dataesr/scientific-paragraphs-categorization huggingface dataset:
Anti-mouse IgD (clone HB250) was in-house produced. Data were acquired on LSR II (Becton and Dickinson) and analyzed with FlowJo version 10. 1r5 (Tree Star). For cell sorting, single-cell suspensions from pooled pLNs or PP were prepared and stained as described above. Cells were sorted into RPMI 1640/10% FCS medium utilizing FACSAria Fusion (Becton Dickinson) or MoFlo XDP (Beckman-Coulter) cell sorters. Cell fractions are pre-gated on singlets and collected as follows: Naïve CD4 T cells (DAPI -B220 -CD4 + GFP -PD1 -GL7 -); TFH GL7 -(DAPI -B220 -CD4 + GFP -PD1 hi GL7 -); TFH GL7 + (DAPI -B220 -CD4 + GFP -PD1 hi GL7 + ); Tregs (DAPI -B220 -CD4 + GFP + PD1 -GL7 -); and TFR cells (DAPI -B220 -CD4 + GFP + PD1 hi GL7 +/-).
All non-control probes of design ID 026655 have been selected to be printed four times onto one area covering 180K probes (yielding in on-chip quadruplicate Features). Control probes required for proper Feature Extraction software operation were determined and placed automatically by eArray using recommended default settings. 25-40 ng of total RNA were used to prepare aminoallyl-UTP-modified (aaUTP) cRNA (Amino Allyl MessageAmp™ II Kit; #AM1753; Life Technologies) as directed by the company (applying one-round of amplification), except that reaction volumes were halved. The labeling of aaUTP-cRNA was performed by use of Alexa Fluor 555 Reactive Dye (#A32756; LifeTechnologies). Prior to the reverse transcription reaction, 1 µl of a 1:100,000 dilution of Agilent's "One-Color spike-in Kit stock solution" (#5188-5282, Agilent Technologies) was added to a subset of total RNA sample.
cRNA fragmentation, hybridization and washing steps were carried-out as recommended in the "One-Color Microarray-Based Gene Expression Analysis Protocol V5. 7, " except that 250-300 ng of each fluorescently labeled cRNA population were used for hybridization. Slides were scanned on the Agilent Micro Array Scanner G2565CA (pixel resolution 3 µm, bit depth 20). Data extraction was performed with the "Feature Extraction Software V10. 7. 3. 1" using the extraction protocol file "GE1_107_Sep09. xml. " Measurements of on-chip replicates (quadruplicates) were averaged using the geometric mean of processed intensity values of the green channel, "gProcessedSignal" (gPS) to retrieve one resulting value per probe and sample. Single Features were excluded from averaging, if they (i) were manually flagged, (ii) were identified as Outliers by the Feature Extraction Software, (iii) lay outside the interval of "1.
Bioinformatics and statistical analysis The heat maps and the PCA analysis shown in Figure  2  were generated by Qlucore Omics Explorer software tool from the normalized gPS values for selected genes. The n values equal 4-7 animals per group in Figures  1A-D, 3A,D, 4, 5B,  and 7C-F , and three animals in Figures  6C-F. In Figures  6E, F , each dot represents single PP. For the transcriptome analysis (Figures  1E, 2, 3B,C, 5A, 6A,  and 7A,B; Figures S1C, S2 , and S4 in Supplementary Material), each cell fraction (where n = cell fraction) is sorted from pooled organs harvest from 7 to 8 mice (n = 2 in PP and day 25, and n = 3 in day 12). All animals in this study were analyzed at the age of 8-14 weeks old. Mean values of each group are shown in all figures and error bars represent ± SD. All statistical analyses were performed with GraphPad Prism 4.
Transcriptome analysis of cD4 Follicular T cells To elucidate potential differences between follicular T cells in pLN versus PP, transcriptome analyses were done by sorting cells from PP or pLN at d12 and d25 p. i. of Foxp3 mice. A gating strategy applied as shown in Figures  1C, D  was used to obtain GFP -PD1 lo GL7 -(naïve T), GFP + PD1 lo GL7 -(Treg), GFP -PD1 hi GL7 + (TFH), and GFP + PD1 hi GL7 +/-(TFR) cells, respectively, from immunized pLN. Cells of PP origin were sorted accordingly but instead of naïve T cells, GFP -PD1 hi GL7 -cells were isolated. RNA from sorted cells was processed, hybridized on Agilent chips (see Materials and Methods) and the results deposited in the GEO database (accession number: GSE106593). The purity of the sorts was usually ≥95% (Figures  S1A, B  in Supplementary Material) and all PD1 hi cells obtained by the applied gating strategy were also CXCR5 + (data not shown, see also Figure  1A ).
