Probiotics Facts and Fancy From pro- and prebiotics to synbiotics

Transcription

1 Probiotics Facts and Fancy From pro- and prebiotics to synbiotics Torkel Wadström Dept Laboratory Medicine University of Lund

2 Probiotic food was consumed by prehistoric man To digest roots and other dietary fibers To ferment fruits ( prehistoric wine and beer ) To protect baby foods To combat food decay, e.g. mycotoxins (also in the gut)

3 Microbes used as probiotics Lactobacillus spp Enterococcus spp Lactococcus, Leukonostoc, Pediococcus spp Bifidobacterium spp Bacillus subtilis, B vietnam and others Saccharomyces boulardii Clostridium butyricum Staphylococcus spp

4 Developing a synbiotic concept for antibiotic associated diarrhoea. Single Probiotic strain Open (LGG/Saccharomyces boulardii/l. casei, Shirota) Single Probiotic strain + Single prebiotic (LGG + Oligo fructose) Multiple Probiotic strains (VSL#3 for DSS induced colitis in Mice) L. Salivarius Ls33, L. plantarum Lp115, L. acidophilus NCFM for TNBS induced colitis in Mice) Multiple Probiotic strains + Multiple prebiotics Qualvivo product For AAD 4

5 Probiotics Open Live microorganisms - Confer a health benefit to the host Lactobacilli, bifidobacteria, Bacillus subtilis, Clostridium butyricum, Saccharomyces Non-digestible food ingredient Prebiotics Resistance to degradation by stomach acid, human digestive enzymes Fermentable by intestinal microbes & Beneficially affects the host by selectively stimulating the growth and/or activity of one or a limited number of bacteria in the colon, thus improving host health Fructooligosaccharides, GOS, XOS etc 5 Journal of Nutrition, 1995, 125:

6

7

8 The barrier effect against GI pathogens

9 What are we doing?? CDAD Probiotics Prebiotics

10 Microbes used as probiotics Lactobacillus spp Enterococcus spp Lactococcus, Leukonostoc, Pediococcus spp Bifidobacterium spp Bacillus subtilis, B vietnam and others Saccharomyces boulardii Clostridium butyricum Staphylococcus spp

11 Some beneficial selection criteria Survival during transport in the GI tract, i.e. acid and bile tolerance Adhesion to intestinal mucus (mucin, ECM) Production of stress proteins Production of β-galactosidase (c.f. lactose intolerance)

12 Selection criteria (contin) Binding of mutagens Production of bacteriocins Specific activity against intestinal Specific activity against intestinal pathogens rotavirus, Salmonella, Campylobacter jejuni, travellers diarrhoea, diarrhoea of piglets and newborn calves

13

14 Probiotics and innate immunity Tighten the mucosal barrier (tight junctions) Inhibition of translocation Stimulate epithelial defensins Immunomodulate cell surface, cell and extracellular molecules Lipoteichoic acid (LTA) and LTA fatty acids, cell surface proteins (CSP:s)

15 Pseudomembranous colitis Open 15 American Family Physician, 2005, 71: 921

16 C difficile Colitis Several case reports and open studies on LGG and on S boulardii in children and adults, no effect or slight beneficial effect L plantarum 299v: recurrence of colitis prevented in 7/11 pts compared to 3/9 controls

17 Antibiotic Associated Diarrhea & Clostridium difficile Associated Diarrhea

18 In vitro Antimicrobial activity of Probiotic strains against C. difficile LAB & Bifidobacteria cultures Liquid assay using microtiter plate 18 18

19 The VSL#3 pioneer product (10 9 CFU/g) inhibits IBD and gut microbe translocation in severe trauma (K Madsen, AGA 2005) Multistrain probiotic, Sigma-Tau

20 Chronic pouchitis in IBD VSL#3 prevented relapses better than placebo, p<0.001 Prevented onset of acute pouchitis, p<0.05 Maintained remission for 1 year p<0.001 Gionchetti et al., 2000, 2003, Mimura et al 2004

21 Inhibition of gut inflammation (IBD and liver) in mouse models: The IL10 -/- mouse SCID Rag IBD and colon cancer (JG Fox, MIT)

22 IL-10 and Inflammatory Bowel Disease IL-10 downregulates the inflammatory response Mice with colitis successfully treated with Lactococcus lactis secreting IL-10 Supply of LAB to IL-10 -/- mice prevented colitis IL-10 -/- mice given L salivarius faecal coliforms, enterococci, C perfringens, and reduction of colon cancer and mucosal inflammation

23 From probiotics to immunobiotics To balance the TNF-α and NFκB pathways we need a LAB `turbo` mix of strains ( LTA, other immunomodulators, ( LTA, other immunomodulators, hydrophobins?)

24 Optimized prophylaxis - therapy to combat antibiotic use Dose and delivery (yoghurts, pills, pro- and prebiotic mixes) Stability (VSL#3, Sigma Tau only one week at 20ºC!!!) C.f. surgery, travellers diarrhoea, antibiotic-induced colitis

25 Probiotics Should strains be of human origin? Live or dead? Should strains colonize the gut >7 day? Dose? Stability in food or tablets

26

27 PCR-DGGE (Denaturing Gradient Gel Electrophoresis) To follow LAB in gut biopsies and faeces Well standardized, cheap (c.f. FISH method) In mouse models Dogs, other pets, volunteer studies

28 Highlight: The gut mucin layer Rich in germ-free (GF) animals Modulated by microbe/lab endo- and exoglycosidases Bile-modifying enzymes and the hydrophobic lining stomach to colon Habitat for many microbes (LAB, microaerophiles c.f. Helicobacter spp etc)

29 Highlight (contin.) Salivary glycoproteins and mucus (trefoil peptides) coaggregate Most lactobacilli adheres to mucus microbes e.g. bypass the stomach (low ph and bile stress) Adhesion of Lactobacillus reuteri to mucus coated Sepharose beads Stefan Roos, Swedish University of Agricultural Sciences

30

31

32 Double-blind, randomized, placebocontrolled studies needed!!!!!! However, EXPENSIVE!!

33 Therapeutic Microbiology J Versalovic, ASM Press 2008