Long term, low-dose TCDD oral exposure changes gut microbial glycosylation patterns in female but not male C57BL/6 mice

TCDD at liver-damaging high doses can cause dysbiosis in the murine gut. We here ask if also a very low dose of TCDD also causes dysbiosis in mice and if there are sex-specific differences. Male and female C57BL/6 mice were orally administered an initial dose of 1 µg TCDD/kg body weight, diluted in corn oil, and maintenance doses every two weeks for 12 weeks; control mice received DMSO/corn oil only. We collected fecal pellets throughout the experiment. Cecal content was collected at the end upon sacrifice of the mice, as well as liver and gut tissue samples. Mice did not suffer weight loss, and the liver weight to body weight ratio remained normal in all groups. Barrier genes` expression was unaffected in all groups. Furthermore, liver histology at the end of the experiment did not reveal abnormalities. 16S sequencing of fecal pellets revealed an increase in alpha diversity over the 12 treatment weeks, irrespective of TCDD treatment or sex. Unifrac analysis of ß-diversity showed that female mice microbiota in both control mice and TCDD-treated mice became “more similar” over time. However, for male mice this increase in similarity was much lower, suggesting a reduced homeostatic capacity. At the 12 week timepoint the ß-diversity differed only for male mice. We next used flow cytometry and analyzed microbiota with our analysis tool FlowSoFine™. Contrary to the sequencing results, we detected significant changes in bacterial community profiles in TCDD-treated female mice. We hypothesized that this could be due to the formation of bacterial clusters or to different glycosylation patterns, both undetectable by sequencing. We therefore stained the fecal material with a panel of seven fluorescent lectins, which detect typical bacterial sugar structures. Significant changes were detected for concanavalin A, soybean agglutinin, and Dolichos biflorus agglutinin binding in the gut microbiota in female but not in male mice. We sorted the bacteria from those areas in the flow cytometry dot plots, which were significantly different between TCDD-treated and control female mice. 16S sequencing of these bacteria revealed a high frequency of Blautia and Acetobacter species in the TCDD-exposed samples. Glycoconjugates govern biofilm formation, infectious behavior of bacteria, and host immune responses, but little is known about diet or xenobiotic induced changes in glycosylation. We conclude that assessing glycosylation parameters in studies of dysbiosis is relevant, as well as stratifying for sex-specific differences.