Isolation and Purification of β-Galactosidase Enzyme From Local Lactic Acid Bacteria Isolates to Overcome The Phenomenon of Non-Degradation of Milk Lactose

This study aim is to purify β-galactosidase from a local isolate of yogurt in Salah al-Din Governorate to overcome the phenomenon of lactose in decomposition. The bacteria were grown on MRS medium supplemented with 1%CaCo3. Twenty isolates of lactic acid bacteria were obtained and conducting culture tests and microscopic examinations were on these isolates. In order to classify them to the level of species, it was found that there were four types, namely: Lactobacilluse acidophilus,LactobacilluseCasei,Lactobacilluse delubrici subsp.bulgaricus,Streptococcus thermophilus, The cultivation and activation steps of the different isolates were carried out forobtaining the most productive and active isolate, which is Lactobacilluse acidophiluse. Beta-galactosidase activation processes were carried out for the enzyme and cell breakdown by lysozyme. Purification and sedimentation processes were carried out using ammonium sulfate and membrane sorting, followed by gel filtration using Lactobacillus G-150. The best extraction rate (L.acidophiluse 70%) was achieved by enzyme precipitation (4,375) units/mol, and the activity increased in the membrane sorting step to (5,900) units/mol, and in gel filtration we obtained activity of the enzyme (15.591) units/mol.

Optimizatiaon of MRS (De Man, Rogosa and Sharpe) Media with an organic waste substitute to assess the potential of Lactobacillus in Ink degradation

Lactobacillus species are normally found in human digestive and urinary tracts. In this study, the capability of probiotic strains to grow in media using banana peel waste as an alternative carbohydrate source was investigated. Lactic acid bacteria were isolated using homemade curd, banana peel powder was made using an alternate drum drier technique and used as a fermentation media on which strain of probiotic, Lactobacillus sp. were grown. The result showed that bacterial growth using banana peel powder obtained at 370C under anaerobic condition (2- 3 days), was comparable to growth seen in MRS medium. It can be concluded that the banana peel powder can be utilized to produce culture medium for cultivation of probiotic bacteria as a substitute to MRS medium which is expensive for cultivating probiotics. This will not only result in cost reduction of probiotic from cheap, inexpensive Agro-waste material, but also helps in solving the environmental pollution problem caused by their extensive disposal. The capability of Lactobacillus sp. to degrade dye was also investigated. Lactobacilli was incorporated under anaerobic condition in the M9 media containing the dye. The dye degrading capability of Lactobacillus sp was estimated colorimetrically at 520nm. The result showed a decrease in the dye concentration obtained at 37oC under anaerobic condition. So, it can be concluded that the Lactobacillus strain can be used for the bioremediation of dyes.

Comparative study of strains of lactic acid bacteria contained in"Lactos" with imported probiotics

We studied some biological properties of the local strains Lactobacillus helveticus LBMA44c and Lactobacillus delbruesckii subsp.bulgaricus LBMA65b contained in the domestic probiotic LACTOSA in compared to imported probiotics Lactobacillus, Linex-malysh and Acidophilus Probiotic Blend. The strains cultivated in liquid and solid selective MRS medium, obtained as a result of subsequent dilutions of a sample of the probiotics under study, perfectly demonstrated the general features of the morphological and cultural properties of lactic acid bacteria. The authors found that the indicators CFU/g of LACTOS (1.9-2.0x108 CFU/g) is not less than that of imported ones, in terms of the oxidation of nutrient media, it is the same as that of Acidophilus Probiotic Blend-and slightly lower than that of Linex and Lacto-Fit. Strains of lactic acid bacteria in all studied probiotics are well cultivated at 37, 45, 60 degrees (0C) and on a selective medium with an addition at the rate of 2, 4, 6 percent of natural bile and grows poorly on a medium with the addition of 4 and 6 percent NaCl. We also noted that the local strains in our LACTOS probiotic were relatively more resistant to antibiotics than imported ones, according to antibiotic resistance estimates. Thus, the local strains of lactic acid bacteria contained in the domestic probiotic under the LACTOS trademark do not differ from the imported ones. “Лактос” пробиотикт агуулагдах сүүн хүчлийн бактерийн шинж чанарыг импортынхтой харьцуулсан судалгаа “Лактос” пробиотикт агуулагдах сүүн хүчлийн бактерийн Lаctobacillus helveticus LBMA44c болон Lаctobacillus delbruesckii subsp.bulgaricus LВMА65b нутгийн омгийн эмнэлэг-биологийн чадавхын зарим үзүүлэлтийг импортын Lасто-Fit, Линекс Малыш №20, Acidophilus Probiotic Blend-ийнхтэй харьцуулан судлав. Судалгаанд хамрагдсан пробиотикуудын дээжний шингэлэлтээс MRS -ийн хатуу, шингэн сонгомол орчинд ургуулахад тэдгээр нь сүүн хүчлийн бактерийн өсгөвөржилт, хэлбэр зүйн нийтлэг шинж байдлыг үзүүлжээ. Пробиотикт агуулагдах амилах чадвартай бактерийн эсийн тоогоор Лактосынх (1,9-2,0х108КҮН/г) импортынхоос дутахгүй, харин орчныг хүчилсэг болгох идэвхээрээ Линекс Малыш, Lасtо-Fit-ээс бага зэрэг доогуур Acidophilus Probiotic Blend -тэй ойролцоо байна. Тэдгээр нь Цельсийн 37, 45, 60 хэмийн дулаанд болон 2, 4, 6 хувиар цөс нэмсэн тэжээлт орчинд сайн өсгөвөржих боловч цэвэр NaCl-ийн нэмэлт 4-6 хувь хүрэхэд пробиотикийн агууламж дахь сүүн хүчлийн бактерийн ургалтын эрчим дундаас доош гэсэн үнэлэмж рүү шилжсэн юм. Лактост агуулагдах сүү хүчлийн бактерийн омгууд нь судалгаанд ашигласан 5 төрлийн антибиотик тэсвэрлэх чадавхын дундаж үзүүлэлтээр импортын пробиотикийнхтой ойролцоо ажээ. Түлхүүр үг: пробиотик, сүүн хүчлийн бактери, тэсвэрт чанар, өсгөвөржилт

