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Leuconostoc genus, which comprise heterofermentative lactic acid bacteria, reduces fructose to mannitol by recycling intracellular NADH. To evaluate the mannitol productivities of different Leuconostoc species, 5 stock cultures and 4 newly isolated strains were cultivated in MRS and simplified media containing glucose and fructose (1:2 ratio). Among them, L. citreum KACC 91348P, which was isolated from kimchi, showed superior result in cell growth rate, mannitol production rate, and yield in both media. The optimal condition for mannitol production of this strain was pH 6.5 and 30 o C. When L. citreum KACC was cultured in simplified medium in a 2 l batch fermenter under optimal conditions, the maximum volumetric productivity was 14.83 g•l-1 h-1 and overall yield was 86.6%. This strain is a novel and efficient mannitol producer originated from foods to be used for fermentation of fructose-containing foods.

In 2004, Leuconostoc mesenteroides DRC was first used as a starter culture for achieving higher organoleptic effects in Korean kimchi manufacture. For a better understanding of starter growth in a mixed culture system, and for predicting starter predominance in kimchi, a monitoring system for the starter was established. The chloramphenicol resistance marker gene (cat) was randomly integrated into chromosomal DNA of L. mesenteroides DRC using a viral transposon and transposase. The DRC mutant, tDRC2, had a similar growth pattern to the host strain, with no major alteration in phenotypic characteristics. The mutant strain was inoculated into real kimchi, and monitoring of the starter population was successfully achieved. The overall predominance of Leuconostoc in kimchi inoculated with DRC followed the general growth pattern of this genus during kimchi fermentation. Our results also demonstrate the competitive ability of the DRC starter against Leuconostoc from natural flora, maintaining its predominance above 88% during the whole fermentation period. Based on this experiment, the random gene integration method using a transposon was shown to be of utility in transferring any commercial starter into a selectable and monitorable strain for simulation purposes.

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Leuconostoc mesenteroides is a commercially important lactic acid bacterium currently used as a starter for kimchi and kefir. However, its sensitivity to acid stress limits its performance. L. mesenteroides was grown in a medium supplemented with 3.2 or 6.4 mM glutathione (GSH), and cell survival rates were measured during a long-term mild acid challenge (pH 4.0). As a result, GSH was imported by the cells and protected against acid stress; thereafter it was consumed as a nutrient. Acid stress resistance of starter cultures of this bacterium can thus be improved by cultivating it in media supplemented with GSH.

To develop a gene expression system for Leuconostoc genus, construction of expression vector and expression of a heterologus protein in Leuconostoc was performed. Alpha-amylase gene from Lactobacillus amylovorus was cloned into a Leuconostoc cloning vector, pLeuCM, with its own signal peptide. pLeuCMamy was introduced into Leuconostoc citreum CB2567 and a successful expression of alpha-amy gene was confirmed by enzyme activity assays. About 90% of alpha-amylase activity was detected in the culture broth, revealing most of expressed alpha-amylase was secreted out cells. The signal sequence of alpha-amy gene is a good candidate for the secretion of heterologous protein by using Leuconostoc host-vector system.