Vaginal Strains Research Articles

Recommendations for strain elastography of the uterine cervix

PurposeConventional vaginal strain ultrasound elastography, not based on shear-wave elastography imaging, can assess the biomechanical properties of the uterine cervix. This assessment may inform the risks of preterm birth and failed induction of labor. However, there is considerable variation in the approaches to strain elastography, including the placement of the region of interest (ROI). Therefore, our aim was to provide recommendations for cervical elastography.MethodsWe conducted a literature review on (1) elastography principles, and (2) the cervical anatomy. Subsequently, we performed elastography scanning using a Voluson™ E10 Expert scanner with the BT18 software of (3) polyacrylamide hydrogel simulators, and (4) pregnant women.ResultsIncreasing the distance between the ROI and probe led to a decrease in the obtained strain value; a 53% decrease was observed at 17.5 mm. Similarly, an increased angle between the ROI and probe-centerline resulted in a 59% decrease for 40° angle. Interposition of soft tissue (e.g., cervical canal) between the ROI and the probe induced an artifact with values from the posterior lip being 54% lower than those from the anterior lip, even after adjusting for probe-ROI distance. Equipment and the recording conductance significantly influenced the results.ConclusionOur findings inform recommendations for future studies on strain cervical elastography.

The phenotypic and genotypic antibiotic susceptibility of vaginal Lactobacillus with potential probiotic properties isolated from healthy women in northern Iran

Background and Objectives: This study assesses the antibiotic susceptibility of vaginal Lactobacillus strains and provides data for determining the prevalence of certain antibiotic resistance genes in the new strains of lactobacilli serving as probiot- ics and selected from healthy women in northern Iran. Materials and Methods: One hundred premenopausal non-pregnant women in the reproductive age range of 22-50 years participated in this study. The potential probiotic vaginal lactobacilli used in the study included Lactobacillus crispatus (34.2%), Lactobacillus gasseri (26.3%), Lactobacillus johnsonii (10.5%), Lactobacillus acidophilus (15.7%) and Lactoba- cillus jensenii (13.1%). The phenotypic antibiotic susceptibility of the strains was determined by E test and DNA extraction and PCR were performed to examine the antibiotic resistance genes. Results: 38 potential probiotic vaginal lactobacilli were isolated. All the strains of lactobacilli were resistant to metronida- zole and trimethoprim/sulfamethoxazole and all of the strains were susceptible to ampicillin and chloramphenicol antibiotics. The results showed that ermB, ermC, and ermA genes were observed in the strains of Lactobacillus acidophilus. Metronida- zole resistance (nim) gene was also found in one strain of Lactobacillus crispatus and Lactobacillus johnsonii. The amino- glycoside resistance (aac6'-aph2") gene was observed in 8% of the strains. Also, tetM, tetK and tetW genes were found in more than 80% of the Lactobacillus strains. Conclusion: The antimicrobial susceptibility of vaginal lactobacilli is an important criterion for establishing whether or not the organism is a probiotic. A high level of resistance to clinical antibiotics, such as metronidazole and aminoglycosides, was demonstrated. Antibiotic resistant genes also appeared widely in vaginal lactobacilli.

Characterization of Vaginal Lactobacilli with Potential Probiotic Properties Isolated from Healthy Women in Northern Iran.

Vaginal lactobacilli protect against bacterial vaginosis and vaginal candidiasis. They may have probiotic properties and help maintain the balance and health of the vaginal ecosystem while the loss of these bacteria predisposes females to urinary and genital infections. The aim of this study was to investigate the probiotic potential of vaginal Lactobacillus among healthy females in northern Iran. The Lactobacillus strains were isolated from vaginal samples and were identified by sequencing of the 16S rRNA fragment. Functional properties such as tolerance to low pH, H2O2 production, adherence ability to Hela cells and antagonistic activity against Candida albicans was examined. A total of 38 vaginal lactobacilli strains from five species, including Lactobacillus crispatus (n = 13), Lactobacillus gasseri (n = 10), Lactobacillus acidophilus (n = 6), Lactobacillus jensenii (n = 5) and Lactobacillus johnsonii (n = 4), were identified. All of the species showed significant tolerance to low pH over 24h (p < 0.001). The best adherence ability to Hela cells was seen in Lactobacillus gasseri strains. Nearly 17 of the strains had higher anti-candida activity compared to the other strains. According to the findings, four lactobacilli strains isolated in the vaginal samples of healthy Iranian women had the best probiotic potential.

