Screening and Evaluation of Human Intestinal Lactobacilli for the Development of Novel Gastrointestinal Probiotics

The aim of this study was to screen intestinal lactobacilli strains for their advantageous properties to select those that could be used for the development of novel gastrointestinal probiotics. Ninety-three isolates were subjected to screening procedures. Fifty-nine percent of the examined lactobacilli showed the ability to auto-aggregate, 97% tolerated a high concentration of bile (2% w/v), 50% survived for 4 h at pH 3.0, and all strains were unaffected by a high concentration of pancreatin (0.5% w/v). One Lactobacillus buchneri strain was resistant to tetracycline. None of the tested strains caused lysis of human erythrocytes. Six potential probiotic strains were selected for safety evaluation in a mouse model. Five of 6 strains caused no translocation, and were considered safe. In conclusion, several strains belonging to different species and fermentation groups were found that have properties required for a potential probiotic strain. This study was the first phase of a multi-phase study aimed to develop a novel, safe and efficient prophylactic and therapeutic treatment system against gastrointestinal infections using genetically modified probiotic lactobacilli.     

A new Lactobacillus plantarum strain, TN8, from the gastro intestinal tract of poultry induces high cytokine production

This study aimed to determine the probiotic potential of 100 strains of Lactic Acid Bacteria (LAB) isolated from different intestinal segments of indigenous poultry in Tunisia. The strains were submitted to a battery of standard tests and criteria commonly used for determining their probiotic properties and attributes. The findings revealed that 19 of the isolates exhibited antimicrobial activity against 4 pathogenic bacteria, and that 4 (TN1, TN8, TN7, and TN13) showed good resistance to pH 3 and 5% bovine bile. Three isolates, namely TN1, TN8, and TN13, showed sensitivity to several antibiotics and were, therefore, selected for further enzymatic activity assays. Two isolates, namely TN1 and TN8, showed high efficacy of adhesion to chicken enterocytes. The cytokines released after stimulation by the two isolates showed high anti-inflammatory profiles, with an increased rate of Interleukin-10 (IL-10) production for the TN8 strain. Showing the highest performance, TN8 was submitted to 16S rRNA gene sequencing, which revealed that the strain was of the species Lactobacillus plantarum. Overall, the findings indicate that the Lactobacilli from poultry intestine has a number of promising properties that make it candidate for application as a probiotic additive in poultry industry.     

Different probiotic properties for Lactobacillus fermentum strains isolated from swine and poultry

Systematic procedures were used to evaluate the probiotic properties of Lactobacillus fermentum (L. fermentum) strains isolated from swine and poultry. The major properties included their capabilities to adhere to the intestinal epithelium of swine and poultry, the inhibition on pathogenic bacteria, and their tolerance to the gastric juice and bile salts. Results showed that L. fermentum strains from poultry digestive tract showed better adherence to the swine intestine and chicken crop epithelial cells as compared to those strains from the swine origin. In addition, six strains from poultry and one strain from swine showed adhesion specificity to their own intestinal epithelium. Four poultry isolates and one swine isolate were able to adhere to the epithelial cells from both swine and chicken. For gastric juice and bile tolerance, most of the strains isolated from swine or poultry were acid tolerant but less strains were bile intolerant. The spent culture supernatant (SCS) of these L. fermentum strains showed antagonistic effect against the indicator bacteria, such as Escherichia coli, Salmonella spp., Shigella sonnei and some enterotoxigenic Staphylococcus aureus. From the above studies, some L. fermentum strains isolated from poultry were found to have the probiotic properties required for use in animal feed supplement. This study suggested that poultry digestive tract may serve as potential source for the isolation of probiotic lactic acid bacteria.     

