Identification of proteoforms of short open reading frame-encoded peptides in Blautia producta under different cultivation conditions.

IMPORTANCE: The identification of short open reading frame-encoded peptides (SEP) and different proteoforms in single cultures of gut microbes offers new insights into a largely neglected part of the microbial proteome landscape. This is of particular importance as SEP provide various predicted functions, such as acting as antimicrobial peptides, maintaining cell homeostasis under stress conditions, or even contributing to the virulence pattern. They are, thus, taking a poorly understood role in structure and function of microbial networks in the human body. A better understanding of SEP in the context of human health requires a precise understanding of the abundance of SEP both in commensal microbes as well as pathogens. For the gut beneficial B. producta, we demonstrate the importance of specific environmental conditions for biosynthesis of SEP expanding previous findings about their role in microbial interactions.

Sorangicin A Is Active against Chlamydia in Cell Culture, Explanted Fallopian Tubes, and Topical In Vivo Treatment.

Current treatment of Chlamydia trachomatis using doxycycline and azithromycin introduces detrimental side effects on the host's microbiota. As a potential alternative treatment, the myxobacterial natural product sorangicin A (SorA) blocks the bacterial RNA polymerase. In this study we analyzed the effectiveness of SorA against C. trachomatis in cell culture, and explanted fallopian tubes and systemic and local treatment in mice, providing also pharmacokinetic data on SorA. Potential side effects of SorA on the vaginal and gut microbiome were assessed in mice and against human-derived Lactobacillus species. SorA showed minimal inhibitory concentrations of 80 ng/mL (normoxia) to 120 ng/mL (hypoxia) against C. trachomatis in vitro and was eradicating C. trachomatis at a concentration of 1 µg/mL from fallopian tubes. In vivo, SorA reduced chlamydial shedding by more than 100-fold within the first days of infection by topical application corresponding with vaginal detection of SorA only upon topical treatment, but not after systemic application. SorA changed gut microbial composition during intraperitoneal application only and did neither alter the vaginal microbiota in mice nor affect growth of human-derived lactobacilli. Additional dose escalations and/or pharmaceutical modifications will be needed to optimize application of SorA and to reach sufficient anti-chlamydial activity in vivo.

Hearing rehabilitation and microbial shift after middle ear surgery with Vibrant Soundbridge in patients with chronic otitis media.

INTRODUCTION: Patients with otitis media (OM) encounter significant functional hearing impairment with conductive, or a combined hearing loss and long-term sequelae involving impaired speech/language development in children, reduced academic achievement and irreversible disorders of middle and inner ear requiring a long time therapy and/or multiple surgeries. In its persistent chronic form, Otitis media (COM) can often only be treated by undergoing ear surgery for hearing restoration. The persistent inflammatory reaction plays a major role, often caused by multi-resistant pathogens in the ear. Herein, we present outcomes of patients implanted with currently the only FDA approved active Middle Ear Implant Vibrant Soundbridge (VSB), suffering from persistent COM. METHODS: The study enrolled 42 patients, treated by performing middle ear (ME) surgery to different extents and implanted with the VSB to various structures in the ME. Included were 17 children and 25 adults that had recurrent and/or persisting OM and significant hearing loss. Preoperative and postoperative patients' audiometric data were evaluated and the benefit with VSB assessed using the Glasgow Benefit Inventory for adults and pediatric cohorts. The microbial spectrum of pathogens was assessed before and after surgery, exploring the colonization of the otopathogens, as well as the intestinal microbiome from individually burdened patients. RESULTS: The mean functional gain is 29.7 dB HL (range from 10 to 56.2 dB HL) with a significant improvement in speech intelligibility in quiet. Following VSB implantation, no significant differences in coupling were observed at low complication rates. Postoperatively patients showed significantly increased benefit with VSB compared to the untreated situation, including less otorrhea, pain, medical visits, and medication intake, with no recurrent OM and significant bacterial shift in otopathogens. The analysis of the intestinal microbiome displayed a high abundance of bacterial strains that might be linked to chronic and persistent inflammation. CONCLUSIONS: Functional ear surgery including rehabilitation with a VSB in patients suffering from COM present to be safe and effective. The successful acceptance accompanied by the improved audiological performance resulted in significant benefit with VSB, with a shift in the ear pathogens and altered microbiome and thus is a great opportunity to be treated.

