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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

Electrical Impedance Spectroscopy (EIS) characterization of saline solutions with a low-cost portable measurement system

Electrical Impedance Spectroscopy (EIS), a powerful technique used for wide range of applications, is usually carried out by means of benchtop instrumentation (LCR meters and ìmpedance analyzers), not suited for in-the-field measurements performed outside a laboratory.In this paper a new portable electronic system for EIS on liquid and semi-liquid media is presented that is capable to produce an electrical fingerprint of the sample under investigation. The proposed system was used for the characterization of four different saline solutions (NaCl, Na2CO3, K2HPO4 and CuSO4). A multi-frequency approach, based on the measurement of maximum value of the impedance imaginary component and its corresponding frequency, was tested for the first time to discriminate different saline solutions. The results show that the proposed method is capable to discriminate the different solutions and to measure the concentration (R2 = 0.9965) independently of the type of saline solution. Keywords: Impedance Spectroscopy, Measurement, Frequency, Sensor, Portable syste

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

Background: Lactobacillus 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. Results: Here, 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. Conclusions: Our 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

Vaginal microbiome and metabolome highlight specific signatures of bacterial vaginosis

In this study, we sought to find novel bacterial and metabolic hallmarks for bacterial vaginosis (BV). We studied the vaginal microbiome and metabolome of vaginal fluids from BV-affected patients (n = 43) and healthy controls (n = 37) by means of an integrated approach based on quantitative polymerase chain reaction (qPCR) and proton nuclear magnetic resonance ((1)H-NMR). The correlations between the clinical condition and vaginal bacterial communities were investigated by principal component analysis (PCA). To define the metabolomics signatures of BV, 100 discriminant analysis by projection on latent structure (PLS-DA) models were calculated. Bacterial signatures distinguishing the health condition and BV were identified by qPCR. Lactobacillus crispatus strongly featured the healthy vagina, while increased concentrations of Prevotella, Atopobium and Mycoplasma hominis specifically marked the infection. (1)H-NMR analysis has led to the identification and quantification of 17 previously unreported molecules. BV was associated with changes in the concentration of metabolites belonging to the families of amines, organic acids, short chain fatty acids, amino acids, nitrogenous bases and monosaccharides. In particular, maltose, kynurenine and NAD(+) primarily characterised the healthy status, while nicotinate, malonate and acetate were the best metabolic hallmarks of BV. This study helps to better understand the role of the vaginal microbiota and metabolome in the development of BV infection. We propose a molecular approach for the diagnosis of BV based on quantitative detection in the vaginal fluids of Atopobium, Prevotella and M. hominis, and nicotinate, malonate and acetate by combining qPCR and (1)H-NMR

Use of Lactobacillus crispatus to produce a probiotic cheese as potential gender food for preventing gynaecological infections

This research is aimed to evaluate the suitability of Squacquerone cheese to support the viability of Lactobacillus crispatus BC4, a vaginal strain endowed with a strong antimicrobial activity against urogenital pathogens and foodborne microorganisms, in order to recommend a gender food for woman wellbeing. The viability of L. crispatus BC4, used as adjunct culture, was evaluated during the refrigerated storage of Squacquerone cheese, as well as when the cheese was subjected to simulated stomach-duodenum passage tested by the patented Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Moreover, the effects of L. crispatus BC4 addition were evaluated on product hydrolytic patterns, in terms of proteolysis, lipolysis and volatile molecule profiles. The data showed that L. crispatus BC4 maintained high viability, also in presence of physiological stress conditions, until the end of the refrigerated storage. Moreover, the inclusion of L. crispatus BC4 gave rise to cheese product with higher score of overall acceptability when compared to control cheese. In addition, the survival of L. crispatus BC4, carried in test cheese, in gastro intestinal conditions was confirmed by SHIME. The results showed that the vaginal Lactobacillus strain was more affected by the low pH of the stomach, simulated by the SHIME reactor, rather than to bile salts and pancreatic juices. Although only in vivo trials will be able to confirm the functionality of the cheese in the vaginal environment, these data represent a first step towards the employment of the Squacquerone cheese as probiotic food able to promote the woman's health by preventing gynaecological infections

Liposomes-in-chitosan hydrogel boosts potential of chlorhexidine in biofilm eradication in vitro

Successful treatment of skin infections requires eradication of biofilms found in up to 90 % of all chronic wounds, causing delayed healing and increased morbidity. We hypothesized that chitosan hydrogel boosts the activity of liposomally-associated membrane active antimicrobials (MAA) and could potentially improve bacterial and biofilm eradication. Therefore, liposomes (∼300 nm) bearing chlorhexidine (CHX; ∼50 μg/mg lipid) as a model MAA were incorporated into chitosan hydrogel. The novel CHX-liposomes-in-hydrogel formulation was optimized for skin therapy. It significantly inhibited the production of nitric oxide (NO) in lipopolysaccharide (LPS)-induced macrophage and almost completely reduced biofilm formation. Moreover, it reduced Staphylococcus aureus and Pseudomonas aeruginosa adherent bacterial cells in biofilm by 64.2–98.1 %. Chitosan hydrogel boosted the anti-inflammatory and antimicrobial properties of CHX

Impact of a synbiotic food on the gut microbial ecology and metabolic profiles

<p>Abstract</p> <p>Background</p> <p>The human gut harbors a diverse community of microorganisms which serve numerous important functions for the host wellbeing. Functional foods are commonly used to modulate the composition of the gut microbiota contributing to the maintenance of the host health or prevention of disease. In the present study, we characterized the impact of one month intake of a synbiotic food, containing fructooligosaccharides and the probiotic strains <it>Lactobacillus helveticus </it>Bar13 and <it>Bifidobacterium longum </it>Bar33, on the gut microbiota composition and metabolic profiles of 20 healthy subjects.</p> <p>Results</p> <p>The synbiotic food did not modify the overall structure of the gut microbiome, as indicated by Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). The ability of the probiotic <it>L. helveticus </it>and <it>B. longum </it>strains to pass through the gastrointestinal tract was hypothesized on the basis of real-time PCR data. In spite of a stable microbiota, the intake of the synbiotic food resulted in a shift of the fecal metabolic profiles, highlighted by the Gas Chromatography Mass Spectrometry Solid Phase Micro-Extraction (GC-MS/SPME) analysis. The extent of short chain fatty acids (SCFA), ketones, carbon disulfide and methyl acetate was significantly affected by the synbiotic food consumption. Furthermore, the Canonical discriminant Analysis of Principal coordinates (CAP) of GC-MS/SPME profiles allowed a separation of the stool samples recovered before and after the consumption of the functional food.</p> <p>Conclusion</p> <p>In this study we investigated the global impact of a dietary intervention on the gut ecology and metabolism in healthy humans. We demonstrated that the intake of a synbiotic food leads to a modulation of the gut metabolic activities with a maintenance of the gut biostructure. In particular, the significant increase of SCFA, ketones, carbon disulfide and methyl acetate following the feeding period suggests potential health promoting effects of the synbiotic food.</p

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 (&gt;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