Nelson Mok ¹ Natalie C Knox ² Feng Zhu ³ Douglas L Arnold ⁴ Amit Bar-Or ⁵ Charles Noah Bernstein ¹ Christine Bonner ² Jessica D Forbes ⁶ Morag Graham ² Janace Hart ⁷ Ruth Ann Marrie ¹ Julia O'Mahony ⁸ E. Ann Yeh ⁸ Yinshan Zhao ³ Gary Van Domselaar ² Brenda Banwell ⁹ Emmanuelle Waubant ⁷ HELEN L TREMLETT ^3*

• ¹ University of Manitoba, Winnipeg, Canada
• ² Public Health Agency of Canada (PHAC), Ottawa, Ontario, Canada
• ³ University of British Columbia, Vancouver, British Columbia, Canada
• ⁴ McGill University, Montreal, Quebec, Canada
• ⁵ University of Pennsylvania, Philadelphia, Pennsylvania, United States
• ⁶ Eastern Ontario Regional Laboratory Association, Ottawa, Canada
• ⁷ University of California, San Francisco, San Francisco, California, United States
• ⁸ Hospital for Sick Children, University of Toronto, Toronto, Ontario, Canada
• ⁹ Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, United States

Evidence suggests that the gut microbiome may play a role in multiple sclerosis (MS). However, most studies have focused on gut bacterial communities; none have examined the fungal microbiota (mycobiota) in persons with pediatric-onset MS (POMS). We examined the gut mycobiota in persons with and without POMS through a cross-sectional examination of the gut mycobiota from 46 participants' stool samples (three groups: 18 POMS, 13 acquired monophasic demyelinating syndromes [monoADS], and 15 unaffected controls). Using metataxonomic sequencing of the fungal internal transcribed spacer region 2, the fungal profiles were compared between participants using visualizations, statistical tests, and predictive analysis. While the mycobiome alpha (Shannon, inverse Simpson) and beta diversity differed across the three groups (ANOVA p < 0.05), further post-hoc analysis of the beta-diversity identified a difference between monoADS versus POMS participants (p = 0.005 [adjusted]). At the genus level of taxonomy, 7/10 of the most abundant genera were similar between all three groups, with Saccharomyces spp. and Candida spp. being in the highest abundance. The Agaricus genus was especially high in POMS participants, dominated primarily due to the species A. bisporus (widely consumed as white button mushrooms). The commonality of high abundance fungi found in our cohort suggests a possible connection to diet. Predictive modeling of differential abundance associated C. albicans, Cyberlindera jadinii, and Fusarium poae most strongly with the POMS participants. Our study provides novel insight into the fungal gut mycobiota in POMS. While findings indicate that the gut mycobiome of participants with POMS may largely comprise fungi considered transient from the diet, the differential predictive analysis suggested rare or under-detected fungal markers being of potential importance, warranting consideration in future mycobiome-MS related studies.