probiotics and mucosal immunity
DESCRIPTION
Living microorganisms, which upon ingestion in certain numbers, exert health effects beyond inherent basic nutrition.TRANSCRIPT
WELCOME
PROBIOTICS AND MUCOSAL IMMUNITY
By:Someshwar Zadbuke and Mayank
Tandon
IVRI, Izatnagar, U.P., India and NDRI, Karnal, Haryana, India
Probiotics
“Living microorganisms, which upon ingestion in certain numbers, exert health effects beyond inherent basic nutrition” (Guarner, 1998)
First reported by Elie Matchnikoff (1907)
Term coined in 1965 (Lilly and Stillwell, 1965)
Lactic acid bacteria (Lactobacilli, Bifidobacteria and Enterococci)
Yeasts Safe as opposed to antibiotics (Reid 2000)
Desirable properties
Survivability in acidic pH of stomach (Dunne et al. 2001)
Resistance to digestion by bile Ability to adhere to intestinal epithelium Antimicrobial substances (Kailaspathy et al. 2000)
Inactivation of procarcinogen (Perdigon et al. 2001)
Self GRAS (Weese and Anderson, 2002)
Mode of action
Antimicrobial substance Competitive exclusion Anticholesterolemic and antilipidemic factors
(Kim and Lee 2005)
Modulation of the immune system Antitumor activity Decrease toxic amines and ammonia Sparing effect Increase GI tone and motility (Dunne et al. 2001)
Effect of probiotics on animal performance Mixed response reported Type of strain Single or multiple? Duration of feeding
Performance of calves fed probiotics
Probiotics Control SE
Initial BW kg
40.0 37.2 1.13
56d BW kg 79.9 73.4 1.85
BW gain kg 40.5 36.2 1.39
Feed conversion
1.59 1.64 .028
Diarrhea, cases
1* 7
Fumiak et al, 1995
The effect of administration of probiotics (Bifidobacrerium pseudolongurn or Lactobacillus acidophilus) on the survival rate of newborn piglets
Performance of WL layers fed 15.3%, 14.3% and 13.8% CP diets containing Lactobacillus CCMS premix
Treatment HEP (%)
FC DFC EM
CCMS(g/kg) Lacto(Mg/kg) Cp (g/kg)
0 0 153 88.6 2.08a 109 52.4a
20 0 153 89.5 2.10a 112 53.1a
20 1100 153 89.1 2.08a 112 53.9b
20 1100 143 88.3 2.11a 112 53.0a
20 1100 138 88.1 2.17b 114 52.5a
(Nahashon et al., 1996a)
Diet DFC g BWG g Feed/Gain
Control 57b 261b 6.11b
C+CCMS 57b 234c 6.82a
C+CCMS Lacto 59a 272a 6.07b
Performance of WL pullets fed corn-soya meal diet with CCMS and Lacto-CCMS premix from 7 to 19 weeks of age (Nahashon et al., 1996b)
Performance of goat kids on supplementation of Probiotics
Parameters Treatment goups
A B C D
DMI kgw0.75/d (g) 68.5 68.5 71.1 71.1
CP Dig % 76.8 75.5 80.1 82.6
CF Dig % 44.1 45.7 55.2 64.3
Wt gain(g/D) 64.3 87.2 94.9 89.6
Feed eff. G/Kg 7.7 5.6 5.6 5.4
Diarrhea cases observed
10 3 6 4
A- Control, B - L. acidophillus, C - S. cerevisiae, D - L. acidophillus + S. cerevisiae
(Mudgal et al. 1995)
The general health score (GHS) of calves per experiment as affected by treatment with a multispecies probiotic (MSPB, white bars) or a calf-specific probiotic (CSPB, gray bars) vs. control treatment (black bars) (Timmerman et al., 2005)
Introduction to host defense and immunity
Gut Mucosal Barrier – Host defense
Normal digestive functions (Sanderson et al. 1993)
Protection- host defense External secretions Intestinal, genital and bronchial fluids Intestinal flora Epithelial cell membranes Peristalsis, Proteolysis Birth
Adoptive changes Maturational changes
(Isolauri et al. 2001)
(Perdigon et al. 2001)
Development of immunity
“Recognition of a foreign material or pathogen and mounting of a reaction to eliminate it”
Innate immunity Adoptive immunity Cells mediating IR:
Lymphocytes Macrophages Epithelial cells Ag presenting cells
(Erikson and Hubbard 2000)
Mucosal immune system (MIS)
GIT, Respiratory tract, Lacrimal, Salivary and Mammary gland (Phillips and Lamm, 1998)
Primary Lymphoid Organs Thymus Bone marrow
Secondary lymphoid organs Spleen Lymph nodes (Tizard 1998)
MALT (Mucosa Associated Lymphoid Tissues) IgA Immune cells (Cytokines and Chemokines)
(Portal, 2003)
Peyer’s patches
Dome region microfold cells (M cells) (Mc Ghee and Kiyono, 1992)
Germinal center- B cells change IgM to IgA Ag presenting cells (APC) – dendritic cells
and macrophages (Perdigon et al. 2001)
IgA
S-IgA Dimeric or polymeric bound by join chain “J” secretory component (Brandtzaeg, 1995)
IgA1 – Small Intestine IgA2 – Colon Part of common MIS (Cebra et al. 1991)
Level depends on presence of microflora (gram -ve) Excretory function (Lamm et al. 1996)
Synthesis of IgA
Ag - immune cells - cytokines Interaction
M cells Epithelial cell Local immune response (Weiner, 1997)
Ag delivery systems
Compounds having adjuvant properties Incorporation of Ag in particles (Michalek et al. 1994)
Effect of Antigen on MIS
Control of Ag absorption in Gut
Immune exclusion Immune elimination Peyer’s patches – covered by M cells
Ag T cells Effector cells
Active immune suppression
Differentiation of IgA secreting B cells
Tolerogenic form
(Sanderson and Walker 1993)
(Strober et al. 1998)
Mechanism of IgA in immune exclusion of Ag (Brandtzaeg P. 2002)
Oral Tolerance
“Immunogenic hyporesponsiveness to Ag’s encountered through the enteric route”
T and B cell mediated (CD8+ T cells) LPS – maturation of T cell precursors (Perdigon et al.
