Membrane inflammasome activation by choriodecidual Ureaplasma parvum infection without intra-amniotic infection in a Non-Human Primate model†.

Intrauterine infection is a significant cause of neonatal morbidity and mortality. Ureaplasma parvum is a microorganism commonly isolated from cases of preterm birth and preterm premature rupture of membranes (pPROM). However, the mechanisms of early stage ascending reproductive tract infection remain poorly understood. To examine inflammation in fetal (chorioamnionic) membranes we utilized a non-human primate (NHP) model of choriodecidual U. parvum infection. Eight chronically catheterized pregnant rhesus macaques underwent maternal-fetal catheterization surgery at ~105-112 days gestation and choriodecidual inoculation with U. parvum (105 CFU/mL, n =4) or sterile media (controls; n = 4) starting at 115-119 days, repeated at 5-day intervals until C-section at 136-140 days (term=167 days). The average inoculation to delivery interval was 21 days, and Ureaplasma infection of the amniotic fluid (AF) was undetectable in all animals. Choriodecidual Ureaplasma infection resulted in increased fetal membrane expression of MMP-9 and PTGS2, but did not result in preterm labor or increased concentrations of AF pro-inflammatory cytokines. However, membrane expression of inflammasome sensors, NLRP3, NLRC4, AIM2, and NOD2, and adaptor ASC (PYCARD) gene expression were significantly increased. Gene expression of IL-1ß, IL-18, IL-18R1  , CASPASE-1, and pro-CASPASE-1 protein increased with Ureaplasma infection. Downstream inflammatory genes MYD88 and NFκB (Nuclear factor kappa-light-chain-enhancer of activated B cells) were also significantly upregulated. These results demonstrate that choriodecidual Ureaplasma infection, can cause activation of inflammasome complexes and pathways associated with pPROM and preterm labor prior to microbes being detectable in the AF.

Polymicrobial stimulation of human fetal membranes induce neutrophil activation and neutrophil extracellular trap release.

Preterm birth is a major contributor to neonatal mortality and morbidity. While the causes of preterm birth remain incompletely understood, infection is a major risk factor, and chorioamnionitis is commonly observed. Chorioamnionitis is characterized by inflammation and neutrophil infiltration of the fetal membranes (FM). We recently reported that human FMs which had been exposed to low levels of bacterial lipopolysaccharide (LPS) recruit neutrophils and activate them, increasing their secretion of pro-inflammatory cytokines, degranulation of myeloperoxidase (MPO), and release of neutrophil extracellular traps (NETs). Herein, we demonstrate that conditioned media (CM) from viral dsRNA (Poly(I:C))-stimulated FMs also increased neutrophil migration, and induced the secretion of inflammatory IL-8 and the release of NETs. Furthermore, CM from FMs stimulated by a combination of bacterial LPS and Poly(I:C) augmented neutrophil NET release, compared to CM from FMs stimulated with either Poly(I:C) or LPS alone. NETs induced by FMs exposed to Poly(I:C), with or without LPS, were released and degraded quicker than those induced by resting or LPS-stimulated FM-CM. These findings indicate that FMs exposed to viral dsRNA promote neutrophil recruitment, activation and NET formation, similar to FMs exposed to bacterial LPS alone. However, in response to FM polymicrobial stimulation the levels and kinetics of NET release are augmented. This work builds upon our understanding of how infections at the maternal-fetal interface may affect neutrophil function.

Evidence for contamination as the origin for bacteria found in human placenta rather than a microbiota.

Until recently the in utero environment of pregnant women was considered sterile. Recent high-sensitivity molecular techniques and high-throughput sequencing lead to some evidence for a low-biomass microbiome associated with the healthy placenta. Other studies failed to reveal evidence for a consistent presence of bacteria using either culture or molecular based techniques. Comparing conflicting "placental microbiome" studies is complicated by the use of varied and inconsistent protocols. Given this situation, we undertook an evaluation of the in utero environment sterility using several controlled methods, in the same study, to evaluate the presence or absence of bacteria and to explain contradictions present in the literature. Healthy pregnant women (n = 38) were recruited in three maternity wards. Placenta were collected after cesarean section with or without Alexis® and vaginal delivery births. For this study we sampled fetal membranes, umbilical cord and chorionic villi. Bacterial presence was analyzed using bacterial culture and qPCR on 34 fetal membranes, umbilical cord and chorionic villi samples. Shotgun metagenomics was performed on seven chorionic villi samples. We showed that the isolation of meaningful quantities of viable bacteria or bacterial DNA was possible only outside the placenta (fetal membranes and umbilical cords) highlighting the importance of sampling methods in studying the in utero environment. Bacterial communities described by metagenomics analysis were similar in chorionic villi samples and in negative controls and were dependent on the database chosen for the analysis. We conclude that the placenta does not harbor a specific, consistent and functional microbiota.

