Monica Modesto

Occurrence of the Family Bifidobacteriaceae in Human Dental Caries and Plaque

Despite the constant presence of members of Bifidobacteriaceae in the human oral cavity, few studies of their occurrence in this habitat have been made. The presence and the distribution in human plaque and dental caries of Bifidobacterium dentium, Scardovia inopinata and Parascardovia denticolens, all belonging to the family Bifidobacteriaceae, were studied. The identification to species level was based on morphological and growth type features, fermentation tests, polyacrylamide gel electrophoresis of the soluble proteins and DNA-DNA homology. The three species, isolated on propionic acid trypticase phytone yeast extract medium, were present in 13 out of 19 and in 11 out of 15 subjects examined for dental caries and plaque, respectively. S. inopinata was the species most frequently isolated in dental caries, whereas B. dentium was more numerous in dental plaque.The prevalence of P. denticolens was similar in the two habitats. In conclusion, the prevalence of bifidobacteria species (B. dentium, P. denticolens, S. inopinata) in relation to non-bifidobacteria isolates was significantly different in caries versus plaque samples (p < 0.0001; χ2 test).

Effect of fructo-oligosaccharides and different doses of Bifidobacterium animalis in a weaning diet on bacterial translocation and Toll-like receptor gene expression in pigs

OBJECTIVE Our aim was to study the possible synergic action of one prebiotic with increasing dietary doses of a probiotic strain of Bifidobacterium animalis on the translocation of bifidobacteria and on Toll-like receptor (TLR) gene expression in different organs of weaned piglets. METHODS Sixty-four pigs, reared from 21 to 35 d of age, were fed eight different diets according to a 2 x 4 factorial design: a control diet or the control diet supplemented with three different levels of B. animalis (10(7), 10(9), 10(11) colony-forming units/d), crossed with 0% or 2% sugar beet fructo-oligosaccharides. Pigs were then sacrificed, and the jejunum mucosa, ileocecal lymph nodes, and liver were sampled to determine the presence of Bifidobacterium spp. DNA and to quantify the expression of TLR2-, TLR4-, and tumor necrosis factor-alpha-encoding genes. RESULTS We found Bifidobacterium spp. genus-specific DNA in lymph nodes of subjects from all dietary treatments, including the control diet, but it increased with the bifidobacteria oral dose (P = 0.065). The linear effect of the dose of B. animalis on the expression of the TLR2-encoding gene in the lymph nodes was observed when fructo-oligosaccharides were added to the diet (P < 0.05). Tumor necrosis factor-alpha-encoding gene expression was positively correlated with TLR4- and TLR2-encoding gene expressions (P < 0.001 and P < 0.01, respectively) and negatively correlated with bifidobacteria DNA (P < 0.05). Moreover, the expression of the TLR4-encoding gene showed a positive correlation with TLR2-encoding gene expression (P < 0.001). In contrast, there was no correlation between expressions of the TLR2- and TLR4-encoding genes with the bifidobacteria DNA. CONCLUSION Soon after weaning, the translocation of the commensal bacteria in the ileocecal lymph nodes is a physiologic process. Moreover, diet affects the expression of the TLR2-encoding gene.

Comparative genomic and phylogenomic analyses of the Bifidobacteriaceae family

BackgroundMembers of the Bifidobacteriaceae family represent both dominant microbial groups that colonize the gut of various animals, especially during the suckling stage of their life, while they also occur as pathogenic bacteria of the urogenital tract. The pan-genome of the genus Bifidobacterium has been explored in detail in recent years, though genomics of the Bifidobacteriaceae family has not yet received much attention. Here, a comparative genomic analyses of 67 Bifidobacteriaceae (sub) species including all currently recognized genera of this family, i.e., Aeriscardovia, Alloscardovia, Bifidobacterium, Bombiscardovia, Gardnerella, Neoscardovia, Parascardovia, Pseudoscardovia and Scardovia, was performed. Furthermore, in order to include a representative of each of the 67 (currently recognized) (sub) species belonging to the Bifidobacteriaceae family, we sequenced the genomes of an additional 11 species from this family, accomplishing the most extensive comparative genomic analysis performed within this family so far.ResultsPhylogenomics-based analyses revealed the deduced evolutionary pathway followed by each member of the Bifidobacteriaceae family, highlighting Aeriscardovia aeriphila LMG 21773 as the deepest branch in the evolutionary tree of this family. Furthermore, functional analyses based on genome content unveil connections between a given member of the family, its carbohydrate utilization abilities and its corresponding host. In this context, bifidobacterial (sub) species isolated from humans and monkeys possess the highest relative number of acquired glycosyl hydrolase-encoding genes, probably in order to enhance their metabolic ability to utilize different carbon sources consumed by the host.ConclusionsWithin the Bifidobacteriaceae family, genomics of the genus Bifidobacterium has been extensively investigated. In contrast, very little is known about the genomics of members of the other eight genera of this family. In this study, we decoded the genome sequences of each member of the Bifidobacteriaceae family. Thanks to subsequent comparative genomic and phylogenetic analyses, the deduced pan-genome of this family, as well as the predicted evolutionary development of each taxon belonging to this family was assessed.