2001) Active suppression and Clonal deletion (Toy and Mayer
1996) Intestinal permeability (Isolauri et al. 1993) Soluble Ag’s Particulate Ag (Metzler et al. 2005) Dose dependent
(Strobel and Mowet, 1998)
Fate of Ag
Ag --- Intestinal wall (Lamina propria and peyer’s patches)
M cells {absorb- apical memb of PP (glucoconjugates) –
coating of Ag (adhesin) and Transport to immune cells}
Lymphoid cells
Immune
response Pathology
(Tizard 1998 and Erikson and Hubbard 2000)
Probiotic modulation of the immune system
altering immunogenecity of Ag
reducing inflammatory mediators
reversing intestinal permeability and
enhancing degradation of Ag
diverting gut Ag uptake toward PP
enhancing mucosal IgA response
normalizing intestinal flora
Non specific immune response
Inflammatory response – PMN and Macrophage
Ag presenting cells (APC) Lymphocytes
Phagocytic activity in blood of Japanese quail after application of L. fermentum AD1
Parameter Control group Experimental group
Leucocyte count (G/l) 16.68 (5.43) 14.40 (2.54)
Heterophils (%) 16.20 (12.50) 26.40 (7.40)
Lymphocytes (%) 83.80 (12.48) 73.40 (7.81)
Monocytes (%) 0.0 (0.0) 0.2 (0.4)
Basophils (%) 0.0 (0.0) 0.0 (0.0)
Eosinophils (%) 0.0 (0.0) 0.0 (0.0)
Phagocytic activity (%)
8.0 (1.6) 16.8 (6.3)*
Index of phagocytic activity
0.56 (0.19) 2.25 (0.95)**
*P < 0.05, **P < 0.01
(Strompfova et al. 2005)
Effects of probiotics on nonspecific immunity
Citation Probiotic Species Assessment Effect
Matsuzaki et al. 2000
L. casei Shirota
Mice Innate immune response
Increased NK cell activity from mesenteric node not of PP or spleen
Gill et al. 2000
L. rhamnosus (HN001, DR20), L. acidophilus (HN017), or B. lactis (HN019, DR10)
Mice responses to concanavalin A and lipopolysaccharide
Increased phagocytic activity with elevated γ-INF
Matsuzaki et al. 1995
L. Acidophylus or B. bifidum oral (Live)
Rat Peritoneal or peripheral blood macrophages
Enhanced phagocytosis
Dalloul et al. 2003
Different srains of Lacto-bacillus
Chicken Challenged with Coccidiosis
Elevated CD3, CD4, CD8 and αβTCR Reduced oocysts shedding Control group- more Abs against recombinant coccidial Ag in intestinal secretions
Specific immune response
Ag - M cells
Peyer’s patches persists in blood
T helper and inducer phenotype
Max conc
B cell response IgA and CD8+
Intraepithelial lymphocytes, T cells with γ,δ receptorsinteract with epithelial cells – attract other immune cellsunexplored area (Kaila 1992, Isolauri et al. 2001)
Response to Bacteria
BacteriaGram + ve
Peptidoglycan (PG) Muramyl dipeptide (MDP)
Gram - vePeptidoglycan (PG)Lypopolysaccharide (LPS)
Development of immune system by cell stimulation
Receptor dependent process involving cell surface CD 14
Toll receptors associated with pathogens
Transduction pathways
Expression of several immune response genes
Macrophages, Endothelial cells, Smooth muscle cells, Neutrophylls
Mediators – Cytokines
CYTOKINE NETWORK
(Yasui and Ohwaki, 1991, Erikson and Hubbard 2000)
Modualtion of specific immunityCitation Probiotic Animal Assessment Effect
Matsuzaki et al. 1995
L. Casei (Live)
Mice Infection and Ab production in malnourished animals
Increased sIgA and reduced enteric infection
Benyacoub et al. 2005
E. faecium Dog- puppies
Assessment of immune response in post weaning stress
Elevated fecal IgA, circulating IgA, IgG Increased mature B cells No difference in CD4+ and CD8+
Haghighi et al. 2005
L. acidophillus, B. bifidium
Chicken Challenged with SRBC BSA TT
Increased IgMNo responseIncreased IgG, IgA
Probiotics (B. subtilis Bs964, Candida utilis BKM-Y74, L. acidophilus LH1F) supplemented orally to chicks from post-hatch to day 3 of age
Enhanced response to Intestinal fluid - IgAat day 7 PP – IgG, IgM forming cells at day 10 and 7 Cecal tonsils –
