dataesr/scientific-paragraphs-categorization huggingface dataset:
Experimental Procedures Reagents-siRNA sequences siSTIM1 (siSTIM1 #1, AAGGG-AAGACCUCAAUUACCAUU; siSTIM1 #2, GGUGGUGUC-UAUCGUUAUUUU), siOrai1 (siOrai1 #1, CUGUCCUCUA-AGAGAAUAAUU; siOrai1 #2, CGUGCACAAUCUCAACU-CGUU), and non-targeting siControl (UGGUUUACAUGUC-GACUAAUU) were from Dharmacon Scientific. Anti-Orai1 (rabbit) antibody was from Sigma (HPA016583), used at a dilution of 1:1000. Anti-STIM1 (mouse, raised using STIM1 amino acids 25-139) antibody is from BD Transduction Laboratories, used at a dilution of 1:250. Anti-␤-actin is from Sigma and used at a dilution of 1:10,000. Anti-GAPDH (mouse) is from Sigma (catalog no. G8795), used at a dilution of 1:3000, and anti-HSP 70/HSC 70 (W27) (mouse) from Santa Cruz Biotechnology (catalog no. sc-24) was used at a dilution of 1:2000. Anti-rabbit IgG-HRP and anti-mouse IgG-HRP are from Calbiochem used at 1:10,000.
Single frequency time collect mode allowed detecting HDMEC response to GPCR agonists with a time resolution of 8 s for 16 wells or 48 s for 96 wells, respectively. For data presentation the resistance, the real part of the impedance data, which is most indicative of paracellular currents via cell-cell junctions was normalized to the last value before agonist addition (normalized resistance) and plotted versus time. For measurements of the complete attachment, growth and cell layer maturation process electrodes were pretreated with 10 mM cysteine solution 1 day before cell seeding. After 10 min of incubation with 150 l of cysteine solution, arrays were rinsed three times with water and incubated in HDMEC medium overnight to support establishment of stable electrode-electrolyte interface. HDMEC attachment, spreading, and cell layer maturation was measured with the multiple frequency time collect mode.
A baseline of the cell-free array with 400 l of medium in each well was measured over at least 1 h. Data acquisition was paused for inoculation with single-cell suspension (100,000 cells/cm 2 ) and for medium changes. Monitoring of HDMEC Response to GPCR Agonists-Thromin (catalog no. T464), histamine (catalog no. 53300) and S1P (catalog no. A8806) were purchased from Sigma-Aldrich. Thrombin stock solutions in MilliQ water (100 M) were stored at Ϫ80 °C. Immediately before use in an experiment, 100 nmol/ liter working solution (for a final concentration of 5 nmol/liter, 20 l/well) was prepared in prewarmed 0. 3% serum medium (50 ml of EBM-2 150 l of FBS, 500 l of 100 ϫ antibioticsantimycotics, and 500 l of 100 ϫ L-glutamine solution). 1 mol/ liter histamine stock solutions in MilliQ water were stored at Ϫ20 °C.
Before use, a 200 mol/liter working 20ϫ solution was prepared in serum-free medium (for a final concentration of 10 mol/liter and addition of 20 l/well). The solution was prewarmed to 37 °C for ϳ5 min before addition to the cells. S1P was solubilized in methanol, aliquoted, dried under nitrogen, and stored at Ϫ20 °C. For use in stimulation experiments, 20 M S1P (20 ϫ) working solution was made in HBSS (Fisher Scientific; catalog no. 14025-092) with 0. 03 mg/ml fatty acidfree BSA (from bovine plasma, Sigma-Aldrich; catalog no. A8806). Solubilization was supported by brief sonication for 5 min at 37 °C. For experiments with thrombin, HDMEC cell layers were used after 4 days in culture. The cells were changed to serumfree medium 12-24 h before start of the experiment. Cell layer quality was checked by phase contrast microscopy and by either recording a single full impedance spectrum or collecting data for 10 -30 min at multiple frequency time mode.
The coverslips with cells were mounted in a Teflon chamber and incubated with 4 M Fura-2/AM in culture medium for 35 min at 37 °C protected from light. Afterward, the cells were gently washed three times with HEPES-buffered saline (140 mM NaCl, 1. 13 mM MgCl 2 , 4. 7 mM KCl, 2 mM CaCl 2 , 10 mM D-glucose, and 10 mM HEPES, pH 7. 4) and let sit for 10 min at room temperature and protected from light. Data acquisition was performed using InCyt Im2 Fluorescence Imaging System and IntCyt Im2 software (Intracellular Imaging Inc. , Cincinnati, OH). Fura-2loaded cells were alternately illuminated with 340 and 380 nm. Fluorescence emission was collected at 510 nm. 340/380 ratio images were obtained on pixel by pixel basis. Figures showing Ca 2ϩ traces show averages from several cells per coverslip and are representative of several independent recordings of several cells as indicated in the respective figure legends.
