Probiotics, prebiotics, and the host microbiome: the science of translation

Recent advances in our understanding of the community structure and function of the human microbiome have implications for the potential role of probiotics and prebiotics in promoting human health. A group of experts recently met to review the latest advances in microbiota/microbiome research and discuss the implications for development of probiotics and prebiotics, primarily as they relate to effects mediated via the intestine. The goals of the meeting were to share recent advances in research on the microbiota, microbiome, probiotics, and prebiotics, and to discuss these findings in the contexts of regulatory barriers, evolving healthcare environments, and potential effects on a variety of health topics, including the development of obesity and diabetes; the long-term consequences of exposure to antibiotics early in life to the gastrointestinal (GI) microbiota; lactose intolerance; and the relationship between the GI microbiota and the central nervous system, with implications for depression, cognition, satiety, and mental health for people living in developed and developing countries. This report provides an overview of these discussions.

Foodomics for personalized nutrition: how far are we?

To effectively prevent the onset and reduce mortality from noncommunicable diseases, we must consider every individual as metabolically unique to allow for a personalized management to take place. Diet and gut microbiota are major components of the exposome that interact together with a genetic make-up in a complex interplay to result in an individual’s metabolic phenotype. In this context, foodomics approaches (such as nutrigenetics, nutrimetabolomics, nutritranscriptomics, nutriproteomics and metagenomics) are essential tools to assess an individual’s optimal metabolic space. These have recently been applied to large human cohorts to identify specific gene-metabolite, diet-metabolite and gene–diet interactions. As the gut microbiota is a key player in metabolic homeostasis, we suggest following a holistic investigation of metagenome–hyperbolome–diet interactions, the findings of which will provide the basis for developing personalized nutrition and personalized functional foods. However, examining these three-way interactions will only be possible when the challenge of large datasets integration will be overcome.

Hydrocarbon biodegradation and soil microbial community response to repeated oil exposure

Bioremediation strategies continue to be developed to mitigate the environmental impact of petroleum hydrocarbon contamination. This study investigated the ability of soil microbiota, adapted by prior exposure, to biodegrade petroleum. Soils from Barrow Is. (W. Australia), a class A nature reserve and home to Australia’s largest onshore oil field, were exposed to Barrow production oil (50 ml/kg soil) and incubated (25 °C) for successive phases of 61 and 100 days. Controls in which oil was not added at Phase I or II were concurrently studied and all treatments were amended with the same levels of additional nutrient and water to promote microbial activity. Prior exposure resulted in accelerated biodegradation of most, but not all, hydrocarbon constituents in the production oil. Molecular biodegradation parameters measured using gas chromatography–mass spectrometry (GC–MS) showed that several aromatic constituents were degraded more slowly with increased oil history. The unique structural response of the soil microbial community was reflected by the response of different phospholipid fatty acid (PLFA) sub-classes (e.g. branched saturated fatty acids of odd or even carbon number) measured using a ratio termed Barrow PLFA ratio (B-PLFAr). The corresponding values of a previously proposed hydrocarbon degrading alteration index showed a negative correlation with hydrocarbon exposure, highlighting the site specificity of PLFA-based ratios and microbial community dynamics. B-PLFAr values increased with each Phase I and II addition of production oil. The different hydrocarbon biodegradation rates and responses of PLFA subclasses to the Barrow production oil probably relate to the relative bioavailability of production oil hydrocarbons. These different effects suggest preferred structural and functional microbial responses to anticipated contaminants may potentially be engineered by controlled pre-exposure to the same or closely related substrates. The bioremediation of soils freshly contaminated with petroleum could benefit from the addition of exhaustively bioremediated soils rich in biota primed for the impacting hydrocarbons.

