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1.
While looking for new means to limit the dissemination of antibiotic resistance, we evaluated the role of potentially probiotic bifidobacteria on the transfer of resistance genes between enterobacteria. Transfers of bla genes encoding extended-spectrum β-lactamases (SHV-5 and CTX-M-15) were studied in the absence or presence of bifidobacteria. In vitro, transfer frequencies of these bla genes decreased significantly in the presence of three of five tested strains, i.e., Bifidobacterium longum CUETM-89-215, Bifidobacterium bifidum CIP-56.7T, and Bifidobacterium pseudocatenulatum CIP-104168T. Four transfer experiments were conducted in the digestive tract of gnotobiotic mice, the first three observing the effect of B. longum CUETM-89-215, B. bifidum CIP-56.7T, and B. pseudocatenulatum CIP-104168T on blaSHV-5 transfer and the fourth experiment studying the effect of B. bifidum CIP-56.7T on blaCTX-M-15 transfer. These experiments revealed significant decreases in the transconjugant levels (up to 3 logs) in mice having received B. bifidum CIP-56.7T or B. pseudocatenulatum CIP-104168T compared to control mice. Bifidobacteria appear to have an inhibitory impact on the transfer of antibiotic resistance genes. The inhibitory effect is associated to specific bifidobacterial strains and may be related to the production of thermostable metabolites by these strains.  相似文献   

2.
Three multiplex polymerase chain reactions (PCRs) targeted on Bifidobacterium and related species were designed to identify human species. The selected primers yielded amplified products of various sizes, each specific for a species. Three to four pairs were gathered in one PCR reaction and their specificity under multiplex conditions was confirmed using DNA from 26 reference strains. Using this technique on unidentified faecal strains, B. bifidum, B. longum and B. breve species were commonly recovered in infants while B. adolescentis, B. catenulatum/B. pseudocatenulatum continuum and B. longum species were predominant in adults. Thus, a single PCR can provide the assignment of a strain to one these species, reducing the number of PCR reactions and hands-on time for the identification of human isolates of bifidobacteria. Moreover, this technique is also applicable for the in situ detection of bifidobacteria in DNA extracts from human stools.  相似文献   

3.
AIMS: The aim of the present study was to compare several molecular methods for the identification and genotyping of bifidobacteria, and further to investigate genetic heterogeneity and functional properties of bifidobacterial isolates from intestinal samples of Finnish adult subjects. METHODS AND RESULTS: A total of 153 intestinal bifidobacterial isolates were included in initial screening and 34 isolates were further characterized. Identification results obtained with PCR-ELISA and ribotyping were well in accordance with each other, while randomly amplified polymorphic DNA (RAPD) gave tentative identification only to Bifidobacterium bifidum and to 65% of the B. longum isolates. The most commonly detected species were B. longum biotype longum followed by B. adolescentis and B. bifidum. In addition, B. animalis (lactis), B. angulatum and B. pseudocatenulatum were found. Ribotyping and pulsed-field gel electrophoresis (PFGE) proved to be discriminatory methods for bifidobacteria, but also RAPD revealed intraspecies heterogeneity. Besides two B. animalis (lactis) isolates with very close similarity to a commercially available probiotic strain, none of the intestinal isolates showed optimal survival in all tolerance (acid, bile and oxygen) or growth performance tests. CONCLUSIONS: Several species/strains of bifidobacteria simultaneously colonize the gastrointestinal tract of healthy Finnish adults and intestinal Bifidobacterium isolates were genetically heterogeneous. Functional properties of bifidobacteria were strain-dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: Applicability of ribotyping with the automated RiboPrinter System for identification and genotyping of bifidobacteria was shown in the present study.  相似文献   