Delayed Microbial Maturation Durably Exacerbates Th17-driven Asthma in Mice.

Microbial maturation disrupted by early-life dysbiosis has been linked with increased asthma risk and severity; however, the immunological mechanisms underpinning this connection are poorly understood. We sought to understand how delaying microbial maturation drives worsened asthma outcomes later in life and its long-term durability. Drinking water was supplemented with antibiotics on Postnatal Days 10-20. To assess the immediate and long-term effects of delaying microbial maturation on experimental asthma, we initiated house dust mite exposure when bacterial diversity was either at a minimum or had recovered. Airway hyperresponsiveness, histology, pulmonary leukocyte recruitment, flow cytometric analysis of cytokine-producing lymphocytes, and assessment of serum IgG1 (Immunoglobulin G1) and IgE (Immunoglobulin E) concentrations were performed. RT-PCR was used to measure IL-13 (Interleukin 13)-induced gene expression in sequentially sorted mesenchymal, epithelial, endothelial, and leukocyte cell populations from the lung. Delayed microbial maturation increased allergen-driven airway hyperresponsiveness and Th17 frequency compared with allergen-exposed control mice, even when allergen exposure began after bacterial diversity recovered. Blockade of IL-17A (Interleukin 17A) reversed the airway hyperresponsiveness phenotype. In addition, allergen exposure in animals that experienced delayed microbial maturation showed signs of synergistic signaling between IL-13 and IL-17A in the pulmonary mesenchymal compartment. Delaying microbial maturation in neonates promotes the development of more severe asthma by increasing Th17 frequency, even if allergen exposure is initiated weeks after microbial diversity is normalized. In addition, IL-17A-aggravated asthma is associated with increased expression of IL-13-induced genes in mesenchymal, but not epithelial cells.

Improving analysis of the vaginal microbiota of women undergoing assisted reproduction using nanopore sequencing.

PURPOSE: Subclinical alterations of the vaginal microbiome have been described to be associated with female infertility and may serve as predictors for failure of in vitro fertilization treatment. While large prospective studies to delineate the role of microbial composition are warranted, integrating microbiome information into clinical management depends on economical and practical feasibility, specifically on a short duration from sampling to final results. The currently most used method for microbiota analysis is either metagenomics sequencing or amplicon-based microbiota analysis using second-generation methods such as sequencing-by-synthesis approaches (Illumina), which is both expensive and time-consuming. Thus, additional approaches are warranted to accelerate the usability of the microbiome as a marker in clinical praxis. METHODS: Herein, we used a set of ten selected vaginal swabs from women undergoing assisted reproduction, comparing and performing critical optimization of nanopore-based microbiota analysis with the results from MiSeq-based data as a quality reference. RESULTS: The analyzed samples carried varying community compositions, as shown by amplicon-based analysis of the V3V4 region of the bacterial 16S rRNA gene by MiSeq sequencing. Using a stepwise procedure to optimize adaptation, we show that a close approximation of the microbial composition can be achieved within a reduced time frame and at a minimum of costs using nanopore sequencing. CONCLUSIONS: Our work highlights the potential of a nanopore-based methodical setup to support the feasibility of interventional studies and contribute to the development of microbiome-based clinical decision-making in assisted reproduction.

The intracellular proteome of the gut bacterium Bacteroides thetaiotaomicron is widely unaffected by a switch from glucose to sucrose as main carbohydrate source.

Bacteroides thetaiotaomicron is a gram negative bacterium within the human gut microbiome that metabolizes a wide range of dietary and mucosal polysaccharides. Here, we analyze the proteome response of B. thetaiotaomicron cultivated on two different carbon sources, glucose and sucrose. Two quantitative LC-MS based proteomics approaches, encompassing label free quantification and isobaric labeling by tandem mass tags were applied. The results obtained by both workflows were compared with respect to the number of identified and quantified proteins, peptides supporting identification and quantification, sequence coverage, and reproducibility. A total of 1719 and 1696 proteins, respectively, were quantified, covering 35 % of the predicted B. thetaiotaomicron proteome. The data show that B. thetaiotaomicron widely maintains its intracellular proteome upon change of the carbohydrates and that major changes are observed solely in the machinery necessary to make use of the carbon sources provided. With respect to the central role of carbohydrates on gut health these data contribute to the understanding of how different carbohydrates contribute to shape bacterial community in the gut microbiome. All proteomics raw data have been uploaded to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD033704.