For bacterial killing, 1 × 10 8 B. burgdorferi/mL were treated with ceftriaxone (MilliporeSigma) for 72 h at 5X the minimum inhibitory concentration  (Veinovic et al. , 2013). Ceftriaxone was chosen as the antibiotic as it is the preferred treatment for neurological symptoms according to both United States and European guidelines for human treatment of Lyme disease  (Mygland et al. , 2010; Hu, 2012). Complete killing of B. burgdorferi was confirmed by darkfield microscopy and subculture into fresh BSKII medium (data not shown). For stimulation experiments performed with B. burgdorferi sonicate, cultures were subjected to five pulses (amplitude 4 for 12-15 s each) with a probe sonicator (Fisher Scientific, Pittsburgh, PA, United States; Model 120). A 10:1 MOI was used to stimulate microglia cells with live B.
Differential Phagocytosis-Related Gene Expression in Human Microglia Cells in Response to B. burgdorferi Primary cultures of human microglia cells were stimulated with exponential phase live, antibiotic-killed and antibiotickilled sonicated spirochetes for 24 h. Again, we utilized commercial qPCR arrays to quantify the transcription of a panel of human phagocytosis-related genes. Surprisingly, no genes were upregulated when stimulated with live B. burgdorferi (data not shown). Because antibiotic-killed B. burgdorferi  had elicited robust chemokine expression, we investigated whether primary human microglia would respond to dead B. burgdorferi. Bacteria were either killed with antibiotics or killed with antibiotics and disrupted by sonication. In contrast to live B. burgdorferi, several genes were upregulated more than threefold by dead B.
Bray-Curtis Similarity between Diets and Common Carp Bray-Curtis similarity was calculated to quantify the compositional similarity between FA profiles of potential diets and common carp. All data of relative FA composition in corresponding diets and carp were log 10 transformed before running Bray-Curtis similarity tests. Log 10 transformation balanced the statistical loading of major and minor FA. Similarity tests were run per treatment and year following the equation: d jk ¼ P n-1 k¼0 j x ji -x ki j P n-1 k¼0 x ji + x ki À Á : ð4Þ In the equation, d jk is the Bray-Curtis similarity between the objects j (FA composition of potential diets) and k (FA composition of common carp), i the index of a variable, and n the total number of variables x. Subsequently, calculated data from Bray-Curtis similarity were plotted against total lipid contents and fitted to a regression model.
Statistical Analysis In this study, natural fish ponds that differed in size were used according to semi-intensive carp aquaculture practice, thus it was not possible to replicate them. As a consequence, we used a continuous variable, that is, total lipid contents in dorsal fish fillets, as a predictor to examine the similarity between dietary and dorsal fillets' FA compositions across all treatments. This continuous predictor variable was a priori independent of fish ponds. Furthermore, we used stable isotopes to assess the feeding difference among the treatments as a measure of diet effects. All statistical tests and curve fits were performed using SigmaPlot 12. Differences between means of total lipids and individual FA in fish, zooplankton, and supplementary feeds of each experimental year, respectively, (Tables  A2-A10 ) were analyzed using one-way ANOVA followed by Tukey 0 s HSD post hoc test after ln-transformation of all data to obtain normal distribution.
Acknowledgements We thank  Z. Changizi-Magrhoor  and  J. Watzke  ( WasserCluster Lunz ), and  M. Watzka  ( University of Vienna ) for analytical support,  Teichwirtschaft Kainz  for managing the experimental ponds and Garant Tiernahrung (Pöchlarn; Austria) for providing commercial fish feeds. Two anonymous reviewers provided helpful comments on how to improve this manuscript. This work was supported by the  Austrian Science Fund  ( FWF  project  L516-B17 ).
Spectral unmixing was performed using ZEN software by Zeiss. Finally, the images were processed using Adobe Photoshop (Adobe Systems, USA). 2. 7. Quantitative Image Analysis. Image stacks were analyzed using Imaris software (Bitplane, Zurich, Switzerland). Neutrophils and A. fumigatus conidia along with the epithelial and smooth muscle layers were identified and processed via "three-dimensional surface rendering" of the appropriate channel, as previously described  [9, 24]. The threshold and filter settings were optimized by visually comparing the result with the maximum-intensity projection. Based on the epithelium and smooth muscle layer position, the "Crop 3D" function was applied to each image to obtain the appropriate region for quantification. Neutrophil numbers were automatically calculated from the respective surface objects.
