Questioning the fetal microbiome illustrates pitfalls of low-biomass microbial studies.

Whether the human fetus and the prenatal intrauterine environment (amniotic fluid and placenta) are stably colonized by microbial communities in a healthy pregnancy remains a subject of debate. Here we evaluate recent studies that characterized microbial populations in human fetuses from the perspectives of reproductive biology, microbial ecology, bioinformatics, immunology, clinical microbiology and gnotobiology, and assess possible mechanisms by which the fetus might interact with microorganisms. Our analysis indicates that the detected microbial signals are likely the result of contamination during the clinical procedures to obtain fetal samples or during DNA extraction and DNA sequencing. Furthermore, the existence of live and replicating microbial populations in healthy fetal tissues is not compatible with fundamental concepts of immunology, clinical microbiology and the derivation of germ-free mammals. These conclusions are important to our understanding of human immune development and illustrate common pitfalls in the microbial analyses of many other low-biomass environments. The pursuit of a fetal microbiome serves as a cautionary example of the challenges of sequence-based microbiome studies when biomass is low or absent, and emphasizes the need for a trans-disciplinary approach that goes beyond contamination controls by also incorporating biological, ecological and mechanistic concepts.

RNA profiles reveal signatures of future health and disease in pregnancy.

Maternal morbidity and mortality continue to rise, and pre-eclampsia is a major driver of this burden1. Yet the ability to assess underlying pathophysiology before clinical presentation to enable identification of pregnancies at risk remains elusive. Here we demonstrate the ability of plasma cell-free RNA (cfRNA) to reveal patterns of normal pregnancy progression and determine the risk of developing pre-eclampsia months before clinical presentation. Our results centre on comprehensive transcriptome data from eight independent prospectively collected cohorts comprising 1,840 racially diverse pregnancies and retrospective analysis of 2,539 banked plasma samples. The pre-eclampsia data include 524 samples (72 cases and 452 non-cases) from two diverse independent cohorts collected 14.5 weeks (s.d., 4.5 weeks) before delivery. We show that cfRNA signatures from a single blood draw can track pregnancy progression at the placental, maternal and fetal levels and can robustly predict pre-eclampsia, with a sensitivity of 75% and a positive predictive value of 32.3% (s.d., 3%), which is superior to the state-of-the-art method2. cfRNA signatures of normal pregnancy progression and pre-eclampsia are independent of clinical factors, such as maternal age, body mass index and race, which cumulatively account for less than 1% of model variance. Further, the cfRNA signature for pre-eclampsia contains gene features linked to biological processes implicated in the underlying pathophysiology of pre-eclampsia.

The stepwise assembly of the neonatal virome is modulated by breastfeeding.

The gut of healthy human neonates is usually devoid of viruses at birth, but quickly becomes colonized, which-in some cases-leads to gastrointestinal disorders1-4. Here we show that the assembly of the viral community in neonates takes place in distinct steps. Fluorescent staining of virus-like particles purified from infant meconium or early stool samples shows few or no particles, but by one month of life particle numbers increase to 109 per gram, and these numbers seem to persist throughout life5-7. We investigated the origin of these viral populations using shotgun metagenomic sequencing of virus-enriched preparations and whole microbial communities, followed by targeted microbiological analyses. Results indicate that, early after birth, pioneer bacteria colonize the infant gut and by one month prophages induced from these bacteria provide the predominant population of virus-like particles. By four months of life, identifiable viruses that replicate in human cells become more prominent. Multiple human viruses were more abundant in stool samples from babies who were exclusively fed on formula milk compared with those fed partially or fully on breast milk, paralleling reports that breast milk can be protective against viral infections8-10. Bacteriophage populations also differed depending on whether or not the infant was breastfed. We show that the colonization of the infant gut is stepwise, first mainly by temperate bacteriophages induced from pioneer bacteria, and later by viruses that replicate in human cells; this second phase is modulated by breastfeeding.

Impact of exposure to air pollution on cervicovaginal microbial communities.

