BackgroundVaginal lactobacilli offer protection against recurrent urinary infections, bacterial vaginosis, and vaginal candidiasis.ObjectiveTo characterise the isolated vaginal lactobacilli strains for their probiotic properties and to compare their probiotic potential.MethodsThe Lactobacillus strains were isolated from vaginal samples by conventional culturing and identified by sequencing of the 16S rDNA fragment. Several functional properties were detected (production of hydrogen peroxide and lactic acid; antagonistic activity against Escherichia coli, Candida albicans, Candida glabrata, and Gardnerella vaginalis; auto-aggregation and adhesiveness) as well as safety (haemolytic activity, antibiotic susceptibility, presence of transferrable resistance genes).ResultsA total of 135 vaginal lactobacilli strains of three species, Lactobacillus crispatus (56%), Lactobacillus jensenii (26%), and Lactobacillus gasseri (18%) were characterised using several functional and safety tests. Most of L. crispatus (89%) and L. jensenii (86%) strains produced H2O2. The best lactic acid producers were L. gasseri (18.2±2.2 mg/ml) compared to L. crispatus (15.6±2.8 mg/ml) and L. jensenii (11.6±2.6 mg/ml) (p<0.0001; p<0.0001, respectively). L. crispatus strains showed significantly higher anti-E. coli activity compared to L. jensenii. L. gasseri strains expressed significantly lower anticandidal activity compared to L. crispatus and L. jensenii (p<0.0001). There was no significant difference between the species in antagonistic activity against G. vaginalis. Nearly a third of the strains were able to auto-aggregate while all the tested strains showed a good ability to adhere to HeLa cells. None of the tested lactobacilli caused haemolysis. Although phenotypical resistance was not found to ampicillin, chloramphenicol, erythromycin, gentamycin, tetracycline, and vancomycin, the erm(B), tet(M), and tet(K) were detected in some strains. All strains were resistant to metronidazole, trimethoprim/sulfamethoxazole, and kanamycin.ConclusionsOur study revealed that the production of different antimicrobial metabolites is highly strain-specific and that the metabolites are not correlated with each other. L. crispatus displays better antagonistic activity against E. coli and Candida spp. than L. gasseri and L. jensenii; therefore; a potential probiotic candidate could be found among L. crispatus strains.

Background: Only a few microbial studies have conducted in IVF (in vitro fertilization), showing the high-variety bacterial contamination of IVF culture media to cause damage to or even loss of cultured oocytes and embryos. We aimed to determine the prevalence and counts of bacteria in IVF samples, and to associate them with clinical outcome. Methods: The studied samples from 50 infertile couples included: raw (n = 48), processed (n = 49) and incubated (n = 50) sperm samples, and IVF culture media (n = 50). The full microbiome was analyzed by 454 pyrosequencing and quantitative analysis by real-time quantitative PCR. Descriptive statistics, t-, Mann-Whitney tests and Spearman's correlation were used for comparison of studied groups. Results: The study involved normozoospermic men. Normal vaginal microbiota was present in 72.0% of female partners, while intermediate microbiota and bacterial vaginosis were diagnosed in 12.0 and 16.0%, respectively. The decreasing bacterial loads were found in raw (35.5%), processed (12.0%) and sperm samples used for oocyte insemination (4.0%), and in 8.0% of IVF culture media. The most abundant genera of bacteria in native semen and IVF culture media were Lactobacillus, while in other samples Alphaproteobacteria prevailed. Staphylococcus sp. was found only in semen from patients with inflammation. Phylum Bacteroidetes was in negative correlation with sperm motility and Alphaproteobacteria with high-quality IVF embryos. Conclusion: Our study demonstrates that IVF does not occur in a sterile environment. The prevalent bacteria include classes Bacilli in raw semen and IVF culture media, Clostridia in processed and Bacteroidia in sperm samples used for insemination. The presence of Staphylococcus sp. and Alphaproteobacteria associated with clinical outcomes, like sperm and embryo quality.

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.

The composition of centenarians’ gut microbiota has consistently been used as a model for healthy aging studies. However, there is an incomplete understanding of how childhood living conditions and eating habits affect the development and composition of gastrointestinal microbiota in centenarians with good cognitive functions. We compared the gut microbiota as well as the living and eating habits of the oldest-old group and the young people group. The richness and diversity of microbiota and the abundance of hereditary and environmental microbes were higher in people with longevity than young people. People with longevity ate more potatoes and cereal products. In their childhood, they had more exposure to farm animals and did not have sewers compared with young people. Young people’s gut microbiota contained more butyrate-producing bacteria and bacteria that characterized an animal-based Western diet. These results expand our understanding of the effects of childhood environment and diet on the development and stability of the microbiota in people with longevity.

