dataesr/scientific-paragraphs-categorization huggingface dataset:
As such, we solved an X-ray crystal structure of B. abortus EipB (residues A30 to K280; PDB ID 6NTR). EipB lacking its signal peptide formed triclinic crystals (a ϭ 47. 4 Å, b ϭ 69. 2 Å, c ϭ 83. 2 Å, ␣ ϭ 90. 1°, ␤ ϭ 90. 0°, ␥ ϭ 78. 7°) that diffracted to 2. 1-Å resolution; we refined this structure to R work ϭ 0. 195 and R free ϭ 0. 245. Crystallographic data and refinement statistics are summarized in Table  S2. Four EipB molecules (chains A to D) are present in the crystallographic asymmetric unit. Each EipB monomer consists of 14 antiparallel ␤-strands (␤1 to ␤14) forming an oval, spiral-like ␤-sheet (minor axis diameter, ϳ25 Å; major axis diameter, ϳ35 Å). Two regions of this ␤-spiral, involving ␤5, ␤6, ␤7, and ␤8, and the hairpin loop connecting ␤9 and ␤10 overlap (Fig. 5 ). Interactions between these two overlapping portions of structure are mostly hydrophobic, though polar contacts are also found in these regions (Fig.
5  and 6 ). One side of the spiral is occluded by the N terminus, a loop connecting ␤-strands 12 and 13, and ␣-helix 1, which form the bottom of this "cup" shaped protein (Fig. 5  and 6A ). The external surface of EipB is positively and negatively charged and also presents small hydrophobic patches (Fig. S5 ); one helix, ␣2, is kinked and positioned at the surface of the cylindrical ␤-spiral (Fig. 5 ). The lumen of EipB is solvent accessible and is partially filled with the side chains of hydrophobic or acidic residues. Hydrophobic residues represent ϳ66% of the residues present inside the EipB cavity (Fig. 5  and 6B ). The size of this cavity suggests that EipB, in this conformation, can accommodate small molecules or ligands in its lumen. We searched the EipB structure against the protein structure database using Dali (  22 ) but failed to identify clear structural homologs.
Pseudomonas aeruginosa PA1994 (PDB ID 2H1T)  (23)  was the closest structural match to EipB (root mean square deviation, ϳ3. 5; Z-score, ϳ11) (Fig. S6A ). Despite very low sequence identity (ϳ8%), PA1994 has noticeable structural similarities to EipB: it adopts a spiral-like ␤-fold involving 15 ␤-strands, which is occluded at one end with a long ␣-helix. Unlike EipB, PA1994 lacks a signal peptide and is predicted to be a cytoplasmic protein. Structural parallels between PA1994 and the periplasmic lipoprotein chaperones LolA/LolB have been noted and a role for PA1994 in glycolipid metabolism has been postulated  (23) , though this prediction remains untested. Like PA1994, EipB has structural similarities to LolA and LolB, in particular the antiparallel and curved ␤-sheet scaffold that engulfs a central ␣-helical plug (Fig.
Reduced stability of EipB lacking its conserved disulfide bond may contribute to the 1-log relative growth defect of ΔeipB strains expressing EipB cysteine mutants on SBA carbenicillin plates (Fig. 7C  and D ). eipB deletion is synthetically lethal with bab1_0430 (ttpA) disruption and synthetically sick with disruption of multiple genes with cell envelope functions. To further characterize how eipB functions in the Brucella cell, we aimed to identify transposon (Tn) insertion mutations that are synthetically lethal with eipB deletion in B. abortus (see Tables  S3  and S4 ). In other words, we sought to discover genes that are dispensable in a wild-type genetic background but that cannot be disrupted in a ⌬eipB background. By sequencing a Tn-Himar insertion library generated in B. abortus ⌬eipB (NCBI Sequence Read Archive accession number SRR8322167) and a Tn-Himar library generated in wild-type B.
abortus (NCBI Sequence Read Archive accession number SRR7943723), we uncovered evidence that disruption of bab1_0430 (RefSeq locus BAB_RS17965) is synthetically lethal with eipB deletion. Specifically, reproducible reads corresponding to insertions in the central 10 to 90% of bab1_0430 were not evident in ⌬eipB but were present in wild-type (Fig. 8A ). bab1_0430 encodes a 621-residue tetratricopeptide repeat-containing (TPR) protein with a predicted signal peptide and signal peptidase site at its N terminus. This protein was previously detected by mass spectrometry analyses of B. abortus extracts and was described as a cell envelopeassociated protein  (24)  or periplasmic protein  (25). Here, we refer to this gene as ttpA (tetratricopeptide repeat protein A gene) based on its similarity to Rhizobium leguminosarum ttpA  (12).