PURPOSE: Vaginal microbial communities can be dominated by anaerobic (community state type IV, CST IV) or Lactobacillus (other CSTs) species. CST IV is a risk factor for spontaneous preterm birth (sPTB) and is more common among Black than White populations. In the US, average air pollution exposures are higher among Black compared to White people and exert systemic health effects. We sought to (1) quantify associations of air pollution, specifically particulate matter <2.5 µm in diameter (PM2.5), with CST IV and (2) explore the extent to which racial disparities in PM2.5 exposure might explain racial differences in the prevalence of CST IV. DESIGN: Methods: We performed a secondary analysis of 566 participants of the Motherhood & Microbiome study. PM2.5 exposures were derived from a machine learning model integrating NASA satellite and EPA ground monitor data. Previously, cervicovaginal swabs from 15 to 20 weeks' gestation were analyzed using 16 S rRNA sequencing and hierarchical clustering assigned CSTs. Multivariable logistic regression models calculated adjusted odds ratios of CST IV (vs. other CSTs) per interquartile range (IQR) increment of PM2.5. Race-stratified and mediation analyses were performed. RESULTS: Higher PM2.5 exposure was associated with CST IV (aOR 1.39, 95% CI 1.02-1.91). Further adjustment for race/ethnicity attenuated the association (aOR 1.34, 95% CI: 0.97-1.83). Black participants (vs. White) had higher median PM2.5 exposure (10.6 vs. 9.6 µg/m3, P < 0.001) and higher prevalence of CST IV (47% vs. 11%, P < 0.001). Mediation analysis revealed that higher PM2.5 exposure may explain 3.9% (P = 0.038) and 3.3% (P = 0.15) of the Black-White disparity in CST IV in unadjusted and adjusted models, respectively. CONCLUSION: PM2.5 was associated with CST IV, a risk factor for sPTB. Additionally, PM2.5 exposure may partially explain racial differences in the prevalence of CST IV. Further research is warranted to discover how environmental exposures affect microbial composition and perpetuate racial health disparities.

The impact of postpartum social support on postpartum mental health outcomes during the COVID-19 pandemic.

Social support is an influential component of postpartum recovery, adjustment, and bonding, which was disrupted by social distancing recommendations related to the COVID-19 pandemic. This study reports on changes in the availability of social support for postpartum women during the pandemic, investigates how those changes may have contributed to postpartum mental health, and probes how specific types of social support buffered against poor postpartum mental health and maternal-infant bonding impairment. Participants were 833 pregnant patients receiving prenatal care in an urban USA setting and using an electronic patient portal to access self-report surveys at two time points, during pregnancy (April-July 2020) and at ~12 weeks postpartum (August 2020-March 2021). Measures included an assessment of COVID-19 pandemic-related change in social support, sources of social support, ratings of emotional and practical support, and postpartum outcomes including depression, anxiety, and maternal-infant bonding. Overall self-reported social support decreased during the pandemic. Decreased social support was associated with an increased risk of postpartum depression, postpartum anxiety, and impaired parent-infant bonding. Among women reporting low practical support, emotional support appeared to protect against clinically significant depressive symptoms and impaired bonding with the infant. Decreases in social support are associated with a risk for poor postpartum mental health outcomes and impaired maternal-infant bonding. Evaluation and promotion of social support are recommended for healthy adjustment and functioning of postpartum women and families.

Association between Low Urban Neighborhood Greenness and Hypertensive Disorders of Pregnancy.

OBJECTIVE: Urban neighborhood greenness is associated with greater cardiovascular health in the general population, and with better pregnancy and neonatal outcomes. Hypertension in pregnancy is a leading cause of maternal mortality and long-term cardiovascular morbidity and mortality in women. We sought to examine the association between greenness and hypertensive disorders of pregnancy. STUDY DESIGN: This study is a secondary analysis of a prospective cohort study of 1,943 women who received prenatal care from December 2013 to December 2016 at a single, urban, and tertiary academic medical center in Philadelphia, PA. Greenness measure was quantified via residential tree canopy cover within circumferential buffers of 100- and 500-meter radii around participants' homes. Associations between greenness and hypertensive disorders of pregnancy (defined as gestational hypertension or preeclampsia) were estimated by using multilevel logistic regression accounting for maternal sociodemographic information (race-ethnicity, insurance status, and age) medical history (diabetes, body mass index, smoking history, and parity), neighborhood deprivation index, and including 1,225 Philadelphia residents for whom key exposure and outcome data were available. RESULTS: At baseline, the participants' mean (SD) age was 27.5 (5.9) years, (range: 14-44 years). The majority of participants were non-Hispanic Black (857, 70.2%). Participants with less residential tree canopy cover were significantly more likely to have hypertensive disorders of pregnancy. The multivariable-adjusted odds ratio of hypertensive disorders of pregnancy among participants with less than 10% compared with those with greater than 30% tree canopy cover was 2.14 (95% confidence interval [CI]: 1.11-4.15) within 100-meter buffer. CONCLUSION: In our cohort, greenness was associated with lower hypertensive disorders of pregnancy odds. Our findings add to evidence that greenness may confer health benefits and warrant further investigations in identifying whether there is a causal pathway through which greenness may be protective against hypertensive disorders of pregnancy. KEY POINTS: · Low residential tree canopy is associated with increased risk of hypertensive disorders of pregnancy. · 100-meter buffers are most sensitive in identifying associations between tree canopy and HDP risk. · The role of greenness against hypertensive disorders of pregnancy should be further studied experimentally.