In Vitro Interaction between Mycoplasma agalactiae and Small Ruminants' Endogenous Bacterial Strains of Enterococcus spp. and Coagulase-Negative Staphylococcus.

Recently, an antimicrobial effect on Mycoplasma agalactiae (Ma), the main etiological agent of contagious agalactia (CA), was reported in vitro with strains of Enterococcus spp. from ovine and caprine milk. The aim of this work was to evaluate the interaction of Ma with the same Enterococcus spp. isolated from other anatomical locations (vagina) and other bacterial populations present in milk, such as coagulase-negative staphylococci (CNS). The vaginal Enterococcus strains and the raw milk CNS were isolated from sheep and goats. Experimental in vitro conditions were prepared to assess the growth of Ma with and without the presence of these strains. The selected vaginal strains were identified as Enterococcus (E.) hirae and E. mundtii, and the strains of CNS were identified as Staphylococcus petrasii. Different interactions of Ma with ovine and caprine wild vaginal strains of Enterococcus and dairy strains of CNS are described for the first time: Ma can grow exponentially during 15 h with the selected strains, although with certain strains, its optimal growth can be negatively affected (p < 0.05). The colonization and/or excretion of Ma could, therefore, be influenced by certain endogenous bacterial strains. Our results increase the knowledge about possible bacterial ecology dynamics surrounding CA.

Vaginal prevention of Candida albicans: synergistic effect of lactobacilli and mannan oligosaccharides (MOS)

Vulvovaginal candidiasis (VVC) affects approximately 30–50% of women at least once during their lifetime, causing uncomfortable symptoms and limitations in their daily quality of life. Antifungal therapy is not very effective, does not prevent recurrencies and usually causes side effects. Therefore, alternative therapies are urgently needed. The goal of this work was to investigate the potential benefits of using mannan oligosaccharides (MOS) extracts together with a Lactobacillus sp. pool, composed by the most significant species present in the vaginal environment, to prevent infections by Candida albicans. Microbial growth of isolated strains of the main vaginal lactobacilli and Candida strains was assessed in the presence of MOS, to screen their impact upon growth. A pool of the lactobacilli was then tested against C. albicans in competition and prophylaxis studies; bacterial and yeast cell numbers were quantified in specific time points, and the above-mentioned studies were assessed in simulated vaginal fluid (SVF). Finally, adhesion to vaginal epithelial cells (HeLa) was also evaluated, once again resorting to simultaneous exposure (competition) or prophylaxis assays, aiming to measure the effect of MOS presence in pathogen adherence. Results demonstrated that MOS extracts have potential to prevent vaginal candidiasis in synergy with vaginal lactobacilli, with improved results than those obtained when using lactobacilli alone.Key pointsPotential benefits of MOS extracts with vaginal lactobacilli to prevent C. albicans infections.MOS impacts on growth of vaginal lactobacilli pool and C. albicans in SVF.MOS extracts in synergy with L. crispatus inhibit C. albicans adhesion in HeLa cells.Graphical

Limosilactobacillus vaginalis Exerts Bifidogenic Effects: A Novel Postbiotic Strategy for Infant Prebiotic Supplementation.

Infant microbiota shaping strictly influences newborns' well-being and long-term health, and babies born by cesarean-section and formula-fed generally show low microbial gut diversity and are more prone to develop various disorders. The supplementation with beneficial microbes of vaginal origin or derivatives (postbiotics, including heat-inactivated cells) represents a valid strategy to drive the correct gut microbiota shaping. Here, we explored for the first time the bifidogenic activity of a heat-killed vaginal strain (Limosilactobacillus vaginalis BC17), in addition to the assessment of its safety. L. vaginalis BC17 whole genome was sequenced by Nanopore technology and highlighted the absence of antibiotic resistance genes and virulence factors, indicating the strain safety profile for human health. MIC values confirmed that L. vaginalis BC17 is susceptible to widely employed antibiotics. Heat-killed BC17 cells significantly enhanced the planktonic growth of Bifidobacterium spp. For the first time, stimulating effects were observed also toward biofilm formation of bifidobacteria and their pre-formed biofilms. Conversely, heat-killed BC17 cells exerted antibacterial and anti-biofilms activities against Gram-positive and Gram-negative pathogens. Lyophilized heat-killed BC17 cells were formulated in a sunflower oil suspension (1010 heat-killed cell/g) intended for infant oral intake. This possessed optimal technological (i.e., re-dispersibility and stability) and functional properties (i.e., bifidogenic activity) that were maintained even after pre-digestion in acidic conditions.