Timing of antimicrobial prophylaxis for cesarean section is critical for gut microbiome development in term born infants.

Animal models imply that the perinatal exposure to antibiotics has a substantial impact on microbiome establishment of the offspring. We aimed to evaluate the effect of timing of antimicrobial prophylaxis for cesarean section before versus after cord clamping on gut microbiome composition of term born infants. We performed an exploratory, single center randomized controlled clinical trial. We included forty pregnant women with elective cesarean section at term. The intervention group received single dose intravenous cefuroxime after cord clamping (n = 19), the control group single dose intravenous cefuroxime 30 minutes before skin incision (n = 21). The primary endpoint was microbiome signature of infants and metabolic prediction in the first days of life as determined in meconium samples by 16S rRNA gene sequencing. Secondary endpoints were microbiome composition at one month and 1 year of life. In meconium samples of the intervention group, the genus Staphylococcus pre-dominated. In the control group, the placental cross-over of cefuroxime was confirmed in cord blood. A higher amino acid and nitrogen metabolism as well as increased abundance of the genera Cutibacterium, Corynebacterium and Streptophyta were noted (indicator families: Cytophagaceae, Lactobacilaceae, Oxalobacteraceae). Predictive models of metabolic function revealed higher 2'fucosyllactose utilization in control group samples. In the follow-up visits, a higher abundance of the genus Clostridium was evident in the intervention group. Our exploratory randomized controlled trial suggests that timing of antimicrobial prophylaxis is critical for early microbiome engraftment but not antimicrobial resistance emergence in term born infants.

Culturomics Approaches Expand the Diagnostic Accuracy for Sexually Transmitted Infections.

Sexually transmitted infections (STIs) are a major health concern with clinical manifestations being acknowledged to cause severe reproductive impairment. Research in infectious diseases has been centered around the known major pathogens for decades. However, we have just begun to understand that the microbiota of the female genital tract is of particular importance for disease initiation, infection progression, and pathological outcome. Thus, we are now aware that many poorly described, partially not yet known, or cultured bacteria may pave the way for an infection and/or contribute to disease severity. While sequencing-based methods are an important step in diagnosing STIs, culture-based methods are still the gold-standard method in diagnostic routine, providing the opportunity to distinguish phenotypic traits of bacteria. However, current diagnostic culture routines suffer from several limitations reducing the content of information about vaginal microbiota. A detailed characterization of microbiota-associated factors is needed to assess the impact of single-bacterial isolates from the vaginal community on vaginal health and the containment of STIs. Here we provide current concepts to enable modern culture routines and create new ideas to improve diagnostic approaches with a conjunct usage of bioinformatics. We aim to enable scientists and physicians alike to overcome long-accepted limitations in culturing bacteria of interest to the human health. Eventually, this may improve the quality of culture-based diagnostics, facilitate a research interface, and lead to a broader understanding of the role of vaginal microbiota in reproductive health and STIs.

Impact of First-Line Antimicrobials on Chlamydia trachomatis-Induced Changes in Host Metabolism and Cytokine Production.

Urogenital infections with Chlamydia trachomatis (C. trachomatis) are the most common bacterial sexually transmitted diseases worldwide. As an obligate intracellular bacterium, chlamydial replication and pathogenesis depends on the host metabolic activity. First-line antimicrobials such as doxycycline (DOX) and azithromycin (AZM) have been recommended for the treatment of C. trachomatis infection. However, accumulating evidence suggests that treatment with AZM causes higher rates of treatment failure than DOX. Here, we show that an inferior efficacy of AZM compared to DOX is associated with the metabolic status of host cells. Chlamydial metabolism and infectious progeny of C. trachomatis were suppressed by therapeutic relevant serum concentrations of DOX or AZM. However, treatment with AZM could not suppress host cell metabolic pathways, such as glycolysis and mitochondrial oxidative phosphorylation, which are manipulated by C. trachomatis. The host cell metabolic activity was associated with a significant reactivation of C. trachomatis after removal of AZM treatment, but not after DOX treatment. Furthermore, AZM insufficiently attenuated interleukin (IL)-8 expression upon C. trachomatis infection and higher concentrations of AZM above therapeutic serum concentration were required for effective suppression of IL-8. Our data highlight that AZM is not as efficient as DOX to revert host metabolism in C. trachomatis infection. Furthermore, insufficient treatment with AZM failed to inhibit chlamydial reactivation as well as C. trachomatis induced cytokine responses. Its functional relevance and the impact on disease progression have to be further elucidated in vivo.