Cervicovaginal Microbial-Immune State and Group B Streptococcus Colonization in Pregnancy.

OBJECTIVE: Maternal colonization with Group B Streptococcus (GBS) is a significant risk factor for serious neonatal morbidity. There are limited data on how the cervicovaginal (CV) microbiota and host immune factor ß-defensin-2 might influence GBS colonization in pregnant individuals. This study sought to determine if the CV microbiota is associated with GBS colonization in pregnant individuals, and if ß-defensin-2 modifies this relationship. STUDY DESIGN: This was a secondary analysis of a prospective cohort study of pregnant individuals with singleton pregnancies who had CV microbiota specimens analyzed at 16 to 20, 20 to 24, and 24 to 28 weeks' gestation, along with a third trimester GBS rectovaginal (RV) culture (n = 492). Microbiota data were analyzed with 16S rRNA gene sequencing and classified into community state types (CSTs). Log-binomial multivariable regression was used to model associations between CST and GBS RV status and to calculate risk ratios. ß-defensin-2, an immune factor known to modulate the relationship between CST and pregnancy outcomes, was examined as an effect modifier. RESULTS: Of 492 individuals, 34.3% were GBS RV + . Compared with individuals with CST I at 16 to 20 weeks, individuals with CST IV-A and CST II had a significantly elevated relative risk of subsequent GBS RV+ status. When stratified by high and low ß-defensin-2 levels, ß-defensin-2 was found to be an effect modifier of the association between CST IV-A and GBS RV+ status. In individuals with low ß-defensin-2 levels, CST VI-A was associated with GBS RV+ status, but among individuals with high ß-defensin-2 levels, there was no such association (interaction p-value = 0.03). CONCLUSION: Pregnant individuals with CV microbiota characterized by CST IV-A and CST II had significantly elevated risk of GBS RV colonization in the third trimester compared with those with CST I, and ß-defensin-2 was an effect modifier of the association between CST IV-A and GBS RV+ status. Future research should investigate if manipulation of the CV microbiota can prevent GBS colonization, thereby reducing intrapartum antibiotic prophylaxis and the risks of neonatal GBS infection. KEY POINTS: · The relationship between the CV microbiota and GBS RV colonization is unknown.. · A Lactobacillus-deficient, anaerobic rich vaginal community, CST IV-A, is associated with increased risk of GBS RV colonization.. · ß-defensin-2 is an effect modifier of the association between CST IV-A and GBS RV+ status..

Racism and perinatal health inequities research: where we have been and where we should go.

For more than a century, substantial racial and ethnic inequities in perinatal health outcomes have persisted despite technical clinical advances and changes in public health practice that lowered the overall incidence of morbidity. Race is a social construct and not an inherent biologic or genetic reality; therefore, racial differences in health outcomes represent the consequences of structural racism or the inequitable distribution of opportunities for health along racialized lines. Clinicians and scientists in obstetrics and gynecology have a responsibility to work to eliminate health inequities for Black, Brown, and Indigenous birthing people, and fulfilling this responsibility requires actionable evidence from high-quality research. To generate this actionable evidence, the research community must realign paradigms, praxis, and infrastructure with an eye directed toward reproductive justice and antiracism. This special report offers a set of key recommendations as a roadmap to transform perinatal health research to achieve health equity. The recommendations are based on expert opinion and evidence presented at the State of the Science Research Symposium at the 41st Annual Pregnancy Meeting of the Society for Maternal-Fetal Medicine in 2021. Recommendations fall into 3 broad categories-changing research paradigms, reforming research praxis, and transforming research infrastructure-and are grounded in a historic foundation of the advances and shortcomings of clinical, public health, and sociologic scholarship in health equity. Changing the research paradigm requires leveraging a multidisciplinary perspective on structural racism; promoting mechanistic research that identifies the biologic pathways perturbed by structural racism; and utilizing conceptual models that account for racism as a factor in adverse perinatal outcomes. Changing praxis approaches to promote and engage multidisciplinary teams and to develop standardized guidelines for data collection will ensure that paradigm shifts center the historically marginalized voices of Black, Brown, and Indigenous birthing people. Finally, infrastructure changes that embed community-centered approaches are required to make shifts in paradigm and praxis possible. Institutional policies that break down silos and support true community partnership, and also the alignment of institutional, funding, and academic publishing objectives with strategic priorities for perinatal health equity, are paramount. Achieving health equity requires shifting the structures that support the ecosystem of racism that Black, Brown, and Indigenous birthing people must navigate before, during, and after childbearing. These structures extend beyond the healthcare system in which clinicians operate day-to-day, but they cannot be excluded from research endeavors to create the actionable evidence needed to achieve perinatal health equity.

Second trimester short cervix is associated with decreased abundance of cervicovaginal lipid metabolites.