Comparative pangenomic analysis of predominant human vaginal lactobacilli strains towards population-specific adaptation: understanding the role in sustaining a balanced and healthy vaginal microenvironment

The vaginal microenvironment of healthy women has a predominance of Lactobacillus crispatus, L. iners, L. gasseri, and L. jensenii. The genomic repertoire of the strains of each of the species associated with the key attributes thereby regulating a healthy vaginal environment needs a substantial understanding.We studied all available human strains of the four lactobacilli across different countries, isolated from vaginal and urinal sources through phylogenetic and pangenomic approaches. The findings showed that L. iners has the highest retention of core genes, and L. crispatus has more gene gain in the evolutionary stratum. Interestingly, L. gasseri and L. jensenii demonstrated major population-specific gene-cluster gain/loss associated with bacteriocin synthesis, iron chelating, adherence, zinc and ATP binding proteins, and hydrolase activity. Gene ontology enrichment analysis revealed that L. crispatus strains showed greater enrichment of functions related to plasma membrane integrity, biosurfactant, hydrogen peroxide synthesis, and iron sequestration as an ancestral derived core function, while bacteriocin and organic acid biosynthesis are strain-specific accessory enriched functions. L. jensenii showed greater enrichment of functions related to adherence, aggregation, and exopolysaccharide synthesis. Notably, the key functionalities are heterogeneously enriched in some specific strains of L. iners and L. gasseri.This study shed light on the genomic features and their variability that provides advantageous attributes to predominant vaginal Lactobacillus species maintaining vaginal homeostasis. These findings evoke the need to consider region-specific candidate strains of Lactobacillus to formulate prophylactic measures against vaginal dysbiosis for women’s health.

Birth Mode Does Not Determine the Presence of Shared Bacterial Strains between the Maternal Vaginal Microbiome and the Infant Stool Microbiome.

Dysbiosis of the neonatal gut microbiome during early life has been suggested as the missing link that may explain higher rates of certain diseases in caesarean section-delivered infants. Many studies report delivery mode-related dysbiosis in infants due to a lack of maternal vaginal microbiome exposure, prompting interventions to correct the neonatal gut microbiome by transferring these missing microbes after caesarean delivery. The maternal vaginal microbiome is among the first microbial exposures that many infants experience, yet little is known about the extent of direct transmission of maternal vaginal microbes. As part of the Maternal Microbiome Legacy Project, we aimed to determine if maternal vaginal bacteria are vertically transmitted to infants. We employed cpn60 microbiome profiling, culture-based screening, molecular strain typing, and whole-genome sequencing to determine whether identical maternal vaginal strains were present in infant stool microbiomes. We identified identical cpn60 sequence variants in both halves of maternal-infant dyads in 204 of 585 Canadian women and their newborn infants (38.9%). The same species of Bifidobacterium and Enterococcus were cultured from maternal and corresponding infant samples in 33 and 13 of these mother-infant dyads, respectively. Pulsed-field gel electrophoresis and whole-genome sequencing determined that near-identical strains were detected in these dyads irrespective of delivery mode, indicating an alternative source in cases of caesarean delivery. Overall, we demonstrated that vertical transmission of maternal vaginal microbiota is likely limited and that transmission from other maternal body sites, such as the gut and breast milk, may compensate for the lack of maternal vaginal microbiome exposure during caesarean delivery. IMPORTANCE The importance of the gut microbiome in human health and disease is widely recognized, and there has been a growing appreciation that alterations in gut microbiome composition during a "critical window" of development may impact health in later life. Attempts to correct gut microbiome dysbiosis related to birth mode are underpinned by the assumption that the lack of exposure to maternal vaginal microbes during caesarean delivery is responsible for dysbiosis. Here, we demonstrate that there is limited transmission of the maternal vaginal microbiome to the neonatal gut, even in cases of vaginal delivery. Furthermore, the presence of identical strains shared between mothers and infants in early life, even in cases of caesarean delivery, highlights compensatory microbial exposures and sources for the neonatal stool microbiome other than the maternal vagina.

Inhibitory effects of vaginal Lactobacilli on Candida albicans growth, hyphal formation, biofilm development, and epithelial cell adhesion.