Extended-spectrum beta-lactamases (ESBL) and AmpC producing-Escherichia coli have spread worldwide, but data about ESBL-producing-E. coli in the Northern and Eastern regions of Europe is scant. The aim of this study has been to describe the phenotypical and molecular epidemiology of different ESBL/AmpC/Carbapenemases genes in E. coli strains isolated from the Baltic States (Estonia, Latvia, and Lithuania), Norway and St. Petersburg (Russia), and to determine the predominant multilocus sequence type and single nucleotide polymorphisms diversity of E. coli isolates deduced by whole genome sequencing (WGS). A total of 10,780 clinical E. coli strains were screened for reduced sensitivity to third-generation cephalosporins. They were collected from 21 hospitals located in Estonia, Latvia, Lithuania, Norway and St. Petersburg during a 5 month period in 2012. The overall prevalence of ESBL/AmpC strains was 4.7% by phenotypical test and 3.9% by sequencing. We found more strains with the ESBL/AmpC phenotype and genotype in St. Petersburg and Latvia than other countries. Of phenotypic E. coli strains, 85% contained confirmed ESBL genes (including blaCTX–M, blaTEM–29, blaTEM–71), AmpC genes (blaCMY–59, blaACT–12/–15/–20, blaESC–6, blaFEC–1, blaDHA–1), or carbapenemase genes (blaNDM–1). blaCTX–M–1, blaCTX–M–14 and blaCTX–M–15 were found in all countries, but blaCTX–M–15 prevalence was higher in Latvia than in St. Petersburg (Russia), Estonia, Norway and Lithuania. The dominating AmpC genes were blaCMY–59 in the Baltic States and Norway, and blaDHA–1 in St. Petersburg. E. coli strains belonged to 83 different sequence types, of which the most prevalent was ST131 (40%). In conclusion, we generally found low ESBL/AmpC/Carbapenemase prevalence in E. coli strains isolated in Northern/Eastern Europe. However, several inter-country differences in distribution of particular genes and multilocus sequence types were found.

Vaginal lactobacilli offer protection against microbiota imbalance and genitourinary tract infections. We compared vaginal lactobacilli in 50 Estonian women of child-bearing age applying culture-based methods, quantitative PCR and next-generation sequencing (NGS). The culture-based methods found three different lactobacilli: Lactobacillus crispatus, Lactobacillus jensenii and Lactobacillus gasseri. Using NGS revealed the presence of L. crispatus in 76%, Lactobacillus iners in 52%, L. jensenii in 47% and L. gasseri in 33% of the samples. According to qPCR, L. iners was present in 67% and L. crispatus in 64% of the samples. The proportions of L. crispatus revealed by qPCR and NGS were in good correlation (R=0.79, P<0.001), while that of L. iners correlated poorly (R=0.13, P>0.05). Good concordance for L. crispatus was also found between the results of the culture-based method and qPCR. Finally, good overlap between the results of the culture-based method and NGS was revealed: in case of a positive NGS result for L. crispatus, the same species was isolated in 95% of samples. The corresponding percentages were 82% for L. jensenii and 86% for L. gasseri. Our data indicate fairly general concordance of the three methods for detecting vaginal lactobacilli, except for L. iners. This points out the importance of standardisation of techniques, and the respective studies should involve cultures applying a medium suitable for the fastidious L. iners.

The spread of carbapenemase-producing Enterobacteriaceae is a global problem; however, no exact data on the epidemiology of carbapenemase in the Baltic countries and St. Petersburg area is available. We aimed to evaluate the epidemiology of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the Baltic States and St. Petersburg, Russia, and to compare the different methods for carbapenemase detection. From January to May 2012, all K. pneumoniae (n = 1983) and E. coli (n = 7774) clinical isolates from 20 institutions in Estonia, Latvia, Lithuania, and St. Petersburg, Russia were screened for carbapenem susceptibility. The IMP, VIM, GIM, NDM, KPC, and OXA-48 genes were detected using real-time PCR and the ability to hydrolyze ertapenem was determined using MALDI-TOF MS. Seventy-seven strains were found to be carbapenem nonsusceptible. From these, 15 K. pneumoniae strains hydrolyzed ertapenem and carried the bla NDM gene. All of these strains carried integron 1 and most carried integron 3 as well as genes of the CTX-M-1 group. No carbapenemase-producing E. coli or K. pneumoniae strains were found in Estonia, Latvia, or Lithuania; however, NDM-positive K. pneumoniae was present in the hospital in St. Petersburg, Russia. A MALDI-TOF MS-based assay is a suitable and cost-effective method for the initial confirmation of carbapenemase production.

ObjectiveThe first aim of this study was to compare the microbiota of different locations (pus, tonsillar fossa, blood) in peritonsillar abscess (PTA) patients in order to optimize the sampling scheme. The second aim was to estimate the occurrence of tonsillitis episodes and macroscopic oropharyngeal signs characteristic of recurrent tonsillitis in PTA patients.MethodsThe study group consisted of 22 consecutive patients with PTA undergoing bilateral tonsillectomy. The PTA was punctured; pus and tonsillar fossa biopsy samples and the peripheral blood cultures were collected. The index of tonsillitis was calculated by multiplying the number of tonsillitis episodes per year by the morbidity period in years. Macroscopic oropharyngeal signs were evaluated and they were as follows: tonsillar sclerosis, obstruction of the tonsillar crypts, scar tissue on tonsils, cryptic debris, and lymphatic tissue aggregates.ResultsThe cultures of the pus were positive in 16 out of 22 patients and the cultures of the tonsillar fossa samples were positive in all cases. In total, 62 different organisms were found from tonsillar fossa, pus, and blood samples, which belonged to 5 different phyla and 18 different families.In the tonsillar fossa, the most frequent bacteria found were Streptococcus spp. In pus samples, the most frequently found bacteria were Streptococcus spp. and bacteria from the Streptococcus milleri group.ConclusionPTA patients had mixed anaerobic and aerobic microbiota both in the tissue of the tonsillar fossa and the pus of the peritonsillar space. We demonstrated that the tonsillar fossa specimen is a better material for microbiological analyses, because it reveals more bacteria per culture. PTA patients usually have a low number of tonsillitis episodes in their previous history, but a relatively high number of macroscopic oropharyngeal signs, indicating the sclerotic process in palatal tonsils.