsonia intracellularis bacteria by Percoll gradient, recovered from livers scraped from infected pigs (Example 2), by the methods described by Anderson et al (11) & Sambrook et al (12).

EXAMPLE 4 IMMUNOEXAMINATION OF GENOMIC GENOTECTS A genomic library of L. intracellularis lambda ZAP II was plated in a cell field of Escherichia coli XLI-Blue (23) at a density of 2,000 plaque forming units (pfu) per 150 mm of L. broth agar plate. The library was examined with a rabbit anti-L. intracellularis serum using the method described in Protoblot Technical Manual (Promega, Wl, USA). The filters were blocked in a buffer containing 10 mM Tris HCl, pH 8.0, 150 mM NaCl, 0.05% Tween 20, 1% w / w gelatin. Positive plaques identified in a primary examination were taken, replatinated at a lower density and retested until individual positive plaques were identified.

EXAMPLE 5 ISOLATION AND SEQUENCING OF I NSERCION IS OF cDNA "phagemid" DNA of phage clones? ZAPII positive was isolated by in vivo excision of "phagemid" pBluescript under the conditions recommended by Stratagene (CA, USA). Plasmid DNA was extracted either by the Brinboim and Doly method as the cDNA insertions sequenced by the chain termination method (21), or by the PEG precipitation method and cyclo-sequenced by the terminator method. dyeing, as recommended by the manufacturer (Applied Biosystems).

EXAMPLE 6 ANTIQUES Antisera for L. intracellularis bacteria were cultured in rabbits and pigs. The rabbits were injected intramuscularly with a preparation of L. intracellularis bacteria purified by Percoll gradient mixed with a double emulsion made by processing with oil auxiliary (incomplete Freund's auxiliary, CSL Limited, Melbourne, Australia), and then with Tween 80 enhancer Two 3 ml injections, containing 9 mg of protein, were provided with four weeks of separation. Blood samples were collected from the marginal vein of the ear before immunization and two weeks after the second injection. A six-week-old pig (395) was hyperimmunized by intramuscular injection of L. intracellularis bacteria purified by Percoll gradient prepared with incomplete Freund's assistant as regards the rabbit. Three injections of prepared antigen were administered at four weeks apart, and blood was collected from the jugular vein two weeks after the final injection. Diluted pig sera (1 ml, 1 in 200) were pre-absorbed with 100 μl of E. coli lysate DH5a (24) for 1 h at room temperature with gentle mixing. The lysate was prepared by freeze-thawing a suspension of E. coli in PBS.

EXAMPLE 7 ELECTROPHORESIS OF DODECIL GEL SODIUM-SULPHATE ACR I LAM I DA Protein samples were resuspended in 50 μl of sample buffer (62.4 mM HCl, 2% w / v SDS, 10% v / v glycerol, 5% v / v 20 mercaptoenaol, 0.002% bromophenol blue, pH 8.6) and were heated at 95 ° C for 5 minutes before separating the electrophoretically solubilized proteins in a vertical plate gel of 0.1% w / v SDS-12% w / v PAGE (13).

EXAMPLE 8"WESTERN BLOTTING" The proteins were electrophoretically transferred to Immobilon-P membranes (Millipore Corporation, MA, USA) in a Trans-Blot Cell (BioRad, CA, USA) at 100 V for 1 h in a buffer containing CAPS (3- [cyclohexylamino] - 1 -propanesulfonic acid, pH 11, Sigma, Ml, USA) and 10% v / v methanol. The membranes were then blocked with 5% w / v Blotto (Diploma skim milk powder, Melbourne, Australia) in PBS for 30 min at room temperature with gentle rocking. The filters were then transferred to antisera diluted in 5% w / v of Blotto, PBS. The pre-absorbed pig antiserum was diluted 1 in 200. The filters were incubated in pig antisera for 1 h followed by washing three times in PBST. Anti-pig conjugated immunoglobulins of HRP (DAKO, CA, USA) were applied at a dilution of 1: 2000. Intensified chemiluminescence (ECL, Amersham, IL, USA) was used to discriminate proteins from L. intracellularis. Before the ECL detection, the spots were washed three times for 7 minutes each. The filters were exposed to autoradiographic film (Agfa, NJ, USA) for less than 1 minute before developing.

EXAMPLE 9 IDENTIFICATION OF GROEL AND GROES Clones found positive according to the immunoexamination method described in Example 4 were sequenced using the protocol detailed in Example 5. An isolated clone represented the GroEL protein. The corresponding nucleotide sequence and amino acid sequence of GroEL are shown in SEQ ID NO: 1 and SEQ ID NO: 2.

Another isolated clone represented the GroES protein. The sequence of GroES nucleotides and corresponding amino acid sequence are shown in SEQ ID NO: 3 and SEQ ID NO: 4.

EXAMPLE 10 IMMUNOFLUORESCENT DETECTION OF BACTERIA THE WSONIA INTRACELLULARIS IN PORTIONS OF PIG Fecal pig plugs were taken using cotton tipped swabs and then the specimen was grafted onto a glass slide. After allowing ten minutes of drying by air, the slides fixed by heat by heating at 60 ° C for about 10 seconds. The slides were then rinsed in PBS. An amount of 30 μl of a 1/200 dilution of mouse ascites containing monoclonal antibody IG4 (see Example 1) was added, a glass cover strip was applied, and the slides were incubated at room temperature for 40 minutes. The cover strip was removed and the slides were washed (PBST for 7 minutes, three times). An amount of 30 μl of a 1/40 dilution of a FITC conjugated anti-mouse antiserum (Silenus, Melbourne, Australia) was added, a glass cover strip was applied and the slides were incubated at room temperature for 40 minutes. The cover strip was removed and the slides were washed (PBST for 7 minutes X3). A final rinse in PBS was given to the slides. A 10% v / v drop of glycerol PBS was added and a glass cover strip was applied. Fluorescent bacteria were visualized under a Lietz laborlux S high power microscope (X1200) at 340 nm. Twenty fields were counted and the results (see Table 1) were expressed as the average number of L. intracellularis bacteria per high power field.

EXAMPLE 11 VACCINE OF L. INTRACELLULARIS DEAD IN FORMALINE The bacterial L. intracellularis pellet purified by Percoll gradient was resuspended in 1 ml of 1% formalin in saline and incubated overnight at 4 ° C. The L. intracellularis bacterium purified by Percoll gradient was then mixed in a double emulsion made by processing with oil auxiliary (incomplete Freund's auxiliary, Commonwealth Serum Laboratories, Melbourne, Australia), and then with Tween 80 enhancer.

EXAMPLE 12 VACCINATION PROTOCOL Twelve weaned pigs (Landrace crossed with Large White) were taken from a Pig Improvement Company pigsty and treated with Neo-Terramycin (0.25 g / kg) for 5 days. Seven days later the pigs (of 40 days) Y10, Y12, Y14 and Y16 were vaccinated as described. The pigs Y3, Y1 1 and Y13 were treated by absecso with long-acting terramycin on day 34.

The twelve pigs were divided into three groups and treated as follows: Group 1 Infected controls Four pigs (Ear Mark No. Y1-Y4) were housed with vaccinated pigs. Group 2 Complete bacterial vaccine Four pigs (Ear Mark No. Y10, Y12, Y14 and Y16) were immunized with 0.5 ml of killed L. intracellularis bacteria in formalin emulsified in 0.5 ml of PBS / incomplete Freunden's assistant on days 33 and 12. Group 3 Controls not infected Four pigs (Ear Mark No. Y9, Y1 1, Y13 and Y15) were not treated and were housed in a separate area from vaccinated pigs and infected control pigs.

EXAMPLE 13 ORAL ASSESSMENTS OF INFECTED PIGS Infected neons were collected from pigs as described in Example 1 and the presence of L. intracellularis was confirmed histologically before storage at -80 ° C. Sections of ileum were thawed and approximately 150 g of infected mucosa was scraped from the intestinal wall. The mucosa was homogenized with an equal volume of sterile PBS at medium speed for 20 s using an omnimixer Sorvall. This suspension was diluted twice with sterile PBS to form the titration suspension. On day 0 each pig of Groups 1 and 2 was dosed with a 5% w / v solution with sodium bicarbonate (10 ml / kg) followed by 30 ml of titration suspension. This was repeated on day 1 and day 2. From day 11 onwards, the number of L. intracellularis bacteria in the feces of each pig was verified by immunofluorescence. The pigs were checked for signs of disease and shedding of L. intracellularis bacteria. The shedding of pigs greater than 100 bacteria per field of high potency and diarrhea were sacrificed for ethical reasons. On day 22 the surviving pigs were humanely slaughtered and the small intestines were recovered. The two sections of small intestine were removed 5 cm and 17 cm proximally from the leocecal junction. These sections were fixed at 10% v / v of formalin, the encrusted wax and sections were sent to an independent veterinary pathologist for analysis.

EXAMPLE 14 LAWSONIA INTRACELLULARIS PROTEINS RECOGNIZED BY VACCINATED PIGS Antibodies raised by pigs for L. intracellularis proteins after vaccination were analyzed by Western blotting followed by ECL detection (Amersham, IL, USA) as described in Example 8. The results are shown in Figure 1. Vaccinated pigs produce antibodies to a range of proteins of L. intracellularis. The most immunodominant proteins recognized were approximately 62.7 Kda, 58.7 Kda, 57.2 Kda, 44 Kda. 36.7 Kda and two lorries of 24-26 Kda and 22-23.5 Kda. The minor immunoreactive bands have approximately the following molecular weights 67 Kda, 52.5 Kda, 50.5 Kda, 48.2 Kda, 47.9 Kda, 44.7 Kda, 43.5 Kda, 42.5 Kda, 41.5 Kda, 40.5 Kda, 39 Kda, 35.3 Kda, 17 Kda, 15.5 Kda, 12 Kda and 7 Kda. The molecular weight of the recognized proteins will vary up to 5% depending on the method used for the estimation.

EXAMPLE 15 SPILLING OF BACTERIA L. INTRACELLULARIS BY PIGS DURING THE TRIAL Three of the pigs from Group 1 (infected control) in Example No. 12 (Y1, Y2 and Y4) shed more than 100 L. intracellularis bacteria per high potency field in their feces by day 19 post oral assessment (Table 1 ). Two of these pigs (Y2 and Y4) had bloody diarrhea. The three pigs were humanely slaughtered on day 20. Y3 shed low levels of L. intracellularis bacteria during the course of the infection test. The maximum bacterial spillage for Y3 was 16 bacteria per high power field. All pigs in group 3 vaccinated with the complete bacterium as set forth in Example 12, never shed more than 3 L. intracellularis by high power field. Vaccination with the killed L. intracellularis vaccine in formalin reduced the total bacterial shedding of L. intracellularis bacteria by pigs vaccinated by 98.5% when compared with the pigs in group 1. None of the pigs in group 3 (controls not infected) shed no L. intracellularis bacteria during the course of the trial. The results of spillage of L. intracellularis bacteria per pig are shown in Table 1.

EXAMPLE 16 GENERAL PATHOLOGY FOR THE PROOF A Group 1 Infected controls Y1 Approximately 5 cm of terminal ileum was thickened excessively. No other sign of PPE was evident macroscopically. The findings are consistent with intestinal adenomatosis (See Figure 2) Y2 It was found that the intestine was thickened excessively and the serosa had the characteristic cerebriform forms (Figure 3). About 2.5 meters of the intestine was involved. It was found that the lumen of the intestine contained fresh meat and fibrinous emptyings were evident. Proliferative hemorrhagic enteropathy. Y3 No excessive sign of PPE was evident. Y4 The intestine was found to have necrotic enteritis (Figure 4). The surface of the mucosa was replaced with a fibrinous pseudomembrane. Mesenteric edema was clearly evident. About 2.0 meters of intestine was involved.

Group 2 Complete cell vaccine of L. intracellularis Y10 No general signs of PPE Y12 No general signs of PPE Y14 No general signs of PPE Y15 No general signs of PPE Group 3 Controls without infection Y9 No general signs of PPE Y1 1 No general signs of PPE Y13 No general signs of PPE Y15 No general signs of PPE EXAMPLE 17 REPORT OF HISTOPATHOLOGY FOR THE TEST The reports are based on the histopathological descriptions established in Jubb et al (20).

