dataesr/scientific-paragraphs-categorization huggingface dataset:
Genetic analysis Genomic DNA was extracted from whole blood using the DNeasy Tissue and Blood kit or from formalin-fixed paraffin-embedded tissue using the GeneRead DNA formalin-fixed paraffin-embedded kit (QIA GEN). WES was performed for P1 and P2 on a HiSeq2000 platform (Illumina), and the process of library preparation, read mapping, and variant analysis has been previously described  (Abolhassani et al. , 2014; Alkhairy et al. , 2015b). For family 2, WGS were performed for P4 (60× mean coverage) and both parents (30× mean coverage) using 300 ng genomic DNA (Broad Institute). Single-nucleotide variant and insertion/ deletion calling were performed using the Genome Analysis Toolkit (3. 4). Existing databases including 1000 Genome, Exome Aggregation Consortium, National Heart, Lung, and Blood Institute exome variant server, and Greater Middle East Variome were used for filtering based on the population frequency for any given variant for both families.
CD70 has a gene damage index of 1. 78 and a mutation significance cut-off of 6. 208  (Itan et al. , 2015 (Itan et al. , , 2016)). The combined annotation-dependent depletion scores  (Kircher et al. , 2014)  for the CD70 variants identified in the two families are 23. 7 and 8. 053 (both above the gene-specific mutation significance cut-off) and are thus predicted to be deleterious. WES and WGS data have been deposited in the Database of Genotypes and Phenotypes under accession no. phs001245. v1. p1. Sanger sequencing was conducted to confirm the presence of the CD70 mutation in the affected siblings and for carrier detection in family 1. Exon 3 of the CD70 gene was PCR amplified (forward, 5′-CCT CAG TTT CCC TAA ACC TCCA-3′ and reverse, 5′-AAG CTC AAT GCC TTC TCT TGTC-3′), and the resulting PCR products were sequenced at Macrogen Inc.
Sequence analysis was performed using the Lasergene software package (DNAStar). The same method was applied to investigate the mutation prevalence in this exon in 251 Iranian blood donors. A nested PCR approach was used to amplify EBNA1 from DNA from tumor biopsy  (Cinque et al. , 1993; Chan et al. , 2001). For variants from family 2, genomic DNA from both affected siblings and parents was PCR amplified (forward primer, 5′-GGC CCC TGT GTG TAC ACT TT-3′ and reverse primer, 5′-TCT CAG CTT CCA CCA AGG TT-3′). Sanger sequencing of purified PCR products was performed by the Genomics Unit of the Rocky Mountain Laboratories Research Technologies Section of the National Institute of Allergy and Infectious Diseases. rNA isolation and quantitative PCr PBMCs were isolated from whole blood by a standard method (Hypaque-Ficoll; GE Healthcare).
Cell proliferation assay PBMCs were thawed and resuspended in sterile PBS/0. 1% BSA containing CFSE (eBioscience) at a final concentration of 2. 5 µM. Cells were incubated at 37°C for 10 min and then washed with five volumes of ice-cold sterile PBS/0. 1% BSA and subsequently plated at ∼1. 8 × 10 5 per well in 96well U-bottom plates containing either RPMI medium/10% FCS alone or 5 µg/ml PHA (Sigma-Aldrich) plus 50 U/ml IL-2 (PeproTech), 20 ng/ml PMA plus ionomycin (Sigma-Aldrich), or T cell activation and expansion beads (one bead to two cells; Miltenyi Biotec) for 4-5 d. Cells were harvested, stained with Zombie aqua fixable viability dye, anti-CD3 Pacific blue, anti-CD8 APC Cy7 (BioLegend), and anti-CD4 BUV395 (BD), fixed, and run on a cell analyzer (LSR II SORP; BD). Data were analyzed using FlowJo software (Tree Star).
HSP90 was used to assess protein loading. (F) Binding of overexpressed WT or mutant CD70 to recombinant human CD27 was evaluated by flow cytometry. Shaded areas represent nontransfected 293T cells. Binding of anti-CD5 mAbs to cells transfected with WT (turquoise) or mutant (yellow) CD70 is shown. Binding of biotinylated CD27/streptavidin-PE in cells transfected with WT (blue) or mutant (red) CD70 is shown. (G) Flow cytometric detection of the flag epitope on cells transfected with WT (blue) or mutant (red) CD70 or on untransfected cells (shaded). (H) Immunoblotting of lysates from untransfected cells or cells transfected with WT or mutant CD70 plasmid, using Abs to CD70 (Ab2), myc tag, or HSP90. All data are representative of at least two independent experiments. Mut, mutant; SSC, side scatter.
