QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tzortzis, G. Articles by Gibson, G. R. Search for Related Content PubMed PubMed Citation Articles by Tzortzis, G. Articles by Gibson, G. R.

---

Nutrient Metabolism

A Novel Galactooligosaccharide Mixture Increases the Bifidobacterial Population Numbers in a Continuous In Vitro Fermentation System and in the Proximal Colonic Contents of Pigs In Vivo¹

School of Food Biosciences, The University of Reading, Whiteknights, Reading RG6 6AP, UK and ^* Institute of Food Research, Norwich Research Park, Colney, Norwich, NR4 7UA, UK

²To whom correspondence should be addressed. E-mail: g.tzortzis{at}reading.ac.uk.

	ABSTRACT

TOP ABSTRACT MATERIALS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Prebiotics are nondigestible food ingredients that encourage proliferation of selected groups of the colonic microflora, thereby altering the composition toward a more beneficial community. In the present study, the prebiotic potential of a novel galactooligosaccharide (GOS) mixture, produced by the activity of galactosyltransferases from Bifidobacterium bifidum 41171 on lactose, was assessed in vitro and in a parallel continuous randomized pig trial. In situ fluorescent hybridization with 16S rRNA-targeted probes was used to investigate changes in total bacteria, bifidobacteria, lactobacilli, bacteroides, and Clostridium histolyticum group in response to supplementing the novel GOS mixture. In a 3-stage continuous culture system, the bifidobacterial numbers for the first 2 vessels, which represented the proximal and traverse colon, increased (P < 0.05) after the addition of the oligosaccharide mixture. In addition, the oligosaccharide mixture strongly inhibited the attachment of enterohepatic Escherichia coli (P < 0.01) and Salmonella enterica serotype Typhimurium (P < 0.01) to HT29 cells. Addition of the novel mixture at 4% (wt:wt) to a commercial diet increased the density of bifidobacteria (P < 0.001) and the acetate concentration (P < 0.001), and decreased the pH (P < 0.001) compared with the control diet and the control diet supplemented with inulin, suggesting a great prebiotic potential for the novel oligosaccharide mixture.

KEY WORDS: • intestinal microflora • prebiotic • galactooligosaccharides

Probiotics, which are live microbial dietary additives thought to confer health advantages, have a long history of use in humans and animals (1). Although numerous publications exist showing that probiotics are active in the gut after ingestion (2,3), inconsistency in scientific reports related to health benefits in human studies are attributed mainly to survivability problems of the organisms after ingestion (4,5). To overcome this survivability issue, the concept of prebiotics was introduced. These are nondigestible food ingredients that beneficially affect the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the colon that can improve host health (6).

Although any dietary material that enters the large intestine, including resistant starch, dietary fibers, proteins, and lipids, can be considered as a candidate prebiotic, in practice, prebiotics are currently confined to oligosaccharides (7), which can induce a specific colonic fermentation by benign or potentially health-promoting indigenous bacteria, e.g., lactic acid–producing flora (5).

Among the various oligosaccharides tested for potential prebiotic application, galactooligosaccharides (GOS)³ have attracted interest, mainly because these are the compounds in human milk that have been associated with the improved colonic health of breast-fed infants (8).

Diets enriched with GOS increase populations of Bifidobacterium and Lactobacillus species (9) and their fermentation products in the colon. The latter are mainly SCFAs, which improve the energy supply to the colonic epithelium, and facilitate calcium and magnesium absorption (9). Moreover, glycoconjugates containing GOS have been implicated in interactions between epithelial and bacterial cells (10), implying that these compounds also have the capacity to inhibit the binding of pathogens to cell surfaces by acting as competitive receptors.

The aim of this work was to evaluate the potential of a novel GOS mixture to modulate the colonic microbiota in a beneficial way and possibly function as "decoy" oligosaccharide binding receptors for 3 gastrointestinal pathogens in vitro. In addition, the prebiotic potential of the mixture was studied in weaned pigs by monitoring microbial changes in the composition of the fecal and colonic microbiota, colonic pH, and colonic SCFA concentration.

