Microbial modification of host long-distance dispersal capacity

Background  

Dispersal plays a key role in shaping biological and ecological processes such as the distribution of spatially-structured populations or the pace and scale of invasion. Here we have studied the relationship between long-distance dispersal behaviour of a pest-controlling money spider,Erigone atra, and the distribution of maternally acquired endosymbionts within the wider meta-population. This spider persists in heterogeneous environments because of its ability to recolonise areas through active long-distance airborne dispersal using silk as a sail, in a process termed 'ballooning'.     

Evidence That Multiple Defects in Lipid Regulation Occur before Hyperglycemia during the Prodrome of Type-2 Diabetes

Background

Blood-vessel dysfunction arises before overt hyperglycemia in type-2 diabetes (T2DM). We hypothesised that a metabolomic approach might identify metabolites/pathways perturbed in this pre-hyperglycemic phase. To test this hypothesis and for specific metabolite hypothesis generation, serum metabolic profiling was performed in young women at increased, intermediate and low risk of subsequent T2DM.

Methods

Participants were stratified by glucose tolerance during a previous index pregnancy into three risk-groups: overt gestational diabetes (GDM; n = 18); those with glucose values in the upper quartile but below GDM levels (UQ group; n = 45); and controls (n = 43, below the median glucose values). Follow-up serum samples were collected at a mean 22 months postnatally. Samples were analysed in a random order using Ultra Performance Liquid Chromatography coupled to an electrospray hybrid LTQ-Orbitrap mass spectrometer. Statistical analysis included principal component (PCA) and multivariate methods.

Findings

Significant between-group differences were observed at follow-up in waist circumference (86, 95%CI (79–91) vs 80 (76–84) cm for GDM vs controls, p<0.05), adiponectin (about 33% lower in GDM group, p = 0.004), fasting glucose, post-prandial glucose and HbA_1c, but the latter 3 all remained within the ‘normal’ range. Substantial differences in metabolite profiles were apparent between the 2 ‘at-risk’ groups and controls, particularly in concentrations of phospholipids (4 metabolites with p≤0.01), acylcarnitines (3 with p≤0.02), short- and long-chain fatty acids (3 with p< = 0.03), and diglycerides (4 with p≤0.05).

Interpretation

Defects in adipocyte function from excess energy storage as relatively hypoxic visceral and hepatic fat, and impaired mitochondrial fatty acid oxidation may initiate the observed perturbations in lipid metabolism. Together with evidence from the failure of glucose-directed treatments to improve cardiovascular outcomes, these data and those of others indicate that a new, quite different definition of type-2 diabetes is required. This definition would incorporate disturbed lipid metabolism prior to hyperglycemia.     

O‐8FALSE NEGATIVE BRONCHIAL CYTOLOGY

Introduction: BK virus nephropathy is an emerging cause of renal transplant failure accounting for allograft loss in 45–50% of recipients between 2–60 months post‐transplantation. The Renal Transplant Unit in Royal Free Hospital has devised a local surveillance programme for screening renal transplant patients at weekly intervals by urinary cytology (UC) and electron microscopy (EM), and confirmation of positive results by plasma PCR and allograft biopsy. Objective: (i) To monitor the implementation of local guidelines; (ii) To compare UC with EM and (iii) To identify areas for improvement. Methods: Decoy cell and EM positive cases were retrieved from the WinPath database for new renal transplant recipients (n = 55) during 1^st November 2004 to 31^st October 2005 in Royal Free Hospital. Plasma PCR was retrieved for positive cases. Results: Up to eight samples were sent for UC at random intervals from 47 patients, and up to 7 were sent for EM from 42 patients. Eleven were UC‐positive and one was EM‐positive. PCR was not requested in UC‐positive cases and 3 out of 10 were positive retrospectively. The EM‐positive case was PCR‐positive but UC‐negative. Discussion: Urine Cytology is more appropriate and cost effective for screening than Electron Microscopy. During the period covered by this audit, samples were sent in an inconsistent fashion after allograft dysfunction rather than for screening. A screening protocol has been agreed and a re‐audit is planned.     

High-throughput metabolic screening of microalgae genetic variation in response to nutrient limitation

Metabolomics - Microalgae produce metabolites that could be useful for applications in food, biofuel or fine chemical production. The identification and development of suitable strains require...     

