In the first days of life, the newborns' intestinal microbiota develops simultaneously with the intestinal gut barrier and follows intestinal immunity. The mode of delivery shows significant impact on microbial development and, thus, the initiation of the tryptophan catabolism pathway. Further antibiotics (ATB) treatment of mothers before or during delivery affects the microbial and tryptophan metabolite composition of stool of the caesarean- and vaginal-delivered newborns. The determination of microbiome and levels of tryptophan microbial metabolites in meconium and stool can characterize intestinal colonization of a newborn. From 134 samples from the Central European Longitudinal Studies of Parents and Children: The Next Generation (CELSPAC: TNG) cohort study, 16S rRNA gene sequencing was performed, and microbial tryptophan metabolites were quantified using ultra-high-performance liquid chromatography with triple-quadrupole mass spectrometry. Microbial diversity and concentrations of tryptophan metabolites were significantly higher in stool compared to meconium. Treatment of mothers with ATB before or during delivery affects metabolite composition and microbial diversity in stool of vaginal- and caesarean-delivered newborns. Correlation of microbial and metabolite composition shows significant positive correlations of indol-3-lactic acid, N-acetyl-tryptophan and indol-3-acetic acid with Bifidobacterium, Bacteroides and Peptoclostridium. The positive effect of vaginal delivery on newborns' microbiome development is degraded when mother is treated with ATB before or during delivery. KEY POINTS: • Antibiotic treatment diminishes the positive effects of vaginal delivery. • Antibiotic treatment affects metabolite and microbial composition in newborns. • Bifidobacterium and Peptoclostridium could be the producer of indole-lactic acid.
- MeSH
- antibakteriální látky * MeSH
- Bifidobacterium metabolismus růst a vývoj MeSH
- feces * mikrobiologie chemie MeSH
- indoly metabolismus MeSH
- kyseliny indoloctové metabolismus MeSH
- lidé MeSH
- longitudinální studie MeSH
- mekonium * mikrobiologie chemie MeSH
- novorozenec MeSH
- RNA ribozomální 16S * genetika MeSH
- střevní mikroflóra * účinky léků MeSH
- tryptofan * metabolismus MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Check Tag
- lidé MeSH
- novorozenec MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
Mucus production is initiated before birth and provides mucin glycans to the infant gut microbiota. Bifidobacteria are the major bacterial group in the feces of vaginally delivered and breast milk-fed infants. Among the bifidobacteria, only Bifidobacterium bifidum is able to degrade mucin and to release monosaccharides which can be used by other gut microbes colonizing the infant gut. Eubacterium hallii is an early occurring commensal that produces butyrate and propionate from fermentation metabolites but that cannot degrade complex oligo- and polysaccharides. We aimed to demonstrate that mucin cross-feeding initiated by B. bifidum enables growth and metabolite formation of E. hallii leading to short-chain fatty acid (SCFA) formation. Growth and metabolite formation of co-cultures of B. bifidum, of Bifidobacterium breve or Bifidobacterium infantis, which use mucin-derived hexoses and fucose, and of E. hallii were determined. Growth of E. hallii in the presence of lactose and mucin monosaccharides was tested. In co-culture fermentations, the presence of B. bifidum enabled growth of the other strains. B. bifidum/B. infantis co-cultures yielded acetate, formate, and lactate while co-cultures of B. bifidum and E. hallii formed acetate, formate, and butyrate. In three-strain co-cultures, B. bifidum, E. hallii, and B. breve or B. infantis produced up to 16 mM acetate, 5 mM formate, and 4 mM butyrate. The formation of propionate (approximately 1 mM) indicated cross-feeding on fucose. Lactose, galactose, and GlcNAc were identified as substrates of E. hallii. This study shows that trophic interactions of bifidobacteria and E. hallii lead to the formation of acetate, butyrate, propionate, and formate, potentially contributing to intestinal SCFA formation with potential benefits for the host and for microbial colonization of the infant gut. The ratios of SCFA formed differed depending on the microbial species involved in mucin cross-feeding.
