A Review on Some Chemical Engineering and Microbiological Aspects Considered in the Production of Highly Concentrated Probiotic Cultures and Bacteriocins by Lactococci and Lactobacilli

The main purpose of this work is to give an overview on the chemical engineering aspects related with the production of probiotic cultures and bacteriocins. Firstly, some evidence of the potential of different Lactococci and Lactobacilli strains for prevention or treatment of different human diseases, or as growth promoters in farm animals is given. In addition, examples of different commercial dietary supplements containing probiotic lactoccocci and lactobacilli in combination or not with other probiotic bacteria are presented. Secondly, the main factors (cultivation method, culture media, and media composition) affecting the production of bacteriocins by lactoccocci and lactobacilli strains are showed. Examples of modelling procedures developed to describe the production of higher concentrations of biomass and bacteriocins in fed-batch cultures subjected or not to successive alkalizations are summarized. Finally, the corresponding mass balance equations performed in the latter cultures are presented to calculate the volumes of feeding substrates and the concentrations of nutrients (lactose, glucose, proteins, nitrogen, and phosphorous) added to the fermentation medium, as well as the concentrations of biomass and products in the fermentor just after each feeding.

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title: Probiotic agents and infectious diseases: a modern perspective on a traditional therapy.
abstract: There is an increasing scientific and commercial interest in the use of beneficial microorganisms, or "probiotics," for the prevention and treatment of disease. The microorganisms most frequently used as probiotic agents are lactic-acid bacteria such as Lactobacillus rhamnosus GG (LGG), which has been extensively studied in recent literature. Multiple mechanisms of action have been postulated, including lactose digestion, production of antimicrobial agents, competition for space or nutrients, and immunomodulation. We have reviewed recent studies of probiotics for the treatment and control of infectious diseases. Studies of pediatric diarrhea show substantial evidence of clinical benefits from probiotic therapy in patients with viral gastroenteritis, and data on LGG treatment for Clostridium difficile diarrhea appear promising. However, data to support use of probiotics for prevention of traveler's diarrhea are more limited. New research suggests potential applications in vaccine development and prevention of sexually transmitted diseases. Further studies are needed to take full advantage of this traditional medical approach and to apply it to the infectious diseases of the new millennium.

title: Production of four potentially probiotic lactic acid bacteria and their evaluation as feed additives for weaned piglets
abstract: The present study was conducted to study the production and evaluation of potentially probiotic additives containing both live lactic acid bacteria (Pediococcus acidilactici NRRL B-5627, Lactococcus lactis subsp. lactis CECT 539, Lactobacillus casei subsp. casei CECT 4043 and Enterococcus faecium CECT 410) and antimicrobial metabolites with could be used as a replacement for antibiotics in weanling pig diets. The gastrointestinal transit tolerance of the four bacteria was determined by exposing washed cell suspensions at 30 °C to acidic conditions (pH 1.0, 2.0, 3.0, 4.0 and 5.0), to a simulated gastric juice (pH 2.0) containing pepsin (3 g/L) and sodium chloride (5 g/L), and to a simulated small intestinal juice (pH 8.0) containing pancreatin (1 g/L) and sodium chloride (5 g/L), mimicking the gastrointestinal environment. These studies showed that the four strains are capable of surviving the passage through the gastrointestinal conditions. Therefore, the production of biomass and antimicrobial products by these bacteria was performed in whey using a fed-batch fermentation technique that provided enhanced productions compared to those obtained in batch fermentations. The obtained fed-batch cultures preserved at −20 °C with skim milk showed a good viability after 3 months of storage. The four cultures exhibited low losses of viability in the piglet feed during their storage at room temperature for 8 days. These results offered the possibility of using the piglet feed as a vehicle to administer the four probiotic bacteria. The effects of the supplementation of separate potentially probiotic cultures and an antibiotic (colistin sulfate) to piglet diets on body weight gain, feed intake, feed efficiency and on the faecal coliform counts of weaned piglets were also studied. Although the best results were obtained in the groups receiving the antibiotic, a significant increase in body weight gain and final body weight was obtained in the groups fed diets supplemented with lactic acid bacteria as compared with the non-treated (controls) groups (P=0.05). The changes in the total coliform population in the control groups over time were not significant (P

title: Performance and intestinal coliform counts in weaned piglets fed a probiotic culture (Lactobacillus casei subsp. casei CECT 4043) or an antibiotic.
abstract: The production of biomass and antibacterial extracellular products by Lactobacillus casei subsp. casei CECT 4043 was followed in both batch and in realkalized fed-batch cultures. Enhanced concentrations of biomass and antibacterial extracellular products were obtained with the use of the latter fermentation technique in comparison with the batch mode. The culture obtained by fed-batch fermentation was mixed with skim milk and used to prepare a probiotic feed for weaned piglets. To test the effect of the potentially probiotic culture of L. casei on body weight gain, feed intake, feed conversion efficiency, and on fecal coliform counts of piglets, two groups of animals received either feed supplemented with the probiotic preparation or avilamycin for 28 days. The control group was fed nonsupplemented feed. At the end of the administration period (day 28), the groups receiving probiotic and avilamycin exhibited the highest average body weight gain values, although the mean feed intake and feed conversion efficiency values were not different among the groups (P > 0.05). For the entire experimental period (42 days), the control group exhibited the lowest feed intake value, the probiotic group exhibited the highest feed conversion efficiency value, and the antibiotic group exhibited the highest body weight gain (P 0.05). Fecal coliform values decreased (although not significantly) by day 28 in the three groups. However, the mean counts returned to pretreatment levels by day 42 in all groups.

title: Novel perspectives in probiotic treatment: the efficacy and unveiled mechanisms of the physiological functions
abstract: Probiotics are defined as “live microorganisms which confer a health benefit on the host” when administered in adequate amounts, and have potential effects for maintaining intestinal development, nutrition, and treating intestinal inflammations, functional disorders, and other extra-intestinal diseases. Although the benefits of probiotics for human health were first noted over 100 years ago, the analysis of probiotic functions began in earnest only 20 years ago. Probiotics, such as some strains of Lactobacillus, Bifidobacterium, Escherichia coli, and Bacillus subtilis, inhibit the growth of pathogenic bacteria, induce competitive effects for the adherent of pathogenic bacteria and their toxins to intestinal epithelia, induce cytoprotective heat shock proteins, enhance the intestinal barrier function, and modulate the host immune responses. The crosstalk between the host and the probiotics appears to be mediated by bacteria-derived effectors, which can be sensed with multiple systems, including the Toll-like receptors and cell membrane transporters. Future analyses will identify more probiotic-derived effectors, the recognition mechanisms of these effectors, and the subsequent changes of the intestinal epithelia and immune cells for each probiotic treatment. For clinical use, a procedure that objectively evaluates the ability of each probiotic effect will help establish a standard for choosing the most valuable strain and its proper dose for each individual patient.

title: Relative Ability of Orally Administered Lactobacillus murinus To Predominate and Persist in the Porcine Gastrointestinal Tract
abstract: Five porcine-derived Lactobacillus or Pediococcus isolates administered to pigs (n = 4), either singly or as a combination at ∼1010 CFU per day varied with respect to intestinal survival and persistence. Two Lactobacillus murinus strains survived best and were excreted at ∼107 to 108 CFU/g of feces. In contrast, Pediococcus pentosaceus DPC6006 had the lowest fecal count at ∼105 CFU/g and was excreted at a significantly lower level than both L. murinus strains. Fecal L. murinus DPC6003 counts were also significantly higher than both Lactobacillus salivarius DPC6005 and Lactobacillus pentosus DPC6004 (∼106 CFU/g). The L. murinus strains persisted for at least 9 days postadministration in both the feces and the cecum. Animals fed a combination of all five strains excreted ∼107 CFU of the administered strains/g, with L. murinus predominating, as determined by randomly amplified polymorphic DNA PCR. Postadministration, variation was observed between animals fed the strain combination, but in general, L. murinus DPC6002 and DPC6003 and L. pentosus DPC6004 predominated in the feces and the cecum while P. pentosaceus DPC6006 was detected only in the cecum. Fifteen days after the start of culture administration, mean fecal Enterobacteriaceae counts were significantly lower in some of the treatment groups. In addition, when mean preadministration counts were compared with those obtained after 21 days of culture administration, Enterobacteriaceae counts were reduced by ∼87 to 98% in pigs fed L. salivarius DPC6005, P. pentosaceus DPC6006, L. pentosus DPC6004, and the culture mix. In conclusion, the porcine intestinal isolates have potential as probiotic feed additives for pigs, with differences in strain performance highlighting the advantages of using culture combinations.

title: Bacillus clausii Probiotic Strains: Antimicrobial and Immunomodulatory Activities
abstract: The clinical benefits observed with probiotic use are mainly attributed to the antimicrobial substances produced by probiotic strains and to their immunomodulatory effects. Currently, the best-documented probiotic bacteria used in human therapy are lactic acid bacteria. In contrast, studies aiming to characterize the mechanisms responsible for the probiotic beneficial effects of Bacillus are rare. The current work seeks to contribute to such characterization by evaluating the antimicrobial and immunomodulatory activities of probiotic B. clausii strains. B. clausii strains release antimicrobial substances in the medium. Moreover, the release of these antimicrobial substances was observed during stationary growth phase and coincided with sporulation. These substances were active against Gram-positive bacteria, in particular against Staphylococcus aureus, Enterococcus faecium, and Clostridium difficile. The antimicrobial activity was resistant to subtilisin, proteinase K, and chymotrypsin treatment, whereas it was sensitive to pronase treatment. The evaluation of the immunomodulatory properties of probiotic B. clausii strains was performed in vitro on Swiss and C57 Bl/6j murine cells. The authors demonstrate that these strains, in their vegetative forms, are able to induce NOS II synthetase activity, IFN-γ production, and CD4+ T-cell proliferation.

title: Prophylactic Lactobacillus GGReduces Antibiotic-Associated Diarrhea in Children With Respiratory Infections: A Randomized Study
abstract: Antimicrobial treatment may disturb the colonization resistance of gastrointestinal microflora, which may induce clinical symptoms, most commonly diarrhea. The severity of antibiotic-associated diarrhea may range from a brief, self-limiting disease to devastating diarrhea with electrolyte disturbances, dehydration, crampy abdominal pain, pseudomembranous colitis, toxic megacolon, or even death. The incidence of diarrhea in children receiving a single antimicrobial treatment is unclear. In addition to more critical use of antimicrobials, adjunctive preventive measures to antibiotic-associated diarrhea are needed. The objective of this study was to evaluate the incidence of diarrhea after antimicrobial treatment in children with no history of antimicrobial use during the previous 3 months. Another aim of this study was to assess the preventive potential of Lactobacillus rhamnosus GG (Lactobacillus GG; American Type Culture Collection 53103), a probiotic strain with a documented safety record and a therapeutic effect in viral gastroenteritis on antibiotic-associated diarrhea.  Oral antimicrobial agents were prescribed for the treatment of acute respiratory infections at the clinics of the Health Care Center of the City of Tampere or Tampere University Hospital, Finland, to 167 patients who were invited to participate in the study. Of the patients, 48 were lost to follow-up; therefore, the final study population consisted of 119 children from 2 weeks to 12.8 years of age (mean: 4.5 years). All study subjects met the inclusion criteria: they had not received any antimicrobial medication during the previous 3 months, they did not suffer from gastrointestinal disorders, and they did not need intravenous antimicrobial treatment. The patients were randomized to receive placebo or 2 × 1010 colony-forming units of Lactobacillus GG in capsules given twice daily during the antimicrobial treatment. Lactobacillus GG and placebo capsules were indistinguishable in appearance and taste. The parents kept a daily symptom diary and recorded stool frequency and consistency at home for 3 months. Diarrhea was defined as at least three watery or loose stools per day for a minimum of 2 consecutive days. In the case of diarrhea, viral (adenovirus, rotavirus, calicivirus and astrovirus) and bacterial (Salmonella, Shigella, Yersinia, Campylobacter, Clostridium difficile, Staphylococcus aureus, and yeasts) analyses were studied in fecal samples. The metabolic activity of the gut microflora was assessed by analysis of fecal urease, β-glucosidase, and β-glucuronidase activities. The primary outcome measure was diarrhea during the first 2 weeks after the beginning of the antimicrobial treatment, because this period most likely reflects the effects of antimicrobial use. Secondary outcome measures were the activities of fecal urease, β-glucuronidase, and β-glucosidase.  On the entire follow-up, 80% of any gastrointestinal symptoms were reported during the first 2 weeks after the beginning of the antimicrobial treatment. The incidence of diarrhea was 5% in the Lactobacillus GG group and 16% in the placebo group within 2 weeks of antimicrobial therapy (χ2 = 3.82). The treatment effect (95% confidence interval) of Lactobacillus GG was −11% (−21%–0%). In diarrheal episodes, the viral and bacterial analyses were positive for Clostridium difficile in 2 cases and for Norwalk-like calicivirus in 3 cases. The age of the patients with diarrhea was between 3 months and 5 years in 75% of cases in both groups. The severity of diarrhea was comparable in the study groups, as evidenced by similar stool frequency (mean: 5 per day; range: 3–6) and the duration of diarrhea (mean: 4 days; range: 2–8). The activities of fecal urease and β-glucuronidase, but not β-glucosidase, changed significantly after the beginning of the antimicrobial treatment in the Lactobacillus GG group and in the placebo group alike. The decrease in urease and β-glucuronidase activities was reversible in patients with no diarrhea, but in patients with diarrhea, the modifications in gut microflora were more profound and prolonged. The activities of the three enzymes were normalized within 3 weeks, evidenced by stable enzyme activities in samples collected 3 weeks, 1 month, and 3 months after the beginning of the antimicrobial treatment, compared with those obtained before treatment.  In the present study, after a single antimicrobial treatment, the incidence of diarrhea was 16%. The higher incidence of antibiotic-associated diarrhea in previous reports may be attributable to a recent antimicrobial therapy that disturbs intestinal flora and exposes to complications. Also, in the present study, changes in the metabolic activity of the intestinal flora were observed, evidenced by a transient decline in fecal enzyme activities. Different probiotic preparations, including lactobacilli, are recommended frequently to prevent antibiotic-associated diarrhea. Although probiotics have been shown to be efficient in the prevention and the treatment of viral gastroenteritis, their usefulness during antimicrobial therapy in children has not been elucidated before. We observed that the administration of Lactobacillus GG to children receiving antimicrobial therapy for respiratory infection reduced the incidence of antibiotic-associated diarrhea to one third. The beneficial effect may be mediated by a number of functions of probiotics, ie, production of antimicrobial substances, local competition of adhesion receptors and nutrients, and stimulation of intestinal antigen specific and nonspecific immune responses.  A probiotic strain, Lactobacillus GG, is effective in the prevention of diarrhea in children receiving antimicrobial treatment to respiratory infections.

title: Silver-coated dressing acticoat caused raised liver enzymes and argyria-like symptoms in burn patient.
abstract: Treatment of acute burn wounds with silver sulfadiazine (SSD) has raised concern about potential silver toxicity. Numerous adverse reactions and side effects have been reported and an increasing resistance to SSD, especially in Pseudomonas strains, have motivated researchers to search for an alternative wound dressing.



Recently, a silver-coated wound dressing Acticoat (Smith & Nephew, Inc.) has become available for use in burn patients. It is a three-ply dressing, consisting of an inner rayon/polyester absorptive core between two layers of silver-coated, high-density polyethylene mesh. In a moist environment, the nanocrystals of silver are released and improve the microbial control in the wound.



After 1 week of local treatment with Acticoat in a young, previously healthy 17-year-old boy with 30% mixed depth burns, hepatotoxicity and argyria-like symptoms, a grayish discoloration of the patient's face, appeared. The silver levels in plasma (107 microg/kg) and urine (28 microg/kg) were clearly elevated, as well as the liver enzymes. As soon as the local application of Acticoat was aborted, the clinical symptoms and liver enzymes returned to the normal values.



This is the first report on silver toxicity in a patient with 30% burns who received Acticoat for local treatment. Due to substantial experiences with adverse SSD reactions and side effects, it is appropriate to keep the possibility of a toxic silver effect in burn patients treated with Acticoat silver-coated wound dressing in mind. The silver levels in plasma and/or urine should be monitored.

title: The assumed assimilation of cholesterol by Lactobacilli and Bifidobacterium bifidum is due to their bile salt-deconjugating activity
abstract: To study the mechanism of the propsed assimilation of cholesterol, we cultured various strains of Lactobacillus acidophilus and a Bifidobacterium sp. in the presence of cholesterol and oxgall. During culturing, both cholesterol and bile salts were precipitated. Because of bacterial bile salt deconjugation, no conjugated bile salts were observed in either the culture fluids or the pellets. During incubation, the cell count and optical density decreased. The degree of precipitation of bile salts and of cholesterol was dependent on the culture conditions. If L. acidophilus RP32 was cultured under acidifying conditions, the degree of precipitation of deconjugated bile salts was higher than if the pH was maintained at 6.0. Under acidifying conditions, cholesterol was coprecipitated with the bile salts, whereas in pH-controlled cultures, no coprecipitation of cholesterol was observed. From control experiments with different mixtures of bile salts, it appeared that coprecipitation of cholesterol during culturing was a result of formation of deconjugated bile salts, which have a decreased solubility at pH values lower than 6.0. It is concluded that the removal of cholesterol from the culture medium by L. acidophilus RP32 and other species is not due to bacterial uptake of cholesterol, but results from bacterial bile salt-deconjugating activity.

title: In vitro selection criteria for probiotic bacteria of human origin: correlation with in vivo findings.
abstract: The enteric flora comprises approximately 95% of the total number of cells in the human body and can elicit immune responses while protecting against microbial pathogens. However, the resident bacterial flora of the gastrointestinal tract may also be implicated in the pathogenesis of diseases such as inflammatory bowel disease (ulcerative colitis and Crohn disease). The objectives of the Probiotic Research Group based at University College Cork were to isolate and identify lactic acid bacteria exhibiting beneficial probiotic traits, such as bile tolerance in the absence of deconjugation activity, acid resistance, adherence to host epithelial tissue, and in vitro antagonism of pathogenic microorganisms or those suspected of promoting inflammation. To isolate potentially effective probiotic bacteria, we screened the microbial population adhering to surgically resected segments of the gastrointestinal tract (the environment in which they may subsequently be reintroduced and required to function). In total, 1500 bacterial strains from resected human terminal ilea were assessed. From among these organisms, Lactobacillus salivarius subsp. salivarius strain UCC118 was selected for further study. In mouse feeding trials, milk-borne L. salivarius strain UCC118 could successfully colonize the murine gastrointestinal tract. A human feeding study conducted in 80 healthy volunteers showed that yogurt can be used as a vehicle for delivery of strain UCC118 to the human gastrointestinal tract with considerable efficacy in influencing gut flora and colonization. In summary, we developed criteria for in vitro selection of probiotic bacteria that may reflect certain in vivo effects on the host such as modulation of gastrointestinal tract microflora.

title: Probiotics: potential pharmaceutical applications.
abstract: Realisation of the importance of human gut microbiota in health restoration and maintenance has kindled an interest in probiotics. Probiotics are defined as the microbial food supplements, which beneficially affect the host by improving its intestinal microbial balance. Probiotics are the health enhancing functional food ingredients used therapeutically to prevent diarrhea, improve lactose tolerance and modulate immunity. They may also have potential to prevent cancer and lower serum cholesterol levels. Lactobacillus, Bifidobacterium and several other microbial species are perceived to exert such effects by changing the composition of the gut microbiota. However, it is important that exogenously administered bacteria reach and establish themselves in the large intestine in an intact form. The use of non-digestible oligosaccharides ('prebiotics') can fortify intestinal microflora and stimulate their growth. The present review encompasses information regarding the probiotics and their proposed uses. It addresses the concepts of prebiotics and synbiotics, the application of genetic engineering to produce newer probiotics. Finally, the list of commercially available products are reviewed with discussion of questions regarding the reliability, utility and the safety of these products.

title: Characterization of indigenous vaginal lactobacilli from healthy women as probiotic candidates.
abstract: The probiotic relevant characteristics of 45 strains of vaginal Lactobacillus isolated from healthy women were analyzed. Of these, 21 strains were classified as L. crispatus, 17 as L. jensenii, six as L. gasseri, and one as L. plantarum. The rate of acidification varied significantly between the strains as did their ability to form biofilms. None used glycogen as a fermentable carbohydrate. H2O2 generation was common, especially among L. jensenii isolates (88%). No bacteriocinogenic strains were detected. Most strains harbored plasmids (from 1 to 7) of various sizes, those in excess of 50 kb being frequent. One of these plasmids was found to be promiscuous since it hybridized with extrachromosomal bands of 15 isolates. All strains were resistant to metronidazole, ciprofloxacin, gentamicin, clindamycin, trimethoprim, and sulfametoxazole and susceptible to a series of beta-lactams, erythromycin, tetracycline, and benzalkonium chloride. Almost half of the strains were highly resistant to nonoxinol 9, which is commonly used as a spermicide. Based on these analyses, strains of all three common species are proposed as new probiotic candidates.

title: A randomized trial of Lactobacillus acidophilus BG2FO4 to treat lactose intolerance.
abstract: Lactose intolerance is the most common disorder of intestinal carbohydrate digestion. Lactobacillus acidophilus BG2FO4 is a strain of lactobacilli with properties of marked intestinal adherence and high beta-galactosidase activity.



