Development of Bacillus subtilis mutants to produce tryptophan in pigs.
OBJECTIVE
To generate tryptophan-overproducing Bacillus subtilis strains for in situ use in pigs, to reduce the feed cost for farmers and nitrogen pollution.


RESULTS
A novel concept has been investigated-to generate B. subtilis strains able to produce tryptophan (Trp) in situ in pigs. Mutagenesis by UV was combined with selection on Trp and purine analogues in an iterative process. Two mutants from different wild types were obtained, mutant 1 (M1) produced 1 mg Trp/l and mutant 2 (M2) 14 mg Trp/l. Genome sequence analysis revealed that M1 had three single nuclear polymorphisms (SNPs) and M2 had two SNPs compared to the wild type strains. In both mutants SNPs were found in genes regulating tryptophan synthesis. Reverse transcription PCR confirmed up-regulation of the tryptophan synthesis genes in both mutants, the expression was up to 3 times higher in M2 than in M1.


CONCLUSIONS
Tryptophan-excreting B. subtilis strains were obtained with UV-mutagenesis and analogue selection and can be used in animal feed applications.
Chr. Hansen A/S, Bøge Allé 10-12, 2970, Hørsholm, Denmark. dkkbj@chr-hansen.com.
Karin
K
Bjerre
Chr. Hansen A/S, Bøge Allé 10-12, 2970, Hørsholm, Denmark.
Mette D
MD
Cantor
Department of Animal Science, Aarhus University, Foulum, 8830, Tjele, Denmark.
Jan V
JV
Nørgaard
Department of Animal Science, Aarhus University, Foulum, 8830, Tjele, Denmark.
Hanne D
HD
Poulsen
Department of Animal Science, Aarhus University, Foulum, 8830, Tjele, Denmark.
Karoline
K
Blaabjerg
Department of Animal Science, Aarhus University, Foulum, 8830, Tjele, Denmark.
Nuria
N
Canibe
Department of Animal Science, Aarhus University, Foulum, 8830, Tjele, Denmark.
Bent B
BB
Jensen
Chr. Hansen A/S, Bøge Allé 10-12, 2970, Hørsholm, Denmark.
Birgitte
B
Stuer-Lauridsen
Chr. Hansen A/S, Bøge Allé 10-12, 2970, Hørsholm, Denmark.
Bea
B
Nielsen
Chr. Hansen A/S, Bøge Allé 10-12, 2970, Hørsholm, Denmark.
Patrick M F
PM
Derkx
D000818:Animals; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D009876:Operon / Q000235:genetics; D013552:Swine; D014364:Tryptophan / Q000378:metabolism*; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D014364:Tryptophan
Bacillus subtilis; Tryptophan; UV-mutagenesis; mtrB; trpS