Non-targeted Way Research Articles

Insights into the molecular mechanisms of pesticide tolerance in the Aporrectodea caliginosa earthworm

Diffuse pollution of the environment by pesticides has become a major soil threat to non-target organisms, such as earthworms for which declines have been reported. However some endogeic species are still abundant and persist in intensively cultivated fields, suggesting they become tolerant to long-term anthropogenic pressure. We thus considered the working hypothesis that populations of Aporrectodea caliginosa earthworms from conventionally managed fields developed a tolerance to pesticides compared with those from organically managed fields. To investigate this hypothesis, we studied earthworm populations of the same genetic lineage from soils that were either lowly or highly contaminated by pesticides to detect any constitutive expression of differentially expressed molecular pathways between these populations. Earthworm populations were then experimentally exposed to a fungicide—epoxiconazole—in the laboratory to identify different molecular responses when newly exposed to a pesticide. State-of-the-art omics technology (RNA sequencing) and bioinformatics were used to characterize molecular mechanisms of tolerance in a non-targeted way. Additional physiological traits (respirometry, growth, bioaccumulation) were monitored to assess tolerance at higher levels of biological organization. In the present study, we generated the de novo assembly transcriptome of A. caliginosa consisting of 64,556 contigs with N50 = 2862 pb. In total, 43,569 Gene Ontology terms were identified for 21,593 annotated sequences under the three main ontologies (biological processes, cellular components and molecular functions). Overall, we revealed that two same lineage populations of A. caliginosa earthworms, inhabiting similar pedo-climatic environment, have distinct gene expression pathways after they long-lived in differently managed agricultural soils with a contrasted pesticide exposure history for more than 22 years. The main difference was observed regarding metabolism, with upregulated pathways linked to proteolytic activities and the mitochondrial respiratory chain in the highly exposed population. This study improves our understanding of the long-term impact of chronic exposure of soil engineers to pesticide residues.

Unwrapping Wrap-around in Gas (or Liquid) Chromatographic Cyclic Ion Mobility-Mass Spectrometry.

Gas chromatography multiplexed with cyclic ion mobility mass spectrometry is a comprehensive two-dimensional separation technique that can resolve compounds that would otherwise coelute in a single-dimension separation. The cyclic geometry of the ion mobility cell enables ions to travel multiple passes, increasing their drift times to the detector and relative separation. However, the quality of the separation may be obfuscated when "wrap-around" occurs, during which speedier ions catch up with slower ion populations when allowed to travel through more than one pass. Consequently, cyclic ion mobility is incorrectly perceived as a targeted approach that requires preselection of ions prior to separation. The present study demonstrates that "wrap-around" can be mitigated by comparing drift times measured during single- and multipass experiments and extrapolating the number of passes experienced by each ion. This straightforward calculation results in the "unwrapping" of cyclic ion mobility data so that the experiments can be interpreted in a nontargeted way while reaping the benefit of peak capacities that rival those achieved using other comprehensive two-dimensional separations.

MO652: Comparison of Two Medium Cut-Off Dialysers Versus Online Haemodiafiltration Regarding Albumin Loss and the Removal of a Broad Spectrum of Molecules

Abstract BACKGROUND AND AIMS Ideally, the goal of haemodialysis (HD) is to maximize the removal of molecules smaller than 50 000 Daltons and minimize albumin loss. The techniques that seem to best achieve this purpose are online haemodiafiltration (HDF) and, recently, HD with medium cut-off membranes. On the other hand, we now have new methods, such as metabolomics, which allow us to detect potential new uraemic toxins in a non-targeted way. The aim of this study is to compare, in terms of efficacy and safety, online HDF using Fx800Cordiax, with 2 medium cut-off dialysers: Theranova 500 and the new Elisio21 HX dialyser. METHOD A total of 14 patients following a baseline treatment with online HDF using Fx800Cordiax were included. Each patient was considered his/her own control and was randomized to receive a consecutive 1-week haemodialysis treatment with Theranova 500 and Elisio 21HX. Pre- and post-HD blood samples were obtained on the middle day of the week with the three dialysers and proportional volumes of dialysate effluent were collected to quantify albumin loss during the session. We compared the reduction rate of differently sized molecules, as well as the variation rate in molecules smaller than 1000 Daltons, detected and quantified by NMR based chemometrics. For the analysis of differences, we used PLS-DA (Partial Least Squares-Discriminant Analysis) and ANOVA with multiple comparisons (Bonferroni). RESULTS The figure shows the result of the PLS-DA analysis carried out via the MetaboAnalyst web platform, showing, in terms of variable importance in projection (VIP) scores, the importance of different metabolites for the discrimination between the pre-HD and post-HD state regarding each of the three dialysis procedures. The metabolites that generate the 30 most important first signals or spectral bins are shown. The right-hand columns show the relative concentrations of the different metabolites on a colour-coded qualitative scale from red to blue. The predominance of dark blue in post-haemodialysis using online HDF (postCntr) indicates a greater elimination of metabolites with this technique, followed in efficacy by ElisioHX(postE), with a predominance of light blue and Theranova 500 (postT) in third place, with a predominance of orange. In any case, these are semi-quantitative differences. The table shows the reduction rate of different molecules and albumin losses. CONCLUSION In this study, haemodialysis with Theranova 500 has proven to be very similar in efficacy to online HDF, although with a significantly higher albumin loss. Haemodialysis with Elisio21HX resulted in lower removal of molecules around 20 000 Da compared to online HDF, but there was no significant difference compared to haemodialysis with Theranova 500. Albumin loss was significantly higher with Theranova 500 than with Elisio21HX. The semi-quantitative metabolomics study suggests that, in terms of clearance of low molecular weight metabolites (<1 kDa), on-line HDF is more effective, followed by haemodialysis with Elisio HX and finally with Theranova 500.

