Method of controlling fungal pathogens, and agents useful for same

Abstract

A method of preventing or treating infection by a range of fungal pathogens in an animal or plant or stored plant product comprising administering an effective amount of a sugar acid, in particular mannonic acid, gluconic acid or galacturonic. The sugar acid can be delivered by a biocontrol that can manufacture the sugar acid. An exemplified biocontrol is Pseudomonas strain AN5, which is characterized by the capacity to convert aldose to sugar acid. Also disclosed are nucleotide sequences from enzymes in the Pseudomonas PQQ enzyme pathway which may be used to transform microorganisms and plants so that they produce sugar acids at levels sufficient to control the growth or viability of pathogens.

1 - 47 . (canceled)  
     
     
         48 . An isolated nucleic acid molecule which comprises a nucleotide sequence which encodes one or more enzymes involved in the biosynthesis of a sugar acid, wherein said nucleotide sequence is selected from the group consisting of: 
 (i) a nucleotide sequence which is at least about 50% identical to at leave about 30 contiguous nucleotides of SEQ ID NO:1 or a complementary sequence thereto;    (ii) a nucleotide sequence which is at least about 50% identical to at least about 30 contiguous nucleotides of the  Pseudomonas  gene sequence contained in the cosmid clone pMN M53 (AGAL Accession No. NM 00/09622);    (iii) a nucleotide sequence which is capable of hybridizing under at least low stringency conditions to at least about 30 contiguous nucleotides of SEQ ID NO:1 or a complementary sequence thereto;    (iv) a nucleotide sequence which is capable of hybridizing under at least low stringency conditions to at least about 30 contiguous nucleotides of the  Pseudomonas  gene sequence contained in the cosmid clone pMN M53 (AGAL Accession No. NM 00/09622); and    (v) a nucleotide sequence which is generate to SEQ ID NO:1 or the  Pseudomonas  gene sequence contained in the cosmid clone pMN M53 (AGAL Accession No. 00/09622).    
     
     
         49 . The isolated nucleic acid molecule of  claim 48  wherein the nucleotide sequence encodes a sugar oxidase enzyme.  
     
     
         50 . The isolated nucleic acid molecule of  claim 49  wherein the sugar oxidase enzyme is a PQQ-dependent sugar oxidase enzyme.  
     
     
         51 . The isolated nucleic acid molecule according to  claim 48  comprising the nucleotide sequence of SEQ ID NO:1.  
     
     
         52 . The isolated nucleic acid molecule according to  claim 48  comprising the nucleotide sequence of the  Pseudomonas  gene sequence contained in the cosmid clone pMN M53 (AGAL Accession No. NM 00/09622).  
     
     
         53 . An isolated nucleic acid molecule comprising a nucleotide sequence encoding one or more enzymes involved in the biosynthesis of PQQ, wherein said nucleotide sequence is selected from the group consisting of: 
 (i) a nucleotide sequence having at least about 50 contiguous nucleotides of any one of SEQ ID NOs:2 to 6 or a complementary sequence thereto;    (ii) a nucleotide sequence having at least about 50 contiguous nucleotides of the  Pseudomonas  gene sequence contained in the cosmid clone pMN L2 (AGAL Accession No. NM 00/09621); and    (iii) a nucleotide sequence that is degenerate to any one of SEQ ID NOs: 2 to 6 or the  Pseudomonas  gene sequence contained in the cosmid clone pMN L2 (AGAL Accession No. NM 00/09621).    
     
     
         54 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of SEQ ID NO:2.  
     
     
         55 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of SEQ ID NO:3.  
     
     
         56 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of SEQ ID NO:4.  
     
     
         57 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of SEQ ID NO:5.  
     
     
         58 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of SEQ ID NO:6.  
     
     
         59 . The isolated nucleic acid molecule according to  claim 53  comprising the nucleotide sequence of the  Pseudomonas  gene sequence contained in the cosmid clone pMN L2 (AGAL Accession No. NM 00/09621).  
     
     
         60 . A method of producing a sugar acid comprising expressing the isolated nucleic acid molecule according to  claim 48  in a cell, tissue, or organism and culturing said cell, tissue or organism in the presence of an aldose substrate for a time and under conditions sufficient to produce a sugar acid.  
     
     
         61 . The method according to  claim 60  further comprising introducing the nucleic acid molecule to the cell, tissue or organ in an expressible format.  
     
     
         62 . The method according to  claim 60  further comprising extracting or purifying the sugar acid produced.  
     
     
         63 . The method according to  claim 60  wherein the cell is a bacterial cell.  
     
     
         64 . The method according to  claim 63  wherein the bacterial cell is a  Pseudomonas  sp.  
     
     
         65 . The method according to  claim 60  wherein the cell, tissue or organ is a plant cell, tissue, or organ.  
     
     
         66 . The method according to  claim 60  further comprising expressing the nucleic acid molecule according to  claim 53  for a time and under conditions sufficient to produce PQQ in the cell, tissue or organ.  
     
     
         67 . A method of enhancing the tolerance of a plant to infection by a fungal pathogen comprising expressing therein the isolated nucleic acid molecule according to  claim 48 , and optionally a second isolated nucleic acid molecule encoding one or more PQQ-biosynthesis enzymes, for a time and under conditions sufficient for a sugar acid to be produced by said plant, or by a cell, tissue or organ of said plant.  
     
     
         68 . The method according to  claim 66  wherein the second isolated nucleic acid molecule is the nucleic acid molecule according to  claim 53 .  
     
     
         69 . A transformed plant comprising the isolated nucleic acid molecule according to  claim 48 .  
     
     
         70 . A progeny plant, cell, tissue or organ of the plant according to  claim 69 , wherein said progeny, cell, tissue or organ comprises the isolated nucleic acid molecule according to  claim 48 .  
     
     
         71 . A transformed plant comprising the isolated nucleic acid molecule of  claim 53 .  
     
     
         72 . A progeny plant, cell, tissue or organ of the plant according to  claim 69 , wherein said progeny, cell, tissue or organ comprises the isolated nucleic acid molecule, wherein the nucleic acid molecule comprises a nucleotide sequence encoding one or more enzymes involved in the biosynthesis of PQQ, wherein said nucleotide sequence is selected from a group consisting of: 
 (i) a nucleotide sequence having at least about 50 contiguous nucleotides of any one of SEQ ID NOs:2 to 6 or a complementary sequence thereto;    (ii) a nucleotide sequence having at least about 50 contiguous nucleotides of the  Pseudomonas  gene sequence contained in the cosmid clone pMN-L2 (AGAL Accession No. NM 00/09621); and    (iii) a nucleotide sequence that is degenerate to any one of SEQ ID NOs:2 to 6 or the  Pseudomonas  gene sequence contained in the cosmid clone pMN-L2 (AGAL Accession No. NM 00/09621).