dataesr/scientific-paragraphs-categorization huggingface dataset:
Leptospira genotyping Three major MLST schemes exist for Leptospira spp. typing worldwide, all supported by the website database http://pubmlst. org/leptospira/. In our study, scheme #3  [40]  was preferred to the other two  [10, 41]  as several molecular data from the SWIO have been made freely available using this same scheme, hence allowing comparison of local STs with those from SWIO islands  [12, 26]. MLST was attempted for adk, icdA, lipL32, rrs2, secY, and lipL41 loci as previously described  [40]  and recently optimized  [26]. All positive domestic animal samples were first submitted to secY sequencing, revealing a nearly clonal population among infected rats (see Results). Hence, a randomly selected subgroup of positive wild animal samples was selected from eleven sites (n = 17; Table  1 ) and submitted to full MLST genotyping.
Phylogenetic analysis Consensus sequence for each sample and multiple alignments between sequences were obtained using Geneious Pro version 5. 4  [45]. In order to provide a comparison with Leptospira strains occurring worldwide, our dataset included sequences from two human clinical isolates from Mayotte  [12]  and 118 human and animal isolates from all over the world  [2]. Sequence alignments were constructed separately for all six considered loci. A five-loci concatenate was generated using SEQMATRIX v1. 7. 8  [46] , excluding icdA locus that was unavailable from the Mayotte clinical dataset and for some of our samples. Bayesian analyses were performed to infer phylogenetic relationships between Leptospira species. The best-fitting model and associated parameters were selected using jModelTest  [47]  and phylogenies were constructed by Bayesian inference.
We performed two independent runs of Metropolis-coupled Markov chain Monte Carlo (MCMCMC) analyses in MrBayes v3. 2. 1  [48]  of all loci independently and of concatenated sequences. Each run included 20,000,000 generations, and trees were sampled every 100 generations. The initial 20,000 trees were discarded as a conservative "burn-in" and the harmonic mean of the likelihood was calculated by combining the two independent runs. The 50% majority-rule consensus tree was then computed from the sampled trees under the best model. Neighbor-joining trees were constructed using Seaview v4. 3 (Kimura's 2-parameter distances, 500 replicates). Trees were visualized in FigTree v1. 3. 1 (http://tree. bio. ecd. ac. uk/). GenBank accession numbers of the sequences produced in the frame of the present study are provided as additional table 1 (S1 Table ).
borgpetersenii infecting the first clinical sample and the only mouse positive for L. borgpetersenii, and between L. borgpetersenii infecting the second clinical sample and one cow sample (based on a partial 434 bp amplification for the cow). The secY locus amplification and sequencing provided sequences for six cow samples, revealing alleles secY-47 (n = 4), secY-49 (n = 1), and a third not yet described allele in one sample (KU183602, see  Fig 3). The allele secY-47 is closely related to secY-48 identified in clinical information). Clinical samples from Mayotte are written in blue, using GenBank accession numbers and corresponding Leptospira species. Li: Leptospira interrogans; Lb: Leptospira borgpetersenii; Lk: Leptospira kirschneri; Lmay: Leptospira mayottensis. doi:10. 1371/journal. pntd.
0004733. g004 samples from Mayotte, whereas secY-49 has been identified in Leptospira-positive Tanzanian rodent samples and in a clinical sample from China (comparison with sequences from  [2] ). The third undescribed secY allele was identified in the cow sample showing a lipL32 sequence common to those from one clinical sample. Noteworthy, twelve secY Leptospira sequences from the insectivorous bat Mormopterus francoismoutoui endemic to Reunion Island (Gen-Bank accession numbers KJ607946 to KJ607957)  [50]  were not related to any of these alleles (Fig  3 ).
Further investigations targeting other loci with higher nucleotide polymorphism are needed to ascertain this point. A different L. borgpetersenii infecting the second clinical case suggested the existence of a second potential reservoir host. A bat species endemic to Reunion Island, Mormopterus francoismoutoui, has been demonstrated as a L. borgpetersenii carrier in urine  [50]. However, the comparison of rrs2 sequences from our two clinical samples infected with L. borgpetersenii to GenBank sequences obtained from this endemic bat species (n = 12) showed a low degree of genetic relatedness (Fig  3 ), thus excluding bats as a source of contamination leading to overt clinical leptospirosis in humans. A serological survey conducted in 2009 showed that up to 32% of cattle were seropositive for leptospirosis in Reunion Island, with Sejroe reported as the main circulating serogroup  [30].
interrogans, referring to sequence types ST02 and ST34 (http://www. pubmlst. org/leptospira/). International isolates previously published on the pubmlst. org database showed ST02 to be isolated from Belgium and Brazil, while ST34 was isolated from Jamaica and India. The second (minor) cluster, ST34, identified in six clinical samples, could not be detected in any of the five wild small mammal species targeted by our study. Additional sampling of domestic animals interestingly highlighted stray dogs as possible carriers of both L. interrogans STs found in human acute cases. Dogs have been repeatedly pinpointed as involved directly or indirectly in human epidemiology (contamination from dogs to humans, or from a common environmental source)  [62] [63] [64] [65] [66]. For instance, a serosurvey conducted on Reunion Island in 2009 identified Canicola and Icterohaemorrhagiae as the major serogroups (43.
