In vitro study of potentially probiotic lactic acid bacteria strains isolated from kimchi

The objective of the present study was to investigate lactic acid bacteria (LAB) isolated from kimchi for their potential probiotic use. Ten preselected LAB strains were evaluated for their functionality and safety. Examined characteristics included acid and bile tolerance, cell adhesion, antimicrobial activity against pathogens, hemolytic activity, undesirable biochemical characteristics, and antibiotic resistance. Results indicated that consumption of these 10 strains does not pose any health risk, as they were not hemolytic and exhibited no undesirable biochemical activity or antibiotic resistance. In particular, three strains, Lactobacillus plantarum NO1, Pediococcus pentosaceus MP1, and Lactobacillus plantarum AF1, showed high degrees of acid and bile tolerance, adherence to Caco-2 and HT-29 cells, and antimicrobial activity against four pathogens (Staphylococcus aureus, Escherichia coli O157:H7, Salmonella typhi, and Listeria monocytogenes). These results suggest that LAB strains from kimchi may have potential use as novel probiotics.     

Growth studies of potentially probiotic lactic acid bacteria in cereal-based substrates

AIMS: The overall growth kinetics of four potentially probiotic strains (Lactobacillus fermentum, Lact. reuteri, Lact. acidophilus and Lact. plantarum) cultured in malt, barley and wheat media were investigated. The objectives were to identify the main factors influencing the growth and metabolic activity of each strain in association with the cereal substrate. METHODS AND RESULTS: All fermentations were performed without pH control. A logistic-type equation, which included a growth inhibition term, was used to describe the experimental data. In the malt medium, all strains attained high maximum cell populations (8.10-10.11 log10 cfu ml(-1), depending on the strain), probably due to the availability of maltose, sucrose, glucose, fructose (approx. 15 g l(-1) total fermentable sugars) and free amino nitrogen (approx. 80 mg l(-1)). The consumption of sugars during the exponential phase (10-12 h) resulted in the accumulation of lactic acid (1.06-1.99 g l(-1)) and acetic acid (0.29-0.59 g l(-1)), which progressively decreased the pH of the medium. Each strain demonstrated a specific preference for one or more sugars. Since small amounts of sugars were consumed by the end of the exponential phase (17-43%), the decisive growth-limiting factor was probably the pH, which at that time ranged between 3.40 and 3.77 for all of the strains. Analysis of the metabolic products confirmed the heterofermentative or homofermentative nature of the strains used, except in the case of Lact. acidophilus which demonstrated a shift towards the heterofermentative pathway. All strains produced acetic acid during the exponential phase, which could be attributed to the presence of oxygen. Lactobacillus plantarum, Lact. reuteri and Lact. fermentum continued to consume the remaining sugars and accumulate metabolic products in the medium, probably due to energy requirements for cell viability, while Lact. acidophilus entered directly into the decline phase. In the barley and wheat media all strains, especially Lact. acidophilus and Lact. reuteri, attained lower maximum cell populations (7.20-9.43 log10 cfu ml(-1)) than in the malt medium. This could be attributed to the low sugar content (3-4 g l(-1) total fermentable sugar for each medium) and the low free amino nitrogen concentration (15.3-26.6 mg l(-1)). In all fermentations, the microbial growth ceased at pH values (3.73-4.88, depending on the strain) lower than those observed for malt fermentations, which suggests that substrate deficiency in sugars and free amino nitrogen contributed to growth limitation. CONCLUSIONS: The malt medium supported the growth of all strains more than barley and wheat media due to its chemical composition, while Lact. plantarum and Lact. fermentum appeared to be less fastidious and more resistant to acidic conditions than Lact. acidophilus and Lact. reuteri. SIGNIFICANCE AND IMPACT OF THE STUDY: Cereals are suitable substrates for the growth of potentially probiotic lactic acid bacteria.     

Identification and probiotic potential of lactic acid bacteria from camel milk

In the present study, a total of 80 presumed lactic acid bacteria (LAB) were isolated from camel milk. Selected LAB were identified as Lactococcus lactis (cam 12), Enterococcus lactis (cam 14) and Lactobacillus plantarum (cam 15) and their potential were tested by tolerance & de-conjugation of bile salts, antimicrobial activity, surface hydrophobicity and adhesion potential) along with this of probiotics were evaluated for curd formation and assessed for sensory properties and syneresis. Selected LABs showed antimicrobial activity against wide range of pathogenic bacteria (Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus cereus and Escherchiaia. coli). LAB (cam 12, cam 14 and cam15) were highly sceptible to chloramphenicol, vancomycin, and tetracyclin. In vitro adhesion studies with Caco-2 cells demonstrated strong adhesion activity with hydrophobicity (99%) was observed. Acute oral toxicity of E. lactis and L. plantarum showed non-toxic, non-virulent and safe for industrial application. The study provides potential LAB which may act as a substitute of functional food, synthetic feed and industrial curd formulation with in the shortest span (240 min at 28–32 °C).     