This study was designed to determine whether oral feeding of Lactobacillus acidophilus BG2FO4 leads to a lactose-tolerant state.



We studied 42 subjects with self-reported lactose intolerance and performed breath-hydrogen tests to determine whether they were lactose maldigesters. Subjects with established lactose maldigestion (n = 24) were invited to be randomly assigned to an omeprazole-treated (hypochlorhydric) group or a non-omeprazole-treated group, but 6 subjects chose not to participate. All randomly assigned subjects (n = 18) ingested Lactobacillus acidophilus BG2FO4 twice per day for 7 d and stool samples were collected. Breath-hydrogen tests were performed and symptom scores were recorded at baseline and after lactobacilli ingestion.



Lactose maldigestion was established in 24 of 42 subjects (57%) with self-reported lactose intolerance. In 18 lactose-maldigesting subjects, overall hydrogen production and symptom scores after ingestion of Lactobacillus acidophilus BG2FO4 were not significantly different from baseline values. Live Lactobacillus acidophilus BG2FO4 was recovered in stool samples from 7 subjects.



Lactose intolerance is overreported in subjects with gastrointestinal symptoms after lactose ingestion. Treatment of lactose-maldigesting subjects with and without hypochlorhydria with Lactobacillus acidophilus BG2FO4 for 7 d failed to change breath-hydrogen excretion significantly after lactose ingestion.

title: Lactobacillus paracasei Strain ST11 Has No Effect on Rotavirus but Ameliorates the Outcome of Nonrotavirus Diarrhea in Children From Bangladesh
abstract: Previous studies have shown that selected strains of lactobacilli that are administered orally result in a modest reduction of diarrhea duration. However, duration alone is not considered optimal for therapeutic evaluation of any agent in diarrhea.  To examine the effect of a new probiotic, Lactobacillus paracasei strain ST11 (ST11), in acute childhood diarrhea by using evaluation criteria recommended by the World Health Organization.  In a randomized, double-blind, placebo-controlled clinical trial, 230 male infants and young children, 4 to 24 months of age, presenting with diarrhea of <2 days' duration were admitted to the metabolic research ward of the International Centre for Diarrheal Disease Research, Bangladesh, and fed 1010 colony-forming units of lyophilized ST11 or placebo daily for 5 days. Stool output and frequency, oral rehydration solution intake, and excretion of rotavirus were monitored daily.  No effect of ST11 treatment on severe rotavirus diarrhea was observed. However, the probiotic treatment did significantly reduce cumulative stool output (225 ± 218 vs 381 ± 240 mL/kg), stool frequency (27.9 ± 17 vs 42.5 ± 26), and oral rehydration solution intake (180 ± 207 vs 331 ± 236 mL/kg) in children with less-severe nonrotavirus diarrhea compared with those receiving placebo treatment. A significantly higher proportion of nonrotavirus children receiving ST11 had their diarrhea resolve within 6 days of therapy (ST11 versus placebo: 76% vs 49%).  ST11 has a clinically significant benefit in the management of children with nonrotavirus-induced diarrhea, but it is ineffective in those with rotavirus diarrhea.

title: Probiotics and prebiotic galacto-oligosaccharides in the prevention of allergic diseases: a randomized, double-blind, placebo-controlled trial.
abstract: The increase in allergic diseases is attributed to a relative lack of microbial stimulation of the infantile gut immune system. Probiotics, live health-promoting microbes, might offer such stimulation.



We studied the effect of a mixture of 4 probiotic bacterial strains along with prebiotic galacto-oligosaccharides in preventing allergic diseases.



We randomized 1223 pregnant women carrying high-risk children to use a probiotic preparation or a placebo for 2 to 4 weeks before delivery. Their infants received the same probiotics plus galacto-oligosaccharides (n = 461) or a placebo (n = 464) for 6 months. At 2 years, we evaluated the cumulative incidence of allergic diseases (food allergy, eczema, asthma, and allergic rhinitis) and IgE sensitization (positive skin prick test response or serum antigen-specific IgE level >0.7 kU/L). Fecal bacteria were analyzed during treatment and at age 2 years.



Probiotic treatment compared with placebo showed no effect on the cumulative incidence of allergic diseases but tended to reduce IgE-associated (atopic) diseases (odds ratio [OR], 0.71; 95% CI, 0.50-1.00; P = .052). Probiotic treatment reduced eczema (OR, 0.74; 95% CI, 0.55-0.98; P = .035) and atopic eczema (OR, 0.66; 95% CI, 0.46-0.95; P = .025). Lactobacilli and bifidobacteria more frequently (P < .001) colonized the guts of supplemented infants.



Probiotic treatment showed no effect on the incidence of all allergic diseases by age 2 years but significantly prevented eczema and especially atopic eczema. The results suggest an inverse association between atopic diseases and colonization of the gut by probiotics.



The prevention of atopic eczema in high-risk infants is possible by modulating the infant's gut microbiota with probiotics and prebiotics.

title: Immunomodulatory effects of probiotics in the intestinal tract.
abstract: The intestinal microbiota is the largest source of microbial stimulation that exerts both harmful and beneficial effects on human health. The interaction between probiotic and enterocytes is the initiating event in immunomodulation and merits particular attention. The effects of probiotic is strain dependent and for each new probiotic strain, profiles of cytokines secreted by lymphocytes, enterocytes or dendritic cells that come in contact with the strain should be systematically established. To evaluate the effects of probiotics on the immune system, models that mimic the mucosa, and thus the physiological reality, should be preferred whenever it is possible. Then, the in vitro observed effects should be backed up by properly conducted randomized double bind clinical studies. More detailed studies are needed to determine the precise action mode of probiotics on both mucosal and systemic immunity.

title: Pathogenic relevance of Lactobacillus: a retrospective review of over 200 cases
abstract: Given that Lactobacillus has been reported to be the causative pathogen in many types of infection despite debate regarding the organism’s clinical significance, a literature review was conducted to investigate the treatments and outcomes of Lactobacillus infections reported to date. In this article, the characteristics of over 200 reported cases of Lactobacillus-associated infections are summarized. Lactobacillus was found to be frequently associated with endocarditis and bacteremia. Lactobacillus was also associated with a variety of other infections including, but not limited to, peritonitis, abscesses, and meningitis. The species casei and rhamnosus were the most common. The isolates tended to be most sensitive to erythromycin and clindamycin and most resistant to vancomycin. The species that was most sensitive to vancomycin was acidophilus. The overall mortality rate was nearly 30%. There was a significant association between mortality and polymicrobial infection (P=0.004). In the subset of patients with bacteremia, increased mortality was associated with inadequate treatment (P=0.001) and polymicrobial bacteremia (P=0.044).

title: Lactobacillus: a Review
abstract: Lactobacilli colonize the gastrointestinal and urinary tracts of humans, making them an integral part of the microbial flora. However, in certain circumstances, they can cause disease. Although lactobacilli are often misidentified as streptococci, it is highly likely that these infections will be attributed to them due to current improvements in laboratory techniques. A recent resurgence in interesting natural products has caused an increased focus on the use of probiotics. Many of these probiotic formulations contain Lactobacillus species. Although there have been reports of lactobacilli providing benefit in infectious diarrhea, the use of these probiotic products in immunosuppressed or critically ill patients is not advised, since these populations are at increased risk of developing infections due to lactobacilli.

title: Lactobacillus endocarditis caused by a probiotic organism.
abstract: originated (data not shown), one would perhaps expect to see mutations being lost or gained within the genes potentially affecting quinolone susceptibility. The stability of mutations in genes known to be associated with fluoroquinolone resistance, and the stable MIC values in clonally related MRSA isolates over time, have important implications for quinolone resistance in S. aureus. From this study of clinical MRSA isolates, it seems likely that once acquired, mutations and the decreased susceptibility to quinolones that they confer are highly stable, suggesting that the high prevalence of fluoroquinolone resistance that we are experiencing today in some parts of the world will remain.

title: Lactobacillus bacteremia during a rapid increase in probiotic use of Lactobacillus rhamnosus GG in Finland.
abstract: Lactobacilli supposedly have low pathogenicity; they are seldom detected in blood culture. Lactobacillus rhamnosus GG, which originates indigenously in the human intestine, became available for use as a probiotic in 1990 in Finland. We evaluated the possible effects of the increased probiotic use of L. rhamnosus GG on the occurrence of bacteremia due to lactobacilli. Lactobacilli were isolated in 0.02% of all blood cultures and 0.2% of all blood cultures with positive results in Helsinki University Central Hospital and in Finland as a whole, and no trends were seen that suggested an increase in Lactobacillus bacteremia. The average incidence was 0.3 cases/100,000 inhabitants/year in 1995-2000 in Finland. Identification to the species level was done for 66 cases of Lactobacillus bacteremia, and 48 isolates were confirmed to be Lactobacillus strains. Twenty-six of these strains were L. rhamnosus, and 11 isolates were identical to L. rhamnosus GG. The results indicate that increased probiotic use of L. rhamnosus GG has not led to an increase in Lactobacillus bacteremia.

title: Probiotics in the Treatment and Prevention of Acute Infectious Diarrhea in Infants and Children: A Systematic Review of Published Randomized, Double-Blind, Placebo-Controlled Trials
abstract: This review was designed to assess the evidence from randomized controlled trials on effects of probiotics in the treatment and prevention of acute infectious diarrhea in infants and children.  A systematic review of published, randomized, double-blind, placebo-controlled trials on probiotics in the treatment or prevention of acute diarrhea defined as >3 loose or watery stools per 24 hours in infants and children.  The use of probiotics as compared with placebo was associated with a significantly reduced risk of diarrhea lasting >3 days. The pooled estimate risk was 0.43 (95% CI, 0.34–0.53) with a fixed-effect model, and remained significant in a random-effect model (0.40; 95% CI, 0.28–0.57). Only Lactobacillus GG showed a consistent effect. Probiotics significantly reduced the duration of diarrhea when compared with placebo, particularly in rotaviral gastroenteritis—the pooled, weighted, mean difference (WMD) assuming the random-effect model was −20.1 hours (95% CI, −26.1 to −14.2) and −24.8 (95% CI, −31.8 to −17.9) respectively. A meta-analysis of the prevention studies was not feasible because of significant clinical and statistical heterogeneity.  There is evidence of a clinically significant benefit of probiotics in the treatment of acute infectious diarrhea in infants and children, particularly in rotaviral gastroenteritis. Lactobacillus GG showed the most consistent effect, although other probiotic strains may also be effective. Further research is needed. Clinical and statistical heterogeneity of the prophylactic interventions preclude drawing firm conclusions about the efficacy of probiotics in preventing acute gastroenteritis.

title: A randomized controlled trial of a probiotic, VSL#3, on gut transit and symptoms in diarrhoea‐predominant irritable bowel syndrome
abstract: To investigate the effects of a probiotic formulation, VSL#3, on gastrointestinal transit and symptoms of patients with Rome II irritable bowel syndrome with predominant diarrhoea.

title: Lactobacillus acidophilus as a dietary adjunct for milk to aid lactose digestion in humans.
abstract: The effect of Lactobacillus acidophilus on lactose utilization of humans was determined by our comparing the amount of hydrogen excreted in their breath after consuming milk containing either 0, 2.5 X 10(6), 2.5 X 10(7), or 2.5 X 10(8) Lactobacillus acidophilus/ml daily for 6 days. Consumption of milk without cells of Lactobacillus acidophilus for 1 wk did not affect lactose utilization. Milk containing either 2.5 X 10(6) or 2.5 X 10(8) Lactobacillus acidophilus/ml improved lactose utilization. However, consumption of milk containing an intermediate 2.5 X 10(7)/ml did not improve utilization based on comparison of group means before and after consuming the test milk. The lack of a significant effect for the latter group of test subjects was probably due to large increases of excreted hydrogen on day 7 as compared to day 0 by two of the six test subjects. The beneficial effect of Lactobacillus acidophilus was immediate and did not require that milk be consumed daily. Improved digestion of lactose was not due to hydrolysis of the lactose prior to consumption, which indicated that the beneficial effect must have occurred in the digestive tract after consumption of milk containing L. acidophilus.

title: Effect of milks inoculated with Lactobacillus acidophilus or a yogurt starter culture in lactose-maldigesting children.
abstract: Dairy products containing live bacteria that possess lactase activity are used for dietary management of lactose maldigestion. The efficacy of acidophilus milk and the effect of consuming unfermented milk that had been inoculated with yogurt bacteria have not been examined in children. We compared scores for breath H2 excretion and symptoms of 20 lactose-maldigesting children following ingestion of 250 ml of uninoculated milk with two identical milks inoculated with 10(10) cells of Lactobacillus acidophilus or with a commercial yogurt starter culture containing 10(8) cells of Lactobacillus lactis and 10(10) cells of Streptococcus thermophilus. Nine of 10 subjects who were symptomatic following ingestion of uninoculated milk experienced a reduction in symptoms following ingestion of milk inoculated with L. acidophilus, without a decline in H2 excretion. Five of 6 subjects who were symptomatic following uninoculated milk had decreased symptoms and a significant reduction in H2 excretion following milk inoculated with the yogurt culture. For lactose-maldigesting children, milks inoculated with L. acidophilus or with a yogurt culture were associated with decreased symptoms compared with those with uninoculated milk.

title: Probiotic bacteria stimulate
virus–specific neutralizing antibodies
following a booster polio vaccination
abstract: SummaryBackgroundOrally
ingested probiotic bacteria may
modulate the immune response
and increase antibody titers against
enteric infections by bacteria or
viruses. Even though positive effects
of probiotics on respiratory
tract infections have been reported,
overall only few studies
have examined effects on virus infections
concerning organs other
than the gastrointestinal tract.
study to investigate whether and
how probiotics affect the immune
response to a standardized enterovirus
challenge (polio) and infections
not limited to the gastrointestinal
tract in healthy adults.MethodsIn a randomized, controlled
and double–blind study 64
volunteers consumed for 5 weeks
chemically acidified clotted milk
without bacteria or with 1010/serving
Lactobacillus rhamnosus GG or
Lactobacillus acidophilus CRL431
added. In the second week subjects
were vaccinated orally against polio
1, 2 and 3. Polio virus neutralizing
serum activity, the primary parameter,
was determined by the
standard neutralization test
(WHO) before and three times after
vaccination. Polio–specific IgA,
IgG and IgM were detected by
ELISAs.ResultsProbiotics increased
poliovirus neutralizing antibody
titers (NT) and affected the
formation of poliovirus–specific
IgA and IgG in serum. The maximum
increase after immunization
was about 2, 2.2, or 4–fold higher,
respectively, for NT, IgG or, IgA, in
volunteers consuming probiotics
instead of placebo. No consistent
difference was noted between bacterial
strains.ConclusionsProbiotics
induce an immunologic response
that may provide enhanced
systemic protection of cells from
virus infections by increasing production
of virus neutralizing antibodies.

title: Probiotics: a novel approach in the management of food allergy.
abstract: The gastrointestinal microflora is an important constituent of the gut mucosal defense barrier. We have previously shown that a human intestinal floral strain, Lactobacillus GG (ATCC 53103), promotes local antigen-specific immune responses (particularly in the IgA class), prevents permeability defects, and confers controlled antigen absorption.



The aim of this study was to evaluate the clinical and immunologic effects of cow's milk elimination without (n = 14) and with (n = 13) the addition of Lactobacillus GG (5 x 10(8) colony-forming units/gm formula) in an extensively hydrolyzed whey formula in infants with atopic eczema and cow's milk allergy. The second part of the study involved 10 breast-fed infants who had atopic eczema and cow's milk allergy. In this group Lactobacillus GG was given to nursing mothers.



The severity of atopic eczema was assessed by clinical scoring. The concentrations of fecal alpha 1- antitrypsin, tumor necrosis factor-alpha, and eosinophil cationic protein were determined as markers of intestinal inflammation before and after dietary intervention.



The clinical score of atopic dermatitis improved significantly during the 1-month study period in infants treated with the extensively hydrolyzed whey formula fortified with Lactobacillus GG. The concentration of alpha 1-antitrypsin decreased significantly in this group (p = 0.03) but not in the group receiving the whey formula without Lactobacillus GG (p = 0.68). In parallel, the median (lower quartile to upper quartile) concentration of fecal tumor necrosis factor-alpha decreased significantly in this group, from 709 pg/gm (91 to 1131 pg/gm) to 34 pg/gm (19 to 103 pg/gm) (p = 0.003), but not in those receiving the extensively hydrolyzed whey formula only (p = 0.38). The concentration of fecal eosinophil cationic protein remained unaltered during therapy.



These results suggest that probiotic bacteria may promote endogenous barrier mechanisms in patients with atopic dermatitis and food allergy, and by alleviating intestinal inflammation, may act as a useful tool in the treatment of food allergy.

title: Intravaginal instillation of lactobacilli for prevention of recurrent urinary tract infections.
abstract: Urinary tract infections remain a common problem, particularly in the female population. New methods are required to manage recurrent cystitis, and extensive research to date has suggested that restoration of the lactobacilli flora of the urogenital tract may prevent these infections. In this study, five females suffering from recurrent urinary tract infections were treated twice weekly with intravaginal and perineal implantation of Lactobacillus casei GR-1. These organisms colonized the epithelium and prevented the emergence of coliform bacteria in most instances, but did not appear to affect enterococcal colonization. In vitro studies showed that L. casei GR-1 inhibited the growth of the coliforms but did not inhibit enterococci. Each of the five patients had infection-free periods ranging from 4 weeks to 6 months. The treatment was well tolerated, had no side effects, led to an improved well-being, and was preferred to antibiotic treatment by all of the patients. These human studies, albeit of a limited nature, are the first to examine the potential for lactobacillus therapy in the prevention of urinary tract infections. The results show that lactobacilli therapy, using carefully selected organisms to treat patients who are closely followed, may be effective in the prevention of recurrent urinary tract infections.

title: Ingestion of yogurt containing Lactobacillus acidophilus as prophylaxis for candidal vaginitis
abstract: Intrauterk devices awl pelvic ioflammatory disease: An international perspective Farley TMM; Rosenberg MJ; Rowe PJ; Chen J-H; Meirik 0 Special Programme of Research,   Training in Human Reproduction, World Health Organisation, CH 1211 Geneva 27, CHE LANCET 1992 33918196 (785-788) The risk of pelvic inflammatory disease (PID) associated with use of an intrauterine device (IUD) has been an important concern that has dominated decisions on its use throughout the world, especially in the USA. Early research that suggested such an association led to both a dramatic decline in use of the method and its withdrawal from the US market by two manufacturers. However, factors other than use of an IUD are now thought to be major determinants of PID risk. To address these concerns, we reviewed the World Health Organisation’s IUD clinical trial data to explore the incidence and patterns of PID risk with use of an IUD. The overall rate of PID among 22 908 IUD insertions and during 51,399 woman-years of follow-up was 1.6 cases per 1000 woman-years of use. After adjustment for confounding factors, PID risk was more than six times higher during the 20 days after insertion than during later times (unadjusted rates, 9.7 vs. 1.4 per 1000 woman-years, respectively); the risk was low and constant for up to eight years of follow-up. Rates varied according to geographical area (highest in Africa and lowest in China) and were inversely associated with age. PID rates were lower among women who had, IUDs inserted more recently. Our findings indicate that PID among IUD users is most strongly related to the insertion process and to background risk of sexually transmissible disease. PID is an infrequent event beyond the first 20 days after insertion. Because of this increased risk with insertion, IUDs should be left in place up to their maximum lifespan and should not routinely be replaced earlier, provided there are no contraindications to continued use and the woman wishes to continue with the device.