BACKGROUND: A short cervix is a risk factor for preterm birth. The molecular drivers of a short cervix remain elusive. Metabolites may function as mediators of pathologic processes. OBJECTIVE: We sought to determine if a distinct cervicovaginal metabolomic profile is associated with a short cervix (<25 mm) to unveil the potential mechanisms by which premature cervical remodeling leads to a short cervix. STUDY DESIGN: This was a secondary analysis of a completed prospective pregnancy cohort. Cervicovaginal fluid was obtained between 20 and 24 weeks' gestation. The participants selected for metabolomic profiling were frequency-matched by birth outcome and cervicovaginal microbiota profile. This analysis included 222 participants with cervical length measured. A short cervix was defined as one having length <25 mm, as measured by transvaginal ultrasound. Unpaired t-tests were performed with a Bonferroni correction for multiple comparisons. RESULTS: There were 27 participants with a short cervix, and 195 with normal cervical length. Of the 637 metabolites detected, 26 differed between those with a short cervix and those with normal cervical lengths; 22 were decreased, of which 21 belonged to the lipid metabolism pathway (all P<.000079). Diethanolamine, erythritol, progesterone, and mannitol or sorbitol were increased in the cases of short cervix. Among participants with Lactobacillus-deficient microbiota, only diethanolamine and mannitol or sorbitol differed between short cervix (n=17) and normal cervical length (n=75), both increased. CONCLUSION: A short cervix is associated with decreased cervicovaginal lipid metabolites, particularly sphingolipids. This class of lipids stabilizes cell membranes and protects against environmental exposures. Increased diethanolamine-an immunostimulatory xenobiotic-is associated with a short cervix. These observations begin to identify the potential mechanisms by which modifiable environmental factors may invoke cell damage in the setting of biological vulnerability, thus promoting premature cervical remodeling in spontaneous preterm birth.

Predictive RNA profiles for early and very early spontaneous preterm birth.

BACKGROUND: Spontaneous preterm birth remains the main driver of childhood morbidity and mortality. Because of an incomplete understanding of the molecular pathways that result in spontaneous preterm birth, accurate predictive markers and target therapeutics remain elusive. OBJECTIVE: This study sought to determine if a cell-free RNA profile could reveal a molecular signature in maternal blood months before the onset of spontaneous preterm birth. STUDY DESIGN: Maternal samples (n=242) were obtained from a prospective cohort of individuals with a singleton pregnancy across 4 clinical sites at 12-24 weeks (nested case-control; n=46 spontaneous preterm birth <35 weeks and n=194 term controls). Plasma was processed via a next-generation sequencing pipeline for cell-free RNA using the Mirvie RNA platform. Transcripts that were differentially expressed in next-generation sequencing cases and controls were identified. Enriched pathways were identified in the Reactome database using overrepresentation analysis. RESULTS: Twenty five transcripts associated with an increased risk of spontaneous preterm birth were identified. A logistic regression model was developed using these transcripts to predict spontaneous preterm birth with an area under the curve =0.80 (95% confidence interval, 0.72-0.87) (sensitivity=0.76, specificity=0.72). The gene discovery and model were validated through leave-one-out cross-validation. A unique set of 39 genes was identified from cases of very early spontaneous preterm birth (<25 weeks, n=14 cases with time to delivery of 2.5±1.8 weeks); a logistic regression classifier on the basis of these genes yielded an area under the curve=0.76 (95% confidence interval, 0.63-0.87) in leave-one-out cross validation. Pathway analysis for the transcripts associated with spontaneous preterm birth revealed enrichment of genes related to collagen or the extracellular matrix in those who ultimately had a spontaneous preterm birth at <35 weeks. Enrichment for genes in insulin-like growth factor transport and amino acid metabolism pathways were associated with spontaneous preterm birth at <25 weeks. CONCLUSION: Second trimester cell-free RNA profiles in maternal blood provide a noninvasive window to future occurrence of spontaneous preterm birth. The systemic finding of changes in collagen and extracellular matrix pathways may serve to identify individuals at risk for premature cervical remodeling, with growth factor and metabolic pathways implicated more often in very early spontaneous preterm birth. The use of cell-free RNA profiles has the potential to accurately identify those at risk for spontaneous preterm birth by revealing the underlying pathophysiology, creating an opportunity for more targeted therapeutics and effective interventions.

Clinician-reported childbirth outcomes, patient-reported childbirth trauma, and risk for postpartum depression.