Fetal membrane bacterial load is increased in histologically confirmed inflammatory chorioamnionitis: A retrospective cohort study.

INTRODUCTION: It is widely debated whether fetal membranes possess a genuine microbiome, and if bacterial presence and load is linked to inflammation. Chorioamnionitis is an inflammation of the fetal membranes. This research focussed on inflammatory diagnosed histological chorioamnionitis (HCA) and aimed to determine whether the bacterial load in fetal membranes correlates to inflammatory response, including histological staging and inflammatory markers in HCA. METHODS: Fetal membrane samples were collected from patients with preterm spontaneous labour and histologically phenotyped chorioamnionitis (HCA; n = 12), or preterm (n = 6) and term labour without HCA (n = 6). The bacterial profile of fetal membranes was analysed by sequencing the V4 region of the 16S rRNA gene. Bacterial load was determined using qPCR copy number/mg of tissue. The association between bacterial load and bacterial profile composition was assessed using correlation analysis. RESULTS: Bacterial load was significantly greater within HCA amnion (p = 0.002) and chorion (p = 0.042), compared to preterm birth without HCA. Increased bacterial load was positively correlated with increased histological staging (p = 0.001) and the expression of five inflammatory markers; IL8, TLR1, TLR2, LY96 and IRAK2 (p=<0.050). Bacterial profiles were significantly different between membranes with and without HCA in amnion (p = 0.012) and chorion (p = 0.001), but no differences between specific genera were detected. DISCUSSION: Inflammatory HCA is associated with infection and increased bacterial load in a dose response relationship. Bacterial load is positively correlated with HCA severity and the TLR signalling pathway. Further research should investigate the bacterial load threshold required to generate an inflammatory response in HCA.

Raman microspectroscopy differentiates perinatal pathogens on ex vivo infected human fetal membrane tissues.

Streptococcus agalactiae, also known as Group B Streptococcus (GBS), is a major cause of chorioamnionitis and neonatal sepsis. This study evaluates Raman spectroscopy (RS) to identify spectral characteristics of infection and differentiate GBS from Escherichia coli and Staphylococcus aureus during ex vivo infection of human fetal membrane tissues. Unique spectral features were identified from colonies grown on agar and infected fetal membrane tissues. Multinomial logistic regression analysis accurately identified GBS infected tissues with 100.0% sensitivity and 88.9% specificity. Together, these findings support further investigation into the use of RS as an emerging microbiologic diagnostic tool and intrapartum screening test for GBS carriage.

Trichomonas vaginalis Transports Virulent Mycoplasma hominis and Transmits the Infection to Human Cells after Metronidazole Treatment: A Potential Role in Bacterial Invasion of Fetal Membranes and Amniotic Fluid.

Mycoplasma hominis is considered an opportunistic pathogen able to colonize the lower urogenital tract; in females the infection is associated with severe pregnancy and postpartum complications, including abortion, endometritis, preterm delivery, and low birth weight. Molecular mechanisms of pathogenicity and virulence effectors remain poorly characterized. A number of studies in the last decade have demonstrated that M. hominis can establish an endosymbiotic relationship with Trichomonas vaginalis, a urogenital parasitic protozoon, also associated with adverse pregnancy outcomes. Recently, two bacterial genes (alr and goiB) associated with amniotic cavity invasion and a single gene (goiC) associated with intra-amniotic infections and high risk of preterm delivery have been identified in M. hominis isolated from a group of pregnant patients. In this work we demonstrate that a high number of M. hominis intracellularly associated with T. vaginalis have goiC gene, in association with alr and goiB. In addition, we demonstrate that metronidazole treatment of M. hominis-infected T. vaginalis allows delivering viable intracellular goiC positive M. hominis from antibiotic-killed protozoa and that free M. hominis can infect human cell cultures. Results suggest that molecular diagnostic strategies to identify both pathogens and their virulence genes should be adopted to prevent severe complications during pregnancy.