- MeSH
- Bifidobacterium růst a vývoj izolace a purifikace metabolismus MeSH
- dospělí MeSH
- Eubacterium růst a vývoj izolace a purifikace metabolismus MeSH
- feces mikrobiologie MeSH
- fermentace MeSH
- kojenec MeSH
- kojení MeSH
- kyseliny mastné těkavé metabolismus MeSH
- lidé MeSH
- muciny metabolismus MeSH
- střeva mikrobiologie MeSH
- střevní mikroflóra MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- kojenec MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
UNLABELLED: The need for suitable selective cultivation media for the isolation of Bifidobacterium spp. continues to be a real concern in the field of intestinal microbiology. Isolation of bifidobacteria from human and animal faecal samples using selective agar plating may be problematic especially in samples with increased clostridial counts than bifidobacterial counts. Due to the absence of anticlostridial agents in existing selective media, clostridia can displace bifidobacteria resulting in incorrect estimation of their counts. Therefore, we supplemented the existing selective medium 'modified Wilkins Chalgren agar with mupirocin' (MWM) with 90 mg l(-1) of 8-hydroxyquinoline (8HQ), which was recently proved to act selectively against clostridia. The newly composed 'modified Wilkins-Chalgren agar with 8HQ' (MWMQ) was tested on pure bifidobacterial and clostridial strains, their mixtures, and using faecal samples of mammalian origin; its selectivity was evaluated by genus-specific identification of isolates. The results demonstrated that the presence of 8HQ in this agar eliminated the growth of nonbifidobacterial strains on MWMQ compared to that on MWM, whereas the recovery of bifidobacterial counts was at satisfactory levels. In conclusion, MWMQ could be recommended for bifidobacterial isolation from human and animal faeces especially when bifidobacteria are not numerically dominant and there are chances of clostridial contamination. SIGNIFICANCE AND IMPACT OF THE STUDY: Routine isolation of bifidobacteria from mammalian faeces does not use a reliable selective agar with an anticlostridial agent. Overgrowth of clostridia may result in incorrect estimation of bifidobacterial counts. Thus, in order to improve the selectivity of existing media for bifidobacterial isolation, we chose the modified Wilkins-Chalgren agar with mupirocin and supplemented it with 8-hydroxyquinoline (8HQ), a molecule that shows anticlostridial activity without affecting the growth of bifidobacteria. This newly composed medium showed enhanced selectivity and specificity compared to the original medium and therefore, can be recommended for the isolation of bifidobacteria from mammal faeces.
- MeSH
- agar farmakologie MeSH
- bakteriální nálož účinky léků MeSH
- Bifidobacterium růst a vývoj izolace a purifikace MeSH
- Clostridium účinky léků růst a vývoj MeSH
- feces mikrobiologie MeSH
- kultivační média farmakologie MeSH
- lidé MeSH
- mupirocin farmakologie MeSH
- oxychinolin farmakologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
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.
- MeSH
- antibakteriální látky metabolismus MeSH
- bakteriální nálož metody MeSH
- Bifidobacterium účinky léků růst a vývoj MeSH
- kultivační média chemie MeSH
- mupirocin metabolismus MeSH
- počet mikrobiálních kolonií metody MeSH
- probiotika analýza MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Prebiotics are used for stimulating the growth of beneficial microorganisms in the gut. However, it is very difficult to find a suitable prebiotic mixture that exclusively supports the growth of beneficial microbes such as bifidobacteria and lactobacilli. We tested the effects of a prebiotic mixture in vitro by incubating it with fecal samples and in vivo by administration of the prebiotic supplement to healthy adult volunteers, followed by analysis of their fecal microbiota. The effect of the oligosaccharides on bacterial metabolism was studied by analyzing short-chain fatty acid (SCFA) production in vitro and the SCFA pattern for the stool samples of volunteers. In the in vitro test, a higher proportion of bifidobacteria (25.77%) was seen in the total bacterial population after cultivation on a prebiotic mixture than on the control medium (7.94%). The gram-negative anaerobe count significantly decreased from 8.70 to 6.40 log CFU/g (from 35.21% to 0.60%) and the Escherichia coli count decreased from 7.41 to 6.27 log CFU/g (from 1.78% to 0.44%). Administration of a prebiotic mixture in vivo (9 g of galactooligosaccharides [GOS]+1 g of maltodextrins; daily for 5 days) significantly increased the fecal bifidobacterial count from 9.45 to 9.83 log CFU/g (from 40.80% to 53.85% of total bacteria) and reduced the E. coli count from 7.23 to 6.28 log CFU/g (from 55.35% to 45.06% of total bacteria). The mixture comprising GOS and maltodextrins thus exhibited bifidogenic properties, promoting the performance of bifidobacteria by boosting their growth and inhibiting the growth of undesirable bacteria.
