Biology subjects

Ravel, J.

Publications and source records attributed to Ravel, J..

28 records · Page 2Linked to original sources

bioRxiv2021

Identification of shared bacterial strains in the vaginal microbiota of reproductive-age mothers and daughters using genome-resolved metagenomics

It has been suggested that the human microbiome might be vertically transmitted from mother to offspring and that early colonizers may play a critical role in development of the immune system. Studies have shown limited support for the vertical transmission of the intestinal microbiota but the derivation of the vaginal microbiota remains largely unknown. Although the vaginal microbiota of children and reproductive age cis women differ in composition, the vaginal microbiota could be vertically transmitted. To determine whether there was any support for this hypothesis, we examined the vaginal microbiota of daughter-mother pairs from the Baltimore metropolitan area (ages 14-27, 32-51; n=39). We assessed whether the daughters microbiota was similar in composition to their mothers using metataxonomics. Permutation tests revealed that while some pairs did have similar vaginal microbiota, the degree of similarity did not exceed that expected by chance. Genome-resolved metagenomics was used to identify shared bacterial strains in a subset of the families (n=22). We found a small number of bacterial strains that were shared between mother-daughter pairs but identified more shared strains between individuals from different families, indicating that vaginal bacteria may display biogeographic patterns. Earlier-in-life studies are needed to demonstrate vertical transmission of the vaginal microbiota. ImportanceEarly colonizers of our microbiota are theorized to play an important role in the development of our immune system, yet we know little about how these communities are established. Vertical transmission from mother to offspring at the time of birth is theorized to be a major source of early colonizers but limited evidence supporting this process has only been shown for the intestinal tract microbiota. The provenance of the vaginal microbiota is largely unknown, although some have posited it is similarly vertically transmitted. We examined the vaginal microbiota of mother-daughter pairs and found limited evidence in support of this hypothesis. However, our analysis also revealed putative biogeographic patterns in the distribution of the strains which comprise the vaginal microbiota. Our results give insight into the role of vertical transmission for the vaginal microbiota and motivate future studies on the biogeography of these bacteria.

microbiology↗

bioRxiv2021

Three-dimensional models of the cervicovaginal epithelia to study host-microbiome interactions and sexually transmitted infections

Two-dimensional (2D) cell culture systems have provided controlled, reproducible means to analyze host-pathogen interactions. Although inexpensive, straightforward, and requiring very short time commitment, these models recapitulate neither the functionality of multi-layered cell types nor the microbial diversity of an infected human. Animal models have commonly been used to recreate the complexity of human infections. However, extensive modifications are commonly required to recreate interactions that resemble those in the human reproductive tract microbiologically and physiologically. Three-dimensional (3D) cell culture models have emerged as alternative means of reproducing key elements of human infections at a fraction of the cost of animal models and on a scale that allows for replicative experiments to be readily performed. Here we describe a new 3D model that utilizes transwells with epithelial cells seeded apically and a basolateral extra cellular matrix (ECM)-like layer containing collagen and fibroblasts. In this system, basal feeding creates a liquid/air interface on the apical side. The model produced tissues with close morphologic and physiological resemblance to human cervical and vaginal epithelia, including observable levels of mucus produced by cervical cells. Infection by both Chlamydia trachomatis and Neisseria gonorrhoeae was demonstrated as well as the growth of bacterial species observed in the human vaginal microbiota, enabling controlled mechanistic analyses of the interactions between host cells, vaginal microbiota and STI pathogens. Future experiments may include immune cells to mimic more closely the genital environment. Finally, the modular set up of the model makes it fully applicable to the analysis of non-genital host-microbiome-pathogen interactions. IMPORTANCEInfected sites in humans are a complex mix of host and microbial cell types interacting with each other to perform specific and necessary functions. The ability to understand the mechanism(s) that facilitate these interactions, and interactions with external factors is paramount to being able to develop preventative therapies. Models that attempt to faithfully replicate the complexity of these interactions are time intensive, costly, and not conducive to high throughput analysis. Two-dimensional (2D) models that have been used as a platform to understand these interactions, while cost effective, are generally limiting in experimental flexibility and structural/physiological relevance. Our three-dimensional (3D) models of the cervicovaginal epithelium can facilitate analysis of interactions between the host epithelium, sexually transmitted pathogens and bacteria present in the vaginal microbiota. Due to the modular design, additional cell types and environmental modulators can be introduced to the system to provide added complexity, approaching conditions in the infected human host.