Periodontal pathogens in the placenta and membranes in term and preterm birth.

INTRODUCTION: Preterm birth is a common cause of adverse neonatal and childhood outcomes. It is commonly associated with infection of the maternal-fetal interface. The relationship between periodontitis and preterm labour is controversial. METHODS: Control placental tissues from uncomplicated term births were compared with those from spontaneous preterm births for incidence of common periodontal bacteria. A chi-square analysis was used to compare the populations, with significance determined at p=<0.05. RESULTS: The study group comprised 29 control women who had an uncomplicated term birth, 25 delivered by caesarean section and 4 vaginal deliveries, and 36 women with a spontaneous preterm labour and subsequent delivery at less than 34 weeks gestation. There were significant (p=<0.05) differences between the preterm and term groups maternal age with 28.7 compared to 32.0 years old respectively. There was no significant (p=>0.05) differences between the groups fetal risk factors or co-morbidities, except the preterm group had a significantly higher (p=<0.05) rate of premature rupture of membrane (PROM). There were significantly (p=<0.01) more Fusobacterium spp. in the placentas from term births than preterm births. DISCUSSION: This study found that the common periodontal pathogen, Fusobacterium spp., is not detected more in placentas from preterm birth and may potentially be lower, possibly resulting from bacterial ecological factors in term placentas.

Inhibition of gingipains prevents Porphyromonas gingivalis-induced preterm birth and fetal death in pregnant mice.

Accumulating epidemiological evidence indicates that infection with Porphyromonas gingivalis which is a major periodontal pathogen, causes preterm birth and low birth weight. However, virulence factors of P. gingivalis responsible for preterm birth/low birth weight remain to be elucidated. In this study, using P. gingivalis-infected pregnant mice as an in vivo model, we investigated whether gingipains-cysteine proteinases produced by P. gingivalis-affect preterm birth and low birth weight. We found that intravenous infection of pregnant mice with P. gingivalis induced higher accumulation of the bacterium in the placenta than that in other organs. Compared to infection with P. gingivalis wild-type, infection with a gingipain-deficient P. gingivalis mutant KDP136 led to significant reduction in preterm birth and pregnancy loss. Although repetitive low-level infections of P. gingivalis failed to induce preterm birth and fetal death, it induced suppressive effects on IFN-γ production. Therapeutically, treatment with ginginpain inhibitors prevented fetal death and preterm birth caused by P. gingivalis infection and resulted in recovery of IFN-γ suppression caused by repetitive chronic P. gingivalis infection. These results indicate that gingipains are major virulence factors of P. gingivalis responsible for preterm birth/low birth, and gingipain inhibitors may be useful not only as a therapeutic agent for periodontal diseases, but also as a preventive medicine for preterm birth/low birth weight.

Group B streptococcus activates transcriptomic pathways related to premature birth in human extraplacental membranes in vitro.

Streptococcus agalactiae (group B streptococcus [GBS]) infection in pregnant women is the leading cause of infectious neonatal morbidity and mortality in the United States. Although inflammation during infection has been associated with preterm birth, the contribution of GBS to preterm birth is less certain. Moreover, the early mechanisms by which GBS interacts with the gestational tissue to affect adverse pregnancy outcomes are poorly understood. We hypothesized that short-term GBS inoculation activates pathways related to inflammation and premature birth in human extraplacental membranes. We tested this hypothesis using GBS-inoculated human extraplacental membranes in vitro. In agreement with our hypothesis, a microarray-based transcriptomics analysis of gene expression changes in GBS-inoculated membranes revealed that GBS activated pathways related to inflammation and preterm birth with significant gene expression changes occurring as early as 4 h postinoculation. In addition, pathways related to DNA replication and repair were downregulated with GBS treatment. Conclusions based on our transcriptomics data were further supported by responses of prostaglandin E2 (PGE2), and matrix metalloproteinases 1 (MMP1) and 3 (MMP3), all of which are known to be involved in parturition and premature rupture of membranes. These results support our initial hypothesis and provide new information on molecular targets of GBS infection in human extraplacental membranes.