Group 1 Infected control group Y1 Numerous microfocal / confluent lesions of Adenomatosis Porcine Intestinal (PIA) are associated with Peyers Patches Y2 Serious generalized (annular) lesions of Porcine Intestinal Adenomatosis. Y3 No conclusive evidence of PIA. Dispersed microfocal lesions suggestive of non-specific mild reactive hyperplasia (reparational) (instead of adenomatous). Y4 Severe generalized (annular) injuries of PIA.

Group 2 Complete cell vaccine of L. intracellularis Y10 No conclusive evidence of PIA. Y12 No conclusive evidence of PIA. Y14 No conclusive evidence of PIA. Y15 No conclusive evidence of PIA. Possible single PIA microfocus is associated with Peyers Patch.

Group 3 Controls not infected Y9 No conclusive evidence of PIA. Y1 1 No conclusive evidence of PIA. Y13 The intestine was not recovered since the pig was killed due to weakness on day 15. Y15 The diagnosis was not possible due to the low quality sections.

EXAMPLE 18 IMMUNOEXAMI NATION OF A GENOTECA OF L. INTRACELLULARIS USING EXPERIMENTAL VACCINATED PIG SUASES Genomic DNA of L. intracellularis was purified as described in Example 3. The DNA was partially digested with the restriction endonuclease Sau3A (Promega) and ligated in Lambda ZAP II Express (Stratagene). The lambda library was plated in an E. coli XLI-Blue cell field at a density of 10,000 pfu per 150 Mm of L. broth agar plate. The library was examined, as described in Example 4, with Y12 Y12 pig was immunized with L. intracellularis killed in formalin, as described in Example 1 1 & 12. The vaccinated pigs produced antibodies for a range of L. intracellularis proteins, as described in Example 14. A number of phage clones expressing L. intracellularis proteins was identified.

EXAMPLE 19 ANALYSIS OF FAGOS CLONES EXPRESSING L. INTRACELLULARIS "phagemid" DNA from phage clones? ZAP II Express was isolated by excision in vivo, using the conditions recommended by the manufacturer (Stratagene). The plasmid DNA, for restriction analysis, was extracted by alkaline lysis, as described by Sambrook et al (12), and for automated sequencing, using the High Purée Plasmid Kit, as recommended by the manufacturer (Boehringer Mannheim). DNA sequencing of insertions was performed using the automated sequencing dye terminator method (ABI Biosystems).

The identified sequences are set forth in SEQ ID NOS: 5-23 (see Example 20).

EXAMPLE 20 IDENTIFICATION OF L. INTRACELLULARIS COMPONENTS The sequence similarity of the DNA molecules encoding putative vaccine candidates identified from Example 18 and 19 was identified using BLAST (27). The nucleotide sequence SEQ ID NO: 6 and its corresponding amino acid sequence SEQ ID NO: 7 have the sequence similarity for a basal body flagellar rod protein. SEQ ID NO: 8 (nucleotides) and SEQ ID NOS: 9 and 10 (amino acids) have sequence similarity to autolysin. SEQ ID NO: 11 (nucleotides) and SEQ ID NO: 12 (amino acids) show sequence similarity to S-adenosylmethionine: tRNA ribosyltransferase-isomerase (queuosin biosynthesis protein queA). SEQ ID NO: 13 (nucleotides) and SEQ ID NO: 14 (amino acids show sequence similarity to enoyl- (acyl-carrier-protein) reductase SEQ ID NO: 15 (nucleotides) and SEQ ID NO: 16 (amino acids) sequence similarity to a glucarate transporter Other nucleotide sequences encoding vaccine candidates are SEQ ID NO: 5, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 20, SEQ ID NO: 21, SEQ ID NO: 22, and SEQ ID NO: 23. Those skilled in the art will appreciate that the invention described herein is susceptible to variations and modifications different from those specifically described. It will be understood that the invention includes all those variations and modifications. The invention also includes all steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of any two or more of said steps or features.

TABLE 1 TABLE 1 Vaccination Assessment Day Day Day Day Day Day Day Day Day Day Day -40 -33 -26 -12 0 1 2 11 12 13 14 15 16 17 18 19 20 21 22 1 Infected controls 1 + 1 + 0 0 5+ 10+ 50+ 100+ 15 5 cn. of thickening 2 Infected Controls 0 1 + 1 + 1 + 3+ 1 + 70+ 100+ 100 PHE 2.5 M 3 Infected controls 0 0 0 0 0 0 1 + 4 16 1+ 0 1 4 Infected controls 1 + 0 0 10+ 0 5+ 60+ 200+ 80 PHE 2.0 4. s.

Bacteria 1ml cel. complete dead 1 ml of cel. complete dead 0 0 0 0 0 1 + 1 + 0 0 0 0 0 complete 12 Bacteria 1 ml cel. complete dead 1 ml of cel. complete dead 1 + 0 0 0 0 2+ 0 0 0 0 0 0 complete 14 Bacteria 1ml cel. complete dead 1 ml of cel. complete dead 0 0 0 0 0 1 + 0 < 1 < 1 or o or complete 16 Bacteria 1 ml of cel. complete dead 1 ml of cel. complete dead 0 0 0 0 0 0 0 3 < 1 or o complete 9 Controls no 0 0 0 0 or 0 0 0 0 0 0 0 infected 11 Controls no 0 0 0 0 0 0 0 0 0 0 0 0 infected 13 Controls no 0 0 0 0 Track dead infected 15 Controls no 0 0 0 0 or 0 0 0 0 0 0 0 infected -4 BIBLIOGRAPHY Barker, I. K. and Van Dreumel, A.A (1985) in "Pathology of Domestic Animáis," 3rd edition, Vol. 2 p. 1-237, eds K.V.F. Jubb, P.C. Kennedy and N. Palmer. (Academic Press: Orlando). Rowland, A.C. and Lawson, G.H. K. (1976) Veterinary Record 97: 178-180. Love, R.J. and Love, D.M. (1977) Veterinary Record 100: 473 Jonsson, L. and Martinsson, K. (1976) Veterinary Act Scandinavica 17: 223-232. O'Neil, I. P.A. (1970) Veterinary Record 87: 742-747. Straw, B. E. (1990). Journal of the American Veterinary Medical Association 197: 355-357. Stills, H.F. (1991). Infection and immunology 59: 3227-3236. Gebhert, C.J. , Ward, G.E., Chang, K. and Kurtz, J.H. (1983). American Journal of Veterinary Research 44: 361-367. Lawson, G.H.K., McOrist, S., Jansi, S. and Mackie, R.A. (1993) Journal of Clinical Microbiology 31: 1 136-1 142. McOrist, S., Boid, R., Lawson, G. H.K. and McConnell, I. (1987) The Veterinary Record 121: 421-422. Anderson, B.J. , M.M. Bills, J. R. Egerton, and J.S. Mattick. (1984) Journal of Bacteriology 160: 748-754. Sambrook, J., E. F. Fritsch, and T. Maniatis. (1989) Molecular cloning. A laboratory manual. Second edition. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. 13 Laemmli, U. K. (1970) Nature 227: 680-685. 14 McOrist, S, Jasni, S, Mackie, RA, MacIntyre, N, Neef, N. and Lawson GHK (1993) Infection and Immunity 61: 4286-4292. 15 Fox, JG, Murphy, JC, Otto, G Pecquet-Goad, ME, Larson, QHK and Scott JA (1989) Veterinary Pathology 26: 515-517. 16 Elwell, MR, Chapman, AL and Frenkel, JK (1981) Veterinary Pathology 18: 136-139. 17 Schodeb, TR and Fox JG (1990) Veterinary Pathology 27: 73-80. 18 Mason, RW, Monkton, P and Hasse D (1995) Australian Veterinary Journal (in press). 19 Manthorpe, M, Cornefert-Jensen, F., Hartikka, J., Felgner, J, Rundell, A, Margalith, M and Dwarki, V. (1993) Human Gene Therapy 4: 419-431. 20 Jubb KVC, Kennedy, PC and Palmer, NC (1993). The Pathology of Domestic Animáis, 4th ed. San Diego, CA, Academic Press, pp 229-233. 21 Birnboim, HC and Doly J (1979) Nucleic Acids Research 7: 1513. 22 Sanger, F, Nicklen, S and Coulson, AR (1977) Proceedings of the National Academy of Science 74: 5463. 23 Block, WO, Femandes, JM and Short, JM (1987) Biothecnics 5: 376-79. 24 Woodcock, DM et al (1989) Nucleic Acids Research 17: 3469-78. 25 Studier, FW et al (1990) Methods in Enzymology 185: 60-89. 26 McOrist, S et al (1995) International Journal of Systematic Bacteriology 45: 820-825. 27 Gish, W and States, D.J. (1993) Nature Genetics 3: 266-272.

