V. Rada

Screening of some Siberian medicinal plants for antimicrobial activity

The antimicrobial activity of crude ethanolic extracts of 16 Siberian medicinal plants was tested against five species of microorganisms: Bacillus cereus, Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans. Of the 16 plants tested, 12 showed antimicrobial activity against one or more species of microorganisms. The most active antimicrobial plants were Bergenia crassifolia, Chelidonium majus, Rhaponticum carthamoides, Sanguisorba officinalis, and Tussilago farfara.

A new selective medium for the isolation of glucose non-fermenting bifidobacteria from hen caeca.

Avian caeca contain a large and diverse population of bacteria. Certain genera, including Bifidobacterium, are thought to exert health-promoting effects. Two media were evaluated to determine their sensitivity and selectivity for bifidobacteria in the hen caecal samples: modified Wilkins-Chalgren agar (MW; Oxoid) with the addition of glacial acetic acid (1 ml/l) and mupirocin (100 mg/l) and modified TPY agar (MTPY; ADSA, Spain) with glacial acetic acid (1 ml/l) and mupirocin (100 mg/l). The colonies arising on the plates inoculated with caecal samples were Gram stained, screened for the presence of fructose-6-phosphate phosphoketolase activity, and tested for fermentation patterns using ANAEROtest (Lachema, Czech Republic) and API 50 CHL (BioMérieux, France) kits. Both agars were selective for bifidobacteria, however, MTPY agar showed higher cfu/g than MW agar. Bifidobacterial counts were higher than 10(10) cfu/g of caecal contents using MTPY agar. Most of strains isolated from this medium fermented melibiose, sucrose, and raffinose, but not glucose. Soya peptone (5 g/l; Oxoid) stimulated the growth of glucose non-fermenting strains in complex liquid media. The results suggest that the media for selective enumeration and isolation of bifidobacteria in poultry caecal samples should not contain glucose as the sole carbon source. It can be concluded that MTPY medium is highly selective and permits the growth of both glucose fermenting and glucose non-fermenting bifidobacteria.

Bifidobacterium bombi sp. nov., from the bumblebee digestive tract.

Gram-positive-staining, anaerobic, non-spore-forming, lactate- and acetate-producing bacterial strains were isolated from the digestive tracts of different bumblebee species (Bombus lucorum, Bombus pascuorum and Bombus lapidarius). All of the isolates produced fructose-6-phosphate phosphoketolase activity. A representative strain, BluCI/TPT, was characterized further. Cells of strain BluCI/TPT showed occasional bifurcation and irregular constrictions. The bacterium utilized a wide range of carbohydrates. Glucose was fermented to acetate and lactate. The DNA base composition was 47.2 mol% G+C. Complete 16S rRNA and partial hsp60 gene sequences were obtained and phylogenetic relationships were determined. Strain BluCI/TPT and related isolates were located in the actinobacterial cluster and were closely related to the genera Bifidobacterium, Scardovia, Aeriscardovia and Parascardovia. The results presented support the proposal of a novel species to accommodate strain BluCI/TPT, with the name Bifidobacterium bombi sp. nov.; the type strain is BluCI/TPT (=DSM 19703T=ATCC BAA-1567T).

Susceptibility ofEscherichia coli to C2-C18 fatty acids

The antimicrobial activity of C2–C18 fatty acids was determinedin vitro in cultures of two strains ofEscherichia coli grown on glucose. Antimicrobial activity was expressed as IC50 (a concentration at which only 50 % of the initial glucose in the cultures was utilized). Utilization of glucose was inhibited by caprylic acid (IC50 0.30–0.85 g/L) and capric acid (IC50 1.25–2.03 g/L). Neither short-chain fatty acids (C2–C6) nor fatty acids with longer chain (C12–C18) influenced substrate utilization. Caproic acid, however, decreased cell yield in cultures ofE. coli in a dose-dependent manner. No inhibition of glucose utilization was produced with unsaturated fatty acids, oleic and linoleic. Calcium ions added in excess reversed the antimicrobial effect of capric acid, but not that of caprylic acid. Antimicrobial activity of caprylic and capric acid decreased when the bacteria were grown in the presence of straw particles, or repeatedly subcultured in a medium containing these compounds at low concentrations. Counts of viable bacteria determined by plating decreased after incubation with caprylic and capric acid (30 min; 1 g/L) at pH 5.2 from >109 to ≈102/mL. A reduction of a mere 0.94–1.96 log10 CFU was observed at pH 6.5–6.6. It can be concluded that caprylic acid, and to a lesser extent also capric acid, has a significant antimicrobial activity towardE. coli. Effects of other fatty acids were not significant or absent.

In vitro growth-inhibitory effect of plant-derived extracts and compounds against Paenibacillus larvae and their acute oral toxicity to adult honey bees.

