Sex steroids are involved in the development of obesity and metabolic disorders. The interrelationship between sex steroid balance and metabolism is complex, and the underlying mechanisms are still unclear. The impact of gut microbiota in development of obesity, inflammation and metabolic dysregulation has been suggested in humans and demonstrated in mouse models. Less is known about the interactions between sex steroids and gut microbiota. Based on available evidence, it is, however, pertinent to ask if sex steroids participate in the regulation of the composition or function of gut microbiota, or if gut microbes regulate sex steroid balance. The aim of the present study was to define the interplay between sex steroid balance and gut microbiota, in the context of obesity and metabolic dysregulation. First, we showed that non-obese human aromatase expressing (AROM+) male mice, with increased circulating estradiol to testosterone ratio and impaired liver lipid metabolism, have a female-type intestinal microbiota and higher ratio of Firmicutes to Bacteroidetes. Treatment with aromatase inhibitor restored estradiol/testosterone levels, followed by a slight improvement in their microbiota ecology. Second, in a mouse model reporting the expression of human aromatase (CYP19A1) gene (hARO-Luc mouse), ovariectomy-induced obesity, accompanied with elevated Firmicutes to Bacteroidetes ratio, altered gut homeostasis and increased aromatase reporter gene expression in the mammary adipose tissue. In conclusion, by applying two gene-modified mouse models, we showed that changes in sex steroid balance are linked to altered gut microbiota composition, and on the other hand, sex steroid –related obesity is linked to increased expression of human aromatase gene reporter. Further experiments, such as fecal transplantation from mice with altered sex steroid balance to WT mice, are needed to confirm the causality between sex steroids, gut microbiota and metabolic disorders.