4.
Twenty-four Bifidobacterium strains were examined for their ability to bind to immobilized human and bovine intestinal mucus glycoproteins. Each of the tested bacteria exhibited its characteristic adhesion to human and bovine fecal mucus. No significant differences were found among the taxonomic species. Among the tested bacteria, B. adolescentis, B. angulatum, B. bifidum, B. breve, B. catenulatum, B. infantis, B. longum and B. pseudocatenulatum adhered to human fecal mucus better than bovine fecal mucus, while the binding of B. animalis and B. lactis was not preferential. These results suggest that the mucosal adhesive properties of bifidobacteria may be a strain dependent feature, and the mucosal binding of the human bifidobacteria may be more host specific.  相似文献   

5.
The immunological study of aqueous buffer extracts obtained from 45 strains of bifidobacteria belonging to the species B. bifidum, B. longum, B. adducens, B. breve, B. infantis and B. parvulorum was made. This study revealed 3 levels of the immunological specificity of soluble bifidobacterial proteins: common to the genus Bifidobacterium, common to a limited number of strains belonging to one or several species of bifidobacteria and strain-specific.  相似文献   

6.
This study reports the development of several cloning vectors for bifidobacteria based on the replicon of pBC1, a cryptic plasmid from Bifidobacterium catenulatum L48 thought to replicate via the theta mode. These vectors, in which antibiotic resistance genes encoding either erythromycin or tetracycline resistance acted as selection markers, were able to replicate in a series of eight Bifidobacterium species at frequencies ranging from 4.0 x 10(1) to 1.0 x 10(5) transformants microg(-1) but not in Lactococcus lactis or Lactobacillus casei. They showed a relative copy number of around 30 molecules per chromosome equivalent and a good segregational stability, with more than 95% of the cells retaining the vectors after 80 to 100 generations in the absence of selection. Vectors contain multiple cloning sites of different lengths, and the lacZalpha peptide gene was introduced into one of the molecules, thus allowing the easy selection of colonies harboring recombinant plasmids in Escherichia coli. The functionality of the vectors for engineering Bifidobacterium strains was assessed by cloning and examining the expression of an alpha-l-arabinofuranosidase gene belonging to Bifidobacterium longum. E. coli and Bifidobacterium pseudocatenulatum recombinant clones were stable and showed an increase in alpha-arabinofuranosidase activity of over 100-fold compared to that of the untransformed hosts.  相似文献   

7.
The composition of vaginal bifidoflora in 56 clinically healthy women of reproductive age was studied. The study revealed that four species of bifidobacteria, viz. Bifidobacterium bifidum, B. breve, B. adolescentis 2 and B. longum, dominated in the composition of this bifidobacterial population. Nine out of 11 isolated strains were found to be capable of inhibiting indicator microorganisms Staphylococcus aureus and Enterococcus faecalis when tested in vitro; in addition, strains B. adolescentis 2 F1, B. bifidum G1, B. breve P2 and B. longum Z4 inhibited Klebsiella ozaenae, Pseudomonas aeruginosa, Escherichia coli and were also active acid producers. Three of these 4 bifidobacterial strains were capable of adhesion to vaginal epitheliocytes, while B. bifidum G1 was practically incapable of adherence to these cells, similarly to B. bifidum strain 791 of intestinal origin. In addition, the spectra of antibiotic susceptibility varied from strain to strain, but all bifidobacterial strains were susceptible to benzylpenicillin and resistant to lomefloxacin, most of them being also resistant to cyprofloxacin and gentamicin. Thus the data presented in this work are indicative of the possibility and advantages of using bifidobacterial strains belonging to this ecological niche as probiotics for the correction of the microflora of the urogenital tract in females.  相似文献   