Fermentation of pomegranate juice by probiotic lactic acid bacteria

In this research, production of probiotic pomegranate juice through its fermentation by four strains of lactic acid bacteria: Lactobacillus plantarum, L. delbruekii, L. paracasei, L. acidophilus was examined. Fermentation was carried out at 30°C for 72 h under microaerophilic conditions. Microbial population, pH, titrable acidity, sugar and organic acid metabolism were measured during the fermentation period and the viability of all strains was also determined during the storage time at 4°C within 4 weeks. The results indicated that L. plantarum and L. delbruekii increased the pH sharply at the initial stages of fermentation and the sugar consumption was also higher in comparison with other strains, better microbial growth was also observed for these two strains during fermentation. Citric acid, as a major organic acid in pomegranate juice was significantly consumed by all probiotic lactic acid bacteria. L. plantarum and L. delbruekii showed higher viability during the storage time. Viable cells remained at their maximum level within 2 weeks but decreased dramatically after 4 weeks. Pomegranate juice was proved to be a suitable media for production of a fermented probiotic drink.     

Boza, a natural source of probiotic lactic acid bacteria

Aims: To evaluate the probiotic properties of strains isolated from boza, a traditional beverage produced from cereals. Methods and Results: The strains survived low pH conditions (pH 3·0), grew well at pH 9·0 and were not inhibited by the presence of 0·3% (w/v) oxbile. Cytotoxicity levels of the bacteriocins, expressed as CC₅₀, ranged from 38 to 3776 μg ml^?1. Bacteriocin bacST284BZ revealed high activity (EC₅₀ = 735 μg ml^?1) against herpes simplex virus type 1. Growth of Mycobacterium tuberculosis was 69% repressed after 5 days in the presence of bacST194BZ. Various levels of auto‐cell aggregation and co‐aggregation with Listeria innocua LMG 13568 were observed. Adhesion of the probiotic strains to HT‐29 cells ranged from 18 to 22%. Conclusions: Boza is a rich source of probiotic lactic acid bacteria. All strains survived conditions simulating the gastrointestinal tract and produced bacteriocins active against a number of pathogens. Adherence to HT‐29 and Caco‐2 cells was within the range reported for Lactobacillus rhamnosus GG, a well‐known probiotic. In addition, the high hydrophobicity readings recorded define the strains as good probiotics. Significance and Impact of the Study: Boza contains a number of different probiotic lactic acid bacteria and could be marketed as a functional food product.     

Isolation and evaluation of probiotic potential of lactic acid bacteria isolated from poultry intestine

The gut is a source of lactic acid bacteria with remarkable functional and technologies properties as well as a potential source of probiotics. In the present study, 13 strains of Lactobacillus were isolated from poultry intestine and identified according to their 16S rDNA sequences, as well as the evaluation of their probiotic potential. The probiotic properties were tested in aspects of antibiotic susceptibility, antimicrobial activity, exopolysaccharide production, lysozyme tolerance, gut condition tolerance (low pH, bile salt tolerance) and adhesion to human colorectal adenocarcinoma cell line (Caco-2). Most isolates were resistance to streptomycin (10 μg/mL), gentamicin (10 μg/mL), kanamycin (30 μg/mL), penicillin (10 μg/mL) and chloramphenicol (30 μg/mL). Isolates shows strong abilities to adhere to Caco-2 cell in the range of (76 to 85%). Isolates SHA101 to SHA113 showed high survival rate under gastrointestinal tract condition (>80%), indicating their potential in application of probiotics. The results of these tests indicate that the lactic acid bacteria isolated from poultry intestine have potential use as probiotic in various products.     

Safety and probiotic functionality of isolated goat milk lactic acid bacteria

Purpose

Lactic acid bacteria (LAB) are traditionally employed in the food industry. LAB strains from goat milk may also present probiotic potential, and it is fundamental to study the safety and functionality aspects which are desirable for their use in food. The objective of this study was to verify the probiotic potential of lactic bacteria isolated from goat milk.