Targeted and non-targeted unexpected food contaminants analysis by LC/HRMS: Feasibility study on rice

A widely applicable analytical LC/HRMS method based on ion source optimization, data treatment optimization on rice matrix was developed. The effects of key parameters of ion source, and their interactions on ESI response were studied on HPLC-QTOF. Compared with center points, 40% and 20% increase of response factors in the positive and negative mode can be achieved by ion source optimization, respectively. Data processing strategies inspired from metabolomics and multi-targeted analysis were compared and developed using case and control rice samples. Highly automated workflow using XCMS achieved highest mass accuracy, highest detection rate of 96% for 5 μg/kg in a non-targeted way. A clear distinction between the control and contaminated samples by PCA and PLS-DA was also achieved by this workflow using XCMS, even for the concentration of 5 μg/kg.

Comprehensive Profiling by Non-targeted Stable Isotope Tracing Capillary Electrophoresis-Mass Spectrometry: A New Tool Complementing Metabolomic Analyses of Polar Metabolites.

Mass spectrometry (MS) driven metabolomics is a frequently used tool in various areas of life sciences; however, the analysis of polar metabolites is less commonly included. In general, metabolomic analyses lead to the detection of the total amount of all covered metabolites. This is currently a major limitation with respect to metabolites showing high turnover rates, but no changes in their concentration. Such metabolites and pathways could be crucial metabolic nodes (e.g., potential drug targets in cancer metabolism). A stable-isotope tracing capillary electrophoresis-mass spectrometry (CE-MS) metabolomic approach was developed to cover both polar metabolites and isotopologues in a non-targeted way. An in-house developed software enables high throughput processing of complex multidimensional data. The practicability is demonstrated analyzing [U-13 C]-glucose exposed prostate cancer and non-cancer cells. This CE-MS-driven analytical strategy complements polar metabolite profiles through isotopologue labeling patterns, thereby improving not only the metabolomic coverage, but also the understanding of metabolism.

Grape and Wine Metabolomics to Develop New Insights Using Untargeted and Targeted Approaches

Chemical analysis of grape juice and wine has been performed for over 50 years in a targeted manner to determine a limited number of compounds using Gas Chromatography, Mass-Spectrometry (GC-MS) and High Pressure Liquid Chromatography (HPLC). Therefore, it only allowed the determination of metabolites that are present in high concentration, including major sugars, amino acids and some important carboxylic acids. Thus, the roles of many significant but less concentrated metabolites during wine making process are still not known. This is where metabolomics shows its enormous potential, mainly because of its capability in analyzing over 1000 metabolites in a single run due to the recent advancements of high resolution and sensitive analytical instruments. Metabolomics has predominantly been adopted by many wine scientists as a hypothesis-generating tool in an unbiased and non-targeted way to address various issues, including characterization of geographical origin (terroir) and wine yeast metabolic traits, determination of biomarkers for aroma compounds, and the monitoring of growth developments of grape vines and grapes. The aim of this review is to explore the published literature that made use of both targeted and untargeted metabolomics to study grapes and wines and also the fermentation process. In addition, insights are also provided into many other possible avenues where metabolomics shows tremendous potential as a question-driven approach in grape and wine research.

