Sarka Musilova

Bifidobacteria from the gastrointestinal tract of animals: differences and similarities

At present, the genus Bifidobacterium includes 48 species and subspecies, and this number is expected to increase. Bifidobacteria are found in different ecological niches. However, most were originally isolated from animals, mainly mammals, especially during the milk feeding period of life. Their presence in high numbers is associated with good health of the host. Moreover, bifidobacteria are often found in poultry and insects that exhibit a social mode of life (honeybees and bumblebees). This review is designed as a summary of currently known species of the genus Bifidobacterium, especially focused on their difference and similarities. The primary focus is on their occurrence in the digestive tract of animals, as well as the specificities of animal strains, with regard to their potential use as probiotics.

Isolation and characterization of bifidobacteria from ovine cheese

Animal products are one of the niches of bifidobacteria, a fact probably attributable to secondary contamination. In this study, 2 species of the genus Bifidobacterium were isolated by culture-dependent methods from ovine cheeses that were made from unpasteurized milk without addition of starter cultures. The isolates were identified as Bifidobacterium crudilactis and Bifidobacterium animalis subsp. lactis using matrix-assisted laser desorption/ionization time-of-flight analysis and sequencing of phylogenetic markers (16S rRNA, hsp60, and fusA).

Comparison of mupirocin-based media for selective enumeration of bifidobacteria in probiotic supplements

An international standard already exists for the selective enumeration of bifidobacteria in milk products. This standard uses Transgalactosylated oligosaccharides (TOS) propionate agar supplemented with mupirocin. However, no such standard method has been described for the selective enumeration of bifidobacteria in probiotic supplements, where the presence of bifidobacteria is much more variable than in milk products. Therefore, we enumerated bifidobacteria by colony count technique in 13 probiotic supplements using three media supplemented with mupirocin (Mup; 100mg/l): TOS, Bifidobacteria selective medium (BSM) and modified Wilkins-Chalgren anaerobe agar with soya peptone (WSP). Moreover, the potential growth of bifidobacterial strains often used in probiotic products was performed in these media. All 13 products contained members of the genus Bifidobacterium, and tested mupirocin media were found to be fully selective for bifidobacteria. However, the type strain Bifidobacterium bifidum DSM 20456 and collection strain B. bifidum DSM 20239 showed statistically significant lower counts on TOS Mup media, compared to BSM Mup and WSP Mup media. Therefore, the TOS Mup medium recommended by the ISO standard cannot be regarded as a fully selective and suitable medium for the genus Bifidobacterium. In contrast, the BSM Mup and WSP Mup media supported the growth of all bifidobacterial species.

Raffinose-Series Oligosaccharides in Soybean Products

Abstract Soybean foods forming a substantial part of Asian diet have still more expanded into European diet. Raffinose-series oligosaccharides (RSO) are important constituents of soya beans and they can be found also in soybean products. These oligosaccharides can be considered potentially prebiotic for their capability of influencing the composition of the host’s intestinal microbiota. The aim of the present paper was to determine the oligosaccharide content in various soybean products. Enzymatic assay has been used for the determination of oligosaccharides. RSO have been found in all tested samples and their content varied from 0.66 g per 100 g in soybean beverage to 5.59 g per 100 g in first clear soybean flour. Generally, the highest content of RSO has been detected in soybean flour in the average amount of 4.83 g per 100 g. There was no statistically significant difference observed in the amount of oligosaccharides in all four types of soybean flour (P < 0.01). Considerably high amounts of RSO have been found in sweet soybean bars and textured soy protein. Foods as soybean flour and soybean bar ‘Sójový suk’ seem to be effective natural sources of prebiotic oligosaccharides for humans.

Colonisation of the gut by bifidobacteria is much more common in vaginal deliveries than Caesarean sections.

