Vaginal colonization by Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a major risk factor for ascending infections, preterm birth, and neonatal sepsis. Current GBS prevention efforts include routine GBS perinatal screening and intrapartum antibiotic prophylaxis, which decrease the rate of early-onset neonatal sepsis, but have drawbacks that include impacting the infant's developing microbiome. Lactobacillus-dominant vaginal microbiomes provide protection against pathogens such as GBS, and using probiotics as an antibiotic-free approach to limit GBS colonization is of increasing interest. In this study, we investigated the ability of Lactobacillus crispatus-loaded electrospun fibers to deliver live L. crispatus cells in an in vitro vaginal epithelial cell model, modulate GBS infection establishment and persistence, and alter vaginal cell inflammatory signaling. Our data demonstrate that electrospun fibers deliver viable L. crispatus to the surface of vaginal epithelial cells and that L. crispatus modulates vaginal cell inflammatory signaling by decreasing inflammatory IL-8 release and increasing anti-inflammatory IL-1RA secretion during established GBS infection. Treatment of pre-established GBS infection with electrospun fibers with or without L. crispatus decreased GBS burden at 24 hours, suggesting L. crispatus-dependent and -independent anti-GBS activity, and L. crispatus elicited an anti-inflammatory response via IL-1RA release. Overall, the data highlight the potential of electrospun fibers as a feasible probiotic delivery platform with antibacterial activity against GBS and which provides commensal lactobacilli capable of modulating host-pathogen interactions and inflammatory signaling of the vaginal epithelium.

We analyzed our challenging experience with a randomized controlled trial of misoprostol for prevention of recurrent C. difficile. Despite careful prescreening and thoughtful protocol modifications to facilitate enrollment, we closed the study early after enrolling just 7 participants over 3 years. We share lessons learned, noting the importance of feasibility studies, inclusion of biomarker outcomes, and dissemination of such findings to inform future research design and implementation successes.

Histone modifications control numerous processes in eukaryotes, including inflammation. Some bacterial pathogens alter the activity or expression of host-derived factors, including sirtuins, to modify histones and induce responses that promote infection. In this study, we identified a deacetylase encoded by Campylobacter jejuni which has sirtuin activities and contributes to activation of human neutrophils by the pathogen. This sirtuin is secreted from the bacterium into neutrophils, where it associates with and deacetylates host histones to promote neutrophil activation and extracellular trap production. Using the murine model of campylobacteriosis, we found that a mutant of this bacterial sirtuin efficiently colonized the gastrointestinal tract but was unable to induce cytokine production, gastrointestinal inflammation, and tissue pathology. In conclusion, these results suggest that secreted bacterial sirtuins represent a previously unreported class of bacterial effector and that bacterial-mediated modification of host histones is responsible for the inflammation and pathology that occurs during campylobacteriosis.

Clostridioides difficile infection (CDI) is a significant cause of antibiotic-associated diarrhea. The Prevent Recurrence of Clostridioides difficile Infection with Misoprostol (PROCLAIM) study was designed to assess efficacy and safety of misoprostol, a prostaglandin analog, in prevention of recurrent CDI. The trial was halted for low enrollment. Misoprostol use during CDI appeared to be safe and well tolerated.

Background: During placental formation, cytotrophoblasts (CTBs) fuse into multinucleate, microvilli-coated syncytiotrophoblasts (STBs), which contact maternal blood, mediating nutrient, metabolite, and gas exchange between mother and fetus, and providing a barrier against fetal infection. Trophoblasts remodel the surrounding extracellular matrix through the secretion of matrix metalloproteinases (MMPs). Maternal obesity and diabetes mellitus can negatively impact fetal development and may impair trophoblast function. We sought to model the impact of metabolic stress on STB function by examining MMP and hormone secretion. Methods: The BeWo CTB cell line was syncytialized to STB-like cells with forskolin. Cell morphology was examined by electron microscopy and immunofluorescence; phenotype was further assessed by ELISA and RT-qPCR. STBs were exposed to a metabolic stress cocktail (MetaC: 30 mM glucose, 10 nM insulin, and 0.1 mM palmitic acid). Results: BeWo syncytialization was demonstrated by increased secretion of HCGβ and progesterone, elevated syncytin gene expression (ERVW-1 and ERVFRD-1), loss of tight junctions, and increased surface microvilli. MetaC suppressed HCGβ and progesterone and altered both MMP-9 and MMP-2. Conclusions: Metabolic stress modeling diabetes and obesity altered BeWo STB hormone and MMP production in vitro. These results compel further study into the potential impact of metabolic stress on trophoblast formation and function.

