Lacticaseibacillus casei is a gram-positive lactic acid bacteria that can adapt to a wide range of environmental conditions. These bacteria are used as probiotics in fermented products and have beneficial effects on human health [1]. L. casei is an attractive research target as it can be used not only for food fermentation but also for various industrial applications such as the production of pharmaceuticals or biofuels [2]. Moreover, although the entire genome of L. casei BL23 has been sequenced [3], there is still not much known about the genes that determine environmental adaptation.
It is widely accepted that small non-coding RNA molecules (sRNAs) play one of the most important roles in the post-transcriptional regulation of gene expression by complementary binding to the target mRNA or protein itself. Changes induced by these molecules usually determine the amount or activity of the protein in the cell, which affects adaptation to the environment [4, 5]. With this in mind, we analysed sLCB2236-sRNA, which is conserved in all L. casei bacteria and is a potential regulatory molecule that could help bacteria adapt to environmental changes. Thus, the aim of this work was to determine the effects of sRNA deletion (sLCB2236-) on bacterial stress adaptation.
For this purpose, an L. casei ΔsLCB2236- deletant was constructed using CRISPR-Cas9D10A nickase and, with wild-type and a deletant with reconstituted sRNA, seeded on agarised MRS media supplemented with different stressors such as ethanol, NaCl, H2O2, EDTA, Penicillin G, lysozyme, and bile acid. Furthermore, the agarised MRS media with bacteria was stored at different temperatures. The results showed that overexpression of sRNA increases bacterial resistance to ethanol stress, although it does not affect bacterial growth under other stresses.
In conclusion, we suggest that sLCB2236- is important for the adaptation of L. casei to ethanol-induced stress. In addition, this means that sLCB2236- could be suitable for research to confer ethanol resistance on other beneficial microorganisms that do not have this ability.