Methods

The presence of safety-related virulence factors (hemolytic activity, gelatinase production, coagulase, and sensitivity to antibiotics) as well as functionality (exopolysaccharide (EPS) production, proteolytic activity, autoaggregation, gas production, survival in the gastrointestinal tract, and antimicrobial activity against bacteria that impair oral health) were determined.

Result

The selected LAB strains are safe against the evaluated parameters and have characteristics of possible probiotic candidates. Especially L. plantarum (DF60Mi) and Lactococcus lactis (DF04Mi) have potential to be added to foods because they have better resistance to simulated gastrointestinal conditions. In addition, they are isolated with already proven antimicrobial activity against Listeria monocytogenes, an important food-borne pathogen. DF60Mi was able to produce EPS (exopolysaccharides). LS2 and DF4Mi strains, both Lactococcus lactis subsp. lactis, demonstrated antimicrobial activity against S. mutans ATCC 25175, a recurrent microorganism in oral pathologies, mainly caries.

Conclusion

This study provides subsidies for future exploration of the potentialities of these LAB strains for both the development of new functional foods and for application in oral health.

     

Potential probiotic properties of lactic acid bacteria isolated from the intestinal mucosa of healthy piglets

In the present study, the probiotic properties of 52 lactic acid bacteria strains, isolated from the intestinal mucosa of 60-day-old healthy piglets, were evaluated in vitro in order to acquire probiotics of potential application. Based on acidic and bile salt resistance, 11 lactic acid bacteria strains were selected, among which 1 was identified as Pediococcus acidilactici, 3 as Enterococcus faecium, 3 as Lactobacillus rhamnosus, 2 as Lactobacillus brevis, and 2 as Lactobacillus plantarum by 16S rRNA gene sequencing. All selected strains were further investigated for transit tolerance in simulated upper gastrointestinal tract, for adhesion capacity to swine intestinal epithelial cells J2 (IPEC-J2), for cell surface characteristics including hydrophobicity, co-aggregation and auto-aggregation, and for antimicrobial activities. Moreover, hemolytic, bile salt hydrolase and biogenic amine-producing abilities were investigated for safety assessment. Two E. faecium (WEI-9 and WEI-10) and one L. plantarum (WEI-51) exhibited good simulated upper gastrointestinal tract tolerance, and showed high auto-aggregation and co-aggregation with Escherichia coli 1570. The strains WEI-9 and WEI-10 demonstrated the highest adherence capacity. The 11 selected strains mentioned above exhibited strong antimicrobial activity against E. coli CVCC1570, Staphylococcus aureus CVCC1882 and Salmonella pullorum AS1.1859. None of the 11 selected strains, except WEI-9 and WEI-33, exhibited bile salt hydrolase, hemolytic or biogenic amine-producing abilities. This work showed that the E. faecium WEI-10 and L. plantarum WEI-51were found to have the probiotic properties required for use as potential probiotics in animal feed supplements.     

Screening of species-specific lactic acid bacteria for veal calves multi-strain probiotic adjuncts

The selection of promising specific species of lactic acid bacteria with potential probiotic characteristics is of particular interest in producing multi species-specific probiotic adjuncts in veal calves rearing. The aim of the present work was to select and evaluate in?vitro the functional activity of lactic acid bacteria, Bifidobacterium longum and Bacillus coagulans strains isolated from veal calves in order to assess their potential use as multi species-specific probiotics for veal calves. For this purpose, bacterial strains isolated from faeces collected from 40 healthy 50-day-calves, were identified by RiboPrinter and 16s rRNA gene sequence. The most frequent strains belonged to the species B. longum, Streptococcus bovis, Lactobacillus animalis and Streptococcus macedonicus. Among these, 7 strains were chosen for testing their probiotic characteristics in?vitro. Three strains, namely L. animalis SB310, Lactobacillus paracasei subsp. paracasei SB137 and B. coagulans SB117 showed varying individual but promising capabilities to survive in the gastrointestinal tract, to adhere, to produce antimicrobial compounds. These three selected species-specific bacteria demonstrated in?vitro, both singularly and mixed, the functional properties needed for their use as potential probiotics in veal calves.     