the a prospective study of Results : This study demonstrates that human breast milk lactoferrin has antimicrobial and anti-biofilm activity against GBS and inhibits its adherence to human gestational membranes largely dependent on iron-chelation. Conclusions : Together, these results indicate that human milk lactoferrin could be used as a prebiotic chemotherapeutic strategy to limit the impact of bacterial adherence and biofilm formation on GBS-associated disease outcomes.

Perinatal infection with Streptococcus agalactiae, or Group B Streptococcus (GBS), is associated with preterm birth, neonatal sepsis, and stillbirth. Here, we study the interactions of GBS with macrophages, essential sentinel immune cells that defend the gravid reproductive tract. Transcriptional analyses of GBS-macrophage co-cultures reveal enhanced expression of a gene encoding a putative metal resistance determinant, cadD. Deletion of cadD reduces GBS survival in macrophages, metal efflux, and resistance to metal toxicity. In a mouse model of ascending infection during pregnancy, the ΔcadD strain displays attenuated bacterial burden, inflammation, and cytokine production in gestational tissues. Furthermore, depletion of host macrophages alters cytokine expression and decreases GBS invasion in a cadD-dependent fashion. Our results indicate that GBS cadD plays an important role in metal detoxification, which promotes immune evasion and bacterial proliferation in the pregnant host.

Histone modifications alter numerous cornerstone processes in eukaryotes, including metabolism, physiology, and immunity. Numerous bacterial pathogens can alter expression of host-derived sirtuins to deacetylate histones in order to promote infection, yet, a bacterial-derived sirtuin has yet to be investigated to deacetylate host histones. Using Campylobacter jejuni, the leading cause of bacterial-derived gastroenteritis, we found a secreted sirtuin, SliP, which binds to and deacetylates neutrophil histones. We found neutrophil activation and extrusion of neutrophil extracellular traps was SliP dependent, whereby sliP mutants are unable to activate neutrophils or promote NETosis. Leveraging the mouse model of campylobacteriosis, we further demonstrate the sliP mutant can efficiently infect IL-10-/- mice, but induction of proinflammatory cytokine production and gastrointestinal pathology is SliP-dependent. In conclusion, we investigate a unique bacterial effector which targets host histones and is responsible for the inflammatory response and tissue pathology observed during campylobacteriosis. Highlights C. jejuni encodes a secreted effector, SliP, which functions as a canonical sirtuin SliP binds to and deacetylates neutrophil histone H3 during bacterial infection C. jejuni-induced neutrophil activation and NETosis are SliP-dependent Inflammation and tissue pathology during C. jejuni infection is SliP-dependent

Group B Streptococcus (GBS) is a gram-positive bacterium that can cause invasive infections in immunocompromised, elderly, pregnant, or neonatal patients. The invertebrate model, Galleria mellonella, has emerged as an effective tool to study GBS-host interactions; specifically, those conserved within the innate arm of the immune system. We sought to determine the role of metal homeostasis functions in GBS infections of G. mellonella larvae and to validate this model as a tool to study GBS-host interactions. Our results indicate that wild-type GBS infects G. mellonella in a dose-dependent manner, replicates in the invertebrate host, induces larval melanization and larval killing. These results were significantly abrogated in cohorts of larvae infected with the isogenic cadD deletion mutant. Additionally, complementation restored GBSdependent infection, bacterial burden, larval melanization, and killing to wild-type levels. Together, these results indicate that the G. mellonella model is a useful tool for studying GBS pathogenesis.