8A ). ttpA contributes to carbenicillin resistance. As ttpA disruption is synthetic lethal with eipB deletion, we postulated that these two genes have complementary functions or are involved in a common physiological process (i. e. , envelope integrity). Thus, to characterize ttpA and the nature of its connection to eipB, we deleted ttpA in B. ovis and evaluated its sensitivity to carbenicillin. All efforts to delete B. ovis ttpA (locus tag bov_0411) using a classic crossover recombination and sacB counterselection approach were unsuccessful, though hundreds of clones were screened. Efforts to delete the chromosomal copy by expressing a copy of ttpA from a plasmid also failed. This result is surprising considering that transposon insertions in B. abortus ttpA (NCBI Sequence Read Archive accession number SRR7943723) and B.
ovis ttpA (NCBI Sequence Read Archive accession number SRR7943724) are tolerated in wild-type backgrounds  (8). As an alternative approach to study the function of this gene, we inactivated ttpA using a single crossover recombination strategy. The resulting strain expressed a truncated version of TtpA containing the first 205 amino acids (including the signal peptide), immediately followed by 22 amino acids from the suicide plasmid. The corresponding B. ovis strain (ΔttpA) was then transformed with a plasmid-borne IPTG-inducible copy of ttpA (pSRK-ttpA) or with an empty plasmid vector (EV). We evaluated the sensitivity of these strains to carbenicillin by plating a dilution series on SBA plates containing 2 or 2. 5 g/ml carbenicillin, with or without IPTG inducer (Fig. 8B  and C ). Compared to the wild-type strain with an empty vector, the B.
In contrast, Brucella EipB does not have a predicted site for lipidation (i. e. , a lipobox) and is therefore unlikely to function as a membrane-anchored protein. The number of unique barcoded Tn-Himar insertions in the apolipoprotein N-acyltransferase gene lnt (bab1_2158; lnt conserved domain database score Ͻ e Ϫ173 ) is lower than expected in a ΔeipB background relative to the wild type (Fig. 8A ). This provides indirect evidence for a link between eipB and lipoproteins. Lnt catalyzes the final acylation step in lipoprotein biogenesis  (32) , which is often considered to be an essential cellular process. However, like Francisella tularensis and Neisseria gonorrhoeae  (33) , B. abortus lnt is dispensable (26) (Fig. 8A  and Table  S4 ). The data presented here suggest that transposon insertions are less tolerated in B.
4-numerical-aperture Ph3 objective. Images were acquired with a mounted Orca-ER digital camera (Hamamatsu) controlled by the Image-Pro software suite (Media Cybernetics). To prepare the different samples, cells were resuspended in PBS containing 4% formaldehyde. Agglutination assay. Agglutination assays were performed as previously described  (8). The different Brucella strains (B. ovis and B. abortus) were harvested and resuspended in sterile PBS at an OD 600 of ϳ0. 5. One milliliter of each cell suspension was loaded in a spectrophotometer cuvette and mixed with 20 l of wild-type B. abortus-infected mouse serum or with acriflavine (final concentration, 5 mM), and the OD 600 was measured at time zero and after 2 h. As a control, 1 ml of each cell suspension was also kept in a spectrophotometer cuvette without serum or acriflavine.
Initial crystallization screening was carried out using the sitting-drop, vapor-diffusion technique. After a week, EipB crystallized in the triclinic space group P1 from the condition 70 (F10) of the MCSG-2 crystallization kit, which contains 24% polyethylene glycol (PEG) 1500 and 20% glycerol. Prior to flash freezing in liquid nitrogen, crystals were cryoprotected by briefly washing them in the crystallization solution containing 25% glycerol. Crystallographic data collection and data processing. Se-Met crystal diffraction was measured at a temperature of 100 K using a 2-s exposure/degree of rotation over 260°. The crystals diffracted to a resolution of 2. 1 Å, and the corresponding diffraction images were collected on the ADSC Q315r detector with an X-ray wavelength near the selenium edge of 12.
