Antifungal paenibacillus strains, fusaricidin-type compounds, and their use

Abstract

The present invention relates to novel isolated bacterial strains, which are members of the genus Paenibacillus , originally isolated from soil and showing antagonistic activity against a broad range of pathogens and being capable of producing antimicrobial metabolites. It was found that the strains Lu16774 and Lu17007 belong to a novel subspecies named Paenibacillus polymyxa ssp. plantarum while the strain Lu17015 belongs to a novel species which is proposed to be Paenibacillus epiphyticus . The present invention also relates to microbial pesticide compositions comprising at least one of such novel bacterial strains, whole culture broth or a cell-free extract or a fraction thereof or at least one metabolite thereof, and/or a mutant of at least one of said novel bacterial strains having all the identifying characteristics of the respective bacterial strain or whole culture broth, cell-free extract, fraction and/or metabolite of the mutant thereof showing antagonistic activity against plant pathogens. The present invention also relates to a method of controlling or suppressing plant pathogens or preventing plant pathogen infections by applying such composition. The present invention also relates to novel fusaricidin-type compounds which are metabolites produced by the strains of the present invention.

1 .- 19 . (canceled) 
     
     
         20 . A  Paenibacillus  strain, which is selected from the group consisting of:
 a) strain Lu16774 deposited with DSMZ under Accession No. DSM 26969;   b) strain Lu17007 deposited with DSMZ under Accession No. DSM 26970;   c) strain Lu17015 deposited with DSMZ under Accession No. DSM 26971; and   d) a strain which comprises a DNA sequence exhibiting
 d1) at least 99.6% nucleotide sequence identity to the DNA sequences SEQ ID NO:4 or SEQ ID NO:9; or 
 d2) at least 99.8% nucleotide sequence identity to the DNA sequence SEQ ID NO:14; or 
 d3) at least 99.9% nucleotide sequence identity to the DNA sequences SEQ ID NO:5 or SEQ ID NO:10; or 
 d4) at least 99.2% nucleotide sequence identity to the DNA sequence SEQ ID NO:15; or 
 d5) at least 99.2% nucleotide sequence identity to the DNA sequences SEQ ID NO:6 or SEQ ID NO:11; or 
 d6) at least 99.8% nucleotide sequence identity to the DNA sequence SEQ ID NO:16; or 
 d7) at least 99.8% nucleotide sequence identity to the DNA sequences SEQ ID NO:7 or SEQ ID NO:12; or 
 d8) at least 99.3% nucleotide sequence identity to the DNA sequence SEQ ID NO:17; or 
 d9) 100.0% nucleotide sequence identity to the DNA sequences SEQ ID NO:8 or SEQ ID NO:13; or 
 d10) 100% nucleotide sequence identity to the DNA sequence SEQ ID NO:18. 
   
     
     
         21 . The  Paenibacillus  strain according to  claim 20 , which is selected from the group consisting of:
 a) strain Lu16774 deposited with DSMZ under Accession No. DSM 26969;   b) strain Lu17007 deposited with DSMZ under Accession No. DSM 26970;   c) strain Lu17015 deposited with DSMZ under Accession No. DSM 26971; and   d) strains which comprise a DNA sequence exhibiting
 d1) at least 99.8% nucleotide sequence identity to any one of the DNA sequences SEQ ID NO:4, SEQ ID NO:9 and SEQ ID NO:14; or 
 d2) 100.0% nucleotide sequence identity to any one of the DNA sequences SEQ ID NO:5, SEQ ID NO:10 and SEQ ID NO:15; or 
 d3) exhibiting at least 99.8% nucleotide sequence identity to any one of the DNA sequences SEQ ID NO:6, SEQ ID NO:11 and SEQ ID NO:16; or 
 d4) at least 99.8% nucleotide sequence identity to any one of the DNA sequences SEQ ID NO:7, SEQ ID NO:12 and SEQ ID NO:17; or 
 d5) 100.0% nucleotide sequence identity to any one of the DNA sequences SEQ ID NO:8, SEQ ID NO:13 and SEQ ID NO:18. 
   
     
     
         22 . The  Paenibacillus  strain according to  claim 20 , wherein said  Paenibacillus  strain has antifungal activity against at least two of the plant pathogen selected from the group consisting of  Alternaria  spp.,  Botrytis cinerea, Phytophthora infestans  and  Sclerotinia sclerotiorum.    
     
     
         23 . The  Paenibacillus  strain according to  claim 20 , wherein said  Paenibacillus  strain produces at least one of the following compounds: 
       
         
           
           
               
               
           
         
       
       in a growth medium comprising at least one source of carbon and one source of nitrogen. 
     
     
         24 . The  Paenibacillus  strain according to  claim 23 , wherein said  Paenibacillus  strain is further capable of producing at least three compounds selected from the group consisting of fusaricidin A, fusaricidin B, fusaricidin C, fusaricidin D, LI-F06a, LI-F06b and LI-F08b. 
     