Antifungal agents are not always efficient in resolving vulvovaginal candidiasis (VVC), a common genital infection caused by the overgrowth of Candida spp., including Candida albicans, or in preventing recurrent infections. Although lactobacilli (which are dominant microorganisms constituting healthy human vaginal microbiota) are important barriers against VVC, the Lactobacillus metabolite concentration needed to suppress VVC is unknown. We quantitatively evaluated Lactobacillus metabolite concentrations to determine their effect on Candida spp., including 27 vaginal strains of Lactobacillus crispatus, L. jensenii, L. gasseri, Lacticaseibacillus rhamnosus, and Limosilactobacillus vaginalis, with inhibitory abilities against biofilms of C. albicans clinical isolates. Lactobacillus culture supernatants suppressed viable fungi by approximately 24%-92% relative to preformed C. albicans biofilms; however, their suppression differed among strains and not species. A moderate negative correlation was found between Lactobacillus lactate production and biofilm formation, but no correlation was observed between hydrogen peroxide production and biofilm formation. Both lactate and hydrogen peroxide were required to suppress C. albicans planktonic cell growth. Lactobacillus strains that significantly inhibited biofilm formation in culture supernatant also inhibited C. albicans adhesion to epithelial cells in an actual live bacterial adhesion competition test. Healthy human microflora and their metabolites may play important roles in the development of new antifungal agent against C. albicans-induced VVC.

In vitro study to assess modulation of Candida biofilm by Escherichia coli from vaginal strains

BackgroundVulvovaginal candidiasis (VVC) is caused by biofilm formation and epithelial invasion. In addition, Escherichia coli (EC) can establish a vaginal intracellular reservoir modulating Candida spp. biofilm production. We aimed to analyze the behavior of Candida albicans (CA) and EC biofilm both in single cultures and in co-cultures. MethodsWe prospectively collected CA and EC isolates from vaginal swabs over 6 months. We selected positive cultures with both CA and EC (cases) and a comparator group with either CA or EC (controls). We analyzed overall biomass production and metabolic activity in single cultures and in co-cultures based on staining assays, confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM) to assess biofilm occupation. We also analyzed clinical manifestations. ResultsWe cultured 455 samples, 16 (3.5%) of which had CA and EC (cases); only CA or EC (controls) was detected, respectively, in 72 (15.8%) and 98 (21.5%). Biomass production and metabolic activity were significantly more pronounced in co-cultures in both groups. CLSM and SEM, on the other hand, showed the biofilm of each species to be significantly reduced when they were cultured together, with higher values in CA (percentage biofilm reduction: CA, 95.8% vs. EC, 36.2%, p < 0.001). There were no clinically significant differences between co-infected patients and patients infected only by C. albicans. ConclusionOurs is the first study assessing co-cultures of CA and EC in a large collection of samples. We observed that coinfection of CA and EC was unusual (3.5%) and promoted high biomass, whereas microscopy enabled us to detect a reduction in biofilm production when microorganisms were co-cultured. No differences in symptoms were observed.

Development and characterization of fermented soy beverages containing encapsulated or non-encapsulated vaginal probiotics

Human microbial niches such as the healthy vagina, are recently emerging as “unconventional” sources of candidate probiotics capable of preventing from different vaginal diseases. These microorganisms could be provided as oral preparations since they can reach the vaginal niche passing through the gastrointestinal tract. However, their use in food would be challenging. The aim of this work was to develop and characterize fermented soy beverages with encapsulated and non-encapsulated vaginal lactobacilli, namely Lactobacillus crispatus BC4 and Lactobacillus gasseri BC9, as future dietary strategies for vaginal dysbiosis. The viability of vaginal strains remained stable at 7 log CFU/mL of product during the entire 28 days of storage, despite the use of encapsulated or non-encapsulated bacteria. Samples containing encapsulated bacteria, especially E-BC4+BC9, showed higher Water Holding Capacity (62.29%), lactic acid content (1.43%), and a remarkable antagonistic activity against enteropathogens. Moreover, encapsulation protected the strains from simulated GIT conditions (>1 Log) but reduced the acceptability of the final products. Overall, strain BC4 and BC9, alone or in mix, demonstrated to be promising co-starter cultures providing a characteristic flavor (pleasant smell and taste) and aroma (lower hexanal, benzaldehyde and higher diacetyl, and 2,3-pentanedione, compared to control) to the fermented soy beverages.

Limosilactobacillus reuteri 29A Cell-Free Supernatant Antibiofilm and Antagonistic Effects in Murine Model of Vulvovaginal Candidiasis.

