Development of a xanthan gum film for the possible treatment of vaginal infections.

Bacterial vaginosis is a vaginal infection that affects 60% of women of reproductive age worldwide. It is mainly caused by the bacterium Gardnerella vaginalis and is a factor that increases the probability of getting sexually transmitted diseases. We aimed to develop a new pharmaceutical form for the treatment of vaginal infections. We employed the solving-casting method to fabricate a polymeric film with Xanthan gum, a natural polymer produced by the bacterium Xanthomonas campestris, and metronidazole, one of the most commonly used drugs for vaginal infections. In order to characterize the film, we measured pH, dose uniformity, dissolution profile, and the percentage of swelling. Moreover, we performed a thermogravimetric analysis and scanning electron microscopy. The results demonstrated a pH suitable for vaginal application and uniform distribution of the drug in the film. Also, the formulation exhibited a high percentage of swelling and a slow release of the drug in a simulated vaginal fluid medium. All these attributes indicated that the manufactured film has ideal characteristics to be used and administered vaginally. It could be an excellent alternative to treat bacterial vaginosis and also improve user adherence.

Gardnerella vaginalis in urinary tract infections, are men spared?

Gardnerella vaginalis in association with anaerobes has been linked to bacterial vaginosis in women, while urinary tract infections (UTIs) in men have rarely been reported. The aim of the review was to reveal the significance of G. vaginalis UTIs in men. Prevalence of G. vaginalis UTIs in men varied from 0.5 to >27% according to patients' groups. Most patients had comorbidity such as urolithiasis or stents, transplants, tumors and diabetes, however, infections can also affect immunocompetent patients. We observed G. vaginalis-associated bacteriuria and leukocyturia in a kidney transplant man. Complications of the UTIs such as bacteremia (in 9/11 cases), hydronephrosis (4/11) and abscesses or septic emboli have been reported. Bacterial vaginosis in female partners has been a risk factor for UTIs in males. In women, biofilm Gardnerella phenotype, stabilized by Atopobium vaginae and Prevotella bivia was linked to ≥6-fold higher antibiotic resistance rates compared with the planktonic phenotype. Non-susceptibility to metronidazole and levofloxacin was found also in males. Therefore, if aerobic urine cultures are negative, urine and blood samples from male patients with predisposing factors and clinical signs of UTIs and bacteremia, can be taken. Plates should be incubated for 2-4 days in capnophilic/microaerophilic conditions, however only anaerobic incubation can help with detecting G. vaginalis strains which grow only anaerobically. Susceptibility testing of the isolates is highly important. Briefly, adherent G. vaginalis phenotype can be sexually transmissible. Despite the infrequency of G. vaginalis UTIs in men, the infections should be considered since they are often linked to severe complications.

Gardnerella vaginalis and Neisseria gonorrhoeae Are Effectively Inhibited by Lactobacilli with Probiotic Properties Isolated from Brazilian Cupuaçu (Theobroma grandiflorum) Fruit.

In recent years, certain Lactobacillus sp. have emerged in health care as an alternative therapy for various diseases. Based on this, this study is aimed at evaluating in vitro the potential probiotics of five lactobacilli strains isolated from pulp of cupuaçu fruit fermentation against Gardnerella vaginalis and Neisseria gonorrhoeae. Our lactobacilli strains were classified as safe for use in humans, and they were tolerant to heat and pH. Our strains were biofilm producers, while hydrophobicity and autoaggregation varied from 13% to 86% and 13% to 25%, respectively. The coaggregation of lactobacilli used in this study with G. vaginalis and N. gonorrhoeae ranged from 15% to 36% and 32% to 52%, respectively. Antimicrobial activity was present in all tested Lactobacillus strains against both pathogens, and the growth of pathogens in coculture was reduced by the presence of our lactobacilli. Also, all tested lactobacilli reduced the pH of the culture, even in incubation with pathogens after 24 hours. The cell-free culture supernatants (CFCS) of all five lactobacilli demonstrated activity against the two pathogens with a halo presence and CFCS characterization assay together with gas chromatography revealed that lactic acid was the most abundant organic acid in the samples (50% to 62%). Our results demonstrated that the organic acid production profile is strain-specific. This study revealed that cupuaçu is a promising source of microorganisms with probiotic properties against genital pathogens. We demonstrated by in vitro tests that our Lactobacillus strains have probiotic properties. However, the absence of in vivo tests is a limitation of our work due to the need to evaluate the interaction of our lactobacilli with pathogens in the vaginal mucosa. We believe that these findings may be useful in developing a product containing our lactobacilli and their supernatants in order to support with vaginal health.