66 keV (0. 97929 Å) for SAD phasing at the 19-ID beamline (SBC-CAT; Advanced Photon Source, Argonne, IL). Diffraction data were processed using the HKL3000 suite  (41). B. abortus EipB crystals were twinned, and the data had to be reprocessed and scaled from the P2 1 space group to the lower symmetry space group P1 with the following cell dimensions: a ϭ 47. 36 Å, b ϭ 69. 24 Å, c ϭ 83. 24 Å, ␣ ϭ 90. 09°, ␤ ϭ 90. 02°, and ␥ ϭ 78. 66°(see Table  S2 ). The structure was determined by SAD phasing using SHELX C/D/E, mlphare, and dm, and initial automatic protein model building with Buccaneer software, all implemented in the HKL3000 software package  (41). The initial model was manually adjusted using COOT  (42)  and iteratively refined using COOT, PHENIX  (43) , and REFMAC (44); 5% of the total reflections were kept out of the refinement in both REFMAC and PHENIX throughout the refinement.
abortus EipB WT or EipB C69SϩC278S , 50ϫ Sypro Orange (Invitrogen), and 2 mM DTT when needed. Each protein sample and solution was prepared with the same dialysis buffer (10 mM Tris [pH 7. 4], 150 mM NaCl, 1 mM EDTA). Plates (96 well; MicroAmp EnduratePlate optical 96-well fast clear reaction plates; Applied Biosystems) were heated from 25 to 95°C with a ramp rate of 0. 05°C/s and read by a thermocycler (QuantumStudio 5 real-time PCR system; Applied Biosystems/ Thermo Fisher Scientific) using excitation and emission wavelengths of 470 Ϯ 15 nm and 558 Ϯ 11 nm, respectively. Protein Thermal Shift software v1. 3 (Applied Biosystems/Thermo Fisher Scientific) was used to calculate the first derivative of the curve to determine the melting temperature. Bacterial two-hybrid protein interaction assay.
The protein was eluted with 200 l of the same buffer containing 20 mM maltose. The different protein samples (elutions and flowthroughs) were subjected SDS-12% PAGE and Coomassie stained. Bioinformatics. Figures of the structures, structural alignments, electrostatic potential representations, and root mean square deviation calculations were performed using PyMOL (PyMOL Molecular Graphics System, version 1. 7. 4; Schrödinger, LLC). Surface hydrophobicity was evaluated using the YRB python script  (47). The XtalPred server  (48)  and Dali server  (49)  were used to identify proteins with the highest structural and sequence relatedness. The BLAST server (https://blast. ncbi. nlm. nih. gov/Blast. cgi) was used to identify homologs of B. abortus EipB in different taxa within the Alphaproteobacteria.
The EipB weblogo was generated by aligning 447 DUF1849 protein sequences of Alphaproteobacteria retrieved from the EMBL-EBI website (https://www. ebi. ac. uk/interpro/entry/IPR015000/proteins -matched). Alignment was generated with Clustal Omega (https://www. ebi. ac. uk/Tools/msa/clustalo/). When necessary, the C termini of sequences were realigned by hand. The Clustal alignment file was converted to a fasta file using http://sequenceconversion. bugaco. com/converter/biology/sequences/ clustal_to_fasta. php. This file was then submitted to the skylign server (http://skylign. org/) to generate a weblogo. The alignment was processed with the following options: remove mostly empty columns/ alignment sequences are full length/score.
0001) and 8 (****, P Ͻ 0. 0001) weeks and had fewer CFU than spleens infected with the wild type at 4 (****, P Ͻ 0. 0001) and 8 (***, P Ͻ 0. 0007) weeks. (C to F) Antibody quantification in mouse sera harvested at 8 weeks postinfection from naive control mice or from mice infected with the wild-type, ΔeipB, or complementation strain. The amounts of total IgG (C), Brucella-specific IgG (D), IgG1 (E), and IgG2a (F) at 8 weeks were determined by ELISA. Each data point (naive, n ϭ 3; WT, n ϭ 2, ΔeipB; complementation, n ϭ 4) indicates the mean Ϯ the standard error of the mean.
ACKNOWLEDGMENTS We thank the members of the  Crosson laboratory  for helpful discussions. We thank members of the  SBC at Argonne National Laboratory  for their help with data collection at the 19-ID beamline. This study was supported by  National Institutes of Health  grants  U19AI107792  and  R01AI107159  to S. C. J. H. , J. W. W. , and S. C. contributed to the design and conceptualization of the study. J. H. , J. W. W. , A. F. , D. M. C. , J. X. C. , E. U. , A. B. , L. B. , G. B. , Y. K. , and S. C. performed the experiments and acquired and analyzed the data. J. H. , J. W. W. , A. F. , and S. C. interpreted the data. J. H. and S. C. wrote the original draft of the manuscript.
