C. bolteae is elevated in NMOSD in India and shares sequence similarity with AQP4

Objective: To understand the role of gut microbiome in influencing the pathogenesis of neuromyelitis optica spectrum disorders (NMOSD) among patients of south Indian origin.Methods: In this case control study, stool and blood samples were collected from 39 NMOSD patients, including 17 with aquaporin 4 IgG antibodies (AQP4+) and 36 matched controls. 16S rRNA sequencing was used to investigate the gut microbiome. Peripheral CD4+ T cells were sorted in 12 healthy controls & 12 AQP4+ NMOSD patients, RNA extracted, and immune gene expression analyzed using Nanostring nCounter human immunology kit code set.Results: Microbiota community structure (beta-diversity) differed between AQP4+ NMOSD and healthy controls (p <0.001, pairwise PERMANOVA test). Linear discriminatory analysis effect size (LEfSe) identified several members of the microbiota that were altered in NMOSD patients, including an increase in Clostridium bolteae (effect size 4.23, p value 0.00007). C.bolteae was significantly more prevalent (p=0.02) among AQP4-IgG + NMOSD (n= 8/17 subjects) compared to seronegative patients (n= 3/22) and was absent among healthy stool samples. C bolteae has a highly conserved glycerol uptake facilitator and related aquaporin protein (p59-71) that shares sequence homology with AQP4 peptide (p92-104), positioned within an immunodominant (AQP4 specific) T cell epitope (p91-110). Presence of C.bolteae correlated with expression of inflammatory genes associated with both innate and adaptive immunity and particularly involved in plasma cell differentiation , B cell chemotaxis and Th17 activation.Conclusion: Our study described elevated levels of C. bolteae associated with AQP4+ NMOSD among Indian patients. It is possible that this organism may be causally related to the immunopathogenesis of this disease in susceptible individuals.