title: Biotherapeutic effects of probiotic bacteria on candidiasis in immunodeficient mice
abstract: Four species of probiotic bacteria were assessed for their capacities to protect athymic bg/bg-nu/nu and euthymic bg/bg-nu/+ mice from mucosal and systemic candidiasis. Each bacterial species and Candida albicans colonized the gastrointestinal tracts of both strains of mice. The presence of probiotic bacteria (Lactobacillus acidophilus, Lactobacillus reuteri, Lactobacillus casei GG, or Bifidobacterium animalis) in the gastrointestinal tracts prolonged the survival of adult and neonatal bg/bg-nu/nu mice compared to that of isogenic mice colonized with C. albicans alone. The incidence of systemic candidiasis in bg/bg-nu/nu mice was significantly reduced by each of the four probiotic bacterial species. The numbers of C. albicans present in the alimentary tracts of euthymic bg/bg-nu/+ mice were significantly reduced by L. casei GG and B. animalis. None of the probiotic bacteria species completely prevented mucosal candidiasis, but B. animalis reduced its incidence and severity. Probiotic bacteria also modulated antibody- and cell-mediated immune responses to C. albicans. The prolonged survival of mice, decreased severity of mucosal and systemic candidiasis, modulation of immune responses, decreased number of C. albicans in the alimentary tract, and reduced numbers of orogastric infections demonstrated not only that probiotic bacteria have biotherapeutic potential for prophylaxis against and therapy of this fungal disease but also that probiotic bacteria protect mice from candidiasis by a variety of immunologic (thymic and extrathymic) and nonimmunologic mechanisms in this model.

title: Lactobacillus farciminis treatment attenuates stress‐induced overexpression of Fos protein in spinal and supraspinal sites after colorectal distension in rats
abstract: Irritable bowel syndrome (IBS), frequently associated with psychological distress, is characterized by hypersensitivity to gut wall distension. Some probiotics are able to alleviate IBS symptoms and reduce visceromotor response to mechanical stimuli in animals. Moreover, we have previously shown that Lactobacillus farciminis treatment abolished the hyperalgesia to colorectal distension (CRD) induced by acute stress. The aims of the present study were to determine whether (i) stress‐induced visceral hyperalgesia modifies the expression of Fos, a marker of general neuronal activation, induced by CRD, (ii) this activation can be modulated by L. farciminis treatment. Female rats were treated by L. farciminis and CRD was performed after partial restraint stress (PRS) or sham‐PRS. The expression of Fos protein was measured by immunohistochemistry. After CRD or PRS, Fos expression was increased in spinal cord section (S1), nucleus tractus solitarius (NTS), paraventricular nucleus (PVN) of the hypothalamus, and in the medial nucleus of the amygdala (MeA). The combination of both stimuli, PRS and CRD, markedly increased this Fos overexpression in the sacral spinal cord section, PVN and MeA, but not in NTS. By contrast, a pretreatment with L. farciminis significantly reduced the number of Fos positive cells in these area. This study shows that PRS enhances Fos protein expression induced by CRD at the spinal and supraspinal levels in rats. Lactobacillus farciminis treatment inhibited this enhancing effect, suggesting that the antinociceptive effect of this probiotic strain results from a decrease of the stress‐induced activation/sensitization of sensory neurons at the spinal and supraspinal level.

title: Lactobacillus and bifidobacterium in irritable bowel syndrome: symptom responses and relationship to cytokine profiles.
abstract: The aim of this study was to compare the response of symptoms and cytokine ratios in irritable bowel syndrome (IBS) with ingestion of probiotic preparations containing a lactobacillus or bifidobacterium strain.



Seventy-seven subjects with IBS were randomized to receive either Lactobacillus salivarius UCC4331 or Bifidobacterium infantis 35624, each in a dose of 1 x 10 10 live bacterial cells in a malted milk drink, or the malted milk drink alone as placebo for 8 weeks. The cardinal symptoms of IBS were recorded on a daily basis and assessed each week. Quality of life assessment, stool microbiologic studies, and blood sampling for estimation of peripheral blood mononuclear cell release of the cytokines interleukin (IL)-10 and IL-12 were performed at the beginning and at the end of the treatment phase.



For all symptoms, with the exception of bowel movement frequency and consistency, those randomized to B infantis 35624 experienced a greater reduction in symptom scores; composite and individual scores for abdominal pain/discomfort, bloating/distention, and bowel movement difficulty were significantly lower than for placebo for those randomized to B infantis 35624 for most weeks of the treatment phase. At baseline, patients with IBS demonstrated an abnormal IL-10/IL-12 ratio, indicative of a proinflammatory, Th-1 state. This ratio was normalized by B infantis 35624 feeding alone.



B infantis 35624 alleviates symptoms in IBS; this symptomatic response was associated with normalization of the ratio of an anti-inflammatory to a proinflammatory cytokine, suggesting an immune-modulating role for this organism, in this disorder.

title: The immune system in healthy adults and patients with atopic dermatitis seems to be affected differently by a probiotic intervention
abstract: Probiotic bacteria are proposed to alleviate atopic dermatitis (AD) in infants. There are few indications about the effect of probiotics on AD in adults.

title: Effect of administration of bifidobacteria and lactic acid bacteria to newborn calves and piglets.
abstract: The effect of oral administration of bifidobacteria and lactic acid bacteria on newborn livestock was investigated. Oral administration of Bifidobacterium pseudolongum or Lactobacillus acidophilus to calves improved BW gain and feed conversion over that of untreated controls. The frequency of diarrhea occurrence was decreased in the group fed probiotics. However, there was no significant difference between calves administered B. pseudolongum and those administered L. acidophilus. Under conditions of feeding without antibiotics, the frequency of diarrhea was decreased markedly by administration of probiotics. The BW gain of piglets was significantly greater during both the suckling and weaning periods for the group fed probiotics than for the control group. The effect was more evident during the suckling period than during the weaning period. Under conditions of feeding without antibiotics, the frequency of piglet mortality was decreased markedly by administration of probiotics. The probiotics tested here had useful effects, including improved BW gain, feed conversion, and fecal condition of both newborn calves and piglets.

title: Response of Piglets to Oral Administration of Lactic Acid Bacteria.
abstract: Three experiments were conducted to determine the effects of orally administered Streptococcus faecium M74, Lactobacillus casei spp., Streptococcus thermophilus spp., and Lactobacillus bulgaricus spp. on growth, development of digestive organs, intestinal microflora, and local mucosal immunity of piglets. Mixed-sex piglets were used in the experiments. Bottle feeding of S. faecium M74 plus L. casei spp. increased weight gain in piglets of different litters, but not in piglets of the same litter. The S. thermophilus spp. plus L. bulgaricus spp. cultures improved only weight gain for 12 to 21 days of age. There was no significant effect of lactic acid bacteria (LAB) administration on plasma glucose, urea, calcium, alkaline phosphatase, total proteins, albumin, and globulins. The number of lactobacilli was higher and the enterococci and coliforms lower in the groups fed LAB than in the control group. The ratio of lactobacilli to coliforms increased in the intestinal contents of the LAB groups. The pH values of the gut contents were not affected by treatments. Although a significant effect on the size of the digestive organs was not found, the liver weight was smaller and the intestinal length shorter in the treated than in the control group. There were no significant differences in the level of interleukin-2 in the tissues of the jejunum and ileum. However, in the group given S. faecium M74 plus L. casei spp. the interleukin-2 concentration in the ileal tissues increased from 3.74 (control) to 7.43 ng/g. The histological structure of the ileal epithelium of the group of piglets given S .faecium M74 plus L. casei spp., showed an increase in the phagocytic activity of the cells. Some differences in the endoplasmic reticulum and mitochondria in the location of the implanted microvilli were observed in the group given S. thermophilus spp. plus L. bulgaricus spp.

title: Selection of a potential probiotic Lactobacillus strain and subsequent in vivo studies
abstract: The probiotic potential of a Lactobacillus strain, isolated from pig faeces, was assessed as a probiotic in piglets. The strain was examined for resistance to pH 2.0, 0.5% oxgall and antibiotics, and antimicrobial activities against enteric pathogenic bacteria. The probiotic strain, L. reuteri BSA131, was administered through the feed to 25 1-month-old Landrace piglets. The piglets were divided into five groups of five piglets each and fed with different diets for 28 days. The daily consumption of L. reuteriBSA131 was assigned into two groups by the concentration of 106 or 108 freeze-dried bacteria. Fecal samples were collected before, during, and after consumption. Lactobacilli and enterobacteria cell counts were determined in the fecal samples. The liveweight gains and feed consumption of the piglets were recorded daily. This study showed that strain BSA 131 enhanced liveweight gains and feed conversion rates in piglets. It also showed a significant increase in lactobacilli cell counts and decreases in enterobacterial numbers in the fecal samples. Strain BSA 131 was considered to be a potential probiotic for piglets, especially after weaning.

title: Probiotic Treatment Using Bifidobacterium lactis HN019 Reduces Weanling Diarrhea Associated With Rotavirus and Escherichia coli Infection in a Piglet Model
abstract: Diarrhea associated with rotavirus and Escherichia coli is one of the major gastrointestinal problems faced by human infants. Using a piglet model, the authors investigated the protective effectiveness of probiotic feeding against naturally acquired diarrhea in weanlings.  Seventeen piglets were allocated into two groups balanced for live weight and litter of origin. The test group was administered Bifidobacterium lactis HN019 (109 colony-forming units (cfu)/piglet/d) orally until the end of the experiment; the control group did not receive probiotic treatment. After 1 week, animals were penned individually and weaned onto a diet for a weaner. The degree of subsequent diarrheal disease was monitored in both groups of animals, feed intake and live weight gain of the piglets were assessed, and blood and fecal samples were collected to measure concurrent systemic and gastrointestinal tract immunologic activity.  Compared with the controls, piglets that received B. lactis HN019 had a lower severity of weanling diarrhea and maintained a greater feed conversion efficiency during weaning. The protective effect of probiotic feeding was associated with lower concentrations of fecal rotavirus and E. coli, higher blood leukocyte phagocytic and T-lymphocyte proliferative responses, and higher gastrointestinal tract pathogen-specific antibody titers.  These results show that dietary treatment using B. lactis HN019 can reduce the severity of weanling diarrhea associated with rotavirus and E. coli, possibly via a mechanism of enhanced immune-mediated protection. This study suggests that probiotic treatment may be an effective dietary means of preventing or limiting diarrhea in human infants.

title: Evaluation of cheddar cheese as a food carrier for delivery of a probiotic strain to the gastrointestinal tract.
abstract: Cheddar cheese was evaluated as a food carrier for the delivery of viable microorganisms of Enterococcus faecium (Fargo 688; Quest Int., Naarden, The Netherlands) to the gastrointestinal tract. This strain had previously been shown to possess properties required of a probiotic microorganism including the ability to relieve irritable bowel syndrome. The strain was found to survive to high numbers in Cheddar cheese during ripening at 8 degrees C for 15 mo (4 x 10(8) cfu/g) and in yogurt during storage at 4 degrees C for 21 d (4 x 10(7) cfu/g). In an in vitro model system, Cheddar cheese was found to have a greater protective effect than yogurt upon exposure of the probiotic culture to porcine gastric juice at pH 2. Subsequently, a feeding trial involving 8 pigs per group was performed in which a rifampicin-resistant variant of the probiotic strain was fed for 21 d at a mean daily intake of 1.3 x 10(10) cfu/d from Cheddar cheese or 3.7 x 10(9) cfu/d from yogurt. During the feeding period, Cheddar cheese yielded a significantly higher mean fecal probiotic count (2 x 10(6) cfu/g of feces) than did yogurt (5.2 x 10(5) cfu/g of feces). These data indicate that mature Cheddar cheese compares very favorably with fresh yogurt as a delivery system for viable probiotic microorganisms to the gastrointestinal tract.

title: Creating and Maintaining the Gastrointestinal Ecosystem: What We Know and Need To Know from Gnotobiology
abstract: Studying the cross talk between nonpathogenic organisms and their mammalian hosts represents an experimental challenge because these interactions are typically subtle and the microbial societies that associate with mammalian hosts are very complex and dynamic. A large, functionally stable, climax community of microbes is maintained in the murine and human gastrointestinal tracts. This open ecosystem exhibits not only regional differences in the composition of its microbiota but also regional differences in the differentiation programs of its epithelial cells and in the spatial distribution of its component immune cells. A key experimental strategy for determining whether “nonpathogenic” microorganisms actively create their own regional habitats in this ecosystem is to define cellular function in germ-free animals and then evaluate the effects of adding single or several microbial species. This review focuses on how gnotobiotics—the study of germ-free animals—has been and needs to be used to examine how the gastrointestinal ecosystem is created and maintained. Areas discussed include the generation of simplified ecosystems by using genetically manipulatable microbes and hosts to determine whether components of the microbiota actively regulate epithelial differentiation to create niches for themselves and for other organisms; the ways in which gnotobiology can help reveal collaborative interactions among the microbiota, epithelium, and mucosal immune system; and the ways in which gnotobiology is and will be useful for identifying host and microbial factors that define the continuum between nonpathogenic and pathogenic. A series of tests of microbial contributions to several pathologic states, using germ-free and ex-germ-free mice, are proposed.

title: Structure of the intestinal flora responsible for development of the gut immune system in a rodent model.
abstract: The intestinal flora comprising indigenous, autochthonous bacteria is constantly present in the alimentary tract of host animals, including humans. The indigenous bacteria greatly affect the structure and functions of the intestinal mucosa. Studies involving gnotobiotic mice or rats have shown that the presence of limited kinds of intestinal bacteria is responsible for the development of the gut immune system, such as secretory IgA, major histocompatibility complex molecules and intraepithelial lymphocytes. Understanding of the structure of the intestinal flora or the organization of the microbial population in the intestine, based on evaluation of the immunological responses, may clarify its functions in the host animal.

title: Effects of a Probiotic Strain of Enterococcus faecium on the Rate of Natural Chlamydia Infection in Swine
abstract: Chlamydiae are obligately intracellular pathogens which cause infections associated with a broad range of diseases in both livestock and humans. In addition, a large proportion of animals may become persistently infected asymptomatic carriers and serve as reservoirs for other animals which also shed these potential zoonotic pathogens. Reducing the chlamydial load of animals is therefore of major importance, and since large-scale antibiotic treatment is neither desired nor feasible, alternative means of prevention are needed. Here we performed a study comparing the efficacy of a probiotic strain of Enterococcus faecium on the reduction of both the rate of natural infection and the shedding of chlamydiae in swine. The presence of Chlamydiaceae was detected by species-specific PCR of fecal samples of sows taken at three times prior to the birth of piglets. Piglets delivered from chlamydia-positive sows in either the control or the probiotic group were also examined for the frequency of chlamydiae at various ages. Eighty-five percent of the piglets from the control group were found to be chlamydia positive, whereas chlamydiae were found in only 60% of piglets from the probiotic group, results confirmed by fluorescence in situ hybridization and immunohistology, which showed higher rates of infection in the control group. In addition to the reduced frequency of chlamydia-positive piglets in the probiotic group, the time of appearance of positive samples was delayed. To our knowledge, these data show for the first time that a probiotic strain of E. faecium can reduce the rate of carryover infections of piglets by obligate intracellular pathogens.

title: Probiotics: Challenging the Traditional Modes of Action
abstract: A trial was conducted to evaluate the performance of broiler birds which supplemented with a commercial inactivated probiotic. Four hundred and fifty day old broiler chickens were allocated into 3 treatments with 6 replicates each. Each replicate was placed on clean wood shavings in floor pens receiving feed and water ad libitum. In treatment 1, feed was supplemented with 500 g/ton of a heat-inactivatedprobiotic containing Lactobacillus spp, Bifidobacterium animalis, Pediococus acidilactici and Enterococcus faecium. In treatment 2, the feed was supplemented with 500 g/ton of the same probiotic without the heat-inactivating process (commercial product). In Treatment 3 the feed was supplemented with Zinc Bacitracin at 100 ppm. During the length of the experiment the birds received the following diets: pre-initial 1 to 7 days; initial 8 to 21 days; growth, 22 to 35 days; and finisher, 36 to 40 days. Data were analyzed following the same dietary periods. In addition, data was arranged in two periods of similar length; from 1 to 21 days and from 22 to 40 days. During the pre-initial phase, treatment 2 had higher weight gain compared to treatment 1. During the initial phase, treatments 1 and 2 had lower feed conversion compared to treatment 3. In the finisher, treatments 1 and 2 had higher body weight and lower feed conversion than treatment 3. In conclusion, regular and inactivated probiotics had a similar performance which under the conditions of this trial were superior to the treatment containing 100 ppm of zinc bacitracin. These data suggest that not all growth promoting effects are mediated by bacterial metabolites or active colonization of the gastrointestinal tract.

title: Production of Angiotensin-I-Converting-Enzyme-Inhibitory Peptides in Fermented Milks Started by Lactobacillus delbrueckiisubsp. bulgaricus SS1 and Lactococcus lactissubsp. cremoris FT4
abstract: Two fermented milks containing angiotensin-I-converting-enzyme (ACE)-inhibitory peptides were produced by using selectedLactobacillus delbrueckii subsp. bulgaricus SS1 and L. lactis subsp. cremoris FT4. The pH 4.6-soluble nitrogen fraction of the two fermented milks was fractionated by reversed-phase fast-protein liquid chromatography. The fractions which showed the highest ACE-inhibitory indexes were further purified, and the related peptides were sequenced by tandem fast atom bombardment-mass spectrometry. The most inhibitory fractions of the milk fermented by L. delbrueckii subsp.bulgaricus SS1 contained the sequences of β-casein (β-CN) fragment 6-14 (f6-14), f7-14, f73-82, f74-82, and f75-82. Those from the milk fermented by L. lactis subsp.cremoris FT4 contained the sequences of β-CN f7-14, f47-52, and f169-175 and κ-CN f155-160 and f152-160. Most of these sequences had features in common with other ACE-inhibitory peptides reported in the literature. In particular, the β-CN f47-52 sequence had high homology with that of angiotensin-II. Some of these peptides were chemically synthesized. The 50% inhibitory concentrations (IC50s) of the crude purified fractions containing the peptide mixture were very low (8.0 to 11.2 mg/liter). When the synthesized peptides were used individually, the ACE-inhibitory activity was confirmed but the IC50s increased considerably. A strengthened inhibitory effect of the peptide mixtures with respect to the activity of individual peptides was presumed. Once generated, the inhibitory peptides were resistant to further proteolysis either during dairy processing or by trypsin and chymotrypsin.

title: Probiotic properties of Lactococcus lactis ssp. lactis HV219, isolated from human vaginal secretions
abstract: To determine the resistance of Lactococcus lactis ssp. lactis HV219 to acids, bile, antibiotics, inflammatory drugs and spermicides, compare adsorption of the strain to bacteria and Caco‐2 cells under stress, and evaluate the antimicrobial activity of bacteriocin HV219.

title: Effect of protectors on the viability of Lactobacillus acidophilus M92 in simulated gastrointestinal conditions.
abstract: L. acidophilus has been considered to be among the predominant lactobacilli in the intestinal tract of healthy humans. The ability of L. acidophilus to survive and retain its viability through the gastrointestinal (GI) tract is one of the main characteristics desirable for probiotic activity. The influence of mucin and dietary constituents, such as casein, whey proteins concentrate (WPC) and skim milk, on survival of L. acidophilus M92 in the simulated GI conditions, was determined. A washed cell suspension was exposed to pepsin (3g/L) and sodium chloride (5g/L) at pH = 1.5, 2.0, 2.5 and 3.0, and to pancreatin (1g/L), sodium chloride (5g/L) and oxgall (1.5, 2.0, 3.0, 5.0 mg/mL) at pH = 8.0, mimicking the GI environment. Much greater tolerance to gastric and small intestinal juice was observed in the presence of mucin and milk proteins, especially with WPC. The influence of WPC, as the best protector, on the cell viability of L. acidophilus M92 in simulated gastric and small intestinal juice was confirmed by a mathematical model. Critical points, indicating GI conditions that L. acidophilus M92 can tolerate, were established. These observations showed that L. acidophilus M92 survives at pH = 1.99 in simulated gastric juice for 2 h and at 3.1 mg/mL oxgall in simulated small intestinal juice for 4 h, while in the presence of WPC it tolerates even lower pH (1.77) and higher concentration of oxgall (4.25 g/L). Furthermore, only 15 % of the L. acidophilus M92 cells survived the direct transit from simulated gastric to simulated small intestinal juice, but with added WPC, 45 % of the L. acidophilus M92 cells survived. These results suggested the addition of WPC as a protector in preparation of L. acidophilus M92 for probiotic use.

