Probiotics in intestinal and non-intestinal infectious diseases--clinical evidence.

There is increasing evidence that certain probiotic strains can be useful in improving human health. The use of probiotics has received attention as a natural way of restoring body's normal microbiota, and an alternative and inexpensive way of preventing or treating infectious diseases without side effects. The best-documented clinical application of probiotics comes from trials on the treatment of gastrointestinal infections, mainly infectious diarrhoea. The enhancement of local as well as systemic immune responses by probiotics also offers new opportunities for probiotics in preventing infections at distal mucosal surfaces, such as those in the oral cavity, respiratory and urogenital tracts. The underlying mechanisms of probiotics are still unclear, but may include strengthening of the non-immunological gut barrier, interference with pathogen adhesion and growth inhibition, and the enhancement of the local mucosal immune system in the gut, as well as of the systemic immune response.

Lactobacillus supplementation for diarrhoea related to chemotherapy of colorectal cancer: a randomised study.

5-Fluorouracil (5-FU)-based chemotherapy is frequently associated with diarrhoea. We compared two 5-FU-based regimens and the effect of Lactobacillus and fibre supplementation on treatment tolerability. Patients diagnosed with colorectal cancer (n=150) were randomly allocated to receive monthly 5-FU and leucovorin bolus injections (the Mayo regimen) or a bimonthly 5-FU bolus plus continuous infusion (the simplified de Gramont regimen) for 24 weeks as postoperative adjuvant therapy. On the basis of random allocation, the study participants did or did not receive Lactobacillus rhamnosus GG supplementation (1-2 x 10(10) per day) and fibre (11 g guar gum per day) during chemotherapy. Patients who received Lactobacillus had less grade 3 or 4 diarrhoea (22 vs 37%, P=0.027), reported less abdominal discomfort, needed less hospital care and had fewer chemotherapy dose reductions due to bowel toxicity. No Lactobacillus-related toxicity was detected. Guar gum supplementation had no influence on chemotherapy tolerability. The simplified de Gramont regimen was associated with fewer grade 3 or 4 adverse effects than the Mayo regimen (45 vs 89%), and with less diarrhoea. We conclude that Lactobacillus GG supplementation is well tolerated and may reduce the frequency of severe diarrhoea and abdominal discomfort related to 5-FU-based chemotherapy.

Lactobacillus bacteremia, species identification, and antimicrobial susceptibility of 85 blood isolates.

BACKGROUND: Data regarding antimicrobial susceptibility of clinical Lactobacillus isolates are scarce, and appropriate interpretation criteria for susceptibility tests are not available. METHODS: We examined 85 cases of Lactobacillus bacteremia, of which 47 cases have been included in our previous studies. Overall, 14 antimicrobial agents were evaluated by the E-test method, and these results were compared with disk diffusion test findings. The clinical outcomes of the patients and their antimicrobial treatments were registered. RESULTS: The antimicrobial susceptibility of Lactobacillus strains was species dependent. The considerable number of Lactobacillus rhamnosus (n=46), Lactobacillus fermentum (n=12), and Lactobacillus casei (n=12) strains available for testing made it possible to compare the susceptibilities within 1 species, as well. Of the 46 L. rhamnosus isolates, 22 were identified as L. rhamnosus GG type by pulsed-field gel electrophoresis. All Lactobacillus isolates demonstrated low minimum inhibitory concentrations (MICs) of imipenem, piperacillin-tazobactam, erythromycin, and clindamycin. MICs of vancomycin were high (>256 microg/mL) for all other species except Lactobacillus gasseri and Lactobacillus jensenii. Disk diffusion and E-test results were concordant. The MICs of cephalosporins varied; cefuroxime demonstrated a higher level of activity than did ceftriaxone. Benzylpenicillin and ampicillin MICs had variable ranges between different species. Combination therapy was given to 83% of the patients, but, in 54% of them, therapy included only 1 microbiologically active agent, according to results of the susceptibility tests. Mortality at 1 week was 12% among patients who presumably were receiving adequate treatment and 27% among patients who were receiving inadequate treatment (P=.131, by E-test). CONCLUSION: Most clinical Lactobacillus blood isolates demonstrated low MICs of imipenem, piperacillin-tazobactam, erythromycin, and clindamycin, but they had variable susceptibility to penicillin and cephalosporins.