Probiotic properties of lactic acid bacteria isolated from stool samples of longevous people in regions of Hotan, Xinjiang and Bama, Guangxi, China

A total of 567 lactic acid bacteria (LAB) strains were isolated from the stool samples of longevous people in regions of Hotan, Xinjiang and Bama, Guangxi, China. In order to reduce the number of strains for further examinations, 36 isolates were screened out for further examination whilst the other strains, which had lower probiotic properties, were not suitable for yogurt production due to the absence of growth in pH 3.5 MRS medium and no curding during fermentation, and so were excluded. The result of identification by API, sequence analysis of 16S rRNA and random amplified polymorphic DNA (RAPD) analysis showed that there were three strains of Lactobacillus acidophilus, 10 strains of Lactobacillus rhamnosus, three strains of Lactobacillus casein, three strains of Lactobacillus brevis, two strains of Enterococcus faecium, two strains of Enterococcus faecalis, four strains of Bifdibacterium infantis, three strains of Bifdibacterium brevise, three strains of Bifdibacterium bifidium, two strains of Bifdibacterium adolecentis and one strain of Bifdibacterium longam among the 36 isolates. These strains were evaluated by in vitro methods including survival upon exposure to pH 2.0, 3.0 and/or 0.3% oxgall and adhesion to the human colon adenocarcinoma cell line Caco-2 as well as antimicrobial activity against potential pathogens. The results presented here show that L. rhamnosus LV108, L. rhamnosus F, B. brevise R39 and B. infantis R42 are acid and bile tolerant, adhere to the cultured human intestinal Caco-2 cell line, antagonistic activity against potential pathogenic bacteria infection in vitro, and so are potential strains for probiotic use.     

Isolation and characterization of Lactobacillus salivarius MTC 1026 as a potential probiotic

Researchers are becoming more interested in studying probiotics at present due to their benefit to human and animal health. In this study, newly isolated strains from human feces were evaluated for probiotic properties. A total of sixty isolated strains were collected from feces and six out of sixty isolated strains could inhibit the growth of Salmonella typhi and Salmonella typhimurium. The strain which gave the best inhibitory effect was selected for further characterization as a probiotic strain. The identification of this strain was analyzed on the basis of morphological and biochemical characteristics and 16S rDNA gene sequence. This strain was Gram-positive, rod shaped, and catalase and oxidase-negative, and produced acids from D-glucose, D-fructose, lactose, mannitol, sorbitol, inulin and starch. It could not hydrolyze esculin or red blood cells. Based on its 16S rDNA gene sequence, it was Lactobacillus salivarius, and so was called L. salivarius MTC 1026 in this study, and was closely related with L. salivarius DSPV 344T isolated from the calf gut. It was able to survive in gastric and small intestinal juices at pH 2.0 and 1.0% bile salt for several hours and also could grow at 45°C. Moreover, this strain showed inhibitory activity against a variety of food-borne pathogens. It was highly sensitive to piperacillin, chloramphenicl, ampicillin, erythromycin and ceftazidime. After plasmid curing, this strain was susceptible to gentamicin, amikacin, norfloxacin and ciprofloxacin. L. salivarius MTC 1026 could significantly inhibit the adhesion process of E. coli ATCC 25922 and S. typhimurium ATCC 13311 on Caco-2 monolayers in a competition assay and also reduced invasion of both pathogens (4-log cfu/ml) in an exclusion and displacement assay. Therefore, it was clearly demonstrated in this study that L. salivarius MTC 1026 has shown promising properties as a candidate for a potential probiotic for applications in humans and animals.     

Probiotic properties of Lactobacillus strains isolated from the feces of breast-fed infants and Taiwanese pickled cabbage