The Role of Progesterone and a Novel Progesterone Receptor, Progesterone Receptor Membrane Component 1, in the Inflammatory Response of Fetal Membranes to Ureaplasma parvum Infection.

Ureaplasma parvum (U. parvum) is gaining recognition as an important pathogen for chorioamnionitis and preterm premature rupture of membranes. We aimed to investigate the roles of progesterone (P4) and a novel progesterone receptor, progesterone receptor membrane component 1 (PGRMC1), in the response of fetal membranes to U. parvum. Fetal membrane cells (amnion, chorion and decidua) were isolated and confirmed to be free of Mycoplasmataceae. Cells were treated with U. parvum (5x106 CFU), and adherence was quantified by qPCR. Amnion and chorion cells were transfected with scrambled siRNA or validated PGRMC1 siRNA for 72h. Cells were then treated with U. parvum for 4h with or without pretreatment with P4 (10-7 M) or ethanol for 1h. Interleukin-8 (IL-8), matrix metalloproteinase 9 (MMP9) and cyclooxygenase (COX-2) mRNA expression were quantified by qRT-PCR. Culture medium was harvested and analyzed for IL-8 and prostaglandin (PGE2) secretion by ELISA and MMP9 activity by zymography. U. parvum had a mean adherence of 15.0±0.6%, 16.9± 3.7% and 4.7±0.3% in cultured amnion, chorion and decidua cells, respectively. Exposure to U. parvum elicited significant inflammatory responses including induction of IL-8, COX-2, PGE2 and MMP9. A possible role of PGRMC1 was identified in the inhibition of U. parvum-stimulated COX-2 and MMP9 mRNA expression in chorion cells and MMP9 activity in amnion cells. On the other hand, it might enhance the U. parvum-stimulated IL-8 protein secretion in amnion cells. P4, mediated through PGRMC1, significantly inhibited U. Parvum-induced MMP9 mRNA and COX-2 mRNA expression in chorion cells. P4 appeared to attenuate U. parvum induced IL-8 mRNA expression in chorion cells, but this P4 effect might not mediated through PGRMC1. In summary, U. parvum preferentially adheres to and induces inflammatory responses in chorion and amnion cells. P4 and PGRMC1 appear to differentially modulate the inflammatory responses induced by U. parvum among amnion and chorion cells.

Combinations and loads of bacteria affect the cytokine production by fetal membranes: An in vitro study.

PROBLEM: The polybacterial invasion and inflammation of the amniotic cavity is a common scenario in PTB, and then, we analyzed the cytokine production by human fetal membranes to better understand the host response to polybacterial infections. METHOD OF STUDY: Fetal membranes were treated by heat-inactivated genital mycoplasmas and Gardnerella vaginalis at 103 or 106 colony/color-forming units/mL alone or in combination. Cytokines/receptors were measured in the medium by immunoassays. RESULTS: Stimulation of genital mycoplasmas did not increase the proinflammatory cytokines, except Ureaplasma urealyticum that increased IL-8 levels. However, U. urealyticum and Mycoplasma hominis significantly increased IL-10 and IL-13 levels. G. vaginalis alone or in combination with genital mycoplasmas showed an increased proinflammatory and anti-inflammatory cytokines. CONCLUSIONS: G. vaginalis sustain a proinflammatory response in the fetal membranes in vitro, while genital mycoplasmas induce a strong control of the inflammatory response. The ability of genital mycoplasmas to control the proinflammatory response may favor their survival in the upper genital tract.

Amniotic membrane sweeping.

Amniotic membrane sweeping or stripping is a safe and effective method of labor induction supported by national obstetrical organizations. While its use dates back to antiquity by both midwives and physicians there are still areas that need further research to define its role in induction of labor. A review of the literature reveals that amniotic membrane sweeping is a safe, effective, and inexpensive method of labor induction. It can be done in the outpatient setting with minimal risks so long as it is avoided in patients with contraindications. Amniotic membrane sweeping can be performed in Group B Streptococcus-positive women with studies showing no increase in untoward outcomes. However, there is no data in women infected with HIV or hepatitis.