Clinical Course in Reproductive Immunology will precede the annual meeting. All of one-day virtual Clinical Course in Reproductive Immunology which will precede the annual meeting. All of the will be and available to our nationally and internationally. the to our and internationally. The American Reproductive Immunology will be presented to a senior investigator who made outstanding clinical or basic research contributions in the area of reproductive immunology. The Dr. John Gusdon Memorial New Investigator Award will be presented to a new investigator with trainee status (graduate student, postdoctoral scientist, or resident) who has made a significant contribution by presenting an outstanding research paper during the annual meeting. This award is given annual in memory of Dr. John Gusdon, a founding member of ASRI and advocate of student participation in ASRI meetings. The J. Herr Award is given annually to a member of ASRI, International Society for Immunology of Reproduction, or European Society for Reproductive Immunology, who has made outstanding achievements in basic or applied research in reproductive immunology, particularly for investigators involved in technology transfer. The Carolyn B. Coulam Memorial Award is given annually to an outstanding female physician-scientist in the field of reproductive immunology who has demonstrated commitments, by publication and/or practice, to scholarly advances in the field of reproductive immunology. The ASRI Distinguished Service Award is given periodically and not more than annually, to a member of the ASRI who has provided distinguished service to advance the goals and mission of the society. The Poster Competition Awards will be presented to the top posters based on the final scores received during the poster session on Monday.

Enteroinvasive Escherichia coli (EIEC), a close relative of Shigella , is implicated in dysenteric diarrhea. EIEC pathogenicity involves cell invasion mediated by effector proteins delivered by a type III secretion system (TTSS) that disrupt the cell cytoskeleton.

65 blood pressurewasmeasured, fetal weight was recorded, and placenta was collected and subjected to immunostaining and RNA sequencing. The kidney was fixed and processed for HE staining for glomerular endotheliosis. Results: A class of disaccharides inhibited the hypoxia-induced accumulation of aggregates and restored impaired autophagy-lysosomal machinery as characterized by evaluation of p62, TFEB, LAMP1/2, and cathepsinD. Disaccharide administration significantly restored normal pregnancy features in PE mice as characterized by normalization of hypertension, proteinuria, growth restriction, and kidney injury. RNAseq identified some novel genes and pathways that are related to preeclampsia and normalized by disaccharides. Conclusions: Certain disaccharides can inhibit protein aggregation in vitro and rescue PE-like features in vivo through normalizing autophagy-lysosomal machinery. These findings reinforce our proposed concept that PE is a disease of protein misfolding and aggregation and that these features can be targeted for therapeutic intervention. SESSION 15: GROUP B STREPTOCOCCUS COLONIZATION AND DISSEMINATION OF THE FEMALE REPRODUCTIVE TRACT S15.1 Modeling GBS perinatal transmission: What have we learned? TaraMRandis University of South Florida, Tampa, FL, USA Despite widespread adoption of intrapartum antibiotic administration, GBS remains a leading cause of neonatal sepsis. Understanding the microbial and host factors that drive perinatal transmission and the newborn host response to GBS exposure are critical for the development of novel prevention strategies. This talk will include a discussion of experimental animal models of GBS transmission in the antepartum, intrapartum and postpartum periods. Pathogen/host interactions that modulate the risk of GBS transmission and progression to invasive diseasewill be reviewed. Existing knowledge gaps andpotential strategies to reduce vertical GBS transmission will be highlighted. S15.2 Characterization of a novel GBS adhesin Laura Cook, Lamar S Thomas Binghamton University, Vestal, NY, USA Problem: Streptococcus agalactiae (Group B Strep, GBS) infections in neonates are often fatal and strongly associated with maternal GBS vaginal colonization. Intrapartumantibiotic use inmaternalwomen are effective at preventing early onset neonatal GBS disease but there are pitfalls involved in their use including adverse effects on the neonatal microbiome. As such, alternative approaches to prevent vaginal colonization are needed and a more complete understanding of mucosal colonization factors is necessary to develop these strategies. Method of Study: We created a genetic knockout of bvaP and tested themutant and complemented strains in a variety of phenotypic assays including examination of bacterial cell morphology using electron and fluorescent microscopy, ability of cells to create biofilms and adherence to cultured epithelial cells. We also examined the role of BvaP in colonization of the vaginal tract using a murine vaginal carriage model. Results: BvaP was previously identified as the most highly upregulated gene in theGBSA909 transcriptomewhen comparing vaginal colonization to growth in liquid culture.We found that expression of BvaP affects GBS adherence to extracellular matrix components and human vaginal epithelial cells and aΔbvaPmutant was significantly decreased in its ability to colonize the murine vaginal tract. Cellular morphological alterations such as changes in cell shape, chain length, and clumping were also observed in a knockout mutant strain. Conclusions: BvaP is involved in mucosal colonization by GBS and, given its high expression in vivo, high degree of conservation among GBS strains, and role in vaginal colonization, BvaP may be an eligible target for GBS vaccination and/or drug therapy. S15.3 Metal homeostasis and Group B Streptococcus Jennifer A. Gaddy1, Michelle Korir2, Ryan Doster1, Jacky Lu1, Jamisha Francis1, MiriamGuevara1, RebeccaMoore3, Sabrina Spicer3, Steven Townsend3, Kristen Noble1, Alison Eastman1, DavidM. Aronoff4, ShannonManning5 1Vanderbilt University Medical Center, Nashville, TN, USA; 2Aurora University, Aurora, IL, USA; 3Vanderbilt University, Nashville, TN, USA; 4Indiana University, Indianapolis, IN, USA; 5Michigan State University, East Lansing, MI, USA Problem: Perinatal infectionwith Streptococcus agalactiae, orGroupB Streptococcus (GBS), is associated with preterm birth, neonatal sepsis, and stillbirth. Method of Study: Here, we study the interactions of GBS with macrophages, essential sentinel immune cells that defend the gravid reproductive tract. Results: Transcriptional analyses of GBS-macrophage co-cultures reveal enhanced expression of a gene encoding a putative metal resistance determinant, cadD. Deletion of cadD reduces GBS survival in macrophages, metal efflux, and resistance to metal toxicity. In a mouse model of ascending infection during pregnancy, the ΔcadD strain displays attenuated bacterial burden, inflammation, and cytokine production in gestational tissues. Furthermore, depletion of host macrophages alters cytokine expression and decreases GBS invasion in a cadD-dependent fashion. Conclusions: Our results indicate that GBS cadD plays an important role in metal detoxification, which promotes immune evasion and bacterial proliferation in the pregnant host.