title: Effects of Feeding of Two Potentially Probiotic Preparations from Lactic Acid Bacteria on the Performance and Faecal Microflora of Broiler Chickens
abstract: The aim of this study was to evaluate the potential of two probiotic preparations, containing live lactic acid bacteria (Lactococcus lactis CECT 539 and Lactobacillus casei CECT 4043) and their products of fermentation (organic acids and bacteriocins), as a replacement for antibiotics in stimulating health and growth of broiler chickens. The effects of the supplementation of both preparations (with proven probiotic effect in weaned piglets) and an antibiotic (avilamycin) on body weight gain (BWG), feed intake (FI), feed consumption efficiency (FCE), relative intestinal weight, and intestinal microbiota counts were studied in 1-day posthatch chickens. The experiments were conducted with medium-growth Sasso X44 chickens housed in cages and with nutritional stressed Ross 308 broiler distributed in pens. Consumption of the different diets did not affect significantly the final coliform counts in Sasso X44 chickens. However, counts of lactic acid bacteria and mesophilic microorganisms were higher in the animals receiving the two probiotic preparations (P < 0.05). In the second experiment, although no differences in BWG were observed between treatments, Ross 308 broilers receiving the probiotic Lactobacillus preparation exhibited the lowest FCE values and were considered the most efficient at converting feed into live weight.

title: A Strain of Enterococcus faecium (18C23) Inhibits Adhesion of Enterotoxigenic Escherichia coli K88 to Porcine Small Intestine Mucus
abstract: Few studies, if any, have addressed the adhesion of enterococci to the intestinal mucosa and their interference with the adhesion of pathogens, although more than 60% of probiotic preparations in the market contain strains of enterococci. The objective of this study was to investigate if Enterococcus faecium 18C23 has the ability to inhibit the adhesion of Escherichia coli K88ac and K88MB to the small intestine mucus of piglets. Approximately 9% ofE. faecium 18C23 organisms adhered to the small intestine mucus, and the adhesion was found to be specific. Living E. faecium 18C23 culture efficiently inhibited the adhesion ofE. coli K88ac and K88MB to the piglet intestine mucus. Inhibition of the adhesion of E. coli K88ac to the small intestine mucus was found to be dose dependent. Inhibition of >90% was observed when 109 CFU or more of living E. faecium 18C23 culture per ml was added simultaneously withE. coli to immobilized mucus. The substances from both the 18C23 cells and the spent culture supernatant contributed to the inhibition of adhesion of E. coli K88 to the small intestine mucus receptors. The inhibiting effect was not solely a pH effect since considerable inhibitory action was demonstrated after neutralizing the mixture or spent culture supernatant to pH 7.0. Part of the inhibition of adhesion of E. coli K88ac by E. faecium 18C23 or its supernatant might occur through steric hindrance.

title: Homemade traditional cheeses for the isolation of probiotic Enterococcus faecium strains.
abstract: One hundred twenty-two strains of Enterococcus faecium isolated from Tafí Cheese, a homemade traditional cheese of the highlands in the province of Tucumán, Argentina, were evaluated for their potential application as starter cultures in the manufacture of this traditional cheese. Eleven of the 122 strains showing limited delays in growth in oxgall were selected for the study of bile salts hydrolase activity (BSH), cholesterol reduction, antimicrobial activity, and virulence determinants. Nine strains were able to remove cholesterol in in vitro assays, a property that was closely related to the bile salt hydrolase activity. Only two strains produced active bacteriocins against Listeria strains although genetic evidence for the bacteriocin structural gene was found in six other enterococci strains. No virulence factors were detected in any of the 11 selected strains of enterococci.

title: Factors Affecting the Protein Quality of Yogurt and Acidophilus Milk
abstract: The apparent protein digestibility in vivo, the protein digestibility in vitro, the net protein ratio, and computed protein efficiency ratio were used to assess the protein quality of yogurt and acidophilus milk as affected by heat treatment, direct acidification, addition of culture, or fermentation. Rats fed diets containing either heated mix or yogurt tended to eat more and gain more weight than rats fed diets containing either unheated mix or the acidified yogurt. Feed efficiency was also significantly higher for rats fed heated mix and yogurt. The apparent protein digestibility in vivo was significantly higher in yogurt diet than in the unheated yogurt mix diet. The net protein ratio of the yogurt diet was significantly higher than that of either the unheated mix or the acidified yogurt but not significantly different from the heated mix. The computed protein efficiency ratio values were also increased with both heating and fermentation. The direct acidified yogurt had a significantly lower net protein ratio and lower computed protein efficiency ratio than the fermented yogurt. Rats fed either sweet acidophilus (unfermented) or acidophilus milk exhibited a significantly higher protein digestibility than rats fed heated milk, indicating that the lactic culture may have a direct effect on protein digestibility. Despite their increased protein digestibilities, the acidophilus products did not display any significant increase in net protein ratio or computed protein efficiency ratio.

title: Therapeutic role of dietary lactobacilli and lactobacillic fermented dairy products
abstract: Lactic acid bacteria present in the human and animal gut are introduced through fermented milk products, food and feed supplements containing viable bacteria. Lactobacillus species, such as L. acidophilus, L. bulgaricus, L. lactis along with B. bifidum and Streptococcus faecium constitute an integral part of the healthy gastro-intestinal microecology and are involved in the host metabolism. They impart nutritional and therapeutic benefits to the consumer. The vitamins and enzymes produced by the lactic acid bacteria contribute to host metabolism. The antimicrobial substances produced by these bacteria control the proliferation of undesired pathogens. They also elaborate enzymes that aid host metabolism. This is particularly true in case of lactase nonpersistent individuals. Their anticholesteremic properties assist in lowering serum cholesterol. It has been suggested that the tumor suppression trait of these microbes reduces the incidence of colon cancer.

title: Review of scientific evidence for efficacy of Lactobacillus acidophilus DDS-1 as a probiotic strain
abstract: For more than 25 years, Lactobacillus acidophilus DDS-1 strain has been marketed commercially worldwide as an effective probiotic strain. It has been the subject of a variety of in vitro, human and animal studies for more than 40 years. The focus of the efficacy research on this strain has been on its nutritional, anti-bacterial and anti-pathogenic, anticarcinogenic and technological properties. At the same time, the microbiological and technological properties of DDS-1 have been studied.

title: The bactericidal effects of Lactobacillus acidophilus, garcinol and Protykin® compared to clarithromycin, on Helicobacter pylori
abstract: Chronic infection with Helicobacter pylori causes peptic ulcers, gastric cancer and lymphoma. We evaluated the inhibitory effects of the probiotic Lactobacillus acidophilus DDS-1J, the antibiotic clarithromycin and the natural antioxidants garcinol and Protykin® (containing 50% trans-resveratrol) on Helicobacter pylori strain ATCC 49503. The findings of this study indicate that Lactobacillus acidophilus DDS-1J exerts a growth inhibitory effect on H. pylori at a ratio of 1:1 or higher in vitro. In the case of clarithromycin, garcinol and resveratrol, the bactericidal effect is time and concentration dependent. Clarithromycin completely inhibited growth at ≥ 62.5 μg/ml at 6 h and at ≥ 31.5 μg/ml at 12 h. For garcinol the highest concentration needed for complete inhibition was 31.5 μg/ml at 6 h and 3.9 μg/ml after 12 h incubation. For resveratrol, significant inhibition was noted at 1000 μg/ml at 12 h only. The bactericidal effect of garcinol was reduced by the addition of resveratrol at all concentrations ≤ 125 μg/ml at 6 and 12 h. We conclude from this study that Lactobacillus acidophilus DDS-1J inhibits H. pylori at 1:1 and higher ratios. Also, between the two antioxidants, garcinol is much more potent than resveratrol as a bactericidal agent against H. pylori, and that resveratrol may antagonize this effect. Finally, our study showed equivalent or better bactericidal activity of garcinol compared to clarithromycin against H. pylori at 6 and 12 h incubation, indicating a potential role for this antioxidant in treatment for H. pylori infection.

title: Lactobacillus acidophilus LA 1 binds to cultured human intestinal cell lines and inhibits cell attachment and cell invasion by enterovirulent bacteria.
abstract: Four human Lactobacillus acidophilus strains were tested for their ability to adhere onto human enterocyte like Caco-2 cells in culture. The LA 1 strain exhibited a high calcium independent adhesive property. This adhesion onto Caco-2 cells required a proteinaceous adhesion promoting factor, which was present in the spent bacterial broth culture supernatant. LA 1 strain also strongly bound to the mucus secreted by the homogeneous cultured human goblet cell line HT29-MTX. The inhibitory effect of LA 1 organisms against Caco-2 cell adhesion and cell invasion by a large variety of diarrhoeagenic bacteria was investigated. As a result, the following dose dependent inhibitions were obtained: (a) against the cell association of enterotoxigenic, diffusely adhering and enteropathogenic Escherichia coli, and Salmonella typhimurium; (b) against the cell invasion by enteropathogenic Escherichia coli, Yersinia pseudotuberculosis, and Salmonella typhimurium. Incubations of L acidophilus LA 1 before and together with enterovirulent E coli were more effective than incubation after infection by E coli.

title: The human Lactobacillus acidophilus strain LA1 secretes a nonbacteriocin antibacterial substance(s) active in vitro and in vivo
abstract: The adhering human Lactobacillus acidophilus strain LA1 inhibits the cell association and cell invasion of enteropathogens in cultured human intestinal Caco-2 cells (M. F. Bernet, D. Brassard, J. R. Neeser, and A. L. Servin, Gut 35:483-489, 1994). Here, we demonstrate that strain LA1 developed its antibacterial activity in conventional or germ-free mouse models orally infected by Salmonella typhimurium. We present evidence that the spent culture supernatant of strain LA1 (LA1-SCS) contained antibacterial components active against S. typhimurium infecting the cultured human intestinal Caco-2 cells. The LA1-SCS antibacterial activity was observed in vitro against a wide range of gram-negative and gram-positive pathogens, such as Staphylococcus aureus, Listeria monocytogenes, S. typhimurium, Shigella flexneri, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Enterobacter cloacae. By contrast, no activity was observed against species of the normal gut flora, such as lactobacilli and bifidobacteria. The LA1-SCS antibacterial activity was insensitive to proteases and independent of lactic acid production.

title: Effect of Lactobacillus acidophilus KFRI342 on the development of chemically induced precancerous growths in the rat colon.
abstract: Lactobacillus acidophilus KFRI342, isolated from the Korean traditional food kimchi, was investigated for its suitability as a dietary probiotic. The effects of L. acidophilus KFRI342 on the development of chemically induced (1,2-dimethylhydrazine; DMH) precancerous cytological changes of the colon were investigated in rats. Forty-five male F344 rats were randomly divided into three dietary groups. The control group received a high-fat diet (HF), a second group received a high-fat diet containing the carcinogen (HFC), and a final group received a high-fat diet containing the carcinogen and L. acidophilus KFRI342 (HFCL). L. acidophilus KFRI342 was administered orally three times per week at 2×10(9) c.f.u. ml(-1). L. acidophilus KFRI342 treatments decreased the number of Escherichia coli in faecal samples, the enzyme activities of β-glucuronidase and β-glucosidase, and plasma triglyceride concentration compared to the HF and HFC treatments (P<0.05). L. acidophilus KFRI342 consumption also decreased the ratio of aberrant crypts to aberrant crypt foci incidence and the number of aberrant crypts in HFCL rats. Therefore, L. acidophilus showed potential probiotic activity as an inhibitor of DMH-induced symptoms in live rats. Our in vivo studies indicate that L. acidophilus from kimchi may be suitable as a probiotic for human use.

title: Lactobacillus acidophilus La-5 increases lactacin B production when it senses live target bacteria.
abstract: Lactobacillus acidophilus La-5 is a probiotic strain used in dairy products. Production of bacteriocin by L. acidophilus La-5 was achieved when it was grown in co-cultures with the yogurt starter species Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus. However, bacteriocin induction was not observed when heat-killed cells were used as inducers. This study demonstrates that L. acidophilus La-5 produces lactacin B and that the bacteriocin expression is controlled by an auto-induction mechanism involving the secreted peptide IP_1800. The transcript level of the lactacin B gene cluster expression was investigated in co-cultures between L. acidophilus La-5 and S. thermophilus STY-31 and a remarkable increase of the bacteriocin structural gene (lbaB) transcription was observed. However, lbaB was transcribed constitutively in uninduced L. acidophilus La-5 cells, but the levels of the secreted bacteriocin were not enough to be detected by the agar diffusion assay. A new method for bacteriocin detection was formulated based on the monitoring on real time of Lactobacillus sakei subsp. sakei growth in presence of the supernatant and the cell wall extracts of pure and induced L. acidophilus La-5. These results showed that part of lactacin B secreted remains adhered to cell envelope.

title: Influence of synbiotic containing Lactobacillus acidophilus La5, Bifidobacterium lactis Bb 12, Streptococcus thermophilus, Lactobacillus bulgaricus and oligofructose on gut barrier function and sepsis in critically ill patients: a randomised controlled trial.
abstract: Infective complications are a common cause of mortality and morbidity in critically ill patients. Many factors affect sepsis, one of which is gut barrier function. The aim of this study was to determine whether the oral administration of a synbiotic preparation could alter gut barrier function in critically ill patients and thus reduce sepsis.



A total of 90 patients admitted to an intensive care unit (ICU) were randomised to receive either synbiotic or placebo preparations (45 into each group). The synbiotic preparation consisted of Lactobacillus acidophilus La5, Bifidobacterium lactis Bb 12, Streptococcus thermophilus and Lactobacillus bulgaricus (probiotics) with oligofructose (prebiotic). Gut barrier function was assessed by measurement of intestinal permeability (lactulose/rhamnose test) and culture of nasogastric aspirate on days 1 and 8. All septic complications and mortality were recorded.



There were no differences between the groups in terms of age, sex, APACHE II or POSSUM scores. After 1 week of therapy, patients in the synbiotic group had a significantly lower incidence of potentially pathogenic bacteria (43% versus 75%, P = 0.05) and multiple organisms (39% versus 75%, P = 0.01) in their nasogastric aspirates than controls. There were no significant differences between the groups in terms of intestinal permeability, septic complications or mortality.



The administration of synbiotic in critically ill patients favourably altered the microbial composition of the upper gastrointestinal tract but had no effect on intestinal permeability and was not associated with measurable clinical benefit.

title: Probiotic treatment of collagenous colitis: A randomized, double‐blind, placebo‐controlled trial with Lactobacillus acidophilus and Bifidobacterium animalis subsp. Lactis
abstract: Probiotic treatment may be effective in diseases involving gut microflora and intestinal inflammation. In collagenous colitis (CC), a potential pathogenic role of the gut microflora has been proposed. The effect of probiotic treatment in CC is unknown. Our aim was to investigate the clinical effect of treatment with Lactobacillus acidophilus LA‐5 and Bifidobacterium animalis subsp. lactis BB‐12 (AB‐Cap‐10) in patients with CC.  Patients with CC and diarrhea were in a double‐blind placebo‐controlled study randomized (2:1) to AB‐Cap‐10 or placebo for 12 weeks. The primary end point was reduction in bowel frequency per week of ≥50%. Secondary end points were changes in bowel frequencies, stool consistency, stool weight, histopathology, and abdominal bloating and pain.  Twenty‐nine patients were randomized: 21 to probiotics and 8 to placebo. Reduction in bowel frequency per week of ≥50% occurred in 6 of 21 (29%) and in 1 of 8 (13%) patients receiving probiotic and placebo, respectively (P = 0.635). No differences between treatments were observed regarding the secondary end points. Post hoc analysis showed a median reduction in bowel frequency per week from 32 (range 18–84) to 23 (range 11–56; P < 0.005), a reduction in number of days with liquid stools per week from 6 days (range 0–7 days) to 1 day (range 0–7 days; P < 0.005), and an increase in number of days with solid stools per week (P < 0.05) in the AB‐Cap‐10 group.  AB‐Cap‐10 had no significant effect on the chosen end points. Post hoc analysis demonstrated amelioration of clinical symptoms in the AB‐Cap‐10 group, indicating that probiotic treatment may potentially influence the disease course of CC.

title: Randomized clinical trial of effect of synbiotics, neomycin and mechanical bowel preparation on intestinal barrier function in patients undergoing colectomy
abstract: The aim of this study was to investigate whether it is possible to modulate gut microflora and preserve intestinal barrier function during elective colorectal surgery by using combinations of oral antibiotics, synbiotics and mechanical bowel preparation (MBP).

title: Effects of ingesting Lactobacillus- and Bifidobacterium-containing yogurt in subjects with colonized Helicobacter pylori.
abstract: Evidence suggests that ingesting lactic acid bacteria exerts a suppressive effect on Helicobacter pylori infection in both animals and humans. Supplementing with Lactobacillus- and Bifidobacterium-containing yogurt (AB-yogurt) was shown to improve the rates of eradication of H. pylori in humans.



We administered AB-yogurt to subjects with asymptomatic H. pylori to test whether the yogurt could inhibit H. pylori growth.



The in vitro inhibition of H. pylori growth was determined by inoculating Lactobacillus acidophilus La5 or Bifidobacterium lactis Bb12 on plates that were inoculated with H. pylori. Assessment of the viability of H. pylori was performed by the mixed culture method with La5 or Bb12. In an intervention study, 59 adult volunteers infected with H. pylori were given AB-yogurt (10(7) colony-forming units of both La5 and Bb12/mL) twice daily after a meal for 6 wk. Eleven subjects positive for H. pylori infection were treated with milk placebo as control subjects. H. pylori bacterial loads were determined with use of the (13)C-urea breath test, which was performed before and 4 and 8 wk after the start of AB-yogurt supplementation.



Bb12 exerted an in vitro inhibitory effect against H. pylori, whereas La5 did not show an effect. Administration of AB-yogurt decreased the urease activity of H. pylori after 6 wk of therapy (P < 0.0001).



Regular intake of yogurt containing Bb12 and La5 effectively suppressed H. pylori infection in humans.

title: Pretreatment with Lactobacillus- and Bifidobacterium-containing yogurt can improve the efficacy of quadruple therapy in eradicating residual Helicobacter pylori infection after failed triple therapy.
abstract: Lactobacillus- and Bifidobacterium-containing yogurt (AB-yogurt) can suppress Helicobacter pylori. Improvement of the eradication rate by quadruple therapy of residual H. pylori after failed triple therapy is needed.



We tested whether prior treatment with AB-yogurt improved the efficacy of quadruple therapy in eradicating residual H. pylori after failed triple therapy.



One hundred thirty-eight patients in whom triple therapy failed were enrolled for a culture study of H. pylori to assess antimicrobial resistance. These patients were then randomly assigned in equal numbers to either a yogurt-plus-quadruple therapy group or a quadruple therapy-only group. The patients received 1 wk of quadruple therapy with or without a 4-wk pretreatment with AB-yogurt (400 mL/d). In the yogurt-plus-quadruple group, excessive delta(13)CO(2)/mL values of the (13)C-urea breath test were collected before and every 2 wk during the 4-wk ingestion of yogurt. For both groups, a (13)C-urea breath test was conducted > or =6 wk after the quadruple therapy to assess the outcome of residual H. pylori eradication.



For the patients in the yogurt-plus-quadruple therapy group infected with either antibiotic-sensitive or -resistant H. pylori, the excessive delta(13)CO(2)/mL values of the (13)C-urea breath test were significantly decreased after the 4-wk ingestion of AB-yogurt (P < 0.0001). The yogurt-plus-quadruple therapy group had a higher H. pylori eradication rate than did the quadruple therapy-only group (intention-to-treat analysis: 85% compared with 71.1%, P < 0.05; per-protocol analysis: 90.8% compared with 76.6%, P < 0.05).



A 4-wk pretreatment with AB-yogurt can decrease H. pylori loads despite antimicrobial resistance, thus improving the efficacy of quadruple therapy in eradicating residual H. pylori.

title: Ingested probiotics reduce nasal colonization with pathogenic bacteria (Staphylococcus aureus, Streptococcus pneumoniae, and beta-hemolytic streptococci).
abstract: As a bacterial reservoir, the nose may harbor potentially pathogenic bacteria (PPB: Staphylococcus aureus, Streptococcus pneumoniae, beta-hemolytic streptococci, and Haemophilus influenzae). In patients carrying PPB, antiseptic regimens could be crucial for infection control after major operations on or injuries of the head, nasal sinuses, or lungs. Such regimens may also be important for diabetic patients and persons receiving hemodialysis, in intensive care units, or with impaired immunity due to various other causes.



We tested a possible effect of the ingestion of probiotics on the bacterial flora of the nose.



In an open, prospective trial, 209 volunteers were randomly assigned to consume either a probiotic, fermented milk drink [65 mL with Lactobacillus GG (ATCC 53103), Bifidobacterium sp B420, Lactobacillus acidophilus 145, and Streptococcus thermophilus; n = 108] or standard yogurt (180 g; n = 101) daily for 3 wk. Nasal microbial flora were analyzed on days 1, 21, and 28. The microbial examination was blinded to the source of the samples.