ACKNOwlEDGMENTs We would like to thank  M. Bartish  for generating the CD70-expressing constructs and for help with Western blotting,  A. Zaravinos  for help with real-time PCR,  H. Ham  and  K. Payne  for technical assistance,  J. Cohen  for reagents,  A. Sönnerborg  for assisting with the interpretation of serology tests,  J. King  for help with editing of the manuscript, and the patients and their families for participating in this study. This study was supported by the  Swedish Research Council  (grant to  L. Hammarström  and  Q. Pan-Hammarström ), the  Swedish Cancer Society  (grant to  Q. Pan-Hammarström ), the  Swedish Childhood Cancer Foundation  (grant to  Q. Pan-Hammarström ), the  Susan and John Freeman Cancer Research Grant  from  Cancer Council NSW (Australia ; grant  RG16-11 ), and the  Intramural Research Program of National Institute of Allergy and Infectious Diseases, National Institutes of Health.
The  laboratory of Human Genetics of Infectious Diseases  is supported by grants from  Institut National de la Santé et de la Recherche Médicale, University Paris Descartes , the  French National Research Agency  under the  Investments for the Future program  (grant n°ANR- 10-IAHU-01 ), the  Rockefeller University , and the  St. Giles Foundation. D. A. Price  is a  Wellcome Trust Senior Investigator. S. G. Tangye  is a recipient of a  Principal Research Fellowship  from the  National Health and Medical Research Council of Australia  (grant  1042925 ) and a  Fulbright Senior Scholarship. The authors declare no competing financial interests. Author contributions:  H. Abolhassani  collected the clinical data and wrote the paper. E. S. J. Edwards ,  H. Jing ,  M. Buggert ,  L. Du ,  M. Matsuda-Lennikov ,  R.
01. All work with infectious RVFV and rNiV was carried out under biosafety level 4 (BSL-4) conditions at the Robert E. Shope BSL-4 laboratory at the University of Texas Medical Branch. Analysis of viral replication. VSV-infected supernatants were removed 24 h. p. i. and titered by determining the 50% tissue culture infectious dose on Vero-E6 cells. RVFV and rNiV-gLuc-P2A-eGFP-infected supernatants were collected at 48 and 24 h. p. i. , respectively, and titered by plaque assays on Vero-CCL81 cells. Fibroblasts infected with rVSV-GFP or rHSV-1-US11-GFP were fixed in 4% PFA 24 h. p. i. and stained with 5 mg ml À 1 Hoechst 33342 in PBS. Cells were imaged and the results were analysed (BioTek Cytation 3; Gen5 software). HCMV-IE2-YFP-infected cells were fixed with 4% PFA 24 h. p. i. , and imaging analysis was performed on an Acumen Explorer plate cytometer.
Acknowledgements This was supported in part by  NIH  grant  R00 AI106942-02  to D. B. ,  NIH  grant  R01 AI101820  to  D. T. , an  American Heart Association pre-doctoral fellowship  and a  USPHS   Institutional Research Training Award   T32-AI07647  to T. J. G. , NRSA  T32 AR07279-30  to M. R. R. ,  NIH  grant  RO1 A1080672  and  Pew Scholar Award  to D. J. L. , funding by the  DFG  ( SFB 944 ) to J. P. ,  NIH  grant  R33 AI102267  to A. N. F. and B. L. ,  CRIP (Center for Research on Influenza Pathogenesis) , and  NIAID funded Center of Excellence for Influenza Research and Surveillance  (contract # HHSN272201400008C ) to AGS. Experimental support was provided by the  Speed Congenics Facility of the Rheumatic Disease Core Center  ( P30 AR048335 ). Work in the Cytokine Signaling Unit was supported by  Institut Pasteur ,  CNRS ,  INSERM  and an  Amgen Scholarship  to E.
from the PasteurParis University International PhD program and supported bytheInstitutCarnot PasteurMaladiesInfectieuses. NH and MJPwere, respectively, supported byfellowships from theVaccine ResearchInstitute(VRI) and Sidaction. Wethank the French AgencyforAIDSResearch(ANRS), theInstitut Pasteur, Sidaction, the«Investissements d"Avenir"programmanaged bythe ANRgrant agreement N° ANR-10-LABX-77 and ANR-11-INBS-0008,as wellasL"Oréal in partnership with UNESCO fortheirsupport. The fundingsources had no influenceon our studies orin the writingof this report.