LIST OF SEQUENCES (1) GENERAL INFORMATION: (i) APPLICANT: (OTHER THAN DIFFERENT TO US) DARATECH PTY LTD and PIG RESEARCH (ONLY US): MICHAEL PANACCIO and DETLEF HASSE (ii) TITLE OF THE INVENTION: THERAPEUTIC COMPOSITIONS AND DIAGNOSTICS (iii) NUMBER OF SEQUENCES: 23 (iv) ADDRESS FOR CORRESPONDENCE: (A) DESTINY: DAVIES COLLISON CAVE (B) STREET: 1 LITTLE CQLLÍNS STREET (C) CITY: MELBOURNE (D) STATE: VICTORIA (D) COUNTRY: AUSTRALIA (E) POSTAL CODE: 3000 (v) LEGIBLE COMPUTER FORM: (A) TYPE OF MEDIA: FLEXIBLE DISC (B) COMPUTER: IBM PC compatible (C) OPERATING SYSTEM: PC-DOS / MS-DOS (D) PACKAGING : Release Patent # 1 .0, version # 1.25 (vi) CURRENT APPLICATION DATA: (A) APPLICATION NUMBER: (B) SUBMISSION DATE: 29-NOV-1996 (vi) PREVIOUS APPLICATION DATA: (A) APPLICATION NUMBER: PN691 1/95 (B) SUBMISSION DATE: 30-NOV-1995 (A) APPLICATION NUMBER: PN6910 / 95 (B) DATE OF SUBMISSION: 30- NOV-1995 (viii) ATTORNEY / AGENT INFORMATION: (A) NAME: HUGHES DR, E JOHN L (C) REFERENCE NUMBER / CASE: EJH / AF (ix) TELECOMM INFORMATION NICATIONS: (A) TELEPHONE: +61 3 9254 2777 (B) TELEFAX: +61 3 9254 2770 (2) INFORMATION FOR SEQ ID NO: 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1647 base pairs (B) TYPE: nucleic acid (C) ) FILAMENT: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 1..1647 (xi) SEQUENCE DESCRIPTION: SEQ ID NO :1 ATG GCT TCT AAA GAA ATC CTT TTT GAT GCT AAA GCC CGT GAA AAA CTT 48 Met Wing Wing Lys Glu lie Leu Phe Asp Wing Lys Wing Arg Glu Lys Leu 1 5. 10 15 TCA CGA GGT GTA GAT AAA CTT GCA AAT GCT GTT AAA GTA ACA CTT GGA 96 Ser Arg Gly Val Asp Lys Leu Ala Asn Ala Val Lys Val Thr Leu Gly 20 25 30 CCT AAA GGC CGT AAT GTC GTT ATT GAA AAG TCT TTT GGT TCC CCA GTT 144 Pro Lys Gly Arg Asn Val Val lie Glu Lys Ser Phe Gly Ser Pro Val 35 40 45 ATT ACA AAA GAT GGT GTA TCT GTT GCA AAA GAA ATT GAA CTT GAA GAT 192 lie Thr Lys Asp Gly Val Ser Val Ala Lys Glu lie Glu Leu Glu Asp 50 55 60 AAG TTT GAA AAT ATG GGC GCT CAG ATG GTT AAA GAA GTA GCT CCC AAA 240 Lys Phe Glu Asn Met Gly Ala Gln Met Val Lys Glu Val Ala Pro Lys 65 70 75 80 ACT AGC GAT ATT GCT GGT GAT GGA ACT ACA ACA GCA ACA GTC CTT GCA 288 Thr Ser Asp lie Wing Gly Asp Gly Thr Thr Thr Wing Thr Val Leu Wing 85 90 95 CAQ GCT ATT TAT CGT GAA GGT GTA AAA CTT GTA GCA GCT GGT CGT AAT 336 Gln Ala lie Tyr Arg Glu Gly Val Lys Leu Val Wing Wing Gly Arg Asn 100 105 110 CCT ATG GCC ATT AAA CGT GGC ATA GAT AAA GCT GTT GTT GCT GTT ACT 38 'Pro Met Ala lie Lys Arg Gly He Asp Lys Wing Val Val Wing Val Thr 115 120 125 AAA GAA CTA AGC GAC ATT ACA AAG CCT ACT CGT GAC CAA AAA GAA ATA 432 Lys Glu Leu Ser Asp He Thr Lys Pro Thr Arg Aßp Gln Lys Glu He 130 135 140 GCT CA GTT GGA ACC ATT TCT GCA AAC TCT GAT ACA ACA ATA GGT AAT 480 Wing Gln Val Gly Thr He Ser Wing Asn Ser Asp Thr Thr He Gly Asn 145 150 155 160 ATC ATA GCT GAA GCT ATG GCT AAA GTT GGA AAA GGA GGT GTT ATC ACA 528 He He Wing Glu Ala Wing Ala Lys Val Gly Lys Gly Gly Val He Thr 165 170 175 GTT GAG GAA GCT AAA GGT CTT GAA ACT ACA T? A GAT GTG GTT GAA GGA 576 Val Glu Glu Ala Lys Gly Leu Glu Thr Thu Leu Asp Val Val Glu Gly 180 185 190 ATG AAG TTT GAC CGT GGC TAC CTC TCT CCA TAC TTT GTA ACT AAT CCT 624 Met Lys Phe Asp Arg Gly Tyr Leu Ser Pro Tyr Phe Val Thr Asn Pro 195 200 205 GAG AAA ATG GTT TGT GAA CTT GAT AAC CCT TAT ATC CTT TGT AAT GAG 672 Glu Lys Met Val Cys Glu Leu Asp Asn Pro Tyr He Leu Cyn Asn Glu 210 215 220 AAA AAG ATT ACT AGC ATG AAA GAC ATG CTA CCA ATC TTA GAA CAA GTT 720 Lys Lys He Thr Ser Met Lys Asp Met Leu Pro He Leu Glu Gln Val 225 230 235 240 GCT AAA GTA AAC CGT CCA CTC CTT ATT ATT GCT GAA GAC GTA GAA GGT 768 Wing Lyß Val Asn Arg Pro Leu Leu He He Wing Glu Aßp Val Glu Gly 245 250 255 GAA GCA CTT GCA ACA CTT GTA GTC AAT AAG CTC CGT GGA GCA CTC CAA 816 Glu Ala Leu Ala Thr Leu Val Val Asn Lys Leu Arg Gly Ala Leu Gln 260 265 270 GTT GTA GCC GTA AAA GCT CCT GGT TTT GGT GAA CGC CGT AAA GCT ATG 864 Val Val Wing Val Lys Wing Pro Gly Phe Gly Glu Arg Arg Lys Wing Met 275 280 285 CTT GAA GAT ATT GCT ATC CTT ACT GGA GGA GAA GCA ATA TTT GAA GAT 912 Leu Glu Asp He Wing He Leu Thr Gly Gly Glu Wing He Phe Glu Asp 290 295 300 CGT GGT ATA AAG CTT GAA AAT GTA AGC TTG TCT TCT TTA GGA ACÁ GCT 960 Arg Gly He Lys Leu Glu Asn Val Ser Leu Ser Ser Leu Gly Thr Ala 305 310 315 320 AAA CGT GTA GTT ATT GAC AAA GAA AAT ACT ACT ATC GTT GAT GGT GCT 1008 Lys Arg Val Val He Aep Lys Glu Asn Thr Thr He Val Asp Gly Ala 325 330 335 GGA AAA TCA GAA GAT ATT AAA GCT CGA GTT AAA CAA ATT CGT GCA CAA 1056 Gly Lys Ser Glu Asp He Lys Wing Arg Val Lys Gln He Arg Wing Gln 340 345 350 ATT GAA ACA AGC TCA GAT TAT GAT CGT GAA AAA CTT CAA GAA CGT 1104 He Glu Glu Thr Ser As Asp Tyr Asp Arg Glu Lys Leu Gln Glu Arg 355 360 365 CTT GTT GGT GGA GTA GCT GTT ATC CAT GTT GGA GCT GCT 1152 Leu Ala Lys Leu Val Gly Gly Val Ala Val He His Val Gly Ala Ala 370 375 380 ACT GAA ACT GAA ATG AAA GAG AAG AAG GAT CGT GTA GAA GAT GCT CTA 1200 Thr Glu Thr Glu Met Lyß Glu Lys Lys Asp Arg Val Glu Aßp Ala Leu 385 390 395 400 AAT GCA ACA AGA GCT GCG GTT GAA GAA GGT ATT GTC CCT GGT GGT GGT 1248 Asn Ala Thr Arg Ala Ala Val Glu Glu Gly He Val Pro Gly Gly Gly 405 410 415 ACT GCT TTT GTC CGC TCC ATT AAA GTC CTT GAT GAT ATT AAA CCT GCT 1296 Thr Wing Phe Val Arg Ser He Lys Val Leu Asp Asp He Lys Pro Wing 420 425 430 GAT GAT GAT GAA CTT GCT GGA CTT AAT ATC ATC CGT CGT TCT CTT GAA 1344 Asp Asp Asp Glu Leu Wing Gly Leu Asn He He Arg Arg Ser Leu Glu 435 440 445 GAC CCT TTA CGT CAA ATT GCT GCA AAT GCT GGC TAT GAA GGT TCT ATT 1392 Glu Pro Leu Arg Gln He Wing Wing Asn Wing Gly Tyr Glu Gly Ser He 450 455 460 GTT GTA GAA AAA GTT CGT GAA CCA AAA GAT GGT TTT GGA TTT AAT GCT 1440 Val Val Glu Lys Val Arg Glu Pro Lys Asp Gl -'- Phe Gly Phe Asn Wing 465 470 475 480 GCA TCA GGA GAA TAT GAA GAC CTT ATT AAA GCT GGT GTC ATT GAT CCT 1488 Wing Ser Gly Glu Tyr Glu Asp Leu He Lys Wing Gly Val He Aep Pro 485 490 495 AAA AAA GTT ACA CGT ATT GCA TTA CAA AAT GCA GCA TCA GTA TCC 1536 Lys Lys Val Thr Arg He Wing Leu Gln Asn Wing Wing Ser Val Wing Ser 500 505 510 TTA CTT CTA ACT ACA GAA TGC GCT ATT GCT GAA AAA CCA GAA CCT AAA 1584 Leu Leu Leu Thr Thr Glu Cys Ala He Wing Glu Lys Pro Glu Pro Lys 515 520 525 AAA GAT ATG CCT ATG CCT GGC GGT GGT ATG GGT GGT ATG GGT GGT ATG 1632 Lys Asp Met Pro Met Pro Gly Gly Gly Met Gly Gly Met Gly Gly Met 530 535 540 GAC GGT ATG TAC TAG_1647_Asp Gly Met Tyr 545 (2) INFORMATION FOR SEQ ID NO: 2: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 548 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 2 Met Wing Ser Lys Glu He Leu Phe Asp Wing Lys Wing Arg Glu Lys Leu 1 5 10 15 Be Arg Gly Val Asp Lys Leu Wing Asn Wing Val Lys Val Thr Leu Gly 20 25 30 Pro Lys Gly Arg Asn Val Val He Glu Lys Ser Phe Gly Ser Pro Val 35 40 45 He Thr Lys Aep Gly Val Ser Val Ala Lys Glu He Glu Leu Glu Asp 50 55 60 Lye Phe Glu Asn Met Gly Wing Gln Met Val Lys Glu Val Wing Pro Lye 65 70 75 80 Thr Ser Asp He Wing Gly Asp Gly Thr Thr Thr Wing Thr Val Leu Wing 85 90 95 Gln Wing He Tyr Arg Glu Gly Val Lys Leu Val Wing Wing Gly Arg Asn 100 105 110 Pro Met Wing He Lyß Arg Gly He Asp Lyß Wing Val Val Wing Val Thr 115 120 125 Lys Glu Leu Be Asp He Thr Lyß Pro Thr Arg Asp Gln Lys Glu He 130 135 140 Wing Gln Val Gly Thr He Ser Wing Asn Being Asp Thr Thr He Gly Asn 145 150 155 160 He He Wing Wing Glu Wing Met Wing Lys Val Gly Lys Gly Val Val Th Th 155 170 175 Val Glu Glu Ala Lys Gly Leu Glu Thr Thr Leu Asp Val Val Glu Gly 180 185 190 Met Lys Phe Asp Arg Gly Tyr Leu Ser Pro Tyr Phe Val Thr Asn Pro 195 200 205 Glu Lys Met Val Cys Glu Leu Asp Asn Pro Tyr He Leu Cys Asn Glu 210 215 220 Lys Lys He Thr Ser Met Lys Aep Met Leu Pro He Leu Glu Gln Val 225 230 235 240 Wing Lys Val Asn Arg Pro Leu Leu He He Wing Glu Asp Val Glu Gly 245 250 255 Glu Ala Leu Ala Thr Leu Val Val Asn Lys Leu Arg Gly Ala Leu Gln 260 265 270 Val Val Ala Val Lys Ala Pro Gly Phe Gly Glu Arg Arg Lys Ala