Effects of Bifidobacterium breve A1 on the cognitive function of older adults with memory complaints: a randomised, double-blind, placebo-controlled trial.

In our previous study, we reported the therapeutic potential of Bifidobacterium breve A1 in preventing cognitive impairment in a mouse model of Alzheimer's disease and participants with mild cognitive impairment; we suggested that probiotic supplementation is an effective therapeutic strategy for managing cognitive function. Accordingly, we conducted a randomised, double-blind, placebo-controlled trial to assess whether 12-week B. breve A1 supplementation could affect the cognitive function of elderly subjects with memory complaints. We assessed cognitive function using the Japanese version of the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) and Mini-Mental State Examination (MMSE) at baseline and after 12 weeks of probiotic supplementation. A total of 121 participants were randomised and received B. breve A1 capsules or placebo daily for 12 weeks; of these, 117 participants completed the study. At 12 weeks, neuropsychological test scores significantly increased in both groups; no significant intergroup difference was observed in terms of changes in scores from the baseline scores. However, a stratified analysis revealed a significant difference between B. breve A1 and placebo groups in terms of the subscale 'immediate memory' of RBANS and MMSE total score in the subjects with low RBANS total score at baseline. No significant differences in terms of blood parameters between the groups or adverse effects caused by B. breve A1 intervention were observed. The results of the present study suggest the safety of B. breve A1 supplementation and its potential in maintaining cognitive function in elderly subjects with memory complaints. However, future large-scale studies on individuals with impaired cognitive function are required to validate the present findings.

Bifidobacterium breve MCC1274 with glycosidic activity enhances in vivo isoflavone bioavailability.

Polyphenols are plant derived compounds that exert many beneficial health effects to the human host. However, associated health benefits of dietary polyphenol are highly dependent on their intestinal metabolism, bioavailability, and absorption. Bifidobacteria, which represent the key members of gut microbiota, have been suggested to promote gut microbial homeostasis and may be involved in the metabolism of polyphenols. In this study, the capabilities of thirteen Bifidobacterium strains in hydrolysing polyphenol glycosides were evaluated. Among the tested strains, Bifidobacterium breve MCC1274 was found to possess the highest ß-glucosidase activity and strong capability to convert daidzin and trans-polydatin to their aglycones; while kinetic analysis revealed that B. breve MCC1274 hydrolysed more than 50% of daidzin and trans-polydatin at less than 3 h of incubation. Further investigation using rats with an antibiotics-disturbed microbiome revealed that following the ingestion of daidzin glycoside, oral administration of B. breve MCC1274 significantly enhanced the plasma concentration of daidzein in rats pre-treated with antibiotics as compared to antibiotics-pre-treated control and non-treated control groups. The relative abundance of Actinobacteria and the total numbers of B. breve were also significantly higher in antibiotics-pre-treated rats administered with B. breve MCC1274 than that of the control groups. These findings suggest that B. breve MCC1274 is effective in enhancing the bioavailability of daidzein in the gut under dysbiosis conditions and may potentially improve intestinal absorption of isoflavones and promote human health.

Deciphering the influence of paraimmunobiotic bifidobacteria on the innate antiviral immune response of bovine intestinal epitheliocytes by transcriptomic analysis.