Vaginal dysbiosis advocates burgeoning of devious human vaginal pathobionts like Candida species that possess multiple virulence properties and metabolic flexibility to cause infections. Inevitably, antifungal resistance may emerge due to their innate nature (e.g., biofilm formation), which assists in their virulence as well as the formation of persister cells after dispersal. In consequence, the phenomenon of biofilm involvement in vulvovaginal candidiasis (VVC) and its recurrence is becoming paramount. Lactic acid bacteria and their derivatives have proven to be hostile to Candida species. Here, we throw more light on the potency of the derivatives, i.e., cell-free supernatant (CFS) produced by an indigenously isolated vaginal Lactobacillus strain, Limosilactobacillus reuteri 29A. In the present study, we investigated the antibiofilm and antagonistic effects of L. reuteri 29A CFS, against biofilms of Candida species and in murine model of vulvovaginal candidiasis. In our in vitro biofilm study, the CFS disrupted and inhibited preformed biofilms of C. albicans and C. glabrata. Scanning electron microscopy displayed the destruction of preformed biofilms and impediment of C. albicans morphogenesis by the CFS. Gas chromatography-mass spectrometry analysis showed multiple key compounds that may act singly or synergistically. In vivo, the CFS showed no collateral damage to uninfected mice; the integrity of infected vaginal tissues was restored by the administration of the CFS as seen from the cytological, histopathological, and electron microscopical analyses. The results of this study document the potential use of CFS as an adjuvant or prophylactic option in addressing vaginal fungal infections.

484. Vaginal Candida albicans: High Frequency of In-vitro Fluconazole Resistance in a Select Clinical Population.

Abstract Background In vitro fluconazole resistance in vaginal C. albicans has rarely been reported in the U.S. Little is known about the characteristics of patients who demonstrate fluconazole resistance vs. sensitivity, or how likely resistant strains are to persist over time. Hence we sought to describe patterns of fluconazole resistance in clinician ordered vaginal cultures positive for C. albicans at our center. We compared patient characteristics between those with fluconazole sensitive versus resistant isolates. Methods We conducted a chart review of patients with clinician ordered vaginal cultures positive for C. albicans undergoing fluconazole susceptibility testing in our medical center’s clinical mycology lab from January 2017 to April 2021. Patient characteristics were compared using chi-square and ANOVA tests and correlated to fluconazole resistance with logistic regression models. Results Of N=92 patients with vaginal C. albicans, 3.3% were sensitive dose-dependent (SDD: minimal inhibitory concentration (MIC) =4) to fluconazole, and 30.4% were resistant (R: MIC &amp;gt; =8). Amongst those with at least 2 previous episodes of vulvovaginal candidiasis (VVC) in the past 6 months (N=45), 50% had R and 6.5% had SDD isolates. Compared to those with sensitive (S: MIC &amp;lt; =2) isolates, patients with SDD or R were younger and more likely to have the following characteristics: Black race, bacterial vaginosis (BV) in the past year, high or multi-dose azole treatment, fluconazole suppressive therapy in the past 6 months, and more VVC and BV episodes in the previous 6 months (Table 1). Of N=10 patients who had a follow-up culture with initial R isolates, 28.6% had a sensitive C. albicans vaginal isolate. Conclusion The frequency of C. albicans vaginal isolates with in vitro resistance to fluconazole in this study was high. Our findings highlight the importance of obtaining cultures and sensitivity testing in patients with recurrent or refractory VVC. However, resistant vaginal strains may not necessarily persist over time. Future, larger studies on those with recurrent VVC are needed to verify factors associated with fluconazole resistance, how frequently azole-resistant strains persist over time, and how well in vitro resistance correlates to in vivo treatment response. Disclosures Susan Tuddenham, MD, MPH, Luca Biologics: Advisor/Consultant|Medscape/WebMD: Honoraria|UpToDate: Royalties.

Persistence and In Vivo Evolution of Vaginal Bacterial Strains over a Multiyear Time Period.