Visual inspection was performed to confirm the accuracy of the automated quantitation results. 2. 8. Statistical Analysis. Data are presented as scattered dot plots of the means or means ± SD for at least 3 mice. Differences between two groups were analyzed with the Mann-Whitney U test using GraphPad Prism version for Windows (GraphPad Software, USA, http://www. graphpad. com). A p value less than 0. 05 was considered statistically significant.
Furthermore, Moses et al. [45]  observed residual neutrophil survival after depletion with anti-Ly6G. Thus, it is also possible that the residual neutrophils observed in the studies described above were safely harbored in the airway wall during immunosuppression and neutrophil depletion. In the present study, we detected neutrophils in the conducting airway wall of mice that were treated with a combination of cyclophosphamide and cortisone acetate or neutrophil-depleting antibodies (anti-Gr-1 or anti-Ly6G). Neutrophil surface renderings were generated based on three-dimensional confocal images, allowing precise quantitation of airway wall neutrophils. These data indicate that the amount of conducting airway wall neutrophils in mice following immunosuppression or neutropenia was significantly lower compared to that in immunocompetent mice.
Acknowledgments The authors thank  Professor Sven Krappmann  ( University Hospital Erlangen, Germany ) for providing the Aspergillus fumigatus conidium strain AfS150. This work was supported by the  Russian Foundation for Basic Research Program  (Project no. 16-04-00989  and no. 16-34-01028 ) and by the personal grant from the  Ministry of Education and Science of the Russian Federation  to  Dr. Valentin Borshchevskiy  (Project no. 6. 9909. 2017/ВУ ).
Acknowledgments The authors would like to thank the patients and their families, and the study investigators, study coordinators, and nurses who assisted with the rituximab clinical program. We would also like to thank  Mundipharma  for provision of bendamustine for use in this study. AS is supported by the  Oxford Partnership Comprehensive Biomedical Research Centre  with funding from the  Department of Health's National Institute of Health Research (NIHR) Biomedical Research Centre funding scheme. The views expressed in this publication are those of the authors and not necessarily those of the UK Department of Health. MABLE was sponsored by  F. Hoffmann-La Roche Ltd , with provision of bendamustine and financial support from  Mundipharma. Thirdparty  medical writing assistance, under the direction of the authors, was provided by  Susan Browne, PhD , of  Gardiner-Caldwell Communications (Macclesfield, UK ) and funded by  F.
FUNDING AP was supported by  INSERM. RM and CJ are supported by  Ecole Normale Supérieure de Lyon. RM is part of the  French Laboratory of Excellence  project  ECOFECT  ( ANR-11-LABX-0048 ). SA and HD are supported by  INSERM. KA is supported by  CNRS  and  Aix-Marseille Université. LR was supported by  Sidaction  (Ph. D. grant). M-IT research was supported by  AXA Research  fund as well as by institutional grants from  Institut Pasteur  and the  Centre National de la Recherche Scientifique (CNRS). The authors acknowledge the support of  ARC, La Ligue Contre le Cancer  (Equipe Labellisée  EL2013-3   Mahieux ) and  ANRS.
This result is coherent with the lysosomal swelling phenomenon, observed with confocal fluorescence imaging in figure  1a  and previously reported data obtained with the same particles in 1321N1 cells  [14]. Similar data were obtained with a different dye combination, namely lysosomotropic dye LTR and viability dye TO-PRO-3, to exclude the artefacts potentially caused by interference between fluorescence dyes. The results (electronic supplementary material, figure  S4 ) also show a population with LTR2/TO-PRO-3-(highlighted in the cyan box) upon 3 h exposure to NH 2 -PS NPs, with an increase of LTR intensity of LTRþ/TO-PRO-3-at the later exposure time points, confirming the above results obtained with LTG/PI staining. We further assessed the destabilization of lysosomes after NP treatment by ultrastructure transmission electron microscopy (TEM) analysis.
To assess the autophagic flux, we have compared the difference of LC3-II levels after NP treatment with or without lysosomal inhibitor HCQ over time. The resulting plot indicates that autophagic flux is slightly induced after 3-6 h treatment of NH 2 -PS NPs; however, after 8 and 24 h exposure to NH 2 -PS NPs, there is a decrease of autophagic flux (figure  4b ). This correlates with the western blot detection of p62 (figure  4a ), a substrate which is degraded by autophagy  [47]. We have first observed a decrease of p62 in the first 6 h of NP treatment (indicative of induced autophagic flux), followed by an increase of its amount after 8 and 24 h (reflecting the blockage of autophagic flux). Together with the fluorescence images shown above (figure  3a, b ), our data strongly indicate that in the first 6 h exposure to NPs, autophagy is functional, while after 8 h NPs induce blockage of autophagic flux, probably due to the fact that lysosomes are severely damaged at this time.