Mitogenic response and probiotic characteristics of lactic acid bacteria isolated from indigenously pickled vegetables and fermented beverages

Lactic acid bacteria from indigenous pickled vegetables and fermented beverages (fermented rice and Madhuca longifolia flowers) were isolated and investigated for their functional characteristics in vitro as potential new probiotic strains. Four isolates (all Lactobacillus spp.) selected on the basis of high tolerance to bile (0.2%) were identified by standard and molecular methods (16S rDNA) as L. helveticus, L. casei, L. delbrueckii and L. bulgaricus from pickled vegetables and fermented beverages respectively. These selected strains had antibiotic resistance, tolerance to artificial gastric juice and phenol (0.4%), enzymatic profile, and antagonistic activity against enteric pathogens (Enterobacter sakazakii, Salmonella typhimurium, Shigella flexneri 2a, Listeria monocytogenes, Yersinia enterocolitica and Aeromonas hydrophila). All strains survived well in artificial gastric juice at low pH (3.0) values for 4 h, possessed bile salt hydrolase activity and were susceptible to most antibiotics including vancomycin. Additionally, the isolates exhibited high tolerance to phenol, high cell surface hydrophobicity (>60%) and induced proliferation of murine splenocytes. All the four strains of present study suppressed the Con A-stimulated proliferation of the mouse spleen cells, although L. casei had the strongest suppressive effect. The results of this study suggest a potential application of the strains (following human clinical trials), for developing probiotic foods.     

Updating the importance of lactic acid bacteria in fish farming: natural occurrence and probiotic treatments

Many recent papers have deepened the state of knowledge about lactic acid bacteria (LAB) in fish gut. In spite of high variability in fish microbiota, LAB are sometimes abundant in the intestine, notably in freshwater fish. Several strains of Streptococcus are pathogenic to fish. Streptococcus iniae and Lactococcus garvieae are major fish pathogens, against which commercial vaccines are available. Fortunately, most LAB are harmless, and some strains have been reported for beneficial effects on fish health. A major step forward in recent years was the converging evidence that LAB can stimulate the immune system in fish. An open question is whether viability can affect immunostimulation. The issue is crucial to commercialize live probiotics rather than inactivated preparations or extracts. There has been a regain of interest in allochthonous strains used as probiotics for terrestrial animals or humans, due to economical and regulatory constraints, but the short survival in sea water may limit application to marine fish. If viability is required, alternative treatments may include the incorporation of prebiotics in feed, and other dietary manipulations that could promote intestinal LAB. Antagonism to pathogens is the other main feature of candidate probiotics, and there are many reports concerning mainly carnobacteria and Enterococcus. Some bacteriocins were characterized which may be of interest not only for aquaculture, but also for food preservation.     

Exopolysaccharides from lactic acid bacteria: perspectives and challenges

Some lactic acid bacteria (LAB) secrete a polysaccharide polymer. This extracellular polysaccharide, or "exopolysaccharide" (EPS), is economically important because it can impart functional effects to foods and confer beneficial health effects. LAB have a "Generally Recognized As Safe" (GRAS) classification and are likely candidates for the production of functional EPSs. Current challenges are to improve the productivity of EPSs from LAB and to produce EPSs of a structure and size that impart the desired functionality. The engineering of improvements in these properties will depend on a deep understanding of the EPS biosynthetic metabolism and of how the structure of EPSs relates to a functional effect when incorporated into a food matrix.     

Purification,kinetic and functional characterization of membrane bound dipeptidyl peptidase-III from NCDC 252: a probiotic lactic acid bacteria

Molecular Biology Reports - Pediococcus acidilactici is a probiotic lactic acid bacteria possessing studied in-vitro probiotic properties. Study of membrane proteins is crucial in developing...     

Screening of potential probiotic lactic acid bacteria and production of amylase and its partial purification

Probiotics are the healthy living bacteria when administered in adequate amounts confers health benefits in the host. The main objective of present study was to screen the bacteria for potential probiotic characters and enzyme production. The probiotic characters like tolerance to low pH, bile salts, antibiotic sensitivity, hydrophobicity and auto-aggregation properties were evaluated. Among all isolates Lactobacillus fermentum and Lactobacillus sp G3_4_1TO2 showed maximum potential probiotic characters and produced amylase enzyme by observing the halo zone around the colonies with the diameter 0.9?mm and 1.23?mm. Lactobacillus sp G3_4_1TO2 produced maximum amylase when compared with Lb. fermentum. The protein yield was 55.4% with the specific activity of 88.9 U/mg and obtained 40.8% purification fold. The molecular weight of amylase enzyme determined by SDS PAGE was 95,000?Da. From the present study it was considered that Lactobacillus sp G3_4_1TO2 was a potential probiotic bacteria producing maximum amylase enzyme.     