2 million people living with HIV in sub-Saharan Africa has been one of the most successful public health interventions ever undertaken  [14] , improving life expectancy  [15] , and reducing transmission  [16, 17]. As a result, ART has been one of the most potent selection pressures on HIV. Given its importance to global health, resistance to ART has been well studied in HIV with many amino acid changes known to lead to DR  [18]. Thus, DR is a useful phenotype for validating GWAS in HIV as findings can be compared to the existing literature as well as to large publically available databases of genes involved in HIV DR. In this study, we aim to identify known variants and validate the applicability of GWAS methods to the HIV genome.
Identification of novel variants As well as known drug resistance variants, additional associations were observed. The first was two associations within RT amino acid 68. The first was between tenofovir failure and SNP 2738G resulting in a change from serine to glycine. Replication was performed using the Stanford University HIV Drug Resistance Database  [19, 20]. Subtype C sequences within the Stanford database from individuals failing to tenofovir or other NRTIs (n = 9,357) all had the reference (serine) amino acid. For sequences showing resistance to tenofovir, however, 5. 5% had glycine at this position (n = 488, p<0. 0001 compared to non exposed distribution). Stavudine resistance was showed associations with a different SNP (2739A) from serine to asparagine (Fig C in S1 File). However further investigation showed this to be an association with the negatively correlated drug tenofovir which had a p-value just below genome-wide significance for this SNP (p = 7.
Whilst not available in the Stanford database, we compared our results to the Los Alamos HIV Sequence Database drug naïve WGS at the amino acid 91 of the matrix region. Interestingly, for the amino acid 91 we observed a high level of genetic variation, with coding for nine amino acids. Focusing on the associated genetic change, we observed significantly different (p<0. 0001) frequencies in the drug naïve sample (37% G vs. 61% A) compared to the tenofovir-exposed sequences in our sample (65% G vs. 35% A). Our WGS tenofovir naïve cases had a same frequency as the publically available sequences (37% G). While not an independent replication, this lends some support to our finding.
Sample description The study sampled 319 HIV-infected adults and 24 children on ART with virological failure in the Hlabisa HIV Treatment and Care Programme in South Africa for which a whole genome of HIV-1 was produced. The inclusion criteria were: ART regimen for at least 12 months followed by virological failure, defined as one viral load >1000 copies/ml. Exclusion criteria were: prior use of nucleoside reverse transcriptase inhibitor (NRTI) monotherapy or dual therapy (not including regimens for the prevention of mother-to-child transmission (pMTCT)). All individuals were seen by a physician, who performed a clinical evaluation and obtained written informed consent for the study. A 5 ml EDTA whole blood sample for HIV DR genotyping was collected during the clinical evaluation. Basic clinical and demographic data, including GPS data on household location, were collected on a clinical form and clinical and treatment information was compared with the records in the Africa Centre's ART Evaluation and Monitoring System (ARTemis), an operational database holding treatment and laboratory monitoring information from the national ART programme in South Africa.
Bioinformatics pipeline Fastq quality control was performed using FASTQC(0. 11. 3) and QUASR(3. 1) software applications. Reads of less than 100bp in length and a quality score lower than 30 were excluded. In addition, the reads were trimmed up to 10bp from 5' and 30bp at the 3' to exclude poor quality sequence at the beginning and end of reads. We noticed that the second pair read of the Illumina Nextera XT was of lower quality and that excluding the last 30bp increased quality score to > 33. We imposed these exclusion criteria in order to decrease the probability of ambiguous read mapping, which occurs when shorter reads of lower accuracy are included in assemblies  [30]. Following these quality control steps, we mapped reads against a subtype C reference sequence (AF411967) with five assembly iterations using Geneious 8 (http://www.
geneious. com) [31]. After assembly, we exported the data as BAM files and exported contigs as FASTA files. In order to determine if there was clustering of sequences (i. e. sequences that were very similar with low genetic diversity), we aligned all of the whole genomes with a reference dataset for HIV-1 subtype C. The tree was constructed with HKY+Gamma site rate variation in a MPI version of RaxML. Reliability of internal nodes was evaluated by 100 bootstrap replicates. Phylogenies were analysed using Phylotype software application  [32]  in order to detect any clustering of sequences with high bootstrap values (>90%) and low sequence diversity (<3%). This was performed to identify pairs of closely related HIV sequences that might confound the analysis and test the sensitivity of the results to their inclusion.
