Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex and debilitating disease with no molecular diagnostics and no treatment options. To identify potential markers of this illness, we profiled 48 patients and 52 controls for standard laboratory tests, plasma metabolomics, blood immuno-phenotyping and transcriptomics, and fecal microbiome analysis. Here, we identified a set of 26 potential molecular markers that distinguished ME/CFS patients from healthy controls. Monocyte number, microbiome abundance, and lipoprotein profiles appeared to be the most informative markers. When we correlated these molecular changes to sleep and cognitive measurements of fatigue, we found that lipoprotein and microbiome profiles most closely correlated with sleep disruption while a different set of markers correlated with a cognitive parameter. Sleep, lipoprotein, and microbiome changes occur early during the course of illness suggesting that these markers can be examined in a larger cohort for potential biomarker application. Our study points to a cluster of sleep-related molecular changes as a prominent feature of ME/CFS in our Japanese cohort.