This study assessed potential probiotic Lactobacillus strains isolated from the feces of breast-fed infants and from Taiwanese pickled cabbage for their possible use in probiotic fermented foods by evaluating their (i) in vitro adhesive ability, resistance to biotic stress, resistance to pathogenic bacteria, and production of β-galactosidase; (ii) milk technological properties; and (iii) in vivo adhesive ability, intestinal survival and microbial changes during and after treatment. Five Lactobacillus isolates identified as Lactobacillus reuteri F03, Lactobacillus paracasei F08, Lactobacillus rhamnosus F14, Lactobacillus plantarum C06, and Lactobacillus acidophilus C11 that showed resistance to gastric juice and bile salts were selected for further evaluation of their probiotic properties. All the strains demonstrated the ability to adhere to Caco-2 cells, particularly, strain L. plantarum C06 and L. reuteri F03 showed satisfactory abilities, which were similar to that of the reference strain L. rhamnosus GG. The strains L. paracasei F08 and L. acidophilus C11 had the highest β-galactosidase activity. Most of the strains were resistant to aminoglycosides and vancomycin but sensitive to ampicillin, erythromycin, and penicillin. All the 5 strains elicited antibacterial activity against both Gram-positive (Bacillus cereus, Listeria monocytogenes and Staphylococcus aureus) and -negative (Escherichia coli and Salmonella enterica) pathogens. Moreover, the strains L. reuteri F03, L. paracasei F08, and L. plantarum C06 could grow rapidly in milk without nutrient supplementation and reached 10? cfu/mL after 24 h of fermentation at 37 °C. The viable cell counts of the 3 strains remained above 10? cfu/mL after 21 d of storage at 4 °C. In the animal feeding trial, the number of intestinal lactobacilli increased significantly after administration of milk fermented with the 3 strains, and the counts of fecal coliforms and Clostridium perfringens were markedly reduced. Lactobacillus strains could also survive in the ileal intestinal tissue of the treated rats. Technologically interesting Lactobacillus isolates may be used in the future as probiotic starter cultures for manufacturing novel fermented foods.     

Characteristics of Bacterial Isolates from the Gut of Freshwater Fish, Labeo rohita that May be Useful as Potential Probiotic Bacteria

In this study, we aimed to evaluate the in vitro probiotic characteristics of three bacteria, Lactobacillus plantarum VSG3, Pseudomonas aeruginosa VSG2, and Bacillus subtilis VSG1, isolated from the gut of Labeo rohita. The bacterial isolates tolerated low pH and high bile concentrations in the fish well. The bacterial adhesion capacity to fish intestinal mucosa revealed that the three potential probiotic isolates had a significantly higher adhesion capacity compared to the pathogenic strains tested. L. plantarum VSG3 exhibited the best adhesion capacity (19.1?%) to the intestinal mucosa. Among the isolates, L. plantarum VSG3 and P. aeruginosa VSG2 showed strong antibacterial activities against fish pathogens as measured in spent culture liquids. Moreover, all the isolates were susceptible to each tested antibiotic, which ensured their inability to exhibit antibiotic-resistance properties. Considering these promising results, selected strains should be further studied to determine their probiotic effects in vivo in fish.     

Adherence and colonization properties of Lactobacillus rhamnosus TB1, a broiler chicken isolate

AIMS: Selected lactic acid bacteria (LAB) isolated from intestinal tract of chicken have been studied in order to investigate their ability to adhere in vitro to Basement Membrane Matrigel (BMM). A selected strain showing a good adherence in BMM test was used for in vivo colonization assays. METHODS AND RESULTS: In vitro assessment of adhesion of broiler chicken isolates was performed using BMM assay. Among LAB strains tested, Lactobacillus rhamnosus TB1 showed a good adherence that was comparable to the one of an Escherichia coli EPEC strain used as positive control. For in vivo colonization assays this strain was fluorescently stained with the carboxyfluorescein diacetate succinimidyl ester (cFDA-SE) thus allowing its detection in different layers of intestinal tract after inoculation in broiler chicken. Further, stained L. rhamnosus were found with a highest value in rectum, jejunum and ileum both 3 and 24 h after administration. CONCLUSIONS: BMM assay is a quick method to test in vitro adhesion properties of bacterial strains and cFDA-SE-stained bacteria may be considered as an alternative method to test in vivo adhesion and colonization properties. SIGNIFICANCE AND IMPACT OF THE STUDY: Lactobacillus rhamnosus TB1 was therefore showed to be able to adhere strongly in vitro to BMM and in vivo to intestinal epithelial cells of chicken and may be considered as a potential probiotic for chicken.     