Quantitative modeling predicts mechanistic links between pre-treatment microbiome composition and metronidazole efficacy in bacterial vaginosis.

Bacterial vaginosis is a condition associated with adverse reproductive outcomes and characterized by a shift from a Lactobacillus-dominant vaginal microbiota to a polymicrobial microbiota, consistently colonized by strains of Gardnerella vaginalis. Metronidazole is the first-line treatment; however, treatment failure and recurrence rates remain high. To understand complex interactions between Gardnerella vaginalis and Lactobacillus involved in efficacy, here we develop an ordinary differential equation model that predicts bacterial growth as a function of metronidazole uptake, sensitivity, and metabolism. The model shows that a critical factor in efficacy is Lactobacillus sequestration of metronidazole, and efficacy decreases when the relative abundance of Lactobacillus is higher pre-treatment. We validate results in Gardnerella and Lactobacillus co-cultures, and in two clinical cohorts, finding women with recurrence have significantly higher pre-treatment levels of Lactobacillus relative to bacterial vaginosis-associated bacteria. Overall results provide mechanistic insight into how personalized differences in microbial communities influence vaginal antibiotic efficacy.

Changes in the vaginal microbiota following antibiotic treatment for Mycoplasma genitalium, Chlamydia trachomatis and bacterial vaginosis.

The human vagina harbor a rich microbiota. The optimal state is dominated by lactobacilli that help to maintain health and prevent various diseases. However, the microbiota may rapidly change to a polymicrobial state that has been linked to a number of diseases. In the present study, the temporal changes of the vaginal microbiota in patients treated for sexually transmitted diseases or bacterial vaginosis (BV) and in untreated controls were studied for 26 days. The patients included 52 women treated with azithromycin, tetracyclines or moxifloxacin for present or suspected infection with Chlamydia trachomatis or Mycoplasma genitalium. Women with concurrent BV were also treated with metronidazole. The controls were 10 healthy women of matching age. The microbiota was analyzed by 16S rRNA gene deep sequencing, specific qPCRs and microscopy. There was generally good correlation between Nugent score and community state type (CST) and qPCR confirmed the sequencing results. By sequencing, more than 600 different taxa were found, but only 33 constituted more than 1 ‰ of the sequences. In both patients and controls the microbiota could be divided into three different community state types, CST-I, CST-III and CST-IV. Without metronidazole, the microbiota remained relatively stable regarding CST although changes were seen during menstrual periods. Administration of metronidazole changed the microbiota from CST-IV to CST-III in approximately 50% of the treated patients. In contrast, the CST was generally unaffected by azithromycin or tetracyclines. In 30% of the BV patients, Gardnerella vaginalis was not eradicated by metronidazole. The majority of women colonized with Ureaplasma parvum remained positive after azithromycin while U. urealyticum was eradicated.

Latencia al parto tras cerclaje de emergencia en gestación múltiple / Latency to delivery after emergency physical-examination indicated cerclage in twin pregnancy

