dataesr/scientific-paragraphs-categorization huggingface dataset:
If identification fails, then speciesspecific PCR or universal PCR targeting Mollicutes 16S rRNA followed by sequence analysis is used  (20). The protocol detects both pathogenic and commensal/opportunistic, cultivable (sub)species. Metadata on the clinical specimens (using a shared terms' list for host animal species, geographic origin, nature of the specimen, clinical signs, age of sampled animals and sampling date, etc. ) is sent by partner laboratories and recorded in an inhouse database (developed by JL Vinard using a MySQL database management system and a Windev Computer Aided Software Engineering from PCSoft to provide a user-friendly operator interface). This database allows easy detection of "duplicates, " i. e. , cultures sourced from the same village or the same animal with a common sampling date.
The reference distributions for M. bovis MICs were retrieved from two previous studies conducted in our laboratory in which AST was performed on Vigimyc isolates using the same methodology  (15, 17). It included between 73 and 170 MIC values, depending on the antimicrobial, that were obtained on both "recent" (between 2000 and 2014) and "older" (after 1978 and before 2000) isolates. Contrary to the situation for M. bovis, no internal reference dataset was available for the M. mycoides-group (sub)species. The reference distribution was established here by testing 58 Mmc, 60 Mcc, and 51 Mp isolates collected between 1977 and 2016, that is 144 "recent" (2011-2016) and  25 "older" (1977-1999)  isolates. To generate this dataset, we determined the MICs of the most commonly-used antimicrobials in small ruminants (Supplementary Figure  1 ).
For routine annual surveillance initiated in 2018, MICs were estimated by testing only two to five concentrations of one antimicrobial per class, chosen according to MIC reference distributions and defined to capture the evolution compared to the reference population (see section Results and Discussion). This strategy aimed at sparing testing-time in order to make yearly surveillance compatible with routine practice and available resources while enabling to quicken early comparison with the reference dataset. A minimum of 10 (and up to 59) isolates per year and per species were chosen from the Vigimyc collection according to their isolation frequency, geographical diversity, clinical history and purity in initial culture (not being part of a subculture containing a mix of Mycoplasma species).
Determination of Epidemiological Cut-Offs MIC distributions obtained with the reference datasets were tentatively used to determine ECOFFs. However, as these MICs did not fulfill all EUCAST SOP 10. 1 requirements (11), we used the term "tentative ECOFF" or "TECOFF" instead of ECOFF. Whenever possible, we estimated both visual and numerical TECOFFs, using the ECOFFinder program (ECOFFinder 2. 1).
One noticeable difference was observed for spectinomycin, where a population with MICs below 64 µg/mL was detected in 2018 (n = 1) and 2019 (n = 11) that was absent from the "recent" strains (2010-2012) of the reference dataset  (15). Furthermore, between 2018 and 2019, there was an increase in isolates with very high MIC values (≥1,024 µg/mL) for tilmicosin (from 79 to 97%) and a potential shift toward higher MICs for florfenicol (from 46 to 83% of MIC ≥ 8 µg/mL) and enrofloxacin (from 17 to 52% of MIC values equal to 0. 5 or 1 µg/mL). These data need to be consolidated by further similar observations over the coming years.
M. mycoides Group: Reference Population and Surveillance Results MIC distributions of 169 isolates of the Mycoides group (sub)species, taken as a reference dataset, are represented in Supplementary Figure  1  and summarized in Table  3. For tylosin, we found a dominant population with low MIC values centered on 0. 125 µg/mL (≤0. 0625-0. 25) that was putatively considered as WT, whereas 14 recent Mcc and 3 recent Mmc isolates showed increased widely-distributed MICs ranging between 1 and 128 µg/mL. The important spread of MICs suggests different, possibly cumulative mechanisms of AMR. This is completely different from M. bovis, for which the shift between WT and non-WT was strictly correlated with homogeneous mutations in the ribosomal target  (17). For lincomycin, two slightly overlapping populations were evidenced, one dominant and centered on 2 µg/mL, and the other with MICs above 8 µg/mL observed for Mcc (recent and older) and Mmc (recent).
125, 1 and 16 µg/mL for tilmicosin, 0. 5 and 2 µg/mL for oxytetracycline, 2 and 16 µg/mL for lincomycin, 64 and 128 µg/mL for spectinomycin, and 0. 125 and 0. 25 µg/mL for enrofloxacin. In 2018 and 2019, 38 and 28 isolates of Mmc, 11 and 17 isolates of Mcc and 10 and 10 isolates of Mp, respectively, were included in the annual AMR monitoring program. This distribution paralleled the frequency of detection of each sub(species) within Vigimyc, although the MICs were aggregated for comparison to the reference dataset (Table  3 ). MICs remained unchanged in 2018 and 2019 compared to the reference population, with (i) for tylosin and lincomycin, a dominant population below 0. 125 and 2 µg/mL and a few strains with MICs exceeding 1 and 16 µg/mL, respectively, (ii) for oxytetracycline and enrofloxacin, monomodal populations centered around 0.
