Determination of the in vivo prebiotic potential of a maize based wholegrain breakfast cereal: a human feeding study

Epidemiological studies have shown an inverse relationship between risk of CVD and intake of whole grain (WG)-rich food. Regular consumption of breakfast cereals can provide not only an increase in dietary WG but also improvements to cardiovascular health. Various mechanisms have been proposed, including prebiotic modulation of the colonic microbiota. In the present study, the prebiotic activity of a maize-derived WG cereal (WGM) was evaluated in a double-blind, placebo-controlled human feeding study (n 32). For a period of 21 d, healthy men and women, mean age 32 (sd 8) years and BMI 23·3 (sd 0·58) kg/m2, consumed either 48 g/d WG cereal (WGM) or 48 g placebo cereal (non-whole grain (NWG)) in a crossover fashion. Faecal samples were collected at five points during the study on days 0, 21, 42, 63 and 84 (representing at baseline, after both treatments and both wash-out periods). Faecal bacteriology was assessed using fluorescence in situ hybridisation with 16S rRNA oligonucleotide probes specific for Bacteroides spp., Bifidobacterium spp., Clostridium histolyticum/perfringens subgroup, Lactobacillus–Enterococcus subgroup and total bacteria. After 21 d consumption of WGM, mean group levels of faecal bifidobacteria increased significantly compared with the control cereal (P = 0·001). After a 3-week wash-out period, bifidobacterial levels returned to pre-intervention levels. No statistically significant changes were observed in serum lipids, glucose or measures of faecal output. In conclusion, this WG maize-enriched breakfast cereal mediated a bifidogenic modulation of the gut microbiota, indicating a possible prebiotic mode of action

Normative, systemic and procedural aspects: a review of indicator‐based sustainability assessments in agriculture

Several methods for assessing the sustainability of agricultural systems have been developed. These methods do not fully: (i) take into account the multi‐functionality of agriculture; (ii) include multidimensionality; (iii) utilize and implement the assessment knowledge; and (iv) identify conflicting goals and trade‐offs. This paper reviews seven recently developed multidisciplinary indicator‐based assessment methods with respect to their contribution to these shortcomings. All approaches include (1) normative aspects such as goal setting, (2) systemic aspects such as a specification of scale of analysis, (3) a reproducible structure of the approach. The approaches can be categorized into three typologies. The top‐down farm assessments focus on field or farm assessment. They have a clear procedure for measuring the indicators and assessing the sustainability of the system, which allows for benchmarking across farms. The degree of participation is low, potentially affecting the implementation of the results negatively. The top‐down regional assessment assesses the on‐farm and the regional effects. They include some participation to increase acceptance of the results. However, they miss the analysis of potential trade‐offs. The bottom‐up, integrated participatory or transdisciplinary approaches focus on a regional scale. Stakeholders are included throughout the whole process assuring the acceptance of the results and increasing the probability of implementation of developed measures. As they include the interaction between the indicators in their system representation, they allow for performing a trade‐off analysis. The bottom‐up, integrated participatory or transdisciplinary approaches seem to better overcome the four shortcomings mentioned above.

Community patterns of soil bacteria and nematodes in relation to geographic distance

Ecosystems consist of aboveground and belowground subsystems and the structure of their communities is known to change with distance. However, most of this knowledge originates from visible, aboveground components, whereas relatively little is known about how soil community structure varies with distance and if this variability depends on the group of organisms considered. In the present study, we analyzed 30 grasslands from three neighboring chalk hill ridges in southern UK to determine the effect of geographic distance (1e198 km) on the similarity of bacterial communities and of nematode communities in the soil. We found that for both groups, community similarity decayed with distance and that this spatial pattern was not related to changes either in plant community composition or soil chemistry. Site history may have contributed to the observed pattern in the case of nematodes, since the distance effect depended on the presence of different nematode taxa at one of the hill ridges. On the other hand, site-related differences in bacterial community composition alone could not explain the spatial turnover, suggesting that other factors, such as biotic gradients and local dispersal processes that we did not include in our analysis, may be involved in the observed pattern. We conclude that, independently of the variety of causal factors that may be involved, the decay in similarity with geographic distance is a characteristic feature of both communities of soil bacteria and nematodes.