High-throughput untargeted screening of veterinary drug residues and metabolites in tilapia using high resolution orbitrap mass spectrometry

An analytical method was developed and validated for simultaneous analysis of one hundred and thirty-seven veterinary drug residues and metabolites from sixteen different classes in tilapia utilizing an improved fully non-targeted way of data acquisition with fragmentation. The automated on-line extraction procedure was achieved in a simple disposable pipet extraction. Ultrahigh-performance liquid chromatography and electrospray ionization quadrupole Orbitrap high-resolution mass spectrometry (UHPLC Q-Orbitrap) was used for the separation and detection of all the analytes. The methodology was validated by taking into consideration the guidelines specified in European SANCO/12571/2013 Guideline 2013 and Commission Decision 2002/657/EC. The extraction recoveries ranged from 81% to 111%. The limits of decision ranged from 0.01 to 2.73 μg kg−1 and the detection capabilities ranged from 0.01 to 4.73 μg kg−1. The one hundred and thirty-seven compounds behave dynamic 0.1–500 μg kg−1, with correlation coefficient >0.99. The fully non-targeted data acquisition way improves both sensitivity and selectivity for the fragments, which is beneficial for screening performance and identification capability. This validated method has been successfully applied on screening of veterinary drug residues and metabolites in muscle of tilapia, an important and intensively produced fish in aquaculture.

Does Encouragement Matter in Improving Gender Imbalances in Technical Fields? Evidence from a Randomized Controlled Trial.

Does encouragement help address gender imbalances in technical fields? We present the results of one of the first and largest randomized controlled trials on the topic. Using an applied statistics conference in the social sciences as our context, we randomly assigned half of a pool of 3,945 graduate students to receive two personalized emails encouraging them to apply (n = 1,976) and the other half to receive nothing (n = 1,969). We find a robust, positive effect associated with this simple intervention and suggestive evidence that women responded more strongly than men. However, we find that women’s conference acceptance rates are higher within the control group than in the treated group. This is not the case for men. The reason appears to be that female applicants in the treated group solicited supporting letters at lower rates. Our findings therefore suggest that “low dose” interventions may promote diversity in STEM fields but may also have the potential to expose underlying disparities when used alone or in a non-targeted way.

Capillary Electrophoresis in Metabolomics.

Metabolomics is an analytical toolbox to describe (all) low-molecular-weight compounds in a biological system, as cells, tissues, urine, and feces, as well as in serum and plasma. To analyze such complex biological samples, high requirements on the analytical technique are needed due to the high variation in compound physico-chemistry (cholesterol derivatives, amino acids, fatty acids as SCFA, MCFA, or LCFA, or pathway-related metabolites belonging to each individual organism) and concentration dynamic range. All main separation techniques (LC-MS, GC-MS) are applied in routine to metabolomics hyphenated or not to mass spectrometry, and capillary electrophoresis is a powerful high-resolving technique but still underused in this field of complex samples. Metabolomics can be performed in the non-targeted way to gain an overview on metabolite profiles in biological samples. Targeted metabolomics is applied to analyze quantitatively pre-selected metabolites. This chapter reviews the use of capillary electrophoresis in the field of metabolomics and exemplifies solutions in metabolite profiling and analysis in urine and plasma.

Simultaneous quantitative determination, identification and qualitative screening of pesticides in fruits and vegetables using LC-Q-Orbitrap™-MS

A method based on QuEChERS extraction and LC-quadrupole-Orbitrap™ MS detection was established utilising an improved fully non-targeted way of data acquisition with and without fragmentation. A full-scan acquisition event without fragmentation (resolving power 70 000) was followed by five consecutive fragmentation events (variable data independent acquisition – vDIA; resolving power 35 000) where all ions from the full-scan range are fragmented. Compared with fragmentation in a single event (all-ion fragmentation – AIF), this improves both selectivity and sensitivity for the fragment ions, which is beneficial for screening performance and identification capability. The method was validated, using the data from the same measurements, for two types of analysis: quantitation/identification and qualitative screening. The quantitative validation, performed according to the guidelines in SANCO/12571/2013, tested the performance of the method for 184 compounds in lettuce and orange at two spiking levels: 10 and 50 ng g−1. The validation showed that the vast majority of the compounds met the criteria for trueness and precision set in the SANCO guidance document. In the qualitative validation the same 184 compounds were used to test the untargeted screening capabilities of the method. In this validation the compounds were spiked at three levels into 11 different fruit and vegetable matrices, which were measured twice on separate days. Taking all data from the qualitative validation together, an overall detection rate of 92% was achieved at the 10 ng g−1 level, increasing to 98% at 200 ng g−1. A screening detection limit (as defined in the SANCO guidelines) of 10 ng g−1 could be achieved for 134 compounds. For 39 and two pesticides the SDL was 50 and 200 ng g−1, respectively. For the other nine compounds no SDL could be established. The identification (ion ratio) criteria as recommended in the SANCO document could be met for 93% of the detected pesticide/matrix/concentration combinations. The outcome of both validations shows that the described method can be used to combine quantitative analysis and the identification of frequently detected pesticides (so far typically done using triple quadrupole MS/MS) with a qualitative screening to be used for a wide range of less frequent detected compounds in one measurement.