cell biology↗

bioRxiv2021

Insight into the ecology of vaginal bacteria through integrative analyses of metagenomic and metatranscriptomic data

Vaginal bacterial communities dominated by Lactobacillus species are associated with a reduced risk to various adverse health outcomes. However, somewhat unexpectedly many healthy women have microbiota that are not dominated by lactobacilli. To determine the factors that drive vaginal community composition we characterized the genetic composition and transcriptional activities of vaginal microbiota in healthy women. We demonstrated that the abundance of a species is not always indicative of its transcriptional activity and that impending changes in community composition can be predicted from metatranscriptomic data. Functional comparisons highlight differences in the metabolic activities of these communities, notably in their degradation of host-produced mucin but not glycogen. Degradation of mucin by communities not dominated by Lactobacillus may play a role in their association with adverse health outcomes. Finally, we show that the transcriptional activities of L. crispatus, L. iners, and G. vaginalis vary with the taxonomic composition of the communities in which they reside. Notably, L. iners and G. vaginalis both demonstrated lower expression of their cholesterol-dependent cytolysins when co-resident with Lactobacillus spp. and higher expression when co-resident with other facultative and obligate anaerobes. The pathogenic potential of these species may depend on the communities in which they reside and thus could be modulated by interventional strategies. Our results provide insight to the functional ecology of vaginal microbiota and reveal strategies for management of these ecosystems.

microbiology↗

bioRxiv2021

Preterm birth is associated with xenobiotics and predicted by the vaginal metabolome

Spontaneous preterm birth (sPTB) is a leading cause of maternal and neonatal morbidity and mortality, yet both its prevention and early risk stratification are limited. The vaginal microbiome has been associated with PTB risk, possibly via metabolic or other interactions with its host. Here, we performed untargeted metabolomics on 232 vaginal samples, in which we have previously profiled the microbiota using 16S rRNA gene sequencing. Samples were collected at 20-24 weeks of gestation from women with singleton pregnancies, of which 80 delivered spontaneously before 37 weeks of gestation. We find that the vaginal metabolome correlates with the microbiome and separates into six clusters, three of which are associated with spontaneous preterm birth (sPTB) in Black women. Furthermore, while we identify five metabolites that associate with sPTB, another five associate with sPTB only when stratifying by race. We identify multiple microbial correlations with metabolites associated with sPTB, including intriguing correlations between vaginal bacteria that are considered sub-optimal and metabolites that were enriched in women who delivered at term. We propose that several sPTB-associated metabolites may be exogenous, and investigate another using metabolic models. Notably, we use machine learning models to predict sPTB risk using metabolite levels, weeks to months in advance, with high accuracy. We show that these predictions are more accurate than microbiome-based and maternal covariates-based models. Altogether, our results demonstrate the potential of vaginal metabolites as early biomarkers of sPTB and highlight exogenous exposures as potential risk factors for prematurity.

systems biology↗

bioRxiv2021

Colonization at birth with human CST IV cervicovaginal microbiota alters development and increases neonatal mortality in mice

Profound racial health disparities contribute to maternal-infant morbidity and mortality. An emergent risk factor is the maternal microbiota, whereby compositional alterations impact maternal health and offspring development during pregnancy and beyond. The presence of a nonoptimal CST IV cervicovaginal microbiota, more common in Black and Hispanic women, is associated with increased risk of preterm birth and adverse birth outcomes. Through examination of the biological mechanisms by which vertical transmission of microbiota from mother to offspring influences postnatal development, we found that exposing cesarean delivered mice with CST IV cervicovaginal microbiota from pregnant women produced lasting effects on offspring metabolic, immune, and neural outcomes. We then examined how compounding effects of a typical high-risk, proinflammatory in utero environment, characterized by a maternal obesogenic state and the presence of G. vaginalis, would affect the offspring response to CST IV microbial gut colonization. The resultant developmental immaturity, coupled with an exaggerated immune response induced by exposure to risk-associated maternal microbiota, resulted in a profound increase in neonatal mortality, supporting the critical importance of elucidating the multifactorial biological mechanisms involved in high-risk pregnancies. HighlightsO_LIGut colonization by nonoptimal human cervicovaginal microbiota had sex-specific effects on postnatal development. C_LIO_LIA high-risk in utero environment increased offspring mortality risk. C_LIO_LIHeightened innate immune response associated with mortality risk. C_LIO_LIDevelopmental immaturity, premature birth, and exposure to CST IV contribute to increased offspring mortality risk. C_LI

systems biology↗

bioRxiv2019

VIRGO, a comprehensive non-redundant gene catalog, reveals extensive within community intraspecies diversity in the human vagina

