Non-human primate models for understanding the impact of the microbiome on pregnancy and the female reproductive tract†.

The microbiome has been shown, or implicated to be involved, in multiple facets of human health and disease, including not only gastrointestinal health but also metabolism, immunity, and neurology. Although the predominant focus of microbiome research has been on the gut, other microbial communities such as the vaginal or cervical microbiome are likely involved in physiological homeostasis. Emerging studies also aim to understand the role of different microbial niches, such as the endometrial or placental microbial communities, on the physiology and pathophysiology of reproduction, including their impact on reproductive success and the etiology of adverse pregnancy outcomes (APOs). The study of the microbiome during pregnancy, specifically how changes in maternal microbial communities can lead to dysfunction and disease, can advance the understanding of reproductive health and the etiology of APOs. In this review, we will discuss the current state of non-human primate (NHP) reproductive microbiome research, highlight the progress with NHP models of reproduction, and the diagnostic potential of microbial alterations in a clinical setting to promote pregnancy health. NHP reproductive biology studies have the potential to expand the knowledge and understanding of female reproductive tract microbial communities and host-microbe or microbe-microbe interactions associated with reproductive health through sequencing and analysis. Furthermore, in this review, we aim to demonstrate that macaques are uniquely suited as high-fidelity models of human female reproductive pathology.

Listeria monocytogenes infection in pregnant macaques alters the maternal gut microbiome†.

OBJECTIVES: The bacterium Listeria monocytogenes (Lm) is associated with adverse pregnancy outcomes. Infection occurs through consumption of contaminated food that is disseminated to the maternal-fetal interface. The influence on the gastrointestinal microbiome during Lm infection remains unexplored in pregnancy. The objective of this study was to determine the impact of listeriosis on the gut microbiota of pregnant macaques. METHODS: A non-human primate model of listeriosis in pregnancy has been previously described. Both pregnant and non-pregnant cynomolgus macaques were inoculated with Lm and bacteremia and fecal shedding were monitored for 14 days. Non-pregnant animal tissues were collected at necropsy to determine bacterial burden, and fecal samples from both pregnant and non-pregnant animals were evaluated by 16S rRNA next-generation sequencing. RESULTS: Unlike pregnant macaques, non-pregnant macaques did not exhibit bacteremia, fecal shedding, or tissue colonization by Lm. Dispersion of Lm during pregnancy was associated with a significant decrease in alpha diversity of the host gut microbiome, compared to non-pregnant counterparts. The combined effects of pregnancy and listeriosis were associated with a significant loss in microbial richness, although there were increases in some genera and decreases in others. CONCLUSIONS: Although pregnancy alone is not associated with gut microbiome disruption, we observed dysbiosis with listeriosis during pregnancy. The macaque model may provide an understanding of the roles that pregnancy and the gut microbiota play in the ability of Lm to establish intestinal infection and disseminate throughout the host, thereby contributing to adverse pregnancy outcomes and risk to the developing fetus.

Listeria monocytogenes infection in intestinal epithelial Caco-2 cells with exposure to progesterone and estradiol-17beta in a gestational infection model.

Listeria monocytogenes (Lm) is a food-borne pathogen associated with serious pregnancy complications, including miscarriage, stillbirth, preterm birth, neonatal sepsis, and meningitis. Although Lm infection within the gastrointestinal (GI) tract is well studied, little is known about the influence sex hormones may have on listeriosis. Estradiol (E2) and progesterone (P4) not only have receptors within the GI tract but are significantly increased during pregnancy. The presence of these hormones may play a role in susceptibility to listeriosis during pregnancy. Caco-2 cell monolayers were grown on trans-well inserts in the presence of E2, P4, both E2 and P4, or no hormones (control). Cells were inoculated with Lm for 1 hour, before rinsing with gentamycin and transfer to fresh media. Trans-epithelial resistance was recorded hourly, and bacterial burden of the apical media, intracellular lysates, and basal media were assessed at 6 hours post inoculation. There were no significant differences in bacterial replication when directly exposed to sex steroids, and Caco-2 cell epithelial barrier function was not impacted during culture with Lm. Addition of P4 significantly reduced intracellular bacterial burden compared to E2 only and no hormone controls. Interestingly, E2 only treatment was associated with significantly increased Lm within the basal compartment, compared to reduction in the intracellular and apical layers. These data indicate that increased circulating sex hormones alone do not significantly impact intestinal epithelial barrier integrity during listeriosis, but that addition of P4 and E2, alone or in combination, was associated with reduced epithelial cell bacterial burden and apical release of Lm.

Acute Exposure to the Food-Borne Pathogen Listeria monocytogenes Does Not Induce α-Synuclein Pathology in the Colonic ENS of Nonhuman Primates.

INTRODUCTION: Gastrointestinal (GI) inflammation elicited by environmental factors is proposed to trigger Parkinson's disease (PD) by stimulating accumulation of pathological α-synuclein (α-syn) in the enteric nervous system (ENS), which then propagates to the central nervous system via the vagus nerve. The goal of this study was to model, in nonhuman primates, an acute exposure to a common food-borne pathogen in order to assess whether the related acute GI inflammation could initiate persistent α-syn pathology in the ENS, ultimately leading to PD. METHODS: Adult female cynomolgus macaques were inoculated by oral gavage with 1×108 colony-forming units (CFUs) Listeria monocytogenes (LM, n=10) or vehicle (mock, n=3) and euthanized 2 weeks later. Evaluations included clinical monitoring, blood and fecal shedding of LM, and postmortem pathological analysis of colonic and cecal tissues. RESULTS: LM inoculation of healthy adult cynomolgus macaques induced minimal to mild clinical signs of infection; LM shedding in feces was not seen in any of the animals nor was bacteremia detected. Colitis varied from none to moderate in LM-treated subjects and none to minimal in mock-treated subjects. Expression of inflammatory markers (HLA-DR, CD3, CD20), oxidative stress (8-OHDG), α-syn, and phosphorylated-α-syn in the enteric ganglia was not significantly different between treatment groups. DISCUSSION: Our results demonstrate that cynomolgus macaques orally inoculated with LM present with a clinical response that resembles human LM exposure. They also suggest that acute exposure to food-borne pathogens is not sufficient to induce significant and persistent α-syn changes in healthy adult female subjects. Based on the results of this limited experimental setting, we propose that, if LM has a role in PD pathology, other underlying factors or conditions, such as male sex, inflammatory bowel disease, exposure to toxins, dysbiosis, and/or aging, are needed to be present.