Cannabinoids stimulate fibroblastic colony formation by bone marrow cells indirectly via CB2 receptors

Recently, the cannabinoid receptors CB1 and CB2 were shown to modulate bone formation and resorption in vivo, although little is known of the mechanisms underlying this. The effects of cannabinoids on mesenchymal stem cell (MSC) recruitment in whole bone marrow were investigated using either the fibroblastic colony-forming unit (CFU-f) assay or high-density cultures of whole bone marrow. Levels of the CB1 and CB2 receptors were assessed by flow cytometry. Treatment of CFU-f cultures with the endocannabinoid 2-arachidonylglycerol (2-AG) dose-dependently increased fibroblastic and differentiated colony formation along with colony size. The nonspecific agonists CP 55,940 and WIN 55,212 both increased colony numbers, as did the CB2 agonists BML190 and JWH015. The CB1-specific agonist ACEA had no effect, whereas the CB2 antagonist AM630 blocked the effect of the natural cannabinoid tetrahydrocannabivarin, confirming mediation via the CB2 receptor. Treatment of primary bone marrow cultures with 2-AG stimulated proliferation and collagen accumulation, whereas treatment of subcultures of MSC had no effect, suggesting that the target cell is not the MSC but an accessory cell present in bone marrow. Subcultures of MSCs were negative for CB1 and CB2 receptors as shown by flow cytometry, whereas whole bone marrow contained a small population of cells positive for both receptors. These data suggest that cannabinoids may stimulate the recruitment of MSCs from the bone marrow indirectly via an accessory cell and mediated via the CB2 receptor. This recruitment may be one mechanism responsible for the increased bone formation seen after cannabinoid treatment in vivo.

Antiproliferative activity of Titanocene Y against tumor colony-forming units

Bis-[(p-methoxybenzyl)cyclopentadienyl] titanium dichloride, better known as Titanocene Y, is a newly synthesized titanium-based anticancer drug. We studied the antitumor activity of Titanocene Y with concentrations of 2.1, 21 and 210 μmol/l against a range of freshly explanted human tumors, using an in-vitro soft agar cloning system. The sensitivity against Titanocene Y was highly remarkable in the case of renal cell, ovarian, nonsmall cell lung and colon cancer. In particular the surprisingly good response of nonsmall cell lung cancer and colon cancer against Titanocene Y at its lowest concentration of 2.1 μmol/l was well comparable or better with respect to cisplatin, given at a concentration of 1.0 μmol/l. Further clinical development of Titanocene Y appears to be warranted because of the broad cytotoxic activity shown and the specific activity of Titanocene Y against renal cell cancer.

Confocal imaging of pseudomonas syringae pv. phaseolicola colony development in bean reveals reduced multiplication of strains containing the genomic island PPHGI-1.

Pseudomonas syringae pv. phaseolicola is the seed borne causative agent of halo blight in the common bean Phaseolus vulgaris. Pseudomonas syringae pv. phaseolicola race 4 strain 1302A contains the avirulence gene hopAR1 (located on a 106-kb genomic island, PPHGI-1, and earlier named avrPphB), which matches resistance gene R3 in P. vulgaris cultivar Tendergreen (TG) and causes a rapid hypersensitive reaction (HR). Here, we have fluorescently labeled selected Pseudomonas syringae pv. phaseolicola 1302A and 1448A strains (with and without PPHGI-1) to enable confocal imaging of in-planta colony formation within the apoplast of resistant (TG) and susceptible (Canadian Wonder [CW]) P. vulgaris leaves. Temporal quantification of fluorescent Pseudomonas syringae pv. phaseolicola colony development correlated with in-planta bacterial multiplication (measured as CFU/ml) and is, therefore, an effective means of monitoring Pseudomonas syringae pv. phaseolicola endophytic colonization and survival in P. vulgaris. We present advances in the application of confocal microscopy for in-planta visualization of Pseudomonas syringae pv. phaseolicola colony development in the leaf mesophyll to show how the HR defense response greatly affects colony morphology and bacterial survival. Unexpectedly, the presence of PPHGI-1 was found to cause a reduction of colony development in susceptible P. vulgaris CW leaf tissue. We discuss the evolutionary consequences that the acquisition and retention of PPHGI-1 brings to Pseudomonas syringae pv. phaseolicola in planta.