Introducción: Las guías clínicas actuales recomiendan el uso del cerclaje de emergencia (CE) como tratamiento de la insuficiencia cervical con exposición de membranas en gestaciones únicas. Sin embargo, el CE en gestación múltiple es tema de controversia dado que no existen ensayos clínicos randomizados que demuestren su eficacia. Algunos estudios retrospectivos sugieren que el CE también podría prolongar la gestación en embarazos múltiples. El objetivo de nuestro estudio es evaluar los resultados de las gestaciones múltiples que se sometieron a un CE en nuestro centro. Materiales y métodos: Se diseñó un estudio retrospectivo que incluyó los CE realizados en gestaciones gemelares en nuestro centro entre 2007-2016. No fueron tributarias de CE gestaciones con malformaciones fetales, monocoriales-monoamnióticas, triple o superior y finalizaciones activas de la gestación. Variables primarias: latencia al parto espontáneo y edad gestacional al parto. Variables secundarias: mortalidad neonatal, ingreso en UCI neonatal, rotura prematura de membranas pretérmino, corioamnionitis y fallo del cerclaje. Resultados. El estudio incluyó 17 pacientes. La edad gestacional mediana (rango intercuartil) al parto fue de 27,1 (24,5-32,3) semanas y la latencia mediana (rango intercuartil) al parto fue de 43 (21-64) días. Hubo 4/17 (23,5%) casos de parto antes de las 24 semanas de gestación y 2/26 (7,7%) de muerte neonatal. Discusión: Estos resultados muestran que la latencia al parto después del CE en gestación múltiple es remarcable, por lo que podría ser considerado como una opción terapéutica. Sin embargo, se requiere evidencia basada en estudios randomizados para hacer una recomendación firme

Introduction: Current guidelines support the use of physical-examination indicated cerclage (PEIC) as a treatment for cervical insufficiency and membrane exposure in single pregnancies. However, PEIC in twin pregnancies is a controversial issue as no data from random clinical trial are available to demonstrate its efficacy. Few studies suggest that PEIC may prolong pregnancy also in twin pregnancies. The aim of this study was to evaluate the results of twin pregnancies that underwent a PEIC in our health centre. Material and methods: A retrospective review was performed on women that underwent a PEIC from 2007-2016 in our centre. Women were not eligible if they were carrying foetuses with major foetal anomalies, more than two foetuses or monochorionic-monoamniotic pregnancies, or three or more foetuses or requesting an elective termination of pregnancy. Primary outcomes: latency to spontaneous delivery and gestational age (GA) at delivery. Secondary outcomes: neonatal mortality and Neonatal Intensive Care Unit admission, preterm premature rupture of membranes (PPROM), chorioamnionitis and cerclage displacement. Results: The study included a total of 17 women. The median (inter-quartile range) gestational age at delivery was 27.1 (24.5-32.3) weeks, and median (inter-quartile range) latency, from cervical cerclage to delivery, was 43 (21-64) days. There were 4/17 (23.5%) cases of delivery before 24 weeks of pregnancy, and 2/26 (7.7%) cases of neonatal death. Discussion: These results suggest that latency to delivery after PEIC in twins is remarkable. Therefore, it could be considered as an optional management. Nevertheless, evidence based on random clinical trial is required to make firm recommendations on its formal use

Long lasting mucoadhesive membrane based on alginate and chitosan for intravaginal drug delivery.

The intravaginal route of administration can be exploited to treat local diseases and for systemic delivery. In this work, we developed an alginate/chitosan membrane sufficiently stable in a simulated vaginal fluid and able to dissolve over time at a very slow and linear rate. The membrane demonstrated good mechanical properties both in its swollen and dry form. As a study case, we evaluated the viability of this potential drug delivery system for the treatment of bacterial vaginosis, a common disease affecting women in their reproductive age. Metronidazole was effectively included in the alginate/chitosan membrane and its bactericide effect was demonstrated against Staphylococcus aureus and Gardnerella vaginalis, simultaneously showing good biocompatibility with a cervix epithelial cell line. Since this alginate/chitosan membrane is stable in a simulated vaginal environment, is easy to fabricate and can be used for the controlled release of a model drug, it represents a promising drug delivery system for local intravaginal applications.

Nonoptimal Vaginal Microbiota After Azithromycin Treatment for Chlamydia trachomatis Infection.

We characterized the composition and structure of the vaginal microbiota in a cohort of 149 women with genital Chlamydia trachomatis infection at baseline who were followed quarterly for 9 months after antibiotic treatment. At time of diagnosis, the vaginal microbiota was dominated by Lactobacillus iners or a diverse array of bacterial vaginosis-associated bacteria including Gardnerella vaginalis. Interestingly, L. iners-dominated communities were most common after azithromycin treatment (1 g monodose), consistent with the observed relative resistance of L. iners to azithromycin. Lactobacillus iners-dominated communities have been associated with increased risk of C. trachomatis infection, suggesting that the impact of antibiotic treatment on the vaginal microbiota could favor reinfections. These results provide support for the dual need to account for the potential perturbing effect(s) of antibiotic treatment on the vaginal microbiota, and to develop strategies to protect and restore optimal vaginal microbiota.

