Zuzana Jiraskova Zakostelska ¹ Michal Kraus ¹ Stepan Coufal ¹ Petra Procházková ¹ Zaneta Slavickova ¹ Tomas Thon ¹ Tomas Hrncir ² Jakub Kreisinger ³ Klara Kostovcikova ¹ Pavlina Kleinova ⁴ Jana Lizrova Preiningerova ⁴ Miluse Pavelcova ⁴ Veronika Ticha ⁴ Ivana Kovarova ⁴ Eva Kubala Havrdova ⁴ Helena Tlaskalova-Hogenova ¹ Miloslav Kverka ^1*

• ¹ Institute of Microbiology, Academy of Sciences of the Czech Republic (ASCR), Prague, Czechia
• ² Laboratory of Gnotobiology, Institute of Microbiology (ASCR), Novy Hradek, Prague, Czechia
• ³ Faculty of Science,Department of Zoology,Charles University, Prague, Czechia
• ⁴ Department of Neurology and Centre of Clinical Neuroscience, First Medical Faculty, Charles University and General Medical Hospital in Prague, Prague, Czechia

The gut microbiota influences the reactivity of the immune system, and Parabacteroides distasonis has recently emerged as an anti-inflammatory commensal. Here, we investigated whether its lysate could prevent severe forms of neuroinflammation in experimental autoimmune encephalomyelitis (EAE) in mice and how this preventive strategy affects the gut microbiota and immune response.Lysate of anaerobically cultured P. distasonis (Pd lysate) was orally administered to C57BL/6 mice in four weekly doses. One week later, EAE was induced and disease severity was assessed three weeks after induction. Fecal microbiota changes in both vehicle-and Pd lysate-treated animals was analyzed by 16S V3-V4 amplicon sequencing and qPCR, defensin production in the intestinal mucosa by RT-qPCR, and cell composition in the mesenteric and inguinal lymph nodes was measured by multicolor flow cytometry.Pd lysate significantly delayed the development of EAE and reduced its severity when administered prior to disease induction. EAE induction was the main factor in altering the gut microbiota, decreasing the abundance of lactobacilli and segmented filamentous bacteria. Pd lysate significantly increased the intestinal abundance of the genera Anaerostipes, Parabacteroides and Prevotella, and altered the expression of antimicrobial peptides in the intestinal mucosa. It significantly increased the frequency of regulatory T cells, induced an anti-inflammatory milieu in mesenteric lymph nodes, and reduced the activation of T cells at the priming site.Pd lysate prevents severe forms of EAE by triggering a T regulatory response and modulating T cell priming to autoantigens. Pd lysate could thus be a future modulator of neuroinflammation that increases the resistance to multiple sclerosis.