Ectomycorrhizal symbiosis can enhance plant nutrition through improved access to discrete organic nutrient patches of high resource quality

It is known that roots can respond to patches of fertility; however, root proliferation is often too slow to exploit resources fully, and organic nutrient patches may be broken down and leached, immobilized or chemically fixed before they are invaded by the root system. The ability of fungal hyphae to exploit resource patches is far greater than that of roots due to their innate physiological and morphological plasticity, which allows comprehensive exploration and rapid colonization of resource patches in soils. The fungal symbionts of ectomycorrhizal plants excrete significant quantities of enzymes such as chitinases, phosphatases and proteases. These might allow the organic residue to be tapped directly for nutrients such as N and P. Pot experiments conducted with nutrient-stressed ectomycorrhizal and control willow plants showed that when high quality organic nutrient patches were added, they were colonized rapidly by the ectomycorrhizal mycelium. These established willows (0.5 m tall) were colonized by Hebeloma syrjense P. Karst. for 1 year prior to nutrient patch addition. Within days after patch addition, colour changes in the leaves of the mycorrhizal plants (reflecting improved nutrition) were apparent, and after I month the concentration of N and P in the foliage of mycorrhizal plants was significantly greater than that in non-mycorrhizal plants subject to the same nutrient addition. It seems likely that the mycorrhizal plants were able to compete effectively with the wider soil microbiota and tap directly into the high quality organic resource patch via their extra-radical mycelium. We hypothesize that ectomycorrhizal plants may reclaim some of the N and P invested in seed production by direct recycling from failed seeds in the soil. The rapid exploitation of similar discrete, transient, high-quality nutrient patches may have led to underestimations when determining the nutritional benefits of ectomycorrhizal colonization.

The commensal infant gut meta-mobilome as a potential reservoir for persistent multidrug resistance integrons

Despite the accumulating knowledge on the development and establishment of the gut microbiota, its role as a reservoir for multidrug resistance is not well understood. This study investigated the prevalence and persistence patterns of an integrase gene (int1), used as a proxy for integrons (which often carry multiple antimicrobial resistance genes), in the fecal microbiota of 147 mothers and their children sampled longitudinally from birth to 2 years. The study showed the int1 gene was detected in 15% of the study population, and apparently more persistent than the microbial community structure itself. We found int1 to be persistent throughout the first two years of life, as well as between mothers and their 2-year-old children. Metagenome sequencing revealed integrons in the gut meta-mobilome that were associated with plasmids and multidrug resistance. In conclusion, the persistent nature of integrons in the infant gut microbiota makes it a potential reservoir of mobile multidrug resistance.

Maternal weaning modulates emotional behavior and regulates the gut-brain axis

Evidence shows that nutritional and environmental stress stimuli during postnatal period influence brain development and interactions between gut and brain. In this study we show that in rats, prevention of weaning from maternal milk results in depressive-like behavior, which is accompanied by changes in the gut bacteria and host metabolism. Depressive-like behavior was studied using the forced-swim test on postnatal day (PND) 25 in rats either weaned on PND 21, or left with their mother until PND 25 (non-weaned). Non-weaned rats showed an increased immobility time consistent with a depressive phenotype. Fluorescence in situ hybridization showed non-weaned rats to harbor significantly lowered Clostridium histolyticum bacterial groups but exhibit marked stress-induced increases. Metabonomic analysis of urine from these animals revealed significant differences in the metabolic profiles, with biochemical phenotypes indicative of depression in the non-weaned animals. In addition, non-weaned rats showed resistance to stress-induced modulation of oxytocin receptors in amygdala nuclei, which is indicative of passive stress-coping mechanism. We conclude that delaying weaning results in alterations to the gut microbiota and global metabolic profiles which may contribute to a depressive phenotype and raise the issue that mood disorders at early developmental ages may reflect interplay between mammalian host and resident bacteria.

Effect of a symbiotic on the response to seasonal influenza vaccination is strongly influenced by degree of immunosenescence