Childbirth trauma is common and increases risk for postpartum depression (PPD). However, we lack brief measures to reliably identify individuals who experience childbirth trauma and who may be at greater prospective risk for PPD. To address this gap, we used data from a racially diverse prospective cohort (n=1082). We collected survey data during pregnancy and at 12 weeks postpartum, as well as clinician-reported data from medical records. A new three-item measure of patient-reported childbirth trauma was a robust and independent risk factor for PPD, above and beyond other known risk factors for PPD, including prenatal anxiety and depression. Cesarean birth, greater blood loss, and preterm birth were each associated with greater patient-reported childbirth trauma. Finally, there were prospective indirect pathways whereby cesarean birth and higher blood loss were related to higher patient-reported childbirth trauma, in turn predicting greater risk for PPD. Early universal postpartum screening for childbirth trauma, targeted attention to individuals with childbirth complications, and continued screening for depression and anxiety can identify individuals at risk for PPD. Such efforts can inform targeted interventions to improve maternal mental health, which plays a vital role in infant development.

Intrauterine Inflammation Leads to Select Sex- and Age-Specific Behavior and Molecular Differences in Mice.

Sex-specific differences in behavior have been observed in anxiety and learning in children exposed to prenatal inflammation; however, whether these behaviors manifest differently by age is unknown. This study assesses possible behavioral changes due to in utero inflammation as a function of age in neonatal, juvenile, and adult animals and presents potential molecular targets for observed differences. CD-1 timed pregnant dams were injected in utero with lipopolysaccharide (LPS, 50 µg/animal) or saline at embryonic day 15. No differences in stress responses were measured by neonatal ultrasonic vocalizations between LPS- and saline-exposed groups of either sex. By contrast, prenatal inflammation caused a male-specific increase in anxiety in mature but not juvenile animals. Juvenile LPS-exposed females had decreased movement in open field testing that was not present in adult animals. We additionally observed improved memory retrieval after in utero LPS in the juvenile animals of both sexes, which in males may be related to a perseverative phenotype. However, there was an impairment of long-term memory in only adult LPS-exposed females. Finally, gene expression analyses revealed that LPS induced sex-specific changes in genes involved in hippocampal neurogenesis. In conclusion, intrauterine inflammation has age- and sex-specific effects on anxiety and learning that may correlate to sex-specific disruption of gene expression associated with neurogenesis in the hippocampus.

Coronavirus disease 2019 vaccine response in pregnant and lactating women: a cohort study.

BACKGROUND: Pregnant and lactating women were excluded from initial coronavirus disease 2019 vaccine trials; thus, data to guide vaccine decision making are lacking. OBJECTIVE: This study aimed to evaluate the immunogenicity and reactogenicity of coronavirus disease 2019 messenger RNA vaccination in pregnant and lactating women compared with: (1) nonpregnant controls and (2) natural coronavirus disease 2019 infection in pregnancy. STUDY DESIGN: A total of 131 reproductive-age vaccine recipients (84 pregnant, 31 lactating, and 16 nonpregnant women) were enrolled in a prospective cohort study at 2 academic medical centers. Titers of severe acute respiratory syndrome coronavirus 2 spike and receptor-binding domain immunoglobulin G, immunoglobulin A, and immunoglobulin M were quantified in participant sera (n=131) and breastmilk (n=31) at baseline, at the second vaccine dose, at 2 to 6 weeks after the second vaccine, and at delivery by Luminex. Umbilical cord sera (n=10) titers were assessed at delivery. Titers were compared with those of pregnant women 4 to 12 weeks from the natural infection (n=37) by enzyme-linked immunosorbent assay. A pseudovirus neutralization assay was used to quantify neutralizing antibody titers for the subset of women who delivered during the study period. Postvaccination symptoms were assessed via questionnaire. Kruskal-Wallis tests and a mixed-effects model, with correction for multiple comparisons, were used to assess differences among groups. RESULTS: Vaccine-induced antibody titers were equivalent in pregnant and lactating compared with nonpregnant women (pregnant, median, 5.59; interquartile range, 4.68-5.89; lactating, median, 5.74; interquartile range, 5.06-6.22; nonpregnant, median, 5.62; interquartile range, 4.77-5.98, P=.24). All titers were significantly higher than those induced by severe acute respiratory syndrome coronavirus 2 infection during pregnancy (P<.0001). Vaccine-generated antibodies were present in all umbilical cord blood and breastmilk samples. Neutralizing antibody titers were lower in umbilical cord than maternal sera, although this finding did not achieve statistical significance (maternal sera, median, 104.7; interquartile range, 61.2-188.2; cord sera, median, 52.3; interquartile range, 11.7-69.6; P=.05). The second vaccine dose (boost dose) increased severe acute respiratory syndrome coronavirus 2-specific immunoglobulin G, but not immunoglobulin A, in maternal blood and breastmilk. No differences were noted in reactogenicity across the groups. CONCLUSION: Coronavirus disease 2019 messenger RNA vaccines generated robust humoral immunity in pregnant and lactating women, with immunogenicity and reactogenicity similar to that observed in nonpregnant women. Vaccine-induced immune responses were statistically significantly greater than the response to natural infection. Immune transfer to neonates occurred via placenta and breastmilk.