Protein Expression Level Changes in Weissella koreensis during Garlic Media Fermentation

This study investigated the changes in Weissella koreensis (WK) protein expression levels during fermentation in MRS medium supplemented with garlic of WK. WK was first discovered as lactic acid bacteria (LAB) in a Korean fermented cabbage dish known as kimchi. The number of WK cells in MRS medium with garlic (MBCG) and without (MB) after 7 days was 3.55 × 1010 and 2.55 × 1010 CFU/mL, respectively. To observe the changes in the carbon sources in the media, we measured the glucose, sucrose, lactic acid, and acetic acid levels in each medium (MB and MBCG). Thus, 67.2 ± 2.4 (MB) and 64.2 ± 4.7 (MBCG) mmol−1 of glucose were consumed. For sucrose, the level was 3.5 ± 2.2 (MB), and 3.4 ± 2.5 (MBCG) mmol−1. There was not much difference in the lactic acid and acetic acid levels at 160.8 ± 0.4 (MB) and 159.2 ± 0.2 (MBCG) and 2.4 ± 0.4 (MB) and 2.2 ± 8.1 (MBCG) mmol−1, respectively. After the 7-day fermentation period, two-dimensional electrophoresis (2DE) was used to confirm the protein expression pattern in the WK strain. The results show that the fusA and ssb1 proteins were reduced, and the clpP protein was increased. Afterwards, the expression patterns of the above proteins were confirmed through qRT-PCR. Thus, this study confirms the changes in protein expression levels in Weissella koreensis when garlic was added to the media. This study provides basic data for future studies on the major biosynthetic pathways of WK.

Mycotoxin Removal by Lactobacillus spp. and Their Application in Animal Liquid Feed

The removal of mycotoxins from contaminated feed using lactic acid bacteria (LAB) has been proposed as an inexpensive, safe, and promising mycotoxin decontamination strategy. In this study, viable and heat-inactivated L. acidophilus CIP 76.13T and L. delbrueckii subsp. bulgaricus CIP 101027T cells were investigated for their ability to remove aflatoxin B1 (AFB1), ochratoxin A (OTA), zearalenone (ZEA), and deoxynivalenol (DON) from MRS medium and PBS buffer over a 24 h period at 37 °C. LAB decontamination activity was also assessed in a ZEA-contaminated liquid feed (LF). Residual mycotoxin concentrations were determined by UHPLC-FLD/DAD analysis. In PBS, viable L. acidophilus CIP 76.13T and L. delbrueckii subsp. bulgaricus CIP 101027T cells removed up to 57% and 30% of ZEA and DON, respectively, while AFB1 and OTA reductions were lower than 15%. In MRS, 28% and 33% of ZEA and AFB1 were removed, respectively; OTA and DON reductions were small (≤15%). Regardless of the medium, heat-inactivated cells produced significantly lower mycotoxin reductions than those obtained with viable cells. An adsorption mechanism was suggested to explain the reductions in AFB1 and OTA, while biodegradation could be responsible for the removal of ZEA and DON. Both viable LAB strains reduced ZEA by 23% in contaminated LF after 48 h of incubation. These findings suggest that LAB strains of L. acidophilus CIP 76.13T and L. delbrueckii subsp. bulgaricus CIP 101027T may be applied in the feed industry to reduce mycotoxin contamination.