Previously, we reported that the non-viable immunomodulatory Bifidobacterium infantis MCC12 and Bifidobacterium breve MCC1274 strains (paraimmunobiotic bifidobacteria) were able to increase the protection against rotavirus infection in bovine intestinal epithelial (BIE) cells. In order to gain insight into the influence of paraimmunobiotic bifidobacteria on the innate antiviral immune response of BIE cells, their effect on the transcriptomic response triggered by Toll-like receptor 3 (TLR3) activation was investigated. By using microarray technology and qPCR analysis, we obtained a global overview of the immune genes involved in the innate antiviral immune response in BIE cells. Activation of TLR3 by poly(I:C) in BIE cells significantly increased the expression of interferon (IFN)-α and IFN-ß, several interferon-stimulated genes, cytokines, and chemokines. It was also observed that both paraimmunobiotic bifidobacteria differently modulated immune genes expression in poly(I:C)-challenged BIE cells. Most notable changes were found in genes involved in antiviral defence (IFN-ß, MX1, OAS1X, MDA5, TLR3, STAT2, STAT3), cytokines (interleukin (IL)-6), and chemokines (CCL2, CXCL2, CXCL6) that were significantly increased in bifidobacteria-treated BIE cells. B. infantis MCC12 and B. breve MCC1274 showed quantitative and qualitative differences in their capacities to modulate the innate antiviral immune response in BIE cells. B. breve MCC1274 was more efficient than the MCC12 strain to improve the production of type I IFNs and antiviral factors, an effect that could be related to its higher ability to protect against rotavirus replication in BIE cells. Interestingly, B. infantis MCC12 showed a remarkable anti-inflammatory effect. The MCC12 strain was more efficient to reduce the expression of inflammatory cytokines and chemokines (IL-16, IL-20, CX3CL1) when compared with B. breve MCC1274. These results provided valuable information for the deeper understanding of the antiviral immune response of intestinal epithelial cells as well as the host-paraimmunobiotic interaction in the bovine host.

Bifidobacterium Breve A1 Supplementation Improved Cognitive Decline in Older Adults with Mild Cognitive Impairment: An Open-Label, Single-Arm Study.

OBJECTIVES: We previously reported the therapeutic potential of Bifidobacterium breve A1 (B. breve A1) for preventing cognitive impairment in Alzheimer's disease model mice, which suggested that supplementation of the probiotics could be an effective therapeutic strategy for managing cognitive function in mild cognitive impairment (MCI). DESIGN AND SETTINGS: We conducted an open-label, single-arm study to examine the effects of 24-week supplementation of B. breve A1 on elderly with MCI in Aki Orthopedics Rehabilitation Clinic in Japan. PARTICIPANTS: 27 participants were screened by their Mini Mental State Examination (MMSE) scores. MEASUREMENTS: Cognitive function was assessed using MMSE and Digit Symbol Substitution Test (DSST) at baseline and every 8 weeks. Mental condition and quality of life for gastrointestinal symptoms were measured using the Profile of Mood States 2nd Edition (POMS2), and the Gastrointestinal Symptom Rating Scale (GSRS). RESULTS: Of the 27 participants enrolled, 19 completed the study. MMSE scores were significantly increased during the intervention by mixed model Dunnett's test and Wilcoxon signed-rank tests (+1.7, P < 0.01). POMS2 and GSRS scores were significantly improved during intervention when analyzed by Wilcoxon signed-rank tests. CONCLUSION: The present study showed that oral supplementation of B. breve A1 in participants with MCI improved cognitive function, thus suggesting the potential of B. breve A1 for improving cognitive function and maintaining quality of life of the elderly. Further randomized, double-blind placebo-controlled studies are worth conducting to examine the beneficial effect of B. breve A1.

Association between blood omega-3 polyunsaturated fatty acids and the gut microbiota among breast cancer survivors.

Omega-3 polyunsaturated fatty acids (PUFAs) are essential nutrients demonstrated to have health benefits, such as decreasing the risk of coronary heart disease, improving parameters associated with metabolic syndrome, and decreasing anxiety symptoms and depression risk. Previous intervention studies indicated the association between blood or tissue PUFA levels and the gut microbiota; however, the details remain incompletely elucidated. We conducted a cross-sectional study to examine the association between PUFAs and the gut microbiota among breast cancer survivors. Adults who had been diagnosed with invasive breast cancer more than one year ago and were not currently undergoing chemotherapy were enrolled. Capillary blood and faecal samples were obtained to assess the blood PUFA levels and gut microbiota compositions. The mean age (n=124) was 58.7 years, and 46% of the participants had a history of chemotherapy. Multiple regression analysis controlling for possible confounders indicated that an increased relative abundance of Actinobacteria was significantly associated with increased levels of docosahexaenoic acid (DHA, beta=0.304, q<0.01). At the genus level, the abundance of Bifidobacterium was positively associated with the level of DHA (beta=0.307, q<0.01). No significant association between omega-6 PUFAs and the relative abundances of gut microbiota members was observed. In addition, analyses stratified by the history of chemotherapy indicated significant associations of PUFA levels with the abundance of some bacterial taxa, including the phylum Actinobacteria (DHA, beta=0.365, q<0.01) and Bacteroidetes (EPA, beta=-0.339, q<0.01) and the genus Bifidobacterium (DHA, beta=0.368, q<0.01) only among participants without a history of chemotherapy. These findings provide the first evidence of positive associations between the abundances of Bifidobacterium among the gut microbiota and the levels of omega-3 PUFAs in the blood. Further studies are required to gain additional insight into these associations in healthy subjects as well as into the causality of the relationship.