- MeSH
- Bifidobacterium účinky léků růst a vývoj metabolismus MeSH
- dospělí MeSH
- Escherichia coli účinky léků růst a vývoj metabolismus MeSH
- feces mikrobiologie MeSH
- fixní kombinace léků MeSH
- galaktosa farmakologie MeSH
- kyseliny mastné těkavé metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- oligosacharidy farmakologie MeSH
- polysacharidy farmakologie MeSH
- prebiotika * MeSH
- střeva účinky léků metabolismus mikrobiologie MeSH
- střevní mikroflóra účinky léků MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- klinické zkoušky kontrolované MeSH
- práce podpořená grantem MeSH
The polysaccharide inulin has great importance in the food and pharmaceutical industries. The degree of polymerization (DP) of inulin influences important properties, such as, solubility, thermal stability, sweetness power and prebiotic activity. Molecules with a high degree of polymerization are obtained through physical techniques for enrichment of the inulin chains because they are not commonly obtained from plants extract. Gas chromatography/Mass Spectrometry and (1)H Nuclear Magnetic Resonance analysis showed that inulin from Stevia rebaudiana roots has a degree of polymerization (DPn 28) higher than the value of DPn 12-15 for inulins from other plant species. Furthermore, the methodology of freeze/thaw to enrich the chains allowed us to increase the DP, similarly to other methodologies used for the enrichment of inulin chains. The prebiotic assays confirm that inulin from S. rebaudiana has a high DP. The combined use of these molecules with low degree of polymerization fructans seems to be advantageous to prolong the prebiotic effect in the colon. Our results suggest that S. rebaudiana roots are a promising source of high degree polymerization inulins.
Various culture media have been proposed for the isolation and selective enumeration of bifidobacteria. Mupirocin is widely used as a selective factor along with glacial acetic acid. TOS (transgalactosylated oligosaccharides) medium supplemented with mupirocin is recommended by the International Dairy Federation for the detection of bifidobacteria in fermented milk products. Mupirocin media with acetic acid are also reliable for intestinal samples in which bifidobacteria predominate. However, for complex samples containing more diverse microbiota, the selectivity of mupirocin media is limited. Resistance to mupirocin has been demonstrated by many anaerobic bacteria, especially clostridia. The objective was to identify an antibiotic that inhibits the growth of clostridia and allows the growth of bifidobacteria, and to use the identified substance to develop a selective cultivation medium for bifidobacteria. The susceptibility of bifidobacteria and clostridia to 12 antibiotics was tested on agar using the disk diffusion method. Only norfloxacin inhibited the growth of clostridia and did not affect the growth of bifidobacteria. Using both pure cultures and faecal samples from infants, adults, calves, lambs, and piglets, the optimal concentration of norfloxacin in solid cultivation media was determined to be 200 mg/L. Our results showed that solid medium containing norfloxacin (200 mg/L) in combination with mupirocin (100 mg/L) and glacial acetic acid (1 mL/L) is suitable for the enumeration and isolation of bifidobacteria from faecal samples of different origins.