Met 275 280 285 Leu Glu Asp He Ala He Leu Thr Gly Gly Glu Wing He Phe Glu Asp 290 295 300 Arg Gly He Lys Leu Glu Asn Val Ser Leu Ser Ser Leu Gly Thr Ala 305 310 315 320 Lys Arg Val Val He Asp Lys Glu Asn Thr Thr He Val Aep Gly Ala 325 330 335 Gly Lys Ser Glu Asp He Lys Wing Arg Val Lys Gln He Arg Wing Gln 340 345 350 He Glu Glu Thr Ser Being Asp Tyr Asp Arg Glu Lys Leu Gln Glu Arg 355 360 365 Leu Ala Lys Leu Val Gly Gly Val Ala Val He His Val Gly Ala Ala 370 375 380 Thr Glu Thr Glu Met Lys Glu Lys Lys Asp Arg Val Glu Asp Ala Leu 385 390 395 400 Asn Ala Thr Arg Ala Ala Val Glu Glu Gly He Val Pro Gly Gly Gly 405 410 415 Thr Ala Phe Val Arg Ser He Lys Val Leu Asp Asp He Lys Pro Wing 420 425 430 Aep Asp Asp Glu Leu Wing Gly Leu Asn He He Arg Arg Ser Leu Glu 435 440 445 Glu Pro Leu Arg Gln He Wing Wing Asn Wing Gly Tyr Glu Gly Ser He 450 455 460 Val Val Glu Lys Val Arg Glu Pro Lyß Asp Gly Phe Gly Phe Asn Wing 465 470 475 480 Wing Ser Gly Glu Tyr Glu Asp Leu He Lys Wing Gly Val He Asp Pro 485 490 495 Lys Lys Val Thr Arg He Wing Leu Gln Asn Wing Wing Ser Val Wing Ser 500 505 510 Leu Leu Leu Thr Thr Glu Cys Wing He Wing Glu Lys Pro Glu Pro Lys 515 520 525 Lys Asp Met Pro Met Pro Gly Gly Gly Met Gly Gly Met Gly Gly Met 530 535 540 Asp Gly Met Tyr 545 (2) IN TRAINING FOR SEQ ID NO: 3: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 306 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) ) TYPE OF MOLECULE: DNA (ix) CHARACTERISTIC: (C) NAME / KEY: CDS (D) LOCATION: 1.306 (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 3 ATG AAC CTG AAA CCT TTG AAT GAC CGT GTT TTA GTA AAA CGT CTT GAA 48 Met Asn Leu Lys Pro Leu Asn Asp Arg Val Leu Val Lys Arg Leu Glu 1 5 10 15 TCT GAA GAA AAA AC GCT GGT GGA CTC TAT ATC CCT GAT ACT GCT AAA 96 Ser Glu Glu Lys Thr Wing Gly Gly Leu Tyr He Pro Asp Thr Wing Lys 20 25 30 GAA AAA CCA TCT CGT GGT GAA GTT GTT GCT GTT GGA CCT GGT AAA CAT 144 Glu Lys Pro Ser Arg Gly Glu Val Val Wing Val Gly Pro Gly Lys His 35 40 45 ACA GAT GAT GGT AAA TTA ATA CCT ATG GCT GTA AAA GCA GGA GAT ACA 192 Thr Asp Asp Gly Lys Leu He Pro Met Wing Val Lys Wing Gly Asp Thr 50 55 60 GTT CTT TTT AAT AAG TAT GCA GGA ACA GAA GTA AAG CTT GAT GGT GTA 240 Val Leu Phe Asn Lys Tyr Ala Gly Thr Glu Val Lys Leu Asp Gly Val 65 70 75 80 GAG CAT CTA GTT ATG CGT GAA GAT GAC ATC CTA GCT GTT ATT ACT GGA 288 Glu His Leu Val Met Arg Glu Asp Asp He Leu Wing Val He Thr Gly 85 90 95 GAA ACT GGC CGC AAG TGA 306 Glu Thr Gly Arg Lys * 100 (2) IN FORMATION FOR SEQ ID NO: 4: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 101 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 4 Met Asn Leu Lys Pro Leu Asn Asp Arg Val Leu Val Lys Arg Leu Glu 1 5 10 15 Ser Glu Glu Lys Thr Wing Gly Gly Leu Tyr He Pro Aßp Thr Wing Lys 20 25 30 Glu Lys Pro Ser Arg Gly Glu Val Val Wing Val Gly Pro Gly Lys His 35 40 45 Thr Asp Asp Gly Lyß Leu He Pro Met Wing Val Lys Wing Gly Asp Thr 50 55 60 Val Leu Phe Asn Lyß Tyr Ala Gly Thr Glu Val Lys Leu Asp Gly Val 65 70 75 80 Glu His Leu Val Met Arg Glu Asp Asp He Leu Wing Val He Thr Gly 85 90 95 Glu Thr Gly Arg Lys 100 (2) INFORMATION FOR SEQ ID NO: 5: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 4972 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simpIe (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 5 AACTCCTGGT CTATCAAGAT CAACTAAAAA ATATTCTTTA TCTAATAGTT 50 GCTCAAAAAT AATTGTACCT ACAGGTAAAT GAAGAATCAA ATCTTCCCCT 100 TTTTTTACCAT GACGCTGGCT CCCTTTACCA CCTTCTCCAT TTTGAGCTCT 150 ATAGTGACGT TGCACACGAA AATCATAAAG GGTTAACAAA CGTGAATCAG 200 CTTTAAAAAT TATATTACCT CCATCTCCTC CATCCCCTCC ATTAGGTCCA 250 CCTTTAGGTA TAAACTTTTC GCGTCTAAAT GAAACACATC CATTTCCACC 300 TTTTCCTGCG CTCACGCTAA TAGTTACTTC ATCAACAAAA CGCATGATTA 350 TCCTTTCAAT AACAAATATC TATTCAATAC TGTTACTAAC TTGTTTACTG 400 TTTTTTTCTAG AAAATTACCT GGCTAATTAT TATAGTTATA TCTAGATTAA 450 TGAAAAAGGA AGAAGTCATT ACACTCCTTC CTTATTAATA GAATCCTGGA 500 ATAATTATTA TACGGTGGGT TGTATATGCA CTCTACTATA TCTTTTACAT 550 TTACGAAAAT ATGTTTCATA AGTTACTATA CCATTAACTT TTGCAAATAA 600 AGTATAGTCT CTTCCCATTC CAACATTTTC TCCAGGATGA ATTTTTGTAC 650 CTAGTTGACG AACAAGGATA TTGCCTGCCA AGACTTTCTG GCCGCCGAAA 700 CGCTTTATAC CACGACGTTG TCCTGGACTA TCTCTACCAT TGCGAGAACT 750 TCCACCAGCT TTCTTATGGG CCATTTTAAT ATCTCCTTAA AGCTGAATAC 800 CTGTTACTTT TAGAGCTGTA TAGTC - TGAC GATGACCTTG GAGTTTACGT 850 GAGTCATTTC TTCTCCACTT TTTAAAAACA AGAATTTTTT TATCACGACC 900 ATGCTCAAGA ACTTTAGCTA TAACTTTAGC ATTATTAATA TATGGTGTTC 950 CAATTTGAGG AGATGAACCA CCAATCATAA AAATTTTATC AAAAAAAATT 1000 TCTGTTCCAA CTTCAGCGTC TATTTTAGAA ACAAAAATTT TAGAACCCTC 1050 TTCAACACAG AATTGTTTTC CACCAGCTTC AATAATTGCG TACATAAATA 1100CAAAAAAGAC AAGAAATACT AATTTGATAT TTTCAATATT 1150 GTCAAGTAGG AACTTTATCT TTAGAATGTT AGATGTAACA ATTTTTTTTAG_1200_AAAAAAAATA TTTTCAATAC AATAGGAAAA GAGGAAAAAA AAAAAGATTT 1250 TTAGAAAAAA TTTTTATTTC TCCAAAAAAT GCAAAAATAT AAAAAATTCT 1300 AATAGGATAG AAGTTATTAC TGTATTGATT TTCAAGACTT ACTTAAAAAT 1350 TTTTATAAAA AAATTTGCAT TCCCCT TTC CCAATTCCCA TAGAGAAGAT 1400 TATTTATCCT AACGATTGGT GGACGCTAAG TCCCTGCTGT TTTGATTATA 1450 TATCAAATGT TGAAACAAAT TTTGTTTAGT TTCTTTTTGT ACTCTAAAAA 1500 GAAGACAAAA AATTCTTTAT AAACTGTACA CTCTAAACAA AATAGTTCAC 1550 AATAAACAGC AATACATTAT AATTAATTGG AGGATACTAT TGTCATGAAC 1600 CTGAAACCTT TGAATGACCG TGTTTTAGTA AAACGTCTTG AATCTGAAGA 1650 AAAAACAGCT GGTGGACTCT ATATCCCTGA TACTGCTAAA GAAAAACCAT 1700 CTCGTGGTGA AGTTGTTGCT GTTGGACCTG GTAAACATAC AGATGATGGT 1750 AAATTAATAC CTATGGCTGT AAAAGCAGGA GATACAGTTC TTTTTAATAA 1800 GTATGCAGGA ACAGAAGTAA AGCTTGATGG TGTAGAGCAT CTAGTTATGC 1850 GTGAAGATGA CATCCTAGCT GTTATTACTG GAGAAACTGG CCGCAAGTGA 1900 AAAAGGCGTA AATAAAAAGA TCGGTGATCT TTAATAATTT TATTCAGTTA 1950 TAATGAAAAC ACTAATTACA CGCACTCTCT GAGAATTTTC TCAGAAAACT 2000 ATATTTAACA ATTCTAAAAT CGATATGTTT TTAGGAGGAA AACCCTAATG 2050 GCTTCTAAAG AAATCCTTTT TGATGCTAAA GCCCGTGAAA AACTTTCACG 2100 AGGTGTAGAT AAACTTGCAA ATGCTGTTAA AGTAACACTT GGACCTAAAG 2150 GCCGTAATGT CGTTATTGAA AAGTCTTTTG GTTCCCCAGT TATTACAAAA 2200 GATGGTGTAT CTGTTGCAAA AGAAATTGAA CTTGAAGATA AGTTTGAAAA 2250 TATGGGCGCT CAAATGGTTA AAGAAGTAGC TCCCAAAACT AGCGATATTG 2300 CTGGTGATGG AACTACAACA GCAACAGTCC TTGCACAAGC TATTTATCGT 2350 GAAGGTGTAA AACTTGTAGC AGCTGGTCGT AATCCTATGG CCATTAAACG 2400 TGGCATAGAT AAAGCTGTTG TTGCTGTTAC TAAAGAACTA AGCGACATTA 2450 CAAAGCCTAC TCGTGACCAA AAAGAAATAG CTCAAGTTGG AACCATTTCT 2500 GCAAACTCTG ATACAACAAT AGGTAATATC ATAGCTGAAG CTATGGCTAA 2550 AGTTGGAAAA GGAGGTGTTA TCACAGTTGA GGAAGCTAAA GGTCTTGAAA 2600 CTACATTAGA TGTGGTTGAA GGAATGAAGT TTGACCGTGG CTACCTCTCT 2650 CCATACTTTG TAACTAATCC TGAGAAAATG GTTTGTGAAC TTGATAACCC 2700 TTATATCCTT TGTAATGAGA AAAAGATTAC TAGCATGAAA GACATGCTAC 2750 CAATCTTAGA ACAAGTTGCT AAAGTAAACC GTCCACTCCT TATTATTGCT 2800 GAAGACGTAG AAGGTGAAGC ACTTGCAACA CTTGTAGTCA ATAAGCTCCG 2850 TGGAGCACTC CAAGTTGTAG CCGTAAAAGC TCCTGGTTTT GGTGAACGCC 2900 GTAAAGCTAT GCTTGAAGAT ATTGCTATCC TTACTGGAGG AGAAGCAATA 2950 TTTGAAGATC GTGGTATAAA GCTTGAAAAT GTAAGCTTGT CTTCTTTAGG 3000 AACAGCTAAA CGTGTAGTTA TTGACAAAGA AAATACTACT ATCGTTGATG 3050 GTGCTGGAAA ATCAGAAGAT ATTAAAGCTC GAGTTAAACA AATTCGTGCA 3100 CAAATTGAAG AAACAAGCTC AGATTATGAT CGTGAAAAAC TTCAAGAACG 3150 TCTTGCAAAA CTTGTTGGTG GAGTAGCTGT TATCCATGTT GGAGCTGCTA 3200 CTGAAACTGA AATGAAAGAG AAGAAGGATC GTGTAGAAGA TGCTCTAAAT 3250 GCAACAAGAG CTGCGGTTGA AGAAGGTATT GTCCCTGGTG GTGGTACTGC 3300 TTTTGTCCGC TCCATTAAAG TCCTTGATGA TATTAAACCT GCTGATGATG 3350 ATGAACTTGC TGGACTTAAT ATCATCCGTC GTTCTCTTGA AGAGCCTTTA 3400 CGTCAAATTG CTGCAAATGC TGGCTATGAA GGTTCTATTG TTGTAGAAAA 3450 AGTTCGTGAA CCAAAAGATG GTTTTGGATT TAATGCTGCA TCAGGAGAAT 3500 ATGAAGACCT TATTAAAGCT GGTGTCATTG ATCCTAAAAA AGTTACACGT 3550 ATTGCATTAC AAAATGCAGC ATCAGTAGCC TCCTTACTTC TAACTACAGA 3600 ATGCGCTATT GCTGAAAAAC CAGAACCTAA AAAAGATATG CCTATGCCTG 3650 GCGGTGGTAT GGGTGGTATG GGTGGTATGG ACGGTATGTA CTAGTCCTAT 3700 CTTCAGTACA ACTTAGATGT ATAAAAACCC CAGAAGCAAT GCTTCCGGGG 3750 TTTTATACTT TCAGCATAAA AAATTAATAT