BackgroundAnalysis of metagenomic and metatranscriptomic data is complicated and typically requires extensive computational resources. Leveraging a curated reference database of genes encoded by members of the target microbiome can make these analyses more tractable. Unfortunately, there is no such reference database available for the vaginal microbiome.\n\nResultsIn this study, we assembled a comprehensive human vaginal non-redundant gene catalog (VIRGO) from 264 vaginal metagenomes and 416 genomes of urogenital bacterial isolates. VIRGO includes 0.95 million non-redundant genes compiled from a total of 5.5 million genes belonging to 318 unique bacterial species. We show that VIRGO covers more than 95% of the vaginal bacterial gene content in metagenomes from North American, African, and Chinese women. The gene catalog was extensively functionally annotated from 17 diverse protein databases, and importantly taxonomy was assigned through in silico binning of genes derived from metagenomic assemblies. To further enable focused analyses of individual genes and proteins, we also clustered the non-redundant genes into vaginal orthologous groups (VOG). The gene-centric design of VIRGO and VOG provides an easily accessible tool to comprehensively characterize the structure and function of vaginal metagenome and metatranscriptome datasets. To highlight the utility of VIRGO, we analyzed 1,507 additional vaginal metagenomes, uncovering an as of yet undetected high degree of intraspecies diversity within and across vaginal microbiota.\n\nConclusionsVIRGO offers a convenient reference database and toolkit that will facilitate a more in-depth understanding of the role of vaginal microorganisms in womens health and reproductive outcomes.

genomics↗

bioRxiv2019

Comparative metagenome-assembled genome analysis of Lachnovaginosum genomospecies, formerly known as BVAB1

Bacterial Vaginosis Associated Bacteria 1 (BVAB1) is an uncultured bacterial species found in the human vagina that belongs to the family Lachnospiraceae within the order Clostridiales, and as its name suggests, is often associated with bacterial vaginosis (BV). In reproductive health, BV is of concern due to its associated risk for HIV, Chlamydia trachomatis, and Neisseria gonorrhoeae acquisition as well as preterm birth. BVAB1 has been shown to be associated with BV persistence after metronidazole treatment and increased vaginal inflammation, and to confer a higher risk of puerperal infections. To date, no genome of BVAB1 is available, which has made it difficult to understand its disease-associated features. We present here a comparative analysis of seven metagenome-assembled genomes (MAGs) of BVAB1, derived from the cervicovaginal lavages of seven separate women. One metagenome was sequenced with long-read technology on a PacBio Sequel II instrument while the others were sequenced using the Illumina HiSeq platform. The MAGs were 1.5-1.7 Mb long, encoding an average of 1,499 genes and had >98% average nucleotide identity to each other. CheckM-estimated genome completion was 98.2%. We propose to rename BVAB1 to Lachnovaginosum genomospecies based on a phylogenetic analysis, and provide genomic evidence that this species may metabolize D-lactate, produce trimethylamine (the chemical responsible for BV-associated odor), and be motile. These MAGs will be a valuable resource and will contribute to our understanding of the heterogenous etiologies of bacterial vaginosis.

microbiology↗

bioRxiv2018

Ultra-high throughput multiplexing and sequencing of >500 bp amplicon regions on the Illumina HiSeq 2500 platform