     
         25 . The  Paenibacillus  strain according to  claim 20 , wherein said  Paenibacillus  strain comprises at least one of the following compounds: 
       
         
           
           
               
               
           
         
       
     
     
         26 . The  Paenibacillus  strain according to  claim 25 , wherein said  Paenibacillus  strain further comprises at least three compounds selected from the group consisting of fusaricidin A, fusaricidin B, fusaricidin C, fusaricidin D, LI-F06a, LI-F06b and LI-F08b. 
     
     
         27 . A substantially purified culture of the  Paenibacillus  strain according to  claim 20 . 
     
     
         28 . A whole culture broth or a cell-free extract of the  Paenibacillus  strain according to  claim 20 . 
     
     
         29 . A compound of formula I 
       
         
           
           
               
               
           
         
         wherein 
         R is selected from 15-guanidino-3-hydroxypentadecanoic acid and 12-guanidinododecanoic acid; 
         X 1  is threonine; 
         X 2  is isoleucine; 
         X 3  is tyrosine; 
         X 4  is threonine; 
         X 5  is selected from glutamine and asparagine; 
         X 6  is alanine; and 
         wherein an arrow defines a single amide bond either between the carbonyl moiety of R and the amino group of the amino acid X 1  or between the carbonyl group of one amino acid and the amino group of a neighboring amino acid,
 wherein the tip of the arrow indicates the attachment to the amino group of 
 said amino acid X 1  or of said neighboring amino acid; and 
 
         wherein the single line without an arrow head defines a single ester bond between the carbonyl group of X 6  and the hydroxyl group of X 1 ; 
         or an agriculturally acceptable salt thereof. 
       
     
     
         30 . The compound according to  claim 29  selected from compounds 1A and 1B: 
       
         
           
           
               
               
           
         
         or an agriculturally acceptable salt thereof. 
       
     
     
         31 . A method of preparing a compound of formula I 
       
         
           
           
               
               
           
         
         wherein 
         R is selected from 15-guanidino-3-hydroxypentadecanoic acid and 12-guanidinododecanoic acid; 
         X 1  is threonine; 
         X 2  is isoleucine; 
         X 3  is tyrosine; 
         X 4  is threonine; 
         X 5  is selected from glutamine and asparagine; 
         X 6  is alanine; and 
         wherein an arrow defines a single amide bond either between the carbonyl moiety of R and the amino group of the amino acid X 1  or between the carbonyl group of one amino acid and the amino group of a neighboring amino acid,
 wherein the tip of the arrow indicates the attachment to the amino group of said amino acid X 1  or of said neighboring amino acid; and 
 
         wherein the single line without an arrow head defines a single ester bond between the carbonyl group of X 6  and the hydroxyl group of X 1 ; 
         or an agriculturally acceptable salt thereof, 
         which method comprises culturing the  Paenibacillus  strain of  claim 20  and recovering said compound or salt thereof from the whole culture broth. 
       
     
     
         32 . A composition comprising
 a) a  Paenibacillus  strain according to  claim 20 ; or   b) a substantially purified culture of a); or   c) a whole culture broth or a cell-free extract of a); or   d) a compound of formula I   
       
         
           
           
               
               
           
         
         wherein 
         R is selected from 15-guanidino-3-hydroxypentadecanoic acid and 12-guanidinododecanoic acid; 
         X 1  is threonine; 
         X 2  is isoleucine; 
         X 3  is tyrosine; 
         X 4  is threonine; 
         X 5  is selected from glutamine and asparagine; 
         X 6  is alanine; and 
         wherein an arrow defines a single amide bond either between the carbonyl moiety of R and the amino group of the amino acid X 1  or between the carbonyl group of one amino acid and the amino group of a neighboring amino acid,
 wherein the tip of the arrow indicates the attachment to the amino group of said amino acid X 1  or of said neighboring amino acid; and 
 
         wherein the single line without an arrow head defines a single ester bond between the carbonyl group of X 6  and the hydroxyl group of X 1 ; 
         or an agriculturally acceptable salt thereof; and 
         an auxiliary. 
       
     
     
         33 . The composition of  claim 32 , further comprising a pesticide. 
     
     
         34 . The composition of  claim 33 , wherein the pesticide is a further biopesticide. 
     
     
         35 . A plant propagation material having a coating comprising the composition according to  claim 32 . 
     
     
         36 . (canceled) 
     
     
         37 . A method of controlling, suppressing plant pathogens or preventing plant pathogen infection, wherein the plant pathogens, their habitat or plants to be protected against plant pathogen attack, or the soil or propagation material are treated with an effective amount of the composition according to  claim 32 . 
     
     
         38 . The method of  claim 37 , wherein the plant pathogens and/or harmful microorganisms are selected from harmful fungi.