TTAATATACA GACACATTAT 3800 TTTGGTATTT ATTATTTATT ATGATCAAAT ATATAGACTG GATACAAAAA 3850 ACAACAATGA TGTTTAAAAA GGCAGGGATA GATTCACCAA AACTCTCTGC 3900 AGAACTTATA TTAAGTCATG TTTTAAATAT TACACGATTA CAAATAATAA 3950 TGACTCCTTT TGAACCTATT CCAACTAATA GCTACTCAAC GCTTAATGAT 4000 ATCATGTTAA GAAGACTCCA TGGAGAACCA ATTGCATATC TCACAGGGAA 4050 AAAAGAATTT TTTTCACGAG AATTTAAAGT CACTCAAGCC ACACTTATCC 4100 CTCGCCCAGA GACAGAGTTA CTTATAGAAT TTGTATTAAA CCATATTAAC 4150 CCAACACAAC AAATATACTT TGCAGACTTA GGTACAGGTA GTGGGTGTAT 4200 TGCAATTACA CTAGCTGCTG AAAGAAAAAA TTGGTTAGGT ATTGCTACTG 4250 ATATCTCTAG TGAAGCATTA AAAATAGCTA AACTTAATAG TTTAAAAAAT 4300 AACACTCATA GTCAACTACA GTTTCTTCAA TCAGATTTTA CACAACCACT 4350 CTGTCTACCC TCTTCATTAG ACTTATATAT CAGTAATCCT CCATATATAA 4400 GTGAAAATGA ACTGACCTCT CTTCCGCATG AAGTAATATC TTTTGAACCT 4450 AAAATAGCTC TTACACCACA TAAATGTATT CATCTTGATG AAATAAATAC 4500 CGTTTTTACAC TGCTATAAAA AAATTATTAC CCAAGCAGAG ATATCCCTTA 4550 AGCCTGGAGG AATAATAATT TTAGAACATG GAGCAACACA AGCAGAAGCT 4600 ATCTTATTGT TGTTAAAAAA CAACATATGG ACAAATGTAA TAAGTCATAC 4650 TGATCTTACA AATAAAAATC GTTTTATTAC AGCATATAAG TATAAAATAT 4700 AACTTAATTA TGTTGkagAa AAAACAAAAA ATAAAAATAA GATATtAAaT 4750 ATTTtttttA aTAAAATTAA GCAAtTACTA ATATCTTTTT TTGGrTCGtt 4800 yaTtGsATwA GAAACTTTGG rGGrTrrCTa TGAACAAACA ACCATnCAAC 4850 GGCCAAnTAC ATnnCAGGnT TGGGGTCATA GGGGCCACGC TTTATGTACG 4900 TACAACCCCn ACTGAAATTC TGGnTTGnTT TGGGGGGnAA nTGGGTATCG 4950 CAACnCTnTC CCCCCCCCCT GG 4972 (2) INFORMATION FOR SEQ ID NO: 6: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 569 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (x) CHARACTERISTICS: (E) NAME / KEY: CDS (F) LOCATION: 209.569 (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 6 GGTTAAAAAG TAAGGAGAAA AGGTTGGTTA AACCAAGTTT AAAAAATTAA TTTTTTTTTA 60 TTACCCAAAA AAGTTTATTA GATTAAGTAA TATTAATTTG GCCCAAAAAT TTTTTTGGGC 120 ATGGGTTTTT TGCTTTTAAA ATAGAGATGT GTAGGTAACA TTTTTTTCCTC CATGAAATTA 180 TTTTTTAGGA GATGTTATCA TGATGGGG AGT TTG TTT ATT GNT GCG AAC AGG 232 Ser Leu Phe He Xaa Wing Asn Arg 1 5 TAT GAA AAC CCA TAG NAC AGG GNT GGT ACT GTC TCC AAT AAT ATT GCT 280 Tyr Glu Asn Pro * Xaa Arg Xaa Gly Thr Val Ser Asn Asn He Wing 10 15 20 AAC GCA AAT ACC ATT GGG TAT AAG CAG CA CAG GTA GTG TTT CAA GAC 328 Asn Wing Asn Thr He Gly Tyr Lys Gln Gln Gln Val Val Phe Gln Aßp 25 30 35 40 CTG TTT AGT CAA GAT TTA GCA ATA GGT TTT ACT GGA AGT CAG GGG CCA 376 Leu Phe Ser Gln Asp Leu Ala He Gly Phe Thr Gly Ser Gln Gly Pro 45 50 55 AAC CAG GCT GGT ATG GGA GCA CAG GTG GGA AGT GTT CGC ACA ATT TTT 424 Asn Gln Wing Gly Met Gly Wing Gln Val Gly Ser Val Arg Thr He Phe 60 65 70 ACA CAG GGT GCT TTT GAA CCT GGC AAT AGT GTA ACA GAT CCT GCT ATT 472 Thr Gln Gly Wing Phe Glu Pro Gly Asn Ser Val Thr Asp Pro Wing He 75 80 85 GGT GGA AAA GGT TTT TTT CAG GTT ACA TTA GAG GAT AAA GTA CAC TAT 520 Gly Gly Lys Gly Phe Phe Gln Val Thr Leu Glu Asp Lys Val His Tyr 90 95 100 ACA CGA GCA GGG AAT TTT CGT TTT ACT CAA GAT GGT TTT TTA AAT GAT C 569 Thr Arg Wing Gly Asn Phe Arg Phe Thr Gln Asp Gly Phe Leu Asn Asp 105 110 115 120 (2) INFORMATION FOR SEQ ID NO: 7: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 123 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 7 Ser Leu Phe He Xaa Wing Asn Arg Tyr Glu Asn Pro * Xaa Arg Xaa 1 5 10 15 Gly Thr Val Ser Asn Asn He Wing Asn Wing Asn Thr He Gly Tyr Lys 20 25 30 Gln Gln Gln Val Val Phe Gln Asp Leu Phe Ser Gln Asp Leu Ala He 35 40 45 Gly Phe Thr Gly Ser Gln Gly Pro Asn Gln Wing Gly Met Gly Ala Gln 50 55 60 Val Gly Ser Val Arg Thr He Phe Thr Gln Gly Wing Phe Glu Pro Gly 65 70 75 80 Asn Ser Val Thr Asp Pro Wing He Gly Gly Lys Gly Phe Phe Gln Val 85 90 95 Thr Leu Glu Asp Lys Val His Tyr Thr Arg Wing Gly Asn Phe Arg Phe 100 105 110 Thr Gln Asp Gly Phe Leu Asn Asp (2) INFORMATION FOR SEQ ID NO: 8: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1450 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple ( D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (ix) CHARACTERISTIC: (A) NAME / KEY: CDS (B) LOCATION: 3..414 (ix) FEATURE: (A) NAME / KEY: CDS (B) ) LOCATION.- 1083..1450 (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 8 GA TCT AAA GAG TCT ACA TAT ATT GCC CGA Í.TT GAA AAT TCT ACA AGT 47 Being Lyß Glu Being Thr Tyr He Wing Arg He Glu Asn Being Thr Ser 1 5 10 15 GAA AAA ACA CTA AAT GAT CTT GAT ATA CTT TTA AAA GAT GTG ATG TTA 95 Glu Lys Thr Leu Asn Asp Leu Asp He Leu Leu Lys Asp Val Met Leu 20 25 30 ACA TCA AAA AAG CAT GAA TCA CGT AGA CTT GCA GAG TCT GTA CAT CAA 143 Thr Ser Lys Lys His Glu Ser Arg Arg Leu Wing Glu Ser Val His Gln 35 40 45 AAT ATT CTT ACC CAC CTT ATA CAA AAA AAT TAT AAT ACT CAC AAT GGT 191 Asn He Leu Thr His Leu He Gln Lys Asn Tyr Aßn Thr His Asn Gly 50 55 60 GGG ATA AAA TCT GCA CCT TTT CAT GTT CTT ATA GGA CCC AAA ATA CCA 239 Gly He Lys Ser Wing Pro Phe His Val Leu He Gly Pro Lys He Pro 65 70 75 AGT ATT CTT GTT GAA GTA GGT TAC TGT AGT AAT AAA GCT GAA GCA CAG 287 Ser He Leu Val Glu Val Gly Tyr Cys Ser Asn Lys Ala Glu Ala Gln 80 85 90 95 CGT CTG GCA TCT AGT AAT TAC CAA AAA GCA TTA ATA GAA GGA TTA GCT 335 Arg Leu Wing Being Ser Asn. Tyr Gln Lys Wing Leu He Glu Gly Leu Wing 100 105 110 AAA GGT ATT TTC TGT TAC CTA AAA AAA CTA CAT CAC CTT GAT ATT TAC 383 Lyß Gly He Phe Cys Tyr Leu Lyß Lys Leu His His Leu Asp He Tyr 115 120 125 TCT AGT TTT ATY CTA TCT AAT TGC ACT TAA T AGCTTGGACA ATTATTATAT 434 Ser Ser Phe He Leu Ser Asn Cys Thr * 130 135 GAAGGGTATC CATGTGAAGG TACCTGGTTA AGCTTTTAAA TGTAAAAATT ATGCAACCAT 494 ACYTTATTCC TTCAGAGGAG CTTCATTATG AAAGTAAAAA CTCTTTCCAT GGCTATTTTA 554 GCTTGTTTAT TAGTAGCTAA CAGTGCATTT TCGGCTGACT TCCCTATTGG TGTCTTTAAT 614 TCTCAATCCA TTGCCATGGA GAGTGAAGCA GCTAAGGCCG CTCAAAAAAA ATTACAATCA 674 GAATTTGGTA ATGAAAAAAC ACAACTTGAA AACAAGCAAA AGWTTGCMAA CAAAAGCTGA 734 TGATTTACAA GCTWAGTCAG CAGCTATGTY TAACCAAGCA CGTGAAGATA AACAAAGAGA 794 ATTTCTTGAA CTTCGTCGTA ATTTCGAAGA AAAATYTCGT GACTTTGCAA TACGTGTCGA 854 ACAAGCTGAA AACACATTAC GTCAATATNT AGCTGAACAA ATNTATNTTG CTGCTGAAAC 914 TATAGCAAAA AAGAAAGGGT TAAACTTGTT TTGATAGTGT TAGGGAAGTG TAATGTACCT 974 TGAAAAAAAT TTAGATATTA CAAAGAAATT YTTGAAGCCA TAAATGCTGC ATGGAAAAAA 1034 GGTGGAAGTA AACTTCCAGA GATGGCAAAC CGGAAAAAAT AACAG ATG CCC CAG TAT 1091 Met Pro Gln Tyr 1 AAA CTT TCA GAA ATT GCT AAA CTT TTA AAC TTA ACA TTA CAA GGT GAT 1139 Lys Leu Ser Glu He Wing Lys Leu Leu Asn Leu Thr Leu Gln Gly Asp 5 10 15 20 GAT ATT GAA GTT GTA GGC GTA AAT ACA CTT CAA GAT GCA TCA CCA AAT 1187 Asp He Glu Val Val Gly Val Asn Thr Leu Gln Asp Ala Ser Pro Asn 25 30 35 GAG ATA AGT TTT CTA GCA AAT GCT AAA TAT ATT CAC CAG CTT GTT TTG 1235 Glu He Ser Phe Leu Wing Asn Wing Lys Tyr He His Gln Leu Val Leu 40 45 50 TCA CAG GCT GGT GCT ATT ATT CTT TCA AAA GAA TAT GCT AGT CGT GTT 1283 Ser Gln Ala Gly Ala He He Leu Ser Ly «« Glu Tyr Ala Ser Arg Val 55 60 65 CCA CGA GCA CTA ATC AGT ACT GAA CCA TAT AGA GAT TTT GGT AGA GTT 1331 Pro Arg Ala Leu He Ser Thr Glu Pro Tyr Arg Asp Phe Gly Arg Val 70 75 80 CTT TCT TTA TTC TCT ATA CCT CAA GGA TGT TTT GAT GGT ATA AGT CAT 1379 Leu Ser Leu Phe Ser He Pro Gln Gly Cys Phe Asp Gly He Ser His 85 90 95 100 CAÁ GCT TAT ATA CAC CCT ACA GCA CAA GTC TCT AAA ACA GCT ACT ATC 1427 Gln Wing Tyr He Hiß Pro Thr Wing Gln Val Ser Lyß Thr Wing Thr lie 105 110 115 TAT CCT TTn GTT TTT ATA GGA TC 1450 Tyr Pro Xaa Val Phe He Gly 120 (2) INFORMATION FOR SEQ ID NO: 9: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 137 amino acids (B) TYPE: amino acid (D) TOPOLOGY: (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 9 Ser Lys Glu Ser Thr Tyr He Wing Arg He Glu Asn Ser Thr Ser Glu 1 5 10 15 Lys Thr Leu Asn Asp Leu Asp He Leu Leu Lys Asp Val Met Leu Thr 20 25 30 Ser Lys Lys His Glu Ser Arg Arg Leu Wing Glu Ser Val His Gln Asn 35 40 45 He Leu Thr His Leu He Gln Lys Asn Tyr Asn Thr His Aen Gly Gly 50 55 60 He Lys Ser Ala Pro Phe His Val Leu He Gly Pro Lys He Pro Ser 65 70 75 80 He Leu Val Glu Val Gly Tyr Cys