125 and 0. 5 µg/mL, respectively, and (iii) for spectinomycin, a dominant population harboring MIC ≤ 64 µg/mL. In 2019, we introduced some supplementary concentrations to check the upper and lower limits. The results confirmed that higher lincomycin MICs (n = 3) do not exceed 128 µg/mL, that most of enrofloxacin MICs above 0. 125 µg/mL were equal to 0. 25 µg/mL (n = 22), and that spectinomycin MICs were mostly between 16 and 64 µg/mL (n = 53). Note, however, that two isolates from 2019 harbored spectinomycin MIC ≤ 8 µg/mL, which was not observed in the reference dataset. Unlike in the reference dataset, there were also increased tilmicosin or lincomycin MICs for Mp (one strain with a tilmicosin MIC between 2 and 16 µg/mL and one with lincomycin MIC >16 µg/mL; data not shown). Our approach enabled to evidence putative multiresistant isolates harboring high MICs for at least 3 antimicrobials.
For instance, in 2019, 4 isolates of Mcc had MICs ≥ 2 µg/mL for tilmicosin, ≥4 µg/mL for lincomycin and ≥32 for spectinomycin (data not shown). These multiresistant patterns were also detected in our reference dataset, and thus require particular vigilance as they could result in emergence and clonal expansion, as shown for M. bovis  (28).
Larger-Scale Contribution and Improvement of the Surveillance Framework To broaden the perspectives of our surveillance framework, we assessed whether our data could contribute to the determination of ECOFFs that could usefully distinguish WT and non-WT populations or serve as a first step toward CBPs. According to EUCAST SOP, the process of setting ECOFFs requires aggregation of at least 5 datasets of different origins, each one containing (i) exact MIC values and (ii) at least 15 isolates belonging to the WT population, that needs to have a normal 3-5 dilutions-wide distribution. The size of the WT population in this aggregated set should be increased to >100 isolates for ECOFF determination using statistical approaches (EUCAST SOP). Our reference dataset was partly compliant with these criteria.
capricolum subsp. capricolum and M. putrefaciens. Number of isolates in each MIC category are shown. "/" means no isolate in the corresponding category. Note that for macrolides, tilmicosin was tested for annual surveillance and tylosin for the reference dataset. ENRO, enrofloxacin; LINCO, lincomycin; OXYTETRA, oxytetracycline; SPECTINO, spectinomycin; TILM, tilmicosin; TYL, tylosin. is interesting as it highlights a limited impact of the chosen methodology. For oxytetracycline, florfenicol and enrofloxacin, the monomodal distribution evidenced on our dataset was also found in other studies  (18, 19, (30) (31) (32) (33) , which rules out a TECOFF. For the Mycoides group (Table  3 ), the situation is more favorable to both visual and statistical TECOFF determination, with lower MICs overall for most antimicrobials and hence a potential WT population.
e. , a unique population with a maximum MIC of 0. 4 µg/mL (34-37, 40), but the truncated lower end again rules out a statistical TECOFF. For lincomycin, the TECOFF visually derived from our data was putatively between 4 and 8 µg/mL and was definitively set at 4 µg/mL using ECOFFinder with a 99. 9% endpoint. This result slightly exceeds those obtained by Tatay et al. (37)  for Mcc, with lincomycin MICs showing a bimodal distribution with a WT upper limit at 1. 6 µg/mL. Visual and statistical TECOFFs for oxytetracycline were estimated as 2 µg/mL on our results, which is similarly slightly higher than the visual TECOFF derived from other studies (≤1 µg/mL)  (40, 41). In conclusion, we were able to use our data to propose several TECOFFs (Tables  2, 3 ), most of which were consistent or only one to two dilutions different to other datasets, even if these were obtained using other AST methodology.
Methods: MIC Determination, Data Reporting, AST Interpretation Isolation was performed according to harmonized guidelines using fertility-controlled culture media that help to standardize the isolate collection process between laboratories belonging to the network [S5]. AST was implemented according to the currently-recommended methods for mycoplasma  (7)  in the absence of reference method  (11)  [S6]. Moreover, AST is centralized in one lab, which limits variability in results [S7] but runs counter to the EUCAST recommendations which promote the inclusion of inter-laboratory variability. However, this situation may prove precarious in the longer term [T1], as several methodological choices are done to accommodate timesaving considerations and end up being non-EUCAST compliant (W2), which could introduce AST biases.
