PRJNA872181
Amplification of Microbial DNA from Bacterial Outer Membrane Vesicles from Human Placenta
The placenta is essential for fetal growth and survival and maintenance of a successful pregnancy. The sterility of the placenta has been challenged recently; however, the presence of a placental microbiome has been controversial. We tested the hypothesis that the outer membrane vesicles (OMVs) from Gram-negative bacteria as an alternate source of microbial DNA, regardless of the existence of a microbial community in the placenta. Placentae from the normal term, not in labor Cesareans deliveries were used for this study and placental specimens were samples randomly from the fetal side. For the first time, we report developing a protocol to isolate OMVs from human tissue (i.e., placenta), and its characterization. The median size of OMVs was 130-140 nm, and the mean concentration was 1.8-5.5x1010 OMVs/g of the wet placenta. OMVs are circular and contain LPS and ompA. Western blots further confirmed ompA but not human exosome markers ALIX confirming the purity of preparations. Taxonomic abundance profiles showed OMV sequence reads above the levels of the negative controls (all reagent controls) whereas the sequence reads in the same placenta were substantially low indicating nothing beyond contamination (low biomass). a-diversity showed the number of detected genera was significantly higher in the OMVs than placenta suggesting OMVs as a likely source of microbial DNA. b-diversity further showed significant overlap in the microbiome between OMV and the placenta confirming that placental OMVs are likely a source of microbial DNA in the placenta. Uptake studies localized OMVs in both maternal (decidual) and placental cells (cytotrophoblast) confirming their ability to enter these cells. Lastly, OMVs significantly increased inflammatory cytokine productions in THP-1 macrophages in a dose-dependent manner but not in the placental or decidual cells. We conclude that the placental OMVs are normal constituents during pregnancy and likely reach the placenta through hematogenous spread from the microbial colonies that are normal in the mother. Their normal presence may result in a low-grade localized inflammation to prime an antigen response in the placenta; however, insufficient to cause a fetal inflammatory response and adverse pregnancy events. Importantly, OMVs can confound placental microbiome studies, but their low biomass in the placenta is unlikely to have any immunologic impact.