Effect of combined bifidobacteria supplementation and resistance training on cognitive function, body composition and bowel habits of healthy elderly subjects.

Physical exercise exerts favourable effects on brain health and quality of life of the elderly; some of these positive health effects are induced by the modulation of microbiota composition. We therefore conducted a randomised, double blind, placebo-controlled trial that assessed whether a combination of Bifidobacterium spp. supplementation and moderate resistance training improved the cognitive function and other health-related parameters in healthy elderly subjects. Over a 12-week period, 38 participants (66-78 years) underwent resistance training and were assigned to the probiotic Bifidobacterium supplementation (n=20; 1.25×1010 cfu each of Bifidobacterium longum subsp. longum BB536, B. longum subsp. infantis M-63, Bifidobacterium breve M-16V and B. breve B-3) or the placebo (n=18) group. At baseline and at 12 weeks, we assessed the cognitive function, using the Japanese version of the Montreal Cognitive Assessment instrument (MoCA-J); modified flanker task scores; depression-anxiety scores; body composition; and bowel habits. At 12 weeks, the MoCA-J scores showed a significant increase in both the groups, while the flanker task scores of the probiotic group increased more significantly than those of the placebo group (0.35±0.9 vs -0.29±1.1, P=0.056). Only the probiotic group showed a significant decrease in the depression-anxiety scores (5.2±6.3 to 3.4±5.5, P=0.012) and body mass index (24.0±2.8 to 23.5±2.8 kg/m2, P<0.001), with a significant increase in the defecation frequency (5.3±2.3 to 6.4±2.3 times/5 days, P=0.023) at 12 weeks. Thus, in healthy elderly subjects, combined probiotic bifidobacteria supplementation and moderate resistance training may improve the mental condition, body weight and bowel movement frequency.

Degradation of food-derived opioid peptides by bifidobacteria.

Some food-derived opioid peptides have been reported to cause diseases, such as gastrointestinal inflammation, celiac disease, and mental disorders. Bifidobacterium is a major member of the dominant human gut microbiota, particularly in the gut of infants. In this study, we evaluated the potential of Bifidobacterium in the degradation of food-derived opioid peptides. All strains tested showed some level of dipeptidyl peptidase activity, which is thought to be involved in the degradation of food-derived opioid peptides. However, this activity was higher in bifidobacterial strains that are commonly found in the intestines of human infants, such as Bifidobacterium longum subsp. longum, B. longum subsp. infantis, Bifidobacterium breve and Bifidobacterium bifidum, than in those of other species, such as Bifidobacterium animalis and Bifidobacterium pseudolongum. In addition, some B. longum subsp. infantis and B. bifidum strains showed degradative activity in food-derived opioid peptides such as human and bovine milk-derived casomorphin-7 and wheat gluten-derived gliadorphin-7. A further screening of B. bifidum strains revealed some bifidobacterial strains that could degrade all three peptides. Our results revealed the potential of Bifidobacterium species in the degradation of food-derived opioid peptides, particularly for species commonly found in the intestine of infants. Selected strains of B. longum subsp. infantis and B. bifidum with high degradative capabilities can be used as probiotic microorganisms to eliminate food-derived opioid peptides and contribute to host health.

Different physiological properties of human-residential and non-human-residential bifidobacteria in human health.