Intraspecific genotypic characterization of Lactobacillus rhamnosus strains intended for probiotic use and isolates of human origin

A set of 118 strains of the species Lactobacillus rhamnosus was collected, including probiotic strains, research strains with potential probiotic properties, food starter cultures, and human isolates. The majority of the strains were collected from companies, hospitals, or culture collections or were obtained after contacting authors who reported clinical case studies in the literature. The present work aimed to reveal the genotypic relationships between strains of these diverse sources. All strains were initially investigated using fluorescent amplified fragment length polymorphism (FAFLP) with three different primer combinations. Numerical analysis of FAFLP data allowed (i) confirmation of the identification of all strains as members of L. rhamnosus and (ii) delineation of seven stable intraspecific FAFLP clusters. Most of these clusters contained both (potentially) probiotic strains and isolates of human origin. For each of the clusters, strains of different sources were selected for pulsed-field gel electrophoresis (PFGE) of macrorestriction fragments obtained with the enzymes NotI and AscI. Analysis of PFGE data indicated that (i) some (potentially) probiotic strains were indistinguishable from other probiotic strains, suggesting that several companies may use duplicate cultures of the same probiotic strain, and (ii) in a number of cases human isolates from sterile body sites were indistinguishable from a particular probiotic strain, suggesting that some of these isolates may be reisolations of commercial strains.     

Species distribution of staphylococci from small wild mammals

A total of 197 isolates of Staphylococcus from small wild animals (insectivores and rodents) were identified by partial sequencing of the rpoB and dnaJ genes. Among the identified isolates the predominant species was S. succinus (28%), followed by S. xylosus (20.8%) and S. stepanovicii (18.3%). The other 14 Staphylococcus species were occasionally isolated. PCR-RFLP of the rpoB gene digested by Hpy8I was a fast and simple method to distinguish the two subspecies of S. succinus. More than 90% of the 55 S. succinus strains isolated belonged to S. succinus subsp. casei and only 9% to S. succinus subsp. succinus. Moreover, the present study describes the first ever isolation of S. fleurettii from healthy animals.     

Characterization of faecal enterococci from rabbits for the selection of probiotic strains

AIMS: To characterize the facultative anaerobic intestinal microbiota of healthy rabbits, especially enterococci, for the selection of potential probiotic strains. METHODS AND RESULTS: Phenotypic and molecular methods were used to identify enterococcal isolates. Results obtained indicated that enterococcal microbiota widely varied among individuals both in size and in composition. Antibacterial and haemolytic activities, and resistance to acid and bile salts were determined. A small group of strains produced bacteriocins active against listeriae and indigenous clostridia and therefore they were selected as potential probiotics. One such strain, 8G, was assayed for colonization capacity. Results obtained suggested that the fate of the introduced strain depended on the composition of the enterococcal indigenous microbiota. CONCLUSIONS: Enterococcus faecalis and Ent. faecium are the predominant enterococcal species in the gut of rabbits. Other species of lactic acid bacteria were not recovered. SIGNIFICANCE AND IMPACT OF THE STUDY: The enterococcal intestinal microbiota of healthy rabbits has been characterized in detail. Monitoring the fate of an introduced probiotic in vivo is required in order to evaluate potential probiotic strains.     

Intraspecific Genotypic Characterization of Lactobacillus rhamnosus Strains Intended for Probiotic Use and Isolates of Human Origin