CAMCR: Computer-Assisted Mixture model analysis for Capture–Recapture count data

Population size estimation with discrete or nonparametric mixture models is considered, and reliable ways of construction of the nonparametric mixture model estimator are reviewed and set into perspective. Construction of the maximum likelihood estimator of the mixing distribution is done for any number of components up to the global nonparametric maximum likelihood bound using the EM algorithm. In addition, the estimators of Chao and Zelterman are considered with some generalisations of Zelterman’s estimator. All computations are done with CAMCR, a special software developed for population size estimation with mixture models. Several examples and data sets are discussed and the estimators illustrated. Problems using the mixture model-based estimators are highlighted.

Assessing the impact of nano- and micro-scale zerovalent iron particles on soil microbial activities: Particle reactivity interferes with assay conditions and interpretation of genuine microbial effects

The effects of nano-scale and micro-scale zerovalent iron (nZVI and mZVI) particles on general (dehydrogenase and hydrolase) and specific (ammonia oxidation potential, AOP) activities mediated by the microbial community in an uncontaminated soil were examined. nZVI (diameter 12.5 nm; 10 mg gÿ1 soil)apparently inhibited AOP and nZVI and mZVI apparently stimulated dehydrogenase activity but had minimal influence on hydrolase activity. Sterile experiments revealed that the apparent inhibition of AOP could not be interpreted as such due to the confounding action of the particles, whereas, the nZVIenhanced dehydrogenase activity could represent the genuine response of a stimulated microbial population or an artifact of ZVI reactivity. Overall, there was no evidence for negative effects of nZVI or mZVI on the processes studied. When examining the impact of redox active particles such as ZVI on microbial oxidation–reduction reactions, potential confounding effects of the test particles on assay conditions should be considered.

Predicting microbial pollution concentrations in UK rivers in response to land use change

The Water Framework Directive has caused a paradigm shift towards the integrated management of recreational water quality through the development of drainage basin-wide programmes of measures. This has increased the need for a cost-effective diagnostic tool capable of accurately predicting riverine faecal indicator organism (FIO) concentrations. This paper outlines the application of models developed to fulfil this need, which represent the first transferrable generic FIO models to be developed for the UK to incorporate direct measures of key FIO sources (namely human and livestock population data) as predictor variables. We apply a recently developed transfer methodology, which enables the quantification of geometric mean presumptive faecal coliforms and presumptive intestinal enterococci concentrations for base- and high-flow during the summer bathing season in unmonitored UK watercourses, to predict FIO concentrations in the Humber river basin district. Because the FIO models incorporate explanatory variables which allow the effects of policy measures which influence livestock stocking rates to be assessed, we carry out empirical analysis of the differential effects of seven land use management and policy instruments (fiscal constraint, production constraint, cost intervention, area intervention, demand-side constraint, input constraint, and micro-level land use management) all of which can be used to reduce riverine FIO concentrations. This research provides insights into FIO source apportionment, explores a selection of pollution remediation strategies and the spatial differentiation of land use policies which could be implemented to deliver river quality improvements. All of the policy tools we model reduce FIO concentrations in rivers but our research suggests that the installation of streamside fencing in intensive milk producing areas may be the single most effective land management strategy to reduce riverine microbial pollution.