A randomised crossover study investigating the effects of galacto-oligosaccharides on the faecal microbiota in men and women over 50 years of age

Faecal microbial changes associated with ageing include reduced bifidobacteria numbers. These changes coincide with an increased risk of disease development. Prebiotics have been observed to increase bifidobacteria numbers within humans. The present study aimed to determine if prebiotic galacto-oligosaccharides (GOS) could benefit a population of men and women of 50 years and above, through modulation of faecal microbiota, fermentation characteristics and faecal water genotoxicity. A total of thirty-seven volunteers completed this randomised, double-blind, placebo-controlled crossover trial. The treatments – juice containing 4 g GOS and placebo – were consumed twice daily for 3 weeks, preceded by 3-week washout periods. To study the effect of GOS on different large bowel regions, three-stage continuous culture systems were conducted in parallel using faecal inocula from three volunteers. Faecal samples were microbially enumerated by quantitative PCR. In vivo, following GOS intervention, bifidobacteria were significantly more compared to post-placebo (P = 0·02). Accordingly, GOS supplementation had a bifidogenic effect in all in vitro system vessels. Furthermore, in vessel 1 (similar to the proximal colon), GOS fermentation led to more lactobacilli and increased butyrate. No changes in faecal water genotoxicity were observed. To conclude, GOS supplementation significantly increased bifidobacteria numbers in vivo and in vitro. Increased butyrate production and elevated bifidobacteria numbers may constitute beneficial modulation of the gut microbiota in a maturing population.

Detection and identification of species-specific bacteria associated with synanthropic mites

Internal bacterial communities of synanthropic mites Acarus siro, Dermatophagoides farinae, Lepidoglyphus destructor, and Tyrophagus putrescentiae (Acari: Astigmata) were analyzed by culturing and culture-independent approaches from specimens obtained from laboratory colonies. Homogenates of surface-sterilized mites were used for cultivation on non-selective agar and DNA extraction. Isolated bacteria were identified by sequencing of the 16S rRNA gene. PCR amplified 16S rRNA genes were analyzed by terminal restriction fragment length polymorphism analysis (T-RFLP) and cloning sequencing. Fluorescence in situ hybridization using universal bacterial probes was used for direct bacterial localization. T-RFLP analysis of 16S rRNA gene revealed distinct species-specific bacterial communities. The results were further confirmed by cloning and sequencing (284 clones). L. destructor and D. farinae showed more diverse communities then A. siro and T. putrescentiae. In the cultivated part of the community, the mean CFUs from four mite species ranged from 5.2 × 102 to 1.4 × 103 per mite. D. farinae had significantly higher CFUs than the other species. Bacteria were located in the digestive and reproductive tract, parenchymatical tissue, and in bacteriocytes. Among the clones, Bartonella-like bacteria occurring in A. siro and T. putresecentiae represented a distinct group related to Bartonellaceae and to Bartonella-like symbionts of ants. The clones of high similarity to Xenorhabdus cabanillasii were found in L. destructor and D. farinae, and one clone related to Photorhabdus temperata in A. siro. Members of Sphingobacteriales cloned from D. farinae and A. siro clustered with the sequences of “Candidatus Cardinium hertigii” and as a separate novel cluster.

In vitro fermentation by human gut bacteria of proteolytically digested caseinomacropeptide nonenzymatically glycosylated with prebiotic carbohydrates

The in vitro fermentation selectivity of hydrolyzed caseinomacropeptide (CMP) glycosylated, via Maillard reaction (MR), with lactulose, galacto-oligosaccharides from lactose (GOSLa), and galacto-oligosaccharides from lactulose (GOSLu) was evaluated, using pH-controlled small-scale batch cultures at 37 °C under anaerobic conditions with human feces. After 10 and 24 h of fermentation, neoglyconjugates exerted a bifidogenic activity, similar to those of the corresponding prebiotic carbohydrates. No significant differences were found in Bacteroides, Lactobacillus�Enterococcus, Clostridium histolyticum subgroup, Atopobium and Clostridium coccoides�Eubacterium rectale populations. Concentrations of lactic acid and short-chain fatty acids (SCFA) produced during the fermentation of prebiotic carbohydrates were similar to those produced for their respective neoglycoconjugates at both fermentation times. These findings, joined with the functional properties attributed to CMP, could open up new applications of MR products involving prebiotics as novel multiple-functional ingredients with potential beneficial effects on human health.