Effect of Cocultivation on Lactobacillus plantarum Strains Growth and Antagonistic Activity

The use of bacterial starters for the production of fermented foods has several advantages over traditional spontaneous fermentation, as it provides a rapid and controlled decrease of pH, improves the microbiological quality of the product, and prolongs the shelf-life. Fermented foods are typically produced using mixed cultures of lactic acid bacteria (LAB) due to the synergism between their constituent bacterial cultures. So, the compatibility of the LAB strains decides the efficacy of a multi-strain starter. The purpose of this study was to investigate the effect of the cocultivation of Lactobacillus plantarum strains on the growth, acidification, and antagonistic activity to determine suitable strain combinations for fermented vegetable production. Methods. The effect of cocultivation on growth characteristics of four L. plantarum strains was determined in MRS medium and cabbage-based medium with 2.5% NaCl. After 8 h of cultivation at 30°C and 37°C, the number of viable cells (CFU/ml) and the pH of the medium were determined. The antagonistic activity of monocultures of L. plantarum and their six compositions against opportunistic pathogenic microorganisms was determined by the method of delayed antagonism. Results. During growth in MRS broth at 30°C cocultivation of L. plantarum 47SM with L. plantarum 691T or L. plantarum 1047K strains led to enhanced rates of growth compared to the monocultures, suggesting some degree of symbiosis between these strains. Viable cell counts of L. plantarum 47SM, 1047K and 691T strains and ΔpH values of L. plantarum 952K, 1047K, and 691T strains were higher after 8 h growth in the cabbage-based medium at 30°C compared to MRS broth. Despite the intensive growth of L. plantarum monocultures in cabbage-based medium, a significant decrease of viable cell counts and ΔpH values during cocultivation at 30°C were found. Cocultivation did not affect the average size of the growth inhibition zones of most of the indicator strains used. However, growth inhibition zones of Shigella flexneri, Escherichia coli, and Proteus vulgaris decreased in some L. plantarum mixed cultures compared to monocultures. Thus, the growth inhibition zones of E. coli and S. flexneri by mixed culture L. plantarum 47 SM+1047K were significantly smaller compared to the growth inhibition zones of L. plantarum monocultures. Conclusions. Thus, based on the data obtained in present work, we can assume that some of these L. plantarum strains used in the work may be bactericinogenic. Although the four L. plantarum strains studied are compatible when cocultivated in a standard rich MRS medium, the results of cocultivation in a cabbage-based medium with 2.5% NaCl does not allow to recommend the use of these L. plantarum strains simultaneously in the starter for vegetable fermentation. Further investigation of bacteriocinogenic properties and mechanisms of growth inhibition under cocultivation in vegetable-like conditions are needed, which will allow combining of some of these L. plantarum strains with LAB strains of other species or genera to create multi-starters for vegetable fermentation.

Comparative Studies of Inhibitory and Antioxidant Activities, and Organic Acids Compositions of Postbiotics Produced by Probiotic Lactiplantibacillus plantarum Strains Isolated From Malaysian Foods

Despite inflammation being a protective natural defense against imbalance stressors in the body, chronic inflammation could lead to the deterioration of immune response, low production, and poor performance in livestock as well as severe economic losses to the farmers. Postbiotics produced by Lactiplantibacillus plantarum has been reported recently to be a natural source of antioxidant, promoting growth performance, anti-inflammation, and immune responses. However, the effects of fermentation media on the compositions of L. plantarum postbiotic have not been reported elsewhere. Hence, a comparative study was conducted to compare the volatile compounds, organic acid composition, and antioxidant and antimicrobial activities of postbiotics produced by six strains of L. plantarum cultivated by using formulated media and the commercial de Man, Rogosa, and Sharpe (MRS) medium as a control. Postbiotics RG14, RI11, and UL4 produced by using formulated media exhibited higher inhibitory activity against Pediococcus acidilactici 446, Escherichia coli E-30, Salmonella enterica CS3, and vancomycin-resistant Enterococci except for Listeria monocytogenes LS55. As for the antioxidant activity, hydroxyl radical scavenging activity was enhanced in formulated media, whereas reducing power activity was the highest in postbiotic RI11. Three organic acids, namely, acetic acid, caproic acid, and lactic acid, were detected in the postbiotic produced by various L. plantarum strains. The concentration of acetic acid was influenced by the fermentation media, whereas caproic acid was detected as the highest in postbiotic RG11. Lactic acid was the predominant compound detected in all the postbiotics and had the significantly highest concentration in postbiotic RS5 when produced by using the MRS medium. Intermediary and pyrrole compounds were the other main compounds that were detected by using GC-MS. Positive correlations were found between organic acid production and inhibitory activity, as well as antioxidant activity exhibited by postbiotics. In conclusion, the compositions and functional characteristics of postbiotics produced by the six strains of L. plantarum were strain-dependent and affected greatly by the fermentation medium. The effects of postbiotic composition on the functional characteristics of postbiotics were elucidated in this study to warrant their applications as a promising beneficial natural growth promoter for the livestock industry.

