The early-life gut microbiome and pediatric asthma Marie-Claire - PowerPoint PPT Presentation

The early-life gut microbiome and pediatric asthma Marie-Claire Arrieta Ph.D Departments of Physiology & Pharmacology and Pediatrics

Graham-Rowe et al . 2011. Nature. Nov.;S2-S4. The global distribution is disproportionate.

Animal models Ovalbumin Model of Allergic Asthma Sensitization Challenge 0 7 14 21 22 23 24 25 26 Sacr. Intranasal OVA OVA IP OVA IP House Dust Mite Model of Allergic Asthma 0 14 21 22 23 25 Sacr. HDM IN HDM IN HDM IN

Epidemiology FARMS C-SECTIONS BREASTFEEDING ANTIBIOTICS HELMINTHS

Low resilience during critical immune development stage Laforest-Lapointe and Arrieta, 2017. Front Immun.

Experimental evidence !"#$%&'()&*+# (((,#$-.#/0123"( /$((( *%"2/0123"( (((4566(0789:(((((((45660789:( Ovalbumin Model of Allergic Asthma Sensitization Challenge * 0 7 14 21 22 23 24 25 26 Birth (Neonatal) Sacr. Intranasal OVA OVA IP OVA IP ------------------ Neonatal Abx Adult Abx 7 wks (Adult)

Neonatal antibiotic treatment affects experimental asthma severity Neonatal Abx Adult Abx A 25 25 * -1 -1 Total cells x 10 ml Total cells x 10 ml 20 20 5 5 15 15 n.s. 10 10 5 5 0 0 PBS OVA OVA + OVA + PBS OVA OVA + OVA + Vanco Strep Vanco Strep OVA-specific IgE Eosinophils 100 25 * ** 20 75 -1 cells x 10 ml -1 ng ml 4 15 n.s. 50 10 25 5 n.d. n.d. 0 0 PBS OVA OVA + OVA + PBS OVA OVA + OVA + Vanco Strep Vanco Strep Russell, et. al. EMBO Rep. 2012.

Hill, et. al. Nature Med. 2012.

Hill, et. al. Nature Med. 2012.

CHILD Study: Atopy Wheeze 87 subjects 136 subjects Atopy Only Wheeze Only 74 subjects Controls ≥ 1 episode of Positive allergen 22 subjects skin prick test at wheezing by 1- Atopy + 1-year Wheeze year of age 16S rRNA analysis at 3 Risk of asthma at school age [Asthma Predictive Index (Odds Ratio)] months and one year of age 15 1 5 10