Quantitative densitometry analysis was performed using ImageJ software. Statistical Analysis-Experimental data are presented as means Ϯ S. D. Only for data sets with very high N numbers were means Ϯ S. E. used, and the values are indicated as such in the figure legends. To test for statistical significance, data sets were subjected to ANOVA analysis. Levels of significance were indicated with asterisks as follows: *, p Ͻ 0. 05; **, p Ͻ 0. 01; ***, p Ͻ 0. 005. A p value of Ͻ0. 05 was considered statistically significant. ANOVA analysis and Pearson's correlation calculations were performed with OriginLab Pro software. Density plots were generated with R software using Kernel density smoothing for histograms. All data were plotted using Origin Pro software.
annulatus Nees, 1834) were both frequently mistreated under E. annulatus  [29, 34]. In the present study, we first provide a reliable molecular phylogeny of the "E. urozonus species group" using a multi-locus approach. Then, for most of the species, we compile host records and data on ovipositor length. We finally carry out a comparative analysis to evaluate the role of phylogenetic constraints in the evolution of ovipositor length and host range as well as the role of the ovipositor's length in determining the host range.
Concatenated datasets analysis Phylogenetic analyses were performed on concatenated nucleotide sequences using both ML and Bayesian methods. Four partitioning schemes were compared: (i) two partitions: one for the two mitochondrial genes (COI and Cytb) and another for all nuclear markers (Wg, EF-1α, Bub3, RpS4 & RpL27a); (ii) six partitions: one for the two mitochondrial markers (COI and Cytb) and one for each nuclear marker (Wg, EF-1α, Bub3, RpS4 and RpL27a); (iii) seven partitions: one for the 1st and 2nd codon positions of the mtDNA, one for the 3rd codon positions of mtDNA, and one for each nuclear gene (Wg, EF-1α, Bub, RpS4 and RpL27a); (iv) nine partitions: same as above with Wg and EF-1α further partitioned by codon position (1st and 2nd codon positions versus 3rd positions). Bayes factors (BF)  [61, 62]  were used to compare the four partitioning schemes.
Evolution models and phylogenetic reconstruction For the separated and concatenated datasets, the bestfitting model was identified using the Akaike information criterion (AIC) as implemented in jModelTest v0. 1. 1  [64]. For both gene-by-gene and concatenated analyses, maximum likelihood analyses and associated bootstrapping were performed using RAxML v8. 0. 9  [65]. The GTRCAT approximation of models was used for ML bootstrapping (1000 replicates). Bootstrap percentages (BP) ≥85 % were considered as strong support and BP < 65 % as weak. Bayesian analyses were performed only on the concatenated dataset using a parallel version of MrBayes v3. 2. 2  [66]. Model parameters for each data partition were independently estimated by unlinking parameters across partitions. Parameter values for the model were initiated with default uniform priors, and branch lengths were approximated using default exponential priors.
Bayesian inferences were estimated using two simultaneous, independent runs of Markov Chain Monte Carlo (MCMC), including three heated and one cold chains. The Metropolis-coupled MCMC algorithm  [67]  was used to improve the mixing of Markov chains. Analyses were run for 20 × 10 6 generations with parameter values sampled every 2000 generations. To ensure convergence, 40 × 10 6 generations were used for the most complex partitioning scheme (9 partitions) with parameter values sampled every 4000 generations. To increase and improve the swap frequencies of states between cold and heated chains, the heating temperature (T) was set to 0. 01 for the most complex partitioning scheme cleaned with Gblocks and to 0. 02 for all other datasets. Convergence was assessed using the standard deviation of split frequencies given by MrBayes and the Effective Sample Size (ESS), as estimated using Tracer v1.
Comparative analysis Evolution of ovipositor length The ovipositor of Hymenoptera is a complex organ that exhibits great interspecific variation (see for instance  [23] ). In species of Eupelmus, part of the ovipositor is easily visible at the extremity of the abdomen (the ovipositor sheaths), while the rest is concealed in the abdomen. The use of this visible part as a "proxy" of the total ovipositor length is a priori tempting in order to avoid damaging of specimens of newly described species known from very few individuals  [32, 33]. In order to validate the use of this proxy, a total of 34 individuals of comparatively common species (e. g. E. azureus, E. confusus, E. gemellus, E. kiefferi, E. pistaciae, and E. urozonus) were dissected and, for each individual, we measured the length of the ovipositor stylet, the visible part of the ovipositor sheath and the metatibia (see dataset on Dryad: doi:10.