Evaluation of metabolomics profiles of grain from maize hybrids derived from near-isogenic GM positive and negative segregant inbreds demonstrates that observed differences cannot be attributed unequivocally to the GM trait

Introduction

Past studies on plant metabolomes have highlighted the influence of growing environments and varietal differences in variation of levels of metabolites yet there remains continued interest in evaluating the effect of genetic modification (GM).

Objectives

Here we test the hypothesis that metabolomics differences in grain from maize hybrids derived from a series of GM (NK603, herbicide tolerance) inbreds and corresponding negative segregants can arise from residual genetic variation associated with backcrossing and that the effect of insertion of the GM trait is negligible.

Methods

Four NK603-positive and negative segregant inbred males were crossed with two different females (testers). The resultant hybrids, as well as conventional comparator hybrids, were then grown at three replicated field sites in Illinois, Minnesota, and Nebraska during the 2013 season. Metabolomics data acquisition using gas chromatography–time of flight-mass spectrometry (GC–TOF-MS) allowed the measurement of 367 unique metabolite features in harvested grain, of which 153 were identified with small molecule standards. Multivariate analyses of these data included multi-block principal component analysis and ANOVA-simultaneous component analysis. Univariate analyses of all 153 identified metabolites was conducted based on significance testing (α = 0.05), effect size evaluation (assessing magnitudes of differences), and variance component analysis.

Results

Results demonstrated that the largest effects on metabolomic variation were associated with different growing locations and the female tester. They further demonstrated that differences observed between GM and non-GM comparators, even in stringent tests utilizing near-isogenic positive and negative segregants, can simply reflect minor genomic differences associated with conventional back-crossing practices.

Conclusion

The effect of GM on metabolomics variation was determined to be negligible and supports that there is no scientific rationale for prioritizing GM as a source of variation.

     

Metabolomic approaches reveal that cell wall modifications play a major role in ethylene-mediated resistance against Botrytis cinerea

In Arabidopsis, resistance to the necrotrophic fungus Botrytis cinerea is conferred by ethylene via poorly understood mechanisms. Metabolomic approaches compared the responses of the wild‐type, the ethylene‐insensitive mutant etr1‐1, which showed increased susceptibility, and the constitutively active ethylene mutants ctr1‐1 and eto2 both exhibited decreased susceptibility to B. cinerea. Fourier transform–infrared (FT‐IR) spectroscopy demonstrated reproducible biochemical differences between treatments and genotypes. To identify discriminatory mass‐to‐charge ratios (m/z) associated with resistance, discriminant function analysis was employed on spectra derived from direct injection electrospray ionisation‐mass spectrometry on the derived principal components of these data. Ethylene‐modulated m/z were mapped onto Arabidopsis biochemical pathways and many were associated with hydroxycinnamate and monolignol biosynthesis, both linked to cell wall modification. A high‐resolution linear triple quadrupole‐Orbitrap hybrid system confirmed the identity of key metabolites in these pathways. The contribution of these pathways to defence against B. cinerea was validated through the use of multiple Arabidopsis mutants. The FT‐IR microspectroscopy indicated that spatial accumulation of hydroxycinnamates and monolignols at the cell wall to confine disease was linked ot ethylene. These data demonstrate the power of metabolomic approaches in elucidating novel biological phenomena, especially when coupled to validation steps exploiting relevant mutant genotypes.     

Metabolite extraction from suspension-cultured mammalian cells for global metabolite profiling

Metabolite profiling of industrially important suspension-cultured mammalian cells is being increasingly used for rational improvement of bioprocesses. This requires the generation of global metabolite profiles that cover a broad range of metabolites and that are representative of the cells at the time of sampling. The protocol described here is a validated method for recovery of physiologically relevant amounts of key metabolites from suspension-cultured mammalian cells. The method is a two-step process consisting of initial quenching of the cells (to stop cellular metabolism and allow isolation of the cells) followed by extraction of the metabolites. The cells are quenched in 60% methanol supplemented with 0.85% (wt/vol) ammonium bicarbonate at -40 °C. Metabolites are then extracted from the quenched cells using two 100% methanol extractions followed by a single water extraction. Metabolite samples generated using this protocol are amenable to analysis by mass spectrometry-based techniques (e.g., gas chromatography-mass spectrometry, liquid chromatography-mass spectrometry), NMR spectroscopy and enzymatic assays.