Data availability. For each Himar insertion library, Tn-seq read data have been deposited in the NCBI Sequence Read Archive under the following accession numbers: B. abortus 2308 wild type, SRR7943723 (BioProject PRJNA493942); B. abortus ΔeipB strain (Δbab1_1186), SRR8322167 (BioProject PRJNA510139). The coordinates of EipB have been deposited in the Protein Data Bank under 6NTR. Crystallographic data and refined model statistics are presented in Table  S2. Diffraction images have been uploaded to the SBGrid diffraction data server (data doi:10. 15785/SBGRID/445).
One-way analysis of variance (ANOVA), followed by Dunnett's posttest (to wild type), supports the conclusion that spleens infected with the eipB deletion strain were significantly smaller at 4 (****, P Ͻ 0. 0001) and 8 (****, P Ͻ 0. 0001) weeks and had fewer CFU than spleens infected with the wild type at 4 (****, P Ͻ 0. 0001) and 8 (***, P Ͻ 0. 0007) weeks. (C to F) Antibody quantification in mouse sera harvested at 8 weeks postinfection from naive control mice or from mice infected with the wild-type, ΔeipB, or complementation strain. The amounts of total IgG (C), Brucella-specific IgG (D), IgG1 (E), and IgG2a (F) at 8 weeks were determined by ELISA. Each data point (naive, n ϭ 3; WT, n ϭ 2, ΔeipB; complementation, n ϭ 4) indicates the mean Ϯ the standard error of the mean.
Virtual world and acquisition of views The virtual environment used in our model was generated by the software Habitat3D  [102] , an open-source tool which provides photorealistic meshes of natural scenes from point clouds acquired with help of a LiDAR scanner (IMAGER 5006i). This environment is mapped on the habitat of Myrmecia ants from Canberra, Australia  [69]. The mesh spans a diameter of around 65 metres and features large eucalyptus trees and distant panorama cues. This dataset can be found on the Insect Vision webpage (https://insectvision. dlr. de/3d-reconstruction-tools/ habitat3d). For speed optimization purposes, we down-sampled the originally high-resolution mesh with the open-source software Blender into a lower number of vertices; the rendering was then realized in OpenGL, with the Python libraries Plyfile and PyOpenGL.
If the CHA was fully effective, the head had empty florets where seeds failed to form due to complete sterilization. If seeds were present, it was assumed that the CHA had not been completely effective and the seed formed from self-pollination. Due to weather conditions, particularly the strong winds of the Great Plains, some bags were lost and those observations affected the balance of the dataset. Entry Number Name Years Included Breeding Program of Chemical hybridizing agent and application. Plots were treated with Croisor 100 at a rate recommended by the manufacturer. ChemSurf 90 (active ingredients: glycerol, sodium xylene sulfonate, alkyl phenol ethoxylate) was used as a surfactant at a rate of 0. 3 ml m -2 (United Suppliers, Eldora, IA, USA). Water was added to the mix for a total spray volume of 30 ml m -2 and applied using a 3.
The cutoff value was selected because the mean number of total seeds on a healthy head of hard winter wheat in the Great Plains is around 30. Thus, seven seeds correspond to a sterility level of about 75% and that value would be acceptable for experimental hybrid seed production. The PROC GLIMMIX and DATA steps of SAS 9. 4 software (SAS Institute, Inc. , Cary, NC, USA) were used to evaluate logit and probit threshold models. The linear predictor was used as follows with a log link: η α α = + + b b ( ) ( 3 ) ijk i j ijk Where η ijk is the frequency of observing seven seeds or fewer for head (αb) ijk of genotype α i in block b j. The random b j and (αb) ijk effects were assumed independent and identically distributed (i. i. d. ) normally with a mean of zero and variances of σ b 2 and σ ab 2 , respectively.
Statistical count models. Counts of seed per bagged head for each genotype were analyzed by mixed effects analysis using PROC UNIVARIATE, PROC GLIMMIX, and PROC NLMIXED procedures in SAS 9. 4 software. Transformed response variables were incorporated into the dataset using a DATA step in SAS 9. 4 software to evaluate the efficacy of transformed data with a normal approximation (SAS Institute, Inc. , Cary, NC, USA). These transformations include logarithmic, square root, and exponential transformations as follows: = + log count seed count " " log ( 1) = + '' sqrt count s eed count " 3 /8 = exp count seedcount " " ( ) 2/3 Initial analyses were done by year, and estimates for seed count were modelled as in Eq. 3, this time letting α i be the estimated seed count for genotype i rather than a binary response.
