HighlightsBacterial composition and community structure in the lower reproductive tract of female rhesus monkeys are significantly different by mid-pregnancy but significant shifts in their gut microbiome do not occur until later near term.By late pregnancy, four species ofLactobacillusandBifidobacterium adolescentiswere among the enriched commensals in the hindgut of female monkeys. The abundance of bacterial genes in the intestines, with potential effects on many metabolic pathways, differed in the last month of pregnancy.Several factors likely contributed to some of the differences in the microbial profiles of pregnant monkeys and women, including the consumption of a standardized diet throughout pregnancy and the comparatively low progesterone levels in rhesus monkeys, which were only 2–3% of the third trimester values in women.AbstractVaginal and rectal specimens were obtained from cycling, pregnant, and nursing rhesus monkeys to assess pregnancy-related changes in the commensal bacteria in their reproductive and intestinal tracts. Using 16S rRNA gene amplicon sequencing, significant differences were found only in the vagina at mid-gestation, not in the hindgut. To verify the apparent stability in gut bacterial composition at mid-gestation, the experiment was repeated with additional monkeys, and similar results were found with both 16S rRNA gene amplicon and metagenomic sequencing. A follow-up study investigated if bacterial changes in the hindgut might occur later in pregnancy. Gravid females were assessed closer to term and compared to nonpregnant females. By late pregnancy, significant differences in bacterial composition, including an increased abundance of 4 species ofLactobacillusandBifidobacterium adolescentis, were detected, but without a shift in the overall community structure. Progesterone levels were assessed as a possible hormone mediator of bacterial change. The relative abundance of only some taxa (e.g.,Bifidobacteriaceae) were specifically associated with progesterone. In summary, pregnancy changes the microbial profiles in monkeys, but the bacterial diversity in their lower reproductive tract is different from women, and the composition of their intestinal symbionts remains stable until late gestation when several Firmicutes become more prominent.