8.
The bifidogenic effect of human milk oligosaccharides (HMOs) has long been known, yet the precise mechanism underlying it remains unresolved. Recent studies show that some species/subspecies of Bifidobacterium are equipped with genetic and enzymatic sets dedicated to the utilization of HMOs, and consequently they can grow on HMOs; however, the ability to metabolize HMOs has not been directly linked to the actual metabolic behavior of the bacteria. In this report, we clarify the fate of each HMO during cultivation of infant gut-associated bifidobacteria. Bifidobacterium bifidum JCM1254, Bifidobacterium longum subsp. infantis JCM1222, Bifidobacterium longum subsp. longum JCM1217, and Bifidobacterium breve JCM1192 were selected for this purpose and were grown on HMO media containing a main neutral oligosaccharide fraction. The mono- and oligosaccharides in the spent media were labeled with 2-anthranilic acid, and their concentrations were determined at various incubation times using normal phase high performance liquid chromatography. The results reflect the metabolic abilities of the respective bifidobacteria. B. bifidum used secretory glycosidases to degrade HMOs, whereas B. longum subsp. infantis assimilated all HMOs by incorporating them in their intact forms. B. longum subsp. longum and B. breve consumed lacto-N-tetraose only. Interestingly, B. bifidum left degraded HMO metabolites outside of the cell even when the cells initiate vegetative growth, which indicates that the different species/subspecies can share the produced sugars. The predominance of type 1 chains in HMOs and the preferential use of type 1 HMO by infant gut-associated bifidobacteria suggest the coevolution of the bacteria with humans.  相似文献   

9.
16SrDNA-targeted genus- and species-specific PCR primers have been developed and used for the identification and detection of bifidobacteria. These primers cover all of the described species that inhabit the human gut, or occur in dairy products. Identification of cultured bifidobacteria using PCR primer pairs is rapid and accurate, being based on nucleic acid sequences. Detection of bifidobacteria can be achieved using DNA extracted from human faeces as template in PCR reactions. We have found that, in adult faeces, the Bifidobacterium catenulatum group was the most commonly detected species, followed by Bifidobacterium longum, Bifidobacterium adolescentis, and Bifidobacterium bifidum. In breastfed infants, Bifidobacterium breve was the most frequently detected species, followed by Bifidobacterium infantis, B. longum and B. bifidum. It was notable that the B. catenulatum group was detected with the highest frequency in adults, although it has often been reported that B. adolescentis is the most common species. Real-time, quantitative PCR using primers targeting 16S rDNA shows promise in the enumeration of bifidobacteria in faecal samples. The approach to detect the target bacteria with quantitative PCR described in this review will contribute to future studies of the composition and dynamics of the intestinal microflora.  相似文献   

10.
AIMS: Bifidobacterium species are known for their beneficial effects on health and their wide use as probiotics. Although various polymerase chain reaction (PCR) methods for the identification of Bifidobacterium species have been published, the reliability of these methods remains open to question. METHODS AND RESULTS: In this study, we evaluated 37 previously reported PCR primer sets designed to amplify 16S rDNA, 23S rDNA, intergenic spacer regions, or repetitive DNA sequences of various Bifidobacterium species. CONCLUSIONS: Ten of 37 experimental primer sets showed specificity for B. adolescentis, B. angulatum, B. pseudocatenulatum, B. breve, B. bifidum, B. longum, B. longum biovar infantis and B. dentium. SIGNIFICANCE AND IMPACT OF THE STUDY: The results suggest that published Bifidobacterium primer sets should be re-evaluated for both reproducibility and specificity for the identification of Bifidobacterium species using PCR. Improvement of existing PCR methods will be needed to facilitate identification of other Bifidobacterium strains, such as B. animalis, B. catenulatum, B. thermophilum and B. subtile.  相似文献   

11.
Twenty-eight exclusively breast-fed healthy infants and 16 infants also exclusively breast-fed with allergic colitis (aged 85 +/- 60 and 98 +/- 58 d, respectively) were screened for differences in fecal flora. Bifidobacteria were detected in 23 healthy infants and only in 4 fecal samples of infants with allergic colitis. All bifidobacteria-free infants possessed Gram-positive regular rods as a major group of their fecal flora. These bacteria were identified as clostridia using genus-specific FISH probe. Infants with allergy colitis possessed significantly lower counts of bifidobacteria and total anaerobes and significantly higher counts of clostridia in their feces. In healthy infants, Bifidobacterium longum was the most frequently found species (54.5% of the samples), followed by B. adolescentis (20.0), B. breve (18.2), B. bifidum (16.4), B. dentium (10.9) and B. pseudocatenulatum (1.80). Bifidobacterial isolates from two babies with allergic colitis were identified as B. longum, one child from patients group contained species B. dentium and one baby B. adolescentis. Our results suggest that there are significantly lower counts of bifidobacteria in infants with allergic colitis than in healthy infants.  相似文献   