	MATERIALS AND METHODS

TOP ABSTRACT MATERIALS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
The GOS mixture used in this study was produced from the activity of galactosyltransferases from Bifidobacterium bifidum NCIMB 41171 on lactose (11). The final product was in dry powder form consisting of (wt:wt) 45% lactose, 9.9% disaccharides [Gal (ß 1–3)- Glc; Gal (ß 1–3)- Gal; Gal (ß 1–6)- Gal; Gal ( 1–6)- Gal], 23.1% trisaccharides [Gal (ß 1–6)-Gal (ß 1–4)- Glc; Gal (ß 1–3)- Gal (ß 1–4)- Glc], 11.55% tetrasaccharides [Gal (ß 1–6)- Gal (ß 1–6)- Gal (ß 1–4)- Glc], and 10.45% pentasaccharides [Gal (ß 1–6)- Gal (ß 1–6)- Gal (ß 1–6)- Gal (ß 1–4)- Glc]. Oligosaccharide fractions for the adhesion assay were purified by gel filtration on a Biogel P2 (Pharmacia) column eluted at 3 mL/min with water. Inulin was supplied by Orafti; all other chemicals and media preparations used in these investigations were from Sigma and Oxoid.

    In vitro gut model. Physicochemical conditions in the colon were replicated in a 3-stage continuous fermenter, representing the proximal, transverse, and distal colon (12), inoculated with 10% (wt:v) fecal homogenate from healthy human volunteers in a growth medium without and with 1% (wt:v) of the synthesized GOS mixture. The growth medium contained the following ingredients: starch, 8 g/L; mucin (porcine gastric), 4 g/L; casein, 3 g/L; peptone water, 5 g/L; tryptone water, 5 g/L; bile #3, 0.4 g/L; yeast, 4.5 g/L; FeSO₄, 0.005 g/L; NaCl, 4.5 g/L; KCl, 4.5 g/L; KH₂PO₄, 0.5 g/L; MgSO₄ · 7H₂O, 1.25 g/L; CaCl₂ · 6H₂O, 0.15 g/L; NaHCO₃, 1.5 g/L; Tween, 80 1 mL; Hemin, 0.05 g/L; and cysteine · HCl, 0.8 g/L. The system was operated at a retention time of 36 h. After inoculation, the gut model was left overnight to equilibrate before the medium pump was switched on, and the system was run for a cycle of at least 10 d before medium containing GOS (6 g/d) was introduced for a further cycle of 10 d. Samples (5 mL) were removed at the beginning and end of each cycle.

    Bacterial enumerations by fluorescence in situ hybridization (FISH). Differences in bacterial populations were assessed by FISH using oligonucleotide probes labeled with the fluorescent dye Cy3 (Eurogentec UK) and designed to target specific diagnostic regions of bacterial 16S rRNA, as previously described (13). The molecular probes for specific bacterial genera used were Bac 303 for Bacteroides spp. (14), Bif 164 for Bifidobacterium spp. (15), Chis 150 for bacteria belonging to the Clostridium histolyticum group (16), and Lab 158 for Lactobacillus/Enterococcus spp. (17). Total bacterial counts were obtained by staining with the nucleic acid stain 4,6-diamidino-2-phenylindole.

    Oligosaccharide sensitivity assay. The HT29 cell line was obtained from European Collection of Cell Cultures for Applied Microbiology and Research. Cell stocks were cultured at 37°C in humidified 5% CO₂ in air in DMEM supplemented with 5% (v:v) fetal bovine serum, 0.1 mol/L penicillin, 0.1 mol/L streptomycin, nonessential amino acids (x100), and 0.2 mol/L -glutamine. Cells were refed every 48 h and passaged before confluence was reached. Serum standard medium (1% v:v) supplemented with different concentrations of oligosaccharides (0.01, 0.1, 1, 10, 100 mmol/L) was used for the oligosaccharide sensitivity assay as described by Olano-Martin et al. (18) to determine any toxic effects of the oligosaccharide mixture on the HT29 cells. The percentage of cell survival was calculated as: % survival = (mean absorbance of treated cells/mean absorbance of control) x 100.