Streptococcus agalactiae, also known as group B Streptococcus (GBS), is a Gram-positive encapsulated bacterium that colonizes the gastrointestinal tract of 30 to 50% of humans. GBS causes invasive infection during pregnancy that can lead to chorioamnionitis, funisitis, preterm prelabor rupture of membranes (PPROM), preterm birth, neonatal sepsis, and maternal and fetal demise. Upon infecting the host, GBS encounters sentinel innate immune cells, such as macrophages, within reproductive tissues. Once phagocytosed by macrophages, GBS upregulates the expression of the gene npx, which encodes an NADH peroxidase. GBS mutants with an npx deletion (Delta npx) are exquisitely sensitive to reactive oxygen stress. Furthermore, we have shown that npx is required for GBS survival in both THP-1 and placental macrophages. In an in vivo murine model of ascending GBS vaginal infection during pregnancy, npx is required for invading reproductive tissues and is critical for inducing disease progression, including PPROM and preterm birth. Reproductive tissue cytokine production was also significantly diminished in Delta npx mutant-infected animals compared to that in animals infected with wild-type (WT) GBS. Complementation in trans reversed this phenotype, indicating that npx is critical for GBS survival and the initiation of proinflammatory signaling in the gravid host.IMPORTANCE This study sheds new light on the way that group B Streptococcus (GBS) defends itself against oxidative stress in the infected host. The enzyme encoded by the GBS gene npx is an NADH peroxidase that, our study reveals, provides defense against macrophage-derived reactive oxygen stress and facilitates infections of the uterus during pregnancy. This enzyme could represent a tractable target for future treatment strategies against invasive GBS infections. This study sheds new light on the way that group B Streptococcus (GBS) defends itself against oxidative stress in the infected host. The enzyme encoded by the GBS gene npx is an NADH peroxidase that, our study reveals, provides defense against macrophage-derived reactive oxygen stress and facilitates infections of the uterus during pregnancy. This enzyme could represent a tractable target for future treatment strategies against invasive GBS infections.