Amplification, sequencing and analysis of the 16S rRNA gene affords characterization of microbial community composition. As this tool has become more popular and amplicon-sequencing applications have grown in the total number of samples, growth in sample multiplexing is becoming necessary while maintaining high sequence quality and sequencing depth. Here, modifications to the Illumina HiSeq 2500 platform are described which produce greater multiplexing capabilities and 300 bp paired-end reads of higher quality than produced by the current Illumina MiSeq platform. To improve the feasibility and flexibility of this method, a 2-Step PCR amplification protocol is also described that allows for targeting of different amplicon regions, thus improving amplification success from low bacterial bioburden samples.\n\nImportanceAmplicon sequencing has become a popular and widespread tool for surveying microbial communities. Lower overall costs associated with high throughput sequencing have made it a widely-adopted approach, especially for projects which necessitate sample multiplexing to eliminate batch effect and reduced time to acquire data. The method for amplicon sequencing on the Illumina HiSeq 2500 platform described here provides improved multiplexing capabilities while simultaneously producing greater quality sequence data and lower per sample cost relative to the Illumina MiSeq platform, without sacrificing amplicon length. To make this method more flexible to various amplicon targeted regions as well as improve amplification from low biomass samples, we also present and validate a 2-Step PCR library preparation method.

microbiology↗

bioRxiv2018

Microbial Biomarkers of Intestinal Barrier Maturation in Preterm Infants

Intestinal barrier immaturity, or \"leaky gut,\" is the proximate cause of susceptibility to necrotizing enterocolitis in preterm neonates. However, the impact of intestinal microbiota development on intestinal mucosal barrier maturation has not been evaluated in this population. In this study, we investigated a longitudinally sampled cohort of 38 preterm infants monitored for intestinal permeability (IP) and fecal microbiota during the first two weeks of life. Rapid decrease in IP indicating intestinal barrier function maturation correlated with significant increase in community diversity. In particular, members of the Clostridiales and Bifidobacterium were highly transcriptionally active, and progressively increasing abundance in Clostridiales was significantly associated with decreased gut permeability. Further, neonatal factors previously identified to promote intestinal barrier maturation, including early exclusive breastmilk feeding and low antibiotic exposure, favor the early colonization of the gut microbiota by members of the Clostridiales, which altogether are associated with improved intestinal barrier function in preterm infants.

microbiology↗

bioRxiv2017

Associations between sexual habits, menstrual hygiene practices, demographics and the vaginal microbiome as revealed by Bayesian network analysis

The vaginal microbiome plays an influential role in several disease states in reproductive age women, including bacterial vaginosis (BV). While demographic characteristics are associated with differences in vaginal microbiome community structure, little is known about the influence of sexual and hygiene habits. Furthermore, associations between the vaginal microbiome and risk symptoms of bacterial vaginosis have not been fully elucidated. Using Bayesian network (BN) analysis of 16S rRNA gene sequence results, demographic and extensive questionnaire data, we describe both novel and previously documented associations between habits of women and their vaginal microbiome. The BN analysis approach shows promise in uncovering complex associations between disparate data types. Our findings based on this approach support published associations between specific microbiome members (e.g., Eggerthella, Gardnerella, Dialister, Sneathia and Ruminococcaceae), the Nugent score (a BV diagnostic) and vaginal pH (a risk symptom of BV). Additionally, we found that several microbiome members were directly connected to other risk symptoms of BV (such as vaginal discharge, odor, itch, irritation, and yeast infection) including L. jensenii, Corynebacteria, and Proteobacteria. No direct connections were found between the Nugent Score and risk symptoms of BV other than pH, indicating that the Nugent Score may not be the most useful criteria for assessment of clinical BV. We also found that demographics (i.e., age, ethnicity, previous pregnancy) were associated with the presence/absence of specific vaginal microbes. The resulting BN revealed several as-yet undocumented associations between birth control usage, menstrual hygiene practices and specific microbiome members. Many of these complex relationships were not identified using common analytical methods, i.e., ordination and PERMANOVA. While these associations require confirmatory follow-up study, our findings strongly suggest that future studies of the vaginal microbiome and vaginal pathologies should include detailed surveys of participants sanitary, sexual and birth control habits, as these can act as confounders in the relationship between the microbiome and disease. Although the BN approach is powerful in revealing complex associations within multidimensional datasets, the need in some cases to discretize the data for use in BN analysis can result in loss of information. Future research is required to alleviate such limitations in constructing BN networks. Large sample sizes are also required in order to allow for the incorporation of a large number of variables (nodes) into the BN, particularly when studying associations between metadata and the microbiome. We believe that this approach is of great value, complementing other methods, to further our understanding of complex associations characteristic of microbiome research.

microbiology↗