Marja I Roslund1, Anirudra Parajuli2, Nan Hui2, Riikka Puhakka2, Mira Grönroos2, Laura Soininen2, Noora Nurminen3, Sami Oikarinen3, Ondřej Cinek4, Lenka Kramná4, Anna-Mari Schroderus5, Olli H Laitinen3, Tuure Kinnunen6, Heikki Hyöty7, Aki Sinkkonen8. 1. Natural Resources Institute Finland (Luke), Helsinki and Turku, Finland; Ecosystems and Environment Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Niemenkatu 73, FI-15140 Lahti, Finland. 2. Ecosystems and Environment Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Niemenkatu 73, FI-15140 Lahti, Finland. 3. Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu 34, FI-33520 Tampere, Finland. 4. Department of Pediatrics, 2nd Faculty of Medicine, Charles University, V Úvalu 84, Praha 5, 150 06 Prague, Czech Republic. 5. Department of Clinical Microbiology, Institute of Clinical Medicine, University of Eastern Finland, Kuopio, Finland. 6. Department of Clinical Microbiology, Institute of Clinical Medicine, University of Eastern Finland, Kuopio, Finland; Eastern Finland Laboratory Centre (ISLAB), Kuopio, Finland. 7. Faculty of Medicine and Health Technology, Tampere University, Arvo Ylpön katu 34, FI-33520 Tampere, Finland; Fimlab Laboratories, Pirkanmaa Hospital District, Tampere, Finland. 8. Natural Resources Institute Finland (Luke), Helsinki and Turku, Finland. Electronic address: aki.sinkkonen@luke.fi.
Abstract
BACKGROUND: According to the biodiversity hypothesis of immune-mediated diseases, lack of microbiological diversity in the everyday living environment is a core reason for dysregulation of immune tolerance and - eventually - the epidemic of immune-mediated diseases in western urban populations. Despite years of intense research, the hypothesis was never tested in a double-blinded and placebo-controlled intervention trial. OBJECTIVE: We aimed to perform the first placebo-controlled double-blinded test that investigates the effect of biodiversity on immune tolerance. METHODS: In the intervention group, children aged 3-5 years were exposed to playground sand enriched with microbially diverse soil, or in the placebo group, visually similar, but microbially poor sand colored with peat (13 participants per treatment group). Children played twice a day for 20 min in the sandbox for 14 days. Sand, skin and gut bacterial, and blood samples were taken at baseline and after 14 days. Bacterial changes were followed for 28 days. Sand, skin and gut metagenome was determined by high throughput sequencing of bacterial 16 S rRNA gene. Cytokines were measured from plasma and the frequency of blood regulatory T cells was defined as a percentage of total CD3 +CD4 + T cells. RESULTS: Bacterial richness (P < 0.001) and diversity (P < 0.05) were higher in the intervention than placebo sand. Skin bacterial community, including Gammaproteobacteria, shifted only in the intervention treatment to resemble the bacterial community in the enriched sand (P < 0.01). Mean change in plasma interleukin-10 (IL-10) concentration and IL-10 to IL-17A ratio supported immunoregulation in the intervention treatment compared to the placebo treatment (P = 0.02). IL-10 levels (P = 0.001) and IL-10 to IL-17A ratio (P = 0.02) were associated with Gammaproteobacterial community on the skin. The change in Treg frequencies was associated with the relative abundance of skin Thermoactinomycetaceae 1 (P = 0.002) and unclassified Alphaproteobacteria (P < 0.001). After 28 days, skin bacterial community still differed in the intervention treatment compared to baseline (P < 0.02). CONCLUSIONS: This is the first double-blinded placebo-controlled study to show that daily exposure to microbial biodiversity is associated with immune modulation in humans. The findings support the biodiversity hypothesis of immune-mediated diseases. We conclude that environmental microbiota may contribute to child health, and that adding microbiological diversity to everyday living environment may support immunoregulation.
BACKGROUND: According to the biodiversity hypothesis of immune-mediated diseases, lack of microbiological diversity in the everyday living environment is a core reason for dysregulation of immune tolerance and - eventually - the epidemic of immune-mediated diseases in western urban populations. Despite years of intense research, the hypothesis was never tested in a double-blinded and placebo-controlled intervention trial. OBJECTIVE: We aimed to perform the first placebo-controlled double-blinded test that investigates the effect of biodiversity on immune tolerance. METHODS: In the intervention group, children aged 3-5 years were exposed to playground sand enriched with microbially diverse soil, or in the placebo group, visually similar, but microbially poor sand colored with peat (13 participants per treatment group). Children played twice a day for 20 min in the sandbox for 14 days. Sand, skin and gut bacterial, and blood samples were taken at baseline and after 14 days. Bacterial changes were followed for 28 days. Sand, skin and gut metagenome was determined by high throughput sequencing of bacterial 16 S rRNA gene. Cytokines were measured from plasma and the frequency of blood regulatory T cells was defined as a percentage of total CD3 +CD4 + T cells. RESULTS: Bacterial richness (P < 0.001) and diversity (P < 0.05) were higher in the intervention than placebo sand. Skin bacterial community, including Gammaproteobacteria, shifted only in the intervention treatment to resemble the bacterial community in the enriched sand (P < 0.01). Mean change in plasma interleukin-10 (IL-10) concentration and IL-10 to IL-17A ratio supported immunoregulation in the intervention treatment compared to the placebo treatment (P = 0.02). IL-10 levels (P = 0.001) and IL-10 to IL-17A ratio (P = 0.02) were associated with Gammaproteobacterial community on the skin. The change in Treg frequencies was associated with the relative abundance of skin Thermoactinomycetaceae 1 (P = 0.002) and unclassified Alphaproteobacteria (P < 0.001). After 28 days, skin bacterial community still differed in the intervention treatment compared to baseline (P < 0.02). CONCLUSIONS: This is the first double-blinded placebo-controlled study to show that daily exposure to microbial biodiversity is associated with immune modulation in humans. The findings support the biodiversity hypothesis of immune-mediated diseases. We conclude that environmental microbiota may contribute to child health, and that adding microbiological diversity to everyday living environment may support immunoregulation.
Authors: Jake M Robinson; James Aronson; Christopher B Daniels; Neva Goodwin; Craig Liddicoat; Laura Orlando; David Phillips; Jessica Stanhope; Philip Weinstein; Adam T Cross; Martin F Breed Journal: Lancet Planet Health Date: 2022-09