A set of 118 strains of the species Lactobacillus rhamnosus was collected, including probiotic strains, research strains with potential probiotic properties, food starter cultures, and human isolates. The majority of the strains were collected from companies, hospitals, or culture collections or were obtained after contacting authors who reported clinical case studies in the literature. The present work aimed to reveal the genotypic relationships between strains of these diverse sources. All strains were initially investigated using fluorescent amplified fragment length polymorphism (FAFLP) with three different primer combinations. Numerical analysis of FAFLP data allowed (i) confirmation of the identification of all strains as members of L. rhamnosus and (ii) delineation of seven stable intraspecific FAFLP clusters. Most of these clusters contained both (potentially) probiotic strains and isolates of human origin. For each of the clusters, strains of different sources were selected for pulsed-field gel electrophoresis (PFGE) of macrorestriction fragments obtained with the enzymes NotI and AscI. Analysis of PFGE data indicated that (i) some (potentially) probiotic strains were indistinguishable from other probiotic strains, suggesting that several companies may use duplicate cultures of the same probiotic strain, and (ii) in a number of cases human isolates from sterile body sites were indistinguishable from a particular probiotic strain, suggesting that some of these isolates may be reisolations of commercial strains.     

Characterization of spore forming Bacilli isolated from the human gastrointestinal tract

Aims: To isolate and characterize spore‐former bacteria able to colonize the human gastrointestinal tract (GIT). Methods and Results: A total of 25 spore‐formers was isolated from faeces and ileal biopsies of healthy human volunteers and identified at the species level. Physiological analysis was performed to evaluate the ability of the various isolates to form biofilms, to swarm, to produce surfactants and molecules that have antimicrobial activity against selected pathogens. To assess the potential probiotic activity of the isolates, we tested the resistance of cells and spores to simulated gastric conditions, the ability to grow and sporulate in anaerobic conditions and the presence of toxin‐encoding genes in their genome. Conclusions: Spore‐formers belonging to various bacterial species have been isolated from the gut of healthy human volunteers. These strains appear to be well adapted to the intestinal environment and we propose them as potential probiotic strains for human use and as oral vaccine vehicles. Significance and Impact of the Study: To our knowledge this is the first detailed characterization of spore‐forming Bacilli from the human GIT. Our data suggest that the isolated species do not transit, but rather colonize this specific habitat and propose them as probiotic strains for human use.     

Evaluation of probiotic properties of Lactobacillus plantarum strains isolated from Chinese sauerkraut

Lactobacillus plantarum strains isolated and identified from naturally-fermented Chinese sauerkraut were examined in vitro for potential probiotic properties and in vivo for cholesterol-lowering effect in mice. Among 7 isolated L. plantarum strains, strains S2-5 and S4-1 were found to possess desirable probiotic properties including ability to survive at pH 2.0 for 60 min, tolerate pancreatin and bile salts, adhere to Caco-2 cells, produce high β-galactosidase activity and antimicrobial activity against Escherichia coli O157 and Shigella flexneri CMCC(B). In addition, strains S2-5 and S4-1 were susceptible to several antibiotics, and capable of reducing cholesterol level in MRS medium by assimilation of cholesterol at 20.39 and 22.28 μg ml^?1, respectively. The in vivo study with L. plantarum S4-1 showed that feeding with fermented milk containing this strain was able to effectively reduce serum cholesterol level in mice, demonstrating its potential as an excellent probiotic candidate for applications in functional products.     

Probiotic properties of Lactobacillus isolates originating from porcine intestine and feces

In this study, a total of 94 lactic acid bacterial (LAB) isolates of porcine small intestinal and fecal origin were screened for their probiotic properties. The aim was to evaluate whether their isolation site and putative species identity play a role in these characteristics and whether either of these can be used as a predictive factor for the probiotic potential of bacterial isolates. The isolates were preliminarily identified by partial 16S rRNA gene sequencing and characterized in vitro for their pH and bile tolerance, adhesion capacity towards porcine enterocytes isolated from five intestinal sites and for antimicrobial activity towards five indicator pathogens. The interdependence of these characteristics was statistically evaluated. The isolates tolerated low pH and bile well. Adherence to the enterocytes of different origins did not correlate with the strain isolation site. In general, higher adherence was observed to colon cells in comparison to the small intestinal enterocytes. Culture filtrates of the isolates caused a decrease of up to three orders of magnitude in the intestinal pathogen cell numbers. The inhibition was mostly due to lactic and other organic acids. The predominating phylotypes identified were Lactobacillus reuteri and Lactobacillus salivarius, of which the former generally had the best adhesion capacity, whereas the latter appeared to be the best inhibitor. Based on the results, several strains of the pig Lactobacillus isolates tested may function as promising candidates for use in probiotic products. However, it was not possible to use the isolation site or the species identity of the isolates as reliable preliminary screening factors.     