    Adhesion assay. HT29 cells were grown in 12-well tissue culture plates to >90% confluence using the standard medium. For the final cell feeding before performing the assay, antibiotic-free medium was used. PBS (0.5 mL), containing each test oligosaccharide (10 mol/L), was added in 3 wells (PBS without any oligosaccharide was included as a control) and 0.5 mL of a 1/1000 dilution of the pathogen culture [enteropathogenic Escherichia coli (EPEC) O26 NCTC 08620, verocytotoxic E. coli (VTEC) O157:H7 VT^– PHLS NCO12900 or Salmonella typhimurium NCFIMB 10248] was added to all wells. The plates were placed on a rocker mixer and incubated aerobically at 37°C for 2 h. The culture was aspirated and after 3 washes with sterile PBS (1 mL/well), 70 µL trypsin/EDTA solution was added to each well, mixed, and allowed to stand for 5 min at 37°C. PBS (1 mL) was added to each well and mixed by pipette to ensure that all of the cells were removed from the bottom of the well and that clumps were broken up. Cell suspension (1 mL) was plated out on plate count agar and incubated at 37°C for 24 h. After incubation, the colonies were enumerated and the inhibition of adhesion was calculated as the ratio of bacteria (cfu/L) present in the sample compared with the control (PBS).

    Animals, diets and feeding study. Weaned male pigs (n = 40; mean live weight of 14.7 ± 2.6 kg; 28 d old) were purchased from JSR Genetics. On arrival, they were housed in 4 groups of 10 for 7 d to allow them time to settle after transport, and to acclimate to the unit and diet. During the acclimation period, pigs were fed the NEG diet⁴ (Deltawean 15 NGP pellets, ABN), a complete, antibiotic-free diet for growing pigs. After this period, the pigs were transferred to individual pens randomly grouped within the same unit. Pigs were bedded on sawdust throughout the study; straw was also provided as an environmental enrichment as were "toys" to help alleviate boredom. The study was conducted in accordance with the guidelines of the Ethics & Research Committee at the University of Reading.

After acclimation, pigs were fed 1 of the following 4 diets: NEG alone, NEG supplemented with the novel GOS mixture (1.6% by weight), the novel GOS mixture (4% by weight), or inulin (1.6% by weight). The novel GOS (55% wt:wt synthesized oligosaccharides) and inulin additives were weighed separately for each pig and stored in sealed mini-grip bags; they were added to the control feed at the time of the morning feeding. This ensured that each individual pig received the prescribed dose and eliminated potential variability due to inadequate mixing.

    Sample collection. The pigs were fed the dietary treatments for a mean of 34 d (33–35 d), after which they were killed by an injection of a lethal dose of sodium barbiturate (15 mL) into an ear vein. Each pig then underwent laparotomy and ties were placed on the large bowel, 2 cm downstream of the ileocecal junction and 2 cm upstream of the rectum. With minimum disturbance, the entire large intestine was removed as quickly as possible to a tray containing ice and transported to an adjacent laboratory. The proximal and distal sections of the large intestine were then freed from the mesentery and clamps placed to isolate the semiliquid contents of the proximal and distal colon. Samples of proximal and distal colonic contents were obtained for bacterial enumeration, SCFA, and pH measurement. Fecal samples from each pig were obtained at the beginning and at the end of the feeding period.

    Analysis of SCFA and lactic acid. Production of lactic, acetic, propionic, and butyric acids in the colonic contents was quantified. Samples were centrifuged at 1500 x g for 15 min and the resultant supernatant used for analysis. A Model 1050 UV HPLC (Hewlett Packard) with an integrated oven compartment (50°C) and data system was used. Sample injection (20 µL) was performed using an autosampler. The column was a prepacked Aminex HPX-87-H strong cation-exchange resin column (150 x 7.8 mm i.d.), fitted with an ion exclusion microguard refill cartridge (Bio-Rad). The eluent was 0.005 mol/L sulfuric acid.

    Statistical analyses. All data were expressed as means ± SD. For the in vitro gut model experiment, means before and after the treatment were analyzed by paired Student’s t test. For the adhesion assay and pig feeding experiment, the data were examined for equality by Levene’s test; if there was a difference (P < 0.05), the data were log-transformed. One-way ANOVA was performed to determine the effect of the test oligosaccharides on the adhesion of the pathogenic bacteria to HT29 cells. Significant differences from the control (PBS) were determined by Dunnett’s test. The effects of diet on 1) proximal colonic microbiota, 2) distal colonic microbiota, 3) fecal microbiota, 4) proximal colonic SCFA concentration, and 5) distal colonic SCFA concentration were determined by 1-way ANOVA. Significant differences between diet groups were determined by Tukey’s Honestly Significant Difference test for comparison of means within a factor. All statistical analyses were performed using the SPSS package program version 11.5.0 and differences were considered significant at P < 0.05.

