Characterization of respiratory physiology in Lactococcus lactis for high-yield production of high-performance starter cultures

Abstract: Fermented food products are consumed world-wide, every day, and the demand is growing as intake increases. Lactococcus lactis is a lactic acid bacterium commonly used as a starter culture to produce fermented dairy products. The quality of starter cultures is linked to the conditions during its production process and affects the culture activity during its use in dairy product fermentation. Therefore, industrial manufacturers of starter cultures strive to not only optimize biomass yield, but also the cell robustness. L. lactis starter cultures are produced by anaerobic fermentation or by aerobic respiration when haemin is available in the cultivation medium. The aim of this work was to investigate the response of L. lactis to respiration-permissive conditions, to enable redesign and optimization of the batch production process. The process parameters were carefully chosen to mimic the industrial process. The effects of the specific growth rate on respiratory metabolism, energetics and cell quality were quantified using chemostat cultivations. Compared to anaerobic metabolism, the respiratory metabolism of L. lactis was remarkably flexible, and could be modulated by controlling the dilution rate. The lowest dilution rate supported full respiratory metabolism, whereas the higher dilution rates caused a shift towards respiro-fermentative metabolism. The inoculation procedures were investigated in detail to gain an understanding of the occurrence of lag phases after inoculation. It was found that the length of the lag phase in subsequent main cultures was related to galactose excretion in lactose-grown pre-cultures. Furthermore, based on lacS gene expression measurements in lactose-grown cultures, it is suggested that LacS is responsible for the galactose excretion as a galactose-lactose antiporter. The quality of frozen and freeze-dried products was investigated and sensitivity to freeze-drying was found to be associated with the physiological state of the cells during cultivation. Cells harvested under respiration-permissive conditions in batch and chemostat cultivations at low dilution rate were less robust to freeze-drying, whereas higher dilution rates led to robust cells performing equally well or better than anaerobic cells. The findings of this work underline the importance of systematically studying the combination of upstream and downstream aspects of production processes. The results indicate that by controlling the specific growth rate and the haemin concentration during the aerobic growth of L. lactis , not only higher biomass yields, but also better cell robustness, can be achieved.