Micro-organisms start to colonise the infant gut during the first days of life and play an important role in human health throughout life (1). More than 10 bacteria per gram of intestinal content present a barrier against colonisation by pathogens and alien microbes. They are involved in metabolism by degrading nondigestible food remnants, producing vitamins B and K and participating in short-chain fatty-acid metabolism. These bacteria also play a role in the stimulation and development of the immune system. Therefore, the colonisation of a newborn infant’s gut is vital and it has a significant influence on the final composition of the resident microbiota in adults. Beneficial bacteria in the intestinal tract, such as bifidobacteria and lactobacilli, contribute to improved health for months, years or even a lifetime (2,3). The colonisation of the human gut is a complicated process that is dependent on a number of factors, which include the duration of pregnancy, the mother’s health, gestational age, antibiotic treatment, hospital hygiene, duration and mode of delivery and type of feeding (4,5). The most significant change in the composition of intestinal microbiota occurs in the first few weeks of life. Bifidobacteria are the predominant group of bacteria in the gut of breastfed and vaginally delivered infants (6). The aim of this study was to determine the influence of the mode of delivery on infant gut colonisation by bifidobacteria. In addition, we sought to identify the predominant gut bacteria of infants born by Caesarean section. The influence of the mode of delivery – Caesarean section versus vaginal delivery – on the microbial composition of faecal samples from infants aged 10–30 days was examined. We analysed 100 faecal samples from healthy infants of both sexes from the Paediatric Department of Motol University Hospital, Prague, Czech Republic. Of these 100 infants, 61 (36 male and 25 female) were born by vaginal delivery and 39 (20 male and 19 female) were born by Caesarean section. All study subjects were full-term infants who were born in the hospital, and all were exclusively breastfed. None of them had been treated with antibiotics or probiotics before sampling. Samples were collected by their mothers, who had also not been treated with antibiotics during pregnancy. Fresh faecal samples were aseptically transferred to tubes containing Wilkins–Chalgren broth (Oxoid), transported to the laboratory and analysed within two hours. The samples were serially diluted in the Wilkins–Chalgren broth under anaerobic conditions. Media were prepared using the rolltube technique in an oxygen-free carbon dioxide environment. Faecal bacteria were detected using selective media, according to the method described by Vlkov a et al. (7). Appropriate dilutions of the sample were transferred to sterile petri dishes that were immediately filled with media selective for total anaerobes (Wilkins–Chalgren agar, Oxoid), bifidobacteria (TPY agar from Scharlau, Spain, modified by the addition of 100 mg/L of mupirocin and 1 mL/L of acetic acid), lactobacilli (Rogosa agar, Oxoid), gram-negative anaerobes (Wilkins–Chalgren agar, supplemented with G-N Anaerobe Selective Supplement, both Oxoid) and Escherichia coli (TBX, Oxoid). Total anaerobes, gram-negative anaerobes and bifidobacteria were incubated in anaerobic jars (Anaerobic Plus System, Oxoid) at 37°C for 48 h. Lactobacilli were cultivated in microaerophilic

Identification of microbiota associated with Pectinatella magnifica in South Bohemia

Abstract The bacterial diversity of Pectinatella magnifica colonies sampled from pounds in South Bohemia during the summer of 2012 was investigated. The bacterial counts determined after cultivation on modified yeast extract-tryptone agar (Oxoid) supplemented with glucose (1 g L−1) varied from 4.22 to 6.61 and from 1.30 to 6.85 log CFU/g for aerobes and anaerobes, respectively. Higher counts were found in the superficial structures of Pectinatella colonies than in the inner gelled mass. Neither a trend in bacterial numbers at the individual site during the season, nor correlations between bacterial counts in P. magnifica and the surrounding water were observed. Fifty-four isolates were identified by sequencing the 16S rRNA gene and through MALDI-TOF MS analysis. Species of Aeromonas and Aquitalea were the predominantly isolated bacteria, but members of Chryseobacterium, Herbaspirillum, Enterobacter, Lactococcus, Leuconostoc, Pseudomonas and Sphingomonas were also found. As listed genera are wildly distributed in different water, soil, and plant samples, we conclude thatPectinatella colonies are inhabited by environmental bacteria. Nevertheless, a symbiotic relationship of these bacteria with P. magnifica cannot be excluded.