Ser Asn Lys Ala Glu Ala Gln Arg 85 90 95 Leu Wing Being Ser Asn Tyr Gln Lys Wing Leu He Glu Gly Leu Wing Lys 100 105 110 Gly He Phe Cys Tyr Leu Lys Lys Leu His His Leu Asp He Tyr Ser 115 120 125 Being Phe He Leu Being Asn Cys Thr * 130 135 (2) INFORMATION FOR SEQ ID NO: 10: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 123 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 10 Pro Gln Tyr Lys Leu Ser Glu He Wing Lys Leu Leu Asn Leu Thr Leu 1 5 10 15 Gln Gly Asp Aep He Glu Val Val Gly Val Asn Thr Leu Gln Asp Wing 20 25 30 Ser Pro Aen Glu He Be Phe Leu Wing Aen Wing Lys Tyr He His Gln 35 40 45 Leu Val Leu Ser Gln Ala Gly Ala He He Leu Ser Lys Glu Tyr Ala 50 55 60 Ser Arg Val Pro Arg Ala Leu He Ser Thr Glu Pro Tyr Arg Asp Phe 65 70 75 80 Gly Arg Val Leu Ser Leu Phe Ser He Pro Gln Gly Cys Phe Asp Gly 85 90 95 He Ser His Gln Ala Tyr He His Pro Thr Ala Gln Val Ser Lys Thr 100 105 110 Wing Thr He Tyr Pro * Val Phe He Gly 115 120 (2) INFORMATION FOR SEQ ID NO: 1 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 559 base pairs (B) TYPE: nucleic acid (C) FILAMENT: simpIe (D) TOPOLOGY: linear (ii) ) TYPE OF MOLECULE: DNA (ix) CHARACTERISTIC: (A) NAME / KEY: CDS (B) LOCATION.-3..557 (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 1 1 GA TCA AAG CCG CAT TTA CNG CAA GAG T.'A GAA ATT GAA GTT TTG AAA 47 Ser Lys Pro His Leu Xaa Gln Glu Leu Glu He Glu Val Leu Lys 1 5 10 15 AAA GAA GAC TTT GGG CGT CAT ATT GTT AAA TTA TGC TGG AAA GGT TCT 95 Lys Glu Asp Phe Gly Arg His He Val Val Leys Cys Trp Lys Gly Ser 20 25 30 TTA TCA AAT ATC TTT TTT TCC TAT GGG GAT ATC CCG CAC CCA CCT TAT 143 Leu Ser Asn He Phe Phe Ser Tyr Gly Asp He Pro His Pro Pro Tyr 35 40 45 ATA CAT CA AGT AAT AAG GTT CAG GAT AAG GAA AGA TAT CNT ACN GTA 191 He His Gln Ser Asn Lys Val Gln Asp Lys Glu Arg Tyr Xaa Xaa Val 50 55 60 TAC TCT ATA TTA CAT AAN CTG GGT TCT GTA GCA GCT CCT ACA GCT GGA 239 Tyr Ser He Leu His Xaa Leu Gly Ser Val Ala Wing Pro Thr Ala Gly 65 70 75 TTA CNC TTT TCT GAA ACT AGC CGT NAT AAA TTA CAC AAA NAT GGT ATT 287 Leu Xaa Phe Ser Glu Thr Ser Arg Xaa Lys Leu His Lyß Xaa Gly He 80 85 90 95 AGT TGG GCA TAA ATC CCT CTT CAC GTG GGA TAT GGA ACA TTC AGT CCC 335 Ser Trp Wing * He Pro Leu Hiß Val Gly Tyr Gly Thr Phe Ser Pro 100 105 110 GTC CTC TGC AAT GAC ATC CCA AAA CAT CTT ATC CNT TCT GAG TTT GTT 383 Val Leu Cyß Aßn Asp He Pro Lys His Leu He Xaa Ser Glu Phe Val 115 120 125 CAC TTT CCT GAA ACT ACN TTT TCC ACT ATA TTA AAT GCA CGG TTT GCA 431 His Phe Pro Glu Thr Xaa Phe Ser Thr He Leu Asn Ala Arg Phe Wing 130 135 140 NGG GAA TAC CTA TGT TCT GCC ATA GGG GAC CCA CTG TTG TCC CCA CCA 479 Xaa Glu Tyr Leu Cys Ser Wing He Gly Asp Pro Leu Leu Ser Pro Pro 145 150 155 TTG GAN GGG TGT TAT CTT ACC CCT TTC GCC CGG GGT TCC CCT CCC CAA 527 Leu Xaa Gly Cys Tyr Leu Thr Pro Phe Wing Arg Gly Ser Pro Pro Gln 160 165 170 175 CCC TAT TCC ATT GNG TTT TCC TCT CAA ATT AT 559 Pro Tyr Ser He Xaa Phe Ser Ser Gln He 180 185 (2) INFORMATION FOR SEQ ID NO: 12: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 185 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (x) SEQUENCE DESCRIPTION: SEQ ID NO: 12 Ser Lys Pro His Leu Xaa Gln Glu Leu Glu He Glu Val Leu Lye Lys 1 5 10 _- > Glu Asp Phe Gly Arg His He Val Val Leys Cys Trp Lys Gly Ser Leu 20 25 30 Being Asn He Phe Phe Being Tyr Gly Aßp He Pro His Pro Pro Tyr He 35 40 45 His Gln Ser Asn Lys Val Gln Asp Lys Glu Arg Tyr Xaa Xaa Val Tyr 50 55 60 Ser He Leu 2-ie Xaa Leu Gly Ser Val Ala Ala Pro Thr Ala Gly Leu 65 70 75 80 Xaa Phe Ser Glu Thr Ser Arg Xaa Lys Leu His Lys Xaa Gly He Ser 85 90 95 Trp Wing * He Pro Leu His Val Gly Tyr Gly Thr Phe Ser Pro Val 100 105 110 Leu Cys Asn Asp He Pro Lyß His Leu He Xaa Ser Glu Phe Val His 115 120 125 Phe Pro Glu Thr Xaa Phe Ser Thr He Leu Aßn Ala Arg Phe Ala Xaa 130 135 140 Glu Tyr Leu Cys Ser Wing He Gly Aßp Pro Leu Leu Ser Pro Pro Leu 145 150 155 160 Xaa Gly Cys Tyr Leu Thr Pro Phß Wing Arg Gly Pro Pro Pro Gln Pro 165 170 175 Tyr Ser He Xaa Phe Ser Ser Gln He 180 185 (2) IN TRAINING FOR SEQ ID NO: 13: (i) SECTION CHARACTERISTICS: (A) LENGTH: 477 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear ( ii) TYPE OF MOLECULE: DNA (ix) CHARACTERISTIC: (A) NAME / KEY: CDS (B) LOCATION: 2.294 (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 13 T ATA AAA CA'i 'TAG CGN CTT TNG TAT TTG GAC TTC AAA AAA ATT TTT 46 He Lys His - * * Leu * Tyr Leu Asp Phe Lys Lys He Phe 1 5 10 15 AAT TAT ATA GGA GAA CAT TCA CCA TTA AAA CGT AAT GTA ANT ATG GAA 94 Asn Tyr He Gly Glu His Ser Pro Leu Lys Arg Asn Val * Met Glu 20 25 30 GAT GTA GGT AAA TCT GCT GTT TTT TTA GCT TCA GAC CTN TCA TCA GGA 142 Asp Val Gly Lys Ser Wing Val Phe Leu Wing Ser Asp * Ser Ser Gly 35 40 45 GTA ACC GGT GAA TTN TTT TTG TTG ATG CTG GNA CAA TAA TTT AGG TAT 190 Val Thr Gly Glu * Phe Leu Leu Met Leu * Gln * Phe Arg Tyr 50 55 60 TTA ACC ATA CAT GCT TTA TAC TAC TAC ATA TTG TGA GTT ACA ATA GCC ATA 238 Leu Thr He Hie Wing Leu Tyr Asn He Leu * Val Thr He Wing He 65 70 75 ACA CAT TTA TAT TCT ATA TAA TAA CAG TAG AAT AAT AAT AGA ATA TTT 286 Thr Hie Leu Tyr Ser He * * Gln * Asn Asn Asn Arg He Phe 80 85 90 95 TTT ATG ACC ATTTGTATCT ATACAATAGT AAATAGATTA ATACATATAA GACTATATTC 344 Phe Met Thr TTTTTGAGAG CAACTTAAAG GAGCGGTTAT GGCTTTAGTT ACAAAAGAAG AAGTACTTCA 404 ATACCATAGT GAACCCCGAC CAGGTAAACT TGAAGTATTT TCTATAAAAC CATGTAAAAC 464 ACAAAAAGAT CC 477 (2) INFORMATION FOR SEQ ID NO: 14: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 97 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 14 He Lys His * Xaa Leu Xaa Tyr Leu Asp Phe Lyß Lys He Phe Asn 10 15 Tyr He Gly Glu His Ser Pro Leu Lys Arg Aßn Val Xaa Met Glu Asp 20 25 30 Val Gly Lyß Ser Ala Val Phe Leu Ala Ser Aßp Xaa Ser Ser Gly Val 35 40 45 Thr Gly Glu Xaa Phe Leu Leu Met Leu Xaa Gln * Phe Arg Tyr Leu 50 55 60 Thr He His Wing Leu Tyr Asn He Leu * Val Thr He Wing He Thr 65 70 75 80 His Leu Tyr Ser He * * Gln * Asn Asn Asn Arg He Phe Phe 85 90 95 Met (2) INFORMATION FOR SEQ ID NO: 15: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 525 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: sim? Le (D) TOPOLOGY: linear ( ii) TYPE OF MOLECULE: DNA (ix) CHARACTERISTIC: (A) NAME / KEY: CDS (B) LOCATION: 2..525 (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 15 G GAA TTG TTA GTA TTC TCC CAG AAC AGA AGC CAA AAT ATT TGG CTA 46 Glu Leu Leu Val Phe Ser Gln Asn Arg Ser Gln Asn He Trp Leu 1 5 10 15 CTT ACA TTA CCT ATT TTT GTG TTA GGT ATA GCA CAA GGT ATA TCA TTT 94 Leu Thr Leu Pro He Phe Val Leu Gly He Wing Gln Gly He Ser Phe 20 25 30 CCT TTA GTA AAC AGC CAC ATT ACA TCA CTT GCA CCA ACA TCC AAC AGA 14"> Pro Leu Val Asn Ser His He Thr Ser Leu Wing Pro Thr Ser Asn Arg 35 40 45 GCT ATT GTT ATG GCT ATA AAC AGT ACA TTT ATG AGG TTA AGT CAG AGT 190 Wing Hem Val Met Wing He Asn Ser Thr Phe Met Arg Leu Ser Gln Ser 50 55 60 ATT TCG CAA ATG GTT TTT GGT ATT GGA TGG TCA TTT TTT GGT TGG CCT 238 Be Ser Gln Met Val Phe Gly He Gly Trp Ser Phe Phe Gly Trp Pro 65 70 75 GGT CCT TTT ATA TTT GGT CTT TTT ACT TCT ATT ATA TTA GCC CTC TTA 286 Gly Pro Phe He Phe Gly Leu Phe Thr Ser He He Leu Ala Leu 'Leu 80 85 90 95 ATT ATG AAG TAT TTT CAA GAT GTA ACC CAA TAT CAC CTA TTT TTG ATA 334 He Met Lys Tyr Phe Gln Asp Val Thr Gln Tyr His Leu Phe Leu He 100 105 110 AGT AGT AAA TTT TAT TAT TAA AAA GCT TAG TTA GTT AAG ATT ACA TAT 382 Ser Ser Lys Phe Tyr Tyr * Lys Ala * Leu Val Lys He Thr Tyr 115 120 125 ATT ATA TAC AAT TAC TAT AAC ATT AAC TAA TTA CTA ACT ATT ACT TCC 430 He He Tyr Asn Tyr Tyr Asn He Asn * Leu Leu Thr He Thr Ser 130 135 140 AAT TGA TTA ATT GAT GCT ATT TAA AGA GGA TAT ATT AAT GAT GTC ATG 478 Asn * Leu He Asp Ala He * Arg Gly Tyr He Asn Asp Val Met 145 150 155 GCT CAC AAT AGG TGT TAT CCT TGG ATT AGT GCA TGG GAT CCA GGT CC 525 Wing His Asn Arg Cys Tyr Pro Trp He Ser Wing Trp Asp Pro Gly 160 165 170 (2) INFORMATION FOR SEQ ID NO: 