Flow cytometry measurement After exposure to NPs for indicated time points or to positive control t-BuOOH for indicated concentrations and time points, cells were harvested by trypsin and stained with 100 nM LTG (Life Technologies) for 15 min at 378C and 20 mg ml 21 PI (Sigma) for 3 min at room temperature. After staining, the cell fluorescence was immediately measured with a Cyan ADP flow cytometer (Beckman Coulter), using a 488 nm laser to excite both fluorophores, FL1 (520 + 20 nm) band pass filter to collect LTG fluorescence and FL2 (613 + 20 nm) band pass filter to collect PI fluorescence. The data analysis was carried out using SUMMIT software (DAKO). Gates were set to discriminate cell debris and cell doublets according to the forward and side scattering. The dot plots of PI versus LTG were compensated with proper controls.
For the other western blot experiments, cells were treated with NPs for indicated time points and cellular proteins were extracted with home-made lysis buffer (50 mM Tris-HCl pH 6. 8, glycerol 10% (v/v), 2% SDS (w/v), 10 mM DTT and 0. 005% bromophenol blue) and subjected to western blot as previously described  [23]. LC3 antibody was purchased from Sigma. Antibodies against p62, pAKT, pS6 were purchased from Cell Signaling (Danvers, MA, USA), and GAPDH from Abcam. Densitometry was performed using IMAGEJ software (Nationcal Institute of Health).
Acknowledgements. We thank  Prof. Kenneth Dawson  ( Center for BioNano Interactions, University College Dublin, UCD ) for supporting previous studies that serve as the base of this work and also giving constructive advice for this study. We thank  Prof. Sylviane Muller  ( CNRS-University of Strasbourg, Biotechnology and Cell Signaling ) for her support. We also thank  Dr Alfonso Blanco  ( Flow Cytometry Core Facilities  of the  UCD Conway Institute ) and  Fernando Escolar  ( Electronic microscopy service of Centro de Investigaciones Biologicas ) for their technical support.
The fluorescence of samples was then read at 480/535 nm using an Enspire Multimode Plate Reader (Perkin Elmer, Courtaboeuf, France). Fluorescence in samples treated with ConA and saliva was compared with fluorescence measured with ConA alone to calculate the relative PBMC activation percentage. Similarly, fluorescence in unstimulated PBMC treated with saliva was compared with fluorescence measured in unstimulated cells. Experiments were performed using cells from three different animals  (1973, 9567, and 9906) , each of the three saliva batches being tested separately. Data are presented as relative PBMC activation, means of values obtained for the three batches of saliva (for each concentration) in triplicate experiments for each of the three animals.
The reaction was stopped with H 2 SO 4 0. 5 M and the plates were read at 450 nm. All the results were determined with standard curves obtained with serial dilutions of recombinant proteins (ref. Cells in the 48-wells plate were collected and labeled with mouse anti bovine monoclonal antibodies for analysis of costimulation surface markers: MHC II (clone J11, kindly provided by ILRI, Kenya), CD40 (clone IL-A156, AbD Serotec, UK), CD80 (clone IL-A159, AbD Serotec, UK), and CD86 (clone IL-A190, AbD Seroteck, UK). Isotypic control was used as non-specific labeling (MCA928, AbD Serotec, UK). Antibodies were revealed using goat anti mouse IgG1-RPE (STAR132PE, AbD Serotec, UK), cells were processed for flow cytometry using a FacsCanto II cytometer (BD Biosciences, USA) and data were analyzed using Diva software (BD Biosciences, USA) for FCS/SSC and FL2 fluorescence intensities.
All MS/MS spectra were recorded using normalized collision energy (33%, activation Q 0. 25 and activation time 10 ms) with an isolation window of 2 m/z. Data were acquired using Xcalibur software (v 2. 2). For all full scan measurements with the Orbitrap detector a lock-mass ion from ambient air (m/z 445. 120024) was used as an internal calibrant as described in  Olsen et al. (2005). MS data were analyzed using the MaxQuant software package (v 1. 5. 0. 0) as described by  Cox and Mann (2008). Tandem mass spectra (MS/MS) were searched for using the Andromeda search engine  (Cox et al. , 2011)  against the UniProtKB database (release 2015_03) for the A. variegatum (AMBVA) taxonomy (753 entries), Amblyomma "all species" taxonomy (22,429 entries), and the UniProtKB proteome UP000001555 database for Ixodes scapularis (20,473 entries) using the following parameters: enzyme specificity was set as Trypsin/P, and a maximum of two missed cleavages and a mass tolerance of 0.