Clinical Course in Reproductive Immunology will precede the annual meeting. All of one-day virtual Clinical Course in Reproductive Immunology which will precede the annual meeting. All of the will be and available to our nationally and internationally. the to our and internationally. The American Reproductive Immunology will be presented to a senior investigator who made outstanding clinical or basic research contributions in the area of reproductive immunology. The Dr. John Gusdon Memorial New Investigator Award will be presented to a new investigator with trainee status (graduate student, postdoctoral scientist, or resident) who has made a significant contribution by presenting an outstanding research paper during the annual meeting. This award is given annual in memory of Dr. John Gusdon, a founding member of ASRI and advocate of student participation in ASRI meetings. The J. Herr Award is given annually to a member of ASRI, International Society for Immunology of Reproduction, or European Society for Reproductive Immunology, who has made outstanding achievements in basic or applied research in reproductive immunology, particularly for investigators involved in technology transfer. The Carolyn B. Coulam Memorial Award is given annually to an outstanding female physician-scientist in the field of reproductive immunology who has demonstrated commitments, by publication and/or practice, to scholarly advances in the field of reproductive immunology. The ASRI Distinguished Service Award is given periodically and not more than annually, to a member of the ASRI who has provided distinguished service to advance the goals and mission of the society. The Poster Competition Awards will be presented to the top posters based on the final scores received during the poster session on Monday.

Group B Streptococcus (GBS) is an encapsulated Gram‐positive human pathogen that causes invasive infections in pregnant hosts and neonates, as well as immunocompromised individuals. Colonization of the human host requires the ability to adhere to mucosal surfaces and circumnavigate the nutritional challenges and antimicrobial defenses associated with the innate immune response. Biofilm formation is a critical process to facilitate GBS survival and establishment of a replicative niche in the vertebrate host. Previous work has shown that the host responds to GBS infection by producing the innate antimicrobial glycoprotein lactoferrin, which has been implicated in repressing bacterial growth and biofilm formation. Additionally, lactoferrin is highly abundant in human breast milk and could serve a protective role against invasive microbial pathogens. This study demonstrates that human breast milk lactoferrin has antimicrobial and anti‐biofilm activity against GBS and inhibits its adherence to human gestational membranes. Together, these results indicate that human milk lactoferrin could be used as a prebiotic chemotherapeutic strategy to limit the impact of bacterial adherence and biofilm formation on GBS‐associated disease outcomes.

The cover feature picture is a composite of a high-resolution scanning electron microscopy (SEM) of Streptococcus agalactiae biofilms at high (10 000×) magnification pseudocolored purple with ribbon diagram structures of the lactoferrin molecule between cells. S. agalactiae, also known as Group B Streptococcus (GBS) is a bacterial pathogen that causes invasive infections during pregnancy that can lead to preterm birth and neonatal sepsis. The work demonstrates that the glycoprotein lactoferrin, which is a dominant component of human breast milk, can inhibit GBS growth, viability, biofilm formation, and adherence to reproductive tissues, such as gestational membranes. More information can be found in the Full Paper by S. D. Townsend, J. A. Gaddy et al.

Chronic infection with Helicobacter pylori increases risk of gastric diseases including gastric cancer. Despite development of a robust immune response, H. pylori persists in the gastric niche. Progression of gastric inflammation to serious disease outcomes is associated with infection with H. pylori strains which encode the cag Type IV Secretion System (cag T4SS). The cag T4SS is responsible for translocating the oncogenic protein CagA into host cells and inducing pro-inflammatory and carcinogenic signaling cascades. Our previous work demonstrated that nutrient iron modulates the activity of the T4SS and biogenesis of T4SS pili. In response to H. pylori infection, the host produces a variety of antimicrobial molecules, including the iron-binding glycoprotein, lactoferrin. Our work shows that apo-lactoferrin exerts antimicrobial activity against H. pylori under iron-limited conditions, while holo-lactoferrin enhances bacterial growth. Culturing H. pylori in the presence of holo-lactoferrin prior to co-culture with gastric epithelial cells, results in repression of the cag T4SS activity. Concomitantly, a decrease in biogenesis of cag T4SS pili at the host-pathogen interface was observed under these culture conditions by high-resolution electron microscopy analyses. Taken together, these results indicate that acquisition of alternate sources of nutrient iron plays a role in regulating the pro-inflammatory activity of a bacterial secretion system and present novel therapeutic targets for the treatment of H. pylori-related disease.