Cationic amphiphiles against Gardnerella vaginalis resistant strains and bacterial vaginosis-associated pathogens.

Antibiotic resistance and infection recurrence are critical issues in treating bacterial vaginosis, the most common vaginal disorder in women of reproductive age. Novel alternatives to traditional antibiotics, such as peptidomimetics, have the potential to address this challenge. Previously, two series of cationic amphiphiles (CAms) were developed with both hydrophilic head groups and non-polar domains, giving them the ability to self-assemble into supramolecular nanostructures with membrane-lytic properties. Those CAms were shown to be effective against biofilms of Gardnerella vaginalis while preserving the commensal microbiota. Two new series of CAms were designed with varying levels of flexibility between the hydrophilic head groups and the hydrophobic domains. Activities against the vaginal pathogen G. vaginalis ranged from 1.3 to 18.5 µM, while the tested vaginal lactobacilli were significantly more tolerant of CAms, with minimal inhibitory concentration values as high as 208 µM. Minimal biofilm bactericidal concentrations of the tested CAms ranged from 21.47 to <388.3 µM, and were lowest against resistant forms of G. vaginalis, while Lactobacillus biofilms were tolerant of concentrations ≥687 µM. Safety aspects of the CAms were also investigated, and they were found to be safe for use against vaginal ectocervical tissue.

Potential vaginal probiotics: safety, tolerability and preliminary effectiveness.

Vaginal discharge is one of the common reasons for gynaecologist consultation, as bacterial vaginosis and candidiasis are the main causes of discharge. These patients frequently experience numerous problems due to recurrent infections, side effects and drug resistance therefore alternative drugs are needed. Our primary aim was to evaluate safety and tolerability of the potentially probiotic Lactobacillus crispatus strains in volunteer women considering themselves healthy. We also monitored the effects of these strains on vaginal health parameters and lactobacilli counts in vagina and intestine. Forty women were recruited into trial. Absence of chronic diseases was confirmed by questionnaire and blood analysis at screening visit. In randomised double-blind placebo-controlled crossover study the eligible participants were randomly allocated to one of four groups and had to consume one of the two study products (Pro I or Pro II) - a capsule containing 3 strains, 109 cfu per strain, or placebo for 1 week. Treatment period was followed by 2-week washout period and continued with second treatment and washout period. Individuals receiving firstly probiotic, received later placebo and vice versa. Blood, vaginal and faecal samples were collected, and self-reported questionnaires were applied. Thirty subjects completed the trial. The probiotic capsules were well-tolerated. The Pro II intake resulted in a significant decrease in Nugent score (from median 3.0 to 2.0, mean 3.9 to 2.6, P=0.002) and reduction in Gardnerella vaginalis counts (log10 3.57 to 2.38; P=0.027). Reduction of total vaginal bacterial counts was revealed in Pro I group (log10 7.99 to 7.72; P=0.048). In conclusion, the selected vaginal L. crispatus strains are well tolerable and Pro II mixture is prospectively effective in reducing Nugent score and vaginal counts of G. vaginalis. Therefore, these strains seem to be promising candidates for development of novel evidence-based well-focused probiotics to target female urogenital tract disorders.

Benzoyl Peroxide Inhibits Quorum Sensing and Biofilm Formation by Gardnerella vaginalis 14018.

Infection recurrence and antibiotic resistance of bacterial vaginosis-associated pathogenic biofilms underline the need for novel and effective treatment strategies. In this study, we evaluated the antimicrobial, antibiofilm, and quorum sensing inhibitory effects of benzoyl peroxide and salicylic acid against Gardnerella vaginalis ATCC 14018, the predominant pathogen of bacterial vaginosis. While the highest tested concentrations of 250 and 125 µg/mL for both compounds were not sufficient in completely inhibiting the growth of G. vaginalis ATCC 14018, they did prevent biofilm formation by inhibiting the bacterial quorum sensing system in the pathogen. To our knowledge, this report is the first evidence that benzoyl peroxide can have a quorum sensing-mediated biofilm controlling effect, as demonstrated using subinhibitory concentrations of this compound in order to reduce the cost, dosage, and negative side effects associated with current antimicrobial treatments.