Metabolomics as a tool for drug discovery and personalised medicine. A review.

Studying the effects of drugs on the metabolome constitutes a huge part of the metabolomics discipline. Whether the approach is associated with drug discovery (altered pathways due to the disease that provide future targets and information into the mechanism of action or resistance, etc.) or pharmacometabolomics (studying the outcome of treatment), there have been many aspiring published articles in this area. With specific experimental design, including fingerprinting analysis with different analytical platforms in a non-targeted way, the approach is advancing towards the discovery of markers for the implication of personalised medicine, while also providing information that could help to improve the efficacy and reduce the side effects associated with a treatment. In this review, the evolution of pharmacometabolomics from other areas of drug efficacy metabolomics studies is explored.

Micron trust is like strawberry pie

Purpose – The paper aims to investigate the relationship between employees’ trust toward a company and their commitment to it. It questions if certain levels of trust presented in each department at Micron Technology, Italy, are interrelated with the levels of commitment. Design/methodology/approach – It reveals that a survey at Micron Technology Italy was conducted. Of the 1,580 employees who received the questionnaire, 892 responses were collected, resulting in a 56.46 per cent response rate. Findings – It shows that for departments where trust values are below 6.8, the commitment level follows the trust value: there is a positive relation between these two factors. However, departments where employee trust reaches values higher than 6.8 are not reciprocated by a proportional commitment level that tends to reach a’satisfaction point’. Practical implications – Advances the view that, since commitment does not seem to continuously increase, after a certain point, it is not cost-effective for a company to implement communication strategies aimed at increasing employee trust in a non-targeted way. The need for customized communication strategies emerges as does the importance of understanding internal communication as integrated with other human-resource management levers. Social implications – It argues that managers must communicate with their employees as honestly and directly as possible, particularly during uncertain times, and should encourage the employees’ participation in the decision-making process. Originality/value – It offers an original contribution to the ongoing discussion concerning employee engagement. Employees’ trust does not always lead to employee commitment behaviors inside the organization.

Vitamin D testing: three important issues

In Liverpool we are auditing vitamin D testing and prescribing in primary care, following guidelines issued to GPs in early 2012 to encourage evidenced based testing and prescribing.1 Our data indicates that some GPs are testing in an increasingly non-targeted way. GPs in Liverpool ordered over £100 000 worth of vitamin D tests in 2012, over 10 times the amount spent in 2007. Though more people were identified as deficient, the …

Does internal communication to generate trust always increase commitment?

Purpose– The aim of this paper is to investigate the relationship between employee trust towards the company and their commitment to it. For this reason, this study questioned if certain levels of trust presented in each department at Micron Technology Italy were interrelated with the levels of commitment.Design/methodology/approach– A survey at Micron Technology Italy was conducted. Of the 1,580 employees who received the questionnaire, 892 responses were collected, resulting in a 56.46 per cent response rate.Findings– The research findings show that for departments where trust values are below 6.8, the commitment level follows the trust value: there is a positive relation between these two factors. However, departments where employee trust reaches values higher than 6.8 are not reciprocated by a proportional commitment level that tends to reach a “satisfaction point”.Research limitations/implications– The results are only partially coherent with previous studies. Further research is needed to determine whether these reports hold true under different situations/companies.Practical implications– Since commitment does not seem to be continuously increasing, after a certain point it is not cost-effective for the company to implement communication strategies aimed at increasing employee trust in a non-targeted way. The need for customized communication strategies emerges as does the importance of understanding internal communication as integrated with other human resource management levers.Originality/value– This paper offers an original contribution to the ongoing discussion concerning employee engagement. Employees' trust does not always lead to employee commitment behaviors inside the organization.