A combination of fibre-rich rye bread and yoghurt containing Lactobacillus GG improves bowel function in women with self-reported constipation.

OBJECTIVE: The aim of the study was to investigate the effects of fibre-rich rye bread and yoghurt containing Lactobacillus GG (LGG) on intestinal transit time and bowel function, and to test whether they have an interaction in cases of self-reported constipation. DESIGN: The study was carried out as a two-by-two factorial design. SETTING: Free-living subjects. SUBJECTS: A total of 59 healthy women with self-reported constipation, recruited by advertisement. INTERVENTIONS: After a baseline period, the subjects were randomized into four diet groups: (1) rye bread+LGG yoghurt, (2) rye bread, (3) LGG yoghurt, and (4) control. The 3-week dietary intervention was followed by a 3-week follow-up period. During each period, total intestinal transit time was measured and the subjects recorded faecal frequency and consistency, difficulty in defecation and gastrointestinal symptoms. RESULTS: The rye bread shortened total intestinal transit time (mean difference, -0.7; CI(95), -1.1 to -0.2; P=0.007), increased faecal frequency (0.3; CI(95), 0.1 to 0.5; P=0.001), softened faeces (-0.3; CI(95), -0.4 to -0.2; P<0.001) and made defecation easier (-0.4; CI(95), -0.5 to -0.2; P<0.001), but also increased gastrointestinal symptoms (1.6; CI(95), 0.7 to 2.4; P<0.001) compared to the low-fibre toast consumed in the LGG and control groups. There were fewer symptoms in the rye bread+LGG group compared to the rye bread group (-1.3; CI(95), -2.4 to -0.2; P=0.027). CONCLUSIONS: Fibre-rich rye bread can be recommended in the treatment of constipation, and the simultaneous consumption of LGG yoghurt relieves the adverse gastrointestinal effects associated with increased intake of fibre. SPONSORSHIP: Valio Ltd, R&D, and Fazer Bakeries Ltd.

Effects of probiotic therapy on the activity and activation of mild rheumatoid arthritis--a pilot study.

OBJECTIVE: To study the effects of Lactobacillus rhamnosus GG (LGG) on rheumatoid arthritis (RA). METHODS: Twenty-one RA patients were randomised to receive 2 capsules of LGG or a placebo twice daily in double-blind fashion for 12 months. Arthritis activity was evaluated by clinical examination, HAQ index, and laboratory tests (e.g. ESR, CRP, pro- and anti-inflammatory cytokines). RESULTS: There were no statistical differences in the clinical parameters, biochemical variables and HAQ index between the study groups over the intervention period. The mean number of tender and swollen joints decreased from 8.3 to 4.6 in the Lactobacillus group and from 5.5 to 4.8 in the placebo group (p = 0.41). According to the global assessment the RA activity was reduced in 71% (LGG group) vs. 30% (controls) (p = 0.15). Serum IL-1 beta increased slightly in the LGG group (p = 0.07), but no differences were seen in IL-6, TNF-alpha, MPO, IL-10 or 1L-12. CONCLUSIONS: Although there were no statistical significant differences in the activity of RA, more subjects in the LGG group reported subjective well being. More studies on the effects of probiotic bacteria in RA are needed.

Effect of Lactobacillus rhamnosus GG on ileal pouch inflammation and microbial flora.

BACKGROUND: Preliminary trials of probiotics in preventing recurrent chronic pouchitis have been encouraging. AIM: To investigate the efficacy of Lactobacillus GG supplementation as primary therapy for ileal pouch inflammation, and its effect on the microbial flora. METHODS: Twenty patients, with a previous history of pouchitis and endoscopic inflammation, were recruited for a prospective, randomized, double-blind, placebo-controlled trial of Lactobacillus GG supplementation (10 LGG, 10 placebo) in two gelatine capsules [(0.5-1) x 10(10) colony-forming units/capsule] b.d. for 3 months. Quantitative bacterial culture of fresh faecal samples and biopsies taken from the pouch and afferent limb was performed before and after supplementation. RESULTS: Lactobacillus GG supplementation changed the pouch intestinal flora by increasing the ratio of total faecal lactobacilli to total faecal anaerobes (P = 0.03) and enhancing the frequency of lactobacilli-positive cultures in the pouch and afferent limb mucosal biopsy samples. However, only 40% of patients were colonized with Lactobacillus GG. No differences were observed between the groups with regard to the mean pouchitis disease activity index or the total anaerobes or aerobes of faecal or tissue biopsy samples. CONCLUSIONS: A single-strain probiotic bacterium supplement of Lactobacillus GG changed the pouch intestinal bacterial flora, but was ineffective as primary therapy for a clinical or endoscopic response. More clinical trials are needed to evaluate the right placement and dosage of probiotics within a treatment regimen for pouchitis.

