Melissa Barker‐Haliski

Objective Brain infection with Theiler's murine encephalomyelitis virus (TMEV) in C57BL/6J mice can induce acquired epileptogenesis. Diet alters acute seizure incidence in TMEV‐infected mice; yet it is unclear whether intestinal dysbiosis may also impact acute or chronic behavioral comorbidities. This study thus assessed the impact of diet formulation and sterilization on acute seizure presentation, gut microbiome composition, and epilepsy‐related chronic behavioral comorbidities. Methods Baseline fecal samples were collected from male C57BL/6J mice (4‐ to 5‐weeks‐old; Jackson Labs) upon facility arrival. Mice were randomized to either autoclaved (AC) or irradiated diet (IR) (Prolab RMH 3000) or IR (Picolab 5053). Three days later, mice underwent intracerebral TMEV or phosphate‐buffered saline (PBS) injection. Fecal samples were collected from a subset of mice at infection (Day 0) and Day 7 post‐infection. Epilepsy‐related working memory deficits and seizure threshold were assessed 6 weeks post‐infection. Gut microbiome diversity was determined by 16S rRNA amplicon sequencing of fecal samples. Results TMEV‐infected mice displayed acute handling‐induced seizures, regardless of diet: 28 of 57 IR Picolab 5053 (49.1%), 30 of 41 IR Prolab RMH 3000 (73.2%), and 47 of 77 AC Prolab RMH 3000 (61%) mice displayed seizures. The number of observed seizures differed significantly by diet: IR Picolab 5053 diet‐fed mice had 2.2 ± 2.8 seizures (mean ± standard deviation), IR Prolab RMH 3000 diet‐fed mice had 3.5 ± 2.9 seizures, and AC Prolab RMH 3000 diet‐fed mice had 4.4 ± 3.8 seizures during the 7‐day monitoring period. Gut microbiome composition differed significantly in TMEV‐infected mice fed the AC Prolab RMH 3000 diet, with measured differences in gram‐positive bacteria. These mice also displayed worsened long‐term working memory deficits. Significance Diet‐induced differences in intestinal dysbiosis in the TMEV model are associated with marked changes in acute seizure presentation, symptomatic recovery, and onset of chronic behavioral comorbidities of epilepsy. Our study reveals a novel disease‐modifying impact of dietary manipulation on intestinal bacterial species after TMEV‐induced acute seizures.