Mupirocin-mucin agar for selective enumeration of Bifidobacterium bifidum.

Bifidobacterium bifidum is a bacterial species exclusively found in the human intestinal tract. This species is becoming increasingly popular as a probiotic organism added to lyophilized products. In this study, porcine mucin was used as the sole carbon source for the selective enumeration of B. bifidum in probiotic food additives. Thirty-six bifidobacterial strains were cultivated in broth with mucin. Only 13 strains of B. bifidum utilized the mucin to produce acids. B. bifidum was selectively enumerated in eight probiotic food supplements using agar (MM agar) containing mupirocin (100 mg/L) and mucin (20 g/L) as the sole carbon source. MM agar was fully selective if the B. bifidum species was presented together with Bifidobacterium animalis subsp. lactis, Bifidobacterium breve, and Bifidobacterium longum subsp. longum species and with lactic acid bacteria (lactobacilli, streptococci). Isolated strains of B. bifidum were identified using biochemical, PCR, MALDI-TOF procedures and 16S rRNA gene sequencing. The novel selective medium was also suitable for the isolation of B. bifidum strains from human fecal samples.

Bifidobacteria, Lactobacilli, and Short Chain Fatty Acids of Vegetarians and Omnivores

Abstract The intestinal microbiota represents the largest and the most complex microbial community inhabiting the human body. Bifidobacteria and lactobacilli represent important commensal bacteria with the ability to utilize complex carbohydrates. The main fermentation products from the breakdown of complex dietary carbohydrates are short chain fatty acids (SCFAs). We examined faecal samples of vegetarians (n = 10) and conventional omnivores (n = 10) to evaluate the counts and occurrence of cultivable bacteria, especially bifidobacteria and lactobacilli, using cultivation on selective media, and matrix-assisted laser desorption/ionization time-of-flight. Moreover, concentrations and molar proportion of SCFAs in faecal samples were measured. Total counts of Gram-negative anaerobic bacteria were significantly lower (P < 0.05) in vegetarian faecal samples, while others (total anaerobic bacteria, Bifidobacterium spp., Lactobacillus spp., Escherichia coli, and presumptive coliforms) were not. Neither total concentrations nor molar proportions of SCFAs in faecal samples differed (P > 0.05) between the diet groups. In total, six Bifidobacterium spp. and thirteen Lactobacillus spp. were detected via culture-dependent methods. Bifidobacteria counts and species composition in faecal samples of both groups were found to be relatively similar, regardless of the diet. Lactobacillus species varied more by individual diet.

Assessment of the synbiotic properites of human milk oligosaccharides and Bifidobacterium longum subsp. infantis in vitro and in humanised mice

The mode of delivery plays a crucial role in infant gastrointestinal tract colonisation, which in the case of caesarean section is characterised by the presence of clostridia and low bifidobacterial counts. Gut colonisation can be modified by probiotics, prebiotics or synbiotics. Human milk oligosaccharides (HMOs) are infant prebiotics that show a bifidogenic effect. Moreover, genome sequencing of Bifidobacterium longum subsp. infantis within the infant microbiome revealed adaptations for milk utilisation. This study aimed to evaluate the synbiotic effect of B. longum subsp. infantis, HMOs and human milk (HM) both in vitro and in vivo (in a humanised mouse model) in the presence of faecal microbiota from infants born by caesarean section. The combination of B. longum and HMOs or HM reduced the clostridia and G-bacteria counts both in vitro and in vivo. The bifidobacterial population in vitro significantly increased and produce high concentrations of acetate and lactate. In vitro competition assays confirmed that the tested bifidobacterial strain is a potential probiotic for infants and, together with HMOs or HM, acts as a synbiotic. It is also able to inhibit potentially pathogenic bacteria. The synbiotic effects identified in vitro were not observed in vivo. However, there was a significant reduction in clostridia counts in both experimental animal groups (HMOs + B. longum and HM + B. longum), and a specific immune response via increased interleukin (IL)-10 and IL-6 production. Animal models do not perfectly mimic human conditions; however, they are essential for testing the safety of functional foods.