In Vitro Evaluation of Probiotic Properties of Two Novel Probiotic Mixtures, Consti-Biome and Sensi-Biome.

Changes in the gut microbiome cause recolonization by pathogens and inflammatory responses, leading to the development of intestinal disorders. Probiotics administration has been proposed for many years to reverse the intestinal dysbiosis and to enhance intestinal health. This study aimed to evaluate the inhibitory effects of two newly designed probiotic mixtures, Consti-Biome and Sensi-Biome, on two enteric pathogens Staphylococcus aureus and Escherichia coli that may cause intestinal disorders. Additionally, the study was designed to evaluate whether Consti-Biome and Sensi-Biome could modulate the immune response, produce short-chain fatty acids (SCFAs), and reduce gas production. Consti-Biome and Sensi-Biome showed superior adhesion ratios to HT-29 cells and competitively suppressed pathogen adhesion. Moreover, the probiotic mixtures decreased the levels of pro-inflammatory cytokines, such as tumor necrosis factor-α, interleukin (IL)-6 and IL-1ß. Cell-free supernatants (CFSs) were used to investigate the inhibitory effects of metabolites on growth and biofilms of pathogens. Consti-Biome and Sensi-Biome CFSs exhibited antimicrobial and anti-biofilm activity, where microscopic analysis confirmed an increase in the number of dead cells and the structural disruption of pathogens. Gas chromatographic analysis of the CFSs revealed their ability to produce SCFAs, including acetic, propionic, and butyric acid. SCFA secretion by probiotics may demonstrate their potential activities against pathogens and gut inflammation. In terms of intestinal symptoms regarding abdominal bloating and discomfort, Consti-Biome and Sensi-Biome also inhibited gas production. Thus, these two probiotic mixtures have great potential to be developed as dietary supplements to alleviate the intestinal disorders.

Complete Genomic Analysis of Enterococcus faecium Heat-Resistant Strain Developed by Two-Step Adaptation Laboratory Evolution Method.

Stress resistance is an important trait expected of lactic acid bacteria used in food manufacturing. Among the various sources of stress, high temperature is a key factor that interrupts bacterial growth. In this regards, constant efforts are made for the development of heat-resistant strains, but few studies were done accompanying genomic analysis to identify the causal factors of the resistance mechanisms. Furthermore, it is also thought that tolerance to multiple stresses are equally important. Herein, we isolated one Enterococcus faecium strain named BIOPOP-3 and completed a full-length genome sequence. Using this strain, a two-step adaptive laboratory evolution (ALE) method was applied to obtain a heat-resistant strain, BIOPOP-3 ALE. After sequencing the whole genome, we compared the two full-length sequences and identified one non-synonymous variant and four indel variants that could potentially confer heat resistance, which were technically validated by resequencing. We experimentally verified that the evolved strain was significantly enhanced in not only heat resistance but also acid and bile resistance. We demonstrated that the developed heat-resistant strain can be applied in animal feed manufacturing processes. The multi-stress-resistant BIOPOP-3 ALE strain developed in this study and the two-step ALE method are expected to be widely applied in industrial and academic fields. In addition, we expect that the identified variants which occurred specifically in heat-resistant strain will enhance molecular biological understanding and be broadly applied to the biological engineering field.

Changes in Cell Membrane Fatty Acid Composition of Streptococcus thermophilus in Response to Gradually Increasing Heat Temperature.

In this study, a method of heat adaptation was implemented in an attempt to increase the upper thermal threshold of two Streptococcus thermophilus found in South Korea and identified the alterations in membrane fatty acid composition to adaptive response to heat. In order to develop heat tolerant lactic acid bacteria, heat treatment was continuously applied to bacteria by increasing temperature from 60°C until the point that no surviving cell was detected. Our results indicated significant increase in heat tolerance of heat-adapted strains compared to the wild type (WT) strains. In particular, the survival ratio of basically low heat-tolerant strain increased even more. In addition, the strains with improved heat tolerance acquired cross protection, which improved their survival ratio in acid, bile salts and osmotic conditions. A relation between heat tolerance and membrane fatty acid composition was identified. As a result of heat adaptation, the ratio of unsaturated to saturated fatty acids (UFA/SFA) and C18:1 relative concentration were decreased. C6:0 in only heatadapted strains and C22:0 in only the naturally high heat tolerant strain were detected. These results support the hypothesis, that the consequent increase of SFA ratio is a cellular response to environmental stresses such as high temperatures, and it is able to protect the cells from acid, bile salts and osmotic conditions via cross protection. This study demonstrated that the increase in heat tolerance can be utilized as a mean to improve bacterial tolerance against various environmental stresses.