Characterization of lactic acid bacteria isolated from seafood

Lactic acid bacteria were isolated from various samples of seafood: fresh pollock, brine shrimp, gravad fish, vacuum-packed seafood (surimi, smoked tuna, salted cod), and fish stored under 100% CO₂ at 5°C (smoked tuna, fresh and salted cod, salmon). Eighty-six independent isolates were obtained and were grouped according to cell morphology, presence or absence of diaminopimelic acid in the cell wall, and lactate configuration. Fifty-four isolates were identified as belonging to the genus Lactococcus and most of them exhibited DNA homologies with L. lactis subsp. lactis. Four strains were identified as Lactobacillus plantarum , eight strains as genus Leuconostoc and 16 belonged to the genus Carnobacterium. One facultative heterofermentative Lactobacillus and three other isolates were not identified. Of the strains 47% showed similar patterns of carbohydrate fermentations especially among strains belonging to the genera Lactococcus and Carnobacterium. Most of the strains (64%) grew at 5°C, in salted media and in fish extract medium without added sugar. Carnobacterium piscicola and Carn. divergens were the only reference strains able to grow in the same conditions as well as psychrotroph strains isolated from seafood. A numerical analysis could not be used because of the divergent properties of isolates of the same genus and strong similarities between different genera.     

In vitro screening of probiotic lactic acid bacteria and prebiotic glucooligosaccharides to select effective synbiotics

Probiotics and prebiotics have been demonstrated to positively modulate the intestinal microflora and could promote host health. Although some studies have been performed on combinations of probiotics and prebiotics, constituting synbiotics, results on the synergistic effects tend to be discordant in the published works. The first aim of our study was to screen some lactic acid bacteria on the basis of probiotic characteristics (resistance to intestinal conditions, inhibition of pathogenic strains). Bifidobacterium was the most resistant genus whereas Lactobacillus farciminis was strongly inhibited. The inhibitory effect on pathogen growth was strain dependent but lactobacilli were the most effective, especially L. farciminis. The second aim of the work was to select glucooligosaccharides for their ability to support the growth of the probiotics tested. We demonstrated the selective fermentability of oligodextran and oligoalternan by probiotic bacteria, especially the bifidobacteria, for shorter degrees of polymerisation and absence of metabolism by pathogenic bacteria. Thus, the observed characteristics confer potential prebiotic properties on these glucooligosaccharides, to be further confirmed in vivo, and suggest some possible applications in synbiotic combinations with the selected probiotics. Furthermore, the distinctive patterns of the different genera suggest a combination of lactobacilli and bifidobacteria with complementary probiotic effects in addition to the prebiotic ones. These associations should be further evaluated for their synbiotic effects through in vitro and in vivo models.     

Characterization of extracellular yeast peptide factors and their stress-protective effect on probiotic lactic acid bacteria

Protective effect of the extracellular peptide fraction (reactivating factors, RF) produced by yeasts of various taxonomic groups (Saccharomyces cerevisiae, Kluyveromyces lactis, Candida utilis, and Yarrowia lipolytica) on probiotic lactic acid bacteria (LAB) Lactobacillus casei, L. acidophilus, and L. reuteri under bile salt (BS)-induced stress was shown. RF of all yeasts were shown to be of peptide nature; the active component of the S. cerevisiae RF was identified as a combination of low-molecular polypeptides with molecular masses of 0.6 to 1.5 kDa. The protective and reactivating effects of the yeast factors were not species-specific and were similar to those of the Luteococcus japonicus subsp. casei RF. In BS-treated cells of the tester bacteria, a protective effect was observed after 10-min preincubation of the LAB cell suspension with yeast RF: the number of surviving cells (CFU) was 2 to 4.5 times higher than in the control. The reactivating effect was observed when RF was added to LAB cell suspensions not later than 15 min after stress treatment. It was less pronounced than the protector effect, with the CFU number 1 to 3 times that of the control. Both the protector and the reactivating effects were most pronounced in the S. cerevisiae and decreased in the row C. utilis > K. lactis > Y. lipolytica. The efficiency of protective action of yeast RF was found to depend on the properties of recipient LAB cells, with the L. casei strain being most sensitive to BS treatment. In both variants, the highest protective effect of RF (increase in the CFU number) was observed for L. acidophilus, while the least pronounced one was observed for L. casei. The reasons for application of the LAB strains combining high stress resistance and high response to stress-protecting metabolites, including RF factors, as probiotics, is discussed.