We found a significant reduction (19%; P < 0.001) in the occurrence of nasal PPB in the group who consumed the probiotic drink but not in the group who consumed yogurt. The effect was mainly on gram-positive bacteria, which decreased significantly (P < 0.05).



The results indicate that regular intake of probiotics can reduce PPB in the upper respiratory tract. The results also indicate a linkage of the lymphoid tissue between the gut and the upper respiratory tract.

title: Dietary intervention with the probiotics Lactobacillus acidophilus 145 and Bifidobacterium longum 913 modulates the potential of human faecal water to induce damage in HT29clone19A cells.
abstract: Probiotics reduce the risk of colon cancer by inhibiting carcinogen-induced DNA damage in animals, but there are no analogous data in human subjects. To enhance knowledge of the effects of probiotics in human subjects, we have investigated the genotoxicity of faecal water after dietary intervention with standard yoghurt or with probiotic yoghurt, which included the strains Lactobacillus acidophilus 145 and Bifidobacterium longum 913. Faeces were collected from nine healthy volunteers after intervention with probiotic yoghurt or standard yoghurt. Faecal water was isolated and incubated with human colon tumour cells HT29clone19A. DNA strand breaks, oxidised DNA bases and damage after challenge with H2O2 were determined by micro-gel-electrophoresis. Faecal water was genotoxic in comparison with NaCl, but protected against H2O2-induced DNA strand breaks. The intervention with probiotic yoghurt significantly lowered faecal water genotoxicity compared with standard yoghurt. However, probiotic intervention also increased oxidative damage; this either reflected prooxidative activity or stimulation of endogenous defence systems. Altogether, the balance of effects favoured protection, since faecal water from the probiotic group reduced overall genetic damage. Thus, there was a reduction of strand break-inducing compounds in human faeces after dietary intervention with probiotic bacteria. This protection reflected results from previous studies in carcinogen-exposed animals where probiotics reduced DNA damage in colon cells.

title: Invited review: the scientific basis of Lactobacillus acidophilus NCFM functionality as a probiotic.
abstract: Lactobacillus acidophilus NCFM is a probiotic strain available in conventional foods (milk, yogurt, and toddler formula) and dietary supplements. Its commercial availability in the United States since the mid-1970s is predicated on its safety, its amenability to commercial manipulation, and its biochemical and physiological attributes presumed to be important to human probiotic functionality. The strain has been characterized in vitro, in animal studies, and in humans. NCFM is the progenitor of the strain being used for complete chromosome sequencing and therefore will be a cornerstone strain for understanding the relationship between genetics and probiotic functionality. Both phenotypic and genotypic techniques have verified its taxonomic status as a type A1 L. acidophilus strain. It adheres to Caco-2 and mucus-secreting HT-29 cell culture systems, produces antimicrobial compounds, and is amenable to genetic manipulation and directed DNA introduction. NCFM survives gastrointestinal tract transit in both healthy and diseased populations. NCFM inhibits aberrant crypt formation in mutagenized rats, indicative of activity that could decrease the risk of colon cancer. A blend of probiotic strains containing NCFM decreased the incidence of pediatric diarrhea. NCFM led to a significant decrease in levels of toxic amines in the blood of dialysis patients with small bowel bacterial overgrowth. At adequate daily feeding levels, NCFM may facilitate lactose digestion in lactose-intolerant subjects. Further validation of the probiotic properties of NCFM in humans and clarification of its mechanisms of probiotic action are needed to better understand the role this strain might play in promoting human health.

title: Antagonistic Action of Lactobacillus acidophilus Toward Intestinal and Foodborne Pathogens in Associative Cultures 1.
abstract: Lactobacillus acidophilus exerted antagonistic actions on growth of Staphylococcus aureus , Salmonella typhimurium , enteropathogenic Escherichia coli , and Clostridium perfringens when grown with each in associative cultures. S. aureus and C. perfringens were more sensitive to the inhibition than were S. typhimurium and E. coli . The amount of the antagonism produced varied among strains of L. acidophilus and could not be directly related to amounts of acid produced; hydrogen peroxide produced by the lactobacilli appeared to be partially responsible for the antagonistic interaction. The inhibitory effect was produced also under anaerobic conditions in a pre-reduced medium.

title: Adherence of Lactobacillus species to human fetal intestinal cells.
abstract: Thirty-two lactobacilli were tested for ability to adhere to a human fetal intestinal epithelial cell line. By an in vitro system, two adherence mechanisms were found. One mechanism, requiring calcium in the adherence reaction, was nonspecific and allowed all lactobacilli tested to adhere. The other system, not requiring calcium, was found in four strains, all human Lactobacillus acidophilus isolates. Colonial morphology, serial broth passage, and exposure of cell crops to freezing or lyophilization did not affect adherence of Lactobacillus acidophilus. In vitro adherence, combined with subsequent in vivo studies, may provide a basis for screening candidate organisms for use in microbiotic supplements.

title: Factors involved in adherence of lactobacilli to human Caco-2 cells
abstract: A quantitative assay performed with bacterial cells labelled with [3H]thymidine was used to investigate factors involved in the adherence of human isolates Lactobacillus acidophilus BG2FO4 and NCFM/N2 and Lactobacillus gasseri ADH to human Caco-2 intestinal cells. For all three strains, adherence was concentration dependent, greater at acidic pH values, and significantly greater than adherence of a control dairy isolate, Lactobacillus delbrueckii subsp. bulgaricus 1489. Adherence of L. acidophilus BG2FO4 and NCFM/N2 was decreased by protease treatment of the bacterial cells, whereas adherence of L. gasseri ADH either was not affected or was enhanced by protease treatment. Putative surface layer proteins were identified on L. acidophilus BG2FO4 and NCFM/N2 cells but were not involved in adherence. Periodate oxidation of bacterial cell surface carbohydrates significantly reduced adherence of L. gasseri ADH, moderately reduced adherence of L. acidophilus BG2FO4, and had no effect on adherence of L. acidophilus NCFM/N2. These results indicate that Lactobacillus species adhere to human intestinal cells via mechanisms which involve different combinations of carbohydrate and protein factors on the bacterial cell surface. The involvement of a secreted bridging protein, which has been proposed as the primary mediator of adherence of L. acidophilus BG2FO4 in spent culture supernatant (M.-H. Coconnier, T. R. Klaenhammer, S. Kernéis, M.-F. Bernet, and A. L. Servin,  1992), was not confirmed in this study. Rather, a pH effect on Caco-2 cells contributed significantly to the adherence of this strain in spent culture supernatant.(ABSTRACT TRUNCATED AT 250 WORDS)

title: Performance of commercial cultures in fluid milk applications.
abstract: Six Lactobacillus acidophilus, 5 Bifidobacterium, and 6 Streptococcus thermophilus strains were studied for characteristics that are important to activity and stability in unfermented fluid milk products. Speciation, strain relatedness, frozen concentrate stability, bile sensitivity, and lactase activity were evaluated. The microbiological stability of a culture-containing fluid milk product was also determined. Two of the bifidobacteria cultures contained > 1 strain. Some strains were shown to be closely related or identical by pulsed-field gel electrophoresis of fragmented chromosomal DNA. Selective media that distinguished among all 3 added genera were identified. All lactobacilli and most of the bifidobacteria were resistant to bile concentrations varying from 1 to 3%, and all streptococci were sensitive to bile. Lactase activities were highest for S. thermophilus strains, supporting use of this species in fluid milk and dairy products to aid in the digestion of lactose by consumers. The experimental product evaluated in this study contained 10(7) cfu/ml of both L. Acidophilus and Bifidobacterium spp. and 5 x 10(7) cfu/ml of S. thermophilus. Lactic, but not psychrotrophic, populations were fairly stable during storage. The results suggest that milk formulated with high concentrations of three different genera of probiotic bacteria can be manufactured with commercial strains.

title: Effect of Lactobacillus acidophilus dietary supplements on 1,2-dimethylhydrazine dihydrochloride-induced intestinal cancer in rats.
abstract: The epidemiologic patterns of large bowel cancer have implicated a diet high in beef as a risk factor. Another study has shown less colon cancer in Finns than in Danes. Among the possible explanations were the consumption of milk and fermented dairy products and the numbers of lactobacilli in the fecal flora, both of which were higher in the Finns. As a test of these hypotheses, 20 mg 1,2-dimethylhydrazine dihydrochloride/kg body weight was administered for 20 weeks to male inbred F344 rats. Rats consuming a beef diet simulating a "Western" diet had higher rates of colon cancer than did grain-fed rats (83% vs. 31%, respectively). Viable cultures of Lactobacillus acidophilus were fed to rats eating the meat diet; the rats were then challenged with 1,2-dimethylhydrazine dihydrochloride. The colon cancer incidence after a 20-week induction period was lower in the animals receiving the L. acidophilus (40% vs. 77% in controls), but no difference in incidence was discerned after a 36-week period. Thus dietary supplements of L. acidophilus seemed to increase the latency or induction time for experimental colon cancer.

title: Lactobacillus acidophilus modulates intestinal pain and induces opioid and cannabinoid receptors
abstract: Abdominal pain is common in the general population and, in patients with irritable bowel syndrome, is attributed to visceral hypersensitivity. We found that oral administration of specific Lactobacillus strains induced the expression of μ-opioid and cannabinoid receptors in intestinal epithelial cells, and mediated analgesic functions in the gut—similar to the effects of morphine. These results suggest that the microbiology of the intestinal tract influences our visceral perception, and suggest new approaches for the treament of abdominal pain and irritable bowel syndrome.

title: Strain-specific inhibition of Helicobacter pylori by Lactobacillus salivarius and other lactobacilli.
abstract: To investigate the anti-Helicobacter pylori activity of 28 strains of Lactobacillus salivarius and 12 other lactobacilli, isolated from different sites and from different geographical regions.



An in vitro agar plate diffusion assay was employed to assess the Lactobacillus anti-H. pylori activity.



Nine out of 28 L. salivarius strains and 3/12 other Lactobacillus species tested inhibited H. pylori growth. There was no correlation between ecological niche/geographical location of isolation of the lactobacilli and their inhibitory capability. Further studies on strain L. salivarius UCC119 showed that this strain could inhibit growth of 6/6 clinical isolates of H. pylori, five of which were antibiotic-resistant. This inhibition was not due to acid production and was not mediated by a protein, but did require the presence of live cells.



Growth inhibition of H. pylori by L. salivarius is strain-dependent and is not linked to any particular environmental niche or geographic location. Strains of L. salivarius showing highest anti-H. pylori activity may be useful as an adjunct in the treatment of strains that are resistant to conventional antibiotics.

title: Adhesion of human Lactobacillus acidophilus strain LB to human enterocyte-like Caco-2 cells.
abstract: Twenty-five strains of lactobacilli were tested for their ability to adhere to human enterocyte-like Caco-2 cells in culture. Seven Lactobacillus strains adhered well to the Caco-2 cells, of which three possessed calcium-independent adhesion properties. A high level of calcium-independent adhesion was observed with the human stool isolate Lactobacillus acidophilus strain LB. Scanning electron microscopy revealed that this strain adhered to the apical brush border of the cells. Adhesion increased in parallel with the morphological and functional differentiation of the Caco-2 cells. Two Lactobacillus components were involved in this adhesion. One was protease-resistant and bacterial-surface-associated; the other was heat-stable, extracellular and protease-sensitive.

title: Inhibition of adhesion of enteroinvasive pathogens to human intestinal Caco-2 cells by Lactobacillus acidophilus strain LB decreases bacterial invasion.
abstract: Salmonella typhimurium and enteropathogenic Escherichia coli (EPEC) were found to adhere to the brush border of differentiated human intestinal epithelial Caco-2 cells in culture, whereas Yersinia pseudotuberculosis and Listeria monocytogenes adhered to the periphery of undifferentiated Caco-2 cells. All these enterovirulent strains invaded the Caco-2 cells. Using a heat-killed human Lactobacillus acidophilus (strain LB) which strongly adheres both to undifferentiated and differentiated Caco-2 cells, we have studied inhibition of cell association with and invasion within Caco-2 cells by enterovirulent bacteria. Living and heat-killed Lactobacillus acidophilus strain LB inhibited both cell association and invasion of Caco-2 cells by enterovirulent bacteria in a concentration-dependent manner. The mechanism of inhibition of both adhesion and invasion appears to be due to steric hindrance of human enterocytic pathogen receptors by whole-cell lactobacilli rather than to a specific blockade of receptors.

title: Antagonistic Activity againstHelicobacter Infection In Vitro and In Vivo by the HumanLactobacillus acidophilus Strain LB
abstract: The purpose of the present study was to examine the activity of the human Lactobacillus acidophilus strain LB, which secretes an antibacterial substance(s) against Helicobacter pyloriin vitro and in vivo. The spent culture supernatant (SCS) of the strain LB (LB-SCS) dramatically decreased the viability of H. pylori in vitro independent of pH and lactic acid levels. Adhesion of H. pylori to the cultured human mucosecreting HT29-MTX cells decreased in parallel with the viability of H. pylori. In conventional mice, oral treatment with the LB-SCS protected against infection with Helicobacter felis. Indeed, at both 8 and 49 days post-LB-SCS treatment (29 and 70 days postinfection), inhibition of stomach colonization by H. felis was observed, and no evidence of gastric histopathological lesions was found. LB-SCS treatment inhibits theH. pylori urease activity in vitro and in H. pylori that remained associated with the cultured human mucosecreting HT29-MTX cells. Moreover, a decrease in urease activity was detected in the stomach of the mice infected with H. felis and treated with LB-SCS.

title: Antagonistic Activity of Lactobacillus acidophilus LB against Intracellular Salmonella enterica Serovar Typhimurium Infecting Human Enterocyte-Like Caco-2/TC-7 Cells
abstract: To gain further insight into the mechanism by which lactobacilli develop antimicrobial activity, we have examined howLactobacillus acidophilus LB inhibits the promoted cellular injuries and intracellular lifestyle of Salmonella entericaserovar Typhimurium SL1344 infecting the cultured, fully differentiated human intestinal cell line Caco-2/TC-7. We showed that the spent culture supernatant of strain LB (LB-SCS) decreases the number of apical serovar Typhimurium-induced F-actin rearrangements in infected cells. LB-SCS treatment efficiently decreased transcellular passage ofS. enterica serovar Typhimurium. Moreover, LB-SCS treatment inhibited intracellular growth of serovar Typhimurium, since treated intracellular bacteria displayed a small, rounded morphology resembling that of resting bacteria. We also showed that LB-SCS treatment inhibits adhesion-dependent serovar Typhimurium-induced interleukin-8 production.

title: Lactobacillus acidophilus (strain LB) from the resident adult human gastrointestinal microflora exerts activity against brush border damage promoted by a diarrhoeagenic Escherichia coli in human enterocyte-like cells
abstract: The normal gastrointestinal microflora exerts a barrier effect against enteropathogens. The aim of this study was to examine whether lactobacilli, a minor genus of the resident gut microflora, exerts a protective effect against the cellular injuries promoted by the diarrhoeagenic Afa/Dr diffusely adhering Escherichia coli (Afa/Dr DAEC) C1845 strain in human intestinal cells.  Cultured human intestinal fully differentiated enterocyte-like Caco-2/TC7 cells were used. Antibacterial activity was examined by measuring the viability of the adhering C1845 bacteria. The distribution of brush border associated cytoskeleton and functional proteins was examined by immunofluorescence labelling coupled to confocal laser scanning microscopy analysis.  The activity of Lactobacillus acidophilus strain LB isolated from the resident human gastrointestinal microflora was examined. A dose dependent decrease in viability of C1845 bacteria was observed after both direct contact in vitro between the spent culture supernatant (LB-SCS) and the bacteria, and when the bacteria were adherent on Caco-2/TC7 cells. Protection against the C1845 induced alterations in expression of F-actin, sucrase-isomaltase, dipeptidylpeptidase IV, alkaline phosphatase, and fructose transporter alterations was observed when the cells were exposed to LB-SCS. L acidophilus strain isolated from the resident adult human gastrointestinal microflora, together with its antimicrobial activity, exerts a protective effect against the brush border lesions promoted by the diarrhoeagenic Afa/Dr DAEC strain C1845.

title: Antimutagenic activity of milk fermented by Streptococcus thermophilus and Lactobacillus bulgaricus.
abstract: Antimutagenic activity of acetone or ethylacetate extracts of skim milk fermented by Streptococcus thermophilus, Lactobacillus bulgaricus, or a combination of both the organisms was studied using Salmonella typhimurium (TA 98 and TA 100). Mutagens used were 4-nitroquinoline-N-oxide (a direct-acting mutagen) and 2-aminofluorene (a mutagen requiring S9 activation). Extracts from all fermented milks showed significant (P less than .05) dose response in suppressing the number of revertants caused by NQNO and 2-aminofluorene in both tester strains, whereas extracts from unfermented milk had no effect. Extracts prepared from milk fermented by L. bulgaricus plus S. thermophilus showed significantly (P less than .05) more antimutagenic activity than extracts prepared from milk fermented by S. thermophilus alone. Solvent (acetone vs. ethyl acetate) effect was not significant with 4-nitroquinoline-N-oxide as mutagen. However, in the case of 2-aminofluorene, acetone extracts showed significantly (P less than .05) higher antimutagenic activity. The results of this and related studies strongly indicate that antimutagenic compounds are produced in milk during fermentation by S. thermophilus and L. bulgaricus.

title: Strains and species of lactic acid bacteria in fermented milks (yogurts): effect on in vivo lactose digestion.
abstract: Lactose in yogurt with live bacteria is better tolerated than lactose in other dairy foods, partly because of the activity of microbial beta-galactosidase (beta-gal), which digests lactose in vivo. To evaluate the ability of different strains and species of lactic acid bacteria to digest lactose in vivo, yogurts (containing mixtures of strains of Streptococcus salivarius subsp thermophilus and Lactobacillus delbrueckii subsp bulgaricus) and fermented milks (containing individual species of S thermophilus, L bulgaricus, L acidophilus, or Bifidobacterium bifidus) that varied in microbial beta-gal activity were produced. Selected products were fed to healthy people who cannot digest lactose, and breath hydrogen production was monitored. All yogurts dramatically and similarly improved lactose digestion, regardless of their total or specific beta-gal activity. The response to fermented milks varied from marginal improvement with B bifidus milk to nearly complete lactose digestion with L bulgaricus milk. The results suggest that total beta-gal was not the limiting factor in promoting lactose digestion, perhaps because of a limited rate of intracellular substrate transport.

title: Immunostimulatory oligonucleotide, CpG-like motif exists in Lactobacillus delbrueckii ssp. bulgaricus NIAI B6.
abstract: The present study was conducted to find an immunostimulatory oligonucleotide derived from yogurt starter cultures. The chromosomal DNA was purified from nine strains of Lactobacillus delbrueckii ssp. bulgaricus and six strains of Streptococcus thermophilus. An immunostimulatory ability of the DNA was examined in a proliferation of peyer's patch and splenic B cells. Only the DNA from L. bulgaricus NIAI B6 induced a significant proliferation of both cells. When the DNA was cloned and amplified using PCR, the mitogenic activities to B cells were significantly increased by 13 of 135 DNA clones. Ten homologous nucleotide sequences were found as possible oligonucleotide sequences of mitogens, and were then chemically synthesized (sOL-LB1 to sOL-LB10). One CpG-like motif (sOL-LB7; 5'-CGGCACGCTCACGATTCTTG-3') was identified as an immunostimulatory oligonucleotide, but it did not contain palindromic CpG structure known as a B cell-specific mitogen. The sOL-LB7 substantially bound to B cells and increased the CD69 positive cells in peyer's patch cells. This study demonstrated that L. bulgaricus NIAI B6 was a good candidate of a starter culture for the production of new functional foods, "Bio-Defense Foods".

title: Probiotics inhibit TNF-α-induced interleukin-8 secretion of HT29 cells
abstract: To study the effect of probiotics on interleukin-8 secretion in intestinal epithelia when stimulated by proinflammatory cytokines. Colonic adenocarcinoma HT29 cells were cultured and divided into four groups:control,TNF-α (group T in short),bifidobacterium (group B), lactobacillus (group L). B. Longum and L. bulgaricus were suspended in culture medium with a concentration of 1×10^8cfu/ml and added into 24 wells respectively. One hour later TNF-α (10ng/ml) was added into each well of groups T, B, L. The supernatants were collected and measured for IL-8 after 3 hours, nuclear factor-κB (NF-κB) p65 was also examined by Western blotting.There was less interleukin-8 secretion in HT29 cells when preincubated with B. Longum or L. bulgaricus compared with group T.Less p65 appeared in nuclei in groups B and L compared with group T,as detected by Westem blot.Probiotics can suppress interleukin-8 secretion in intestinal epithelia when stimulated by proinflammatory cytokines, which is most likely mediated by NF-κB.