It is not clear whether the bacterial strains that comprise our microbiota are mostly long-term colonizers or transient residents. Studies have demonstrated decades-long persistence of bacterial strains within the gut, but persistence at other body sites has yet to be determined. The vaginal microbiota (VMB) is often dominated by Lactobacillus, although it is also commonly comprised of a more diverse set of other facultative and obligate anaerobes. Longitudinal studies have demonstrated that these communities can be stable over several menstrual cycles or can fluctuate temporally in species composition. We sought to determine whether the bacterial strains that comprise the VMB were capable of persisting over longer time periods. We performed shotgun metagenomics on paired samples from 10 participants collected 1 and 2 years apart. The resulting sequences were de novo assembled and binned into high-quality metagenome assembled genomes. Persistent strains were identified based on the sequence similarity between the genomes present at the two time points and were found in the VMB of six of the participants, three of which had multiple persistent strains. The VMB of the remaining four participants was similar in species composition at the two time points but was comprised of different strains. For the persistent strains, we were able to identify the mutations that were fixed in the populations over the observed time period, giving insight into the evolution of these bacteria. These results indicate that bacterial strains can persist in the vagina for extended periods of time, providing an opportunity for them to evolve in the host microenvironment. IMPORTANCE The stability of strains within the vaginal microbiota is largely uncharacterized. Should these strains be capable of persisting for extended periods of time, they could evolve within their host in response to selective pressures exerted by the host or by other members of the community. Here, we present preliminary findings demonstrating that bacterial strains can persist in the vagina for at least 1 year. We further characterized in vivo evolution of the persistent strains. Several participants were also found to not have persistent strains, despite having a vaginal microbiota (VMB) with similar species composition at the two time points. Our observations motivate future studies that collect samples from more participants, at more time points, and over even longer periods of time. Understanding which strains persist, what factors drive their persistence, and what selective pressures they face will inform the development and delivery of rationally designed live biotherapeutics for the vagina.

Lactobacilli extracellular vesicles: potential postbiotics to support the vaginal microbiota homeostasis

BackgroundLactobacillus species dominate the vaginal microflora performing a first-line defense against vaginal infections. Extracellular vesicles (EVs) released by lactobacilli are considered mediators of their beneficial effects affecting cellular communication, homeostasis, microbial balance, and host immune system pathways. Up to now, very little is known about the role played by Lactobacillus EVs in the vaginal microenvironment, and mechanisms of action remain poorly understood.ResultsHere, we hypothesized that EVs can mediate lactobacilli beneficial effects to the host by modulating the vaginal microbiota colonization. We recovered and characterized EVs produced by two vaginal strains, namely Lactobacillus crispatus BC5 and Lactobacillus gasseri BC12. EVs were isolated by ultracentrifugation and physically characterized by Nanoparticle Tracking Analysis (NTA) and Dynamic Light Scattering (DLS). EVs protein and nucleic acids (DNA and RNA) content was also evaluated. We explored the role of EVs on bacterial adhesion and colonization, using a cervical cell line (HeLa) as an in vitro model. Specifically, we evaluated the effect of EVs on the adhesion of both vaginal beneficial lactobacilli and opportunistic pathogens (i.e., Escherichia coli, Staphylococcus aureus, Streptococcus agalactiae, and Enterococcus faecalis). We demonstrated that EVs from L. crispatus BC5 and L. gasseri BC12 significantly enhanced the cellular adhesion of all tested lactobacilli, reaching the maximum stimulation effect on strains belonging to L. crispatus species (335% and 269% of average adhesion, respectively). At the same time, EVs reduced the adhesion of all tested pathogens, being EVs from L. gasseri BC12 the most efficient.ConclusionsOur observations suggest for the first time that EVs released by symbiotic Lactobacillus strains favor healthy vaginal homeostasis by supporting the colonization of beneficial species and preventing pathogens attachment. This study reinforces the concept of EVs as valid postbiotics and opens the perspective of developing postbiotics from vaginal strains to maintain microbiota homeostasis and promote women’s health.

Functional and safety characterization of beneficial vaginal lactic acid bacteria for the design of vaginal hygiene products.

The aim was to complement the evaluation of functional and safety properties of beneficial vaginal lactic acid bacteria to select the most adequate strains as potential probiotics for their inclusion in a vaginal probiotic formula. Twenty-four beneficial vaginal lactic acid bacteria strains previously isolated from healthy women were characterized by applying phenotypical and genetic techniques. The biofilm formation and exopolysaccharide production by the different strains showed a strong influence of the growth medium on the expression of these properties. The evaluated strains evidenced different antibiotic susceptibility patterns by phenotypic tests, while the resistance genes were not always correlated with the phenotypic profiles. None of the evaluated strains showed lecithinase, gelatinase, or β-hemolytic activity. Based on the results obtained, Lactobacillus gasseri CRL 1320 and Lacticaseibacillus rhamnosus CRL 1332 were selected to advance in the design of the products. A wide variability in the properties expressed by beneficial vaginal lactic acid bacteria strains were shown, evidencing a strain-specific and independent behaviour of the species/metabolic group where classified. The selection criteria applied provided a valuable tool for the selection of the best vaginal lactobacilli strains for their inclusion in formulas for vaginal application. Since probiotics are an important tool for improving human health, the designed formula are an excellent alternative to reconstitute the vaginal microbiota and prevent/treat urogenital tract infections in women. The results obtained contribute to deepen the characterization of homologous beneficial vaginal lactic acid bacteria and advance in the design of vaginal probiotic products.