	RESULTS

TOP ABSTRACT MATERIALS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
In vitro gut model system

The in vitro gut model is a system for screening the effect of dietary intervention on the colonic microbiota composition. GOS supplementation did not affect population numbers of total bacteria, lactobacilli, bacteroides, or clostridia (Table 1). Numbers of bifidobacteria increased by 0.9 log₁₀ cells in the first vessel (P < 0.01) (representing the proximal colon), and similarly in the second vessel, the transverse colon (0.7 log₁₀ cells; P < 0.01) (Table 1). Numbers of lactobacilli (0.2 log₁₀ cells) tended to be greater (P = 0.16) in the second vessel after addition of the novel GOS.

Addition of the GOS mixture, up to 10 mmol/L, to the growth medium had no toxic effect on HT29 cells after 24 and 48 h of incubation. Cell viability was reduced in the presence of 100 mmol/L of GOS after 24 h (P < 0.001) and 48 h (P < 0.001) of incubation

Adhesion assay

The GOS addition, as a mixture and as individual fractions, did not affect the adhesion of E. coli 0157:H7 (VT^–) (P = 0.194). A very strong inhibition (almost 90%) of attachment of EPEC (P < 0.001) and Salmonella enterica serotype Typhimurium (P < 0.001) to HT29 cells occurred in the presence of the novel GOS mixture. We attributed this inhibition to the disaccharide fraction of the mixture, which had identical anti-adhesive properties (P = 0.001 and P < 0.001, respectively). Apart from the disaccharide fraction, only the fraction containing tetra- and pentasaccharides inhibited attachment (60% against S. enterica serotype Typhimurium P < 0.05).

Clinical measurements

    Dietary intake and pig growth. All pigs consumed the entire volume of feed provided each day. The final body weight was 42.5 ± 3 kg and did not differ among experimental groups at any time of the study. One pig was euthanized on welfare grounds after 3 wk of the study.

    Bacterial population changes in colonic contents and feces. Major bacterial genera in the proximal and distal colonic contents (Table 2), and feces of pigs were measured (Table 3). Total numbers of bacteria for the same colonic regions did not differ among treatments (proximal colon P = 0.876; distal colon P = 0.994; feces P = 0.989), but did differ among sample type (P < 0.05).

Pigs fed 4% GOS had greater numbers of bifidobacteria than pigs fed NEG (P < 0.05) or 1.6% GOS (P < 0.05), but did not differ from pigs fed inulin (P = 0.533). The numbers of lactobacilli did not differ among the 1.6% GOS-, 4% GOS-, and inulin-fed groups (P = 0.752), all of which had more lactobacilli than the NEG group (P < 0.01) (Table 2).

Fecal clostridia population numbers did not differ among the groups (P = 0.072). Pigs fed inulin had higher numbers of fecal bacteroides than those fed NEG (P < 0.05), but did not differ from those fed 1.6% GOS (P = 0.207) or 4% GOS groups (P = 0.245). Fecal lactobacilli did not differ among the 1.6% GOS-, 4% GOS-, and inulin-fed groups, all of which had higher numbers than pigs fed the NEG diet (P < 0.05). Fecal bifidobacteria populations in pigs fed inulin were higher than in pigs fed NEG (P < 0.001) or 1.6% GOS (P < 0.05), but did not differ from those in pigs fed 4% GOS (P = 0.735) (Table 3).

Lactic acid and SCFA concentration in colonic contents

Lactic, acetic, propionic, and butyric acid concentrations in the proximal and distal colonic contents of pigs were determined (Table 4). In the proximal colonic contents, lactic acid concentration did not differ between pigs fed the NEG diet and those fed the inulin supplement (P = 0.814). Pigs fed 4% GOS had greater lactic acid concentration than pigs fed NEG (P < 0.05) and inulin (P = 0.058). Addition of 1.6% GOS resulted in the highest lactic acid concentration (6.62 mmol/L) among the treatment groups (P < 0.001). No lactic acid was detected in the distal colonic samples.