title: Colonization of the porcine gastrointestinal tract by lactobacilli
abstract: Eight strains of lactobacillus isolated from the porcine gastrointestinal tract were tested for their ability to adhere in vitro to cells collected from stratified squamous epithelium in the digestive tracts of newborn piglets. Piglets were inoculated with individual strains, and their digestive tracts were sampled at intervals to determine the colonizing ability of the lactobacilli. The results of the in vitro test did not predict whether a lactobacillus strain would associate with stratified squamous epithelium in the piglet digestive tract, but epithelial association in vivo appeared to be an important factor in the maintenance of lactobacillus populations in the tract. None of the lactobacillus strains used as inocula was numerically dominant in the tract 7 days after inoculation of the piglets with a single dose of the bacteria.

title: Transfer of Lactic Acid Bacterial Strains from the Feed to the Sow, the Environment, and the Piglets
abstract: The spread of lactic acid bacterial strains to the environment and to newborn piglets was investigated after feeding of such strains to sows. Rifampicin resistant bacterial strains were fed to sows, 1010 c.f.u. per day, during the period from 1 week before expected farrowing until 1 week after farrowing. Fecal samples from the sows and samples of litter were collected for bacteriological examination together with swabs from the pens, the skin of the sows, and from the rectum of the piglets. The test strains were only excreted in relatively low amounts in the feces of the sows, approximately 103 −106 c.f.u. per gram. They were not able to displace the normal lactic acid bacterial flora in the sows nor were they transmitted to the intestinal tract of the piglets to any significant extent. After the last administration the test strains disappeared from both feces, skin, and environment, indicating that no permanent colonization had taken place, although considerable differences in duration of persistence were noticed between test strains.

title: The survival and persistence of Lactobacillus acidophilus LF221 in different ecosystems.
abstract: Being a human isolate and bacterocin producer with wide antimicrobial spectrum, Lactobacillus acidophilus LF221 fulfills preliminary conditions as a potential probiotic strain. To investigate the LF221 strain with respect to its persistence in complex ecosystems, viable LF221 cells were orally administered to laboratory animals (mice and piglets) and added to cheese milk. During the 10-day feeding of mice with LF221 viable cells, 0.9% of lactobacilli isolated from faeces were identical with LF221 strain. Within the piglets' feeding trial, 920 lactobacilli colonies randomly selected from faecal samples were screened for the presence of LF221-like colonies. During the six times of sampling of piglets' faeces, LF221 colonies were always detected. Even the final sampling 10 days after the last LF221 cell suspension was dosed into piglets revealed the presence of LF221 strain at a concentration of about 3 x 10(8) cells/g of faeces. LF221 intake did not significantly influence the total count of lactobacilli in faecal samples and it had no negative health effects on animals. Analyses of cheese demonstrated that the LF221 strain maintained a high viability during ripening and that it could be detected for at least 6 weeks at a level of about 6.8 x 10(6) cells/g of cheese. According to these results, semihard cheese represents a suitable medium for introducing a potential probiotic LF221 into a diet.

title: In Vitro and In Vivo Inhibition of Helicobacter pylori by Lactobacillus casei Strain Shirota
abstract: We studied the potential inhibitory effect of Lactobacillus casei strain Shirota (from the fermented milk product Yakult [Yakult Ltd., Tokyo, Japan]) on Helicobacter pylori by using (i) in vitro inhibition assays with H. pylori SS1 (Sydney strain 1) and nine H. pylori clinical isolates and (ii) the in vivo H. pylori SS1 mouse model of infection over a period of 9 months. In vitro activity against H. pylori SS1 and all of the clinical isolates was observed in the presence of viable L. casei strain Shirota cells but not in the cell-free culture supernatant, although there was profound inhibition of urease activity. In vivo experiments were performed by oral administration of L. casei strain Shirota in the water supply over a period of 9 months to 6-week-old C57BL/6 mice previously infected with H. pylori SS1 (study group; n = 25). Appropriate control groups of H. pylori-infected but untreated animals (n = 25) and uninfected animals given L. casei strain Shirota (n = 25) also were included in the study. H. pylori colonization and development of gastritis were assessed at 1, 2, 3, 6, and 9 months postinfection. A significant reduction in the levels of H. pylori colonization was observed in the antrum and body mucosa in vivo in the lactobacillus-treated study group, as assessed by viable cultures, compared to the levels in the H. pylori-infected control group. This reduction was accompanied by a significant decline in the associated chronic and active gastric mucosal inflammation observed at each time point throughout the observation period. A trend toward a decrease in the anti-H. pylori immunoglobulin G response was measured in the serum of the animals treated with lactobacillus, although this decrease was not significant.

title: Effects of a fermented milk drink containing Lactobacillus casei strain Shirota on the human NK-cell activity.
abstract: Nine healthy middle-aged and 10 elderly volunteers drank fermented milk containing 4 x 10(10) live cells of Lactobacillus casei strain Shirota daily for 3 wk, and their natural killer (NK) activity and other immunological functions were examined. In the experiments with middle-aged volunteers, NK activity significantly increased (P<0.01) 3 wk after the start of intake, elevated NK cell activity remained for the next 3 wk, and this effect was particularly prominent in the low-NK-activity individuals. In the experiments with elderly volunteers, NK activity significantly decreased (P<0.01) in the control group 3 wk after the start of intake; however, the intake of Lactobacillus casei strain Shirota maintained the NK activity. These results suggest that daily intake of Lactobacillus casei strain Shirota provides a positive effect on NK-cell activity.

title: Effect of Lactobacillus strains and Saccharomyces boulardii on persistent diarrhea in children.
abstract: The efficacy of probiotics on persistent diarrhea remains uncertain. The purpose of this study was to evaluate the effect of Lactobacillus sp and Saccharomyces boulardii on persistent diarrhea in children. In a double-blind trial eighty-nine children, aged 6-24 months were randomly distributed to receive pasteurized cow milk containing 2 viable lyophilized strains Lactobacillus casei and Lactobacillus acidophillus strains CERELA, (10(10)-10(12) colony-forming units per g) (n = 30), or lyophilized S. boulardii, (10(10)-10(12) colony forming units per g) (n = 30) or pasteurized cow milk as placebo (n = 29); on each diet 175 g was given twice a day for a 5 day period. Number of depositions, duration of illness and frequency of vomiting were considered. Enteric pathogens were isolated from stools in 40% of the patients, 27% had rotavirus. Lactobacillus and S. boulardii significantly reduced the number of depositions (p < 0.001) and diarrheal duration (p < 0.005). Similarly both significantly (p < 0.002) reduced vomiting as compared with placebo. There was no difference between treatments depending on rotavirus status. In conclusion, L. casei and L. acidophillus strains CERELA and S. boulardii are useful in the management of persistent diarrhea in children.

title: The Probiotic Bacterium Lactobacillus casei Induces Activation of the Gut Mucosal Immune System through Innate Immunity
abstract: The mechanisms by which probiotic bacteria affect the immune system are unknown yet, but many of them are attributed to an increase in the innate or in the acquired immune response. To study the influence of the probiotic bacterium Lactobacillus casei in the expression of receptors involved in the innate immune response, this bacterium was orally administered to BALB/c mice. After, they were sacrificed; the small intestine and intestinal fluids were collected to measure secretory immunoglobulin A (IgA) specific for L. casei. Mononuclear cells from Peyer's patches were isolated to determine the CD-206 and TLR-2 receptors. In histological slices we determined the number of IgA+, CD4+, CD8+, and CD3+ cells and two cytokines (interleulin-5 [IL-5] and IL-6). CD-206 and TLR-2 increased with respect to the untreated control. We did not observe an increase in the T population or in the IL-5-positive cells. IgA+ cells and IL-6-producing cells increased after 7 days of L. casei administration. We did not find specific antibodies against L. casei. The main immune cells activated after oral L. casei administration were those of the innate immune response, with an increase in the specific markers of these cells (CD-206 and TLR-2), with no modification in the number of T cells.

title: Induction by a Lactic Acid Bacterium of a Population of CD4+ T Cells with Low Proliferative Capacity That Produce Transforming Growth Factor β and Interleukin-10
abstract: We investigated whether certain strains of lactic acid bacteria (LAB) could antagonize specific T-helper functions in vitro and thus have the potential to prevent inflammatory intestinal immunopathologies. All strains tested induced various levels of both interleukin-12 (IL-12) and IL-10 in murine splenocytes. In particular,Lactobacillus paracasei (strain NCC2461) induced the highest levels of these cytokines. Since IL-12 and IL-10 have the potential to induce and suppress Th1 functions, respectively, we addressed the impact of this bacterium on the outcome of CD4+ T-cell differentiation. For this purpose, bacteria were added to mixed lymphocyte cultures where CD4+ T-cells from naive BALB/c mice were stimulated weekly in the presence of irradiated allogeneic splenocytes. In these cultures, L. paracasei NCC2461 strongly inhibited the proliferative activity of CD4+ T cells in a dose-dependent fashion. This was accompanied by a marked decrease of both Th1 and Th2 effector cytokines, including gamma interferon, IL-4, and IL-5. In contrast, IL-10 was maintained and transforming growth factor β (TGF-β) was markedly induced in a dose-dependent manner. The bacteria were not cytotoxic, because cell viability was not affected after two rounds of stimulation. Thus, unidentified bacterial components from L. paracasei NCC2461 induced the development of a population of CD4+ T cells with low proliferative capacity that produced TGF-β and IL-10, reminiscent of previously described subsets of regulatory cells implicated in oral tolerance and gut homeostasis.

title: Specific probiotic therapy attenuates antibiotic induced visceral hypersensitivity in mice
abstract: Abdominal pain and discomfort are common symptoms in functional disorders and are attributed to visceral hypersensitivity. These symptoms fluctuate over time but the basis for this is unknown. Here we examine the impact of changes in gut flora and gut inflammatory cell activity on visceral sensitivity.  Visceral sensitivity to colorectal distension (CRD) was assessed at intervals in healthy mice for up to 12 weeks, and in mice before and after administration of dexamethasone or non-absorbable antibiotics with or without supplementation with Lactobacillus paracasei (NCC2461). Tissue was obtained for measurement of myeloperoxidase activity (MPO), histology, microbiota analysis, and substance P (SP) immunolabelling.  Visceral hypersensitivity developed over time in healthy mice maintained without sterile precautions. This was accompanied by a small increase in MPO activity. Dexamethasone treatment normalised MPO and CRD responses. Antibiotic treatment perturbed gut flora, increased MPO and SP immunoreactivity in the colon, and produced visceral hypersensitivity. Administration of Lactobacillus paracasei in spent culture medium normalised visceral sensitivity and SP immunolabelling, but not intestinal microbiota counts.  Perturbations in gut flora and in inflammatory cell activity alter sensory neurotransmitter content in the colon, and result in altered visceral perception. Changes in gut flora may be a basis for the variability of abdominal symptoms observed in functional gastrointestinal disorders and may be prevented by specific probiotic administration.

title: Nutritional Approach to Restore Impaired Intestinal Barrier Function and Growth After Neonatal Stress in Rats
abstract: Psychological stress during the neonatal period results in intestinal barrier dysfunction and growth alterations later in life. We aimed to restore impaired barrier function and growth rate by a nutritional intervention.  Male rat pups (n = 84) were assigned to 1 of 2 rearing conditions from postnatal day (PND) 2 to PND14: S, separated 3 h/d from their mothers, or H, 15 min/d handled controls. From PND15 to PND35, rats received a control diet or a similar diet adapted to contain arachidonic and docosahexaenoic acids, galacto- and fructo-oligosaccharides and Lactobacillus paracasei NCC2461.  Maternal separation had only a minor impact on the measured gut barrier parameters at PND15, whereas it severely affected them at PND35. At this age, intestinal permeability to macromolecules was higher, mucin content in small intestinal tissues was lower and microbiota composition was altered in S compared with H animals. Feeding the adapted diet normalized the intestinal permeability, although it did not restore intestinal mucin content or microbiota. In addition, the adapted diet improved the growth rate recovery of the S animals after weaning and resulted in increased villus length in small intestine.  Our results suggest that an adapted diet containing specific long-chain polyunsaturated fatty acids, prebiotics and probiotics can revert the negative imprinting of neonatal stress on both intestinal barrier function and growth.

title: Probiotics and immune response.
abstract: Current evidence supports the concept that oral administration of probiotic lactobacilli may be therapeutic in preventing antibiotic-associated diarrhea in children and in reestablishing normal flora in the gastrointestinal tract. Children with human immunodeficiency virus (HIV) infections may have episodes of diarrhea and frequently experience malabsorption associated with possible bacterial overgrowth; together these may interact to produce the growth abnormalities characteristic of this group. The overall objective of this investigation has been to determine whether oral administration of the probiotic Lactobacillus plantarum 299v could improve nutrient status and promote growth in children congenitally exposed to HIV. In addition, the possible beneficial effect of Lactobacillus plantarum 299v in modulating immune response was evaluated. In preliminary results described here, we report on the ability of Lactobacillus plantarum 299v to colonize children with HIV and to elicit specific systemic immune response after oral supplementation.

title: Alteration of intestinal microflora is associated with reduction in abdominal bloating and pain in patients with irritable bowel syndrome
abstract: Alteration of intestinal microflora is associated with reduction in abdominal bloating and pain in patients with irritable bowel syndrome

title: A controlled, double-blind, randomized study on the efficacy of Lactobacillus plantarum 299V in patients with irritable bowel syndrome
abstract: Irritable bowel syndrome (IBS) is a widespread functional disorder of the digestive tract. Its aetiology is unknown and therapeutic options are limited. Recent reports suggest that probiotics may have a role in regulating the motility of the digestive tract.  To assess the efficacy of Lactobacillus plantarum 299V (LP299V) in patients with IBS.  Forty patients were randomized to receive either LP299V in liquid suspension (20 patients) or placebo (20 patients) over a period of 4 weeks. Clinical examination was performed at baseline and at the end of the study. Additionally, patients assessed their symptoms by applying a scoring system.  All patients treated with LP299V reported resolution of their abdominal pain as compared to 11 patients from a placebo group (P = 0.0012). There was also a trend towards normalization of stools frequency in constipated patients in six out of 10 patients treated with LP299V compared with two out of 11 treated with placebo (P = 0.17). With regards to all IBS symptoms an improvement was noted in 95% of patients in the LP299V group vs 15% of patients in the placebo group (P < 0.0001).  LP299V seems to have a beneficial effect in patients with IBS. Further studies on larger cohorts of patients and with longer duration of therapy are required in order to establish the place of L. plantarum in the treatment of IBS.

title: T2030 Evaluation of Lactobacillus Plantarum 299v Efficacy in IBS: Results of a Randomized Placebo-Controlled Trial in 200 Patients
abstract: Lactobacillus plantarum 299v is a probiotic strain indicated to treat IBS patients. No other trial so far has tested the symptomatic effect of this strain in such a large population sample.  The aim of this multicenter double blind randomized placebocontrolled trial was to determine the efficacy of L.plantarum 299v in a powerful trial. Patients from 19 to 70 years old and suffering from chronic IBS according to Rome III criteria were recruited in 3 Indian centers. Treatment duration was 4 weeks long with per day one cap of placebo or one cap of 1010 cfu of L.plantarum 299v. Both frequency and intensity of abdominal pain, bloating, feeling of incomplete rectal evacuation and the number of stools were assessed by a VAS. The results are expressed as the percentage of improvement between baseline and the end of the treatment period. Overall patients' assessment was expressed on a 4-point rated scale. Significance analyses were made by using Chi-square or ANOVAR tests.  The ITT population comprised 200 patients and only 10 patients dropped out (PP population: 190). Neither the demographic characteristics nor the diet were different between the two treatment arms (see table 1). At the end of treatment, L.plantarum 299v improved significantly the symptoms both compared to baseline and to placebo (see table 2). Overall patients' assessment was significantly better in the L.plantarum 299v group compared to the placebo group (see table 2). Pulse and respiratory rates, blood pressure and internal temperature remained normal throughout the study without any significant changes in both arms. No side-effect was reported in both groups.  A 4 weeks treatment with Lactobacillus plantarum 299v has shown to be very effective for the relief of symptoms, particularly abdominal pain and bloating in IBS patients fulfilling the Rome III criteria.  Clinical Evaluation and Patients' assessment

title: Randomized clinical trial of specific lactobacillus and fibre supplement to early enteral nutrition in patients with acute pancreatitis
abstract: Microbial infection of the pancreatic tissue in patients with severe acute pancreatitis increases the morbidity and mortality rates. Colonization of the lower gastrointestinal tract and oropharynx with Gram‐negative, but sometimes also Gram‐positive, bacteria precedes contamination of the pancreas. The aim of this study was to determine whether lactic acid bacteria such as Lactobacillus plantarum 299 could prevent colonization of the gut by potential pathogens and thus reduce the endotoxaemia associated with acute pancreatitis.

title: Early enteral supply of lactobacillus and fiber versus selective bowel decontamination: a controlled trial in liver transplant recipients
abstract: Early enteral nutrition with solutions containing prebiotics (fiber) and probiotics (Lactobacillus) is suggested to reduce bacterial translocation and minimize the incidence of infections after liver transplantation.  In a prospective, randomized placebo-controlled trial consisting of 95 patients, we compared the incidence of postoperative infections and other complications after liver transplantation among three different groups, all supplied with early enteral nutrition: (a) standard formula plus selective bowel decontamination (SBD), (b) fiber-containing formula plus living Lactobacillus plantarum 299, and (c) fiber-containing formula plus heat-killed L plantarum 299.  The groups were comparable regarding preoperative American Society of Anesthesiologists classification, Child-Pugh classification of cirrhosis, operative data, and degree of immunosuppression. The patients who received living lactobacilli plus fiber developed significantly fewer bacterial infections (13%) than the patients with SBD (48%). The incidence of infections was 34% in the group with inactivated lactobacilli and fiber. Cholangitis and pneumonia were the leading infections and enterococci the most commonly isolated bacteria. In the living Lactobacillus group, the mean duration of antibiotic therapy, the mean total hospital stay, and the stay on the intensive care unit were also shorter than in the groups with inactivated lactobacilli and fiber as well as with SBD. However, these differences did not reach statistical significance.  Early enteral nutrition with fiber-containing solutions and living L plantarum 299 was well tolerated. It decreases markedly the rate of postoperative infections both in comparison with inactivated L plantarum 299 and significantly with SBD and a standard enteral nutrition formula. As it is a cheap and feasible alternative to SBD, further studies should evaluate whether this ecoimmunonutrition should be already started while patients are on the waiting list for transplantation.

title: Early enteral supply of fiber and Lactobacilli versus conventional nutrition: a controlled trial in patients with major abdominal surgery.
abstract: Early enteral nutrition with fiber-containing solutions plus Lactobacillus may reduce bacterial translocation and minimize the incidence of infections after surgery.



In a prospective, randomized trial in three groups (n = 30/group) of patients after major abdominal surgery, we compared our previous regimen with parenteral nutrition or fiber-free enteral nutrition (group A) with enteral fiber-containing nutrition with living Lactobacillus (group B) and heat-killed Lactobacillus (group C). The main endpoint was the development of bacterial infection. Other analyzed parameters were the durations of antibiotic therapy and hospital stay, non-infectious complications, side effects of the nutrition, and onset of bowel movement. Routine parameters, nutritional parameters, and cellular immune status in the blood were measured preoperatively and on 1, 5, and 10 d postoperatively.



The incidence of infections was significantly lower (P = 0.01) in groups B and C with enteral nutrition containing fibers (10% each) than in group A (30%). Patients in group B received antibiotics for a significantly shorter time (P = 0.04) than did the patients in groups A and C. The length of hospital stay and the incidence of non-infectious complications did not differ significantly. Fibers and lactobacilli were well tolerated. There were no general benefits of living Lactobacillus as opposed to heat-killed Lactobacillus in the entire study population, but benefits were observed in the patients with gastric and pancreas resections, although no statistical analysis was done due to their small numbers.