5061/dryad. 115m1). Measurements of the length of the ovipositor sheaths and hind tibia followed Al khatib et al. [32]  (Additional file 2: Figure  S18 A  and C ). The length of the ovipositor stylet (first and second valvulae) was measured from the articulation of the second valvula with the articulating bulb to the apex of the second valvula (Additional file 2: Figure  S18 B ). Using this dataset, we found evidence of linear relationships between the ovipositor sheath (response variable) and either the ovipositor stylet or the metatibia as predictors (data not shown). Moreover, no interaction was found between these two predictors and the host species (respectively F 5df,20df = 1. 23 with p = 0. 34 and F 5df,22df = 1. 20 with p = 0. 34). This suggests that the visible part of the ovipositor sheath can indeed be used as a reliable proxy of the entire ovipositor.
As a consequence, a first analysis was performed on the 19 species of the "E. urozonus species group" for which information about the ovipositor sheaths and the metatibia were available. This analysis includes a total of 121 individuals, with at least 2 individuals/species except for E. priotoni and E. simizonus (only one individual in each case). In most of the cases, we tried to select individuals from at least two geographical locations and/or, for generalist species, two host species (see dataset on Dryad: doi:10. 5061/dryad. 115m1). Both the absolute length of the ovipositor sheath ("AOS") and the ratio ("ROS") between the ovipositor sheaths and the metatibia were taken into account, the second one being potentially less sensitive to environmental-induced phenotypic plasticity (host and/or abiotic conditions).
Influence of phylogeny and ovipositor length on host range A second analysis was restricted to a subset of 13 species for which host range was also available. Most of the information about host range was obtained from Al khatib et al. [32]  and from Gibson and Fusu (in prep). Jean Lecomte (comm. pers. ) communicated the rearing of E. confusus from curculionid larvae. Taken as a whole, our host survey is probably not exhaustive but nevertheless encompassed a total of several thousands of individuals of the "E. urozonus species group" and, with regard to the host's diversity, 95 insect species representing 22 families and 6 orders (see dataset on Dryad: doi:10. 5061/ dryad. 115m1). Taken as a whole, these host insects were distributed on 18 plant families. Dissimilarities in host range were calculated-at three taxonomic levels (species, family and order) for the host insect and at one level (family) for the host plant-using the Bray-Curtis distance, each host taxon being treated qualitatively (at least one record versus none).
Software and packages Manipulations of files and statistical tests were conducted using the software R (http://www. R-project. org -version 3. 0. 3 -2014-03-06) with the following packages "ade4" (Euclidian transformation of matrices)  [73] , "ape" (phylogeny)  [74] , "caper" (comparative analysis), "ecodist" (Mantel tests)  [75]  and "vegan" (similarities between host ranges)  [76].
Alignments and single-marker analyses Successful amplification and sequencing was completed for all gene regions used in this study. However, sequencing failures occurred for some markers for a few individuals. Genbank accessions of the sequences obtained for all analysed genes are given in Additional file 1: Table  S2. The final matrix contained 91 specimens. No stop codons, frame shifts, insertions or deletions were observed in coding gene regions. The numbers of aligned base pairs, variable sites and parsimony-informative sites for each gene are summarized in Table  2. As expected, mitochondrial genes showed more parsimony-informative sites compared to nuclear markers (472 out of 1085 bp). Among the nuclear markers, EF-1α exhibited the lowest number of variable and parsimony-informative sites (respectively 116 and 106 out of 517 bp).
Evolution models and partitions in the concatenated dataset Alignment lengths of the concatenated datasets with or without the exclusion of highly divergent blocks were 3197 bp and 5000 bp respectively. For all partitions, the best-fitting substitution model was the general time reversible model (GTR) with among-sites rate variation (ASRV) modelled by a discrete gamma distribution (Γ)  [77]  for which we used four categories. For all Bayesian analyses, after discarding 25 % of the samples as burn-in, the ESS value of each parameter largely exceeded 200, which indicated that convergence of runs was reached. Sixteen combined trees were obtained (Additional file 2: Figures  S1 -S8 ). For all combined datasets, Bayes factors showed that the most complex partitioning scenario (9 partitions) was preferred over the three less complex ones (Table  3 ).
