Health promotion and prevention of disorders through daily diet is an ideal approach. The use of probiotics, whose safety is well-established, has been increasing annually for this purpose. Much research about probiotic properties has been performed, however, it is pointed out that even the effectiveness of the same probiotic strain varies depending on the individual host because of the environmental background and gut microbiota. In this context, developmental research on probiotics based on designing and/or breeding technology is gaining attention. In this review, we provide an application overview of genetic engineering and ribosome engineering to lactic acid bacteria as a developmental study to propose probiotics with robust functionality.

Lactic acid bacteria (LAB) are usually cultivated in anaerobic batch culture. In this environment, since LAB produce significant amounts of lactate, it is difficult to cultivate them at high cell-densities and the cell yield is low. LAB produce lactate to regenerate NAD+ consumed in the glycolytic system by lactate dehydrogenase (LDH). Therefore, lactate production can be suppressed if NAD+ regeneration systems other than LDH are available or if ATP can be obtained in systems that do not consume NAD+. However, these alternative pathways, in addition to not being able to fully complement the consumption of NAD+ in the glycolytic system, are mostly repressed by glucose. To suppress lactate production, therefore, it is important to derepress the alternative pathways by gradually feeding sugar that match the capacity of the alternative pathways. Among the alternative pathways, the pathway that regenerates NAD+ using oxygen as an electron acceptor does not waste carbon and is expected to improve cell yield. It will be possible to cultivate LAB in high yield and high cell-density by fed-batch culture under aerobic conditions with a support for a formation of a respiratory chain by adding heme and menaquinone.

MALDI-MS has begun to spread to various fields, mainly clinical microbiology tests. In this article, we introduce the characteristics of MALDI-MS, the knack of performing microbial analysis by MALDI-MS, and the subspecies-level classification of Lactococcus lactis using ribosomal proteins as biomarkers as an example of MALDI-MS proteotyping.