Amniocentesis for threatened preterm labor with intact membranes and the impact on adverse outcome in infants born at 22 to 28 weeks of gestation.

BACKGROUND: It remains unclear whether performing amniocentesis to detect intra-amniotic infection is useful for improving neonatal outcomes. AIMS: To determine the efficacy of amniocentesis on the neonatal outcomes in women exhibiting threatened preterm labor and intact membranes. STUDY DESIGN: Retrospective cohort study SUBJECTS: A total of 174 women with threatened preterm labor and intact membranes at 22 to 33 weeks of gestation. Women with obvious clinical chorioamnionitis, multifetal pregnancy and/or major anomalies were excluded. OUTCOME MEASURES: Neonatal short- and long-term outcomes RESULTS: Sixty-seven women underwent amniocentesis (Tap group), while the remaining 107 did not. The prevalence of a positive Gram stain or a positive culture result was 10% in the Tap group. The overall outcomes were not statistically different between the two groups, with the exception of borderline significance (p=0.052) in long-term outcomes, favoring the Tap group. We performed a subgroup analysis focusing on infants born at 22-28 weeks of gestation. Consequently, the Tap group had better neonatal outcomes than the no-Tap group with respect to both short-term (OR 0.19, 95%CI 0.07-0.55) and long-term (OR 0.15, 0.05-0.46) outcomes. A multivariate analysis revealed that after adjusting confounding factors, the gestational age at delivery (OR 0.4, 0.3-0.7) and amniocentesis (OR 0.1, 0.02-0.3) remained significantly different. CONCLUSIONS: Amniocentesis is useful for improving neonatal outcomes in infants born at 22-28 weeks of gestation to women exhibiting preterm labor and intact membranes.

The Effect of Simvastatin on Infection-Induced Inflammatory Response of Human Fetal Membranes.

PROBLEM: Inflammatory cytokines and matrix metalloproteinases (MMPs) contribute to preterm labor pathophysiology. The objective of this study was to test anti-inflammatory properties of simvastatin in human fetal membranes exposed to lipopolysaccharide (LPS). METHOD OF STUDY: Normal term human fetal membrane explants (n = 11) were allocated to one of the six study groups: control, LPS only (100 ng/mL), simvastatin only (125 ng/mL), simvastatin given 6 hrs prior to LPS (S-L), simvastatin given 6 hrs post-LPS (L-S), and simvastatin and LPS given simultaneously (L+S). Explants were incubated for 24 hrs. Multiplex ELISA for cytokines: IL-1ß, IL-6, IL-10, and TNF-α; soluble cytokine receptors: sIL-1R2, sIL-6R, sTNFR1, and R2; MMPs (1, 2, 7, 9, and 10); and tissue inhibitor of metalloproteinase-2 (TIMP-2) was performed on tissue culture supernatants. Pairwise comparison between different groups was conducted by least square mean estimates. RESULTS: Compared with controls, LPS stimulation increased cytokine production and their tissue bioavailability (measured as the molar ratio of cytokine to its soluble receptor), thus confirming membrane immune reactivity (P < 0.01). Pre-treatment with simvastatin (S-L) reduced IL-6 (P = 0.02), TNF-α (P = 0.02), and MMP-9 (P = 0.01); post-treatment (L-S) reduced IL-1ß (P = 0.02) and TNF-α (P = 0.04), while simultaneous treatment (L+S) did not reduce any of the cytokines tested. Simvastatin reduced the molar ratio of TNF-α/sTNFR1 or R2 and IL-1ß/sIL-1R2 (P = 0.01 and 0.04 in S-L group; P = 0.01 and 0.004 in L-S group, respectively). S-L additionally reduced MMP-9/TIMP-2 molar ratio (P = 0.0007). CONCLUSIONS: Simvastatin downregulates LPS-induced inflammatory response and restores homeostasis between pro- and anti-inflammatory processes. Simvastatin may reduce fetal inflammatory response associated with infection-induced preterm birth.