Background Ageing increases risk of respiratory infections and impairs the response to influenza vaccination. Pre- and probiotics offer an opportunity to modulate anti-viral defenses and the response to vaccination via alteration of the gut microbiota. This study investigated the effect of a novel probiotic, Bifidobacterium longum bv. infantis CCUG 52486, combined with a prebiotic, gluco-oligosaccharide (B. longum + Gl-OS), on the response to seasonal influenza vaccination in young and older subjects in a double-blind, randomized controlled trial, taking into account the influence of immunosenescence markers at baseline. Results Vaccination resulted in a significant increase in total antibody titres, vaccine-specific IgA, IgM and IgG and seroprotection to all three subunits of the vaccine in both young and older subjects, and in general, the increases in young subjects were greater. There was little effect of the synbiotic, although it tended to reduce seroconversion to the Brisbane subunit of the vaccine and the vaccine-specific IgG response in older subjects. Immunological characterization revealed that older subjects randomized to the synbiotic had a significantly higher number of senescent (CD28-CD57+) helper T cells at baseline compared with those randomized to the placebo, and they also had significantly higher plasma levels of anti-CMV IgG and a greater tendency for CMV seropositivity. Moreover, higher numbers of CD28-CD57+ helper T cells were associated with failure to seroconvert to Brisbane, strongly suggesting that the subjects randomized to the synbiotic were already at a significant disadvantage in terms of likely ability to respond to the vaccine compared with those randomized to the placebo. Conclusions Ageing was associated with marked impairment of the antibody response to influenza vaccination in older subjects and the synbiotic failed to reverse this impairment. However, the older subjects randomized to the synbiotic were at a significant disadvantage due to a greater degree of immunosenscence at baseline compared with those randomized to the placebo. Thus, baseline differences in immunosenescence between the randomized groups are likely to have influenced the outcome of the intervention, highlighting the need for detailed immunological characterization of subjects prior to interventions.

Microbial culture and checkerboard DNA-DNA hybridization assessment of bacteria in root canals of primary teeth pre- and post-endodontic therapy with a calcium hydroxide/chlorhexidine paste

Aim. To investigate the root canal microbiota of primary teeth with apical periodontitis and the in vivo antimicrobial effects of a calcium hydroxide/chlorhexidine paste used as root canal dressing. Design. Baseline samples were collected from 30 root canals of primary teeth with apical periodontitis. Then, the root canals were filled with a calcium hydroxide paste containing 1% chlorhexidine for 14 days and the second bacteriologic samples were taken prior to root canal filling. Samples were submitted to microbiologic culture procedure to detect root canal bacteria and processed for checkerboard DNA-DNA hybridization. Results. Baseline microbial culture revealed high prevalence and cfu number of anaerobic, black-pigmented bacteroides, Streptococcus, and aerobic microorganisms. Following root canal dressing, the overall number of cfu was dramatically diminished compared to initial contamination (P < 0.05), although prevalence did not change (P > 0.05). Of 35 probes used for checkerboard DNA-DNA hybridization, 31 (88.57%) were present at baseline, and following root canal dressing, the number of positive probes reduced to 13 (37.14%). Similarly, the number of bacterial cells diminished folowing application of calcium hydroxide/chlorhexidine root canal dressing (P = 0.006). Conclusion. Apical periodontitis is caused by a polymicrobial infection, and a calcium hydroxide/chlorhexidine paste is effective in reducing the number of bacteria inside root canals when applied as a root canal dressing.

In vitro antibacterial action of Tetraclean, MTAD and five experimental irrigation solutions

P>Aim To investigate the antibacterial effect of Tetraclean, MTAD and five experimental irrigants using both direct exposure test with planktonic cultures and mixed-species in vitro biofilm model. Methodology Tetraclean, MTAD and five experimental solutions that were modifications of existing formulae including MTAD + 0.01% cetrimide (CTR), MTAD + 0.1% CTR, MTAC-1 (Tween 80 replaced by 0.01% CTR in MTAD), MTAC-2 (Tween 80 replaced by 0.1% CTR) and MTAD-D (MTAD without the Tween 80 and no CTR added) were used as disinfectants in the experiments. In the direct exposure test, a suspension of Enterococcus faecalis was mixed with each of the solutions. After 0.5, 1, 3 and 10 min, an inactivator was added and the number of surviving bacteria was calculated. A mixed-species biofilm from subgingival plaque bacteria was grown in brain heart infusion broth in anaerobic conditions on synthetic hydroxyapatite discs. Two-week-old biofilms were exposed to the solutions for 0.5, 1 and 3 min. The samples were observed by confocal laser scanning microscopy after bacterial viability staining. The scans were quantitatively analysed, and the volume of killed cells of all cells was calculated for each medicament. Results Tetraclean and MTAC-2 (0.1% CTR) killed planktonic E. faecalis in < 30 s. Complete killing of bacteria required 1 min by MTAC-1, 3 min by MTAD + 0.1% CTR and 10 min by MTAD, MTAD-D and MTAD + 0.01% CTR. In the biofilm test, there were significant differences in microbial killing between the different solutions and times of exposure (P < 0.005). MTAC-2 showed the best performance, killing 71% of the biofilm bacteria in 3 min, followed by MTAC-1 and Tetraclean. MTAD and the three MTAD modifications demonstrated the lowest antibacterial activity. Conclusion Tetraclean was more effective than MTAD against E. faecalis in planktonic culture and in mixed-species in vitro biofilm. CTR improved the antimicrobial properties of the solutions, whereas Tween 80 seemed to have a neutral or negative impact on their antimicrobial effectiveness.