In vitro fermentation of alternansucrase raffinose-derived oligosaccharides by human gut bacteria

In this work, in vitro fermentation of alternansucrase raffinose-derived oligosaccharides, previously fractionated according to their degree of polymerization (DP; from DP4 to DP10), was carried out using small-scale pH-controlled batch cultures at 37 °C under anaerobic conditions with human feces. Bifidogenic activity of oligosaccharides with DP4�6 similar to that of lactulose was observed; however, in general, a significant growth of lactic acid bacteria Bacteroides, Atopobium cluster, and Clostridium histolyticum group was not shown during incubation. Acetic acid was the main short chain fatty acid (SCFA) produced during the fermentation process; the highest levels of this acid were shown by alternansucrase raffinose acceptor pentasaccharides at 10 h (63.11 mM) and heptasaccharides at 24 h (54.71 mM). No significant differences between the gas volume produced by the mixture of raffinose-based oligosaccharides (DP5�DP10) and inulin after 24 h of incubation were detected, whereas lower gas volume was generated by DP4 oligosaccharides. These findings indicate that novel raffinose-derived oligosaccharides (DP4�DP10) could be a new source of prebiotic carbohydrates.

Effect of dextransucrase cellobiose acceptor products on the growth of human gut bacteria

The selective fermentation by human gut bacteria of gluco-oligosaccharides obtained from the reaction between the glucosyl group of sucrose and cellobiose, catalyzed by dextransucrases (DSR) from Leuconostoc mesenteroides, has been evaluated. Oligosaccharides were fractionated according to their molecular weight, and their effect on the growth of different bacterial groups was studied. To determine the structure (position and configuration of glycosidic linkages)�function relationship, their properties were compared to those of DSR maltose acceptor products (DSRMal) and of recognized prebiotic carbohydrates (fructo-oligosaccharides, FOS). Cellobiose acceptor products (DSRCel) showed bifidogenic properties similar to those of FOS. However, no significant differences related to molecular weight or isomeric configurations were found for DSRCel and DSRMal products.

Gut microbial catabolism of grape seed flavan-3-ols by human faecal microbiota. Targetted analysis of precursor compounds, intermediate metabolites and end-products.

In vitro batch culture fermentations were conducted with grape seed polyphenols and human faecal microbiota, in order to monitor both changes in precursor flavan-3-ols and the formation of microbial-derived metabolites. By the application of UPLC-DAD-ESI-TQ MS, monomers, and dimeric and trimeric procyanidins were shown to be degraded during the first 10 h of fermentation, with notable inter-individual differences being observed between fermentations. This period (10 h) also coincided with the maximum formation of intermediate metabolites, such as 5-(3′,4′-dihydroxyphenyl)-γ-valerolactone and 4-hydroxy-5-(3′,4′-dihydroxyphenyl)-valeric acid, and of several phenolic acids, including 3-(3,4-dihydroxyphenyl)-propionic acid, 3,4-dihydroxyphenylacetic acid, 4-hydroxymandelic acid, and gallic acid (5–10 h maximum formation). Later phases of the incubations (10–48 h) were characterised by the appearance of mono- and non-hydroxylated forms of previous metabolites by dehydroxylation reactions. Of particular interest was the detection of γ-valerolactone, which was seen for the first time as a metabolite from the microbial catabolism of flavan-3-ols. Changes registered during fermentation were finally summarised by a principal component analysis (PCA). Results revealed that 5-(3′,4′-dihydroxyphenyl)-γ-valerolactone was a key metabolite in explaining inter-individual differences and delineating the rate and extent of the microbial catabolism of flavan-3-ols, which could finally affect absorption and bioactivity of these compounds.

An objective, niche-based approach to indicator species selection

1. Species-based indices are frequently employed as surrogates for wider biodiversity health and measures of environmental condition. Species selection is crucial in determining an indicators metric value and hence the validity of the interpretation of ecosystem condition and function it provides, yet an objective process to identify appropriate indicator species is frequently lacking. 2. An effective indicator needs to (i) be representative, reflecting the status of wider biodiversity; (ii) be reactive, acting as early-warning systems for detrimental changes in environmental conditions; (iii) respond to change in a predictable way. We present an objective, niche-based approach for species' selection, founded on a coarse categorisation of species' niche space and key resource requirements, which ensures the resultant indicator has these key attributes. 3. We use UK farmland birds as a case study to demonstrate this approach, identifying an optimal indicator set containing 12 species. In contrast to the 19 species included in the farmland bird index (FBI), a key UK biodiversity indicator that contributes to one of the UK Government's headline indicators of sustainability, the niche space occupied by these species fully encompasses that occupied by the wider community of 62 species. 4. We demonstrate that the response of these 12 species to land-use change is a strong correlate to that of the wider farmland bird community. Furthermore, the temporal dynamics of the index based on their population trends closely matches the population dynamics of the wider community. However, in both analyses, the magnitude of the change in our indicator was significantly greater, allowing this indicator to act as an early-warning system. 5. Ecological indicators are embedded in environmental management, sustainable development and biodiversity conservation policy and practice where they act as metrics against which progress towards national, regional and global targets can be measured. Adopting this niche-based approach for objective selection of indicator species will facilitate the development of sensitive and representative indices for a range of taxonomic groups, habitats and spatial scales.