Vaginal Lactobacillus Impair Candida Dimorphic Switching and Biofilm Formation.

Lactobacillus spp. generally dominate the vaginal microbiota and prevent pathogen adhesion and overgrowth, including Candida spp., by various mechanisms. Although Candida spp. can be commensal, in certain conditions they can become pathogenic, causing vulvovaginal candidiasis. The insurgence of candidiasis is related to the expression of Candida virulence factors, including morphologic switching and biofilm formation. Germ tubes, pseudohyphae, and hyphae promote Candida tissue invasion, biofilms increase persistence and are often resistant to antifungals and host immune response. Here, we explored the inhibitory activity of vaginal Lactobacillus strains belonging to Lactobacillus crispatus, Lactobacillus gasseri, Limosilactobacillus vaginalis, and Lactiplantibacillus plantarum species towards Candida virulence factors. With the aim to investigate the interrelation between mode of growth and functionality, supernatants were collected from lactobacilli planktonic cultures and, for the first time, from adherent ones, and were evaluated towards Candida dimorphic switching and biofilm. Candida biofilms were analyzed by multiple methodologies, i.e., crystal violet staining, MTT assay, and confocal microscopy. Lactobacillus supernatants reduce Candida switching and biofilm formation. Importantly, L. crispatus supernatants showed the best profile of virulence suppression, especially when grown in adherence. These results highlight the role of such species as a hallmark of vaginal eubiosis and prompt its employment in new probiotics for women’s health.

Genotypic Characterization and Biofilm Production of Group B Streptococcus Strains Isolated from Bone and Joint Infections.

ABSTRACTBone and joint infections (BJI) represent the second cause of invasive Group B Streptococcus (GBS) infections. Biofilm formation plays a major role in BJI. This study’s aim was to analyze the genetic features and biofilm production of GBS strains. In six French laboratories, 77 GBS strains isolated from BJI and 57 strains from vaginal human colonization (Hcol) were characterized and compared by Multi-Locus Sequence Typing (MLST). PCR was used to search for the adhesins (bsaB, lmb, scpB, fbsA, fbsB, hvgA, bibA, bca, srr-1, and srr-2) and Pilus Islands (PI) related genes (PI-1, PI-2a, PI-2b). Biofilm production was studied by crystal violet assay. Strains were categorized into three groups, based on Specific Biofilm Formation (SBF) values defined as: weak, moderate, or strong producers. Molecular study revealed three major clonal complexes (CC) in BJI strains: CC1 (42%), CC23 (22%) and CC10 (14%). Several associations between CC and adhesin/pili were identified: CC1 with srr2, PI-1 + 2a; CC10 with srr-1, bca, PI-1 + 2a; CC17 with fbsB, hvgA, srr-2, PI-1+PI-2b; CC19 with bibA, srr-1, PI-1 + 2a; CC23 with fbsB, bibA, srr-1, PI-2a. The biofilm production was significantly different according to CC, adhesins and pili gene detection. CC10, CC23 and strains harboring fbsB produce more biofilm than CC1, PI-1 + 2a (independently). Finally, SBF values were significantly stronger for Hcol strains rather than for BJI strains (76% versus 40%). This study revealed that Hcol strains appeared to produce stronger biofilm than BJI strains, though they belonged to similar CCs and had the same adhesin and pili content.IMPORTANCE Bone and joint infections (BJI) are pathologies that can be life-threatening and result in compromised functional prognosis for patients. Relapses are common and often related to biofilm formation. Group B streptococci (GBS) BJI increased since the last decade. However, few data are available on this subject in the literature. Our study aims to highlight genotype and biofilm production of GBS isolates from BJI. Seventy-seven GBS strains isolated from BJI and 57 from asymptomatic human vaginal colonization were characterized by multilocus sequence typing (MLST), adhesins content, nature of the pili and the ability to form biofilm. Our results revealed that vaginal human colonization strains produced stronger biofilm than BJI strains, despite belonging to the same phylogenetic lineage and having the same adhesin and pili content.