Associations of public water system trihalomethane exposure during pregnancy with spontaneous preterm birth and the cervicovaginal microbial-immune state.

BACKGROUND: Water total trihalomethanes (TTHMs) are disinfectant byproducts found in municipal water supplies. TTHM exposure has been linked to cancer and may be associated with adverse reproductive outcomes. A non-optimal cervicovaginal microbiota and low cervicovaginal beta-defensin-2 levels are associated with increased risk of spontaneous preterm birth. Whether TTHM exposure increases the risk of spontaneous preterm birth or alters the cervicovaginal microbial or immune state is unknown. OBJECTIVE: Investigate associations of water TTHM levels with spontaneous preterm birth, a non-optimal cervicovaginal microbiota, and beta-defensin-2 levels in a completed, diverse, urban pregnancy cohort. We hypothesized that higher TTHM levels would be associated with spontaneous preterm birth, a non-optimal cervicovaginal microbiota, and lower beta-defensin-2 levels. DESIGN: Methods: This was a secondary analysis of participants (n = 474) in the Motherhood & Microbiome (M&M) study (n = 2000), who lived in Philadelphia and had cervicovaginal samples analyzed for cervicovaginal microbiota composition and beta-defensin-2 levels. The microbiota was classified into community state types (CSTs). CST IV (non-optimal microbiota) is characterized by a paucity of Lactobacillus species and wide array of anaerobes. Municipal water TTHM levels were obtained from 16 sites monthly across the city of Philadelphia to establish mean residential water supply levels for each participant for the first four months of pregnancy (prior to vaginal swab collection at 16-20 weeks' gestation). Associations of water TTHM levels with spontaneous preterm birth and a non-optimal cervicovaginal microbiota birth were analyzed using multivariable logistic regression. Multivariable linear regression was used to model associations of water TTHM levels with log-transformed cervicovaginal beta-defensin-2 levels. Since water TTHM levels vary by season and beta-defensin-2 levels have been shown to differ by race, stratified models by warm (April-September) and cold (October-March) seasons as well as by self-identified race were utilized. RESULTS: Participants' water supply TTHM levels (mean µg/L [SD]) were higher in the warm (53.5 [9.4]) than cold (33.4 [7.5]) season (p < 0.0001). TTHM levels were non-significantly higher among Black participants than non-Black participants (44.8 [13.5] vs. 41.8 [11.8], p = 0.07). No associations were detected between TTHM with spontaneous preterm birth (per SD increment of TTHM, aOR 0.94, 95%CI: 0.66, 1.34) or with CST IV (aOR 0.94, 95%CI: 0.86, 1.16). Counter to our hypothesis, we observed positive associations of water TTHM with log-transformed cervicovaginal beta-defensin-2 levels in unadjusted models (ß 0.20 [95%CI: 0.02, 0.39]) per SD increment of TTHM), but the association was null after adjustment for season. However, in models adjusted for covariates including season and stratified by race, TTHM was significantly associated with lower beta-defensin-2 levels among non-Black participants (ß -0.75 [95%CI: -1.43, -0.08]) but not among Black participants (ß 0.17 [95%CI: -0.15, 0.49]), interaction p = 0.013). CONCLUSION: We did not detect associations of water TTHM levels with spontaneous preterm birth or the structure of the cervicovaginal microbiota. However, the finding of a significant interaction between TTHM and race on beta-defensin-2 levels suggest that environmental exposures may contribute to differences in reproductive tract innate immune function by race. Future studies to delineate environmental contributions to the cervicovaginal microbial-immune state, a potentially important biologic underpinning for preterm birth, are warranted.

Urban residential tree canopy and perceived stress among pregnant women.