The inhibitory effect of natural microflora of food on growth of Listeria monocytogenes in enrichment broths

The aims of this study were to (i) compare the inhibitory effects of the natural microflora of different foods on the growth of Listeria monocytogenes during enrichment in selective and non-selective broths; (ii) to isolate and identify components of the microflora of the most inhibitory food; and (iii) to determine which of these components was most inhibitory to growth of L. monocytogenes in co-culture studies. Growth of an antibioticresistant marker strain of L. monocytogenes was examined during enrichment of a range of different foods in Tryptone Soya Broth (TSB), Half Fraser Broth (HFB) and Oxoid Novel Enrichment (ONE) Broth. Inhibition of L. monocytogenes was greatest in the presence of minced beef, salami and soft cheese and least with prepared fresh salad and chicken pâté. For any particular food the numbers of L. monocytogenes present after 24 h enrichment in different broths increased in the order: TSB, HFB and ONE Broth. Numbers of L. monocytogenes recovered after enrichment in TSB were inversely related to the initial aerobic plate count (APC) in the food but with only a moderate coefficient of determination (R2) of 0.51 implying that microbial numbers and the composition of the microflora both influenced the degree of inhibition of L. monocytogenes. In HFB and ONE Broth the relationship between APC and final L. monocytogenes counts was weaker. The microflora of TSB after 24 h enrichment of minced beef consisted of lactic acid bacteria, Brochothrix thermosphacta, Pseudomonas spp., Enterobacteriaceae, and enterococci. In co-culture studies of L. monocytogenes with different components of the microflora in TSB, the lactic acid bacteria were the most inhibitory followed by the Enterobacteriaceae. The least inhibitory organisms were Pseudomonas sp., enterococci and B. thermosphacta. In HFB and ONE Broth the growth of Gram-negative organisms was inhibited but lactic acid bacteria still reached high numbers after 24 h. A more detailed study of the growth of low numbers of L. monocytogenes during enrichment of minced beef in TSB revealed that growth of L. monocytogenes ceased at a cell concentration of about 102 cfu/ml when lactic acid bacteria entered stationary phase. However in ONE Broth growth of lactic acid bacteria was slower than in TSB with a longer lag time allowing L. monocytogenes to achieve much higher numbers before lactic acid bacteria reached stationary phase. This work has identified the relative inhibitory effects of different components of a natural food microflora and shown that the ability of low numbers of L. monocytogenes to achieve high cell concentrations is highly dependent on the extent to which enrichment media are able to inhibit or delay growth of the more effective competitors.

Variation in antibiotic-induced microbial recolonization impacts on the host metabolic phenotypes of rats

The interaction between the gut microbiota and their mammalian host is known to have far-reaching consequences with respect to metabolism and health. We investigated the effects of eight days of oral antibiotic exposure (penicillin and streptomycin sulfate) on gut microbial composition and host metabolic phenotype in male Han-Wistar rats (n = 6) compared to matched controls. Early recolonization was assessed in a third group exposed to antibiotics for four days followed by four days recovery (n = 6). Fluorescence in situ hybridization analysis of the intestinal contents collected at eight days showed a significant reduction in all bacterial groups measured (control, 1010.7 cells/g feces; antibiotic-treated, 108.4). Bacterial suppression reduced the excretion of mammalian-microbial urinary cometabolites including hippurate, phenylpropionic acid, phenylacetylglycine and indoxyl-sulfate whereas taurine, glycine, citrate, 2-oxoglutarate, and fumarate excretion was elevated. While total bacterial counts remained notably lower in the recolonized animals (109.1 cells/g faeces) compared to the controls, two cage-dependent subgroups emerged with Lactobacillus/Enterococcus probe counts dominant in one subgroup. This dichotomous profile manifested in the metabolic phenotypes with subgroup differences in tricarboxylic acid cycle metabolites and indoxyl-sulfate excretion. Fecal short chain fatty acids were diminished in all treated animals. Antibiotic treatment induced a profound effect on the microbiome structure, which was reflected in the metabotype. Moreover, the recolonization process was sensitive to the microenvironment, which may impact on understanding downstream consequences of antibiotic consumption in human populations.