Evolutionary properties of the markers Model parameter estimates for each partition of the Bayesian analysis of the "9 partitions without Gblocks cleaning dataset" are depicted in Table  4. As expected, the mitochondrial partitions showed high base compositional bias (71. 4 and 89. 8 % of A/T for the first two positions and the third codon position respectively). Among the nuclear gene partitions, RpL27a, Bub3 and RpS4 were A/T-biased (77. 9, 70 and 68. 8 %) while (See figure on previous page. ) Fig. 1  Phylogram of relationships among species of the "Eupelmus urozonus species group" obtained from the concatenated dataset alignment (5000 bp and 9 partitions) without the Gblocks cleaning of divergent blocks. Uppercase letters refer to clades discussed in the text. Nodes with likelihood bootstrap (BP) values <65 have been collapsed.
Phylogenetic trees inferred from concatenated datasets Impacts of alignment strategy and reconstruction methods ML and Bayesian topologies obtained from the concatenated alignments without Gblocks cleaning were more resolved than those obtained with removal of poorly aligned blocks. Whatever the partitioning scheme and regardless of whether or not divergent blocks were included in the analyses, most internal nodes were nevertheless statistically supported (BP value ≥ 65, PP value ≥ 90). Moreover, the 18 species recently defined by Al khatib et al. [32]  and E. vindex were recovered as a monophyletic group. Overall, topologies showed three major clades (A, B, C) that emerge on highly supported basal nodes (Figs. 1  and 2  and Additional file 2: Figures  S1-S8 ). Three topological conflicts were observed depending on whether or not the Gblocks cleaning step was performed: (i) Clade A was not supported in topologies inferred from the datasets cleaned using Gblocks (Fig.
) Fig. 2  Phylogram of relationships among species of the "Eupelmus urozonus species group" obtained from the concatenated dataset alignment (3197 bp and 9 partitions) with Gblocks-default parameters. Uppercase letters refer to clades discussed in the text. Nodes with likelihood bootstrap (BP) values <65 have been collapsed. BP (≥65) and Bayesian posterior probabilities (≥0. 90) are indicated at nodes. Each line represents a sequenced individual with information in the following order: molecular code, species, and country AOS: r = 0. 09, p = 0. 39 -Mantel test for ROS: r = 0. 08, p = 0. 44). When ovipositor length was treated as a binary variable with "long" ovipositors being those above the third quartile (4 or 5 cases among the 19 species), the observed D-statistics for AOS (0. 13) and ROS (1.
urozonus species group". Although the phylogenetic resolution was proven to be sensitive to inclusion or exclusion of divergent blocks by using Gblocks procedure from the sequence alignments, we obtained a reliable phylogeny which strongly supported the monophyly of our focus group of Eupelmus, including the 18 species treated in Al khatib et al. [32]  and E. vindex, which is morphologically distinct from other members of the group in the shape of the syntergum and the anterior displacement of the ovipositor sheaths  (Gibson & Fusu, in prep). Additionally, the included species of the "E. urozonus species group" were distributed in three strongly supported clades, referred here as A, B and C (Fig. 1 ). The molecular monophyly of the Palaearctic "E. urozonus species group" reflected in our concatenated datasets can be also supported through morphology.
Acknowledgements This research was directly supported by the projects  "EUPELMUS" (Scientific Department "Santé des Plantes et Environnement " of  INRA -grant  2012-04-05-20 ), " INULA" (APR Pesticides 2011: "Changer les pratiques agricoles pour préserver les services écosystémiques ") and ' BIOFIS ' (number  1001-001 ) allocated by the  French Agropolis Fondation (Montpellier). This project was supported by the  network Bibliothèque du Vivant  funded by the  CNRS , the  Muséum National d'Histoire Naturelle (MNHN)  and the  Institut National de la Recherche Agronomique (INRA) , and technically supported by the Genoscope. This study also took benefits from field work conducted in the frame of the projects " BIOINV-4I " ( ANR-06-BDIV-008  -Coordinator: Thomas Guillemaud), " Lutte biologique contre le Cynips du Châtaignier " (ONEMA 2011 -Coordinator:  Nicolas Borowiec & Jean-Claude Malausa ), and the  Swedish Malaise Trap Project (SMTP).
The depository's acronyms of voucher specimens are included in (Additional file 1: Table S2; Additional file Dryad: doi:10. 5061/dryad. 115m1). Sampling information (host-plants, collection dates, and localities) is listed in Table -Thirteen species were used as outgroup including (i) species belonging to the three subgenera of Eupelmus sensu Gibson (1995): Eupelmus [E. : E. acinellus Askew, 2009, E. annulatus, E. azureus, E. cerris Förster, 1860, E. confusus, E. fulvipes Förster, 1860, E. gemellus, E. janstai Delvare and Gibson, 2015, E. kiefferi, E. longicalvus Al khatib & Fusu, 2015, E. minozonus Delvare, 2015, E. opacus Delvare, 2015, E. pistaciae Al khatib, 2015, E. priotoni Delvare, 2015, E. purpuricollis Fusu & Al khatib, 2015, E. simizonus Al khatib, 2015, E. tibicinis Bouček, 1963 and E.
