Over the past two decades, Lactococcus lactis has vastly extended its application from food to being a successful microbial cell factory.Not only can it produce lactic acid, vitamins, bacteriocins and antimicrobial peptide (nisin) in traditional fermentation, it can also express heterologous antigens, thus allowing the bacteria to act as live vaccines. Using L. lactis as vaccine carriers is appealing as it is able to induce both mucosal and systemic immune responses, has adjuvant properties, and is free from risks associated with the use of conventional attenuated live pathogens. With lyophilization, L. lactis does not require storage at low temperature, and administration of the preparation does not require specialized personnel.
However, L. lactis suffers from various stress conditions including acid, osmotic and oxygen during fermentation processing, among which acid stress and cold stress are great survival challenges. During fermentation, the decrease of environmental pH will inhibit the activity of intracellular enzymes, which will also affect the normal growth and metabolism of microbial cells. Furthermore, as live vaccines, L.lactis has an advantage in the low pH condition of gastric juice compared with other chassis microorganism. During the process of lyophilization, it suffers from severe cold stress, which rapidly reduce their survival rate. Therefore, the strain resistant to acid and cold needs to be constructed as soon as possible.
Therefore, we aim to create an ideal chassis microorganism with good resistance and food-safe characteristics by synthetic biology methods. We added genes msmk, cspD2, promoter ppepN, gfp and a kill-switch (→Basic parts) respectively to model strain Lactococcus lactis NZ3900 to give it the anti-acid, anti-cold characteristics.