Bifidobacteria have increasingly been shown to exert positive health benefits to humans, which are clearly reflected by their application in various commercialised dairy products and supplements. Bifidobacteria naturally inhabit a range of ecological niches and display substantial differences in their ecological adaptation among species. In general, bifidobacteria could be categorised into two major groups; bifidobacterial species of human origins as human-residential bifidobacteria (HRB) while other species which are the natural inhabitants of animals or environment as non-HRB. Current research has focused on the differential physiological features of HRB and non-HRB, such as metabolic capabilities, whilst comparative and functional genomic investigations have revealed the genetic attributes of bifidobacteria that may explain their colonisation affinities in human gut. It is becoming more apparent that distinct residential origins of bifidobacteria are likely contributed to their comparable adaptive health attributes on human host. Notably, debate still remains about the nature of bifidobacteria for use as human probiotics. Clinical evaluations involving supplementation of bifidobacteria of different origins point out the superiority of HRB in human host. Evidence also suggests that HRB especially infant-type HRB may exert better health-promoting effects and therefore serve as a better probiotic candidate for infant use. In this review, we aim to provide an overview of the genotypic and physiological differences of bifidobacteria associated with different residential origins and to shed light on the practical considerations for selection of bifidobacteria as probiotics in order to establish a healthy gut microbial community in humans.

Bifidobacterium longum BB536 alleviated upper respiratory illnesses and modulated gut microbiota profiles in Malaysian pre-school children.

This 10-months randomised, double-blind, parallel and placebo-controlled study evaluated the effects of Bifidobacterium longum BB536 on diarrhoea and/or upper respiratory illnesses in 520 healthy Malaysian pre-school children aged 2-6 years old. The subjects randomly received a one-gram sachet containing either BB536 (5×109 cfu) or placebo daily. Data analysis was performed on 219 subjects who fully complied over 10-months (placebo n=110, BB536 n=109). While BB536 did not exert significant effects against diarrhoea in children, Poisson regression with generalised estimating equations model indicated significant intergroup difference in the mean number of times of respiratory illnesses over 10 months. The duration of sore throat was reduced by 46% (P=0.018), with marginal reduction for duration of fever (reduced by 27%, P=0.084), runny nose (reduced by 15%, P=0.087) and cough (reduced by 16%, P=0.087) as compared to the placebo. Principal coordinate analysis at genus level of the gut microbiota revealed significant differences between 0 and 10 months in the BB536 group (P<0.01) but not in placebo group (P>0.05). The abundance of the genus Faecalibacterium which is associated with anti-inflammatory and immuno-modulatory properties was significantly higher in the BB536 group (P<0.05) compared to the placebo group. Altogether, our present study illustrated the potential protective effects of BB536 against upper respiratory illnesses in pre-school Malaysian children, with gut microbiota modulating properties.

Differences between live and heat-killed bifidobacteria in the regulation of immune function and the intestinal environment.

Probiotics are live microorganisms that confer a health benefit on the host, such as improvement of the intestinal environment, modulation of immune function and energy metabolism. Heat-killed probiotic strains have also been known to exhibit some physiological functions; however, the differences between live and heat-killed probiotics have not been well elucidated. In this study, we investigated the differences between live and heat-killed Bifidobacterium breve M-16V, a probiotic strain, in the regulation of immune function, intestinal metabolism and intestinal gene expression of the host using gnotobiotic mouse model and omics approaches. Both live and heat-killed cells of B. breve M-16V showed immune-modulating effects that suppressed pro-inflammatory cytokine production in spleen cells and affected intestinal metabolism; however, live cells exhibited a more remarkable effect in the regulation of intestinal metabolism and intestinal gene expression involved in nutrient metabolism. Our findings are valuable for considering the health benefits of live and heat-killed bacteria and the usefulness of different forms of probiotics.

Development of an in vitro immunobiotic evaluation system against rotavirus infection in bovine intestinal epitheliocytes.