In total, 26 natural compounds of various chemical classes (flavonoids, alkaloids, terpenoids) and 19 crude extracts from selected plants were tested in vitro for antibacterial activity against three strains of P. larvae, the causal agent of American Foulbrood Disease of honey bees (AFB) by the broth microdilution method. Among the individual substances, sanguinarine (MIC 4 microg/ml), followed by thymoquinone, capsaicin, trans-2-hexenal and nordihydroguaiaretic acid (MIC 4-32 microg/ml) possessed the strongest antibacterial effect. In case of extracts, common hop (Humulus lupulus L.) and myrtle (Myrtus communis L.) methanolic-dichloromethane extracts exhibited the highest growth-inhibitory effect with MICs ranging from 2 to 8 microg/ml. Acute oral toxicity of the most active natural products was determined on adult honey bees, showing them as non-toxic at concentrations as high as 100 microg peer bee. Our study leads to identification of highly potent natural products effective against AFB in vitro with very low MICs compared to those reported in literature, low toxicity to adult honey bees and commercial availability suggesting them as perspective, low cost and consumer-acceptable agents for control of AFB.

Distribution of bifidobacteria in the gastrointestinal tract of calves

Development of gastrointestinal microflora of calves with special reference to bifidobacteria was investigated; fecal bacteria were enumerated in calves aged 3 days to 7 weeks. Bacteria were detected by using selective media, bifidobacteria using modified TPY agar with an addition of mupirocin and acetic acid and by fluorescencein situ hybridization (FISH). Bifidobacteria were dominant group of fecal flora of calves after 7 d of life, constituting 10 % of total bacterial counts. The highest bacterial concentrations were observed in rumen, cecum, and colon, the lowest in abomasum and duodenum. Bifidobacteria and lactobacilli exhibited the highest survival ability during stomach passage and dominated in all parts of the digestive tract. Bifidobacteria counts determined by FISH were significantly higher than those provided by cultivation. Modified TPY agar was highly selective and suitable for bifidobacteria isolation but FISH was shown to be a more precise method for their enumeration. Our results show that gastrointestinal microflora of calves in the milk-feeding period is similar to breast-fed infants with respect to the occurrence of bifidobacteria as a dominant bacterial group. The use ofBifidobacterium strains offers a promising way for providing beneficial effectors for calves in the milk-feeding period.

Xanthine oxidase inhibitory properties of Czech medicinal plants.

AIM OF THE STUDY To investigate in vitro xanthine oxidase inhibitory properties of plants traditionally used in Czech Republic and Central-East Europe region for gout, arthritis or rheumatism treatment. MATERIALS AND METHODS Methylene chloride-methanolic and two ethanolic extracts of 27 plant species were screened for in vitro xanthine oxidase inhibitory activity using a spectrophotometric method. RESULTS Around 50% of the species exhibited some degree of xanthine oxidase inhibitory properties at 200 μg/mL, showing a moderate correlation (r=0.59) with total phenol content. The most active were methylene chloride-methanolic extracts of Populus nigra and Betula pendula, with IC(50) of 8.3 and 25.9 μg/mL, respectively, followed by 80% ethanolic extract of Caryophyllus aromaticus and Hypericum perforatum, both under 50 μg/mL. CONCLUSIONS Populus nigra and Betula pendula were identified as species with the highest xanthine oxidase inhibitory potential in our study. This correlates with the ethnobotanical data on their use in Central European folklore and provides the basis for further investigation on these plants.

Cross-talk of human gut with bifidobacteria.

The gut constitutes a prominent part of the immune system. Its commensal microflora plays an important role in defense and in tolerance to diet allergens. Disturbances in immune regulations may lead to food allergy. Among commensal bacteria, bifidobacteria are able to induce mechanisms of immune tolerance. Comprehension of their mutual cross-talk with the host is necessary for understanding their role in the diet and in food supplements.

Auto-Aggregation and Co-Aggregation Ability in Bifidobacteria and Clostridia

A total of 142 human and 88 calf bifidobacteria were isolated and identified; ≈12 % of all isolated strains exhibited auto-aggregation (Agg) phenotype (Agg+). Properties considered to be predicting for their adhesion to intestine, i.e. auto-aggregation, and hydrophobicity were determined by xylene extraction in 18 human and 8 calf origin bifidobacteria. Co-aggregation of 8 human bifidobacteria with 8 clostridia was also evaluated. Agg varied between 16.3 and 96.4 %, hydrophobicity values ranged from 0 to 82.8 %. The strongest Agg and hydrophobicity were observed in B. bifidum and B. merycicum isolates. However, there were no statistically significant correlations between these two properties. Variability in the percentage of Agg and hydrophobicity was observed after cultivation of bifidobacteria on different carbon sources. All bifidobacteria showed co-aggregation ability with clostridia tested but there were remarkable differences depending on specific combinations of strains. The bifidobacterial strains with the highest ability to co-aggregate with clostridia were B. bifidum I4 and B. longum I10 isolated from infants; these strains gave also high values of Agg. Agg properties together with co-aggregation ability with potential pathogen can be used for preliminary selection of probiotic bacteria.

Modulation of natural immunity in the gut by Escherichia coli strain Nissle 1917

The beneficial effect of probiotic Escherichia coli strain Nissle 1917 (EcN) suggests the gut epithelium plays a basic role in immune interactions with bacteria. Contrary to other commensal strains of Escherichia coli, EcN profoundly modulates the gut barrier to elevate its resistance to microbial pathogens. The present review documents the properties of EcN that have led to the protection of gnotobiotic pigs against lethal enteric infections. This effect could be important in light of the growing number of acquired deficiencies that paralyze gut immunity in humans.