D 3-months 1-year 3-months 1-year Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. Veillonella spp. *** *** 10000 1000 100 *** 1000 * 1.0 × 10 00 100 10 1.0 × 10 00 100 10 10 1 1.0 × 10 -02 1.0 × 10 -02 1 1 0.1 0.1 0.1 1.0 × 10 -04 0.01 0.01 1.0 × 10 -04 0.01 0.001 0.001 0.001 1.0 × 10 -06 0.0001 0.0001 1.0 × 10 -06 0.0001 0.00001 0.000001 1.0 × 10 -08 0.00001 0.00001 e s e e s s z l z l 1.0 × 10 -08 atopy wheeze e o controls z o o l Rothia spp. r e e e t e Rothia spp. r r h Rothia spp. n e h t t w o h Rothia spp. n n w w o o Rothia spp. c 1.0 × 10 -10 c y Rothia spp. c + + p o y CTRL AW y Controls AW t p a *** p o 1000 1000 o t *** t a 1000 10000 a Lachnospira spp. Lachnospira spp. Lachnospira spp. Lachnospira spp. 100 100 10 * 10 *** 100 10 1.0 × 10 00 1 1.0 × 10 -02 Relative Quantification (log10) 0.1 1 0.1 1 1.0 × 10 -02 0.01 1.0 × 10 -04 0.1 0.001 1.0 × 10 -04 0.01 0.001 0.01 0.0001 Relative Quantification 1.0 × 10 -06 0.001 0.00001 1.0 × 10 -06 Atopy + Wheeze Controls e s z l 0.00001 o 0.0001 e r Faecalibacterium spp. e t atopy wheeze h n controls Atopy + Wheeze Controls Faecalibacterium spp. w o 1.0 × 10 -08 c Faecalibacterium spp. 1.0 × 10 -08 y p Faecalibacterium spp. Faecalibacterium spp. o Faecalibacterium spp. t a 10000 * 1.0 × 10 -10 * 1.0 × 10 -10 1000 10000 10000 10000 L W Controls AW 100 1000 R A T 10 Rothia spp. Rothia spp. 100 Rothia spp. C 100 100 1 Rothia spp. 10 0.1 *** 1 1.0 × 10 -04 0.01 1.0 × 10 00 1 1 0.001 0.1 0.01 0.0001 1.0 × 10 -02 0.01 0.00001 0.01 1.0 × 10 -06 0.0001 0.001 0.000001 1.0 × 10 -04 e s z l atopy + wheeze controls e o e r h n t w o 0.000001 0.0001 c y Lachnospira spp. Lachnospira spp. Lachnospira spp. e s atopy wheeze p controls o Lachnospira spp. z l 1.0 × 10 -06 t e o a r 1.0 × 10 -08 e t h n Lachnospira spp. Lachnospira spp. 10000 w o c + 10000 Lachnospira spp. y 1.0 × 10 -08 1000 p o 1000 100000 t a 100 100000 100 1.0 × 10 -10 1.0 × 10 -10 10 10 Controls AW 1000 CTRL AW 1 1000 1 Faecalibacterium spp. Faecalibacterium spp. Faecalibacterium spp. 0.1 0.1 10 0.01 10 Faecalibacterium spp. 0.01 *** 0.001 0.001 1.0 × 10 -02 1.0 × 10 00 0.1 0.1 0.0001 0.0001 0.00001 0.00001 atopy wheeze controls e 1.0 × 10 -02 0.001 z s 0.001 e l 1.0 × 10 -04 o e r Bifidobacterium spp. Bifidobacterium spp. h t Bifidobacterium spp. Bifidobacterium spp. n W o C + 1.0 × 10 -04 0.00001 0.00001 y p 100000 e s 1000 o e t s 1.0 × 10 -06 z o l A z l e e o r e e t r 10000 h n Bifidobacterium spp. h t Bifidobacterium spp. n w o 1.0 × 10 -06 W o c 100 C y 1000 p + o y t 1.0 × 10 -08 a 10000 p 1000000 100 10 o 1.0 × 10 -08 t A 10 1 10000 1 1.0 × 10 -10 100 1.0 × 10 -10 0.1 CTRL AW Controls AW 0.1 100 Bifidobacterium spp. 0.01 Bifidobacterium spp. Bifidobacterium spp. Bifidobacterium spp. 1 0.01 0.001 1 1.0 × 10 -02 1.0 × 10 00 0.0001 0.001 atopy wheeze controls Atopy + Wheeze Controls 0.01 0.01 1.0 × 10 -02 1.0 × 10 -04 0.0001 0.0001 1.0 × 10 -04 Atopy + Wheeze Atopy + Wheeze Controls Controls Controls Atopy + Wheeze atopy wheeze controls 1.0 × 10 -06 1.0 × 10 -06 1.0 × 10 -08 1.0 × 10 -08 1.0 × 10 -10 1.0 × 10 -10 Controls AW CTRL AW Arrieta et al. 2015 Sci. Transl. Medicine .

Gut bacterial metabolism: short-chain fatty acids (SCFA) SCFA: • Used as energy source by intestinal and mucosal immune cells. • Transported to the liver where they are involved in numerous biochemical pathways of lipid and energy metabolism. • Involved in immune response regulation.