The bovine intestinal epithelial cell line (BIE cells) expresses the Toll-like receptor (TLR)3 and is able to mount an antiviral immune response after the stimulation with poly(I:C). In the present study, we aimed to further characterise the antiviral defence mechanisms in BIE cells by evaluating the innate immune response triggered by rotavirus (RV) infection. In addition, we attempted to determine whether immunobiotic bifidobacteria are able to confer protection of BIE cells against RV infection by beneficially modulating the antiviral immune response. RV OSU (porcine) and UK (bovine) effectively infected BIE cells, while a significant lower capacity to infect BIE cells was observed for human (Wa) and murine (EW) RV. We observed that viral infection in BIE cells triggered TLR3/RIG-I-mediated immune responses with activation of IRF3 and TRAF3, induction of interferon beta (IFN-ß) and up-regulation of inflammatory cytokines. Our results also demonstrated that preventive treatments with Bifidobacterium infantis MCC12 or Bifidobacterium breve MCC1274 significantly reduced RV titres in infected BIE cells and differentially modulated the innate immune response. Of note, both strains significantly improved the production of the antiviral factor IFN-ß in RV-infected BIE cells. In conclusion, this work provides comprehensive information on the antiviral immune response of BIE cells against RV, that can be further studied for the development of strategies aimed to improve antiviral defences in bovine intestinal epithelial cells. Our results also demonstrate that BIE cells could be used as a newly immunobiotic evaluation system against RV infection for application in the bovine host.

[Endoplasmic reticulum stress in INS-1-3 cell associated with the expression changes of MODY gene pathway].

Objective: To profile the gene expression changes associated with endoplasmic reticulum stress in INS-1-3 cells induced by thapsigargin (TG) and tunicamycin (TM). Methods: Normal cultured INS-1-3 cells were used as a control. TG and TM were used to induce endoplasmic reticulum stress in INS-1-3 cells. Digital gene expression profiling technique was used to detect differentially expressed gene. The changes of gene expression were detected by expression pattern clustering analysis, gene ontology (GO) function and pathway enrichment analysis. Real time polymerase chain reaction (RT-PCR) was used to verify the key changes of gene expression. Results: Compared with the control group, there were 57 (45 up-regulated, 12 down-regulated) and 135 (99 up-regulated, 36 down-regulated) differentially expressed genes in TG and TM group, respectively. GO function enrichment analyses indicated that the main enrichment was in the endoplasmic reticulum. In signaling pathway analysis, the identified pathways were related with endoplasmic reticulum stress, antigen processing and presentation, protein export, and most of all, the maturity onset diabetes of the young (MODY) pathway. Conclusion: Under the condition of endoplasmic reticulum stress, the related expression changes of transcriptional factors in MODY signaling pathway may be related with the impaired function in islet beta cells.

Effect of probiotic yoghurt on animal-based diet-induced change in gut microbiota: an open, randomised, parallel-group study.

Diet has a significant influence on the intestinal environment. In this study, we assessed changes in the faecal microbiota induced by an animal-based diet and the effect of the ingestion of yoghurt supplemented with a probiotic strain on these changes. In total, 33 subjects were enrolled in an open, randomised, parallel-group study. After a seven-day pre-observation period, the subjects were allocated into three groups (11 subjects in each group). All of the subjects were provided with an animal-based diet for five days, followed by a balanced diet for 14 days. Subjects in the first group ingested dairy in the form of 200 g of yoghurt supplemented with Bifidobacterium longum during both the animal-based and balanced diet periods (YAB group). Subjects in the second group ingested yoghurt only during the balanced diet period (YB group). Subjects who did not ingest yoghurt throughout the intervention were used as the control (CTR) group. Faecal samples were collected before and after the animal-based diet was provided and after the balanced diet was provided, followed by analysis by high-throughput sequencing of amplicons derived from the V3-V4 region of the 16S rRNA gene. In the YB and CTR groups, the animal-based diet caused a significant increase in the relative abundance of Bilophila, Odoribacter, Dorea and Ruminococcus (belonging to Lachnospiraceae) and a significant decrease in the level of Bifidobacterium after five days of intake. With the exception of Ruminococcus, these changes were not observed in the YAB group. No significant effect was induced by yoghurt supplementation following an animal-based diet (YB group vs CTR group). These results suggest that the intake of yoghurt supplemented with bifidobacteria played a role in maintaining a normal microbiota composition during the ingestion of a meat-based diet. This study protocol was registered in the University Hospital Medical Information Network: UMIN000014164.

Bifidobacterium breve prevents necrotising enterocolitis by suppressing inflammatory responses in a preterm rat model.