Recurrent Urinary Tract Infections: Unraveling the Complicated Environment of Uncomplicated rUTIs.

Urinary tract infections (UTIs) are frequent in humans, affecting the upper and lower urinary tract. Present diagnosis relies on the positive culture of uropathogenic bacteria from urine and clinical markers of inflammation of the urinary tract. The bladder is constantly challenged by adverse environmental stimuli which influence urinary tract physiology, contributing to a dysbiotic environment. Simultaneously, pathogens are primed by environmental stressors such as antibiotics, favoring recurrent UTIs (rUTIs), resulting in chronic illness. Due to different confounders for UTI onset, a greater understanding of the fundamental environmental mechanisms and microbial ecology of the human urinary tract is required. Such advancements could promote the tandem translation of bench and computational studies for precision treatments and clinical management of UTIs. Therefore, there is an urgent need to understand the ecological interactions of the human urogenital microbial communities which precede rUTIs. This review aims to outline the mechanistic aspects of rUTI ecology underlying dysbiosis between both the human microbiome and host physiology which predisposes humans to rUTIs. By assessing the applications of next generation and systems level methods, we also recommend novel approaches to elucidate the systemic consequences of rUTIs which requires an integrated approach for successful treatment. To this end, we will provide an outlook towards the so-called 'uncomplicated environment of UTIs', a holistic and systems view that applies ecological principles to define patient-specific UTIs. This perspective illustrates the need to withdraw from traditional reductionist perspectives in infection biology and instead, a move towards a systems-view revolving around patient-specific pathophysiology during UTIs.

S100A8 and S100A9 Are Important for Postnatal Development of Gut Microbiota and Immune System in Mice and Infants.

BACKGROUND & AIMS: After birth, the immune system matures via interactions with microbes in the gut. The S100 calcium binding proteins S100A8 and S100A9, and their extracellular complex form, S100A8-A9, are found in high amounts in human breast milk. We studied levels of S100A8-A9 in fecal samples (also called fecal calprotectin) from newborns and during infancy, and their effects on development of the intestinal microbiota and mucosal immune system. METHODS: We collected stool samples (n = 517) from full-term (n = 72) and preterm infants (n = 49) at different timepoints over the first year of life (days 1, 3, 10, 30, 90, 180, and 360). We measured levels of S100A8-A9 by enzyme-linked immunosorbent assay and analyzed fecal microbiomes by 16S sRNA gene sequencing. We also obtained small and large intestine biopsies from 8 adults and 10 newborn infants without inflammatory bowel diseases (controls) and 8 infants with necrotizing enterocolitis and measured levels of S100A8 by immunofluorescence microscopy. Children were followed for 2.5 years and anthropometric data and medical information on infections were collected. We performed studies with newborn C57BL/6J wild-type and S100a9-/- mice (which also lack S100A8). Some mice were fed or given intraperitoneal injections of S100A8 or subcutaneous injections of Staphylococcus aureus. Blood and intestine, mesenterial and celiac lymph nodes were collected; cells and cytokines were measured by flow cytometry and studied in cell culture assays. Colon contents from mice were analyzed by culture-based microbiology assays. RESULTS: Loss of S100A8 and S100A9 in mice altered the phenotypes of colonic lamina propria macrophages, compared with wild-type mice. Intestinal tissues from neonatal S100-knockout mice had reduced levels of CX3CR1 protein, and Il10 and Tgfb1 mRNAs, compared with wild-type mice, and fewer T-regulatory cells. S100-knockout mice weighed 21% more than wild-type mice at age 8 weeks and a higher proportion developed fatal sepsis during the neonatal period. S100-knockout mice had alterations in their fecal microbiomes, with higher abundance of Enterobacteriaceae. Feeding mice S100 at birth prevented the expansion of Enterobacteriaceae, increased numbers of T-regulatory cells and levels of CX3CR1 protein and Il10 mRNA in intestine tissues, and reduced body weight and death from neonatal sepsis. Fecal samples from term infants, but not preterm infants, had significantly higher levels of S100A8-A9 during the first 3 months of life than fecal samples from adults; levels decreased to adult levels after weaning. Fecal samples from infants born by cesarean delivery had lower levels of S100A8-A9 than from infants born by vaginal delivery. S100 proteins were expressed by lamina propria macrophages in intestinal tissues from infants, at higher levels than in intestinal tissues from adults. High fecal levels of S100 proteins, from 30 days to 1 year of age, were associated with higher abundance of Actinobacteria and Bifidobacteriaceae, and lower abundance of Gammaproteobacteria-particularly opportunistic Enterobacteriaceae. A low level of S100 proteins in infants' fecal samples associated with development of sepsis and obesity by age 2 years. CONCLUSION: S100A8 and S100A9 regulate development of the intestinal microbiota and immune system in neonates. Nutritional supplementation with these proteins might aide in development of preterm infants and prevent microbiota-associated disorders in later years.