Hence, serological data and PCR data on kidneys may be discrepant. Second, the kidney samples used in our study were collected from growing-finishing pigs under one year old, a potential age bias that might have affected our findings. Indeed, a proper assessment of pigs as reservoirs of leptospires at risk for human health requires the screening of older animals sampled from different farms as well as from family breedings throughout the entire island. It is hypothesized that Leptospira were introduced onto islands with their animal hosts, and that a variable number of introduced strains have adapted to the new local environment and available hosts  [14]. On Reunion Island, the fact that L. interrogans is almost clonal in rats is in favour of a recent leptospiral introduction on the island concomitant with the introduction of rats, back to the 16 th century.
Acknowledgments The authors thank the  Direction de de l'Aménagement et du Logement (DEAL Reunion)  for issuing permits, the  Office National des Forêts (ONF) , the  Parc National de La Reunion  and the  Réserve Naturelle Nationale de l'Etang St Paul , which gave authorization for the trapping of Tenrec ecaudatus ( DIR/I/2013/008 ) and allowed us to sample inside protected areas. We also acknowledge  Jean-Paul Dussud  from  Services Vétérinaires de La Réunion  for providing cow samples, and CIREST and intercommunal pound of  St André  for allowing the dog sampling. We thank  Amélie Desvars  for providing four cow samples collected in 2009 during her PhD and  Jean-Sébastien Dehecq  for providing Reunion Island maps. This paper is dedicated to the memory of our late colleague  Alain Michault , who disappeared the 5 th August 2015 during the preparation of this manuscript.
Imaging and image analysis All microscopic images were acquired using a Zeiss 780 Meta confocal microscope (Carl Zeiss), equipped with a Zeiss Plan-Apochromat 63× oil immersion objective lens. For each sample, several images were captured from randomly chosen regions. All recorded images were analysed with BlobFinder software (Centre for Image Analysis, Uppsala University). PLA signals (dots/cells) were counted from at least 100 cells for each sample.
Acknowledgements We thank  Peter Nielsen  and  Lise Leclercq  for critical reading of this manuscript. We thank  Hassan Jumaa  for the 3046 pro-B-cell line. We also thank  Klaus Rajewsky  and  Margaret Hibbs  for the B1-8 transgenic mice and the Lyndeficient mice, respectively. This study was supported by the  Excellence Initiative of the German Federal and State Governments  ( EXC 294 ), by  ERC  Grant  322972 , by the  Deutsche Forschungsgemeinschaft  through  TRR130  and by the  Freiburg Institute for Advanced Studies (FRIAS)  and the  University of Strasbourg Institute for Advanced Study (USIAS)  through the  Joint Fellowship Programme Strasbourg/Freiburg.
92 between both methods (D. Saulnier, data not shown). All qPCR assays were conducted on a MX3000 Thermocycler (Stratagene) using SYBR-Green. Each reaction was amplified in duplicate using a final volume of 25 lL containing: 12. 5 lL of Brillant â SYBR-Green Master Mix reagent, 2. 5 lL of both reverse and forward primers diluted at the concentration of 4 lmol/L, and 10 lL of DNA sample, all diluted (NanoDrop â ND-1000 spectrophotometer) to a concentration 1 ng/lL (host and Symbiodinium quantifications) or 5 ng/lL (Vibrio quantifications). The following protocol was performed for each sample: 1 cycle of pre-incubation at 95°C for 10 min, followed by 40 cycles of amplification at 95°C for 30 sec, followed by 60°C/64°C 1 min incubations for the Symbiodinium and polyp primers respectively, followed by a 72°C 1 min incubation, and finally a melting temperature curve analysis at 95°C for 1 min, 60°C for 30 sec, and 95°C for 30 sec.