CONCLUSION AMR surveillance was successfully integrated into the clinicaldedicated Vigimyc network and enabled us to track resistance trends in different pathogenic mycoplasma species in a way that meets most criteria for surveillance guidelines and that link into clinical evolution. This new AMR surveillance framework further benefited from the multi-species and broad-geographical coverage inherent to Vigimyc. This monitoring system meets the needs for surveillance in a context of concern around AMR where mycoplasmas have so far been neglected despite being significant drivers of antimicrobial use. However, the perspectives for a larger-scale contribution of Vigimyc to surveillance remain limited as of now. Our methodological choices mean that our data was only partially compliant with EUCAST requirements, and aggregation with other datasets for ECOFF setting was limited.
MICs distributions of 5 antimicrobials for M. bovis isolates either from the reference populations or from 2018-2019 annual monitoring. Number of isolates in each MIC category are shown. "/" means no isolate in the corresponding category. Pasteurellaceae clinical intermediate breakpoints for cattle (CLSI 2015) are represented by a horizontal red separator line. REF1 and REF2 correspond to reference datasets of isolates collected between 1978-2014 (15, 17) and 1978-2012 (15), respectively. ENRO, enrofloxacin; FLOR, florfenicol; OXYTETRA, oxytetracycline; SPECTINO, spectinomycin; TILM, tilmicosin.
The Sedimentation Rate (SR) was determined before and at the end of treatment. The swelling was assessed using a tape measure before treatment and at the end of treatment; stiffness was measured using a goniometer. Observations were noted at the end of treatment according to the following criteria: aggravated state if there is an increase in at least one of the signs of inflammation with or without an increase in SR; steady state if there is no noticeable change; improved condition if there is a decrease in inflammatory signs with or without a decrease in SR; and cure: absence of inflammatory signs with or without return of SR to normal. In order to assess the tolerance of the product, clinical symptoms of intolerance were sought and data concerning all possible adverse events were recorded.
Libraries were prepared using a Nextera XT DNA Library Preparation Kit and Nextera XT Index Kit (Illumina, Paris, France). Samples were then sequenced with a NextSeq 500 machine using a NextSeq 500 Mid Output Kit v2 (300 cycles), (Illumina, France). Paired-end raw reads were deposited on the EnteroBase database platform for Salmonella (http://enterobase. warwick. ac. uk/, accessed on 16 April 2021), and were automatically de novo assembled using SPAdes  [25]  once the sequences were uploaded. Genomic comparison of the isolated strains was carried out using the cgMLST scheme with 3002 genes included, available on EnteroBase  [26]. Similar but nonidentical strains (strains showing different core genome Sequence Type (cgST)) were identified in EnteroBase by using the hierarchical clustering method (HierCC) that allows for grouping of strains into hierarchical clusters (HCs) that can differ up to a specified and fixed number of cgMLST alleles.
This number is indicated by the suffix following "HC" (e. g. , HC5 for 5 cgMLST allelic differences). To assess the genetic relationship between strains of the same serotype and the population structure of Salmonella isolates, a neighbor-joining tree was created from cgMLST allelic differences in EnteroBase using GrapeTree  [27]  and the RapidNJ algorithm  [28]. The assembly sequences are publicly available from EnteroBase; their accession numbers (barcodes) are listed in Table  3.
Materials and Methods Cell culture and reagents. Huh7. 5 cells (kind gift of C. Rice, Rockefeller University, New York, USA), Vero E6 cells (ATCC CRL-1586) and 293T kidney (ATCC CRL-1573) cells were grown in Dulbecco's modified minimal essential medium (DMEM, Invitrogen, France) supplemented with 100U/ml of penicillin, 100μg/ml of streptomycin, and 10% fetal bovine serum. Plasmids. Homo sapiens codon optimized SARS-CoV-2 S (Wuhan-Hu-1, GenBank: QHD43419. 1) was cloned into pVAX1 vector. The delta 19 truncation of S form was generated by site directed mutagenesis introducing a stop codon after Cys1254  [45]. SARS-CoV-2 E, M and N genes (Wuhan-Hu-1, GenBank: QHD43419. 1) were synthesized and cloned into pCDNA3. 1(+) vector. The plasmid pEGFP-N3-VSV-Gts (kind gift from K. Konan, Albany Medical College, USA).