Colonization-induced host-gut microbial metabolic interaction

The gut microbiota enhances the host's metabolic capacity for processing nutrients and drugs and modulate the activities of multiple pathways in a variety of organ systems. We have probed the systemic metabolic adaptation to gut colonization for 20 days following exposure of axenic mice (n = 35) to a typical environmental microbial background using high-resolution (1)H nuclear magnetic resonance (NMR) spectroscopy to analyze urine, plasma, liver, kidney, and colon (5 time points) metabolic profiles. Acquisition of the gut microbiota was associated with rapid increase in body weight (4%) over the first 5 days of colonization with parallel changes in multiple pathways in all compartments analyzed. The colonization process stimulated glycogenesis in the liver prior to triggering increases in hepatic triglyceride synthesis. These changes were associated with modifications of hepatic Cyp8b1 expression and the subsequent alteration of bile acid metabolites, including taurocholate and tauromuricholate, which are essential regulators of lipid absorption. Expression and activity of major drug-metabolizing enzymes (Cyp3a11 and Cyp2c29) were also significantly stimulated. Remarkably, statistical modeling of the interactions between hepatic metabolic profiles and microbial composition analyzed by 16S rRNA gene pyrosequencing revealed strong associations of the Coriobacteriaceae family with both the hepatic triglyceride, glucose, and glycogen levels and the metabolism of xenobiotics. These data demonstrate the importance of microbial activity in metabolic phenotype development, indicating that microbiota manipulation is a useful tool for beneficially modulating xenobiotic metabolism and pharmacokinetics in personalized health care. IMPORTANCE: Gut bacteria have been associated with various essential biological functions in humans such as energy harvest and regulation of blood pressure. Furthermore, gut microbial colonization occurs after birth in parallel with other critical processes such as immune and cognitive development. Thus, it is essential to understand the bidirectional interaction between the host metabolism and its symbionts. Here, we describe the first evidence of an in vivo association between a family of bacteria and hepatic lipid metabolism. These results provide new insights into the fundamental mechanisms that regulate host-gut microbiota interactions and are thus of wide interest to microbiological, nutrition, metabolic, systems biology, and pharmaceutical research communities. This work will also contribute to developing novel strategies in the alteration of host-gut microbiota relationships which can in turn beneficially modulate the host metabolism.

Microbial utilization and selectivity of pectin fractions with various structures

To evaluate the fermentation properties of oligosaccharides derived from pectins and their parent polysaccharides, a 5-ml-working-volume, pH- and temperature-controlled fermentor was tested. Six pectic oligosaccharides representing specific substructures found within pectins were prepared. These consisted of oligogalacturonides (average degrees of polymerization [DP] of 5 and 9), methylated oligogalacturonides (average DP of 5), oligorhamnogalacturonides (average DP of 10 as a disaccharide unit of galacturonic acid and rhamnose), oligogalactosides (average DP of 5), and oligoarabinosides (average DP of 6). The influence of these carbohydrates on the human fecal microbiota was evaluated. Use of neutral sugar fractions resulted in an increase in Bifidobacterium populations and gave higher organic acid yields. The Bacteroides-Prevotella group significantly increased on all oligosaccharides except oligogalacturonides with an average DP of 5. The most selective substrates for bifidobacteria were arabinan, galactan, oligoarabinosides, and oligogalactosides.

Benchmarking and setting intervention targets for key cell count parameters

Herd Companion uses routine milk‐recording records to generate twelve‐month rolling averages that indicate performance trends. This article looks at Herd Somatic Cell Count (SCC) and four other SCC‐related parameters from 252 National Milk Records (NMR) recorded herds to assess how each parameter correlates with the Herd SCC. The analysis provides evidence for the importance of targeting individual cows with high SCC recordings (>200,000 cells/ml and >500,000 cells/ml) and/or individual cows with repeatedly high SCC recordings (chronic high SCC) and/or cows that begin lactation with a high SCC recording (dry period infection) in order to achieve bulk milk Herd SCC below 200,000 cells/ml.

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.