Escherichia coli-induced temporal and differential secretion of heat-shock protein 70 and interleukin-1ß by human fetal membranes in a two-compartment culture system.

INTRODUCTION: Escherichia coli is recognized as an etiological bacteria associated with chorioamnionitis and the preterm premature rupture of fetal membranes. This pathological condition induces pro-inflammatory cytokines and degradative metalloproteinases, which are considered biological markers secreted in an acute stage of infection. Heat-shock proteins (HSPs) are an important component of the innate immunity response and are found in different pathological conditions. They have not been previously measured in human fetal membranes in response to infectious conditions. We hypothesized that the choriodecidual tissue and amniotic epithelium secreted temporal and differential Hsp-60, Hsp-70, and interleukin (IL)-1ß mediated by E. coli infection. METHODS: Fetal membranes were mounted in a two-compartment culture system and infected with two passes of live E. coli at different doses (10², 104, 105, and 106 colony-forming units (CFU)/mL) and intervals of incubation (3, 6, and 24 h). The culture medium was collected, and Hsp-60, Hsp-70, and IL-1ß were assessed using the enzyme-linked immunosorbent assay (ELISA) method. RESULTS: After 3 and 6 h of infection, E. coli induced an increase in Hsp-70 secretion in the choriodecidual tissue. However, after 24 h of incubation, Hsp-70 was downregulated and we observed an increase in IL-1ß secretion. By contrast, E. coli induced a lower Hsp-60 secretion in the amnion compared to Hsp-70. DISCUSSION: Human fetal membranes responded actively to E. coli infection, with an increase in Hsp-70 during the first hours of infection. After 24 h, there was an increase in the liberation of IL-1ß.

Antepartum Membrane Stripping in GBS Carriers, Is It Safe? (The STRIP-G Study).

OBJECTIVE: Stripping of the membranes is an established and widely utilized obstetric procedure associated with higher spontaneous vaginal delivery rates, reduced need for formal induction of labor and a lower likelihood of post-term pregnancy. Due to the theoretical concern of bacterial seeding during the procedure many practitioners choose not to sweep the membranes in Group B Streptococcus (GBS) colonized patients. We conducted 'the STRIP-G study' in order to determine whether maternal and neonatal outcomes are affected by GBS carrier status in women undergoing membrane stripping. STUDY DESIGN: We conducted a prospective study in a tertiary referral center, comparing maternal and neonatal outcomes following membrane stripping among GBS-positive, GBS-negative, and GBS-unknown patients. We compared the incidence of composite adverse neonatal outcomes (primary outcome) among the three study groups, while secondary outcome measure was composite adverse maternal outcomes. RESULTS: A total of 542 women were included in the study, of which 135 were GBS-positive, 361 GBS-negative, and 46 GBS-unknown status. Demographic, obstetric, and intra-partum characteristics were similar for all groups. Adverse neonatal outcomes were observed in 8 (5.9%), 31 (8.6%), and 2 (4.3%) in the GBS-positive, GBS-negative, and unknown groups, respectively (P = 0.530), (Odds Ratio between GBS-Positive vs. GBS-Negative groups 0.67 (95%, CI = 0.30-1.50)); while composite adverse maternal outcomes occurred in 9 (6.66%), 31 (8.59%), and 5 (10.87%) in the GBS-positive, GBS-negative, and unknown groups, respectively (P = 0.617). CONCLUSIONS: Antepartum membrane stripping in GBS carriers appears to be a safe obstetrical procedure that does not adversely affect maternal or neonatal outcomes.

The extraembryonic serosa is a frontier epithelium providing the insect egg with a full-range innate immune response.

Drosophila larvae and adults possess a potent innate immune response, but the response of Drosophila eggs is poor. In contrast to Drosophila, eggs of the beetle Tribolium are protected by a serosa, an extraembryonic epithelium that is present in all insects except higher flies. In this study, we test a possible immune function of this frontier epithelium using Tc-zen1 RNAi-mediated deletion. First, we show that bacteria propagate twice as fast in serosa-less eggs. Then, we compare the complete transcriptomes of wild-type, control RNAi, and Tc-zen1 RNAi eggs before and after sterile or septic injury. Infection induces genes involved in Toll and IMD-signaling, melanisation, production of reactive oxygen species and antimicrobial peptides in wild-type eggs but not in serosa-less eggs. Finally, we demonstrate constitutive and induced immune gene expression in the serosal epithelium using in situ hybridization. We conclude that the serosa provides insect eggs with a full-range innate immune response.