12.
AIMS: The anti-infectious activity of Bifidobacteria in combination with transgalactosylated oligosaccharides (TOS) against Salmonella enterica serovar Typhimurium LT-2 in an opportunistic antibiotic-induced murine infection model in mice was examined. METHODS AND RESULTS: B. breve (strain Yakult) with natural resistance to streptomycin sulphate (SM, MIC: > 4 mg ml(-1)), when given daily at a dose of 108 cfu/mouse orally under SM treatment was constantly excreted at 10(10) cfu g(-1) faeces so long as SM was administered, even at 2 weeks after discontinuing administration of B. breve. Explosive intestinal growth and subsequent extra-intestinal translocation of orally infected LT-2 under SM treatment were inhibited by B. breve colonization, and this anti-infectious activity was strengthened by synbiotic administration of TOS with B. breve. Comparison of anti-Salmonella activity among several Bifidobacterium strains with natural resistance to SM revealed that strains such as B. bifidum ATCC 15696 and B. catenulatum ATCC 27539T conferred no activity, even when they reached high population levels similar those of effective strains such as strain Yakult and B. pseudocatenulatum DSM 20439. Both the increase in the concentration of organic acids and the lowered pH in the intestine due to bifidobacterial colonization correlated with the anti-infectious activity. Moreover, the crude cecal extract of B. breve-colonized mice exerted growth-inhibitory activity against LT-2 in vitro, whereas that of the ineffective B. bifidum-colonized cecum showed much lower activity. CONCLUSIONS: Intestinal colonization by bifidobacteria given exogenously together with TOS during antibiotic treatment prevents the antibiotic-induced disruption of colonization resistance to oral infection with S. enterica serovar Typhimurium, and the metabolic activity needed to produce organic acids and lower the intestinal pH is important in the anti-infectious activity of synbiotics against enteric infection with Salmonella. SIGNIFICANCE AND IMPACT OF THE STUDY: These results indicate that certain bifidobacteria together with prebiotics may be used for the prophylaxis against opportunistic intestinal infections with antibiotic-resistant pathogens.  相似文献   

13.
The possibility of the formation of exoenzymes, such as DNAase, RNAase and hemolysin, by bifidobacteria was studied in comparison with their acid-forming and adhesive activity. Bifidobacterium reference strains, originally isolated from healthy adults and children, were studied. The study involved altogether 73 strains of bifidobacteria, including 24 B. bifidum strains, 13 B. adolescentis strains, 7 B. infantis strains, 10 B. breve strains and 19 B. longum strains. The bifidobacteria under study were shown to differ not only in the presence and activity of properties useful for macroorganisms, but also in the presence of enzymes having depolymerizing activity (DNAase, hemolysin). Thus, out of 73 strains under study 9 proved to be DNAase-positive and 6, hemolysin positive. At the same time a specific feature of bifidobacteria was their high acid-forming activity with the complete absence of RNAase activity and insignificant DNAase- and hemolysin-forming activity.  相似文献   

14.
BACKGROUND: Cell surface pili in Gram positive bacteria have been reported to orchestrate the colonization of host tissues, evasion of immunity and the development of biofilms. So far, little if any information is available on the presence of pilus-like structures in human gut commensals like bifidobacteria. RESULTS AND DISCUSSION: In this report, Atomic Force Microscopy (AFM) of various bifidobacterial strains belonging to Bifidobacterium bifidum, Bifidobacterium longum subsp. longum, Bifidobacterium dentium, Bifidobacterium adolescentis and Bifidobacterium animalis subsp. lactis revealed the existence of appendages resembling pilus-like structures. Interestingly, these microorganisms harbour two to six predicted pilus gene clusters in their genome, with each organized in an operon encompassing the major pilin subunit-encoding gene (designated fimA or fimP) together with one or two minor pilin subunit-encoding genes (designated as fimB and/or fimQ), and a gene encoding a sortase enzyme (strA). Quantitative Real Time (qRT)-PCR analysis and RT-PCR experiments revealed a polycistronic mRNA, encompassing the fimA/P and fimB/Q genes, which are differentially expressed upon cultivation of bifidobacteria on various glycans.  相似文献   