Induction of nitric oxide synthesis by probiotic Lactobacillus rhamnosus GG in J774 macrophages and human T84 intestinal epithelial cells.

BACKGROUNDS AND AIMS: Probiotic Lactobacillus rhamnosus GG (LGG) has proved to be beneficial in the treatment of viral- and antibiotic-associated diarrhea but the mechanisms of action remain unknown. Nitric oxide (NO) is involved in the protective mechanisms in the gastrointestinal tract and may contribute to some of the beneficial effects of probiotics. The aim of the present study was to investigate if induction of NO synthesis is involved in the cellular actions of LGG. METHODS: NO synthesis and its regulation were measured in cultures of J774 murine macrophages and human T84 colon epithelial cells. NO production was measured as its metabolite nitrite accumulated into the culture medium. Inducible nitric oxide synthase (iNOS) protein and iNOS mRNA were detected by Western blot and RT-PCR, respectively. RESULTS: In J774 macrophages, LGG induced a low level production of NO in the presence of gamma interferon (IFNgamma) and it was inhibited by NOS inhibitors, cycloheximide and by a NF-kappa B inhibitor pyrrolidinedithiocarbamate. Accordingly, LGG and IFNgamma-stimulation increased iNOS mRNA and protein levels. T84 cells produced NO in response to LGG when first primed with a combination of IL-1beta, TNFalpha and IFNgamma. Lipoteichoic acid (LA), an antigenic structure in gram-positive bacteria, also induced NO formation in J774 cells in the presence of IFNgamma suggesting that LA may be the active component in LGG. CONCLUSIONS: LGG induces NO production in J774 macrophages and in human T84 colon epithelial cells through induction of iNOS by a mechanism involving activation of transcription factor NF-kappaB. Induction of iNOS and low-level synthesis of NO may be involved in the protective actions of LGG in the gastrointestinal tract.

Good adhesion properties of probiotics: a potential risk for bacteremia?

The ability to adhere to human intestinal mucus was tested for lactic acid bacteria of clinical blood culture, human fecal and dairy origin. The blood culture isolates were found to adhere better than the dairy strains. Of the Lactobacillus rhamnosus strains (nine clinical, 10 fecal and three dairy), blood culture isolates adhered better than the fecal strains. Although these results indicate a trend for blood culture isolates to bind to intestinal mucus in higher numbers than strains of dairy and human fecal origin, other factors are also likely to be involved in the etiology of lactobacillemia since some of the clinical Lactobacillus isolates exhibited a relatively low level of adhesion.

Effect of long term consumption of probiotic milk on infections in children attending day care centres: double blind, randomised trial.

OBJECTIVE: To examine whether long term consumption of a probiotic milk could reduce gastrointestinal and respiratory infections in children in day care centres. DESIGN: Randomised, double blind, placebo controlled study over seven months. SETTING: 18 day care centres in Helsinki, Finland. PARTICIPANTS: 571 healthy children aged 1-6 years: 282 (mean (SD) age 4.6 (1.5) years) in the intervention group and 289 (mean (SD) age 4.4 (1.5) years) in the control group. INTERVENTION: Milk with or without Lactobacillus GG. Average daily consumption of milk in both groups was 260 ml. MAIN OUTCOME MEASURES: Number of days with respiratory and gastrointestinal symptoms, absences from day care because of illness, respiratory tract infections diagnosed by a doctor, and course of antibiotics. RESULTS: Children in the Lactobacillus group had fewer days of absence from day care because of illness (4.9 (95% confidence interval 4.4 to 5.5) v 5.8 (5.3 to 6.4) days, 16% difference, P=0.03; age adjusted 5.1 (4.6 to 5.6) v 5.7 (5.2 to 6.3) days, 11% difference, P=0.09). There was also a relative reduction of 17% in the number of children suffering from respiratory infections with complications and lower respiratory tract infections (unadjusted absolute % reduction -8.6 (-17.2 to -0.1), P=0.05; age adjusted odds ratio 0.75 (0.52 to 1.09), P=0.13) and a 19% relative reduction in antibiotic treatments for respiratory infection (unadjusted absolute % reduction -9.6 (-18.2 to -1.0), P=0.03; adjusted odds ratio 0.72 (0.50 to 1.03), P=0.08) in the Lactobacillus group. CONCLUSIONS: Lactobacillus GG may reduce respiratory infections and their severity among children in day care. The effects of the probiotic Lactobacillus GG were modest but consistently in the same direction.