Sequencing and symbiont identification The 28S DNA region of some samples (580 pb) was amplified by PCR with the universal Symbiodinium clade primer set  (Richter et al. 2008). Amplified fragments were then purified (Promega Wizard â SV Gel and PCR Clean-Up System) and directly sequenced (Macrogen, Korea) either with the universal primer set, when only one Symbiodinium clade was previously detected by qPCR in the colony, or with specific-clade primers sets A-D when several Symbiodinium clades were detected. Symbiodinium clades were then characterized by aligning DNA sequences using "clustalw," included in the MEGA 6. 06 software package, and referencing the GenBank library. Vibrionaceae were amplified (~90 bp) from some coral DNA samples using genus-specific primers and conditions for Vibrio spp.
Statistical analyses All S/H ratios were estimated in 28S copies per polyp for Symbiodinium CFU for Vibrio and were log+1-transformed for statistical analyses. To estimate the qualitative correlation of clades with coral health and Vibrio infection, a chi-square (v 2 ) test of homogeneity was performed, using a Monte Carlo simulation (v 2 MC ) in case of low sample size (n ≤ 5). The quantitative variations of Symbiodinium were estimated with a two-factor ANOVA on delta values of 28S copies per polyp calculated between periods t n and t nÀ1 according to the different clades and the presence (1) or absence (0) of Vibrio spp. Discriminant analysis of principal components (DAPC) on the delta values of log (S/H ratios) for each Symbiodinium clade associated with coral colonies at different sampling times was performed using R statistical software with the FactoMineR package.
Acknowledgments We thank  Nathan Spindel  for providing English language editing as well as constructive comments which improved the manuscript. We also thank  Herman Wirshing  for providing editing and constructive insights on the manuscript. We are grateful to the technicians from  CRIOBE  ( B. Espiau ,  P. Ung , and  F. Lerouvreur ) and from  IFREMER from Taravao  ( J. Fievet ,  C. Belliard , and  A. Santini ) and Plan ete Urgence volunteers for their technical help. We also thank  M. A. Coffroth  and her laboratory (BURR; http://www. nsm. buffalo. edu/Bio/burr/)for providing us Symbiodinium cultures. H. R. has been supported by the  Proscience and TeMana o teMoana (French Polynesia ) associations. This work was funded by the D el egation a la  Recherche of French Polynesia , the  Minist ere de l'Outre-Mer , and the contrat de projet Etat-Polyn esie franc ßaise.
ACKNOWLEDGMENTS We thank all members of our laboratories and  Helen Pickersgill  for critical reading and expert advice, and we thank  Cecile Pickart  and  Marc Timmers  for the bacterial ubiquitin E2-expressing plasmids. We are grateful to the members of the  IMP-IMBA Biooptics facility  for assistance in cell sorting. SMAC mimetic  BV6  and  RIP3  kinase mutant mice were generously obtained by  Genentech. The work was supported by the  Austrian Academy of Sciences  and an advanced  European Research Council (ERC)  grant (to J. M. P. ), the  Helmsley Foundation (VEO-IBD , to J. M. P. ), an  Innovator award  by  Era of Hope  (to J. M. P. ), the  Max Planck Society  (to A. P. ),  WWTF LS05003  (to A. P. ),  DFG PI 917/1-1  (to A. P. ), and by the  Swiss National Science Foundation  ( PBEZP3_145993  to L.
T. ). M. J. M. B. has a tenure track position in the  Multidisciplinary Research Program of Ghent University  (GROUP-ID). G. K. is supported by the  Ligue Nationale contre le Cancer, Agence Nationale pour la Recherche, European Commission (ArtForce) ,  Institut National du Cancer (INCa) ,  Canceropôle Ile-de-France ,  Fondation Bettencourt-Schueller ,  LabEx Immuno-Oncology , and PACRI. P. V. is supported by European grants ( FP7 EC RTD Integrated Project ,  Apo-Sys ,  FP7-200767 ;  Euregional PACT II), Belgian grants (Interuniversity Attraction Poles ,  IAP 6/18 ), Flemish grants ( Research Foundation Flanders ,  FWO G. 0875. 11  and  FWO G. 0973. 11 ),  Ghent University  grants (MRP, GROUP-ID consortium), and grants from  Flanders Institute for Biotechnology (VIB). P. V. also holds a  Methusalem grant  ( BOF09/01M00709 ) from the  Flemish Government.
Data Availability Statement: All relevant data are either within the paper and its Supporting Information files or available from the NCBI; MIseq reads used for de novo genome assembly and Hiseq reads used for RNAseq de novo have been deposited at SRA under accession numbers SRX883538 (http://www. ncbi. nlm. nih. gov/sra/?term=SRX883538) and SRX969055 (http://www. ncbi. nlm. nih. gov/sra/?term=SRX969055), respectively. This Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession JYHR00000000 (http://www. ncbi. nlm.
