handouts seminar

1. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 1
Het effect van het microbioom op gedrag –
van wetenschap naar praktijk (en weer terug)
Koen Venema
Beneficial Microbes Consultancy, Wageningen, The
Netherlands, koen.venema@outlook.com

2. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 2
• initiator and organisor of the
Beneficial Microbes Conference series
• editor-in-chief of the journal
Beneficial Microbes
• founder and CEO of
Beneficial Microbes Consultancy
2008
2010
2012
10-12 Oct. 2016
2015

3. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 3
Waar gaan we het vandaag over hebben?

4. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 4
Tijdens deze gast lezing zal ik een uitleggen wat microbiota en
probiotica zijn en doen en tevens uitleggen waarom het nodig is om in
multidisciplinaire teams de rol van de microbiota en probiotica op
moderne ziektes en aandoeningen, zoals overgewicht, allergiën,
inflammatoire ziektes, autisme, gedrag, depressie, etc., te onderzoeken.
Verwacht een combi van een klein beetje hard-core science met enkele
interessante feiten en af en toe wat humor.
“If you don't like bacteria, you're on the wrong planet.”
― Stewart Brand
“Wanneer je al het meercellige leven van de aarde zou
wegvagen, zou het microbiële leven waarschijnlijk een
klein beetje verschuiven. Verdwijnt het microbiële
leven, dan betekent dat de onmiddelijke dood voor
onze planeet.”
― Carl Woese

5. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 5
Als kinderen geboren worden, komen ze uit
een relatief steriele omgeving van de
baarmoeder in een wereld vol met bacteriën…

6. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 6
Script van deel 1 van de presentatie
 Microbiota
 aandacht in de wetenschap
 rol van microbiota bij ziekte en gezondheid
 hoe ziet onze microbiota er nu uit?
 ontwikkeling microbiota van jong tot oud
 probiotica
 prebiotica
 feces transplantatie (repoopulate)
 niet alle probiotica zijn hetzelfde
 Take home messages deel 1

7. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 7
Microbiota in de (wetenschappelijke) pers

8. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 8

9. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 9

10. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 10
.... want de microbiota is betrokken bij:
- colon kanker
- inflammatory bowel disease (IBD)
- allergie (huid, luchtwegen)
- overgewicht
- type I en II diabetes
- levensduur
- prikkelbaar darm syndroom (IBS)
- angst en depressie
- brein ontwikkeling en cognitie
- autisme
- Parkinson’s
- Alzheimer’s
- .....
Probiotics and Prebiotics
Current Research and Future Trends
2015. Eds. K. Venema & A.P. do Carmo

11. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 11
Het aantal bacteriën in de darm is 10x zo veel als onze
eigen lichaamscellen
Volwassenen kunnen ~ 1 kg darmbacteriën met zich
meedragen
We deponeren per jaar ons eigen gewicht aan bacteriën
in het toilet
Als we 70 zijn komt dat overeen met 12 olifanten
> 50% van feces is bacteriën (natgewicht)
Elk persoon heeft meer darm bacteriën dan het aantal
mensen dat er ooit op deze planeet geleefd heeft
400 - >1000 species
“Soundbites”
Courtesy of Roderick Mackie / Glenn Gibson

12. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 12
De darm microbiota is noodzakelijk voor:
- ontwikkeling van de darm
- ontwikkelen en trainen van het immuun systeem (70% van het immuun systeem
zit in de darmen)
- het produceren van vitamines (o.a. vit. K)
- het produceren van zogenaamde korte-keten vetzuren (SCFA; azijnzuur,
propionzuur en boterzuur); waarvan boterzuur als meest belangrijkste geacht
wordt, omdat microbieel boterzuur verantwoordelijk is voor ~70% van de totale
energiebron van het epitheel van de dikke darm
- kolonisatie resistentie
- door competitie voor nutrienten
- door competitie voor aanhechtingsplaatsen
- door productie van SCFA, waardoor lage pH
- door productie bacteriocines

13. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 13

14. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 14
“Exposed to a world of micro-organisms”
• Totale oppervlak van de darm is de grootte van een tennis court
• Dus dit moet goed verdedigd worden tegen indringers

15. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 15
Dit is waar uw lunch
nu is, met uw levende
bacteriele gasten

16. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 16

17. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 17
Bacteriën strijden om de aanhechting aan darm-epitheel

18. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 18
Als kinderen geboren worden, komen ze uit
een relatief steriele omgeving van de
baarmoeder in een wereld vol met bacteriën…

19. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 19

20. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 20
Moedermelk als een bron van gezonde bacteriën
Probiotica

21. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 21

22. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 22
microbiota
verstoring
microbiota
verstoring
ontsteking

23. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 23
Ziekte van Crohn
gereduceerde microbiele
genen diversiteit

24. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 24
Microbiota en overgewicht/obesitas

25. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 25
microbiota
verstoring
microbiota
verstoring
overgewicht

26. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 26
Kunnen we (een aantal van) die ziektes managen?

27. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 27
Probiotics: definition
live microorganisms which, when administered in
adequate amounts, confer a health benefit on the host
FAO/WHO 2001

28. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 28

29. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 29
<105
106
107
108
109
CFU/ml
Niet alle probiotica zijn gelijk! Slide van Ger Rijkers
#55
#22
#36
0 1 2 3 4 5 6
Time (hrs)
Lactobacillus acidophilus

30. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 30
Hoe werken probiotica?
Hoe kunnen op het brein inwerken als ze in de darm zitten?
barrier/
mucosal
integrity
lumen/
resilience
immune system
dendritic cell
epithelial
cell
microbiota
probiotics

31. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 31

32. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 32
Hoe kies je dan het juiste probiotica?
Courtesy of Ger Rijkers

33. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 33
Hoe blijf je op de hoogte?
Welke publicaties zijn van goede kwaliteit?
Wat voor type bewijs is voldoende?

34. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 34
Graad A
Graad B
Graad C
Graad D
Aanbeveling op basis van bewijs
Systematische review of meta-analyse
(minstens) 1 RCT
(minstens) 1 goed opgezette en
gecontroleerde studie
(minstens) 1 cohort studie
Goed opgezette beschrijvende
studies, case control, case series
Verslagen van expert comitees,
klinishe ervaring van experts

35. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 35
Zijn DBPCRT altijd nodig?
Double-Blind Placebo-Controlled Randomized Trials
Courtesy of Eric Claassen

36. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 36
Zijn DBPCRT altijd nodig?
Double-Blind Placebo-Controlled Randomized Trials
DBPCRT leveren graad A bewijs en zijn het meest overtuigend....
.... als ze op de juiste manier uitgevoerd worden!

37. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 37

38. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 38
Prebiotics: definition
a selectively fermented ingredient *
that results in specific changes in
the composition and/or activity of
the gastrointestinal microbiota, thus
conferring benefit(s) upon host
health
* largely restricted to carbohydrates until now
Blatchford, Venema and others 2014, Intl J Probiotics Prebiotics 8(4)

39. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 39

40. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 40
Effect van probiotica op maagdarm ziektes

41. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 41
Effect van soort probiotica n

42. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 42

43. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 43
■ genome stability
Sybesma et al. AEM 2013

44. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 44
Antibiotica gebruik & C. difficile
C. difficile
bacterie

45. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 45
• C. difficile vaak ongevoelig voor antibiotica
• dat is juist de reden dat na een antibioticum kuur C.diff
uitgroeit
• en is tegelijk de reden dat een volgende antibioticum kuur niet
goed helpt
• veel patienten hebben recidiverende C.diff infecties
• soms/vaak zijn die levensbedreigend
• vaak is dan de enige optie nog .....
• FECES TRANSPLANTATIE
• cure-rate van ~90% !!
• geen medicijn is zo goed !
Antibiotica gebruik & C. difficile

46. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 46
Feces transplantaties.....
....

47. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 47
Graad A
Graad B
Graad C
Graad D

48. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 48
Do you (have to) believe what a company tells you?
Wanneer geen probiotica?
immunocompromised?
kritiek zieke mensen?
zeer jong kinderen?
ouderen met een verminderde
weerstand?
zwangere vrouwen?
In veel van deze gevallen worden probiotica al
gebruikt.

49. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 49

50. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 50

51. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 51
understatement
Bij veel van de hedendaagse problemen kunnen probiotica een
gezonde bijdrage leveren

52. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 52
Wrap up and
Take home messages

53. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 53
Conclusies Deel 1
• Probiotica werken! Maar het maakt
wel uit welke stam (of mengsel) je
kiest.

54. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 54
Script van deel 2 van de presentatie
 gut-brain axis
 rol van microbiota
 preclinische aanwijzingen
 klinische studies
 niet alle probiotica zijn hetzelfde
 Take home messages deel 2

55. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 55
.... want de microbiota is betrokken bij:
- colon kanker
- inflammatory bowel disease (IBD)
- allergie (huid, luchtwegen)
- overgewicht
- type I en II diabetes
- levensduur
- prikkelbaar darm syndroom (IBS)
- brein ontwikkeling en cognitie
- autisme
- .....
control rat autistic rat

56. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 56
Anxiety
BALB/c SPF mice w/o antibiotics
ATM = neomycin 5 mg/mL, bacitracin 5 mg/mL, and pimaricin 1.25 μg/mLAfter ATM: more explorative

57. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 57
Anxiety
BALB/c SPF mice w/o antibiotics
After ATM: biochemical
changes in the brain.
BDNF: Brain-derived neurotrophic factor,
a protein which is active in areas vital to
learning, memory, and higher thinking

58. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 58
Anxiety – microbiota transplant

59. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 59
Anxiety – microbiota transplant

60. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 60
Birth delivery mode alters anxiety-like behaviour (8-9 wks)
Original setup
Natural birth Cross-fostered C-section
marble bury test

61. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 61
Stress alters the
gut-brain axis
cortisol
cytokines
adrenocorticotropic hormone
serotonin (5-HT)
barrier function of the gut
Increased permeability
translocation of endotoxin
increased colonic transit
mucosal inflammation
changes in colonic morphology
etc.
Altered levels and metabolism of:
Altered:
bi-directional
crosstalk

62. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 62
Immune
O’Mahony et al. Psychopharmacology (2011)
Endocrine
Visceral Pain
Open Field Increased colonic transit, intestinal
permeability, mucosal inflammation, colonic
morphology (O’Malley et al., 2010, Stress)

63. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 63
Autism Spectrum Disorders
control rat autistic rat + PA
Single intracerebroventricular (ICV)
infusions (4 µL of 0.26 M solution over
4 min) of propionic acid (PA)

64. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 64
http://apc.ucc.ie
Desbonnet et al 2014

65. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 65
Social deficit after propionate infusion
control rat autistic rat + PA
Single intracerebroventricular (ICV)
infusions (4 µL of 0.26 M solution over
4 min) of propionic acid (PA)

66. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 66
Odds Ratio: 1.23
(1.07, 1.40)
1,191,920
participants,
12,897 of which
had ASD

67. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 67
Parkinson’s disease
Parkinson’s disease is the second most common
neurodegenerative disease and is characterized by region-
specific loss of dopaminergic neuromelanin-containing
neurons in the brain
Parkinson’s disease patients suffer from gastrointestinal
discomfort
nearly half of all Parkinson’s disease patients report long-
term constipation
Parkinson’s disease patients had a significantly increased
intestinal permeability
Future studies might elucidate further links between gut and
brain function in Parkinson’s disease, and should preferably
comprise investigations of the gut microbiota.
Pro- and/or prebiotics that strengthen the epithelial barrier
function might be beneficial to these patients

68. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 68
Amyloid protein precursor (APP) is a pivotal protein in the
formation of the plaque-inducing amyloid-β protein, which
causes neuronal degeneration in the brain of Alzheimer’s
disease patients
systemic administration of LPS in mice has been shown to
increase APP expression and caused neuronal cell death
significantly increased plasma levels of LPS in Alzheimer’s
disease patients compared with healthy controls were
detected, which might suggest that Alzheimer’s disease
patients suffer from elevated intestinal permeability
Pro and/or prebiotics that have an anti-inflammatory effect or
reduce bacterial and/or LPS translocation might be beneficial
for these patients
Alzheimer’s disease

69. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 69
Probiotic alters GABA
receptors in various
brain areas
Bravo et al., PNAS Sept 2011
L. rhamnosus JB-1 reduces anxiety and
behavioural despair

70. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 70
Defensive Marble Burying
Elevated Plus Maze
174 1205
Esc = escitalopram

71. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 71
Log10[Latency](s)
W
ater/Sal
W
ater/LPS
B
G
O
S/Sal
B
G
O
S/LPS
0.0
0.5
1.0
1.5
2.0
*
*
*
Timeinlight(s)
W
ater/Sal
W
ater/LPS
B
G
O
S/Sal
B
G
O
S/LPS
0
25
50
75
100
*
*
*
Prebiotic BGOS feeding is anxiolytic
less anxious

72. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 72
[BDNF](pg/mgprotien)
frontal cortex hippocampus
0
50
100
150
200 *water
FOS
GOS
Anxiolytic changes coincided with
changes in brain biochemistry, and....

73. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 73
0.0
0.2
0.4
0.6 *
*
[TNFa](pg/mgprotein)
0
2
4
6
8
** *
[IL1b](pg/mgprotein)
0.00
1.75
3.50
5.25
7.00
*
[IL6](pg/mgprotein)
0.0
3.5
7.0
10.5
14.0
[IL10](pg/mgprotein)
.... a reduction in inflammatory cytokines

74. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 74
Human trials

75. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 75
Human trials (1)
in order of publication date
• 124 healthy volunteers; mean age 61.8 years
• Double-blind, placebo (milk) controlled study
• Lactobacillus casei Shirota in milk-drink for 3
weeks
• Mood and cognition were measured at baseline,
and after 10 and 20 days of consumption.

76. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 76
Human trials (1); LcS
POMS = profile of mood state
• An unexpected finding was that
the consumption of probiotics
resulted in a slightly-poorer
performance on two measures
of memory.

77. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 77
• 31 chronic fatigue syndrome patients
• RDBPC study
• Lactobacillus casei Shirota in sachet-format for 8
weeks
• Beck Depression Inventory (BDI) and the Beck
Anxiety Inventory (BAI) at baseline and after 8 w.
Human trials (2)

78. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 78
Human trials (2); LcS
• there was a significant improvement in anxiety among those taking the
active LcS compared to the placebo. The differences as assessed by the
BDI did not reach statistical significance.
BDI: a 21-question multiple-choice self-report inventory, one of the most widely used psychometric tests for
measuring the severity of depression
BAI: is a 21-question multiple-choice self-report inventory that is used for measuring the severity of anxiety in
children and adults

79. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 79
• 55 volunteers with a score of ≤12 in the HADS-
anxiety subscale and in the HADS-depression
subscale and ≤ 20 in the HADS total score on initial
examination
• RDBPC study
• L.helveticus R0052 + B. longum R0175 in a stick
format, or placebo (xylitol, maltodextrin, plum
flavour and malic acid) for 30 days
• Tests: HSCL-90; HADS; PSS, CCL
• Urinary cortisol
Human trials (3)

80. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 80
Human trials (3); L.h R0052 / B.l R0175
Hopkins Symptom Checklist-90.The HSCL-90 is a 90-item self-reported
multidimensional questionnaire screening a broad range of psychopathological
disorders
Hospital Anxiety and Depression Scale: The HADS, is a 14-item self-assessment
instrument, often applied and convenient for measuring psychological distress in
subjects with somatic or psychosomatic disorders
Perceived Stress Scale: The PSS is a 14-item self-reported questionnaire
assessing the degree to which recent life situations are appraised as stressful
Coping Checklist: The CCL is a validated 29-item questionnaire measuring five
types of coping strategies when confronting an adverse event: ‘problem solving’;
‘avoidance with wishful thinking’; ‘seeks social support’; ‘positive re-evaluation’;
‘self-blamed’.

81. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 81
Human trials (3); L.h R0052 / B.l R0175
Probiotics-treated subjects had a lower global severity index of the HSCL-90 over
time, due to lower values for somatisation, depression and anger–hostility.
The potential usefulness of the probiotics as an anti-stress/anti-anxiety agent was
supported by diminished HADS global scores over time.
Taken together, probiotics appears to show a beneficial effect on general signs of
anxiety and depression, but this did not generalise to the PSS, although all three
tests comprise self-reported measures.
In the CCL test the two groups differed in emotional reactivity, with subjects
administered probiotics reducing their self-blame score.
L. helveticus R0052 and B. longum R0175 taken in combination display
beneficial psychological effects in healthy human volunteers

82. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 82
• Same study as the previous one, but now in a
subpopulation of 25 subjects with urinary free
cortisol (UFC) levels less than 50 ng/ml at baseline
• More scores were beneficially affected by the
probiotics in this subpopulation
Human trials (3a)

83. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 83
• 36 healthy women, age 30 y +/- 10.4
• RDBPC study
• B. animalis subsp lactis DN-173 010, Streptococcus
thermophilus CNCM I-1630, 2 strains of L. bulgaricus,
CNCM I-1632 and CNCM I-1519 and Lactococcus lactis
subsp lactis CNCM I-1631 fermented milk, non-
fermented milk or no intervention for 4 weeks
• functional magnetic resonance imaging before and after
the intervention to measure brain response to an
emotional faces attention task and resting brain activity.
Human trials (4)

84. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 84
Human trials (4)
A fermented milk product with probiotic resulted in robust alterations in the
response of a widely distributed brain network to a validated task probing
attention to negative context, which have been associated with the dampening
of emotional and sensory responsiveness.

85. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 85
• 44 university students who performed strenuous
exercise daily
• RDBPC study
• Lactobacillus gasseri OLL2809, OLL2809 + α-
lactalbumin, or placebo for 4 weeks
• Profile of Mood States (POMS) questionnaire
Human trials (5)

86. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 86
Human trials (5)
LG2809 ingestion was effective in elevating mood from a depressed state.

87. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 87
• 224 healthy volunteers
• RDBPC study
• yogurt containing L. gasseri SBT2055 and B.
longum SBT2928, or control yoghurt for 12 weeks
• levels of two stress-induced hormones,
adrenocorticotrophic hormone (ACTH) and cortisol
Human trials (6)

88. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 88
Human trials (6)
These results suggest that the probiotic-containing test yogurt confers
health benefits by alleviating stress.

89. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 89
• 50 healthy participants, mean age 23.22 years
• RDBPC study
• two strains of L. acidophilus CUL60 and CUL21,
B.lactis CUL34 and B.bifidum CUL20 in a capsule, or
placebo for 6 weeks
• anxiety levels decreased in the probiotic condition
whilst increasing in the placebo condition (p=0.042).
• ‘continuity of attention’ increased in response to the
probiotic treatment and decreased with the placebo
• group (p=0.035)
Human trials (7)
Abstract

90. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 90
• 75 infants
• RDBPC study
• L. rhamnosus GG or placebo during the first 6 months
of life
• attention deficit hyperactivity disorder (ADHD) and
Asperger syndrome (AS)
• At the age of 13 y, ADHD or AS was diagnosed in 6/35
(17.1%) children in the placebo and none in the
probiotic group (P = 0.008).
Human trials (8)

91. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 91
• 40 healthy participants without current mood disorder
• RTBPC study
• a multispecies probiotic containing B. bifidum W23,
B.lactis W52, L.acidophilus W37, L.brevis W63,
L.casei W56, L.salivarius W24, and 2 strains of
Lactococcus lactis W19 and W58 in a sachet, or
placebo
• cognitive reactivity to sad mood was assessed using
the revised Leiden index of depression sensitivity
scale
Human trials (9)

92. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 92
Human trials (9)
The LEIDS-r is a self-reported
questionnaire with 34 items that
assesses to what extent
dysfunctional thoughts are
activated when experiencing mild
dysphoria
(i.e., it measures cognitive
reactivity to sad mood, also
referred to as vulnerability to
depression).
Rumination is the compulsively focused attention on the
symptoms of one's distress, and on its possible causes
and consequences, as opposed to its solutions.

93. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 93
• 30 patients with laryngeal cancer
• RDBPC study
• Unknown strain of Clostridium butyricum for 2
weeks, twice a day in a capsule, or placebo
• Serum corticotropin-releasing factor
• The degree of anxiety assessed by the Hamilton
Anxiety Scale (HAMA)
Human trials (10)

94. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 94
Human trials (10) m
Probiotics can ameliorate the clinical anxiety and biochemical features
of stress in patients scheduled for laryngectomy (and other surgery?)

95. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 95
• 45 healthy medical students under academic
examination stress
• RDBPC study
• L.casei Shirota fermented milk, or placebo for 8
weeks prior to taking national exam
• Subjective anxiety scores
• Salivary cortisol levels
• The presence of physical symptoms
Human trials (11)
[Epub ahead of print]

96. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 96
Human trials (11)
LcS may prevent hypersecretion of cortisol and physical symptoms (not shown
in this presentation) under stressful conditions, possibly through vagal afferent
signaling to the brain and reduced stress reactivity in the brain (the latter based
on a parallel rat study)

97. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 97
• 45 healthy volunteers with no history of psychiatric
disorders
• Double-blind, placebo (maltodextrin) controlled
study
• FOS, Bimuno®-galacto-oligosaccharides, or
maltodextrin, daily for 3 weeks
• Awakening salivary cortisol measured on day 0 and
day 21
• Emotional test battery on day 21
Human trials (12); prebiotic

98. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 98
Dot-probe task:
Attention to positive
stimuli is increased
BGOS reduces awakening cortisol and increases attention to positive
stimuli in healthy volunteers
Human trials (12); prebiotic

99. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 99
Other things that could not be
covered in this presentation
The gut microbiota produces other neuromodulatory molecules, such as
serotonin, GABA, melatonin
The gut microbiota is involved in brain development and cognition
The gut microbiota affects the blood brain barrier
Since pro- & prebiotics modulate the gut microbiota composition and
activity, they may also be involved in happiness, trustworthiness,
depression, sleeping, etc. … **
… in addition to anxiety, stress and autism
There is still room for mouse-models to study these gut microbiota-brain
interactions
** time for an anekdote?

100. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 100
Wrap up and
Take home messages

101. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 101
Conclusions Part 2
• The gut microbiota is involved in brain functioning
• Pro-, pre- (and synbiotics) modulate the gut microbiota and
thereby can influence brain functioning as well
• Although mostly evidenced in animal studies, some clinical trials
show very promising results

102. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 102
Script van deel 3 van de presentatie
 hoe werkt dat nou?
 wat kan ik er mee in de praktijk
 take home messages deel 3

103. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 103
Bepalen onze bacteriën in de darm wat we
in de supermarkt uit de schappen halen?

104. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 104
Hoe werken probiotica?
Hoe kunnen op het brein inwerken als ze in de darm zitten?

105. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 105
bi-directional
crosstalk
100 trillion bacteria
60-70% of immune cells
100 billion neuronal cells
communication from gut to brain
communication from brain to gut

106. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 106
1. Stress
a) barrier
b) transit
c) microbiota
2. Food intake
a) satiety
b) mood
3. Immunomodulation
a) cytokines via blood
4. Metabolite production
a) SCFA, GABA, 5-HT, etc
b) via blood, immune cells, ENS
5. Interaction with ENS
a) via vagus nerve
b) via blood 12 34 5

107. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 107

108. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 108

109. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 109

110. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 110
Potential effects of pro- and prebiotics also, e.g., obesity, allergy, infection
also, e.g, inflammation
also, e.g., obesity, allergy, inflammation
but also, role in prevention of weight gain
also, e.g., kidney failure
e.g., IBS

111. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 111

112. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 112
Saccharolytic and proteolytic activity
• Saccharolysis leads primarily to SCFA and gasses
• SCFA: acetate, propionate, butyrate
• lactate -> only accumulates when there
is a fast fermentation
• CO2, CH4, H2
• Proteolysis in addition leads to toxic metabolites
• BCFA: iso-butyrate, iso-valerate
• ammonia
• phenolics: phenol, indol, p-cresol, skatol
• H2S, CH3SH, etc

113. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 113
Colonic metabolites
some examples
• Butyrate:
• 70% of the fuel of colonocytes
• lack -> epithelial atrophy
• reduces cancer promotion in transformed tissue
• possibly reverses transformation by induction of
differentiation
• H2S, CH3SH:
• block butyrate oxidation
• ammonia:
• liver cirrhosis
• etc.

114. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 114
Prolonged carbohydrate fermentation
proximal colon: primarily saccharolytic
fermentation
transverse colon: combination of saccharolytic
and proteolytic fermentation
distal colon: primarily proteolytic fermentation
distal colon: colon cancer and IBD

115. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 115
microbiota host
Host – microbe interaction :
relevance for brain health

116. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 116
diet
fermentation by
gut microbiota
microbial
metabolites
tissue and organ
metabolism
host physiological
response

117. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 117
Dieet-adviezen?
Afhankelijk van achterliggend probleem
overgewicht/obesitas
prikkelbare darm
coeliakie
darmontsteking
allergie
In bijna alle gevallen (behalve autisme)
vezels (groeibron voor microbiota -> SCFA, etc.)
Probiotica
diarree, constipatie
ontstekingsproblemen (huid, luchtwegen, darm)

118. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 118
Dieet-adviezen? (2)
Ouderen
goed verteerbaar eiwit
vitaminen
vezels
Brein
mood/humeur -> tryptofaan (aminozuur in eiwit)
memory/geheugen -> probiotica?
ontwikkeling -> PUFAs
Stress (allerlei mogelijke vormen) -> verstoorde darm barriere
probiotica
glucose/glutamine

119. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 119
Wrap up and
Take home messages

120. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 120
Conclusions Part 3
• Pro- en prebiotica kunnen op veel verschillende manieren de
darm-brein interactie moduleren.
• Effecten zijn stam-specifiek
• Dieet-advies sterk afhankelijk van achterliggende probleem
• For more information:
koen.venema@outlook.com
k.venema@maastrichtuniversity.nl

121. Yakult seminar “Gut – brain – microbiota” – Almere – 13-Oct-2016 © Beneficial Microbes Consultancy 121
Acknowledgements
• Henrike Hamer
• Steven Vanhoutvin
• Freddy Troost
• Daisy Jonkers
• Robert-Jan Brummer
• Tao He
• Roel Vonk
April 20, 2011TIFN Satiety
screenings
study
121
Dank u hartelijk voor uw aandacht!