Effect of long-term consumption of a probiotic bacterium, Lactobacillus rhamnosus GG, in milk on dental caries and caries risk in children.

Lactobacillus rhamnosus GG, ATCC (LGG), has shown antagonism to many bacteria including mutans streptococci. This randomized, double-blind, placebo-controlled intervention study was designed to examine whether milk containing LGG has an effect on caries and the risk of caries in children when compared with normal milk. 594 children, 1-6 years old, from 18 municipal day-care centres were included. The children received the milk with meals from coded containers 5 days a week in the day-care centres for 7 months. The children's oral health was recorded at baseline and at the end, using WHO criteria. The caries risk was calculated based on clinical and microbiological data, comprising mutans streptococcus levels from dental plaque and saliva. The risk was classified as high if the child had a dmft/DMFT or initial caries score >0, and a mutans streptococcus count > or = 10(5) CFU/ml. The results showed less dental caries in the LGG group and lower mutans streptococcus counts at the end of the study. LGG was found to reduce the risk of caries significantly (OR = 0.56, p = 0.01; controlled for age and gender, OR = 0.51, p = 0.004). The effect was particularly clear in the 3- to 4-year-olds. Thus, milk containing the probiotic LGG bacteria may have beneficial effects on children's dental health.

Probiotic bacteria and intestinal health: new methods of investigation.

This paper highlights some new methods in the probiotic research based on the use of colonic biopsies and molecular biological techniques for strain identification.

Antiproliferative effects of homogenates derived from five strains of candidate probiotic bacteria.

Unheated and heat-treated homogenates were separately prepared from candidate probiotic bacteria, including Lactobacillus rhamnosus GG, Bifidobacterium lactis, Lactobacillus acidophilus, Lactobacillus delbrueckii subsp. bulgaricus, and Streptococcus thermophilus. We compared the phytohemagglutinin-induced proliferation of mononuclear cells in the presence of homogenates and in the presence of a control containing no homogenate by assessing thymidine incorporation in cell cultures. All homogenates suppressed proliferation, whether the enzymatic activity was inactivated or not inactivated by heating. When the proliferation assays were repeated with cytoplasmic and cell wall extracts derived from the homogenate of L. rhamnosus GG, the cytoplasmic extract but not the cell wall extract was suppressive. These findings indicate that candidate probiotic bacteria possess a heat-stable antiproliferative component(s). These bacteria may be used to generate microbiologically nonviable yet immunologically active probiotic food products that are easier to store and have a longer shelf life.

Comparison of ribotyping, randomly amplified polymorphic DNA analysis, and pulsed-field gel electrophoresis in typing of Lactobacillus rhamnosus and L. casei strains.

A total of 24 strains, biochemically identified as members of the Lactobacillus casei group, were identified by PCR with species-specific primers. The same set of strains was typed by randomly amplified polymorphic DNA (RAPD) analysis, ribotyping, and pulsed-field gel electrophoresis (PFGE) in order to compare the discriminatory power of the methods. Species-specific primers for L. rhamnosus and L. casei identified the type strain L. rhamnosus ATCC 7469 and the neotype strain L. casei ATCC 334, respectively, but did not give any signal with the recently revived species L. zeae, which contains the type strain ATCC 15820 and the strain ATCC 393, which was previously classified as L. casei. Our results are in accordance with the suggested new classification of the L. casei group. Altogether, 21 of the 24 strains studied were identified with the species-specific primers. In strain typing, PFGE was the most discriminatory method, revealing 17 genotypes for the 24 strains studied. Ribotyping and RAPD analysis yielded 15 and 12 genotypes, respectively.