Necrotising enterocolitis (NEC) is associated with inflammatory responses and barrier dysfunction in the gut. In this study, we investigated the effect of Bifidobacterium breve M-16V on factors related to NEC development using an experimental rat model. Caesarean-sectioned rats were given formula milk with or without B. breve M-16V by oral gavage thrice daily, and experimental NEC was induced by exposing the rats to hypoxic conditions. Naturally delivered rats that were reared by their mother were used as healthy controls. The pathological score of NEC and the expression of molecules related to inflammatory responses and the barrier function were assessed in the ileum. B. breve M-16V reduced the pathological scores of NEC and resulted in some improvement in survivability. B. breve M-16V suppressed the increased expression of molecules related to inflammation and barrier function that resulted from NEC induction. B. breve M-16V normalised Toll-like receptor (TRL)4 expression and enhanced TLR2 expression. Our data suggest that B. breve M-16V prevents NEC development by modulating TLR expressions and suppressing inflammatory responses in a rat model.

Lysozyme in breast milk is a selection factor for bifidobacterial colonisation in the infant intestine.

The objective of this work was to study the residential characteristics of bifidobacteria, which can be classified as either human-residential bifidobacteria (HRB) or non-HRB. We investigated the growth of different strains of HRB and non-HRB in human breast milk with the aim of understanding the mechanisms involved in the unique habitation of each taxon. The growth of 37 strains of different bifidobacterial species or subspecies in breast milk was investigated by incubating each under anaerobic conditions at 37 °C. The tolerance of each strain to either egg white or human lysozyme was compared. Among the infant-type HRB strains, all strains of Bifidobacterium longum subsp. infantis and Bifidobacterium breve grew well in breast milk, but the growth characteristics of B. longum subsp. longum and B. bifidum were strain-dependent. In contrast, the tested strains of adult-type HRB and non-HRB generally failed to grow and died after incubation in breast milk. Most infant-type HRB strains were tolerant to high concentrations of lysozyme, while adult-type HRB strains possessed intermediate tolerance to lysozyme, and non-HRB strains were susceptible to lysozymes of egg white or human origin. These data suggest that breast milk lysozyme content plays a central role in the exclusion of non-HRB, while other factors, together with lysozyme content, are involved in the growth inhibition of adult-type strains in human milk. Our results suggest that infant-type HRB strains would be suitable candidates for use as infant probiotics.

Effect of Bifidobacterium breve B-3 on skin photoaging induced by chronic UV irradiation in mice.

Probiotics have been shown to have a preventative effect on skin photoaging induced by short term UV irradiation, however, the underlying mechanisms and the effect of probiotics on skin photoaging induced by chronic UV irradiation remain unclear. In this study, we investigated the effect of Bifidobacterium breve B-3 on skin photoaging induced by chronic UV irradiation in hairless mice. Mice were irradiated with UVB three times weekly and orally administered B. breve B-3 (2×10(9) cfu/mouse /day) for 7 weeks. Nonirradiated mice and UVB-irradiated mice without probiotic treatment were used as controls. B. breve B-3 significantly suppressed the changes of transepidermal water loss, skin hydration, epidermal thickening and attenuated the damage to the tight junction structure and basement membrane induced by chronic UVB irradiation. Administration of B. breve B-3 tended to suppress the UV-induced interleukin-1ß production in skin (P=0.09). These results suggest that B. breve B-3 could potentially be used to prevent photoaging induced by chronic UV irradiation.

Expression and diagnostic value of proteins in Mycobacterium tuberculosis.