High prevalence of multidrug-tolerant bacteria and associated antimicrobial resistance genes isolated from ornamental fish and their carriage water

Background: Antimicrobials are used to directly control bacterial infections in pet (ornamental) fish and are routinely added to the water these fish are shipped in to suppress the growth of potential pathogens during transport. Methodology/Principal Findings: To assess the potential effects of this sustained selection pressure, 127 Aeromonas spp. isolated from warm and cold water ornamental fish species were screened for tolerance to 34 antimicrobials. Representative isolates were also examined for the presence of 54 resistance genes by a combination of miniaturized microarray and conventional PCR. Forty-seven of 94 Aeromonas spp. isolates recovered from tropical ornamental fish and their carriage water were tolerant to >= 15 antibiotics, representing seven or more different classes of antimicrobial. The quinolone and fluoroquinolone resistance gene, qnrS2, was detected at high frequency (37% tested recent isolates were positive by PCR). Class 1 integrons, IncA/C broad host range plasmids and a range of other antibiotic resistance genes, including floR, blaTEM21, tet(A), tet(D), tet(E), qacE2, sul1, and a number of different dihydrofolate reductase and aminoglycoside transferase coding genes were also detected in carriage water samples and bacterial isolates. Conclusions: These data suggest that ornamental fish and their carriage water act as a reservoir for both multi-resistant bacteria and resistance genes.

Development of a miniaturised microarray-based assay for the rapid identification of antimicrobial resistance genes in Gram-negative bacteria

We describe the development of a miniaturised microarray for the detection of antimicrobial resistance genes in Gram-negative bacteria. Included on the array are genes encoding resistance to aminoglycosides, trimethoprim, sulphonamides, tetracyclines and beta-lactams, including extended-spectrum beta-lactamases. Validation of the array with control strains demonstrated a 99% correlation between polymerase chain reaction and array results. There was also good correlation between phenotypic and genotypic results for a large panel of Escherichia coli and Salmonella isolates. Some differences were also seen in the number and type of resistance genes harboured by E. coli and Salmonella strains. The array provides an effective, fast and simple method for detection of resistance genes in clinical isolates suitable for use in diagnostic laboratories, which in future will help to understand the epidemiology of isolates and to detect gene linkage in bacterial populations. (C) 2008 Published by Elsevier B.V. and the International Society of Chemotherapy.

A randomised, double-blind, placebo controlled cross-over study to determine the gastrointestinal effects of consumption of arabinoxylanoligosaccharides enriched bread in healthy volunteers

BACKGROUND: Prebiotics are food ingredients, usually non-digestible oligosaccharides, that are selectively fermented by populations of beneficial gut bacteria. Endoxylanases, altering the naturally present cereal arabinoxylans, are commonly used in the bread industry to improve dough and bread characteristics. Recently, an in situ method has been developed to produce arabinoxylan-oligosaccharides (AXOS) at high levels in breads through the use of a thermophilic endoxylanase. AXOS have demonstrated potentially prebiotic properties in that they have been observed to lead to beneficial shifts in the microbiota in vitro and in murine, poultry and human studies. METHODS: A double-blind, placebo controlled human intervention study was undertaken with 40 healthy adult volunteers to assess the impact of consumption of breads with in situ produced AXOS (containing 2.2 g AXOS) compared to non-endoxylanase treated breads. Volatile fatty acid concentrations in faeces were assessed and fluorescence in situ hybridisation was used to assess changes in gut microbial groups. Secretory immunoglobulin A (sIgA) levels in saliva were also measured. RESULTS: Consumption of AXOS-enriched breads led to increased faecal butyrate and a trend for reduced iso-valerate and fatty acids associated with protein fermentation. Faecal levels of bifidobacteria increased following initial control breads and remained elevated throughout the study. Lactobacilli levels were elevated following both placebo and AXOS-breads. No changes in salivary secretory IgA levels were observed during the study. Furthermore, no adverse effects on gastrointestinal symptoms were reported during AXOS-bread intake. CONCLUSIONS: AXOS-breads led to a potentially beneficial shift in fermentation end products and are well tolerated.