Vaginal microbiome modulates topical antiretroviral drug pharmacokinetics.

Tenofovir gel and dapivirine ring provided variable HIV protection in clinical trials, reflecting poor adherence and possibly biological factors. We hypothesized that vaginal microbiota modulates pharmacokinetics and tested the effects of pH, individual bacteria, and vaginal swabs from women on pharmacokinetics and antiviral activity. Tenofovir, but not dapivirine, uptake by human cells was reduced as pH increased. Lactobacillus crispatus actively transported tenofovir leading to a loss in drug bioavailability and culture supernatants from Gardnerella vaginalis, but not Atopobium vaginae, blocked tenofovir endocytosis. The inhibition of endocytosis mapped to adenine. Adenine increased from 65.5 µM in broth to 246 µM in Gardnerella, but decreased to 9.5 µM in Atopobium supernatants. This translated into a decrease in anti-HIV activity when Gardnerella supernatants or adenine were added to cultures. Dapivirine was also impacted by microbiota, as drug bound irreversibly to bacteria, resulting in decreased antiviral activity. When drugs were incubated with vaginal swabs, 30.7% ± 5.7% of dapivirine and 63.9% ± 8.8% of tenofovir were recovered in supernatants after centrifugation of the bacterial cell pellet. In contrast, no impact of microbiota on the pharmacokinetics of the prodrugs, tenofovir disoproxil fumarate or tenofovir alafenamide, was observed. Together, these results demonstrate that microbiota may impact pharmacokinetics and contribute to inconsistent efficacy.

Inhibition of sialidase activity and cellular invasion by the bacterial vaginosis pathogen Gardnerella vaginalis.

Bacterial vaginosis is a genital tract infection, thought to be caused by transformation of a lactobacillus-rich flora to a dysbiotic microbiota enriched in mixed anaerobes. The most prominent of these is Gardnerella vaginalis (GV), an anaerobic pathogen that produces sialidase enzyme to cleave terminal sialic acid residues from human glycans. Notably, high sialidase activity is associated with preterm birth and low birthweight. We explored the potential of the sialidase inhibitor Zanamavir against GV whole cell sialidase activity using methyl-umbelliferyl neuraminic acid (MU-NANA) cleavage assays, with Zanamavir causing a 30% reduction in whole cell GV sialidase activity (p < 0.05). Furthermore, cellular invasion assays using HeLa cervical epithelial cells, infected with GV, demonstrated that Zanamivir elicited a 50% reduction in cell association and invasion (p < 0.05). Our data thus highlight that pharmacological sialidase inhibitors are able to modify BV-associated sialidase activity and influence host-pathogen interactions and may represent novel therapeutic adjuncts.

Bacteramia Caused by Gardnerella Vaginalis in a Cesarean Section Patient.

BACKGROUND: Gardnerella vaginalis is a facultative anaerobic and small gram-variable rod bacterium. G. vaginalis, which can be transmitted through sexual contact, is the common pathogen for the feminine bacterial pathogen (BV). Here we describe a case of bacteremia in a patient after cesarean section caused by G. vaginalis in China. Case presentation: A 35-year-old woman suffered bacteremia caused by G. vaginalis after cesarean section. This patient, without evidence of polymicrobial infection, was treated with cefuroxime and had a good outcome. CONCLUSIONS: G. vaginalis bacteremia is rarely reported. Our report expands the range of infection caused by G. vaginalis.

Anti-inflammatory effect of two Lactobacillus strains during infection with Gardnerella vaginalis and Candida albicans in a HeLa cell culture model.