OBJECTIVE: To examine the association of urban residential tree canopy cover with perceived stress in a cohort of pregnant women in Philadelphia, PA, and explore whether this association differed among participants with a history of anxiety and depression. STUDY DESIGN: We performed a secondary analysis of 1294 participants of the Motherhood & Microbiome (M&M) pregnancy cohort who lived in Philadelphia, with first visit perceived stress (Cohen's Perceived Stress Scale, PSS-14), and key covariate data. Tree canopy cover was calculated as percent cover within 100 and 500 m radii buffers around participants' homes. We performed multilevel mixed effects linear regression models, with perceived stress as the dependent variable. The main independent variable was tree canopy coverage. Individual-level covariates included season of last menstrual period, history of depression or anxiety, race/ethnicity, insurance, parity, and age. Census tract neighborhood deprivation index was used to account for area-level socioeconomic confounding variables. We also examined whether a history of anxiety or depression, modified the association between tree canopy coverage and perceived stress. RESULTS: Most participants were non-Hispanic Black (70.6%, n = 913), on Medicaid or uninsured (60.4%, n = 781), and 15.8% (n = 204) of participants had a prior history of depression or anxiety. We did not detect associations between tree canopy coverage and perceived stress overall. However, we detected effect modification; among participants with a history of depression or anxiety, each standard deviation increase in tree canopy cover was associated with lower PSS-14 in 100 m buffers (ß -1.0, 95% CI -1.8, -0.2), but not among participants with no histories of depression or anxiety (ß 0.2, 95% CI -0.3, 0.7) (interaction P = 0.007). Results were similar in directionality but not statistically significant within 500 m buffers. CONCLUSION: Residential tree canopy coverage was associated with reduced perceived stress among urban-dwelling pregnant women with history of anxiety or depression. Future studies of the effects of greenness and other stress-reducing efforts should consider underlying mental health conditions as effect modifiers.

Effect of a Nonoptimal Cervicovaginal Microbiota and Psychosocial Stress on Recurrent Spontaneous Preterm Birth.

OBJECTIVE: While select cervicovaginal microbiota and psychosocial factors have been associated with spontaneous preterm birth, their effect on the risk of recurrence remains unclear. It is also unknown whether psychosocial factors amplify underlying biologic risk. This study sought to determine the effect of nonoptimal cervicovaginal microbiota and perceived stress on the risk of recurrent spontaneous preterm birth. STUDY DESIGN: This was a secondary analysis of a prospective pregnancy cohort, Motherhood and Microbiome. The Cohen's Perceived Stress Scale (PSS-14) was administered and cervical swabs were obtained between 16 and 20 weeks of gestation. PSS-14 scores ≥30 reflected high perceived stress. We analyzed cervicovaginal microbiota using 16S rRNA sequencing and classified microbial communities into community state types (CSTs). CST IV is a nonoptimal cervicovaginal microbial community characterized by anaerobes and a lack of Lactobacillus. The final cohort included a predominantly non-Hispanic Black population of women with prior spontaneous preterm birth who had recurrent spontaneous preterm birth or term birth and had stress measurements (n = 181). A subanalysis was performed in the subset of these women with cervicovaginal microbiota data (n = 74). Multivariable logistic regression modeled adjusted associations between CST IV and recurrent spontaneous preterm birth, high stress and recurrent spontaneous preterm birth, as well as high stress and CST IV. RESULTS: Among the 181 women with prior spontaneous preterm birth, 45 (24.9%) had high perceived stress. We did not detect a significant association between high stress and recurrent spontaneous preterm birth (adjusted odds ratio [aOR] 1.67, 95% confidence interval [CI]: 0.73-3.85). Among the 74 women with prior spontaneous preterm birth and cervicovaginal microbiota analyzed, 29 (39.2%) had CST IV; this proportion differed significantly among women with recurrent spontaneous preterm birth (51.4%) compared with women with term birth (28.2%) (p = 0.04). In models adjusted for race and marital status, the association between CST IV and recurrent spontaneous preterm birth persisted (aOR 3.58, 95% CI: 1.25-10.24). There was no significant interaction between stress and CST IV on the odds of spontaneous preterm birth (p = 0.328). When both stress and CST IV were introduced into the model, their associations with recurrent spontaneous preterm birth were slightly stronger than when they were in the model alone. The aOR for stress with recurrent spontaneous preterm birth was 2.02 (95% CI: 0.61-6.71) and for CST IV the aOR was 3.83 (95% CI: 1.30-11.33). Compared to women with neither of the two exposures, women with both high stress and CST IV had the highest odds of recurrent spontaneous preterm birth (aOR = 6.01, 95% CI: 1.002-36.03). CONCLUSION: Among a predominantly non-Hispanic Black cohort of women with a prior spontaneous preterm birth, a nonoptimal cervicovaginal microbiota is associated with increased odds of recurrent spontaneous preterm birth. Adjustment for perceived stress may amplify associations between CST IV and recurrent spontaneous preterm birth. Identification of modifiable social or behavioral factors may unveil novel nonpharmacologic interventions to decrease recurrent spontaneous preterm birth among women with underlying biologic risk. KEY POINTS: · CST IV, a nonoptimal microbiota, is associated with increased odds of recurrent spontaneous preterm birth.. · Adjustment for perceived stress amplified associations between CST IV and recurrent spontaneous preterm birth.. · Identification of modifiable psychosocial factors may unveil novel nonpharmacologic interventions to decrease recurrent preterm birth..

Cervicovaginal microbial communities deficient in Lactobacillus species are associated with second trimester short cervix.