The plasmids encoding HCV ΔE2p7(JFH1) was described previously  [18]. Antibodies. Mouse anti-actin (clone AC74, Sigma-Aldrich), rabbit anti-SARS-CoV2 S2, mouse anti-SARS-CoV2 S1 and mouse anti-SARS-CoV2 N (Sino Biological), mouse anti-GFP (Roche), anti-VSV-G 41A1 and rabbit anti-GM130 (clone EP892Y, Abcam) were used according to the providers' instructions. SARS-CoV-2 particles (kind gift of B. Lina, CIRI, Lyon) are referenced in GISAID EpiCoVTM database (reference BetaCoV/France/IDF0571/2020, accession ID EPI_ISL_411218) and were amplified on VeroE6 cells  [46]. Briefly, for stock production, cells were infected with MOI=0. 01 in DMEM for 90min at 37°C. Then, medium was replaced with DMEM-2%FCS. Supernatant fluids were collected after two days at 37°C, clarified by centrifugation (400xg, 5min), aliquoted and titrated in plaque forming unit by classic dilution limit assay on the same Vero E6 cells.
1% Triton X-100. After a saturation step with 3% BSA/PBS, cells were incubated for 1 hour with rabbit anti-GM130 and mouse anti-SARS-CoV2 S1 antibodies at 1/200 dilution in 1% BSA/PBS, washed 3 times with 1%BSA/PBS, and stained for 1 hour with donkey anti-rabbit AlexaFluor-488 and donkey anti-mouse AlexaFluor-555 secondary antibodies (Molecular Probes) diluted 1/2,000 in 1% BSA/PBS. Cells were then washed 3 times with PBS, stained for nuclei with Hoechst (Molecular Probes) for 5 min, washed and mounted in Mowiol (Fluka) before image acquisition with LSM-710 or LSM-800 confocal microscopes. Images were analyzed with the ImageJ software (imagj. nih. gov) and the Manders' overlap coefficients were calculated by using the JACoP plugin. Cell-cell fusion assay. The cell-cell fusion assay was adapted from  [48].
Statistical analysis Significance values were calculated by applying the Kruskal-Wallis test and Dunn's multiple comparison test using the GraphPad Prism 6 software (GraphPad Software, USA). For fusion index, a two-tailed, unpaired Mann-Whitney test was applied. P values under 0. 05 were considered statistically significant and the following denotations were used: ****, P≤0. 0001; ***, P≤0. 001; **, P≤0. 01; *, P≤0. 05; ns (not significant), P>0. 05. (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Figure legends The copyright holder for this preprint this version posted August 24, 2020. ; https://doi. org/10. 1101/2020. 08. 24. 260901 doi: bioRxiv preprint.
Acknowledgments We thank to  Konan Kouacou  for the VSV-Gts construct and  Charles Rice  for the Huh7. 5 cells. We also thank  B. Lina  for the SARS-CoV-2 particles. We thank  Lu LU  and  Moumita MONDAL  for technical help in the generation of SARS-CoV2 derived plasmids We acknowledge the contribution of  SFR Biosciences  ( UMS3444/CNRS ,  US8/Inserm ,  ENS de Lyon, UCBL)  facilities:  LBI-PLATIM-Microscopy, ANIRA-Cytometry  for excellent technical assistance and support. We than  Didier Décimo  for support with the BSL3 facility. (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
Fundings FLC received financial support from the  LabEx Ecofect  ( ANR-11-LABX-0048 ) of the '' Université de Lyon" , within the  program ''Investissements d'Avenir"  ( ANR-11-IDEX0007 ) operated by the  French National Research Agency (ANR) , the  ANR  (grant from  RA-Covid-19 ), and the  Fondation pour la Recherche Médicale  (FRM). DL received financial support from  National Key R&D Program 376 of China  ( 2020YFC0845900 ,  D. La ),  Shanghai Municipal Science and Technology Major Project  ( 20431900402 ,  D. La ). theNational  Natural Science Foundation of China  ( 31870153 D. La ),  Chinese academy of Sciences PIFI program  ( D. La ). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
| Assessment of novel variations Variations were searched in databases, including the NCBI (NCBI. nlm. nih. gov), the Leiden Open Variation Database (LOVD3), the Exome Variant Server (evs. gs. washington. edu/EVS/), and the UMD-TREAT-NMD DMD mutations database (Universal Mutation Database-Translational Research in Europe for the Assessment and Treatment of Neuromuscular Disorders). The corroborated novel variants were deposited at the NCBI site and analyzed for pathogenicity using in silico web tools such as PredictSNP2, MutationTaster Variant Effect Predictor (VEP), and SIFT indel LoF Score Human Splice Finder  (Bendl et al. , 2014; Desmet et al. , 2009; Hu & Ng, 2013; McLaren et al. , 2016; Pagel et al. , 2017; Schwarz et al. , 2014). The 2015 American College of Medical Genetics and Genomics criteria were also used for variant classification and correct nomenclature  (Richards et al.