Lactobacilli are the dominant bacteria of the vaginal tract of healthy women and they play a major role in the maintenance of mucosal homeostasis, preventing genital infections, such as bacterial vaginosis (BV) and vulvovaginal candidiasis (VVC). It is now known that one mechanism of this protection is the influence that lactobacilli can exert on host immune responses. In this context, we evaluated two Lactobacillus strains (L. plantarum 59 and L. fermentum 137) for their immunomodulatory properties in response to Gardnerella vaginalis (BV) or Candida albicans (VVC) infections in a HeLa cell infection model. G. vaginalis and C. albicans triggered the secretion of pro-inflammatory cytokines (TNF-α, IL-1ß, IL-6 and IL-8) and the activation of NF-κB in HeLa cells, in contrast to L. plantarum 59 and L. fermentum 137. Treatments with the Lactobacillus strains or their cell-free supernatants before (pre-treatment) or after (post-treatment) the challenge with the pathogens resulted in decreased secretion of pro-inflammatory cytokines and decreased activation of NF-κB. The treatments with Lactobacillus strains not only decreased the secretion of IL-8, but also its expression, as confirmed by gene reporter luciferase assay, suggesting transcription-level control by lactobacilli. In conclusion, L. plantarum 59 and L. fermentum 137 were confirmed to have an anti-inflammatory effect against G. vaginalis and C. albicans and they were able to influence signalling in NF-κB pathway, making them interesting candidates as probiotics for the prevention or treatment of BV and VVC.

The Effect of Lysozyme on Reducing Biofilms by Staphylococcus aureus, Pseudomonas aeruginosa, and Gardnerella vaginalis: An In Vitro Examination.

Two basic questions about lysozyme activities on the selected microorganisms were investigated, namely whether lysozyme inhibits biofilm production and which concentrations of the enzyme have the ability to change the natural biofilm producing capacity of different strains of Staphylococcus aureus (methicillin sensitive and resistant), Streptococcus pyogenes, Pseudomonas aeruginosa, and Gardnerella vaginalis. The effect of lysozyme on biofilm formation capacities of 16 strains of selected microorganisms was investigated, whereby four testing replicates have been performed in vitro using the Test Tube method, and the potential of lysozyme to change biofilm forming capacities depending on its concentration, species, and strains of microorganisms is demonstrated. A lysozyme concentration of 30 µg/ml indicated to have the highest inhibiting effect on all tested microorganisms. Furthermore, G. vaginalis was the most sensitive of them all, as its biofilm formation was inhibited in the presence of as low as 2.5 µg/ml of lysozyme. At enzyme concentrations of 7.5-50 µg/ml (with the exception of 30 µg/ml) the biofilm forming capacities of P. aeruginosa were enhanced. Depending on the strain of P. aeruginosa, the total biofilm quantity was either reduced or unaffected at lysozyme concentrations of 2.5, 5, 7.5, and 30 µg/ml. In contrast, lysozyme concentrations below 15 or 20 µg/ml did not affect or increase the volume of biofilm formation, while higher concentrations (15, 20, 25 µg/ml) reduced biofilm formation by 50% (3/6) and 30 µg/ml of biofilm reduced biofilm forming capacity of S. aureus by 100% (6/6). The results of this study are a strong foundation for further research on lysozyme as a modulator of the biofilm forming capacity of different species with the potential to aid in the development of new drugs for the treatment of oral and vaginal infections.

Sustained Simultaneous Delivery of Metronidazole and Doxycycline From Polycaprolactone Matrices Designed for Intravaginal Treatment of Pelvic Inflammatory Disease.

Poly(ɛ-caprolactone) (PCL) intravaginal matrices were produced for local delivery of a combination of antibacterials, by rapidly cooling a mixture of drug powders dispersed in PCL solution. Matrices loaded with different combinations of metronidazole (10%, 15%, and 20% w/w) and doxycycline (10% w/w) were evaluated in vitro for release behavior and antibacterial activity. Rapid "burst release" of 8%-15% of the doxycycline content and 31%-37% of the metronidazole content occurred within 24 h when matrices were immersed in simulated vaginal fluid at 37°C. The remaining drug was extracted gradually over 14 days to a maximum of 65%-73% for doxycycline and 62%-71% for metronidazole. High levels of antibacterial activity up to 89%-91% against Gardnerella vaginalis and 84%-92% against Neisseria gonorrhoeae were recorded in vitro for release media collected on day 14, compared to "nonformulated" metronidazole and doxycycline solutions. Based on the in vitro data, the minimum levels of doxycycline and metronidazole released from PCL matrices in the form of intravaginal rings into vaginal fluid in vivo were predicted to exceed the minimum inhibitory concentrations for N. gonorrhea (reported range 0.5-4.0 µg/mL) and G. vaginalis (reported range 2-12.8 µg/mL) respectively, which are 2 of the major causative agents for pelvic inflammatory disease.