BACKGROUND: The cervix functions as a barrier to ascending pathogens in pregnancy. Short cervical length and lack of cervicovaginal Lactobacillus species are risk factors for spontaneous preterm birth; however, whether they interact to increase risk remains unknown. OBJECTIVE: We sought to examine the relationship between cervicovaginal microbiota and short cervix as well as their combined impact on spontaneous preterm birth risk. STUDY DESIGN: This was a secondary analysis of a prospective nested, case-control pregnancy study. Cervical swabs were collected between 16 and 20 weeks of gestation. Cervical length was measured per standard clinical care during a clinically indicated ultrasound at approximately 20 weeks of gestation. Cervicovaginal microbiota were analyzed with 16S ribosomal RNA gene sequencing and classified into community state types among 67 cases of spontaneous preterm birth, 47 cases of medically indicated preterm birth, and 358 cases of term births. Logistic regression was used to model associations of community state type IV, a community characterized by a paucity of Lactobacillus species and a wide array of anaerobic bacteria, and short cervix (<25 mm) as well as to model the association of a combination of short cervix and community state type IV with the odds of spontaneous preterm birth. RESULTS: Among the 472 women in the data set, there were 38 with short cervix (8.1%) and 177 with community state type IV (37.5%). Short cervix was associated with spontaneous preterm birth (adjusted odds ratio, 15.59; 95% confidence interval, 6.77-35.92). Women with community state type IV had higher odds of short cervix (adjusted odds ratio, 2.17; 95% confidence interval, 1.04-4.53) as well as spontaneous preterm birth (adjusted odds ratio, 1.97; 95% confidence interval, 1.06-3.65). While the interaction of community state type IV and short cervix was not significant (P = .771), women with both short cervix and community state type IV (n = 20) had higher odds of spontaneous preterm birth compared with women with both normal cervical length and community state types I, II, III, or V (n = 277) (adjusted odds ratio, 21.8; 95% confidence interval, 6.78-70.2). CONCLUSION: Community state type IV, characterized by a diverse set of strict and facultative anaerobes and a paucity of Lactobacillus species, is associated with increased odds of short cervix. Women with both community state type IV and short cervix have higher odds of spontaneous preterm birth than women with either factor alone. Determining the cascade of events leading to premature cervical shortening, including dysbiosis, may be critical in preventing spontaneous preterm birth.

The promise and pitfalls of precision medicine to resolve black-white racial disparities in preterm birth.

Differences in preterm birth rates between black and white women are the largest contributor to racial disparities in infant mortality. In today's age of precision medicine, analysis of the genome, epigenome, metabolome, and microbiome has generated interest in determining whether these biomarkers can help explain racial disparities. We propose that there are pitfalls as well as opportunities when using precision medicine analyses to interrogate disparities in health. To conclude that racial disparities in complex conditions are genetic in origin ignores robust evidence that social and environmental factors that track with race are major contributors to disparities. Biomarkers measured in omic assays that may be more environmentally responsive than genomics, such as the epigenome or metabolome, may be on the causal pathway of race and preterm birth, but omic observational studies suffer from the same limitations as traditional cohort studies. Confounding can lead to false conclusions about the causal relationship between omics and preterm birth. Methodological strategies (including stratification and causal mediation analyses) may help to ensure that associations between biomarkers and exposures, as well as between biomarkers and outcomes, are valid signals. These epidemiologic strategies present opportunities to assess whether precision medicine biomarkers can uncover biology underlying perinatal health disparities.

The Metabolomic Signature of the Placenta in Spontaneous Preterm Birth.

The placenta is metabolically active and supports the growth of the fetus. We hypothesize that deficits in the capacity of the placenta to maintain bioenergetic and metabolic stability during pregnancy may result in spontaneous preterm birth (SPTB). To explore this hypothesis, we performed a nested cased control study of metabolomic signatures in placentas from women with SPTB (<36 weeks gestation) compared to normal pregnancies (≥38 weeks gestation). To control for the effects of gestational age on placenta metabolism, we also studied a subset of metabolites in non-laboring preterm and term Rhesus monkeys. Comprehensive quantification of metabolites demonstrated a significant elevation in the levels of amino acids, prostaglandins, sphingolipids, lysolipids, and acylcarnitines in SPTB placenta compared to term placenta. Additional quantification of placental acylcarnitines by tandem mass spectrometry confirmed the significant elevation in SPTB human, with no significant differences between midgestation and term placenta in Rhesus macaque. Fatty acid oxidation as measured by the flux of 3H-palmitate in SPTB placenta was lower than term. Collectively, significant and biologically relevant alterations in the placenta metabolome were identified in SPTB placenta. Altered acylcarnitine levels and fatty acid oxidation suggest that disruption in normal substrate metabolism is associated with SPTB.