Acetate Propionate Butyrate * A * * 2.0 40 8 umol/g feces umol/g feces umol/g feces 1.5 30 6 4 1.0 20 2 0.5 10 0 0.0 0 L W CTRL A + W CTRL A + W R + T A C Urine metabolites B glycolithocholate glycocholenate glycohyocholate tauroursodeoxycholate sulfate sulfate * ** * ** 6 3 × 10 5 10 * 8 * * * 8 6 Pathway 4 2 × 10 5 glycohyocholate 6 oursodeoxycholate 4 4 Secondary bile acid metabolism 2 1 × 10 5 2 2 Hemoglobin metabolism 0 0 0 0 Phenylalanine metabolism phenylacetylglutamine imidazole propionate I-urobilinogen ferulate ** * * 1.5 6 25 P = 0.052 * * 80 * Histidine metabolism * 20 60 1.0 4 15 Food component 10 0.5 2 20 5 0 0 0.0 0 CTRL AW CTRL AW CTRL AW L W R A Arrieta, et al 2015 Sci. Transl. Medicine . T C

From correlation to causation Hu Human man mic microb obiot iota a tran ansfer er (inoculated with feces from AW subject) Ovalbumin breeding offspring mice OVA (i) Control Asthmatic Microbiota birth 3 wks 7 wks (AW) (AW) OVA (ii) FLVR AW + FLVR birth 3 wks 7 wks 14

Microbial intestinal colonization in F1 mice B ����������� A AW ���� 60 ���� AW + FLVR �� Fa Family 100 40 �� 100 PC 2 (24.8%) 20 Bacteroidaceae Bacteroidaceae �� �������������� 0 Clostridiaceae � Clostr -20 ��� 75 Enterobacteriaceae 75 Enterobacter -40 ��� Enterococcaceae Enterococcaceae -60 ��� ce (%) Lachnospiraceae Lachnospir -60 -40 -20 0 20 40 60 ��� ��� ��� � �� �� �� 50 �������������� 50 Abundance PC 1 (41.5%) Micrococcaceae C Micrococcaceae Peptostreptococcaceae Peptostreptococcaceae 1.5 60 0.6 60 Prevotellaceae Pre ce (%) 25 25 1.0 Abundance 40 Ruminococcaceae 0.4 Ruminococcaceae 40 S24-7 S24 0.5 20 0.2 20 Turicibacteraceae 0 Tur 0 CTRL VLRF AW AW AW AW + 0 0 0.0 0 Lach chnosp spira Faeca calibact cterium Ro Rothia Ve Veillonella FLVR FLV Faecalibacterium Rothia Veillonella Lachnospira Arrieta, et al 2015 Sci. Transl. Medicine .

Na Naive FLVR reduces lung inflammation Bronchoalveolar lavage AW AW BAL Total Cell Differential BAL Counts Eosinophils * Neutrophils Lymphocytes ** * 80 Macrophages 200 **** cells(x10 4 )/ml 60 cells (x10 3 )ml 150 40 100 20 50 0 0 AW AW + FLVR AW + Naive AW Naive AW AW + FLVR FLVR Arrieta, et al 2015 Sci. Transl. Medicine . 16

What about asthma elsewhere? Las Esmeraldas, Ecuador Prof. Phil Cooper 29 AW 71 Controls 3 months Forno et al. Thorax 2015

Ru Rural t to u urban tr transiti tion

Ecuador cohort: epidemiology Categorical variable Odds P ratio Maternal antibiotics during 2.39 0.02 pregnancy Vaginal birth -2.42 0.03 Breastfeeding over 5 months -0.17 0.02 Child Ascaris infection 3.82 0.05 Child Trichuris infection -8.18 0.004 Respiratory infections 2.21 0.001 Potable water 2.49 0.02

Microbiota in babies at risk of asthma Bacteria Phylum Firmicutes Bacteroidetes Veillonella_4 Acetate Actinobacteria Veillonela_26 150 µ mol/g feces Streptococcus_5 * 100 Streptococcus_9 Streptococcus_10 50 Ruminococcus_gnavus_18 0 Bacteroides_7 AW CTRL Bifidobacterium_179 8 6 4 2 0 2 4 6 8 - - - - log 2 Fold Change CTRL:AW Arrieta, et al 2017 J Allergy Clin Immunol.