]; the threshold of logarithmic LDA score was 2.0. Spearman’s correlation coefficient was calculated for bacterial species that have more than 0.5% relative abundance in 12-month-old recipient mice using the psych R package.To examine the cognitive response of mice, a passive avoidance test was performed. A mouse was placed in a light chamber for 30 s and allowed to move to a dark chamber upon gate opening.
For the intraocular pressure test, mice were anesthetized using an intraperitoneal injection of xylazine and zolazepam and tiletamine . IOP was measured using a rebound tonometer , according to the manufacturer’s instructions. One trial result was obtained after six consecutive measurements, and the mean of consecutive trials was used for analyses.
Electroretinogram analysis was performed using Micron Ganzfeld ERG . The mouse was dark-adapted at least 12 h before the experiment for scotopic testing . After anesthesia, the pupil dilated as previously described. Once the pupil was fully dilated, we applied hypromellose 2.5% and inserted the electrodes. ERG was recorded using Micron Ganzfeld ERG, according to the manufacturer’s instructions. Scotopic ERG was obtained with increasing flash intensity at a range of −1.7 log cd/s/m.
For immunofluorescence, tissue sample preparation and blocking were performed in the same manner as described in immunohistochemistry. Then, the slides were probed at 4 °C overnight with the primary antibodies , labeled for 2 h with Cy3-conjugated goat anti-rabbit IgG or A488 donkey anti-mouse IgG , and stained with DAPI for 15 min.
QuantSeq 3′ mRNA-Seq reads were aligned using Bowtie2. Bowtie2 indices were either generated from the genome assembly sequence or representative transcript sequences for aligning to the genome and transcriptome. The alignment file was used for assembling transcripts, estimating their abundances, and detecting differential expression of genes. Raw read counts were normalized and differentially expressed genes were identified using the edgeR package .
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The gut microbiota in multiple sclerosis varies with disease activity - Genome MedicineBackground Multiple sclerosis is a chronic immune-mediated disease of the brain and spinal cord resulting in physical and cognitive impairment in young adults. It is hypothesized that a disrupted bacterial and viral gut microbiota is a part of the pathogenesis mediating disease impact through an altered gut microbiota-brain axis. The aim of this study is to explore the characteristics of gut microbiota in multiple sclerosis and to associate it with disease variables, as the etiology of the disease remains only partially known. Methods Here, in a case-control setting involving 148 Danish cases with multiple sclerosis and 148 matched healthy control subjects, we performed shotgun sequencing of fecal microbial DNA and associated bacterial and viral microbiota findings with plasma cytokines, blood cell gene expression profiles, and disease activity. Results We found 61 bacterial species that were differentially abundant when comparing all multiple sclerosis cases with healthy controls, among which 31 species were enriched in cases. A cluster of inflammation markers composed of blood leukocytes, CRP, and blood cell gene expression of IL17A and IL6 was positively associated with a cluster of multiple sclerosis-related species. Bacterial species that were more abundant in cases with disease-active treatment-naïve multiple sclerosis were positively linked to a group of plasma cytokines including IL-22, IL-17A, IFN-β, IL-33, and TNF-α. The bacterial species richness of treatment-naïve multiple sclerosis cases was associated with number of relapses over a follow-up period of 2 years. However, in non-disease-active cases, we identified two bacterial species, Faecalibacterium prausnitzii and Gordonibacter urolithinfaciens, whose absolute abundance was enriched. These bacteria are known to produce anti-inflammatory metabolites including butyrate and urolithin. In addition, cases with multiple sclerosis had a higher viral species diversity and a higher abundance of Caudovirales b
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