Dissimilarity of Airway and Lung Tissue Microbiota in Smokers undergoing Surgery for Lung Cancer.

Human airways are continuously colonized by microaspiration of microbiota. Less is known about the presence, origin and composition of microbiota in the lung parenchyma. In a study of 13 patients undergoing surgery for peripheral lung cancer microbiota composition was comparatively evaluated in upper airway, lower airway and lung tissue samples using 16S rDNA analysis. Bacterial density decreased stepwise from upper to lower airways and tissue. On a taxonomic level upper and lower airway microbiota were similar whereas lung tissue showed marked dissimilarities compared to lower airways that may reflect different environmental conditions shaping local microbiota and host immunity.

Elaborations on Corallopyronin A as a Novel Treatment Strategy Against Genital Chlamydial Infections.

Ascending Chlamydia trachomatis infection causes functional damage to the fallopian tubes, which may lead to ectopic pregnancy and infertility in women. Treatment failures using the standard regimens of doxycycline and azithromycin have been observed. We tested the polyketide-derived α-pyrone antibiotic Corallopyronin A (CorA) that inhibits the bacterial DNA dependent RNA polymerase and has strong activity against various extracellular and some intracellular bacteria. Extensive testing in cell culture infection models and in an ex vivo human fallopian tube model under different oxygen concentrations was performed to assess the anti-chlamydial efficacy of CorA at physiological conditions. CorA showed high efficacy against C. trachomatis (MICN/H: 0.5 µg/mL for serovar D and L2), C. muridarum (MICN/H: 0.5 µg/mL), and C. pneumoniae (MICN/H: 1 µg/mL) under normoxic (N) and hypoxic (H) conditions. Recoverable inclusion forming units were significantly lower already at 0.25 µg/mL for all tested chlamydiae. CorA at a concentration of 1 µg/mL was also effective against already established C. trachomatis and C. pneumoniae infections (up to 24 h.p.i.) in epithelial cells, while efficacy against C. muridarum was limited to earlier time points. A preliminary study using a C. muridarum genital infection model revealed corresponding limitations in the efficacy. Importantly, in an ex vivo human fallopian tube model, the growth of C. trachomatis was significantly inhibited by CorA at concentrations of 1-2 µg/mL under normoxic and hypoxic conditions. The overall high efficacies of CorA against C. trachomatis in cell culture and an ex vivo human fallopian tube model under physiological oxygen concentrations qualifies this drug as a candidate that should be further investigated.

Dietary ursolic acid improves health span and life span in male Drosophila melanogaster.

The health and life span of Drosophila melanogaster are partly determined by intestinal barrier integrity, metabolic rate as well as stress response and the expression of longevity-associated genes, depending on genetic and dietary factors. Ursolic acid (UA) is a naturally occurring triterpenoid exhibiting potential antimicrobial, anti-inflammatory, and antiobesity activity and counteracting age-related deficits in muscle strength. In this study, UA was dietarily administered to w1118 D. melanogaster which significantly elongated the health and life span of males. Spargel (srl) is the Drosophila orthologue of mammalian peroxisome proliferator-activated receptor-gamma coactivator 1 α(PGC1α), an important regulator of energy homeostasis and mitochondrial function. Our results indicate that the health-promoting effect of UA, demonstrated by a significant increase in climbing activity, occurs via an upregulation of srl expression leading to a metabolic shift in the fly without reducing fecundity or gut integrity. Moreover, UA affected the flies' microbiota in a manner that contributed to life span extension. Srl expression and microbiota both seem to be affected by UA, as we determined by using srl-mutant and axenic flies. © 2018 BioFactors, 45(2):169-186, 2019.