Multilocus Sequence Typing and Antifungal Susceptibility of Vaginal and Non-vaginal Candida glabrata Isolates From China.

Candida glabrata is a common cause of Candida infections. In our present study, we investigated the antifungal susceptibility and molecular epidemiology of vaginal and non-vaginal C. glabrata isolates. Seventy-six vaginal C. glabrata strains isolated from patients with vulvovaginal candidiasis and 57 non-vaginal C. glabrata isolates were collected at two hospitals in Shanghai, China. Antifungal susceptibility was examined using a broth microdilution method. Multilocus sequence typing was used for genotyping. Overall, 28 (21.1%), 28 (21.1%), and 29 (21.8%) C. glabrata isolates were resistant to fluconazole, itraconazole, and voriconazole, respectively. Briefly, 18 (23.7%), 18 (23.7%), and 19 (25%) vaginal strains were resistant to fluconazole, itraconazole, and voriconazole. While the resistance to these antifungals were all 17.5% (10/57) in non-vaginal strains. All isolates retained susceptibility to amphotericin B, and only four non-vaginal isolates were caspofungin resistant. Genotyping identified 17 ST patterns. In non-vaginal samples, the same genotypes appear as in the vaginal samples, except for one genotype (ST-182), while in the vaginal samples more genotypes appear (ST8, ST19, ST45, ST55, ST66, ST80, ST138, and ST17). The most common genotype was ST7 (81 strains), followed by ST10 (14 strains) and ST15 (11 strains). The majority of resistant phenotype strains (25/30, 83.3%) correlated to the predominant genotype (ST7), and the rest belonged to ST3 (2/30, 6.7%), ST10 (1/30, 3.3%), ST19 (1/30, 3.3%), and ST45 (1/30, 3.3%). Our survey revealed cross-resistance in vaginal and non-vaginal C. glabrata isolates. Moreover, there is no genotype associated with the resistance phenotype.

Antifungal Activity of Natural Compounds vs. Candida spp.: A Mixture of Cinnamaldehyde and Eugenol Shows Promising In Vitro Results.

Candida spp. are commensal organisms of the skin, mucous membranes, gastrointestinal tract, blood, and vagina of animals and humans. In recent decades, the incidence of human fungal infections has increased, with Candida spp. (mainly C. albicans) infections being the most frequent, and the treatment of fungal infections is still a clinical challenge. Colonization of the human gastrointestinal tract by Candida spp. is significant because infections (e.g., candidemia and vulvovaginal candidiasis) frequently arise from commensal microorganisms. The aim of this study was to test in vitro the antifungal activity and the eventual synergistic effect of five pure components of essential oils: cinnamaldehyde, α-pinene, limonene, eucalyptol, and eugenol. These compounds were tested on 18 Candida strains (15 C. albicans, 2 C. glabrata, and 1 C. lusitaniae) derived from a culture collection of vaginal clinical strains. Methods: Fungistatic activity was evaluated using the disk diffusion method. For fungicidal activity, microdilution and time–kill curve protocols were set up. The checkerboard method was chosen to evaluate a possible synergistic effect of these compounds when mixed. Results: Cinnamaldehyde and eugenol gave the best results, inhibiting all the Candida strains and showing a highly additive effect (FICI 0.625). The cinnamaldehyde inhibition zone (IZ), MIC, and MFC means were 69 mm, 50.05 mg/L, and 109.26 mg/L respectively. Cinnamaldehyde led to the total loss of viable Candida cells within 4 h. Eugenol IZ, MIC, and MFC means were 35.2 mm, 455.42 mg/L, and 690.09 mg/L, respectively. Eugenol led to the total loss of viable fungal cells within 1 h. Treatment with α-pinene inhibited 88.9% of Candida strains, with an IZ mean of 21.2 mm, a MIC mean of 195.41 mg/L, and a MFC mean of 251.27 mg/L; this compound led to the total loss of viable fungal cells only after 24 h. Limonene inhibited only 33.3% of the tested strains and eucalyptol did not produce an inhibition halo, so these compounds were not tested further. Conclusions: These characteristics, together with the well-known safety of cinnamaldehyde and eugenol for human use, make these two natural compounds the perfect candidates for the treatment of candidiasis. This was a pilot study, the purpose of which was to evaluate the best composition of a mixture to be used against intestinal and vulvovaginal candidiasis; in vivo studies are needed to confirm these very encouraging results.