It takes guts for tolerance: The phenomenon of oral tolerance and the regulation of autoimmune response

The intestinal tract is a peculiar environment due to its constant contact with the microbiota agents, food antigens and other molecules. Such exposure requires the establishment of important regulatory mechanisms in order to avoid inflammatory response and self aggression. In this context, the GALT plays a very relevant role due to the presence of several different cellular populations which are the main players in this phenomenon. Moreover, it was described a while ago that the oral ingestion of a given molecule is able to induce systemic tolerance to the same molecule when it is used as an immunogen by parenteral route, known as oral tolerance. This observation led researches to use these mechanisms to induce tolerance against cognate antigens of different autoimmune diseases. In this context, in this review we focused on several tolerance inducing mechanisms which are relevant not only for the maintenance of intestinal tract but also for the suppression of T effector cells, such as Th1, Th2 and the newly described Th17 cells. To name a few, CD103(+) dendritic cells, Tr1 cells derived IL-10 secretion, Foxp3 conversion and CD4(+)LAP(+) regulatory cells induction are among the recently described features of the tolerogenic environment of the intestinal tract. (C) 2009 Elsevier B.V. All rights reserved.

Nd : YAG laser clinical assisted in class II furcation treatment

The Nd:YAG laser efficacy associated with conventional treatment for bacterial reduction has been investigated throughout literature. The purpose of this study was to evaluate the bacterial reduction after Nd:YAG laser irradiation associated with scaling and root planning in class II furcation defects in patients with chronic periodontitis. Thirty-four furcation lesions were selected from 17 subjects. The control group received conventional treatment, and the experimental group received the same treatment followed by Nd:YAG laser irradiation (100 mJ/pulse; 15 Hz; 1.5 W, 60 s, 141.5 J/cm(2)). Both treatments resulted in improvements of most clinical parameters. A significant reduction of colony forming unit (CFU) of total bacteria number was observed in both groups. The highest reduction was noted in the experimental group immediately after the treatment. The number of dark pigmented bacteria and the percentage of patients with Porphyromonas gingivalis, Prevotella intermedia, and Actinobacillus actinomycetemcomitans reduced immediately after the treatment and returned to values close to the initial ones 6 weeks after the baseline for both groups. The Nd:YAG laser associated with conventional treatment promoted significant bacterial reduction in class II furcation immediately after irradiation, although this reduction was not observed 6 weeks after the baseline.

Adherence and invasion of Bacteroidales isolated from the human intestinal tract

Members of the genera Bacteroides and Parabacteroides are important constituents of both human and animal intestinal microbiota, and are significant facultative pathogens. In this study, the ability of Bacteroides spp. and Parabacteroides distasonis isolated from both diarrhoeal and normal stools (n = 114) to adhere to and invade HEp-2 cells was evaluated. The presence of putative virulence factors such as capsule and fimbriae was also investigated. Adherence to HEp-2 cells was observed in 75.4% of the strains, which displayed non-localized clusters. Invasion was observed in 37.5% and 26% of the strains isolated from diarrhoeal and non-diarrhoeal stools, respectively. All strains displayed a capsule, whereas none of them showed fimbriae-like structures. This is the first report of the ability of Bacteroides spp. and P. distasonis to adhere to and invade cultured HEp-2 epithelial cells.