Quantitative Appraisal of the Probiotic Attributes and In Vitro Adhesion Potential of Anti-listerial Bacteriocin-producing Lactic Acid Bacteria

Estimation of bile tolerance, endurance to gastric and intestinal environment and adhesion potential to intestinal cells are significant selection criteria for probiotic lactic acid bacteria (LAB). In this paper, the probiotic potential of native bacteriocin-producing LAB isolated previously from indigenous source has been determined through quantitative approaches. Among fifteen anti-listerial bacteriocin-producing native LAB, ten strains were found to be bile tolerant. The presence of bile salt hydrolase (bsh) gene in native Lactobacillus plantarum strains was detected by PCR and confirmed by nucleic acid sequencing of a representative amplicon. Interestingly, three native LAB strains exhibited significant viability in simulated gastric fluid, analogous to the standard LAB Lactobacillus rhamnosus GG, while an overwhelming majority of the native LAB strains demonstrated the ability to survive and remain viable in simulated intestinal fluid. Quantitative adhesion assays based on conventional plating method and a fluorescence-based method revealed that the LAB isolates obtained from dried fish displayed significant in vitro adhesion potential to human adenocarcinoma HT-29 cells, and the adhesion level was comparable to some of the standard probiotic LAB strains. The present study unravels putative probiotic attributes in certain bacteriocin-producing LAB strains of non-human origin, which on further in vivo characterization could find specific applications in probiotic food formulations targeted for health benefits.     

Probiotic Potential of Bacillus Strains Isolated from an Acidic Fermented Food Idli

Present study is intended to assess the probiotic properties of Bacillus spp. isolated from idli batter, a traditional fermented food of Southern India and Sri Lanka. A total of 32 isolates were screened for potential pathogenic behaviour through haemolysis assay, DNase activity and antibiotics sensitivity. Two of the isolates were found to be potentially safe and identified as Bacillus spp. These strains were characterized for in vitro probiotic attributes and antioxidant activity. Both the strains showed strong acid and bile tolerance, transit tolerance, lysozyme tolerance, cell surface hydrophobicity, auto-aggregation, co-aggregation, biofilm formation potential and adhesion to human colon adenocarcinoma (HT 29) cell line demonstrating potential probiotic ability. These strains also exhibited considerable cholesterol binding, thermostability, β-galactosidase production, proteolytic, amylolytic and lipolytic activity. Cell-free supernatant inhibited the biofilm formation by Pseudomonas aeruginosa (KT266804) to 90%. Intact cells showed significant DPPH (41%), hydroxyl (31%), radical scavenging activity and lipid peroxidation inhibition (20.38%), while cell-free extracts exhibited significant superoxide anion radical scavenging activity (16.25%). Results revealed that isolates could be potential probiotic candidate after further assessment of in vivo probiotic properties and safety evaluation and could be utilised as starter cultures in functional foods.

     

Production of antibacterial substances by bifidobacterial isolates from infant stool active against Listeria monocytogenes