16: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 174 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 16 Glu Leu Leu Val Phe Ser Gln Asn Arg Ser Gln Asn He Trp Leu Leu 1 5 10 15 Thr Leu Pro He Phe Val Leu Gly He Wing Gln Gly He Ser Phe Pro 20 25"30 Leu Val Asn Ser His He Thr Ser Leu Wing Pro Thr Ser Asn Arg Wing 35 40 45 He Val Met Wing He Asn Being Thr Phe Met Arg Leu Being Gln Being He 50 55 60 Being Gln Met Val Phe Gly He Gly Trp Being Phe Phe Gly Trp Pro Gly 65 70 75 80 Pro Phe He Phe Gly Leu Phe Thr Ser He He Leu Ala Leu Leu He 85 90 95 Met Lyß Tyr Phe Gln Asp Val Thr Gln Tyr His Leu Phe Leu He Ser 100 105 110 Being Lyß Phe Tyr Tyr * Lyß Ala * Leu Val Lyß He Thr Tyr He 115 120 125 He Tyr Asn Tyr Tyr Aßn He Asn * Leu Leu Thr He Thr Ser Asn 130 135 140 * Leu He Asp Ala He * Arg Gly Tyr He Asn Asp Val Met Wing 145 150 155 160 His Asn Arg Cye Tyr Pro Trp He Ser Wing Trp Asp Pro Gly 165 170 (2) INFORMATION FOR SEQ ID NO: 17: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 846 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 17 TATTTACTCG CGCGGCCGGG CGTCTTACAC AAATGGATCC CTTGCANTAA TCCAAGGATA 60 ACNCCTATTG TGANCCATGA ACATCATCAN NATATCCTCT TTANATAGCA TCNANNNNTC 120 AANNGGAATT AACAGTTACT ANNTAGTTAA TGTCATAGTA ATTGTCNATA ATATATGTAA 180 TCTTAACTAA CTAAGCTNNT TAATAATAAA ATTNACTACT TATCAANAAT AGGTGATATN 240 GGGTTACATC TTGAAAATAC TTNCCATAAT TANGAGGGCT AATATAATNG AANTAAAAAG 300 ACCANATATA AAAGGACCAG GCCAACCAAA AAATGACCAT CCAATACCNA AAACAATTGG 360 CGAAAATACT CTGACTTAAC CTCANAAATG TACTGTTTAT AGCCATATCA ATAGCTCTGT 420 TGGATGTNGG NGCAATTGAT GTAATGTGGC TGTNTACTAN ANGAAATGAT NTACCTCGTG 480 CTATNCCTAN NACAANAATA NGTAATGTAA GTANCCNAAT ATCTTGGCTT TGTAATGGGA 540 GAATAATNNC AAGTCCTTGG GAAATNAANT TACNNCCAGC CAGCTATNNT AAGCAGTTCT 600 NTGGTGACTA TACGTCCTAC TNAANTCGTG CCAAAGATTA AATANNCGAT AATCGCNCTN 660 CCTAAANCAN GCAATACTAA AATGGTTTCT NCCTANCTTG GNATANGGTG GAAGCNCGGA 720 CAGAATTNAN TTCGCNANTT TANANNGGAA NATNCGTNAA NTTANTCGGG GCCCANNCCN 780 AAATTCCTNA NTCNATANAN NAACTNNCTN CTNTAAAANG GCCNACTGGA NTNGTTAAAT 840 GAAATA 846 (2) INFORMATION FOR SEQ ID NO: 18: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 855 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 18 GATTNTTTAT CGATCACTNT AGACGCGATT TGGGNAACAC TTACCTGGTA NCCACCCGGG 60 TGGAAAAATC GATGGGCCCG CGGCCGCTCT AGAAGTACTC TCGAGAAGCT TTTTGAATTC 120 TTTGGATCCT CAACACAGGG TATGGATTAA AACAACTTTA GCTCTAACAG GAGCATTTTA 180 TAATATATTC CCTGGTAGAA CAATATCTAC TCAAGAAAAT CTGTCTATTG GTTTTCAACT 240 AAAAAAAACT TTTAAACCTT TTCATTGGAC CATCTTACTC TTAGATGAAC ATTATATGTC 300 TTCGCCAAGA ATTGCAGCAG CAATTATGCC TGCACAGCTT GCTGGAGTTA AAAACATTAT 360 AGCTGTTTGG ACCAGTAAAA ATAACCGACT GACCGCTGAA AAAATCTCAC CTGCTTTACT 420 AACAACATTA GAACTTTCAG GAGTTAACAT AGCCCTAACA CTTACCCACA CTGAAACTGA 480 ACTTCTTATT CATCAATTAA TGAAAATAGG TATTGGAAAC CTGTTATATT TTTTAAAAGA 540 AGAAGACATA CTACATATAT CTACTATACC TGTACTACCT TTCTGGAAAG AATATACTTC 600 TCATCGACTT GTTATAGAAA AAGATGCTGG CNTTAATACA GAAATCCTCC AATGGGCNCA 660 TCCTCATTCA ATTATTGAAC AAATAGCAAC AGAACCATAC TCTGAAANAT ATCCCAGATG 720 CACTTTACTG TGCTAGCTCA TCCANTAAAA ACTATNCTCA TANAGNATCC CCAGAATTTT 780 TCATNATGGA CTTGAACCTA TTTGGATTCA NCCCAACNCT TCCTCCAANC CTCCTTTCTC 840 CATACACCAT GGGGA 855 (2) IN TRAINING FOR SEQ ID NO: 19: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1082 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) ) TYPE OF MOLECULE: DNA (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 19 TATCTNGTTG ANTCAATAAA ACTTTTGGGG CCCNTNAAAN TTTCATNANN AAAAAAACAA 60 NATTNCTGGG GGNCCCNTCC CAAAAAANNC AATCANTNNG AANCTTGNCT TCTTATTNNG 120 NTTTTNANAC TATAATATNT NTTATCNATA ATNN? TCNNT ATACTNATTT CTNATTCANT 180 NACANNGGNN AGNAANNTTA ATCTNAAANA CTNCNAAGGG GGNNNTNATA NTNTTTNTTT 240 NTTTNTCCCN TNNAATNNAT AACCNNNCAC CCNNATTANT TNNAATNNAT ACCATANCNN 300 CCTTTCAAAC TGTACACATA NTANNNAANN ACACTCNANC NTTTTNCATC CTCTCTANTN 360 CCNACTCCNA TNNANCTNTT CCCCCATNCC TATNTNTCNC TGCTTCCCAG NTTNNACNTN 420 NCTTNNTTTC ACANTATTCC TATCCAANCT AACATNTNTN NTNTCNTNCT CCTTNTNTNT 480 TATNTNTTTC TNNTACCTNN CACTGACANT CTATNANTNA NNTCNNATAC TNNTATANCT 540 NTANGCNANT NTATCTANAA NTNTANCNNN NNATCNTNAC NGCCGTNNAT NTNNNNNCAN 600 TTANNTANNN CTANCNTNNC CAANNNCNTA TNTATNAATA ACNACTATCC NATATTNNAT 660 TNNNTNNTNT CNTANNCAAA TNATTTANGC NCACNNCACT ANGTNATATN ANNATTNTAT 720 ATTNTGAANC TTCTNGGCTT CNCNAATANT ACCANTNNNC ANCNTCNNNT NCATCTNNNT 780 NTACTTCNTA CCATANCGCT CTCNAGNNTC ACTACTTCTA NTAGTNATCN TCTACTGCCN 840 ATGGCNNNNN GCNNNNCGAN AGNTATNCAC NTACANTNNC NTCTACTATN TANATCTANN 900 NCNTCCGNNG CCTNCNGTAC GNNTNGGCNA ANTCGNNTAC TTTNCNTNTA TCTAGTCNCA 960 TCAGNNNTNG ANTCCTCAAN CNNGCTCTAN TTACATGTNN NNTNATGCNC TANANCGNNA 1020 CNTCTATCCT TCNANTCTGC NCTNANTNTA TANACTCTNN NNNATCNNCN AANCTATNTC 1080 CC 1082 (2) IN TRAINING FOR SEQ ID NO: 20: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 354 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) ) TYPE OF MOLECULE: DNA (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 20 CTCCCNTNNC NCTAAGTGGA TCGCGCGCT GCAGGTCGAC ACTAGTGGAT CTTGATATAC 60 TTTTAAAAGA TGTGATGTTA ACATCAAAAA AGCATGAATC ACGTTAGACT TGCAGAGTCT 120 GTACATCAAA ATATTCTTTA CCCACCTTAA TACGAAAANA AATNNTTATN CNCCNCNATG 180 GGTGGGGNTN AAATCCTNGC CCCNTTNCCC TGTTCNTTTA GGGAACCCCC NAATTCCCCN 240 NGTTATTCCT CTGTTTGAAA NTTCTGGTTN CCCGGCCCTN TNACCAANAG CTTGANNNCC 300 NCCCCGTCCT GGGGCATCCT CNTGTTTATT TTCCCTCNAN CNCCCCCTTN ACTN 354 (2) INFORMATION FOR SEQ ID NO: 21: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 477 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) ) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 21 GGATCTTTTT GTGTTTTACA TGGTTTTATA GGAAATACTT CAAGTTTACC TGGTCGGGGT 60 TCACTATGGT ATTGAAGTAC TTCTTCTTTT GTNACTAAAG CCATAACCGC TCCTTTAAGT 120 TGTTCTCAAA AAGAATATAG TCTTATATGT ATTAATCTAT TTACTATTGT ATAGATACAA 180 TAGGTCATAA AAAATATTCT ATTATTATTC TACTGTTATT ATATAGAATA TAAATGTGTT 240 ATGGCTATTG TAACTCACAA TATGTTGTAT AAAGCATGTA TGGTTAAATA CCTAAATTAT 300 TGTNCCAGCA TCAACAAAAA NAATTCACCG GTTACTCCTG ATGANAGGTC TGAAGCTAAA 360 AAAACAGCAG ATTTACCTAC ATCTTCCATA NTTACATTAC GTTTTAATGG TGAATGTTCT 4 0 CCTATATAAT TAAAAATTTT TTTGAAGTCC Z-AATACNAAA GNCGCTAATG TTTTATA 477 (2) INFORMATION FOR SEQ ID NO: 22: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 568 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (i) ) TYPE OF MOLECULE: DNA (xi) SEQUENCE DESCRIPTION: SEQ ID NO: 22 GATCATTTAA AAAACCATCT TGAGTAAAAC GAAAATTCCC TGCTCGTGTA TAGTGTACTT 60 TATCCTCTAA TGTAACCTGA AAAAAACCTT TTCCACCAAT AGCAAGATCT GTTACACTAT 120 TGCCAGGTTC AAAAGCACCC TGTGTAAAAA TTGTGCGAAC ACTTCCAACC TGTGCTCCCA 180 TACCAGCCTG GTTTGGCCCC TGACTTCCAG TAAAACCTAT TGCTAAATCT TGACTAAACA 240 GGTCTTGAAA CACTACCTGT TGCTGCTTAT ACCCAATGGT ATTTGCGTTA GCAATATTAT 300 TGGAGACAGT ACCANCCCTG TNCTATGGGT TTTCATACCT GTTGGCANCA ATAAACAAAC 360 TCCCCATCAT GATAACATCT CCTAAAAAAT AATTTCATGG NGGNAAAAAT GTTACCTACA 420 CATCTCTATT TTNAAAGCAA AAAACCCATG CCCAANAAAA TTTTTGGGCC NAATTAATAT 480 ACTTAATCTA ATAAACTTTT TTGGGTAATN AAAAAAAATT AATTTTTTAA ACTTGGTTTN 540 ACCAACCTTT TCTCCTTACT TTTTAACC (2) INFORMATION FOR SEQ ID NO: 23: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 477 base pairs (B) TYPE: nucleic acid (C) FILAMENTO: simple (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: DNA (xi) DESCRIPTION OF SEQUENCE: SEQ ID NO: 23 GGTACCCCAC CCGC 7T-.C - AA AATCGATGGG CCCGCGGCCG CTCTAAAANT 50 ACTCTCGAGA AGCTTTTTGA ATTCTTTGGA TCCCCAGGAA TAACTTGTTG 100 ACGGAATTTT ACATTTTCTA TCCCTGCAAA TANAAAAACT TTACCTTGTA 150 GTTCATTAAT AGGAAAAGAT TGGAGTACTG TGATTCCACC TGATTGCGCC 200 ATAGCTTCTA AAATTAGAAC TCCAGGCATG ACAGGAAATC CAGGGGAAAT 250 GACCCNGAAA AAATGGTTCA TTAATACTAA CATTTTTATA AGCTTTAATA 300 TATTTGCCAG CATTAAATTC AATAACTCTA TCTACAATTA AAAAGGGATA 350 ACGGTGGGGA ATTTACTGTA AAATTTCTTG GATATTTTGG AGGTATGGAT 400 GGGGACATTA ATTTTCCTAT ATATATGCTC TTTTTCTTTT CNAAAATTTT 450 TCAGCTTTTT TATCCCNTAA AAACCTC 467