5 Da for fragment ion were applied. A second database of known contaminants provided with the MaxQuant suite was also used. The "match between runs" option was checked. Oxidation (M) was specified as variable modification and carbamidomethyl (C) as fixed modification. Database searches were performed with a mass tolerance of 20 ppm for precursor ion for mass calibration, and with a 4. 5 ppm tolerance after calibration. The maximum false peptide and protein discovery rate was set at 0. 01. The MaxQuant software generates several output files that contain information about the peptides and proteins identified. The "proteinGroups. txt" file is dedicated to identified proteins: each single row collapses all proteins that cannot be distinguished based on identified peptides into protein groups.
In addition, other properties like molecular weight, bulky residue content, small residue content, average hydrophobicity, average charge a. s. o. and the content of selected dipeptidegroups are also calculated from the sequence. A total of 144 such properties are weighted individually and used as query vectors. The weights have been trained on a set of protein families with known structures, using a genetic algorithm. Sequences in the database are also transformed into vectors, and the Euclidian distance between the query and database sequences is calculated. Distances are rank ordered, and sequences with lowest distance are reported on top  (Hobohm and Sander, 1995). We used Propsearch first to better characterize proteins identified as "Putative Uncharacterized Protein" and second to improve the potential functional information on the identified proteins.
Molecular Characterization of A. variegatum Saliva Proteomics analysis of A. variegatum crude saliva led to the identification of 336 proteins, with no significant qualitative and quantitative variation between individuals (PRIDE PXD007821), allowing us to use pools of saliva for both cell biology assays and molecular characterization. The protein sequence data, consisting of the Amblyomma taxonomy (22,429 entries) and the I. scapularis proteome (UP000001555, 20,473 entries), were retrieved from UniProtKB (release 2015_03) for identification purposes. Among the 336 proteins identified, only 58 (17. 3%) A. variegatum specific proteins were identified, knowing that the UniProtKB A. variegatum specific/particular database contained 753 entries (UniProtKB release 2015_03). Other identified proteins were divided into 236 Amblyomma (70.
control unstimulated cells were measured by flow cytometry. Blue lines and shading show the linear regression fit and the 0. 95 confidence interval, respectively. The p-value of the model is indicated on the panel. Supplementary Figure  2  | Effect of A. variegatum saliva on the production of cytokines by bovine unstimulated macrophages. Bovine blood monocyte-derived macrophages were stimulated for 24 h with different concentrations of A. variegatum saliva. Cell culture supernatants were collected and IL-10, IL-12, and TNF-α production was titrated by ELISA. (Data are results obtained for one experiment performed on three different animals; note different scales on graphs). Blue lines and shading show the linear regression fit and the 0. 95 confidence interval, respectively. The p-value of the model is indicated on the panel.
ACKNOWLEDGMENTS This research was supported by  CIRAD  and  INRA  and a collaborative project on A. variegatum with  USDA-Agricultural Research Service. This study was partly conducted in the framework of the project  MALIN  " Surveillance , diagnosis, control and impact of infectious diseases of humans, animals and plants in tropical islands" supported by the  European Regional Development Fund (ERDF)  and the  Regional Council of Guadeloupe. Mass spectrometry experiments were carried out using facilities of the  Functional Proteomics Platform of Montpellier.
Added value of this study To our knowledge, we report the first use of WGS in conjunction with relevant epidemiological and phenotypic data in an international programme for regional surveillance of sexually transmitted infections and we improve the understanding of the distribution of resistant clones in different risk groups nationally and regionally. For simplified, appropriate analysis in gonococcal genomic projects, a publicly available and user-friendly web application was adapted and optimised for N gonorrhoeae and used for all analyses and dissemination of results to participating countries. We used WGS to investigate 1054 Euro-GASP isolates (consecutive susceptible and resistant isolates) that were collected in 20 European countries in 2013 and we describe the genomic baseline of the European gonococcal population, identify antimicrobial resistant clades and clones and their associations with patient metadata, predict antimicrobial resistance, and compare our results with a European molecular survey from 2009-10.