Multiple sclerosis (MS) is a chronic neurodegenerative disease that is the leading cause of disability in young adults. The most severe clinical form of MS is the primary progressive (PP). We and others have shown that melatonin (MLT), improves the disease modulating the immune response in the Experimental Autoimmune Encephalomyelitis (EAE) monofasic mouse model of MS. Thus, the objective of this study was to evaluate the therapeutic use of MLT in the neuroinflammation, neurodegeneration and gut dysbiosis in the preclinical model of MS-PP, the progressive EAE (p-EAE).p-EAE was induced in 10-weeks NOD/ShiLtJ mice and daily MLT (80 mg/kg) and vehicle were administered intraperitoneally from the onset of the clinical symptoms. The profile of infiltrated immune cells in the CNS of animals in the progressive phase of the disease was evaluated by multiparametric flow cytometry and RT-qPCR. Stool samples were collected, total DNA was extracted and 16S rRNA was sequenced in order to analyze gut microbiota composition.Treatment with MLT significantly reduced the severity of p-EAE, increased the animal survival as well as significant decreased the access of immune cells into the CNS, compared to controls. Decreased Th1 responses in CD4, CD8 and B lymphocytes, as well as an increase in the production of IL-10 were detected. Finally, MLT treatment altered gut microbiota population. Significant recovery of richness and diversity was observed in the MLT-supplemented group. Relative abundance of Akkermansia muciniphila and Ruminoccoccus genus was depleted in MLT treated mice when compared to non-treated animals.