Variant calling and GWAS software adaptation The processing of WGS data to the performing of GWAS is outlined in Fig  1 , with comparison to human GWAS steps. BAM files were converted to VCF format variant calls individually for each sequence in PILON  [33]. A threshold of a depth of 50 reads per base was used for a variant to be called. As GWAS software was originally designed for diploid organisms (i. e. those with two chromosomes and so two copies of any given loci), each sample can be called either as homozygous for an allele (e. g. AA or TT) or as heterozygous (e. g. AT). While heterozygosity is incorrect in the sense that HIV is haploid, it captures an important reality of viral infection: genetic differences within the host's viral population. We wanted to retain the feature of diploidy to account for samples with diversity at a given DR loci.
Replication Genome-wide significant SNPs within the Pol region were able to be taken forward for replication in a publically available independent sample. This was the Stanford University HIV Drug Resistance Database  [19, 20] , where information on amino acid frequencies were available for sequences exposed to different drugs. This analysis was restricted to the 13,676 subtype C sequences. Additional analyses also made use of a subset of all publically available subtype C WGS (n = 505) from the Los Alamos HIV Sequence Database (http://www. hiv. lanl. gov/). This was done to ensure our variant frequencies were in agreement with those observed elsewhere.
The Study As part of our mission as a World Health Organization collaborating center, we performed MLST on N. meningitidis serogroup X isolates sent to the Institut Pasteur during 1998-2005 (Table  1 ). We further performed whole-genome sequencing on 8 invasive serogroup X isolates obtained during 1998-2005 from several countries within the meningitis belt and 3 invasive isolates from France (Table  1 ). Genomic DNA was extracted by using a Genomictip 20/G kit (QIAGEN, Valencia CA, USA) from culture grown overnight on gonococcal medium base agar plates with Kellogg supplements  (9). Whole-genome sequencing was performed by using an Illumina HiSeq 2000 sequencer (Illumina, San Diego, CA, USA) and assembled as described  (10). Sequences are available through the PubMLST database (http://pubmlst.
org/neisseria/), which runs on the Bacterial Isolate Genome Sequence Database (BIGSdb) platform  (11) ; identification numbers are 34731-34732, 34734-34741, and 34745. We analyzed the 11 isolates from sub-Saharan Africa and France and all other serogroup X genomes available in BIGSdb by using the BIGSdb genome comparator tool. The isolates from Africa were genetically related, belonged to CC181, and were separated from all the other isolates that belonged to other CCs (Table  1 ; Figure  1 , panel A). The CC181 isolates formed a single main lineage comprising 2 sublineages, 181. 1 and 182. 2. Sublineage 181. 1 was detected only in isolates obtained in 2006, but sublineage 181. 2 was detected in isolates from both periods (1990s and since 2006). Genomes from the 3 isolates from France and those available in the PubMLST database were highly diverse with no close clustering (Figure  1, panel A ).
This work was supported by the Institut Pasteur and by funding from the Fondation Total including a fellowship to A. A. This study made use of the Neisseria Multilocus Sequence Typing website (http://pubmlst. org/neisseria/), developed by Keith Jolley and sited at the University of Oxford, and the dynamic imaging facilities of Imagopole at the Institut Pasteur, Paris. The development of this site has been funded by the Wellcome Trust and European Union. This study also made use of the Meningitis Research Foundation Meningococcus Genome Library (http://www. meningitis. org/research/genome) developed by Public Health England, the Wellcome Trust Sanger Institute, and the University of Oxford. Dr. Agnememel is a microbiologist at the Institut Pasteur, Paris, and works on development of bacterial meningitis rapid diagnostic tools.
HIV-specific cells were stained with human leucocyte antigen (HLA) B*57, B*27 and B*08 restricted Gag-and Nef-derived tetramers containing the following peptides: KAFSPEVIPMF (Gag162-72), KRWIILGLNK (Gag 131-140), FLKEKGGL (Nef 90-97) and EIYKRWII (Gag 260-267). For each marker of interest, fluorescence-minus-one controls (FMO) were prepared. At least 1 million events per sample were acquired on an LSR II flow cytometer (BD Biosciences). Raw data were analysed with fluorescence activated cell sorter (FACS) Diva version 5 (BD Biosciences, Heidelberg, Germany) and FlowJo software (TreeStar Inc. , Ashland, OR, USA).
Statistical analysis All groups were tested for normal distribution with the Kolmogorov-Smirnov test and were compared by the adequate test. For normally distributed data, parametric tests were applied: for two paired groups, paired t-tests; for more than two groups, analysis of variance (ANOVA) followed by Tukey's multiple comparisons test. Data that were not distributed normally were tested by the Wilcoxon test for paired groups or the Kruskal-Wallis test followed by Dunn's multiple comparisons test for more than two groups, respectively. All statistical analysis was carried out using Graphpad Prism version 5 software. A P-value of < 0Á05 was considered significant. The following symbols were applied to denote statistically significant findings: ***P < 0Á001, **P < 0Á01 and *P < 0Á05.