We constructed a prokaryotic expression vector expressing the Mycobacterium tuberculosis protein TB16.3, as well as 3 other proteins, including TB15.3, CFP-10, and Rv2626C, which were purified and analyzed for their effectiveness as detection antibodies. The TB16.3 genes of M. tuberculosis H37Rv genomic DNA were amplified by polymerase chain reaction, inserted into the expression vector pET-30a, and expressed in Escherichia coli. An enzyme-linked immunosorbent assay was used to detect the 4 M. tuberculosis antibodies. Engineered E. coli bacteria expressing TB16.3 and the 3 other proteins were constructed and found mainly to be soluble. For recombinant TB16.3 proteins, serum samples of 118 tuberculosis (TB) patients and 96 healthy controls were analyzed. Sensitivity, specificity, and adjusted concordance rate for the TB16.3 antibody were 72.9, 86.5, and 79.6%, respectively. The positive rate of Rv2626C antibody in TB patients (44.1%) was significantly lower than that in normal controls (75.0%, χ(2) = 20.8, P < 0.01). TB15.3 and TB16.3 were used for simultaneous detection and showed sensitivity, specificity, and repeatability rates of 69.4, 96.9, and 83.7%. The antibody positive rate and specificity for patients with lung disease was 9.6 and 90.4%, respectively. TB15.3 and TB16.3 were mixed and detected simultaneously. Combined with the results for TB15.3, the sensitivity, specificity, and concordance rates were 82.2, 95.9, and 88.9%, respectively. The concordance rate was the highest value observed. Target genes were cloned into a host strain and expressed successfully. The TB16.3 recombinant protein may be used as a new serological antigen for tuberculosis diagnosis.

Expression and serological diagnosis of Mycobacterium tuberculosis CFP-10 and Rv2626c proteins.

We constructed a Mycobacterium tuberculosis vector expressing CFP-10 and Rv2626c to examine the expression of these proteins in Escherichia coli as well as their immunoreactivity. The CFP-10 and Rv2626c genes were amplified from tuberculosis H37Rv genomic DNA using polymerase chain reaction. They were ligated into the expression vector PET30a and expressed in E. coli. Histidine tag nickel column chromatography was used to purify the recombinant protein. An enzyme-linked immunosorbent assay (ELISA) was used for detection. In our E. coli-engineered bacteria containing a CFP10 and Rv2626c plasmid, the target protein was found mainly to be in the soluble form. We formed mixed antigens of the recombinant CFP10 and Rv2626c proteins. ELISA results showed that in 214 blood samples, the positive rate was 77.1%. The target gene was successfully expressed in the host strain. Mixed antigens of the recombinant CFP-10 and Rv2626c proteins can be used as a combination antigen in the serological diagnosis of tuberculosis.

Bifidobacterium breve B-3 exerts metabolic syndrome-suppressing effects in the liver of diet-induced obese mice: a DNA microarray analysis.

We previously reported that supplementation with Bifidobacterium breve B-3 reduced body weight gain and accumulation of visceral fat in a dose-dependent manner, and improved serum levels of total cholesterol, glucose and insulin in a mouse model of diet-induced obesity. In this study, we investigated the expression of genes in the liver using DNA microarray analysis and q-PCR to reveal the mechanism of these anti-obesity effects in this mouse model. Administration of B. breve B-3 led to regulated gene expression of pathways involved in lipid metabolism and response to stress. The results indicate that these regulations in the liver are related to the anti-metabolic syndrome effects of B. breve B-3.

New screening methods for probiotics with adhesion properties to sialic acid and sulphate residues in human colonic mucin using the Biacore assay.

AIMS: To determine the relationship between adhesive ability of probiotics and acidic residues in human colonic mucin, we developed a new screening method using Biacore to evaluate adherence of bacteria before and after sialic acid or sulphate residues were blocked or removed from mucin. METHODS AND RESULTS: Ten strains of lactobacilli and three strains of bifidobacteria isolated from human faeces were evaluated for their adhesive properties to soluble human colonic mucin (sHCM) using the Biacore binding assay. Three strains (Lactobacillus strain ME-522, Lact. gasseri ME-527 and Bifidobacterium bifidum MCC1092) showing significant adherence were selected. Decreased binding activities were observed after removing sialic acid of sHCM using sialidase. However, after removing the sulphate residue using sulphatase, the adhesion of ME-527 decreased; whereas the remaining two strains had increased adhesion. The adhesion of three probiotics significantly decreased after the sulphate residue was blocked by elution with barium chloride. CONCLUSIONS: A new evaluation method using the Biacore assay was developed to observe binding properties to the acidic residues of sHCM. Results indicated that there was a strong relationship between probiotic adhesion and acidic residues of sHCM. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report showing a screening method that quantitatively measures the binding between bacteria and acidic residues in sHCM using the Biacore binding assay; and provides a new method for the selection of probiotics in the future.