15.
To characterize the ability of bifidobacteria to affect the production of macrophage-derived cytokines, a murine macrophage-like cell line, J774.1, was cultured in the presence of 27 strains of heat-inactivated bifidobacteria. Bifidobacterium adolescentis and B. longum, known as adult-type bifidobacteria, induced significantly more pro-inflammatory cytokine secretion, IL-12 and TNF-alpha, by J774.1 cells, than did the infant-type bifidobacteria, B. bifidum, B. breve, and B. infantis (P<0.01). In contrast, B. adolescentis did not stimulate the production of anti-inflammatory IL-10 from J774.1 cells as the other tested bacteria did. The results suggest that the adult-type bifidobacteria, especially B. adolescentis, may be more potent to amplify but less able to down-regulate the inflammatory response.  相似文献   

16.
Quantitative real-time PCR assays targeting the groEL gene for the specific enumeration of 12 human fecal Bifidobacterium species were developed. The housekeeping gene groEL (HSP60 in eukaryotes) was used as a discriminative marker for the differentiation of Bifidobacterium adolescentis, B. angulatum, B. animalis, B. bifidum, B. breve, B. catenulatum, B. dentium, B. gallicum, B. longum, B. pseudocatenulatum, B. pseudolongum, and B. thermophilum. The bifidobacterial chromosome contains a single copy of the groEL gene, allowing the determination of the cell number by quantification of the groEL copy number. Real-time PCR assays were validated by comparing fecal samples spiked with known numbers of a given Bifidobacterium species. Independent of the Bifidobacterium species tested, the proportion of groEL copies recovered from fecal samples spiked with 5 to 9 log(10) cells/g feces was approximately 50%. The quantification limit was 5 to 6 log(10) groEL copies/g feces. The interassay variability was less than 10%, and variability between different DNA extractions was less than 23%. The method developed was applied to fecal samples from healthy adults and full-term breast-fed infants. Bifidobacterial diversity in both adults and infants was low, with mostly ≤3 Bifidobacterium species and B. longum frequently detected. The predominant species in infant and adult fecal samples were B. breve and B. adolescentis, respectively. It was possible to distinguish B. catenulatum and B. pseudocatenulatum. We conclude that the groEL gene is a suitable molecular marker for the specific and accurate quantification of human fecal Bifidobacterium species by real-time PCR.  相似文献   

17.
We investigated the effects of lactoferrin on the growth of L. acidophilus CH-2, Bifidobacterium breve ATCC 15700, B. longum ATCC 15707, B. infantis ATCC 15697, and B. bifidum ATCC 15696. The growth of L. acidophilus was stimulated by bovine holo-lactoferrin but not by apo-lactoferrin. With bifidobacteria, bovine lactoferrin stimulated growth of three strains: B. breve, B. infantis and B. bifidum under certain conditions. Both apoprotein and holoprotein had similar effects. However, B. longum growth was not affected by lactoferrin. Thus, the mechanism of stimulating growth of bifidobacteria may be different from that of L. acidophilus. By far-western blotting using biotinylated lactoferrin and horseradish peroxidase-conjugated streptavidin, lactoferrin-binding proteins were detected in the membrane protein fraction of L. acidophilus, B. bifidum, B. infantis and B. breve. The molecular weights of lactoferrin-binding proteins of L. acidophilus were estimated from SDS-polyacrylamide gel electrophoresis to be 27, 41 and 67 kDa, and those of the three bifidobacterial strains were estimated to be 67-69 kDa. However, no such lactoferrin-binding components were detected in the membrane fraction of B. longum. It is interesting that the appearance of lactoferrin-binding proteins in the membrane fraction of these species corresponds to their growth stimulation by lactoferrin.  相似文献   