Using the set parameters from which the data were simulated, the confidence intervals were tested for coverage-to see if the parameter was contained within the interval. Coverage was recorded as a binary variable, 1 for true and 0 for false, where 'false' indicated that the true mean was not in the 95% confidence interval. The mean coverage was calculated for each model tested across the 500 experiments in duplicate for a total of 1000 experiments. The empirical coverage proportion indicates how well the models tested are able to estimate the mean count for each genotype if the counts do arise from a zero-inflated or hurdle process.
The relevant fit statistics included the AICc, BIC, χ 2 /d. f. , and variance estimates. Models were selected based on lowest values for the AICc and BIC fit statistics. However, comparing fit statistics is only appropriate between similar models (i. e. Poisson with NB or hurdle models). Additionally, the transformed data sets give variance estimates on a scale not compatible with the GLMM models, making direct comparisons difficult. Comparing models within the 2015 dataset showed variation between the relevant fit statistics (Table  S5 ). Next the Pearson χ 2 /d. f. value was examined for the Poisson and negative binomial models. For the Poisson model, this value was 4. 48, indicating over-dispersion. For the negative binomial model, the observed value of 0. 98 could be considered acceptable as it is close to one  15.
Simulation experiments. If data such as these counts arise from a process with zero-inflation, it is helpful to know if the zero-inflated models can capture that process well. It is also critical to determine how much information and precision is lost when using models that do not account for the excess zeros. Thus, a dataset known or designed to be zero-inflated can be used to show how much is lost if one uses other modeling strategies. To demonstrate this, five hundred experiments were simulated two times from a zero-inflated process, and the models used to test the experimental crossing block data were used on each of the simulated datasets to monitor overall trends of fit. 95% confidence intervals for the count estimates were constructed on the same scale as the original data. Empirical coverage probability was calculated as the proportion of confidence intervals that included the true mean of the count for that genotype.
Discussion We collected seed count data to determine the frequency of sterility for CHA-treated wheat. The initial data indicated a mean of 5. 7 seeds head -1 in 2015 and 2. 6 seeds head -1 in 2016. For most wheat in the Great Plains, the average head of wheat has 30 seeds  27. Using these estimates, the combined sterility across the genotypes tested was greater than 80% in both years. Researchers in hybrid wheat have not yet set a rule regarding the proportions of sterility required for producing hybrid wheat seed for testing, but it is anticipated that it will be around 75%. Our target number of selfed seeds per head would therefore be seven seeds. As such, this was the cutoff point set for our logit and probit models. Certified hybrid seed for small grains produced using chemical hybridization must be 95% pure hybrid with a maximum of 5% of self-fertilized female parent seed  28.
Obtaining more detailed information directly relating to the estimated seed count then became more useful for 2015 and count-based models provided the best information from a biological perspective. The estimates of seed count from the count models can be used in downstream analyses to determine which genotypes are better suited to chemical hybridization if hybrid performance is similar. Using a number of statistical models, we measured estimated genotypic differences to CHA sterilization on the count of seeds directly. A standard approach in agronomic research to data that are not normally distributed is to apply a variance-stabilizing transformation to the data. The analysis then uses the transformed response variable and assumes it to be normally distributed  11. The data in this study showed evidence of over-dispersion, and after transformation the variance was not well-controlled.
Because the seed count data are collected the year prior to the planting of the hybrid seed, the estimates of seed count could be helpful in explaining differences in vigor or uniformity of the hybrid plots. For manufacturing process analysis, results from zero-inflated models were shown to be more conservative in their estimates than directly applying Poisson or Negative Binomial GLMMs, thus lowering Type I error  21. Thus it was unsurprising that confidence intervals for the estimates of seed counts from zero-inflated and hurdle models were wider than those from the Poisson and Negative Binomial models. This represents a decrease in precision of the estimates, but simulations indicated that use of the wrong model on data with excess zeros gave confidence intervals that did not include the true value of the parameter, and hence were less accurate.
Acknowledgements The authors wish to thank  Dr. Mengyuan Wang ,  Mr. Gregory Dorn , and  Mr. Mitch Montgomery  for technical assistance with this research. The corresponding author would also like to thank  Ms. Johnna Baller  and  Ms. Jessica Hauschild  for meaningful discussions on statistical programming aspects of this project. Graduate student support for A. C. Easterly was provided by the  Daugherty Water for Food Global Institute at the University of Nebraska. Partial funding for P. S. Baenziger is from  Hatch  Project  NEB-22-328 ; the  National Institute of Food and Agriculture  as part of the  International Wheat Yield Partnership, U. S. Department of Agriculture , under award number  2017-67007-25939 ; the  CERES Trust Organic Research Initiative ; and  USDA  under Agreement No. 59-0790-4-092 , a cooperative project with the  U.