- MeSH
- bakteriologické techniky metody MeSH
- Bifidobacterium růst a vývoj izolace a purifikace MeSH
- dospělí MeSH
- kojenec MeSH
- kultivační média chemie MeSH
- kyselina octová metabolismus MeSH
- lidé středního věku MeSH
- lidé MeSH
- lodě MeSH
- mikrobiální testy citlivosti MeSH
- mupirocin metabolismus MeSH
- norfloxacin metabolismus MeSH
- prasata MeSH
- skot MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- kojenec MeSH
- lidé středního věku MeSH
- lidé MeSH
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
The major risk factor for Clostridium difficile infection (CDI) is the use of antibiotics owing to the disruption of the equilibrium of the host gut microbiota. To preserve the beneficial resident probiotic bacteria during infection treatment, the use of molecules with selective antibacterial activity enhances the efficacy by selectively removing C. difficile. One of them is the plant alkaloid 8-hydroxyquinoline (8HQ), which has been shown to selectively inhibit clostridia without repressing bifidobacteria. Selective antimicrobial activity is generally tested by culture techniques of individual bacterial strains. However, the main limitation of these techniques is the inability to describe differential growth dynamics of more bacterial strains in co-culture within the same experiment. In the present study, we combined fluorescent in situ hybridization and flow cytometry to describe the changes in active and non-active cells of a mixed culture formed by the opportunistic pathogen C. difficile CECT 531 and the beneficial Bifidobacterium longum subsp. longum CCMDMND BL1 after exposure to 8HQ. It was observed that without 8HQ, the proportion of both strains was almost equal, oscillating between 22.7 and 77.9 % during a time lapse of 12 h, whereas with 8HQ the proportion of active C. difficile decreased after 4 h, and persisted only between 8.8 and 17.5 %. In contrast, bifidobacterial growth was not disturbed by 8HQ. The results of this study showed the selective inhibitory effect of 8HQ on clostridial and bifidobacterial growth dynamics, and the potential of this compound for the development of selective agents to control CDIs.
- MeSH
- antibakteriální látky farmakologie MeSH
- Bifidobacterium účinky léků růst a vývoj MeSH
- Clostridioides difficile účinky léků růst a vývoj MeSH
- hybridizace in situ fluorescenční MeSH
- mikrobiální interakce MeSH
- oxychinolin farmakologie MeSH
- průtoková cytometrie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- Bifidobacterium růst a vývoj MeSH
- fermentace MeSH
- funkční potraviny MeSH
- geneticky modifikované potraviny * škodlivé účinky využití MeSH
- jogurt * mikrobiologie MeSH
- lidé MeSH
- probiotika klasifikace terapeutické užití MeSH
- škrob analýza aplikace a dávkování škodlivé účinky MeSH
- Check Tag
- lidé MeSH
The main goal in reversing the allergy epidemic is the development of effective prophylactic strategies. We investigated the prophylactic effect of neonatal mother-to-offspring mono-colonization with Bifidobacterium longum ssp. longum CCM 7952 on subsequent allergic sensitization. Adult male and female germ-free (GF) mice were mono-colonized with B. longum, mated and their offspring, as well as age-matched GF controls, were sensitized with the major birch pollen allergen Bet v 1. Furthermore, signaling pathways involved in the recognition of B. longum were investigated in vitro. Neonatal mono-colonization of GF mice with B. longum suppressed Bet v 1-specific IgE-dependent β-hexosaminidase release as well as levels of total IgE and allergen-specific IgG2a in serum compared to sensitized GF controls. Accordingly, Bet v 1-induced production of both Th1- and Th2-associated cytokines in spleen cell cultures was significantly reduced in these mice. The general suppression of Bet v 1-specific immune responses in B. longum-colonized mice was associated with increased levels of regulatory cytokines IL-10 and TGF-β in serum. In vitro, B. longum induced low maturation status of bone marrow-derived dendritic cells and production of IL-10 in TLR2-, MyD88-, and MAPK-dependent manner. Our data demonstrate that neonatal mono-colonization with B. longum reduces allergic sensitization, likely by activation of regulatory responses via TLR2, MyD88, and MAPK signaling pathways. Thus, B. longum might be a promising candidate for perinatal intervention strategies against the onset of allergic diseases in humans.
- MeSH
- alergie prevence a kontrola MeSH
- antigeny rostlinné imunologie MeSH
- Bifidobacterium růst a vývoj MeSH
- cytokiny sekrece MeSH
- gastrointestinální trakt mikrobiologie MeSH
- gnotobiologické modely MeSH
- hexosaminidasy metabolismus MeSH
- imunizace MeSH
- imunoglobulin E krev MeSH
- imunologická tolerance MeSH
- modely nemocí na zvířatech MeSH
- myši inbrední BALB C MeSH
- myši MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