IgA forming cells at day 7 to 10 IgG forming cells at day 7 IgM forming cells at day 4 to 7 T lymphocytes at day 7 Increased density of microvilli and length of cecal tonsils
Decreased efficiency with ageing of the chicken
Effect of probiotics on intestinal mucosal immunity and ultrastructure of the cecal tonsils of chicken
(Yurong et al. 2005)
Total IgA levels in feces of control and probiotic-fed mice on d 0, 7, 14, and 28 of the trial.P < 0.05
Benyacoub et al, 2005a
Specific anti-canine distemper virus immunoglobulin IgG (upper panel) and IgA (lower panel) in the plasma collected at wk 0, 10, 18, 31 and 44 from puppies fed diets with or without SF68
Benyacoub et al, 2005b
Effect of Bifidobacterium brave, LPS and Con A on the proliferation of PP cells in mice
Mitogen PP cells Unfractionated cells
B cell enriched fraction
T cell enriched fraction
Control 4,529 401 69 1,299
B. Breve 9,119** 2,376** 815*** 1,914
LPS 19,588*** 9,228*** 3,643*** 2,272*
Con A 4,815 83329*** 937*** 156,423***
*P<0.02, **P<0.01, ***P<0.001
(Yasui and Ohwaki, 1991)
Recirculation of MIS (Common mucosal system) ~2% lymphocytes pool recirculating each hr MALT and the systemic lymphoid system Lymphocte cells adhesion molecules - spc for mucosal post
capillary venulePeyer’s patches
Lymphocytes
Regional lymphnode
Mescentric group
Thoracic duct
Blood vascular system
Intestinal lamina propria
Other secreting tissues
(Brandtzaeg et al. 1999)
(Cebra et al. 1991)
Probiotic effects in rodent models of some diseases
Disease model Probiotic Assessment Effect
Insulin dependent diabetes mellitus
L. casei, oral (live)
T-cell markers, splenic cytokines
Decreased CD4+ cells, IFN-γ and IL-2
Imnsulin dependent diabetes mellitus
L. casei, oral (Heat killed)
Splenic T and B cell number and production of cytokines
Decreased incidence of diabetes, increased CD45+ b-cells and IL-2 decreased CD8+ T- cells and IFN- γ
Influenza immunization
B. Bifidus oral
Resp. tract infection and antiinfluenza virus IgG
Protection against lower respiratory tract infections Higher serum IgG levels
(Errikson and Hubbard 2000)
Mean antibody titer against Newcastle disease vaccine on day 42 in chicken
Strain Control OTC L. Casei
HH 2.42b 3.01ab 3.37a
SS 3.19 3.01 3.22
(Zulkifli et al. 2000)
Challeges faced by MIS
Microbial infections Route of entry of Ag Dietary indiscretions Allergies and food intolerances Oral antibiotics Disruption of lipid and FA metabolism Ageing Inflammation – level of IgA
Intraluminal Ag
Beneficial/ own
microflora- probiotics
Enhance immune response
Non-beneficial Ag
Adhered to epithelial cells
Entry via enterocyte
Escape tolerance induction of peyer’s patches
promote OT
Balance of metabolic activity and gut microflora
Balance of gut microflora
Choosing probiotics
Type of immune cells stimulated (Inflammatory or specific immune response)
Most active strain Dose required for maximum effect? When to be administered? Is it safe to use probiotics in immunosuppressed
host? Storage quality
Future prospects
Species identification and their use needed to be quantified
Identification of direct cause and effect needed
Reduced inflammation either at local or systemic level?
Can they be used as adjuvants for oral immunisation?
Modulation of MALT or a systemic immune response?
Conclusions
No clear cut evidence observed on the effect of probiotics on production aspect of animals
Competitive exclusion is beneficial in early phase of life to prevent diarrheal occurrences
Probiotics modulate immunity of host through enhanced mucosal immunity (non specific as well as specific)
Can be used as prophylactic measures to enhance health status of the animals