Drechmeria coniospora produces non-motile spores (conidia) that stick to the nematode cuticle, via a specialized adhesive bud  [10, 11]. Shortly after, the spores germinate, producing an appresorium that allows the fungus to pierce the nematode cuticle and send hyphae into its epidermis  [10]. Until now, there has been essentially no molecular characterisation of D. coniospora. Thus, at the start of this project, nothing was known about its genetic makeup, apart from 1. 05 kb of rRNA sequence in Genbank (GI:16763389; AF106012) that had been used to assign D. coniospora to the hypocrealean family, Clavicipitaceae, which includes many fungal pathogens of arthropods, such as Beauveria bassiana  [12]. The same single sequence was used in a subsequent analysis that removed Drechmeria from the Clavicipitaceae and recognized it as one of six genera within the Ophiocordycipitaceae  [13].
Results Sequencing output processing, de novo genome assembly and scaffolding D. coniospora genomic DNA was sequenced on an Illumina MIseq sequencer as 2 x 150 bp paired-end reads. After filtering, we obtained 11. 3 million reads, for a 100X coverage of a genome originally estimated to be 30 Mb. To determine which frequently used de novo genome assembly program performed best with this set of data, we tested four, Velvet  [34] , SPAdes  [35] , SOAPdenovo2  [36]  and ABySS  [37]. Each assembly was scaffolded with SSPACE  [38] , using two libraries of mate-paired 2 x 60 bp SOLiD reads. We applied a stringent filter, keeping only very high-quality reads, to limit errors during scaffolding. The libraries finally contained 23. 2 and 23. 6 million mate-paired reads, with insert sizes of 1. 5 kb and 3 kb, respectively.
Optical mapping data integration We then used optical mapping to test further the quality of the different assemblies generated before and after SSPACE scaffolding. Individual chromosomes were stretched on a glass slide and cut in situ with a restriction enzyme. The resulting fragments were visualized using a fluorescent microscope and their lengths measured. These lengths were compared to the predicted lengths of fragments from the longer scaffolds (i. e. with a length of at least 20 kb). The same approach was applied for each assembly. The nine distinct maps that were identified by optical mapping are indicative of a genome organization into 9 distinct chromosomes, with sizes ranging from 0. 58 to 11. 3 Mb (S1 Table ). The assembly obtained with ABySS (kmer = 96) was selected since it maximized the remapping of the scaffolds on the optical map whilst at the same time minimizing the number of misassemblies observed in the optical map analysis (4 versus 20 for Spades and 9 for ABySS at a kmer value of 64; S1 Table ).
coniospora genome. Transposable elements, repeat sequences, tRNAs, rRNAs and mtDNA Amplification of transposable elements (TE) can contribute to chromosomal rearrangements, altering genomic structure and gene expression. It is believed to be an important route to speciation in some fungi. We therefore characterized the set of TEs predicted in the D. coniospora genome using TransposonPSI (http://transposonpsi. sourceforge. net/). A total of 600 elements were detected, falling into 12 classes (Table  2 ), and covering 1. 6% of the genome (516 kb). We compared these results with the number of TEs found in the genomes of 11 other fungi, chosen on the basis of their phylogenetic position and/or lifestyle, using the name assigned by the NCBI Taxonomy Database: (1) Arthrobotrys oligospora, Hirsutella minnesotensis, Monacrosporium haptotylum and Pochonia chlamydospora, 4 nematophagous fungi that feed on different species and stages of nematode worms; (2) the entomopathogenic fungi Metarhizium acridum, Metarhizium anisopliae and Ophiocordyceps sinensis; (3) the plant pathogens Fusarium graminearum and Fusarium oxysporum; (4) the mycoparasite Tolypocladium ophioglossoides; (5) Trichoderma reesei, a model mesophilic and filamentous fungus.
We also searched for predicted tRNAs and found a total of 123 nuclear tRNA loci (Fig  1 ). Eighteen mitochondrial-specific tRNAs were located on Scaffold43 (23. 8 kb, G+C 25. 7%) after a tRNAscan search  [43]  with the option "organelle". The 141 predicted tRNAs correspond to 49 out of 64 codons. This degree of coverage compares very favourably with other sequenced fungal genomes (S3 Table ). A more comprehensive investigation of Scaffold43 with TBLASTX  [44]  against a database of mitochondrial genes led to the identification of homologues for multiple mitochondrial proteins and established it as the mitochondrial DNA (S1 Text).