Early enteral nutrition with fiber-containing solutions reduced the rate of postoperative infections in comparison with parenteral nutrition and fiber-free enteral formula. Addition of living Lactobacillus seemed to increase the benefits in patients with gastric and pancreatic resections.

title: Lactobacillus plantarum 299v Inhibits Escherichia coli-Induced Intestinal Permeability
abstract: The purpose of this work was to investigate whether a probiotic bacterium, Lactobacillus plantarum 299v, could affect Escherichia coli-induced passage of mannitol across the intestinal wall. Sprague-Dawley rats were pretreated for one week by either tube feeding with L. plantarum 299v twice daily, free access to L. plantarum 299v by adding the bacterium in the drinking water, or negative control receiving regular feeding. Intestinal segments were mounted in Ussing chambers and the mucosa was exposed to control medium, E. coli, and L. plantarum 299v (alone or together). [14C]Mannitol was added as a marker of intestinal permeability and samples were taken from the serosal side. E. coli exposure induced a 53% increase in mannitol passage across the intestinal wall (P < 0.05). One week of pretreatment with L. plantarum 299v in the drinking water abolished the E. coli-induced increase in permeability. Tube feeding for one week or short-term addition of L. plantarum 299v in the Ussing chambers had no effect on the permeability provoked by E. coli challenge. Notably, L. plantarum 299v itself did not change the intestinal passage of mannitol. These data demonstrate that pretreatment with L. plantarum 299v, which is a probiotic bacterium, protects against E. coli-induced increase in intestinal permeability, and that L. plantarum 299v alone has no influence on the intestinal permeability. Thus, this study supports the concept that probiotics may exert beneficial effects in the gastrointestinal tract.

title: FEEDING OF A PROBIOTIC FOR THE PREVENTION OF COMMUNITY-ACQUIRED DIARRHEA IN YOUNG MEXICAN CHILDREN. † 1089
abstract: FEEDING OF A PROBIOTIC FOR THE PREVENTION OF COMMUNITY-ACQUIRED DIARRHEA IN YOUNG MEXICAN CHILDREN. † 1089

title: Lactobacillus reuteri as a therapeutic agent in acute diarrhea in young children.
abstract: Certain strains of lactobacilli may promote recovery from acute diarrhea. Lactobacillus reuteri is of human origin and is a natural colonizer of gastrointestinal tract. In this trial, exogenously administered L. reuteri was studied as a therapeutic agent in acute diarrhea.



Forty patients between 6 and 36 months of age hospitalized with acute diarrhea (75% rotavirus) were studied. After parental consent, the patients were randomized to one of two treatment groups to receive either 10(10) to 10(11) colony-forming units of L. reuteri or a matching placebo daily for the length of hospitalization or up to 5 days. The clinical outcome of diarrhea and colonization of L. reuteri were evaluated.



The mean (SD) duration of watery diarrhea after treatment was 1.7 (1.6) days in the L. reuteri group and 2.9 (2.3) days in the placebo group (p = 0.07). On the second day of treatment only 26% of patients receiving L. reuteri had watery diarrhea, compared with 81% of those receiving placebo (p = 0.0005). Cultures of lactobacilli from stool samples demonstrated that administration of L. reuteri resulted in colonization of the gastrointestinal tract. Lactobacillus reuteri accounted for > 75% of the total lactobacilli found in children fed with this product.



Lactobacillus reuteri is effective as a therapeutic agent in acute rotavirus diarrhea in children. Further studies are warranted to confirm the present finding and to explore the full therapeutic potential of L. reuteri in acute viral diarrhea.

title: Bacteriotherapy with Lactobacillus reuteri in rotavirus gastroenteritis.
abstract: Certain lactic acid bacteria may accelerate recovery from acute diarrhea. Lactobacillus reuteri is a commonly occurring Lactobacillus species with therapeutic potential in diarrhea.



Prospective, randomized, placebo-controlled trial in two hospitals.



Children between 6 and 36 months of age admitted for rotavirus-associated diarrhea were randomized into three groups to receive either 10(10) or 10(7) colony-forming units (cfu) of L. reuteri or a matching placebo once a day for up to 5 days.



The main effect of L. reuteri was on the duration of watery diarrhea. The mean (+/-SD) duration of watery diarrhea after initiation of treatment was 2.5 (1.5) days in the placebo group (n = 25) vs. 1.9 (0.9) days in the small dosage (n = 20) and 1.5 (1.1) days in the large dosage (n = 21) L. reuteri recipients (P = 0.01). By the second day of treatment watery diarrhea persisted in 80% of the placebo, 70% of the small dosage and 48% of the large dosage L. reuteri recipients (P = 0.04, large dosage vs. placebo). Stool cultures for lactobacilli confirmed that administration of L. reuteri resulted in good colonization of the GI tract. The mean (+/-SD) of total Lactobacillus count 2 days after treatment initiation was 2.8 (1.6) log 10 cfu/g in the placebo group, 4.5 (2.0) log 10 cfu/g in the small dosage L. reuteri group and 6.1 (1.2) log 10 cfu/g in the large dosage L. reuteri group (P = 0.0004).



L. reuteri effectively colonized the gastrointestinal tract after administration and significantly shortened the duration of watery diarrhea associated with rotavirus. There was a correlation between the dosage of L. reuteri and the clinical effect.

title: Effect of a Probiotic Infant Formula on Infections in Child Care Centers: Comparison of Two Probiotic Agents
abstract: To investigate the effect of 2 different species of probiotics in preventing infections in infants attending child care centers.  A double-blind, placebo-controlled, randomized trial was conducted from December 1, 2000, to September 30, 2002, at 14 child care centers in the Beer-Sheva area of Israel in healthy term infants 4 to 10 months old. Infants were assigned randomly to formula supplemented with Bifidobacterium lactis (BB-12), Lactobacillus reuteri , or no probiotics. Duration of feeding, including follow-up, for each participant was 12 weeks. All infants were fed only the assigned formula and were not breastfed due to parental decision before recruitment to the study. Probiotic or prebiotic food products or supplements were not allowed. Main outcome measures were number of days and number of episodes with fever (>38°C) and number of days and number of episodes with diarrhea or respiratory illness.  Participants (n = 201) were similar regarding gestational age, birth weight, gender, and previous breastfeeding. The controls (n = 60), compared with those fed B lactis (n = 73) or L reuteri (n = 68), had significantly more febrile episodes (mean [95% confidence interval]: 0.41 [0.28–0.54] vs 0.27 [0.17–0.37] vs 0.11 [0.04–0.18], respectively). The controls also had more diarrhea episodes (0.31 [0.22–0.40] vs 0.13 [0.05–0.21] vs 0.02 [0.01–0.05], respectively) and episodes of longer duration (0.59 [0.34–0.84] vs 0.37 [0.08–0.66] vs 0.15 [0.12–0.18] days, respectively). The L reuteri group, compared with BB-12 or controls, had a significant decrease of number of days with fever, clinic visits, child care absences, and antibiotic prescriptions. Rate and duration of respiratory illnesses did not differ significantly between groups.  Child care infants fed a formula supplemented with L reuteri or B lactis had fewer and shorter episodes of diarrhea, with no effect on respiratory illnesses. These effects were more prominent with L reuteri, which was also the only supplement to improve additional morbidity parameters.

title: Lactobacillus reuteri therapy to reduce side‐effects during anti‐Helicobacter pylori treatment in children: a randomized placebo controlled trial
abstract: Helicobacter pylori eradication fails in about 25–30% of children, particularly because of the occurrence of resistance to antibiotics and side‐effects.

title: Lactobacillus reuteri (American Type Culture Collection Strain 55730) Versus Simethicone in the Treatment of Infantile Colic: A Prospective Randomized Study
abstract: The goal was to test the hypothesis that oral administration of Lactobacillus reuteri in a prospective randomized study would improve symptoms of infantile colic.  Ninety breastfed colicky infants were assigned randomly to receive either the probiotic L reuteri (108 live bacteria per day) or simethicone (60 mg/day) each day for 28 days. The mothers avoided cow's milk in their diet. Parents monitored daily crying times and adverse effects by using a questionnaire.  Eighty-three infants completed the trial: 41 in the probiotic group and 42 in the simethicone group. The infants were similar regarding gestational age, birth weight, gender, and crying time at baseline. Daily median crying times in the probiotic and simethicone groups were 159 minutes/day and 177 minutes/day, respectively, on the seventh day and 51 minutes/day and 145 minutes/day on the 28th day. On day 28, 39 patients (95%) were responders in the probiotic group and 3 patients (7%) were responders in the simethicone group. No adverse effects were reported.  In our cohort, L reuteri improved colicky symptoms in breastfed infants within 1 week of treatment, compared with simethicone, which suggests that probiotics may have a role in the treatment of infantile colic.

title: Colonization and Immunomodulation by Lactobacillus reuteri ATCC 55730 in the Human Gastrointestinal Tract
abstract: is a probiotic (health-promoting) bacterium widely used as a dietary supplement. This study was designed to examine local colonization of the human gastrointestinal mucosa after dietary supplementation with L. reuteri ATCC 55730 and to determine subsequent immune responses at the colonized sites. In this open clinical investigation, 10 healthy volunteers and 9 volunteers with ileostomy underwent gastroscopy or ileoscopy and biopsy samples were taken from the stomach, duodenum, or ileum before and after supplementation with 4 × 108 CFU of live L. reuteri ATCC 55730 lactobacilli per day for 28 days. Biopsy specimen colonization was analyzed using fluorescence in situ hybridization with a molecular beacon probe, and immune cell populations were determined by immunostaining. Endogenous L. reuteri was detected in the stomach of 1 subject and the duodenum of 3 subjects (out of 10 subjects). After L. reuteri ATCC 55730 supplementation, the stomachs of 8 and the duodenums of all 10 subjects were colonized. Three ileostomy subjects (of six tested) had endogenous L. reuteri at baseline, while all six displayed colonization after L. reuteri supplementation. Gastric mucosal histiocyte numbers were reduced and duodenal B-lymphocyte numbers were increased by L. reuteri ATCC 55730 administration. Furthermore, L. reuteri administration induced a significantly higher amount of CD4-positive T-lymphocytes in the ileal epithelium. Dietary supplementation with the probiotic L. reuteri ATCC 55730 induces significant colonization of the stomach, duodenum, and ileum of healthy humans, and this is associated with significant alterations of the immune response in the gastrointestinal mucosa. These responses may be key components of a mechanism by which L. reuteri ATCC 55730 exerts its well-documented probiotic effects in humans.

title: Lactobacillus reuteri in bovine milk fermented decreases the oral carriage of mutans streptococci.
abstract: The effect of Lactobacillus reuteri against one of the major cariogenic organism, Streptococcus mutans, was studied. Yogurt products containing L.reuteri showed a significant growth inhibitory effect against S. mutans, whilst yoghurts with lactobaccilli other than L. reuteri did not show such inhibition. Further, double-blind, placebo-controlled trial demonstrated that consuming yogurt with L. reuteri significantly reduced the oral carriage of mutans streptococci, compared with the placebo yogurt.

title: Inhibition of Helicobacter pylori Infection in Humans by Lactobacillus reuteri ATCC 55730 and Effect on Eradication Therapy: A Pilot Study
abstract: Several studies report an inhibitory effect of probiotics on Helicobacter pylori.

title: Nutritional factors affecting the production of two bacteriocins from lactic acid bacteria on whey.
abstract: The ability of Lactococcus lactis subsp. lactis CECT 539 and Pediococcus acidilactici NRRL B-5627 to produce bacteriocins on both diluted and concentrated whey was investigated in batch fermentations. Both strains produced the higher amounts of biomass and bacteriocin titres on diluted whey. Luedeking and Piret expression was able to model the production of nisin, which was produced as a primary metabolite on both culture media. However, the pediocin production could not be typified in any case due to the negligible growth of P. acidilactici. Although the whey supported the growth and bacteriocin production by the two strains, both biomass and bacteriocin productions were lower than those obtained on MRS broth. The effect of total sugar, nitrogen, phosphorous and buffer concentrations on the production of nisin and pediocin was studied in diluted whey using factorial experiments and empirical modelling. The production of nisin was greatly inhibited by the increase in nitrogen, buffer, and to a lesser extent, sugar concentration in the medium, nevertheless, the used phosphorous source produced a light stimulatory effect on bacteriocin synthesis. In addition, the growth of Lc 1.04 was mainly affected by the nitrogen source used. On the other hand, pediocin was inhibited by the increase in buffer, phosphorous, and to a lesser degree, by the sugar and nitrogen concentration. The inhibitory activity of pediocin disappeared almost totally after 15 min of treatment with trypsin, papain, subtilisin and pepsin. The activity of nisin was drastically reduced by treatment with trypsin, subtilisin and pepsin. Nevertheless, 50% of the initial activity was retained when nisin was treated with papain. Both bacteriocins showed the highest heat stability at acidic pH and short incubation times.

title: Bacteriocins of lactic acid bacteria.
abstract: Lactic acid bacteria produce a variety of antagonistic factors that include metabolic end products, antibiotic-like substances and bactericidal proteins, termed bacteriocins. The range of inhibitory activity by bacteriocins of lactic acid bacteria can be either narrow, inhibiting only those strains that are closely related to the producer organism, or wide, inhibiting a diverse group of Gram-positive microorganisms. The following review will discuss biochemical and genetic aspects of bacteriocins that have been identified and characterized from lactic acid bacteria.

title: Production of bacteriocins from Lactococcus lactis subsp. lactis CECT 539 and Pediococcus acidilactici NRRL B‐5627 using mussel‐processing wastes
abstract: The growth and bacteriocin production by Lactococcus lactis subsp. lactis CECT 539 and Pediococcus acidilactici NRRL B‐5627 were investigated on mussel‐processing wastes. Both bacteriocin productions were satisfactorily modelled using a modified form of the Luedeking and Piret expression, which includes a term for the influence of the pH reduction rate. Experimental data from cultures buffered at different initial concentrations (0, 0.03, 0.10 and 0.25 M) of both bacteria were used to fit and verify the model. The influence of total sugars, nitrogen, phosphorus and buffer concentration on nisin and pediocin production was also studied using response‐surface methodology and empirical modelling. Enhanced nisin production (33 BU/ml) was achieved in media buffered with 0.10 M potassium hydrogen phthalate/NaOH. However, the highest levels of pediocin (368 BU/ml) were obtained in the non‐buffered media.

title: Bacteriocins of gram-positive bacteria.
abstract: In recent years, a group of antibacterial proteins produced by gram-positive bacteria have attracted great interest in their potential use as food preservatives and as antibacterial agents to combat certain infections due to gram-positive pathogenic bacteria. They are ribosomally synthesized peptides of 30 to less than 60 amino acids, with a narrow to wide antibacterial spectrum against gram-positive bacteria; the antibacterial property is heat stable, and a producer strain displays a degree of specific self-protection against its own antibacterial peptide. In many respects, these proteins are quite different from the colicins and other bacteriocins produced by gram-negative bacteria, yet customarily they also are grouped as bacteriocins. Although a large number of these bacteriocins (or bacteriocin-like inhibitory substances) have been reported, only a few have been studied in detail for their mode of action, amino acid sequence, genetic characteristics, and biosynthesis mechanisms. Nevertheless, in general, they appear to be translated as inactive prepeptides containing an N-terminal leader sequence and a C-terminal propeptide component. During posttranslational modifications, the leader peptide is removed. In addition, depending on the particular type, some amino acids in the propeptide components may undergo either dehydration and thioether ring formation to produce lanthionine and beta-methyl lanthionine (as in lantibiotics) or thio ester ring formation to form cystine (as in thiolbiotics). Some of these steps, as well as the translocation of the molecules through the cytoplasmic membrane and producer self-protection against the homologous bacteriocin, are mediated through specific proteins (enzymes). Limited genetic studies have shown that the structural gene for such a bacteriocin and the genes encoding proteins associated with immunity, translocation, and processing are present in a cluster in either a plasmid, the chromosome, or a transposon. Following posttranslational modification and depending on the pH, the molecules may either be released into the environment or remain bound to the cell wall. The antibacterial action against a sensitive cell of a gram-positive strain is produced principally by destabilization of membrane functions. Under certain conditions, gram-negative bacterial cells can also be sensitive to some of these molecules. By application of site-specific mutagenesis, bacteriocin variants which may differ in their antimicrobial spectrum and physicochemical characteristics can be produced. Research activity in this field has grown remarkably but sometimes with an undisciplined regard for conformity in the definition, naming, and categorization of these molecules and their genetic effectors. Some suggestions for improved standardization of nomenclature are offered.

title: Detection and activity of lactacin B, a bacteriocin produced by Lactobacillus acidophilus.
abstract: A total of 52 strains of Lactobacillus acidophilus were examined for production of bacteriocins. A majority (63%) demonstrated inhibitory activity against all members of a four-species grouping of Lactobacillus leichmannii, Lactobacillus bulgaricus, Lactobacillus helveticus, and Lactobacillus lactis. Four L. acidophilus strains with this activity also inhibited Streptococcus faecalis and Lactobacillus fermentum, suggesting a second system of antagonism. Under conditions eliminating the effects of organic acids and hydrogen peroxide, no inhibition of other gram-positive or -negative genera was demonstrated by L. acidophilus. The agent produced by L. acidophilus N2 and responsible for inhibition of L. leichmannii, L. bulgaricus, L. helveticus, and L. lactis was investigated. Ultrafiltration studies indicated a molecular weight of approximately 100,000 for the crude inhibitor. The agent was sensitive to proteolytic enzymes and retained full activity after 60 min at 100 degrees C (pH 5). Activity against sensitive cells was bactericidal but not bacteriolytic. These characteristics identified the inhibitory agent as a bacteriocin, designated lactacin B. Examination of strains of L. acidophilus within the six homology groupings of Johnson et al. ( 1980) demonstrated that production of the bacteriocin lactacin B could not be used in classification of neotype L. acidophilus strains. However, the usefulness of employing sensitivity to lactacin B in classification of dairy lactobacilli is suggested.

title: Modelling the influence of pH on the kinetics of both nisin and pediocin production and characterization of their functional properties
abstract: A kinetic model of the production of bacteriocins on MRS medium by Lactococcus lactis subsp. lactis CECT 539 and Pediococcus acidilactici NRRL B-5627 has been developed. The model is a modification of the Luedeking and Piret expression, including a term for the influence of pH on both nisin and pediocin production. With this model it was demonstrated that these bacteriocins are produced as primary metabolites depending on the pH. By modeling the effects of temperature and pH on their activities, the high heat stability of both bacteriocins was demonstrated, with optimum values at acidic pH and short incubation times. The initial activity of both nisin and nisaplin was drastically reduced by treatment with trypsin, subtilisin and pepsin, but 50% activity was retained when both antibacterial substances were treated with papain. The inhibitory activity of pediocin almost totally disappeared after 15 min of treatment with these four proteases.

title: Enhancement of nisin production by Lactococcus lactis in periodically re‐alkalized cultures
abstract: Synthesis of nisin as well as biomass production by Lactococcus lactis subsp. lactis CECT (Colección Española de Cultivos Tipo) 539 on both hydrolysed mussel‐processing waste and whey medium were followed in three fixed volume fed‐batch fermentations, with re‐alkalization cycles. The two cultures on mussel‐processing waste (MPW) were fed with a 240 g/l concentrated glucose and with a concentrated MPW (about 100 g of glucose/l). The culture on whey was fed with a mixture of concentrated whey (48 g of total sugars/l) and a 400 g/l concentrated lactose. The three cultures were mainly characterized with higher nisin titres [49.7, 109.6 and 124.7 bacteriocin activity units (AU)/ml respectively] compared with the batch process on de Man, Rogosa and Sharpe [(1960)  23, 130–135] medium (49.6 AU/ml), MPW (9.5 AU/ml) and whey (22.5 AU/ml) [1 AU/ml is the amount of antibacterial compound needed to obtain 50% growth inhibition (LD50) compared with control tubes]. In the three fed‐batch cultures a shift from homolactic to mixed‐acid fermentation was observed, and other products (acetic acid, butane‐2,3‐diol or ethanol) in addition to lactic acid were detectable in the medium. However, their contributions to the total antibacterial activity of the post‐incubates (the cell‐free culture supernatant obtained at the end of the fermentation process) of L. lactis CECT 539 against Carnobacterium piscicola CECT 4020 were very low.

title: Production of a potentially probiotic culture of Lactobacillus casei subsp. casei CECT 4043 in whey
abstract: Batch and fed-batch fermentations for the production of a highly concentrated culture of Lactobacillus casei CECT 4043 were carried out in whey-based media. In batch fermentation, the maximum concentrations of biomass (0.33 g L −1 ) and antibacterial activity (3.42 AU mL −1 ) were obtained in diluted whey medium (2% w/v lactose, initial pH 7.0) after 12 h. Based on these results, biomass and antimicrobial metabolites production in two re-alkalized fed-batch fermentations were carried out in diluted whey medium. The first fed-batch culture was fed with concentrated whey (5% w/v lactose) and a 400 g L −1 concentrated lactose solution. The second culture, which was fed with a concentrated mussel processing waste (10% w/v glucose) and a 310 g L −1 concentrated glucose solution, provided the highest concentrations of biomass (1.7 g L −1 ), viable cells (∼1.7 × 10 10  cfu mL −1 ), lactic acid (27.8 g L −1 ), acetic acid (3.9 g L −1 ) and antibacterial activity (74.2 AU mL −1 ).

