Additional file 2 of Detoxified synthetic bacterial membrane vesicles as a vaccine platform against bacteria and SARS-CoV-2
Bibliographic record
Abstract
Additional file 1: figure S1: Schematic overview of OMV and SyBV isolation from cultured bacteria. Figure S2. Characterization of RNA isolated from OMV and SyBV. Representative electropherograms of RNA molecules isolated from SyBV in comparison to those from OMV. The filled triangle indicates internal marker. Figure S3. Quantification of DNA isolated from OMV and SyBV (n = 3 independent samples). Data are presented as the mean ± s.e.m. ns, not significant by unpaired two-tailed Student’s t-test. Figure S4. Comparison of protein composition between P. aeruginosa OMV and SyBV. (a) Principle component analysis of OMV and SyBV proteome. Two biological replicates per sample and three technical replicates for each biological replicate. (b) Partial view of heatmap of the hierarchical clustering based on the relative proteins abundance of OMV and SyBV proteome. Color code shows the normalized median abundance of proteins belonging to the category (red, most abundant; blue, least abundant). Figure S5. SyBV can be efficiently taken up by macrophages comparable to OMV. DiO-labelled vesicles (1 × 109) were incubated with MH-S cells for 6 h. And, the uptake of SyBV by cells was compared with OMV by flow cytometry, and the results are shown as the percentage of DiO-positive cells (n = 3). Data are presented as the mean ± SEM. ***P < 0.001; ns, not significant, by one-way ANOVA with Tukey’s post test. Figure S6. Measurement of body temperature during immunization with P. aeruginosa OMV or SyBV. The temperature of mice was investigated at 24 h after each immunization (n = 5). Data are presented as mean ± s.e.m. ns, not significant; by one-way ANOVA with Tukey’s post test versus the sham group. Figure S7. Differential cell count and chemokine level in BAL fluid of mice immunized with P. aeruginosa OMV or SyBV. (a) The effect of OMV and SyBV on the numbers of BAL cells (macrophage, neutrophil and lymphocyte) was investigated at 48 h after the last challenge (n = 5). (b) The level of neutrophil-chemoattractant chemokine (KC) in BAL fluid was evaluated at 48 h after the last challenge (n = 5). Data are presented as mean ± s.e.m. ***P < 0.001; ns, not significant; by two-way ANOVA with Tukey’s post test versus the sham group. Figure S8. The level of P. aeruginosa-specific CD4+ T-cell-derived IL-4 after CD4+ T-cells were isolated from immunized spleens (three independent samples). All data are presented as the mean ± s.e.m. ns, not significant by two-way ANOVA with Tukey’s post-test versus the sham group. Figure S9. Survival curve of mice immunized with heat-inactivated SyBV from E. coli. The result was monitored for 5 days after intraperitoneal challenge with lethal dose of E. coli (n = 10). For survival curve, log-rank (Mantel-Cox) test was used to compare with sham group (*P < 0.05). Figure S10. Quantification of germinal center B cells. Graph shows cell counts per germinal center cross-section in the mice spleens immunized with P. aeruginosa or E. coli SyBV. Data are presented as mean ± s.e.m. ***P < 0.001 by one-way ANOVA with Tukey’s post test. Figure S11. The level of S1-specific CD4+ T-cell-derived IL-4 after CD4+ T-cells were isolated from immunized spleens (three independent samples). All data are presented as the mean ± s.e.m. ns, not significant by two-way ANOVA with Tukey’s post-test versus the sham group. Figure S12. Western blot analysis of IPTG-induced OMVS1 and SyBVS1 with anti-His Tag antibody. The filled triangle indicates the Lpp-OmpA-S1 complex. Table S1. Immunogenic outer membrane protein markers expressed in SyBV in comparison to OMV.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.002 | 0.025 |
| Meta-epidemiology (narrow) | 0.002 | 0.001 |
| Meta-epidemiology (broad) | 0.002 | 0.001 |
| Bibliometrics | 0.002 | 0.003 |
| Science and technology studies | 0.001 | 0.000 |
| Scholarly communication | 0.003 | 0.002 |
| Open science | 0.002 | 0.002 |
| Research integrity | 0.002 | 0.001 |
| Insufficient payload (model declined to judge) | 0.876 | 0.182 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".