In Vitro Activity of Lactobacilli with Probiotic Potential Isolated from Cocoa Fermentation against Gardnerella vaginalis.

Study of the probiotic potential of microorganisms isolated from fermented foods has been increasing, especially studies related to lactobacilli. In intestinal models, lactobacilli have demonstrated beneficial properties, such as anti-inflammatory activity and increased antibody production, but the molecular mechanisms involving probiotic and antagonistic action as well as their effect on human vaginal cells have not yet been fully elucidated. The aim of this study was to evaluate the functional and antagonistic properties of three strains of lactobacilli isolated from cocoa fermentation (Lactobacillus fermentum 5.2, L. plantarum 6.2, and L. plantarum 7.1) against Gardnerella vaginalis. Our results show that the lactobacilli have potential use as probiotics, since they have high hydrophobicity and autoaggregation properties and effectively adhere to vaginal cells. Metabolites secreted into the culture medium and whole cells of the strains under study are capable of interfering with the growth of G. vaginalis to different degrees. The elucidation of the antagonistic mechanisms as well as their effect on human cells may be useful in the development of a product containing such microorganisms or products secreted by them.

Method of Selection of Bacteria Antibiotic Resistance Genes Based on Clustering of Similar Nucleotide Sequences.

A new method for selection of bacterium antibiotic resistance genes is proposed and tested for solving the problems related to selection of primers for PCR assay. The method implies clustering of similar nucleotide sequences and selection of group primers for all genes of each cluster. Clustering of resistance genes for six groups of antibiotics (aminoglycosides, ß-lactams, fluoroquinolones, glycopeptides, macrolides and lincosamides, and fusidic acid) was performed. The method was tested for 81 strains of bacteria of different genera isolated from patients (K. pneumoniae, Staphylococcus spp., S. agalactiae, E. faecalis, E. coli, and G. vaginalis). The results obtained by us are comparable to those in the selection of individual genes; this allows reducing the number of primers necessary for maximum coverage of the known antibiotic resistance genes during PCR analysis.

Gemini Cationic Amphiphiles Control Biofilm Formation by Bacterial Vaginosis Pathogens.

Antibiotic resistance and recurrence of bacterial vaginosis (BV), a polymicrobial infection, justify the need for novel antimicrobials to counteract microbial resistance to conventional antibiotics. Previously, two series of cationic amphiphiles (CAms) which self-assemble into supramolecular nanostructures with membrane-lytic properties were designed with hydrophilic head groups and nonpolar domains. The combination of CAms and commonly prescribed antibiotics is suggested as a promising strategy for targeting microorganisms that are resistant to conventional antibiotics. Activities of the CAms against Gardnerella vaginalis ATCC 14018, a representative BV pathogen, ranged from 1.1 to 24.4 µM. Interestingly, the tested healthy Lactobacillus species, especially Lactobacillus plantarum ATCC 39268, were significantly more tolerant of CAms than the selected pathogens. In addition, CAms prevented biofilm formation at concentrations which did not influence the normal growth ability of G. vaginalis ATCC 14018. Furthermore, the biofilm minimum bactericidal concentration (MBC-Bs) of CAms against G. vaginalis ATCC 14018 ranged from 58.8 to 425.6 µM, while much higher concentrations (≥850 µM) were required to produce ≥3-log reductions in the number of biofilm-associated lactobacilli. The conventional antibiotic metronidazole synergized strongly with all tested CAms against planktonic cells and biofilms of G. vaginalis ATCC 14018. The synergism between CAms and the tested conventional antibiotic may be considered a new, effective, and beneficial method of controlling biofilm-associated bacterial vaginosis.