Propionic acid concentration did not differ among the treatment groups in the distal colonic contents (P = 0.311). In the proximal colonic contents, the propionic acid concentration was lower in pigs fed 1.6% GOS than in pigs fed NEG (P < 0.01), 4% GOS (P = 0.129), or inulin (P = 0.582). Concentrations of butyric acid concentration (10–11 mmol/L) did not change in response to the dietary treatments (P = 0.944) in the proximal colonic content. In the distal colonic contents, pigs fed inulin had a higher butyric acid concentration than pigs fed NEG (P = 0.068), 1.6% GOS (P < 0.05) or 4% GOS (P = 0.234).

pH of colonic contents

Pigs fed 4% GOS had lower proximal colonic pH than pigs fed NEG (P < 0.05), 1.6% GOS (P < 0.05), or inulin (P < 0.001). Pigs fed inulin had higher pH than pigs fed 1.6% GOS (P < 0.05). The distal colonic pH did not differ among the treatment groups (P = 0.995) (Table 5).

	DISCUSSION

TOP ABSTRACT MATERIALS AND METHODS RESULTS DISCUSSION LITERATURE CITED

Oligosaccharides may also have therapeutic potential to reduce Salmonella spp. and E. coli–associated enterotoxigenic infections. The hypothesis is that these oligosaccharides act as functional mimics for bacterial receptors on intestinal cells; therefore, they competitively bind the bacteria, thereby preventing attachment to the gut epithelium. Previous studies reported that glycolipid receptors containing the galabiose disaccharide can mediate the binding of P-fibriated E. coli to uroepithelial cells (19) and that oligosaccharides were able to block the adherence of P-fimbriated E. coli to cells in vitro (20,21). Here, we used the synthesized oligosaccharides (GOS), as a mixture and as oligosaccharide fractions to study the adhesion of an EPEC, a VTEC O157:H7 (VT^–), and S. enterica serotype Typhimurium to HT29 adenocarcinoma cells in vitro. Addition of the novel GOS mixture to the assay system reduced the attachment of EPEC (P < 0.01) and S. enterica serotype Typhimurium (P < 0.01) but not of the VTEC (P = 0.194). When various fractions of the mixture were tested, only the disaccharide fraction, which also contained an anomeric configuration, had a similar effect, although larger molecules of the tetra- and pentasaccharides showed a degree of inhibition of S. enterica serotype Typhimurium attachment.

The prebiotic capacity of the novel GOS was further investigated in a pig feeding trial. Pigs are considered to be a good experimental model for humans due to physiologic and metabolic similarities (22). For a food ingredient to be considered to be a prebiotic, in addition to promoting a healthier colonic microbiota, it must not be hydrolyzed or absorbed in the upper part of the gastrointestinal tract (6). The reduced pH (P < 0.05) in the proximal colon combined with significant increases in SCFA production (P < 0.01) strongly suggest that the GOS mixture was not absorbed in the small intestine, but was delivered to the colon intact. The increase in SCFA in the proximal colon was due almost entirely to acetic acid production (P < 0.01), the main fermentation product of bifidobacteria. This was in turn confirmed by an increase in bifidobacterial numbers, consistent with prebiotic activity. Generally, the performance of the novel mixture (high concentration) in manipulating the bacterial composition was comparable to that of inulin in the distal colon and fecal samples, but offered a significant improvement in the proximal colon.

Nondigestible oligosaccharides were suggested to benefit the health of the host by selectively stimulating the growth and activity of potentially health promoting bacteria. It has been proposed that the "second generation" prebiotics should be designed to have improved functionality by providing a more selective fermentation and/or added functionality. In this context, we showed in vitro and in vivo that this novel GOS mixture not only offers a prebiotic effect, but also selectivity toward bifidobacteria. Moreover, the GOS mixture (in vitro) has the potential to inhibit the attachment of 2 common gastrointestinal pathogens, enteropathogenic E. coli and S. typhimurium, to the colonic epithelium by acting as a decoy for their receptor sites. However, further in vivo studies are warranted to evaluate that potential.

	FOOTNOTES

 
¹ Funded by a grant from Clasado, Incorporated.

³ Abbreviations used: EPEC, enteropathogenic E. coli; FISH, fluorescent in situ hybridization; Gal, galactose; Glc, glucose; GOS, galactooligosaccharide; VTEC, verocytotoxic E. coli.

