Lawsonia intracellularis proteins, and related methods and materials

Abstract

Isolated polynucleotide molecules contain a nucleotide sequence that encodes a L. intracellularis HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein, a substantial portion of the sequences, or a homologous sequence. Related polypeptides, immunogenic compositions and assays are described.

What is claimed is:  
     
         1 . An isolated polynucleotide molecule comprising a nucleotide sequence that is selected from the group consisting of 
 a) a nucleotide sequence encoding  L. intracellularis  HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein;    b) a nucleotide sequence that is a substantial part of said nucleotide sequence encoding said  L. intracellularis  HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein; and    c) a nucleotide sequence that is homologous to the nucleotide sequence of a) or b).    
     
     
         2 . The isolated polynucleotide molecule of  claim 1  comprising said nucleotide sequence encoding said  L. intracellularis  HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein.  
     
     
         3 . The isolated polynucleotide molecule of  claim 1  comprising a nucleotide sequence consisting of the open reading frame selected from the group consisting of the open reading frame of SEQ ID NO: 1 from about nt 165 to about nt 1745, the open reading frame of SEQ ID NO: 1 from about nt 3031 to about nt 3738, the open reading frame of SEQ ID NO: 1 from about nt 3695 to about nt 6385, the open reading frame of SEQ ID NO: 2 from about nt 252 to about nt 2690. the open reading frame of SEQ ID NO: 2 from about nt 2891 to about nt 4315, and the open reading frame of SEQ ID NO: 2 from about nt 4581 to about nt 4829.  
     
     
         4 . A polynucleotide molecule comprising a nucleotide sequence of greater than 20 nucleotides having promoter activity and found within SEQ ID NO: 2 from about nt 2691 to about nt 2890, or its complement.  
     
     
         5 . A recombinant vector comprising the polynucleotide molecule of  claim 1 .  
     
     
         6 . The recombinant vector of  claim 5  consisting of a plasmid selected from the group consisting of plasmid pER432 containing the ponA gene (ATCC accession number PTA-635), plasmid pER434 containing the htrA gene (ATCC accession number PTA-636), plasmid pER436 containing the hypC gene (ATCC accession number PTA-637), plasmid pT068 containing the lysS and ycfW genes (ATCC accession number PTA-2232), plasmid pER438 containing the ycfW and abc1 genes (ATCC accession number PTA-638), and plasmid pER440 containing the Omp100 gene (ATCC accession number PTA-639).  
     
     
         7 . A transformed host cell comprising the recombinant vector of  claim 5 .  
     
     
         8 . A polypeptide produced by the transformed host cell of  claim 7 .  
     
     
         9 . A genetic construct comprising a polynucleotide molecule that can be used to alter a Lawsonia gene, comprising: (a) a polynucleotide molecule comprising a nucleotide sequence that is otherwise the same as a nucleotide sequence of a htrA, ponA, hypC, lysS, ycfW, abc1, or omp100 gene, or that is otherwise the same as a nucleotide sequence that is homologous thereto, or a substantial portion of said nucleotide sequence, but which nucleotide sequence further comprises one or more mutations capable of altering the htrA, ponA, hypC, lysS, ycfW, abc1, or omp100 gene; or (b) a polynucleotide molecule comprising a nucleotide sequence that naturally flanks in situ the ORF of said htrA, ponA, hypC, lysS, ycfW, abc1, or omp100 gene, or a nucleotide sequence that is homologous to said flanking sequence; such that transformation of a Lawsonia cell with the genetic construct of (a) or (b) results in altering the htrA, ponA, hypC, lysS, ycfW, abc1, or omp100 gene.  
     
     
         10 . A transformed cell comprising the genetic construct of  claim 9 .  
     
     
         11 . An isolated polypeptide, said polypeptide being selected from the group consisting of: 
 (a)  L. intracellularis  HtrA, PonA, HypC, YcfW, ABC1, or Omp100 protein;    (b) a polypeptide having an amino acid sequence that is homologous to that of said  L. intracellularis  HtrA, PonA, HypC, YcfW, ABC1, or Omp100 protein;    (c) a polypeptide consisting of a substantial portion of said  L. intracellularis  HtrA, PonA, HypC, YcfW, ABC1, or Omp100 protein or of said polypeptide having an amino acid sequence that is homologous to that of said  L. intracellularis  HtrA, PonA, HypC, YcfW, ABC1, or Omp100 protein;    (d) a fusion protein comprising the protein or polypeptide of (a), (b) or (c) fused to another protein or polypeptide; and    (e) an analog or derivative of the protein or polypeptide of (a), (b), (c) or (d).    
     
     
         12 . The isolated polypeptide of  claim 11  wherein said  L. intracellularis  protein has an amino acid sequence that comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 102.  
     
     
         13 . The isolated polypeptide of  claim 11  comprising a polypeptide that consists of said  L. intracellularis  HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein having between 1 and 10 amino acids inserted, deleted, or substituted, including combinations thereof.  
     
     
         14 . A substantially pure polypeptide comprising an epitope of HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein that is specifically reactive with anti-Lawsonia antibodies.  
     
     
         15 . An isolated polypeptide comprising an amino acid sequence encoded by the polynucleotide molecule of  claim 1 .  
     
     
         16 . An isolated antibody that specifically reacts with  L. intracellularis  HtrA, PonA, HypC, LysS, YcfW, ABC1, or Omp100 protein.  
     
     
         17 . A live attenuated vaccine comprising the transformed cell of  claim 10 .  
     
     
         18 . A killed cell vaccine comprising transformed cells of  claim 10  in killed form.  
     
     
         19 . An immunogenic composition comprising an immunogenically effective amount of the polypeptide of  claim 11  in combination with a pharmaceutically acceptable carrier.  
     
     
         20 . An immunogenic composition comprising an immunogenically effective amount of a polynucleotide molecule comprising a nucleotide sequence encoding the polypeptide of  claim 11  in combination with a pharmaceutically acceptable carrier.