Bifidobacterium myosotis sp. nov., Bifidobacterium tissieri sp. nov. and Bifidobacterium hapali sp. nov., isolated from faeces of baby common marmosets (Callithrix jacchus L.).

In a previous study on bifidobacterial distribution in New World monkeys, six strains belonging to the Bifidobacteriaceae were isolated from faecal samples of baby common marmosets (Callithrix jacchus L.). All the isolates were Gram-positive-staining, anaerobic, asporogenous and fructose-6-phosphate phosphoketolase-positive. Comparative analysis of 16S rRNA gene sequences revealed relatively low levels of similarity (maximum identity 96 %) to members of the genus Bifidobacterium, and placed the isolates in three independent clusters: strains of cluster I (MRM_5.9T and MRM_5.10) and cluster III (MRM_5.18T and MRM_9.02) respectively showed 96.4 and 96.7 % 16S rRNA gene sequence similarity to Bifidobacterium callitrichos DSM 23973T, while strains of cluster II (MRM_8.14T and MRM_9.14) showed 95.4 % similarity to Bifidobacterium stellenboschense DSM 23968T. Phylogenetic analysis of partial hsp60 and clpC gene sequences supported an independent phylogenetic position of each cluster from each other and from the related type strains B. callitrichos DSM 23973T and B. stellenboschense DSM 23968T. Clusters I, II and III respectively showed DNA G+C contents of 64.9-65.1, 56.4-56.7 and 63.1-63.7 mol%. The major cellular fatty acids of MRM_5.9T were C14 : 0, C16 : 0 and C18 : 1ω9c dimethylacetal, while C16 : 0 was prominent in strains MRM_5.18T and MRM_8.14T, followed by C18 : 1ω9c and C14 : 0. Biochemical profiles and growth parameters were recorded for all the isolates. Based on the data provided, the clusters represent three novel species, for which the names Bifidobacterium myosotis sp. nov. (type strain MRM_5.9T = DSM 100196T = JCM 30796T), Bifidobacterium hapali sp. nov. (type strain MRM_8.14T = DSM 100202T = JCM 30799T) and Bifidobacterium tissieri sp. nov. (type strain MRM_5.18T = DSM 100201T = JCM 30798T) are proposed.

Bifidobacterium aerophilum sp. nov., Bifidobacterium avesanii sp. nov. and Bifidobacterium ramosum sp. nov.: Three novel taxa from the faeces of cotton-top tamarin (Saguinus oedipus L.) ☆

Forty-five microorganisms were isolated on bifidobacteria selective medium from one faecal sample of an adult subject of the cotton-top tamarin (Saguinus oedipus L.). All isolates were Gram-positive, catalase-negative, anaerobic, fructose-6-phosphate phosphoketolase positive, and asporogenous rod-shaped bacteria. In this study, only eight out of the forty-five strains were characterized more deeply, whereas the others are still currently under investigation. They were grouped by BOX-PCR into three clusters: Cluster I (TRE 17(T), TRE 7, TRE 26, TRE 32, TRE 33, TRE I), Cluster II (TRE C(T)), and Cluster III (TRE M(T)). Comparative analysis of 16S rRNA gene sequences confirmed the results from the cluster analysis and revealed relatively low level similarities to each other (mean value 95%) and to members of the genus Bifidobacterium. All eight isolates showed the highest level of 16S rRNA gene sequence similarities with Bifidobacterium scardovii DSM 13734(T) (mean value 96.6%). Multilocus sequence analysis (MLSA) of five housekeeping genes (hsp60, rpoB, clpC, dnaJ and dnaG) supported their independent phylogenetic position to each other and to related species of Bifidobacterium. The G+C contents were 63.2%, 65.9% and 63.0% for Cluster I, Cluster II and Cluster III, respectively. Peptidoglycan types were A3α l-Lys-l-Thr-l-Ala, A4β l-Orn (Lys)-d-Ser-d-Glu and A3β l-Orn-l-Ser-l-Ala in Clusters I, II and III, respectively. Based on the data provided, each cluster represented a novel taxon for which the names Bifidobacterium aerophilum sp. nov. (TRE 17(T)=DSM 100689=JCM 30941; TRE 26=DSM 100690=JCM 30942), Bifidobacterium avesanii sp. nov. (TRE C(T)=DSM 100685=JCM 30943) and Bifidobacterium ramosum sp. nov. (TRE M=DSM 100688=JCM 30944) are proposed.

Isolation and identification of cultivable Bifidobacterium spp. from the faeces of 5 baby common marmosets (Callithrix jacchus L.)

Ninety-two bifidobacterial strains were obtained from the faeces of 5 baby common marmosets, three known species Bifidobacterium aesculapii, Bifidobacterium callithricos and Bifidobacterium reuteri and 4 novel putative bifidobacterial species were retrieved. The occurrence of bifidobacteria in non-human primate babies is described for the first time.

