Consortium Probiotic Fermented Milk using Bifidobacterium sp. and Lactobacillus acidophilus Protects against Salmonella typhimurium and Repairs the Intestine

Background: This research investigates the implication effect of consortium microbiota probiotic Lactobacillus bulgaricus, Streptococcus thermophilus, Lactobacillus acidophilus and Bifidobacterium sp. in yogurt on the zone of inhibition against Salmonella typhimurium in yogurt, also the activities of the enzyme. Methods: The experiment used a randomized complete design (CRD) with four treatments and two replicates. T1: Yogurt A with Streptococcus thermophilus , Lactobacillus acidophilus, Bifidobacterium spp and Lactobacillus bulgaricus.; T2: Yoghurt A + S from cow milk + skim milk 5%.; T3: Pure cow milk; and T4: Amoxicillin 100 ppm. Analysis of variance was used to look at the data numerically and then Duncan’s multiple range test was done. This study was carried out at Laboratory of Biochemistry Mathematics and Natural Sciences. The research started from February 2 nd until 20 th , 2023. Result: Results indicated that the probiotic microbiota produced higher enzyme activities and showed the inhibition zone better than milk, even though there is no significant effect compared to antibiotic control.


INTRODUCTION
The nutritional value of yogurt is determined by the milk as the sources and processing factors. These variables include temperature, heat duration and storage conditions. In addition, the nutritional and physiological properties of the product are contingent upon the specific probiotic strain utilised during the fermentation process (Mazza, 1998;. Milk is a valuable dietary source of essential vitamins, including B-6, B-12, riboflavin, niacin and folic acid. Folate is a B vitamin that can be produced by several species of lactic acid bacteria. Several bacterial species are utilized for milk fermentation and the potential of yogurt production to biosynthesize S. thermophilus and Bifidobacteria, which are known to generate folate, was evaluated. W hen Bifidobacterium sp. and S. thermophilus are used together, folate can increase six-fold (Crittenden et al., 2003). Yoghurt, besides vitamins, provides 3-4 times the calcium intake through the feed. Essential minerals such as potassium, magnesium, phosphorus and zinc are found in yogurt (Kerry et al., 2001).Calcium is involved in the processes of bone development and mineralization. According to the Recommended Dietary Allowance (RDA), adults' recommended daily calcium intake is approximately 900 mg, while teenagers and the elderly need 1200 m g/day (Gueguen and Pointillart, 2000).
In terms of disease prevention and health advantages, probiotics are acquiring popularity. Growth, adhesion and invasion of Salmonella cells can be inhibited by Bifidobacterium sp. according to in vitro studies. Likewise, it alters the immune response by reducing the secretion of interleukin (IL)-8 and the production of tumour necrosis factor (TNF) in the small intestine.
This study aims to determine how gut health and function are affected by the content of yogurt and Lactic Acid Producing Bacteria (LAB). Salmonella induced small intestinal damage may reduce villi-associated enzyme activity. This research is necessary because cases of fever due to Salmonella typhi or typhimurium in Indonesia range between 350-810 per 100,000 population. This disease affects 1.6% of Indonesia, the fifth most common infectious illness affecting all ages in Indonesia, which is 6.0% and ranks 15 th for causes of death for all ages in Indonesia.

Materials
The milk is from Koperasi Peternak Sapi Perah Bandung Utara (KPSBU) Lembang, W est Java, Indonesia. Streptococcus thermophilus, Lactobacillus acidophilus, Bifidobacterium sp. and Lactobacillus bulgaricus are all pure cultivated bacteria. Milk and Amoxicillin is the control treatment. Milk is a base for making yogurt. Amoxicillin is an antibiotic frequently used to kill pathogenic bacteria and Salmonella sp. in this research.

Statistical analysis
A Completely Randomized Design (CRD) was used in the trial, with four treatments, each treatments has two replications. Analysis of Variance was used to look at the numbers of the data and then Duncan's multiple range test was done. This study was carried out at Laboratory of Biochemistry Mathematics and Natural Sciences. The research started from February 2 to 20, 2023. The following are four treatments: T1: Yogurt A with Streptococcus thermophilus, Lactobacillus acidophilus, Bifidobacterium sp. and Lactobacillus bulgaricus. T2: Yoghurt A + S from cow milk + skim milk 5%.
T3: Pure cow milk. T4: Amoxicillin 100 ppm. Table 1 shows the results. The results showed that Yoghurt A with the consortium amount of Streptococcus thermophilus, Lactobacillus acidophilus, Bifidobacterium sp. and Lactobacillus bulgaricus. Yogurt A+S is fermented milk with the addition of 5% skim milk. The results showed that yoghurt, made only from fermented milk with a mixture of 5% skim milk, showed better results than consuming only unfermented milk. However, the use of 100 ppm amoxicillin showed the best results. Yogurt with a mixture of skim milk showed a higher amount of lactose than youghurt without the addition of skim milk.

RESULTS AND DISCUSSION
Salmonella is a bacteria that causes gastroenteritis, which disrupts normal gut function and causes diarrhea. However, Salmonella will induce an unknown disease when it interacts with its host.
Brush border enzyme activity is reduced by Salmonella typhimurium. These alterations are most likely the result of a combination of minor intestinal morphological changes and the inflammatory response of the host immune system, which may alter enzyme gene expression and protein activity. LPS administration showed physiological changes, decreased body weight and the administration of probiotics containing Bifidobacterium.
Lactic acid bacteria (LAB) used in fermentation contribute to sustaining the nutritional value of various foods. The starter culture is a source of proteolytic enzymes during fermentation, ultimately contributing to product preservation as it produces inhibitory metabolites (O'Keeffe and Hill, 1999). In the digestive tract, lactic acid bacteria are important microbes (Holzapfel et al., 2001). Lactic acid and acetic acid production were produced more by Lactobacillus bulgaricus and Streptococcus thermophilus than Lactobacillus acidophilus and Bifidobacterium sp. (Adriani and Lengkey, Note: The diameter of disc used is 6 mm. *Inhibitation zone are not clear.  , Adriani et al., 2021, also blood lipid . Antibacterial compounds produced by probiotics are able to control intestinal pathogens as competition for nutrients and adhesion sites (Fig 1). In addition, probiotics produce lactic acid, acetic acid and substances such as antibiotics resulting in increase the enzyme activity and antibody levels (Hose and Sozzi, 1991). Based on Table 2

CONCLUSION
It is believed that consuming yogurt, including consortium lactic acid bacteria with probiotics, could inhibit the growth of Salmonella typhimurium, which causes typhus and enhances gut health. Several of these beliefs are supported by numerous studies on the potential health benefits of yogurt in preventing gut-related diseases.In addition to the health benefits of yogurt with live and active cultures such as L. bulgaricus, S. thermophilus, L. acidophilus and B. bifidum, the protection against Salmonella typhimurium is one of the most intriguing preventive effects because it produces protease and lipase enzyme. These results are exciting and should lead to more studies into how yoghurt does what it does and what parts of yoghurt are most important for that to happen.