A combination of the probiotic and prebiotic product can prevent the germination of Clostridium difficile spores and infection.

Clostridium difficile infection (CDI) is one of the most prevalent healthcare associated infections in hospitals and nursing homes. Different approaches are used for prevention of CDI. Absence of intestinal lactobacilli and bifidobacteria has been associated with C. difficile colonization in hospitalized patients. Our aim was to test a) the susceptibility of C. difficile strains of different origin and the intestinal probiotic Lactobacillus plantarum Inducia (DSM 21379) to various antimicrobial preparations incl. metronidazole, vancomycin; b) the susceptibility of C. difficile strains to antagonistic effects of the probiotic L. plantarum Inducia, prebiotic xylitol (Xyl) and their combination as a synbiotic (Syn) product; c) the suppression of germination of C. difficile spores in vitro and in vivo in animal model of C. difficile infection with Inducia, Xyl and Syn treatment. The VPI strain 10463 (ATCC 43255), epidemic strain (M 13042) and clinical isolates (n = 12) of C. difficile from Norway and Estonia were susceptible and contrarily L. plantarum Inducia resistant to vancomycin, metronidazole and ciprofloxacin. The intact cells of Inducia, natural and neutralized cell free supernatant inhibited in vitro the growth of tested C. difficile reference strain VPI and Estonian and Norwegian clinical isolates of C. difficile after co-cultivation. This effect against C. difficile sustained in liquid media under ampicillin (0.75 µg/ml) and Xyl (5%) application. Further, incubation of Inducia in the media with 5% Xyl fully stopped germination of spores of C. difficile VPI strain after 48 h. In infection model the 48 hamsters were administered ampicillin (30 mg/kg) and 10-30 spores of C. difficile VPI strain. They also received five days before and after the challenge a pretreatment with a synbiotic (single daily dose of L. plantarum Inducia 1 ml of 1010 CFU/ml and 20% xylitol in 1 ml by orogastric gavage). The survival rate of hamsters was increased to 78% compared to 13% (p = 0.003) survival rate of hamsters who received no treatment. When administered Xyl the survival rate of hamsters reached 56% vs.13% (p = 0.06). In both Syn (6/9, p = 0.003) and Xyl (3/9, p = 0.042) groups the number of animals not colonized with C. difficile significantly increased. In conclusion, the combination of xylitol with L. plantarum Inducia suppresses the germination of spores and outgrowth into vegetative toxin producing cells of C. difficile and reduces the colonization of gut with the pathogen. Putative therapeutical approach includes usage of the synbiotic during antimicrobial therapy for prevention of CDI and its potential to reduce recurrences of CDI.

Comparison of detection methods for vaginal lactobacilli.

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.

Safety of a probiotic cheese containing Lactobacillus plantarum Tensia according to a variety of health indices in different age groups.

Safety of the probiotic Lactobacillus plantarum strain Tensia (DSM 21380) was tested in vitro, in semihard Edam-type cheese, in an animal model and after consumption of the probiotic cheese in double-blind randomized placebo-controlled human intervention studies with different age groups. The susceptibility of L. plantarum Tensia to 8 antibiotics, and the presence of tetracycline (tet M, S, O, K, L) genes and class 1 integron was assessed by applying epsilometer-test and PCR-based methods. Production of biogenic amines by the probiotic strain in decarboxylation medium containing 1% of l-histidine, l-glutamine, l-ornithine, l-arginine, or l-lysine and in cheese was tested by gas chromatography. The biosafety of L. plantarum Tensia was evaluated on National Institutes of Health-line mice fed cheese containing Tensia at a concentration of 9.6 log cfu/g for 30 consecutive days. In human intervention trials in adults and the elderly, the effects of different doses of Edam-type cheese and the probiotic bacterium on BW, gut functionality indices, and host metabolism were evaluated. The strain L. plantarum Tensia was susceptible to all tested antibiotics and did not possess the tetracycline resistance-determining genes tet(L), tet(S) and tet(O), nor did it contain the integron (Int1) gene. However, the strain was tet(K) and tet(M) positive. Lactobacillus plantarum Tensia did not produce potentially harmful biogenic amines, such as histamine or cadaverine. The amount of tyramine produced in the cheese environment during ripening and after 15 wk of storage was below the clinically significant content. In the animal model, no translocation of the administered strain or other microbes into the blood or organs of mice was detected. No harmful effect was observed on body mass index, inflammatory markers, or serum lipidograms during human intervention trials with different age groups at a daily dose of 10.3 or 8.17 log cfu/serving for 3 wk. No negative effect on gastrointestinal welfare was observed, but the consumption of 100g/d for 3 wk caused hard stools from the second week of the trial. The content of total lactobacilli increased in feces, and the presence of the ingested probiotic strain was confirmed after the consumption of cheese. Thus, L. plantarum strain Tensia is suitable for generally recognized as safe (GRAS) and qualified presumption of safety (QPS) criteria because it did not have any undesirable characteristics. The regular semihard Edam-type cheese (fat content of 26%) with the probiotic additive at a daily dose of 50 g or in excess (100g) and with a probiotic daily dose of 10 log cfu for 3 wk was safe.

Antibiotic susceptibility patterns of community- and hospital-acquired Staphylococcus aureus and Escherichia coli in Estonia.

This study compares the susceptibility patterns of Staphylococcus aureus and Escherichia coli isolated from patients with hospital-acquired and outpatient infections. A total of 902 isolates of S. aureus and 1,114 of E. coli were collected in five different Estonian medical centers between January 1997 and November 1997. Strains were grouped into two different categories, depending on whether they had been obtained from inpatients or outpatients. Compared to S. aureus strains isolated from inpatients, the strains from outpatients were significantly more resistant to erythromycin (25.3% vs. 17.9%), tetracycline (33.5% vs. 22.4%) and trimethoprim-sulfamethoxazole (13.9% vs. 7.9%). The overall prevalence of oxacillin-resistant S. aureus was 10.4%, with no significant differences noted between isolates recovered from inpatients and outpatients. In the case of E. coli, significantly more isolates from inpatients (42.8%) than from outpatients (34.4%) were ampicillin-resistant. Inpatient isolates of E. coli were also more resistant to cefotaxime (9.3%) and nitrofurantoin (11.2%) than outpatient strains (0% and 3.1%, respectively). Analysis showed remarkable co-resistance among both inpatient and outpatient strains of S. aureus and E. coli. Multiple resistant S. aureus and E. coli strains represented 15.1% and 17.3%, respectively of the organisms examined in this study. With respect to E. coli, significantly more multiresistant isolates were found in inpatient than outpatient isolates (20.4% vs. 8.9%). Our results indicate that the distinction between community-acquired and hospital infections due to S. aureus and E. coli may not be valid in Estonia.