⁴ A commercial diet containing (g/kg): oil 33, protein 192, lysine 13.2, fiber 28, ash 48, moisture 138, vitamin premix 1.2 (vitamin A, 9500 IU; vitamin E, 100 IU; cholecalciferol, 1850 IU; sodium selenite, 0.0003; cupric sulfate, 0.17; antioxidant premix, 6 (BHA, 1; BHT, 1; ethoxyquin, 4).

Manuscript received 18 February 2005. Initial review completed 31 March 2005. Revision accepted 27 April 2005.

	LITERATURE CITED

TOP ABSTRACT MATERIALS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 

1. Fuller R., Gibson G. R. Modification of the intestinal microflora using probiotics and prebiotics. Scand. J. Gastroenterol. 1997;32:28-31.[Medline]

2. Bartram H. P., Scheppach W., Gerlach S., Ruckdeschel G. Does yoghurt enriched with Bifidobacterium longum affect colonic microbiology and fecal metabolites in healthy subjects?. Am. J. Clin. Nutr. 1994;59:428-432.[Abstract/Free Full Text]

3. Ling W. H., Korpela R., Mykkanen H., Salminen S. Lactobacillus strain GG supplementation decreases colonic hydrolytic and reductive enzyme activities in healthy female adults. J. Nutr. 1994;124:18-23.

4. Roberfroid M. D., Van Loo J.A.E., Gibson G. R. The bifidogenic nature of chicory inulin and its hydrolysis products. J. Nutr. 1998;128:11-19.[Abstract/Free Full Text]

5. Collins M. D., Gibson G. R. Probiotics, prebiotics and synbiotics: approaches for the nutritional modulation of microbial ecology. Am. J. Clin. Nutr. 1999;69:1052S-1057S.[Abstract/Free Full Text]

6. Gibson G. R., Roberfroid M. B. Dietary modulation of the human colonic microflora—introducing the concept of prebiotics. J. Nutr. 1995;125:1402-1412.

7. Gibson G. R., Ottaway P. B., Rastall R. A. Dietary modulation of the human colonic microflora—introducing the concept of prebiotics. Prebiotics: New Developments in Functional Foods. 2000 Chandos Publishing Oxford, UK.

8. Boehm G., Jelinek J., Stahl B., Laere van K., Knol J., Fanaro S., Moro G., Vigi V. prebiotics in infant formulas. J. Clin. Gastroenterol. 2004;38:S76-S79.[Medline]

9. Sako T., Matsumoto K., Tanaka R. Recent progress on research and application of non-digestible galacto-oligosaccharides. Int. Dairy J. 1999;9:69-80.

10. Schnaar R. L. Glycosphingolipids in cell surface recognition. Glycobiology. 1991;1:477-485.[Free Full Text]

11. Tzortzis G., Goulas A. K., Gibson G. R. Synthesis of prebiotic galactooligosaccharides using whole cells of a novel strain, Bifidobacterium bifidum NCIMB 4117. Appl. Microbiol. Biotechnol. 2005; (in press).

12. Macfarlane G. T., Macfarlane S., Gibson G. R. Validation of a three-stage compound continuous culture system for investigating the effect of retention time on the ecology and metabolism of bacteria in the human colon. Microb. Ecol. 1998;35:180-187.[Medline]

13. Vulevic J., Rastall R. A., Gibson G. R. Developing a quantitative approach for determining the in vitro prebiotic potential of dietary oligosaccharides. FEMS Microbiol. Lett. 2004;236:153-159.[Medline]

14. Langendijk P. S., Schut F., Jansen G. J., Raangs G. W., Kamphuis G. R., Wilkinson M.H.F., Welling G. W. Quantitative fluorescent in situ hybridization of Bifidobacterium spp. with genus specific 16S rRNA targeted probes and its application in fecal samples. Appl. Environ. Microbiol. 1995;61:3069-3075.[Abstract]

15. Manz W., Amann R., Ludwig W., Vancanneyt M., Schleifer K. H. Application of a suite of 16s rRNA-specific oligonucleotide probes designed to investigate bacteria of the phylum cytophage-flavobacter-bacteroides in the natural environment. Microbiology. 1996;142:1097-1106.[Abstract/Free Full Text]

16. Franks A. H., Harmsen H.J.M., Raangs G. C., Jansen G. J., Schut F., Welling G. W. Variation of bacterial populations in human feces measured by fluorescent in situ hybridization with group specific 16S rRNA-targeted oligonucleotide probes. Appl. Environ. Microbiol. 1998;64:3336-3345.[Abstract/Free Full Text]

17. Harmsen H.J.M., Elfferich P., Schut F., Welling G. W. A 16S rRNA-targeted probe for detection of lactobacilli and enterococci in fecal samples by fluorescent in situ hybridization. Microb. Ecol. Health Dis. 1999;11:3-12.

