الفهرس 
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Abstract
Evaluation of some Bacteria and yeast for production of probiotic
The aim of this study was to obtain potential probiotic microorganisms isolated from different food products for their safety and functional properties.
To achieve this goal, the following experiments were conducted:
1. Isolation of probiotic microorganisms (PBM). 120 bacteria and yeast isolates were obtained from 6 different food product samples, namely, Egyptian artisanal cheeses, fermented chickpeas, fermented rice, natural yogurt, pickles, and raw milk were used as isolation sources of PBM.
2. Screening and selection of PBM isolates through hemolytic activity. PBM isolates were selected according to the hemolytic activity to select the generally recognized as safe (GRAS) isolates for safe application and human consumption. Out of 120 isolates, 75 isolates (19 yeast & 56 bacteria) exhibited γ-hemolysis, and thus considered GRAS, and selected for complementary characterization and identification.
3. Physiochemical characterization. The selected PBMs were tested for resistance to gastric conditions of low acidity and bile salt concentration, pathogenicity, toxicity, antibiotics resistance, synthesis of antimicrobial compounds, antimicrobial activity, and inhibition of enteric pathogens, auto-aggregation and co-aggregation.
a. The 75 PBM isolates were varied for pH tolerance from pH 2 to 6.5.
b. The 75 GRAS-PBM bacteria and yeast isolates exhibited significant tolerance to bile salts at 0.3% -1.5%, and the most suitable bile salt concentration was found to be 0.3% for more than 53% of the total PBM isolates.
c. Antagonistic activity. Among 56 bacterial isolates, 8 isolates were effective against all the five tested pathogens, 45 isolates inhibited Bacillus cereus, E. coli O157:H7, Salmonella typhi and 48 isolates inhibited Candida albicans and 46 isolate inhibited Listeria monocytogenes, while 6 isolates, namely exhibited no antagonistic effect.
d. Total acidity by PBM isolates. Total acidity as milli equivalent per liter (meq/l) was estimated for all PBM isolates ranged from 50 to 200 meq/l for five isolates and acidity ranged from 2 to 1.57 %. For the yeast PBM isolates, the lowest pH was 3.8-4.0 and total acidity of 130 and 150 meq/l with 1.35 and 1.11 % acidity, respectively.
e. Carbohydrates fermentation by tested PBM isolates. The selected. 30 PBM isolates were tested for fermentation nine sugars (Arabinose, Glucose, Fructose, Maltose, Mannitol, Lactose, Ribose, Sucrose, and Raffinose). Results showed that only 15 PBM isolates (one of which is yeast) could ferment all nine sugars, 2 isolates were able to ferment 88.9%, and the rest fermented between 77 to 22% of the sugars.
f. Antibiotic sensitivity. 18 PBM isolates, from the carbohydrates fermentation assay, were tested for antibiotic sensitivity, and results showed that 6 were sensitive for Ampicillin, Ciprophloxacin, Nystatin, Streptomycin, Tetracycline, Cotrim, and Chloramphenicol, with susceptibility of resistant with 44.4%, 22.2%, 11.1%, 22.2%, 27.8%, 22.2%, and 72.2%, respectively. Chloramphenicol was shown to be the most effective antibiotic, while nystatin was the lowest effective antibiotic. Based on these results, the four isolates were selected for identification.
4. Phenotypic and genotypic identification of the pioneer PBM cultures
The selected four isolated, PB1, PB7, PC2, and PY8, were gone through phenotypic identification based on metabolic fingerprint principle conducted by Biolog system, and isolates were identified as Bacillus, Lacticaseibacillus, Pediococcus, and Saccharomyces species, respectively.
5. Genotypic characteristics and the phylogenetic tree. Molecular identification and classification based on 16S and 18S rDNA sequence analysis were performed for PB1, PB7, PC2, and PY8 isolates and results also showed that these isolates were identified as Bacillus bingmayongesis, Lacticaseibacillus paracasei, Pediococcus pentosaceus, and Saccharomyces cerevisiae.
6. In vitro adhesion assay of PBM strains. The four selected PBM strains of L. paracasei, B. bingmayongesis, P. pentosaceus, and S. cerevisiae were tested for their ability to adhere to colon mucus epithelial cells. Scanning electron micrographs showed that all the selected PBM strains exhibited excellent adhesion to the colon epithelial cell Caco-2 cell lines. L. paracasei has the most significant adhesion better followed by B. bingmayongesis then P. pentosaceus, and S. cerevisiae.
7. In-vitro auto-aggregation and co-aggregation assays for PBM strains. Auto-aggregation and co-aggregation were investigated for the four selected PBM strains and five indicator pathogenic strains on the basis of their sedimentation characteristics. Results showed that L. paracasei had the strongest auto-aggregation among the other strains. Results of coaggregation ranged from 81 to 95%.
8.Specific activities were tests for the selected PBM strains, and results showed that the selected strains were able to produce EPS, organic acids and bacteriocins and have antioxidant and antitumor activities.
9. Substances produced by the selected PBM strains were identified by GC-MS technique.
Conclusions:
Based on all the performed tests, the selected PBM strains showed to be safe for human use, based on their γ-hemolysis activity, efficient probiotic, due to their significant production of antioxidant, antitumor and exopolysaccharides, adhesion and carbohydrates fermentation abilities, and antibiotic sensitivity. Therefore, these four strains have potential to be applies as food-additive probiotic for human consumption.