18.
An immuno-culture method has been developed by combination of specific monoclonal antibodies and plate culture to allow detection of viable bifidobacteria. Cell wall proteins were selected as surface antigen to produce antibodies against bifidobacteria. The cell wall proteins were extracted and purified from six ATCC strains of bifidobacteria grown in MRS broth using an anaerobic system. To compare the profile of the protein extracts, all the protein solutions obtained were analyzed by SDS-PAGE. Similar bands corresponding to the major proteins of each species of bifidobacteria were observed. The proteins were tested for their immunogenicity in Balb/c mice after immunization and subsequent analysis using ELISA procedures. High immune responses were generated in mice immunized by proteins from Bifidobacterium bifidum and Bifidobacterium longum. Monoclonal antibodies were produced against B. longum and tested for their specificity, sensitivity and cross reactivity with other bifidobacteria species. All the hybridoma cells selected produced anti-B. longum antibodies cross-reacting with native and purified proteins from five other bifidobacteria species. An epitope supported by a cross-reacting protein of 58 kDa shared by bifidobacteria was revealed by western blot. This was confirmed by immune-transmission electron microscopy observations which showed the specific interaction of these antibodies with bifidobacterial cell wall proteins. Also, the antibody obtained was found to be specific for the genus Bifidobacterium and sensitive, allowing the detection of at least 10(5) target cells/ml. An immuno-culture detection approach was then developed using the selected anti-B. longum antibodies. This method was shown to be very efficient for the detection of viable cells of bifidobacteria suggesting the possibility of its use to quantify these bacteria in various food matrices.  相似文献   

19.

Objectives

Bifidobacterium species are one of the major components of the infant''s intestine microbiota. Colonization with bifidobacteria in early infancy is suggested to be important for health in later life. However, information remains limited regarding the source of these microbes. Here, we investigated whether specific strains of bifidobacteria in the maternal intestinal flora are transmitted to their infant''s intestine.

Materials and Methods

Fecal samples were collected from healthy 17 mother and infant pairs (Vaginal delivery: 12; Cesarean section delivery: 5). Mother''s feces were collected twice before delivery. Infant''s feces were collected at 0 (meconium), 3, 7, 30, 90 days after birth. Bifidobacteria isolated from feces were genotyped by multilocus sequencing typing, and the transitions of bifidobacteria counts in infant''s feces were analyzed by quantitative real-time PCR.

Results

Stains belonging to Bifidobacterium adolescentis, Bifidobacterium bifidum, Bifidobacterium catenulatum, Bifidobacterium longum subsp. longum, and Bifidobacterium pseudocatenulatum, were identified to be monophyletic between mother''s and infant''s intestine. Eleven out of 12 vaginal delivered infants carried at least one monophyletic strain. The bifidobacterial counts of the species to which the monophyletic strains belong, increased predominantly in the infant''s intestine within 3 days after birth. Among infants delivered by C-section, monophyletic strains were not observed. Moreover, the bifidobacterial counts were significantly lower than the vaginal delivered infants until 7 days of age.

Conclusions

Among infants born vaginally, several Bifidobacterium strains transmit from the mother and colonize the infant''s intestine shortly after birth. Our data suggest that the mother''s intestine is an important source for the vaginal delivered infant''s intestinal microbiota.  相似文献   

20.
为筛选具有潜在益生功能的双歧杆菌,本研究采用改良MRS培养基,从婴幼儿粪便中分离到5株菌株:AR499,AR667,AR668,AR669和AR610。通过16S r DNA测序分别鉴定为两歧双歧杆菌,短双歧杆菌,动物双歧杆菌,长双歧杆菌和假小链双歧杆菌。对其耐酸耐胆盐能力和黏附性能进行测试,结果表明,菌株AR499耐酸性较好,菌株AR610有较强的耐胆盐能力,菌株AR499的自动聚集能力和细胞表面疏水性都较高。实验表明,菌株AR499可作为一株耐受性和黏附性能较好的益生菌进一步深入研究,以期应用于优良双歧杆菌产品的开发。  相似文献   

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