Term and preterm labour are associated with distinct microbial community structures in placental membranes which are independent of mode of delivery.

Infection is considered a possible trigger for preterm labour, supported by evidence showing the presence of bacteria in the placenta and placental membranes from preterm births. In this study, 16S rDNA pyrosequencing was used to identify bacteria in placental membranes. Caesarean sections and vaginal deliveries at term were found to harbour common genera. Mycoplasma hominis, Aerococcus christensenii, Gardnerella vaginalis and Fusobacterium nucleatum were either only present in preterm membranes or in greater abundance than at term. These data support previous studies that used either targeted qPCR or broad-range 16S rDNA PCR and cloning but not a recent microbiome analysis of placental tissue using high-throughput sequencing.

Histologic chorioamnionitis: different histologic features at different gestational ages.

OBJECTIVE: The aim of this study was to investigate the rate of the different histological chorioamnionitis (HCA) grade in relation to the gestational age in term and preterm delivery. METHODS: Three hundred and ninety-two women with singleton pregnancy with spontaneous onset of labor either prematurely or at term, with histologic diagnosis of HCA, were enrolled. Placentas were classified as: deciduitis and/or histologic chorioamnionitis within the membranes (HCA1); amnionitis or inflammation of the chorionic plate without funisitis (HCA2); and histologic chorioamnionitis with funisitis (HCA3). Microbiological culture was performed on both placental and fetal membrane samples. RESULTS: HCA1 was more frequent in women delivering at term than in preterm (p < 0.001). HCA2 was more represented in women delivering between 32 and 36 weeks (p < 0.001) and HCA3 occurred more frequently in those delivering within 32 weeks (p < 0.001). The positive bacterial culture was higher (p = 0.008) in presence of HCA3 in comparison with HCA1 and HCA2. CONCLUSIONS: This study showed a significantly different distribution of HCA grades in relation to gestational age at delivery. HCA may represent the expression of different subtending etiologies and may also reflect specific immune competence of gestational tissues at different gestational ages, strengthening as pregnancy advances.

Ureaplasma parvum undergoes selection in utero resulting in genetically diverse isolates colonizing the chorioamnion of fetal sheep.

Ureaplasmas are the microorganisms most frequently isolated from the amniotic fluid of pregnant women and can cause chronic intrauterine infections. These tiny bacteria are thought to undergo rapid evolution and exhibit a hypermutatable phenotype; however, little is known about how ureaplasmas respond to selective pressures in utero. Using an ovine model of chronic intraamniotic infection, we investigated if exposure of ureaplasmas to subinhibitory concentrations of erythromycin could induce phenotypic or genetic indicators of macrolide resistance. At 55 days gestation, 12 pregnant ewes received an intraamniotic injection of a nonclonal, clinical Ureaplasma parvum strain followed by (i) erythromycin treatment (intramuscularly, 30 mg/kg/day, n = 6) or (ii) saline (intramuscularly, n = 6) at 100 days gestation. Fetuses were then delivered surgically at 125 days gestation. Despite injecting the same inoculum into all the ewes, significant differences between amniotic fluid and chorioamnion ureaplasmas were detected following chronic intraamniotic infection. Numerous polymorphisms were observed in domain V of the 23S rRNA gene of ureaplasmas isolated from the chorioamnion (but not the amniotic fluid), resulting in a mosaiclike sequence. Chorioamnion isolates also harbored the macrolide resistance genes erm(B) and msr(D) and were associated with variable roxithromycin minimum inhibitory concentrations. Remarkably, this variability occurred independently of exposure of ureaplasmas to erythromycin, suggesting that low-level erythromycin exposure does not induce ureaplasmal macrolide resistance in utero. Rather, the significant differences observed between amniotic fluid and chorioamnion ureaplasmas suggest that different anatomical sites may select for ureaplasma subtypes within nonclonal, clinical strains. This may have implications for the treatment of intrauterine ureaplasma infections.