Pre-adaptation of selected probiotic strains to multiple stress factors : consequent effect on their stability and probiotic properties

Abstract

The interest in the use of probiotics has escalated in the last decade, this is consequent to an increase in the number of studies showing that these microorganisms have beneficial effects on the host’s health. Probiotics can be administered in different ways, as capsules (pharmaceuticals) and also incorporated in different food products. In the definition of probiotics, it is highlighted that for them to be beneficial, they have to be administered alive and in adequate numbers. Nevertheless, there have been a number of problems associated with the number of viable cells of probiotics, with reports that viable numbers decline drastically on exposure to various stresses including those that prevail in the GIT. This raised an interest in the probiotics research, focusing on the techniques that can yield stress resistant or tolerant probiotics. These techniques are aimed to increase the number of the surviving cells after the exposure to technological and gastrointestinal stress factors. There has, therefore, been an increase in the studies focusing specifically on how to adapt the probiotic cells to different stress factors. The mechanism of pre- adaptation or cross- protection has been one of the most studied areas. With these mechanisms, researchers pre- expose probiotics to different stress factors so that they can survive better when they are later exposed to the same stress factor. Pre- adaptation of probiotics to multiple stress factors will therefore offer tolerance to more stress factors. Taking that into consideration, the present study aimed at determining whether probiotic cells that have been pre- exposed to multiple stress factors (acid, bile and temperature) will have better tolerance to different gastric and intestinal conditions when compared to non- adapted cells. The first part of the research followed a stepwise stress adaptation mechanism for six probiotics (Bifidobacterium bifidum LMG 11041, B. longum LMG 13197, B. longum Bb46, Lactobacillus acidophilus La14 150B, L. fermentum and L. plantarum). The results obtained show that the stability of the probiotic cells improves when the cells are further adapted to more stress factors. After the probiotics were exposed to stress factors, the tolerance of these probiotics towards acid and bile was investigated. These are the stress factors the probiotics encounter through their GIT following their consumption. The acid and bile tolerances of the stress exposed cells were higher than those of the cells that were not exposed to stress factors. After sequential exposure of the cells to the simulated gastric and intestinal conditions, viability of the three Lactobacilli cells and B. bifidum LMG 11041 were higher than their non- adapted counterparts. The bile salt hydrolase (BSH) activity and the antibiotic profiles of the probiotics remained unchanged. From these results it was evident that multi- stress pre- adaptation of probiotics increases the chances of survival for these probiotics in the gastrointestinal tract, without negatively affecting their antibiotic sensitivity profile or their ability to produce the enzyme BSH, which is one criterion used for selection of probiotics, specifically those with cholesterol lowering properties. The observed better survival of multi- stress pre- adapted cells when exposed to simulated gastrointestinal conditions raised an interest in another study that was used in the treatment of diseases using probiotics, the use of multiple probiotics. This part of the study aimed to determine first the survival of multiple cells when exposed to acid and bile, and then investigating their ability to inhibit growth of enteric pathogens, specifically Staphylococcus aureus and Escherichia coli, when used individually as single- or multiple- stress adapted cells, combinations of multi-stress adapted cells and comparing them to a combination of the non- adapted cells. A cocktail containing L. plantarum, L. fermentum and B. longum Bb46 and the one containing all the six adapted cells survived better in 2% bile and pH 2, respectively. Interestingly, for both the acid and bile tolerance studies, a cocktail containing all the six non- adapted cells was the least resistant. In the antipathogenic tests, a combination containing L. plantarum, B. longum Bb46 and B. longum 13197 inhibited S. aureus better and combination containing all the six stress adapted cells inhibited E. coli better. It was evident that although the stress adapted single cells inhibited both pathogens, there was an increase in the inhibition when the stress- adapted combinations were used. In all cases the combination containing all the six non- adapted cells was the least effective of all cocktails in inhibition of E. coli and S. aureus. The results of this study revealed that multi- stress pre- adapted probiotics survive better than the single stress adapted cells and above all, the use of non- adapted cells. This was even demonstrated in the use of combinations, where the stress adapted combinations had better results than the non- adapted combinations. This study is of importance to consumers, food industries, pharmaceutical and the probiotic industry as a whole. This study shows the increase in surviving cells after the exposure to stress factors. This information can be used in the production of different products. For an increase in the number of surviving cells, they can use the pre- adaptation technique before the production of any products. The pharmaceutical industry can also apply the mechanisms of using multi- stress pre- adapted cells for their treatment of different diseases. The pre- adaptation of probiotic cells to multiple stress factors will be beneficial to the consumer because they will be getting the adequate number of live cells when they ingest probiotic products.