169 (0. 141-0. 197) 0. 349 (0. 313-0. 385) 0. 222 (0. 166-0. 279) 0. 190 (0. 152-0. 229) RpS4 0. 332 (0. 308-0. 354) 0. 162 (0. 146-0. 180) 0. 147 (0. 131-0. 163) 0. 357 (0. 334-0. 380) 0. 427 (0. 364-0. 496) 0. 262 (0. 224-0. 303) RpL27a 0. 390 (0. 367-0. 412) 0. 109 (0. 096-0. 123) 0. 111 (0. 097-0. 124) 0. 389 (0. 366-0. 410) 0. 820 (0. 693-0. 946) 0. 536 (0. 455-0. 619) Mean and 95 % credibility intervals of the model parameters for each partition included in the Bayesian analyses of concatenated datasets without Gblocks cleaning (9 partitions) are reported.
Results The genomes of 75 isolates of T. b. gambiense Group 1 (Supplementary file 1) were sequenced, including multiple samples from geographically separated disease foci within Guinea (n=37), Cote d'Ivoire (n=36) and Cameroon (n=2) collected over fifty years. For comparative purposes, isolates of T. b. rhodesiense (n=4), T. b. gambiense Group 2 (n=4) and T. b. brucei (n=2) were also sequenced. A total of 230,891 single nucleotide polymorphisms (SNPs) were identified compared to the haploid consensus assembly of the T. b. brucei reference genome  (Berriman et al. , 2005). These were evenly distributed over the eleven major chromosomes, covering 85% of the genome (Figure  1 -figure supplement 1). T. b. gambiense Group 1 showed a 5-10 fold lower number of  SNPs (11, 398)  and SNP density compared to the other groups (Figure  1 -source data 1), despite an over-representation in terms of the number of samples.
To test whether haplotypes have evolved independently at the whole-chromosomal level, the sequence dataset for all sub-species was computationally phased and haplotypes inferred for each isolate. Excluding LOH regions, phylogenetic trees for each chromosome were generated, revealing that A and B haplotypes separate into distinctive clusters (example in Figure  4A ) and for every chromosome this pattern was maintained (Figure  4 -figure supplement 1). Co-phylogenetic analysis revealed congruence between the A and B haplotype trees across different chromosomes (Figure  4B ; Figure  4 -figure supplements 2 and 3) illustrating that these haplotypes are evolving in parallel. The time of emergence of T. b. gambiense Group 1 was determined. We estimated the genomewide mutation rate using the number of accumulated mutations (both genome-wide and on Chromosome 9) together with the year of isolation for each isolate, using root-to-tip linear regression  (Drummond et al.
Illumina sequencing and SNP calling The T. brucei genome is approximately 26 Mb in size and comprises eleven major chromosomes between one and six megabases together with an array of intermediate and mini-chromosomes  (Ogbadoyi et al. , 2000). Illumina paired-end sequencing libraries were prepared from genomic DNA and sequenced by standard procedures on Illumina HiSeq machines, to yield paired sequence reads of 75 bases in length. For each parasite strain, the data yield from the sequencing machines passing the default purity filter was between 7. 4 million and 40. 4 million read pairs (median of 15. 3 million), which corresponds to a nominal genome coverage of between 37. 9-fold and 207. 1-fold (median of 78. 2-fold). For the purpose of calling SNPs, mapping of the paired sequencing reads to the genome reference sequence from GeneDB (Trypanosoma brucei brucei TREU927, referred to here as Tb927) was carried out with SMALT (www.
sanger. ac. uk/resources/software/smalt/) version 0. 7. 4 using the following parameters: word length = 13, skip step = 3, maximum insert size = 800, minimum Smith-Waterman score = 60, and with the exhaustive search option enabled. A median fraction of 79. 8% of sequencing reads were mapped and a median fraction of 64. 9% of sequencing reads were classified as 'proper pairs', i. e. with the two mates of a sequencing read pair mapped within the expected distance and in the correct orientation. Only sequence reads mapped as 'proper pairs' were used for SNP calling, and the first 5 and last 15 nucleotides were clipped from all reads prior to subsequent analysis. Genotypes for every genomic position were determined using SAMtools version 0. 1. 19 (Li et al. , 2009 ) by using the 'samtoolsmpileup' command with minimum baseQ/BAQ ratio of 15 (-Q) followed by SAMtools' 'bcftools view' command with options -c and -g enabled.