18. Olano-Martin E., Williams M. R., Gibson G. R., Rastall R. A. Pectins and pectic-oligosaccharides inhibit Escherichia coli O157: H7 Shiga toxin as directed towards the human colonic cell line HT29. FEMS Microbiol. Lett. 2003;218:101-105.[Medline]

19. Karlsson K. A. Meaning and therapeutic potential of microbial recognition of host glycoconjugates. Mol. Microbiol. 1989;29:1-11.

20. Svanborg-Eden C., Andersson B., Aniansson G., Lindstedt R., Man de P., Nielsen A., Leffler H., Wold A. Inhibition of bacterial attachment: examples from the urinary and respiratory tracts. Curr. Top. Microbiol. Immunol. 1990;121:167-184.

21. Guggenbichler J. P., Bettignies-Dutz de A., Meissner P., Schellmoser S., Jurenitsch J. Acidic oligosaccharides from natural sources block adherence of Escherichia coli on uroepithelial cells. Pharm. Pharmacol. Lett. 1997;1:35-38.

22. Anzenbacherova E., Anzenbacher P., Svoboda Z., Ulricova J., Kvetina J., Zoulova J., Perlik F., Martinkova J. Minipig as a model for drug metabolism in man: comparison of in vitro and in vivo metabolism of propafenone. Biomed. Papers. 2003;147:155-159.

This article has been cited by other articles:

				R. Marti, P. Dabert, and A.-M. Pourcher Pig Manure Contamination Marker Selection Based on the Influence of Biological Treatment on the Dominant Fecal Microbial Groups Appl. Envir. Microbiol., August 1, 2009; 75(15): 4967 - 4974. [Abstract] [Full Text] [PDF]

				L. E. J. Searle, A. Best, A. Nunez, F. J. Salguero, L. Johnson, U. Weyer, A. H. Dugdale, W. A. Cooley, B. Carter, G. Jones, et al. A mixture containing galactooligosaccharide, produced by the enzymic activity of Bifidobacterium bifidum, reduces Salmonella enterica serovar Typhimurium infection in mice J. Med. Microbiol., January 1, 2009; 58(1): 37 - 48. [Abstract] [Full Text] [PDF]

				J. Vulevic, A. Drakoularakou, P. Yaqoob, G. Tzortzis, and G. R Gibson Modulation of the fecal microflora profile and immune function by a novel trans-galactooligosaccharide mixture (B-GOS) in healthy elderly volunteers Am. J. Clinical Nutrition, November 1, 2008; 88(5): 1438 - 1446. [Abstract] [Full Text] [PDF]

				F. Depeint, G. Tzortzis, J. Vulevic, K. I'Anson, and G. R Gibson Prebiotic evaluation of a novel galactooligosaccharide mixture produced by the enzymatic activity of Bifidobacterium bifidum NCIMB 41171, in healthy humans: a randomized, double-blind, crossover, placebo-controlled intervention study Am. J. Clinical Nutrition, March 1, 2008; 87(3): 785 - 791. [Abstract] [Full Text] [PDF]

				A. Amaretti, T. Bernardi, E. Tamburini, S. Zanoni, M. Lomma, D. Matteuzzi, and M. Rossi Kinetics and Metabolism of Bifidobacterium adolescentis MB 239 Growing on Glucose, Galactose, Lactose, and Galactooligosaccharides Appl. Envir. Microbiol., June 1, 2007; 73(11): 3637 - 3644. [Abstract] [Full Text] [PDF]

				G. Loh, M. Eberhard, R. M. Brunner, U. Hennig, S. Kuhla, B. Kleessen, and C. C. Metges Inulin Alters the Intestinal Microbiota and Short-Chain Fatty Acid Concentrations in Growing Pigs Regardless of Their Basal Diet J. Nutr., May 1, 2006; 136(5): 1198 - 1202. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tzortzis, G. Articles by Gibson, G. R. Search for Related Content PubMed PubMed Citation Articles by Tzortzis, G. Articles by Gibson, G. R.