Identification of low abundant secreted proteins and peptides from primary culture supernatants of human T-cells

Live cells continually communicate with their surroundings by the secretion of biomolecules, among which proteins and/or peptides are an important class. As such, these protein/peptide signals which end up in the extracellular medium, reflect the state of a cell in a certain condition, and as by definition are potential biomarkers indicative for specific physiological/pathological processes. We here report on a mass spectrometry based method for the detection and analysis of peptides and proteins secreted in a highly complex background, such as cell culture supernatant. Our method, which combines chromatography, high duty cycle tandem mass spectrometry and bio-informatics, enables the detection of interleukin-2 (IL-2), a cytokine secreted by activated T-cells, present in cell supernatant while representing only 0.006‰ of the total protein content. Moreover, the method allows the mass spectrometric analysis of signaling proteins in a non-targeted way and without any prior immunodepletion of the highest abundant cell culture medium proteins. In this study this is exemplified by the detection of yet two other secretory peptides, i.e., the granulins A and B, in the primary culture supernatant of non-activated T-cells.

Imaging Mass Spectrometry Unravels the Mystery of Life

Imaging mass spectrometry (IMS) visualizes the distribution of a wide range of biomolecules in cells and tissue sections. IMS can analyze a variety of biomolecules including unexpected metabolites, without labels in a non-targeted way. We developed AP-MALDI (atmospheric pressure matrix assisted laser desorption/ionization) which is equipped with an atmospheric pressure ion-source chamber. It allows us to analyze flesh samples with minimal loss of intrinsic water or volatile compounds.

Metabolomics-oriented isolation and structure elucidation of 37 compounds including two anthocyanins from Arabidopsis thaliana

In order to conduct metabolomic studies in a model plant for genome research, such as Arabidopsis thaliana (Arabidopsis), it is a prerequisite to obtain structural information for the isolated metabolites from the plant of interest. In this study, we isolated metabolites of Arabidopsis in a relatively non-targeted way, aiming at the construction of metabolite standards and chemotaxonomic comparison. Anthocyanins ( 5 and 7) called A8 and A10 were isolated and their structures were elucidated as cyanidin 3- O-[2- O-(β- d-xylopyranosyl)-6- O-(4- O-(β- d-glucopyranosyl)- E- p-coumaroyl)-β- d-glucopyranoside]-5- O-[6- O-(malonyl)-β- d-glucopyranoside] and cyanidin 3- O-[2- O-(2- O-( E-sinapoyl)-β- d-xylopyranosyl)-6- O-(4- O-(β- d-glucopyranosyl)- E- p-coumaroyl)-β- d-glucopyranoside]-5- O-[β- d-glucopyranoside] from analyses of 1D NMR, 2D NMR ( 1H NMR, NOE, 13C NMR, HMBC and HMQC), HRFABMS, FT-ESI-MS and GC–TOF-MS data. In addition, 35 known compounds, including six anthocyanins, eight flavonols, one nucleoside, one indole glucosinolate, four phenylpropanoids and a derivative, together with three indoles, one carotenoid, one apocarotenoid, three galactolipids, two chlorophyll derivatives, one steroid, one hydrocarbon, and two dicarboxylic acids, were also isolated and identified from their spectroscopic data.

Sharing the wealth. The mechanics of a data release from industry.

In this era of “-omics,” the approach to data collection has widened and become less focused. Researchers will often collect data in a very non-targeted way, generating a large resource that can later be mined to address specific questions. The creation of these resources has created a new

Tailoring product development to strategy: case of a European technology manufacturer

A large body of work on new product development (NPD) has identified a number of general success drivers, which imply `best practice' approaches in NPD. In particular, a newer market or a newer technology renders a project riskier and, therefore, requires a higher hurdle rate on returns. Furthermore, a good NPD process exhibits customer orientation and demand pull, cross-functional co-operation, top management support, existence of a champion, good planning and execution with a strong project manager, and the use of a well-defined process with formal measures. Radically new NPD projects require less structure and more exploration than incremental projects. This article examines whether such general `best practices' are directly applicable to a specific company. We study 90 NPD projects across many different business units in a large diversified European technology manufacturer. We find that the market positioning of new products depends more on the specific portfolio needs of our host company than on general principles of riskiness. In addition, we identify three predominant NPD process approaches in the company, but success differences among them are `muddled' by the non-targeted way they are used. We conclude that there is no `best practice' NPD process. Rather, a company should develop a customized NPD project portfolio and a corresponding mixture of processes, which together meet its strategic innovation needs. We develop a systematic procedure that can help a company to achieve such strategic alignment.