title: Modelling the fed-batch production of pediocin using mussel processing wastes
abstract: Cell growth and pediocin production by Pediococcus acidilactici NRRL B-5627 were compared using batch (on MRS broth and a culture medium from mussel processing wastes (MPW)) and two fed-batch fermentations on MPW with re-alkalization cycles. This last fermentation technique yielded the highest biomass and pediocin productions as compared to batch fermentations. Mathematical models were set up to describe fed-batch production of biomass and pediocin by P. acidilactici. While cell growth was dependent on pH change, nitrogen and phosphorous availability and product inhibition (lactic acid, ethanol and butane-2,3-diol), pediocin production was dependent on both growth and the final pH reached in each re-alkalization period. The models developed offer a better fit and a more realistic description of the experimental biomass and pediocin production data than the logistic and Luedeking and Piret model. Therefore, they could be used to design feeding strategies for enhancing and controlling fed-batch pediocin production.

title: Fed-batch pediocin production on whey using different feeding media
abstract: Pediocin production by Pediococcus acidilactici NRRL B-5627 was followed in both batch and re-alkalized fed-batch fermentations on diluted whey (DW) supplemented with 2% (w/v) yeast extract (DWYE2 medium). In the first fed-batch culture, the fermentor was fed with a mixture of a 400 g/L concentrated lactose and concentrated whey (CW) supplemented with 2% (w/v) yeast extract. The maximum pediocin concentration obtained was only 1.2-fold higher than in batch fermentation. Since supplementing the DW with increasing amounts of glucose inhibited pediocin production, a second re-alkalized fed-batch culture was carried by using a 400 g/L concentrated glucose as feeding substrate. The pediocin concentration obtained was 2.2-fold higher than that in the first fed-batch culture. The third fed-batch fermentation was carried out by feeding the growing culture with a mixture of a 400 g/L concentrated glucose and CW medium supplemented with 4% (w/v) yeast extract. The concentration of biomass and the volumetric pediocin productivity obtained in this last culture were higher than those obtained in both the batch and the two fed-batch cultures (P < 0.05). Therefore, the use of feeding substrates containing glucose instead of lactose could be an appropriate alternative for increasing fed-batch production of pediocin in DWYE2 medium.

title: Continuous nisin production in laboratory media and whey permeate by immobilized Lactococcus lactis
abstract: Continuous production of nisin in laboratory media and whey permeate was investigated using a packed-bed bioreactor. Lactococcus lactis subsp. lactis ATCC 11454 was immobilized by natural attachment to fibre surfaces and entrapment in the void volume within spiral wound fibrous matrix. The inoculated bioreactor was fed M17 medium and effect of pH, temperature, dilution rate, and substrate concentrations on nisin production was examined. Bioreactor performance was monitored by checking cell density, lactic acid production, lactose consumption, and nisin activity. Optimal nisin activity was observed at pH 5.5, 31 °C, 0.2–0.3/h dilution rate, and 30 g/l lactose in M17. The maximum nisin titre was 2.6×10 4  AU/ml against a nisin sensitive strain, Lactobacillus leichmannii ATCC 4797. The bioreactor was fed whey permeate, supplemented with casein hydrolysate, and growth of L. lactis and associated nisin production were monitored. Optimal conditions for continuous nisin production in whey permeate were pH 5.5, 31 °C, 10–20 g/l casein hydrolysate, and 0.2/h dilution rate. Under these conditions, a maximum nisin titre of 5.1×10 4  AU/ml was observed. The bioreactor was operated continuously for 6 months without encountering any clogging, degeneration, or contamination problems. The cell density in the bioreactor was 52.0 g/l with 96.4% of the cells immobilized. This study illustrates the possibility of continuous production of high concentration of bacteriocins by lactic acid bacteria for use as food biopreservatives.

title: The influence of pH and growth rate on production of the bacteriocin, bavaricin MN, in batch and continuous fermentations
abstract: Bavaricin MN, a bacteriocin produced by Lactobacillus bavaricus MN, reached titres of 2000 AU ml-1 in APT broth maintained at pH 6.0, 30°C in a batch fermenter. Levels of bavaricin MN at pH 5.5 and 6.5 were lower despite comparable levels of producer cells. The addition of 3.0 g l-1 beef extract to APT broth resulted in increases in both the growth rate of the culture and the production of bavaracin MN. The titre of bavaricin MN in batch fermenters controlled at pH 6.0 in APT broth plus 3 g l-1 beef extract reached 3200 AU ml-1 at 30°C. This level was reduced to 800 AU ml-1 by 76 h. Glucose-limited continuous culture of Lact. bavaricus MN under the same conditions resulted in an increase in the titre of bavaricin MN to 6400 AU ml-1. This level was maintained, independent of growth rate, for 345 h. Growth rates of 0.205, 0.118, 0.169 and 0.058 h-1 were examined.

title: Effects of aeration and pH gradient on nisin production.: A mathematical model
abstract: A study on the effects of aeration and pH on nisin production by Lactococcus lactis showed significant increases as either aeration or pH drop gradient were risen. Nisin production at the maximum biomass point quadrupled when the oxygen saturation percentage increased from 50 to 100%. This suggests the association of this bioproduction with an oxidative metabolic pathway. A procedure based on successive re-alkalizations of the culture with feeding with glucose at regular intervals was proposed. It was shown that pH drop gradient (VpH) enhances nisin production, which increased approximately four-fold, and increases the efficiency of nutrient consumption. By using conventional equations of microbial kinetics, with the only additional assumption that VpH has some effect on the growth rate of the microorganism, a model that describes the results successfully is proposed. This model enables nisin to be typified as a primary metabolite, which tends to acquire secondary nature when the productive period of the culture is prolonged by imposing a stepwise-pH profile.

title: Variable Volume Fed-Batch Fermentation for Nisin Production by Lactococcus lactis subsp. lactis W28
abstract: A feeding technology that was suitable for improving the nisin production by Lactococcus lactis subsp. lactis W28 was established. The effects of initial sucrose concentration (ISC) in the fermentation broth, feeding time, and feeding rate on the fermentation were studied. It was observed that a fed-batch culture (ISC = 10 g l−1) with 100 ml sucrose solution (190 g l−1) being evenly fed (9–10 ml h−1) into the fermenter after 3-h fermentation gave the best performance in terms of biomass and nisin yield. Under these conditions, the total biomass and the total nisin yield were approximately 23% and 51% higher than those in batch fermentation, respectively. When the sucrose concentration was controlled at 5–10 g l−1 in variable volume intermittent fed-batch fermentation (VVIF) with ISC = 10 g l−1, the total biomass and the total nisin yield were 29% and 60% above those in batch fermentation, respectively. The VVIF proved to be effective to eliminate the substrate inhibition by maintaining sucrose at appropriate levels. It is also easy to be scaled up, since various parameters involved in industrial production were taken into account.

title: Bacteriocin Production with Lactobacillus amylovorus DCE 471 Is Improved and Stabilized by Fed-Batch Fermentation
abstract: Amylovorin L471 is a small, heat-stable, and hydrophobic bacteriocin produced by Lactobacillus amylovorus DCE 471. The nutritional requirements for amylovorin L471 production were studied with fed-batch fermentations. A twofold increase in bacteriocin titer was obtained when substrate addition was controlled by the acidification rate of the culture, compared with the titers reached with constant substrate addition or pH-controlled batch cultures carried out under the same conditions. An interesting feature of fed-batch cultures observed under certain culture conditions (constant feed rate) is the apparent stabilization of bacteriocin activity after obtaining maximum production. Finally, a mathematical model was set up to simulate cell growth, glucose and complex nitrogen source consumption, and lactic acid and bacteriocin production kinetics. The model showed that bacterial growth was dependent on both the energy and the complex nitrogen source. Bacteriocin production was growth associated, with a simultaneous bacteriocin adsorption on the producer cells dependent on the lactic acid accumulated and hence the viability of the cells. Both bacteriocin production and adsorption were inhibited by high concentrations of the complex nitrogen source.

title: Influence of the phosphorus and nitrogen source on nisin production in Lactococcus lactis subsp. lactis batch fermentations using a complex medium
abstract: The influence of different phosphorus and nitrogen sources on Lactococcus lactis subsp. lactis NIZO 22186 growth and nisin production was studied in batch fermentations using a complex medium. KH2PO4 was found to be the best phosphorus source for nisin production. Increasing initial phosphate concentrations from 0 to 5% KH2PO4 exerted a double effect, creating favourable pH conditions and particularly stimulating the nisin production levels, which were highest at 5% KH2PO4. Up to now, no such high initial phosphate concentrations have been reported for the production of other antibiotics or bacteriocins. Nisin, a lanthionine-containing peptide antibiotic with bacteriocin properties, clearly behaved as a primary metabolite, since its formation was linked with active growth and was not suppressed by phosphate concentrations up to 5%. A complex medium supplemented with cotton seed meal as nitrogen source also gave very high nisin yields.

title: Nisin and pediocin production on mussel‐processing waste supplemented with glucose and five nitrogen sources
abstract: Optimization of bacteriocin production by L. lactis subsp. lactis CECT 539 and Ped. acidilactici NRRL B‐5627 on mussel‐processing wastes.

title: Influence of pH, lactose and lactic acid on the growth of Streptococcus cremoris: a kinetic study
abstract: SummaryThe effect of various culture conditions on growth kinetics of an homofermentative strain of the lactic acid bacterium Streptococcus cremoris were investigated in batch cultures, in order to facilitate the production of this organism as a starter culture for the dairy industry. An optimal pH range of 6.3–6.9 was found and a lactose concentration of 37 g·l-1 was shown to be sufficient to cover the energetic demand for biomass formation, using the recommended medium. The study of the effect of lactic acid concentration on growth kinetics revealed that the end-product was not the sole factor affecting growth. The strain was characterized for its tolerance towards lactic acid and a critical concentration of 70 g·l-1 demonstrated. With the product yield of 0.9 g·g-1 at non-lactose limiting conditions the lactic acid concentration of 33 g·l-1 could not explain the low growth rates obtained, implicating a nutritional limitation.

title: Relation of growth of Streptococcus lactis and Streptococcus cremoris to amino acid transport
abstract: The maximum specific growth rate of Streptococcus lactis and Streptococcus cremoris on synthetic medium containing glutamate but no glutamine decreases rapidly above pH 7. Growth of these organisms is extended to pH values in excess of 8 in the presence of glutamine. These results can be explained by the kinetic properties of glutamate and glutamine transport (B. Poolman, E. J. Smid, and W. N. Konings, J. Bacteriol. 169:2755-2761, 1987). At alkaline pH the rate of growth in the absence of glutamine is limited by the capacity to accumulate glutamate due to the decreased availability of glutamic acid, the transported species of the glutamate-glutamine transport system. Kinetic analysis of leucine and valine transport shows that the maximal rate of uptake of these amino acids by the branched-chain amino acid transport system is 10 times higher in S. lactis cells grown on synthetic medium containing amino acids than in cells grown in complex broth. For cells grown on synthetic medium, the maximal rate of transport exceeds by about 5 times the requirements at maximum specific growth rates for leucine, isoleucine, and valine (on the basis of the amino acid composition of the cell). The maximal rate of phenylalanine uptake by the aromatic amino acid transport system is in small excess of the requirement for this amino acid at maximum specific growth rates. Analysis of the internal amino acid pools of chemostat-grown cells indicates that passive influx of (some) aromatic amino acids may contribute to the net uptake at high dilution rates.

title: Novel method to extract large amounts of bacteriocins from lactic acid bacteria
abstract: Antimicrobial peptides, bacteriocins, produced by lactic acid bacteria were adsorbed on the cells of producing strains and other gram-positive bacteria. pH was a crucial factor in determining the degree of adsorption of these peptides onto cell surfaces. In general, between 93 and 100% of the bacteriocin molecules were adsorbed at pHs near 6.0, and the lowest (< or = 5%) adsorption took place at pH 1.5 to 2.0. On the basis of this property, a novel isolation method was developed for bacteriocins from four genera of lactic acid bacteria. By using this method we made preparations of pediocin AcH, nisin, sakacin A, and leuconocin Lcm1 that were potent and concentrated. This method produced a higher yield than isolation procedures, which rely on precipitation of the bacteriocins from the cell-free culture liquor. It is simple and can be used to produce large quantities of bacteriocins from lactic acid bacteria to be used as food biopreservatives.

title: Factors influencing production of bacteriocins by lactic acid bacteria
abstract: Bacteriocins of lactic acid bacteria have potential for use as food biopreservatives to control spoilage and pathogenic bacteria. For economical use in food, the bacteriocins have to be produced in large amounts and preferably by growing the strains in media containing food grade ingredients. The influence of several factors on production of nisin by Lactococcus lactis, pediocin AcH by Pediococcus acidilactici, leuconocin Lcm1 by Leuconostoc carnosum Lm1 and sakacin A by Lactobacillus sake Lb 706 were studied. Among the several strains studied in each species, only some strains produced higher amounts of the bacteriocins. Production of a bacteriocin in a simple medium can be increased by growing the cells at optimum pH and supplementing with nutrients specific for a species/strain. Conditions that provide high cell density favor high bacteriocin production. Growing cells under an optimum environment for 16 h facilitates high bacteriocin production. For high yield of a bacteriocin, these aspects need to be considered.

title: Influence of pH drop on both nisin and pediocin production by Lactococcus lactis and Pediococcus acidilactici
abstract: To develop a kinetic model for describing the specific effect of pH drop on nisin and pediocin production in whey.

title: Lantibiotic nisin Z fermentative production by Lactococcus lactis IO-1: relationship between production of the lantibiotic and lactate and cell growth
abstract: The influence of several parameters on the fermentative production of nisin Z by Lactococcus lactis IO-1 was studied. Considerable attention has been focused on the relationship between the primary metabolite production of bacteriocin and lactate and cell growth, which has so far not been clarified in detail. Production of nisin Z was optimal at 30°C and in the pH range 5.0–5.5. The addition of Ca2+ to the medium showed a stimulating effect on the production of nisin Z. A maximum activity of 3150 IU/ml was obtained during pH-controlled batch fermentation in the medium supplemented with 0.1 M CaCl2. It was about three times higher than that obtained under the optimal conditions for cell growth and lactic acid production.

title: A comparison of factors affecting the production of two bacteriocins from lactic acid bacteria
abstract: The effect of tryptone, yeast extract, Tween 80 and initial pH on the production of enterocin 1146 and lactocin D, two bacteriocins produced by lactic acid bacteria, was studied in a basal buffered medium (tryptone-yeast extract-tween, TYT) using factorial experiments and empirical modelling. Production of enterocin 1146 was affected by pH, yeast extract and Tween 80 and to a lesser degree, by the initial pH of the medium. On the basis of the predictions of the models developed, three TYT media (TYT10, TYT11 and TYT30) were designed to maximize bacteriocin production while minimizing the amount of peptides in the medium. Growth and bacteriocin production by Enterococcus faecium DPC 1146 (enterocin 1146), Lactococcus lactis subsp. lactis biovar diacetylactis DPC 3286 (lactocin D) and Lact. lactis subsp. cremoris LMG2130 (lactococcin A) was compared in TYT media and seven other culture media (Elliker lactic broth, M17, M17 dialysate, MRS, tryptose phosphate, tryptone yeast extract broth, yeast glucose Lemco broth). Bacteriocin production in TYT media was comparable with that in M17 and MRS, which had a higher peptide content. TYT30 allowed good production of enterocin 1146 and lactocin D while TYT11 proved acceptable for all the strains tested.

title: An empirical model for describing the effects of nitrogen sources on nisin production
abstract: A study on the effects of tryptone and yeast extract on nisin production by Lactococcus lactis was carried out using a second order rotatable factorial design.

title: Control of fed-batch fermentations.
abstract: Fed-batch fermentation is used to prevent or reduce substrate-associated growth inhibition by controlling nutrient supply. Here we review the advances in control of fed-batch fermentations. Simple exponential feeding and inferential methods are examined, as are newer methods based on fuzzy control and neural networks. Considerable interest has developed in these more advanced methods that hold promise for optimizing fed-batch techniques for complex fermentation systems.

title: Dynamic mathematical models to describe the growth and nisin production by Lactococcus lactis subsp. lactis CECT 539 in both batch and re-alkalized fed-batch cultures
abstract: The growth and bacteriocin production by Lactococcus lactis subsp. lactis CECT 539 was studied in batch and in re-alkalized fed-batch fermentations in media prepared with whey and mussel-processing wastes. From these cultures, mathematical models were developed to describe the growth and nisin production. The growth model developed which was based on Monod kinetics, has clearly more mechanistic approach and provides a major biological interpretability of the parameters than the logistic growth equation. This is the first study presenting a pseudomechanistic model to describe the growth kinetics of lactic acid bacteria in re-alkalized fed-batch fermentations. Nisin production was modeled with a modified form of the Luedeking and Piret model, which includes a term for the effect of the optimal final pH value in bacteriocin synthesis. Both models were demonstrated for three fed-batch cultivations of L. lactis using different culture media, feeding substrates and times of re-alkalization and feeding.

title: A kinetic study of the lactic acid fermentation. Batch process at controlled pH. Reprinted from Journal of Biochemical and Microbiological Technology Engineering Vol. I, No. 4. Pages 393-412 (1959).
abstract: Kinetic data are needed to develop basic understanding of fermentation processes and to permit rational design of continuous fermentation processes. The kinetics of the fermentation of glucose to lactic acid have been studied at six constant pH levels between 4. 5 and 6.0 by measuring the instantaneous rates of bacterial growth and of lactic acid formation throughout each fermentation. It was found that the instantaneous rate of acid formation dP/dt, should be related to the instantaneous rate of bacterial growth dN/dt, and to the bacterial density N, throughout a fermentation at a given pH, by the expression when the constants alpha and beta are determined by the pH of the fermentation.

title: Substrate inhibition of Pediococcus acidilactici by glucose on a waste medium. Simulations and experimental results
abstract: The possibility of substrate inhibition by glucose on biomass and pediocin production was studied in cultures of Pediococcus acidilactici on a residual medium.

title: Growth of the Bacteriocin-ProducingLactobacillus sakei Strain CTC 494 in MRS Broth Is Strongly Reduced Due to Nutrient Exhaustion: a Nutrient Depletion Model for the Growth of Lactic Acid Bacteria
abstract: Although commercial MRS broth has been designed to allow excellent growth of lactobacilli, most of these bacteria are still subjected to a self-inhibiting process. The most likely explanation is the accumulation of lactic acid or other toxic end products and the depletion of nutrients. In this study, the self-inhibition ofLactobacillus sakei CTC 494 was analyzed in a kinetic way, and a nutrient depletion model was set up to describe the growth inhibition process. This simple model has considerable advantages compared to commonly used descriptive models such as the logistic growth equation. It offers a better fit and a more realistic description of the growth data by taking into account both growth inhibition due to lactic acid production and changes in growth rates due to nutrient depletion. Depending on the fermentation conditions, in MRS broth there appears to be a strong decrease of the specific growth rate over time. Some undefined compounds present in the complex nitrogen source of MRS broth appear to be of crucial importance because of their limited availability. Moreover, nutrient availability affects bacteriocin production through its effect on cell growth as well as on the bacteriocin production per cell. A plateau value for the bacteriocin production by L. sakei CTC 494 was observed.

title: Modelling the Biphasic Growth and Product Formation by Enterococcus faecium CECT 410 in Realkalized Fed-Batch Fermentations in Whey
abstract: The influence of initial pH on growth and nutrient (total sugars, nitrogen, and phosphorous) consumption by Enterococcus faecium CECT 410 was studied during batch cultures in whey. With these data, two realkalized fed-batch fermentations were developed using different feeding substrates. The shift from homolactic to mixed acid fermentation, the biphasic kinetics observed for cell growth and nitrogen consumption and the increase in the concentrations of biomass and products (lactic acid, acetic acid, ethanol, and butane-2,3-diol) were the most noteworthy observations of these cultures. Modelling the fed-batch growth of Ent. faecium with the Logistic and bi-Logistic models was not satisfactory. However, biomass production was best mathematically described with the use of a double Monod model, which was expressed in terms of biomass, product accumulation, and nitrogen utilization. Product formation was successfully modelled with a modified form of the Luedeking and Piret model developed in this study.