Persistence of colonization of human colonic mucosa by a probiotic strain, Lactobacillus rhamnosus GG, after oral consumption.

Lactobacillus rhamnosus GG is one of the most thoroughly studied probiotic strains. Its advantages in the treatment of gastrointestinal disorders are well documented. The aim of the present study was to demonstrate with colonic biopsies the attachment of strain GG to human intestinal mucosae and the persistence of the attachment after discontinuation of GG administration. A whey drink fermented with strain GG was fed to human volunteers for 12 days. Fecal samples were collected before, during, and after consumption. L. rhamnosus GG-like colonies were detected in both fecal and colonic biopsy samples. Strain GG was identified by its characteristic colony morphology, a lactose fermentation test, and PCR. This study showed that strain GG was able to attach in vivo to colonic mucosae and, although the attachment was temporary, to remain for more than a week after discontinuation of GG administration. The results demonstrate that the study of fecal samples alone is not sufficient in evaluating colonization by a probiotic strain.

Demonstration of safety of probiotics -- a review.

Probiotics are commonly defined as viable microorganisms (bacteria or yeasts) that exhibit a beneficial effect on the health of the host when they are ingested. They are used in foods, especially in fermented dairy products, but also in pharmaceutical preparations. The development of new probiotic strains aims at more active beneficial organisms. In the case of novel microorganisms and modified organisms the question of their safety and the risk to benefit ratio have to be assessed. Lactic acid bacteria (LAB) in foods have a long history of safe use. Members of the genera Lactococcus and Lactobacillus are most commonly given generally-recognised-as-safe (GRAS) status whilst members of the genera Streptococcus and Enterococcus and some other genera of LAB contain some opportunistic pathogens. Lactic acid bacteria are intrinsically resistant to many antibiotics. In many cases resistances are not, however, transmissible, and the species are also sensitive to many clinically used antibiotics even in the case of a lactic acid bacteria- associated opportunistic infection. Therefore no particular safety concern is associated with intrinsic type of resistance. Plasmid-associated antibiotic resistance, which occasionally occurs, is another matter because of the possibility of the resistance spreading to other, more harmful species and genera. The transmissible enterococcal resistance against glycopeptide antibiotics (vancomycin and teicoplanin) is particularly noteworthy, as vancomycin is one of the last effective antibiotics left in the treatment of certain multidrug-resistant pathogens. New species and more specific strains of probiotic bacteria are constantly identified. Prior to incorporating new strains into products their efficacy should be carefully assessed, and a case by case evaluation as to whether they share the safety status of traditional food-grade organisms should be made. The current documentation of adverse effects in the literature is reviewed. Future recommendations for the safety of already existing and new probiotics will be given.

Increased fecal frequency and gastrointestinal symptoms following ingestion of galacto-oligosaccharide-containing yogurt.

Gastrointestinal symptoms and fecal frequency following ingestion of yogurt containing 15 g of galacto-oligosaccharides (GOS) per d were observed in 12 healthy volunteers. The effect of GOS on intestinal microflora was also studied in six volunteers. Defecation frequency increased during the administration period, but gastrointestinal symptoms, especially flatulence, also increased. The level of fecal bifidobacteria did not increase by the yogurt intake, but a significant increase was observed in the fecal bacteria growing on MRS media. The results indicate that dietary GOS increase gastrointestinal symptoms and fecal frequency in normal adults and have an effect on intestinal microecosystem.

Lactobacillus rhamnosus GG (ATCC 53103) and platelet aggregation in vitro.

Lactobacillus rhamnosus GG is an experimentally and clinically well documented probiotic used in different dairy products. The present study aimed to investigate the safety aspects of Lactobacillus rhamnosus GG, particularly with respect to platelet aggregation, the initiating event in thrombosis. Platelet rich plasma was separated from the blood of healthy volunteers, and the effects of Lactobacillus rhamnosus GG (ATCC 53103), Lactobacillus rhamnosus (ATCC 7469) and Enterococcus faecium T2L6 in different dilutions on spontaneous, ADP- and adrenaline-induced aggregation were tested. The bacteria did not influence spontaneous aggregation. Only Enterococcus faecium T2L6 enhanced the adrenaline-induced aggregation, with a less clear effect on ADP-induced aggregation.