5x10 6 (Fig  2). The live parasite level dramatically decreased by 23-fold in mice that were immunized with PL together with WT CNF1 compared with controls. To evaluate the effects of CNF1 activity, we also quantified the parasite levels in a group of mice immunized with PL + mCNF1. The results revealed that the WT CNF1 catalytic activity produced a marked 6-fold increase of protection compared with mCNF1. Together, these experiments suggest that mice immunized against PL together with active WT CNF1 develop a strong resistance to infection. In parallel, we assessed the IgG-titer against PL in infected mice (Fig  3 ). The results showed a 3-fold increase in the IgG level in PL + WT CNF1 immunized mice compared with naïve and PL + mCNF1 conditions (Fig  3 ). Our data are in good agreement with our findings that active CNF1 together with promastigote lysate conferred a high resistance to infection in vaccinated mice.
WT CNF1 shows curative activity against L. infantum The above data revealed a previously unknown property of WT CNF1 in stimulating a cytokine response, thus demonstrating its strong capacity to stimulate pro T-helper Th1 immune responses. This prompted us to assess whether the CNF1 activity might also be endowed with adjuvant curative properties. Mice were first infected and later treated with or without PL in the presence or absence of either WT CNF1 or the catalytically inactive mutant, mCNF1, as a control. Therapeutic vaccination was repeated twice at one-week intervals, and the infection was monitored at day 42 post-immunization. Fig  5  depicts the parasite burden in the spleen. The infected mice treated with WT CNF1 had a reduced parasite burden compared with the mCNF1 and control mice.
Acknowledgments We are grateful to  Dr Mery Tulic  and  Carmelo Luci  for critical reading of the manuscript. This work was supported by the  Groupe d'Action Contre la Leishmaniose  (GACL), by institutional funding from  INSERM , grants from the  Fondation Infectiopôle Sud , the  Agence Nationale de la Recherche  ( ANR 11BSV3 004 01 ), the " Investments for the Future" LABEX SIGNALIFE   ANR-11-LABX-0028-01  and the C3M animal facility. NM was supported by  DGA  and the joint ministerial program of R&D against CBRNE threats. SIMOPRO is a member of the laboratory of Excellence LERMIT supported by the grant from the  Agence Nationale de la Recherche  ( ANR-10-LABEX-33 ). We thank  Veronique Corcelle  and the C3M animal facility team ( Yannick Michelle ,  Sandrine Verger  and  Alexandre Ipekdjian ) for their help and valuable assistance with animal care.
This work was supported by the  Groupe d'Action Contre la Leishmaniose  (GACL), by institutional funding from  INSERM , grants from the  Fondation Infectiopôle Sud , the  Agence Nationale de la Recherche  ( ANR 11BSV3 004 01 ), the " Investments for the Future" LABEX SIGNALIFE   ANR-11-LABX-0028-01  and the C3M animal facility. NM was supported by  DGA  and the joint ministerial program of R&D against CBRNE threats. SIMOPRO is a member of the laboratory of Excellence LERMIT supported by the grant from the  Agence Nationale de la Recherche  ( ANR-10-LABEX-33 ).
Wald statistics, which have a χ 2 distribution, were established. Days were considered to be frost days when the minimum daily temperature fell below 0 °C. An ice day or permanent frosts were designated when even the highest daily temperature remained below 0 °C. Lagged monthly temperature and precipitation data were also examined for any lag effects on monthly nymph density. Lag effects were considered to be important because of the potential gradual depletion of the existing nymph stock  [10]. All univariate analysis variables with a P-value ≤ 0. 25 associated with nymph density were included in the multivariate analysis. Model simplification following fitting of the full model of the multivariate analysis was based on the change in the Akaike Information Criterion (AIC). Individual parameters were sequentially removed and then replaced and the new model that generated the largest drop in AIC, following removal of a variable, was retained.
Acknowledgments We gratefully thank all who participated in collecting ticks throughout the study:  M. Chaumien ,  E. Devillers ,  F. Femenia ,  N. Galais ,  D. Huet ,  X. Liu ,  S. Moutailler ,  C. Reis  and  L. Simo. We are very grateful to  Jean-Louis Chapuis  for his critical reading of the manuscript. Thanks are also due to the " Tiques et Maladies à Tiques" group  ( REID, Réseau Ecologie des Interactions Durables ) for stimulating discussions.