AIMS: This study aimed to characterize new isolates of human bifidobacteria, evaluate some of their probiotic potential and to screen these isolates for their effectiveness at inhibiting Listeria monocytogenes in vitro. METHODS AND RESULTS: Thirty-four Bifidobacterium isolates from infant faeces were identified by fructose-6-phosphate phosphoketolase and PCR. Six isolates, coded RBL67, RBL68, RBL69, RBL70, RBL85 and RBL86, showed higher antagonistic activity against L. monocytogenes. Neutralized culture supernatants of these strains did not inhibit L. monocytogenes when tested by agar diffusion method. However, the concentration of supernatant by speed-vac resulted in the formation of an inhibitory effect with supernatants from strains RBL67, RBL68 and RBL70. This effect was shown to be related to heat-stable proteinaceous compound(s) which were resistant to heating at 100 degrees C for 5 min but not to pronase-E, proteinase-K or trypsin. The extraction of the inhibitory compounds by methanol-acetone extraction procedure indicated that four strains (RBL67, RBL68, RBL69 and RBL70) were mostly soluble in acetone. However, strain RBL85 produced inhibitory substances that were soluble in methanol. CONCLUSION: Infant bifidobacterial isolates produce heat-stable proteinaceous compounds active against L. monocytogenes. SIGNIFICANCE AND IMPACT OF THE STUDY: Production of antibacterial substances by bifidobacteria would improve intestinal bacterial ecology and inhibit intestinal pathogens.     

Characterization of Bacillus probiotics available for human use

Bacillus species (Bacillus cereus, Bacillus clausii, Bacillus pumilus) carried in five commercial probiotic products consisting of bacterial spores were characterized for potential attributes (colonization, immunostimulation, and antimicrobial activity) that could account for their claimed probiotic properties. Three B. cereus strains were shown to persist in the mouse gastrointestinal tract for up to 18 days postadministration, demonstrating that these organisms have some ability to colonize. Spores of one B. cereus strain were extremely sensitive to simulated gastric conditions and simulated intestinal fluids. Spores of all strains were immunogenic when they were given orally to mice, but the B. pumilus strain was found to generate particularly high anti-spore immunoglobulin G titers. Spores of B. pumilus and of a laboratory strain of B. subtilis were found to induce the proinflammatory cytokine interleukin-6 in a cultured macrophage cell line, and in vivo, spores of B. pumilus and B. subtilis induced the proinflammatory cytokine tumor necrosis factor alpha and the Th1 cytokine gamma interferon. The B. pumilus strain and one B. cereus strain (B. cereus var. vietnami) were found to produce a bacteriocin-like activity against other Bacillus species. The results that provided evidence of colonization, immunostimulation, and antimicrobial activity support the hypothesis that the organisms have a potential probiotic effect. However, the three B. cereus strains were also found to produce the Hbl and Nhe enterotoxins, which makes them unsafe for human use.     

Detection and distribution of probiotic Escherichia coli Nissle 1917 clones in swine herds in Germany

AIMS: To verify the presence of Escherichia coli Nissle 1917 as a natural isolate in swine and to characterize in vitro probiotic properties as well as in vivo persistence in a feeding experiment. METHODS AND RESULTS: During studies on the intestinal microflora of pigs, we isolated E. coli Nissle 1917 sporadically from a pig population over a period of 1 year. The identity of the isolates as E. coli Nissle 1917 was verified by serotyping, Nissle-specific PCR, macrorestriction analysis (pulsed field gel electrophoresis) and the determination of in vitro probiotic properties in invasion and adhesion assays using a porcine intestinal epithelial cell line. Both the E. coli isolates and the E. coli Nissle 1917 strain showed strong reductions in adhesion of porcine enteropathogenic E. coli and invasion of Salmonella typhimurium with epithelial cells in vitro, with a probiotic effect. Screening of five epidemiologically unlinked swine farms and two wild boar groups showed one farm positive for E. coli Nissle 1917. A feeding experiment with four piglets showed viable E. coli Nissle 1917 in the intestine of three animals. CONCLUSIONS: The results of this study suggest that the E. coli Nissle 1917 strain is already partially established in swine herds, but the colonization of individual animals is variable. SIGNIFICANCE AND IMPACT OF THE STUDY: We report natural, long-term colonization and transmission of the probiotic E. coli Nissle 1917 strain in a swine herd, characterized individual persistence and colonization properties in swine and established an in vitro porcine intestinal epithelial cell model of probiotic action. The results of this study would have implications in the use of this strain as a probiotic in swine and contribute to a better understanding of the individual nature of intestinal bacterial persistence and establishment.