Vibrios dominate as culturable nitrogen-fixing bacteria of the Brazilian coral Mussismilia hispida

Taxonomic characterization was performed on the putative N-2-fixing microbiota associated with the coral species Mussismilia hispida, and with its sympatric species Palythoa caribaeorum, P. variabilis, and Zoanthus solanderi, off the coast of Sao Sebastiao (Sao Paulo State, Brazil). The 95 isolates belonged to the Gammaproteobacteria according to the 16S rDNA gene sequences. In order to identify the isolates unambiguously, pyrH gene sequencing was carried out. The majority of the isolates (n = 76) fell within the Vibrio core group, with the highest gene sequence similarity being towards Vibrio harveyi and Vibrio alginolyticus. Nineteen representative isolates belonging to V. harveyi (n = 7), V. alginolyticus (n = 8), V. campbellii (n = 3), and V parahaemolyticus (n = 1) were capable of growing six successive times in nitrogen-free medium and some of them showed strong nitrogenase activity by means of the acetylene reduction assay (ARA). It was concluded that nitrogen fixation is a common phenotypic trait among Vibrio species of the core group. The fact that different Vibrio species can fix N, might explain why they are so abundant in the mucus of different coral species. (C) 2008 Published by Elsevier GmbH.

Occurrence of herpes simplex virus 1 and three periodontal bacteria in patients with chronic periodontitis and necrotic pulp

Viral and bacterial associations appear to be implicated in the development of periodontal infections. Little information is available describing the periodontopathic agents in root canals with necrotic pulp. In this study, the occurrence and the combinations among herpes simplex virus type 1 (HSV-1) and Dialister pneumosintes, Tannerella forsythia.. and Treponema denticola in patients with chronic periodontitis and necrotic pulp were evaluated. Clinical samples from healthy subjects and patients with periodontal or pulp infections were analyzed using a nested polymerase chain reaction PCR to detect HSV and PCR to detect the 3 periodontal bacteria. The presence of Tannerella forsythia and Treponema denticola was observed in healthy, periodontitis, and necrotic pulp patients. HSV was observed in periodontitis and necrotic pulp patients, and no healthy subject harbored D. pneumosintes or HSV. The occurrence of Tannerella forsythia was not statistically significant in patients with necrotic pulp (P = 0.704). Periodontal bacteria were observed varying from 10.3% to 20.7% in periodontitis and necrotic pulp patients. The presence of Treponema denticola - HSV association was predominant in patients showing necrotic pulp (24.1%); however, HSV alone was observed in one patient with periodontitis and in another patient with necrotic pulp. The presence of double association among bacteria or bacteria - HSV could indicate a role in both periodontitis and necrotic pulp, and Tannerella forsythia - Treponenta denticola - HSV and Tannerella forsythia - D. pneumosintes - Treponema denticola - HSV associations might be important in periodontitis.

Prevalence and microbiological diversity of Archaea in peri-implantitis subjects by 16S ribosomal RNA clonal analysis

Background and Objective: This study evaluated the prevalence and the molecular diversity of Archaea in the subgingival biofilm samples of subjects with peri-implantitis. Material and Methods: Fifty subjects were assigned into two groups: Control (n = 25), consisting of subjects with healthy implants; and Test (n = 25), consisting of subjects with peri-implantitis sites, as well as a healthy implant. In the Test group, subgingival biofilm samples were taken from the deepest sites of the diseased implant. In both groups, subgingival biofilm was collected from one site with a healthy implant and from one site with a periodontally healthy tooth. DNA was extracted and the 16S ribosomal RNA gene was amplified with universal primer pairs for Archaea. Amplified genes were cloned and sequenced, and the phylotypes were identified by comparison with known 16S ribosomal RNA sequences. Results: In the Control group, Archaea were detected in two and three sites of the implant and the tooth, respectively. In the Test group, Archaea were detected in 12, 4 and 2 sites of diseased implants, healthy implants and teeth, respectively. Diseased implants presented a significantly higher prevalence of Archaea in comparison with healthy implants and natural teeth, irrespective of group. Over 90% of the clone libraries were formed by Methanobrevibacter oralis, which was detected in both groups. Methanobacterium congelense/curvum was detected in four subjects from the Test group and in two subjects from the Control group. Conclusion: Although M. oralis was the main species of Archaea associated with both healthy and diseased implant sites, the data indicated an increased prevalence of Archaea in peri-implantitis sites, and their role in pathogenesis should be further investigated.