, 1988). The patients studied in the present study are part of a majority around the globe that is underrepresented by the "European bias" and is missing diversity data in human genetics studies  (Sirugo et al. , 2019). To the present authors' knowledge, this is the first molecular report to use MLPA and targeted NGS of DMD variants to study dystrophinopathies on a Peruvian population. Out of 125 patients with DMD mutations, we found that 41. 6% of participants exhibited exon deletions, 16% duplications, and 42. 4% small mutations. Percentages differ from results reported for other populations, which show a higher number of exon deletions/duplications and a smaller mutation prevalence of <25%. For example, reports of the LOVD3, and the UMD-TREAT-NMD DMD variation database, each with several thousands of mutations, report 66%-69% exon deletions and 22%-20% small mutations, respectively  (Aartsma-Rus et al.
Regarding therapies for specific mutations-such as drugs that read through premature stop codons, antisense oligonucleotide-exon skipping, and stem cells-there are, as yet, no local trials but increased information related to molecular diagnoses will help health authorities better understand the issue. Creating a database with these variations will become a priority for physicians and health authorities to depict the spectrum of variations found in Peru, and certainly, it will have an impact in other Andean countries due to our common history. In our sample, it would ideally help 34 patients who could benefit from the readthrough stop codon therapy and nine patients with the exon skipping of exon 51 strategy. In conclusion, this is the first report of an analysis performed among Peruvian patients with clinical diagnoses of dystrophinopathy from different health institutions in Lima.
None of these variables was retained in the nal analysis as alcohol and smoking were not associated with health outcome and level of education was closely correlated to the jobs and exposures under study. Separate analyses were run for each histological type. We used 2-sided statistical tests and a 5% signi cance level. Analyses were performed with the SAS ® software, version 9. 2 (SAS Institute Inc, Cary, NC, United States).
The result showed that the LOH was found to contain a high number of phenols (740. 06 ± 1. 2 µg GAE/mg). The total flavonoid content of LOH was determined by utilizing aluminum chloride colorimetric method. The tested LOH was found to contain high amounts of flavonoids (550 ± 2 µg RE/mg). Phytochemical screening of LOH by means of UPLC-MS analysis revealed the presence primarily of phenolic acids and flavones C-glycosides compounds. A variety of these phenolic compounds were identified, and these relative proportions are different (data not shown). The results obtained were compared with those obtained by  Tchoumtchoua et al. (2019)   [27]  and confirmed that the aerial parts of flax contain a significant source of phenolic compounds.
Spectral Characterization of the Synthesized Alg/nHA/LOH FTIR analysis of Alg/nHA/LOH samples, sodium alginate, and nHA powder was carried out by accumulating 16 scans at a resolution of 4 cm -1 over a wavenumber region of 400-4000 cm -1 using an FTIR spectrometer (Spectrum RX1 FTIR system, Perkin-Elmer, Texas City, TX, USA). The Powder X-ray diffraction (XRD) characteristics of the nHA were carried out using a Philips Xpert instrument. X-ray diffraction data were collected using Cu-Kα radiation operating at 40 kV and 30 mA with 2θ ranging from 10 to 90 • at a scan speed of 0. 08 s -1. For each sample, peak intensity changes were analyzed relative to the reference band within the same spectra. The particle size, polydispersity index (PDI), and zeta potentials of nHA and synthesized samples were measured using a dynamic light scattering (DLS) instrument (Zetasizer Nano ZS90, Malvern, Worcestershire, UK).
Release Study The release kinetics of LOH from hydrogel in different releasing mediums of PBS and deionized water were compared. In any solution, certain quantities of formulated gels were immersed in a 20 mL releasing medium at room temperature. The release rate of the LOH from Alg/nHA/LOH 21% was determined periodically via withdrawing 2 mL of supernatant solution and depositing it into an Eppendorf tube at appropriate time intervals up to 2 h. Two ml of fresh releasing media was added at each time point to maintain the solution volumes in 20 mL and the sink condition. Cumulative release of LOH in the aliquots was quantified by measuring the absorption band centered at 270 nm by UV-VIS spectroscopy and calibration curve axis. The data represent triplicate determinations. 4. 9. In Vitro Evaluations 4.