In Vitro Activity of Tea Tree Oil Vaginal Suppositories against Candida spp. and Probiotic Vaginal Microbiota

The aim of this work is to evaluate the in vitro microbicidal activity of vaginal suppositories (VS) containing tea tree oil (TTO‐VS) towards Candida spp. and vaginal probiotics. A total of 20 Candida spp. strains, taken from patients with vaginitis and from an established type collection, including reference strains, were analysed by using the CLSI microdilution method. To study the action of VS towards the beneficial vaginal microbiota, the sensitivity of Bifidobacterium animalis subsp. lactis (DSM 10140) and Lactobacillus spp. (Lactobacillus casei R‐215 and Lactobacillus acidophilus R‐52) was tested. Both TTO‐VS and TTO showed fungicidal activity against all strains of Candida spp. whereas placebo‐VS or the Aloe gel used as controls were ineffective. The study of fractional fungicidal concentrations (FFC) showed synergistic interaction with the association between Amphotericin B and TTO (0.25 to 0.08 µg/ml, respectively) against Candida albicans. Instead, the probiotics were only affected by TTO concentration ≥ 4% v/v, while, at concentrations < 2% v/v, they remained viable. TTO‐VS exhibits, in vitro, a selective fungicidal action, slightly affecting only the Bifidobacteriun animalis strain growth belonging to the vaginal microbiota. In vivo studies are needed to confirm the efficacy to prevent acute or recurrent vaginal candidiasis. Copyright

Bifidobacterium. eulemuris sp. nov. isolated from the faeces of the black lemur (Eulemur macaco).

Forty-three strains of bifidobacteria were isolated from the faeces of two adult black lemurs, Eulemur macaco. Thirty-four were identified as Bifidobacterium lemurum, recently described in Lemur catta. The nine remaining isolates were Gram-positive-staining, non-spore-forming, fructose-6-phosphate phosphoketolase-positive, microaerophilic, irregular rod-shaped bacteria that often presented Y- or V-shaped cells. Typing techniques revealed that these isolates were nearly identical, and strain LMM_E3T was chosen as a representative and characterized further. Phylogenetic analysis based on 16S rRNA gene sequences clustered this isolate inside the genus Bifidobacterium and showed the highest levels of sequence similarity with B. lemurum DSM 28807T (99.3 %), with Bifidobacterium pullorum LMG 21816T and Bifidobacterium longum subsp. infantis ATCC 15697T (96.4 and 96.3 %, respectively) as the next most similar strains. The hsp60 gene sequence of strain LMM_E3T showed the highest similarity to that of Bifidobacterium stellenboschense DSM 23968T (93.3 %), and 91.0 % similarity to that of the type strain of B. lemurum. DNA-DNA reassociation with the closest neighbour B. lemurum DSM 28807T was found to be 65.4 %. The DNA G+C content was 62.3 mol%. Strain LMM_E3T showed a peptidoglycan structure that has not been detected in bifidobacteria so far: A3α l-Lys-l-Ser-l-Thr-l-Ala. Based on the phylogenetic, genotypic and phenotypic data, strain LMM_E3T represents a novel species within the genus Bifidobacterium, for which the name Bifidobacterium eulemuris sp. nov. is proposed; the type strain is LMM_E3T ( = DSM 100216T = JCM 30801T).

Identification of Bifidobacterium spp. using hsp60 PCR-RFLP analysis: An update

A PCR-RFLP technique has been applied on 13 species of Bifidobacterium in order to update a previous study carried out by Baffoni et al. This method is based on the restriction endonuclease activity of HaeIII on the PCR-amplified hsp60 partial gene sequence, and allows a rapid and efficient identification of Bifidobacterium spp. strains at species and subspecies level.

Bifidobacterium callitrichidarum sp. nov. from the faeces of the emperor tamarin (Saguinus imperator)

Three Gram-stain-positive, non-spore-forming, microaerophilic and fructose-6-phosphate phosphoketolase positive strains were isolated from a faecal sample of an adult subject of the emperor tamarin (Saguinus imperator). Given that the isolates revealed identical BOX PCR profiles, strain TRI 5T was selected as a representative and characterized further. Comparative analysis of 16S rRNA gene sequence similarity revealed that strain TRI 5T was closely related to Bifidobacterium saguini DSM 23967T (96.4 %) and to Bifidobacterium longum subsp. longum ATCC 15708 (96.2 %). Multilocus sequence analyses of five housekeeping genes showed the close phylogenetic relatedness of this strain to Bifidobacterium breve DSM 20213T (hsp60 94.1 %), Bifidobacterium saguini DSM 23967T (clpC 91 %), Bifidobacterium avesanii DSM 100685T (dnaG 80.3 %), Bifidobacterium longumsubsp. infantis ATCC 15697T (dnaJ 85.3 %) and Bifidobacterium longumsubsp. longum ATCC 15708 (rpoB 93 %), respectively. The peptidoglycan type was A3β, with an interpeptide bridge comprising l-Orn (Lys) - l-Ser - l-Ala - l-Thr - l-Ala. The DNA G+C content of strain TRI 5T was 60.9 mol%. Based on the data provided, strain TRI 5T represents a novel species of the genus Bifidobacterium for which the name Bifidobacteriumcallitrichidarum sp. nov. is proposed. The type strain is TRI 5T (=DSM 103152T=JCM 31790T).