Linoleic acid isomerization ability of Lactobacillus spp. isolated from Vietnamese human intestinal origins

Conjugated linoleic acid (CLA) have been shown to exert numerous health benefits, including anti-carcinogenic, anti-atherogenic, anti-diabetic, antiobesity, cholesterol reducing, antioxidant, anti-microbial, immune system modulator and growth-stimulating properties. In human, CLA is produced from Linoleic acid (LA) by gut bacteria. In this study, nineteen Lactobacillus (Lac.) strains isolated from human feces were studied to determine their ability to metabolize LA. The bacteria were grown in the liquid form of anaerobic MRS medium supplemented with 0.5 mg/mL LA. The linoleate isomerase activity in bacteria grown on MRS medium was determined by Gas chromatograpy. The results indicated that 4 out of 19 strains, including strains Lac.02, Lac.05, Lac.14 and Lac.16 are capable of producing about 40-50 μg/mL CLA from LA. Among them, the highest ability to produce CLA from LA is Lac.02 strain. In the production of CLA from LA, enzymes involved in this metabolism in Lactobacillus act as catalysts of hydration/dehydration (CLA-HY), oxidation of hydroxy groups/reduction of oxo groups (CLA-DH), migration of carbon-carbon double bonds (CLA-DC), and saturation of carbon-carbon double bonds (CLA-ER). The cla-dh, cla-dc, cla-hy and cla-er genes that encode enzymes CLA-DH, CLA-DC, and CLA-ER had been found in all Lac.02, Lac.05, Lac.14 and Lac.16 strains. Gas chromatography traces indicated that these strains produced the same compounds, which was subsequently identified as cis-9, trans-11, and trans-10, cis-12 CLA. In the next study, we will optimize the conditions such as substrate concentrations, pH values, temperature and culture time of each strain to obtain the best rerults.

Mixed fruit wastes as a potential source for Bacteriocin Producing Lactic acid bacterium – Lactobacillus saniviri NKSS1

Mixed fruit juices contain microflora on the surface of fruits during the harvest and post-harvest practices. The presence of useful organisms like lactic acid bacteria from the mixed fruit wastes was explored in this study since these microbes use these wastes as a nutrient source for their growth. The lactic acid bacteria isolated using MRS medium was identified as Lactobacillus saniviri NKSS1 by 16s rRNA analysis. The bacteriocin produced by Lactobacillus saniviri NKSS1 showed inhibitory effect against the food pathogen (Listeria monocytogenes) and clinical pathogen (Acinetobacter baumannii). Optimization of bacteriocin production from Lactobacillus saniviri NKSS1 was achieved at 24 h of incubation, temperature at 35 °C with the initial medium pH of 6.5. The carbon & nitrogen sources like dextrose (3% w/v) and yeast extract (0.75% w/v) enhanced the production of bacteriocin in MRS medium. Antimicrobial activity was reduced in the partially purified bacteriocin when incubated at 95 °C for 2 h but it retained its activity in the pH range of 5.5 to 8.5. Whereas, metals like CuSO4 and MgSO4 at (0.5 % w/v) interfered with the antagonistic activity of partially purified bacteriocin. Ionic detergents like SDS and CTAB partially decreased the antimicrobial activity, while other non-ionic detergents inhibited the antimicrobial activity completely. The molecular weight of partially purified bacteriocin from Lactobacillus saniviri NKSS1 was found to be 10.9 kDa. It can be concluded that bacteriocin of L. saniviri NKSS1 holds a promising potential for extension of shelf-life and improvement of microbiological safety in food industries.

Production of Lactobacillus rhamnosus BRM 029693 in feed-batch fermentation

The objective of this work was to produce Lactobacillus rhamnosus BRM 029693 biomass in a fermenter with the aim to expand its production scale. Lactobacillus rhamnosus BRM 029693 was grown in a fermenter under different aeration conditions (in surface and depth) using a commercial substrate (MRS broth) and simplified substrate formulated in the laboratory itself, and under different pH conditions (4.7, 5.2, 5.7). The highest biomass production on a fermenter scale occurred using synthetic MRS, 1 v.v.m. aeration on the medium surface and without pH control (3.2 g/L) among all the tests. The biomass production was only 0.7 g/L when the MRS medium was replaced by simplified MRS. The pH also influenced the biomass production, reaching 0.9 g/L when the fermentation pH was controlled at pH 4.7.