THE RELATION BETWEEN ICHTHYOSIS VULGARIS, HOST GENOTYPE AND SKIN MICROBIOTA Microbiome R&D congress 3rd of April in Amsterdam Sacha van Hijum Principal scientist microbiomics at NIZO Associate professor at Radboud UMC sacha.vanhijum@nizo.com

FROM SCIENCE TO SOLUTIONS

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Applied science Fundamental science

core business contract

research

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joint research

academic research

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NIZO MICROBIOMICS PLATFORM - FROM UNDERSTANDING TO MODULATOR DISCOVERY -

Upscaling & production microbiome modulators

Discovery Platform

Microbiome Modulators

MicrobiOMICs Identification Microbiome

Markers Human POC, Challenge

Field trial

Microbiome markers of health/disease

DNA/RNA extraction

Sample collection & logistics

Expert consultation design

Sequencing

Data-analytics & Expert Interpretation In silico

In vitro strains

In vitro community

In vitro Host-microbe interactomics

PROFILING THE SKIN, VAGINAL AND GASTROINTESTINAL MICROBIOTA - TOWARDS CAUSALITY AND INTERVENTION STUDIES -

SKIN

Microbiota in health & disease: • Psoriasis, AD • Acne, dandruff, (mal)odour • Microskin • 3D skin models: host-microbe interactions

8/9 Lowerback

3 inner - elbow 1 Fronthead

Fronthead ~ P. acnes

inner - elbow Armpit

Fronthead

Forehead

Armpit Left

Inner Elbow Left

Lowerback

Women’s health GI-tract

Microbiota management - Dietary interventions studies - Microbial challenge experiments - Microbiota & Immunity - Microbiota & GI comfort - Infant formula studies - In vitro high throughput screening

Manage and modulate vaginal microbiota - Clinical study design and sampling - (Longitudinal) microbiota studies - In silico prediction of modulators - In vitro assays to test/validate modulators - Markers for monitoring women’s health

• Filaggrin knock-out (FLG-/-) causes ichthyosis vulgaris

• FLG: histidine-rich epidermal protein

• Degradation products serve as “natural moisturizing factors” (NMFs)

• Lower leg skin microbiome of 13 FLG-/- and 10 FLG+/+ subjects studied

HOW DOES FILAGGRIN GENOTYPE INFLUENCE CUTANEOUS MICROBIOTA?

About 50% of FLG-/- subjects also had atopic dermatitis

“Scaly skin”

Zeeuwen et al., J Allergy Clin Immunol, 2016

WORKFLOW MICROBIOME ANALYSIS

Study design, sample collection (skin swab, scraping, or tape strip)

Generation of DNA (DNA isolation; purification)

Amplification of the library by PCR (16S rRNA gene; V3-V4 region)

Data generation via NGS sequencing (Illumina)

Bioinformatics data analysis & expert interpretation QIIME, classification, visualization & statistical

analysis

(Barcoded) primer design and PCR optimization

16S microbiome profiling Contrast analysis Red: bacteria present in FLG+/+

Blue: bacteria present in FLG-/-

Peptoniphilic GPAC bacteria (G+ anaerobe cocci) are present in FLG+/+

HOW TO TEST SKIN MICROBES AND MODULATORS IN VITRO?

Benefits callus model • Grow various species, strain

combinations and microbiota • Test modulators together with

microbes

HOW TO TEST SKIN MICROBES AND MODULATORS IN VITRO?

Strain survival on callus model

IN VITRO EXPERIMENTS LINK HOST GENOTYPE, MODULATOR AND SKIN COMMENSAL

FLG+/+ callus has more NMFs F. magna grows better on FLG+/+ callus

(HIS, histidine; PCA, pyrrolidone carboxylic acid; UCA, urocanic acid)

• The microbial Hut pathway is predicted to be more abundant in FLG+/+ microbiome

• Benefits understanding pathways

• Establish causality

• Claim substantiation

• Design microbiome modulators

MICROBIOME PATHWAY PREDICTION TOWARDS CAUSALITY

Hut: histidine utilization pathway to degrade NMFs (PICRUSt applied to 16s data & KEGG based)

NIZO MICROBIOMICS PLATFORM - ICHTHYOSIS VULGARIS, SKIN MICROBIOTA & MODULATORS -

Upscaling & production microbiome modulators

Discovery Platform

Microbiome Modulators

MicrobiOMICs Identification Microbiome

Markers Human POC, Challenge

Field trial

Microbiome markers of health/disease

DNA/RNA extraction

Sample collection & logistics

Expert consultation design

Sequencing

Data-analytics & Expert Interpretation In silico

In vitro strains

In vitro community

In vitro Host-microbe interactomics

THANKS TO

Joost Schalkwijk

Patrick Zeeuwen

Tom Ederveen

Harro Timmerman

Jos Boekhorst

Sabina Lukovac

Saskia Schalkwijk