While overall, 60% (5248/ 8733) of the predicted proteins have a homologue in the curated UniprotKB/Swissprot database, 74. 8% of the proteins as long or longer than the median were assigned a homologue, while only 34. 6% of the proteins less than 250 amino acids were. It should be noted that these are conservative estimates of homology since although of high quality, the UniprotKB/Swissprot database is not exhaustive. Indeed, for example, on the basis of BLASTP analyses using the current NCBI non-redundant database, many (19/31) of the predicted proteins longer than 1000 amino acids but without any annotation have homologues in other fungi. The closest homologue was most often found in Tolypocladium ophioglossoides (S5 Table ), consistent with the phylogenetic analysis. To define gene families and assign proteins to orthologous and paralogous groups, we performed an OrthoMCL analysis  [52].
The presence of the Saposin A domain (PF02199) in 2 predicted proteins (g3895. t1 and g1982. t1, with 3 and 2 domains respectively) is equally atypical (S5 and S7 Tables). We address its possible role below. On the basis of their constituent domains, several other atypical or highly represented protein families  (Fig 4 and S7 Table)  are also potentially linked to virulence. These include the single iron-sequestrating lipocalin (PF13924) and the deuterolysin M35 metalloprotease (PF02102; called here M35) domains. The M35 domain is unusually highly represented compared to other fungi  [58, 59] , being present in 10 predicted D. coniospora proteins. Of a total of 777 domains absent from D. coniospora, 32 were present in all 11 other species (S7 Table ). As a most striking example, D. coniospora lacks proteins containing the NACHT domain (PF05729), which is present between 8 and 117 times in the other species, suggesting that this is unlikely to simply be a problem of sequence coverage or gene prediction.
In ascomycete fungi, the NACHT domain can be found together with the HET domain (PF06985) in heterokaryon incompatibility proteins. It acts as a death effector domain, preventing viable heterokaryotic cells from being formed by the fusion of filaments from different wild-type strains  ). (B) All-against-all dot-plot representation  [55]  of the alignment of the predicted protein sequences from g4180. t1 and from 5 neighbouring genes on scaffold omap6908, all from the OrthoMCL paralogous group 2. Dots represent regions of sequence similarity (within a 100 a. a. sliding window). The intensity of each dot is proportional to the corresponding alignment score. The ". t1" suffix has been removed from all sequence names. doi:10. 1371/journal. pgen. 1006017. g003 doi:10. 1371/journal. pgen. 1006017.
g004 [60]. In D. coniospora, of the 6 proteins predicted to contain a HET domain, 5 (g3856. t1, g5845. t1, g5969. t1, g6809. t1, g7038. t1) do not contain any other domains, while in common with many other fungal proteins of this class, one (g215. t1) is coupled to an ankyrin repeat (PF12796) domain. No sexual stage has been described for D. coniospora, and in common with 3 of the 4 other nematophagous fungi, no mating type protein (MATα, PF04769) domain was predicted (S7 Table ), making it unlikely that these different proteins play any role in heterokaryon incompatibility. To see whether there was any general pattern for the 32 domains absent from D. coniospora suggestive of a coordinate evolutionary process, we used dcGO analysis to look for ontology term enrichment at the domain level  [61].
This revealed a potential involvement of the NACHT domain together with 4 others (PF00931; PF01966; PF02178; PF09273) in the biological process, "regulation of response to stress" and of 4 of them (PF00931; PF01966; PF05729; PF09273) in "regulation of defense response". This is discussed further below. Submitting all 777 domains absent from D. coniospora to dcGO analysis highlighted the absence of 16 related domains, all annotated as being involved in hydrolase activity, acting on glycosyl bonds, including PF00331 that corresponds to the Carbohydrate-Active Enzyme (CAZyme) glycosyl hydrolase (GH) family 10. There was also a significant (p = 2. 5x10 -4 ) enrichment for the more specific ontology term "alpha-N-arabinofuranosidase activity" (PF05270, PF06964, PF09206), suggesting an alteration in the capacity of D.
PHI-base analysis To gain a general view of proteins potentially involved in virulence, we made use of the pathogen-host interactions database, PHI-base (phi-base. org; [65] ). Of the 2104 proteins matching a PHI-base entry, 990 had the annotation "reduced virulence" or "loss of pathogenicity", indicating that a homologous protein in at least one other species plays a demonstrated role as a virulence factor in a particular model of infection (S5 Table ). Among the highly represented (!5) hits in PHI-base, several were characterised by the presence of ABC transporter domains, and so predicted to be involved in ATP-dependent export of organic anions or drugs from the cytoplasm (S10 Table  and Fig 1). For predicted NRPS proteins, 9/20 members of the orthoMCL group OG5_126718 were assigned a PHI-base annotation (PHI:2511).