Lithocholic Acid Improves the Survival of Drosophila Melanogaster.

SCOPE: Primary bile acids are produced in the liver, whereas secondary bile acids, such as lithocholic acid (LCA), are generated by gut bacteria from primary bile acids that escape the ileal absorption. Besides their well-known function as detergents in lipid digestion, bile acids are important signaling molecules mediating effects on the host's metabolism. METHODS AND RESULTS: Fruit flies (Drosophila melanogaster) are supplemented with 50 µmol L-1 LCA either for 30 days or throughout their lifetime. LCA supplementation results in a significant induction of the mean (+12 days), median (+10 days), and maximum lifespan (+ 11 days) in comparison to untreated control flies. This lifespan extension is accompanied by an induction of spargel (srl), the fly homolog of mammalian PPAR-γ co-activator 1α (PGC1α). In wild-type flies, the administration of antibiotics abrogates both the LCA-mediated lifespan induction as well as the upregulation of srl. CONCLUSION: It is shown that the secondary bile acid LCA significantly induces the mean, the median, and the maximum survival in D. melanogaster. Our data suggest that besides an upregulation of the PGC1α-homolog srl, unidentified alterations in the structure or metabolism of the gut microbiota contribute to the longevity effect mediated by LCA.

Selection of validated hypervariable regions is crucial in 16S-based microbiota studies of the female genital tract.

Next-generation sequencing-based methods are extensively applied in studies of the human microbiota using partial 16 S rRNA gene amplicons. However, they carry drawbacks that are critical to consider when interpreting results, including differences in outcome based on the hypervariable region(s) used. Here, we show that primers spanning the V3/V4 region identify a greater number of taxa in the vaginal microbiota than those spanning the V1/V2 region. In particular, taxa such as Gardnerella vaginalis, Bifidobacterium bifidum and Chlamydia trachomatis, all species that influence vaginal health and disease, are not represented in V1/V2-based community profiles. Accordingly, missing or underestimating the frequency of these species overestimates the abundance of other taxa and fails to correctly assess the bacterial diversity in the urogenital tract. We elaborate that covering these taxa using the V3/V4 region leads to profound changes in the assignment of community state types. Altogether, we show that the choice of primers used for studying the vaginal microbiota has deep implications on the biological evaluation of the results.

Microbiota changes impact on sexually transmitted infections and the development of pelvic inflammatory disease.

The integrity of the human urogenital microbiome is crucial for women's health and well-being. An imbalance of the urogenital microbiota increases the risk for sexually transmitted infections. In this review, we discuss the microbiota composition of the female urogenital tract and its role in protecting from sexually transmitted infections and the emergence of pelvic inflammatory disease.

Microbiota-based analysis reveals specific bacterial traits and a novel strategy for the diagnosis of infectious infertility.

Tubal factor infertility (TFI) accounts for more than 30% of the cases of female infertility and mostly resides from an inflammatory process triggered by an infection. Clinical appearances largely differ, and very often infections are not recognized or remain completely asymptomatic over time. Here, we characterized the microbial pattern in females diagnosed with infectious infertility (ININF) in comparison to females with non-infectious infertility (nININF), female sex workers (FSW) and healthy controls (fertile). Females diagnosed with infectious infertility differed significantly in the seroprevalence of IgG antibodies against the C. trachomatis proteins MOMP, OMP2, CPAF and HSP60 when compared to fertile females. Microbiota analysis using 16S amplicon sequencing of cervical swabs revealed significant differences between ININF and fertile controls in the relative read count of Gardnerella (10.08% vs. 5.43%). Alpha diversity varies among groups, which are characterized by community state types including Lactobacillus-dominated communities in fertile females, an increase in diversity in all the other groups and Gardnerella-dominated communities occurring more often in ININF. While all single parameters did not allow predicting infections as the cause of infertility, including C. trachomatis IgG/IgA status together with 16S rRNA gene analysis of the ten most frequent taxa a total of 93.8% of the females were correctly classified. Further studies are needed to unravel the impact of the cervical microbiota in the pathogenesis of infectious infertility and its potential for identifying females at risk earlier in life.