Method to improve
Lactococcus lactis strains with improved preservation characteristics, and improved acid and bile salt tolerance are disclosed. More specifically, a L. lactis strain having a heterologous trehalose-6-phosphate synthase gene and/or a trehalose-6-phosphate phosphatase gene, results in an accumulation of trehalose in the cytoplasm and/or in the cytoplasmic membrane. A L. lactis strain, having an internal trehalose concentration of at least 10 mg per gram cells (w/w) is disclosed.
1. An isolated Lactococcus lactis strain, that expresses a heterologous trehalose-6-phosphate phosphatase gene, wherein the heterologous trehalose-6-phosphate phosphatase gene is the OtsB gene from Escherichia coli , and wherein trehalose-6-phosphate phosphorylase activity is inactivated in the Lactococcus lactis.
2. The isolated Lactococcus lactis strain according to claim 1 , wherein the gene is operably linked to the inducible L. lactis nisin promoter.
3. The isolated Lactococcus lactis strain according to claim 1 , wherein the gene is functionally linked to the constitutive P1 promoter.
4. The isolated Lactococcus lactis strain according to claim 1 , wherein the strain further comprises a heterologous trehalose-6-phosphate synthase gene, wherein the heterologous trehalose-6-phosphate synthase gene is the OtsA gene from Escherichia coli.
5. The isolated Lactococcus lactis strain according to claim 4 wherein one or both genes are operably linked to the inducible L. lactis nisin promoter.
6. The isolated Lactococcus lactis strain according to claim 4 wherein one or both genes are functionally linked to the constitutive P1 promoter.
7. A composition that expresses:
i) an isolated Lactococcus lactis strain, comprising a heterologous trehalose-6-phosphate phosphatase gene, wherein the heterologous trehalose-6-phosphate phosphatase gene is the OtsB gene from Escherichia coli , and wherein trehalose-6-phosphate phosphorylase activity is inactivated in the Lactococcus lactis , and
ii) a growth medium comprising trehalose and another carbon source.