Claims (90)

1. A vaccine composition for the prophylaxis or treatment of infection in an animal or bird by Lawsonia intracellularis or related microorganism, said vaccine composition comprising an immunogenic, non-pathogenic form of L. intracellularis or related microorganism or an immunogenic component thereof and one or more carriers, diluents and / or auxiliaries suitable for veterinary or pharmaceutical use.

2. A vaccine composition according to claim 1, wherein the composition is for the prophylaxis or treatment of infection in pigs by L. intracellularis or related microorganism.

3. A vaccine composition according to claim 2, wherein the non-pathogenic form of L. intracellularis or related microorganism is an attenuated species of the microorganism.

4. A vaccine composition according to claim 2, wherein the non-pathogenic form of L. intracellularis or related microorganism is a dead preparation of the microorganism.

A vaccine composition according to claim 2, wherein the non-pathogenic form of L. intracellularis or related microorganism is a dead preparation in formalin of the microorganism.

6. A vaccine composition according to claim 1 or 2, wherein said composition comprises a molecule of peptide, polypeptide, protein, carbohydrate, lipid or nucleic acid or a combination thereof of L. intracellularis or related microorganism in a effective amount to induce a protective immune response agent of L. intracellularis or related microorganism.

7. A vaccine composition according to claim 6, wherein the composition comprises a peptide, polypeptide, protein or a derivative thereof of L. intracellularis or related microorganism.

8. A vaccine composition according to claim 7, wherein the peptide, polypeptide or protein is in recombinant form.

9. A vaccine composition according to claim 7 or 8, wherein the composition comprises a redoubled protein / heat shock, a basal body protein flagellar, S-adenosylmethionine: tRNA ribosyltransferase-isomerase, autolysin, enoyl- (acyl-carrier-protein) reductase or a glucarate transporter or derivative thereof.

10. A vaccine according to claim 9, wherein the protein is GroEL having an amino acid sequence set forth in SEQ ID NO: 2 or is a protein having at least approximately similarity thereto. eleven .

A vaccine according to claim 9, wherein the protein is GroES having an amino acid sequence set forth in SEQ ID NO: 4 or is a protein having at least approximately similarity thereto.

A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 1 or a sequence having at least about 40% similarity to the same.

A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 3 or a sequence having at least about 40% similarity to the same.

A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 5 or a sequence having at least about 40% similarity to the same.

A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 6 or a sequence having at least about 40% similarity to the same.

16. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 8 or a sequence having at least about 40% similarity thereto.

17. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 1 1 or a sequence having at least about 40% similarity with the same.

18. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 13 or a sequence having at least about 40% similarity to the same.

19. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 15 or a sequence having at least about 40% similarity to the same.

20. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 17 or a sequence having at least about 40% similarity to the same.

21. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 18 or a sequence having at least about 40% similarity to the same.

22. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 19 or a sequence having at least about 40% similarity to the same.

23. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 20 or a sequence having at least about 40% similarity to the same.

24. A vaccine composition according to claim 8. wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 21 or a sequence having at least about 40% similarity to the same.

25. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 22 or a sequence having at least about 40% similarity to the same.

26. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 7 or a sequence having at least 40% similarity.

27. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 9 or a sequence having at least 40% similarity.

28. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 10 or a sequence having at least 40% similarity.

29. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 12 or a sequence having at least 40% similarity.

30. A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 14 or a sequence having at least 40% similarity.

31 A vaccine composition according to claim 8, wherein the composition comprises a peptide, polypeptide or protein having an amino acid sequence of SEQ ID NO: 16 or a sequence having at least 40% similarity.

32. A method for vaccinating an animal or bird against infection with L. intracellularis or related microorganism or treating an animal or bird infected with L. intracellularis, said method comprising administering to said animal or bird an effective amount of a non-pathogenic form of L. intracellularis or related microorganism or an immunogenic component thereof for a time and under conditions sufficient to induce a protective immune response against L. intracellularis or related microorganism.

33. A method according to claim 32, wherein the animal is a pig.

34. A method according to claim 33, wherein the non-pathogenic form of L. intracellularis or related microorganism is an attenuated species of the microorganism.

35. A method according to claim 33, wherein the non-pathogenic form of L. intracellularis or related microorganism is a dead preparation of the microorganism.

36. A method according to claim 33, wherein the non-pathogenic form of L. intracellularis or related microorganism is a dead preparation in formalin of the microorganism.

37. A method according to claim 32 and 33, wherein said immunogenic component comprises a molecule of peptide, polypeptide, protein, carbohydrate, lipid or nucleic acid or a combination thereof of L. intracellularis or related microorganism in an amount effective to induce a protective immune response against L. intracellularis or related microorganism.

38. A method according to claim 37, wherein said component comprises a peptide, polypeptide, protein or a derivative thereof of L. intracellularis or related microorganism.

39. A method according to claim 38, wherein the peptide, polypeptide or protein is in recombinant form.

40. A method according to claim 29 or 30, wherein the immunogenic component is a redoubled protein / heat shock, a basal body protein flagellar, S-adenosylmethionine: tRNA ribosyltransferase-isomerase, autolysin, enoyl- ( acyl-carrier-protein) reductase or a glucarate transporter or derivative thereof.

41. A method according to claim 40, wherein the protein is GroEL having an amino acid sequence set forth in SE ID NO: 2 or is a protein having at least about 40% similarity thereto.

42. A method according to claim 40, wherein the protein is GroES having an amino acid sequence set forth in SEQ ID NO: 4 or is a protein having at least about 40% similarity thereto.

43. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 1 or a sequence having at least about 40% similarity thereto. .

44. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 3 or a sequence having at least about 40% similarity thereto.

45. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 5 or a sequence having at least about 40% similarity thereto. .

46. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 6 or a sequence having at least about 40% similarity thereto.

47. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising 3EQ ID NO: 8 or a sequence having at least about 40% similarity thereto. .

48. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 1 1 or a sequence having at least about 40% similarity to she.

49. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 13 or a sequence having at least about 40% similarity thereto. .

50. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 15 or a sequence having at least about 40% similarity thereto. .

51. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 17 or a sequence having at least about 40% similarity thereto.

52. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleic acid sequence comprising SEQ ID NO: 18 or a sequence having at least about 40% similarity thereto. .

53. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 19 or a sequence having at least about 40% similarity thereto. .

54. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 20 or a sequence having at least about 40% similarity thereto. .

55. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 21 or a sequence having at least about 40% similarity thereto. .

56. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein encoded by a nucleotide sequence comprising SEQ ID NO: 22 or a sequence having at least about 40% similarity thereto.

57. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 7 or having at least 40% similarity thereto.

58. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 9 or having at least 40% similarity thereto.

59. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 10 or having at least 40% similarity thereto.

60. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 12 or having at least 40% similarity thereto.

61. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 14 or having at least 40% similarity thereto.

62. A method according to claim 38, wherein the immunogenic component comprises a peptide, polypeptide or protein comprising an amino acid sequence set forth in SEQ ID NO: 16 or having at least 40% similarity thereto.

63. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 1 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 1, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

64. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 3 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 3, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

65. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 5 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 5, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

66. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 6 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 6, and which is capable of hybridizing thereto under conditions of low stringency and which encodes a peptide, polypeptide or immunogenic protein of L. intracellularis or related microorganism.

67. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 8 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 8, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

68. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 11 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 11, and which is able to hybridize thereto under conditions of low stringency and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

69. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 13 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 13, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

70. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 15 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 15, and which is able to hybridize to it under conditions of low severity and which encodes a peptide, polypeptide or immunogenic protein of L. intracellularis or related microorganism.

71 An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 17 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 17, and which is able to hybridize to it under conditions of low severity and which encodes a peptide, immunogenic protein or polypeptide of L. intracellularis or related microorganism.

72. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 18 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 18, and which is able to hybridize thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

73. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 19 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 19, and which is capable of hybridizing thereto under conditions of low stringency and which encodes a peptide, polypeptide or immunogenic protein of L. intracellularis or related microorganism.

74. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 20 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 20, and which is capable of hybridizing thereto under conditions of low stringency and which encodes a peptide, polypeptide or immunogenic protein of L. intracellularis or related microorganism.

75. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 21 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 21, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

76. An isolated nucleic acid molecule comprising a sequence as set forth in SEQ ID NO: 22 or a nucleotide sequence having at least 40% similarity to the nucleotide sequence set forth in SEQ ID NO: 22, and which is capable of hybridizing thereto under conditions of low severity and which encodes an immunogenic peptide, polypeptide or protein of L. intracellularis or related microorganism.

77. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 1, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 1 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

78. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 3, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 3 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

79. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 5, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 5 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

80. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 6, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 6 under conditions of low severity , said expression DNA sequence being capable in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

81. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 8, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 8 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

82. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 1 1, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 1 1 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

83. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 13, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 13 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

84. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 15, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 15 under conditions of low stringency , said expression DNA sequence being capable in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

85. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 17, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 17 under conditions of low severity , said expression DNA sequence being capable in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

86. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 18, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 18 under conditions of low stringency , said expression DNA sequence being capable in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

87. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 19, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 19 under conditions of low stringency , said expression DNA sequence being capable in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

88. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 20, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 20 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

89. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 21, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 21 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.

90. A genetic vaccine comprising a DNA sequence having a nucleotide sequence set forth in SEQ ID NO: 22, or having at least 40% similarity to it or capable of hybridizing to SEQ ID NO: 22 under conditions of low severity, said DNA sequence being capable of expression in a pig to produce an amount of a peptide, polypeptide or protein effective to induce a protective immune response against L. intracellularis or related microorganism.