TCRs from additional LTBI cohorts enhance the ability of GLIPH2 to distinguish Mtb-specific clonotypes. (A and B) Pie charts comparing percentage (and number) of (A) GLIPH2 groups or (B) unique TCRβs linked to a response to Mtb-infected macrophages (green), MTB300 only (blue), or lysate only (yellow). (C) Bar graph of GLIPH2 groups estimated to be Mtb-specific (x axis) containing TCRs contributed by ≥3 participants, rank-ordered by the sum of the highest number of TCR copies recovered in an experiment (y axis). Responses to infected macrophages (green), MTB300 only (blue), both (blue stripes), or lysate only (yellow) are indicated. GLIPH2 groups containing TCRs previously annotated as Mtb antigen–specific in IEDB are in red font; % indicates any amino acid substitution. (D and E) Pie charts comparing (D) total or (E) percentage of total CD4+ T cell TCRβs sequenced from 10 × 106 unstimulated PBMCs after cross-referencing GLIPH2 groups from Cleveland participants’ responses to infected macrophages (± MTB300 or lysate) to determine natural circulating frequency. (F) Bar graph of the mean number of each TCRβ clonotypes (different symbols) sequenced from unstimulated PBMCs (for Cleveland participants only), corresponding to each GLIPH2 group. (G) Bar graphs of the number (and percentage) of unique TCRs within GLIPH2 groups estimated to be Mtb-specific or non–Mtb-specific bystander GLIPH2 groups that contain either 100% (left) or 97–99% (right) CDR3α homology with MAIT cells. (H) Comparison of TCRdist3 meta-clonotypes generated from all TCRs in respective GLIPH2 groups estimated to be Mtb-specific in C. Thickness of each line corresponds to the number of TCR sequences across all samples. For meta-clonotypes, “.” indicates any amino acid substitution (analogous to % in GLIPH2), “?” indicates an optional amino acid, and brackets indicate an either–or substitution. (I) Chi-square statistics comparing membership for all TCRs (All) or only the GLIPH2 groups in C (GLIPH2 [Mtb]). (J) Proportional Venn diagram showing the number of unique TCRs clustered by GLIPH2 or TCRdist3 within groups (or meta-clonotypes) with TCRs from ≥2 participants. The green circle represents TCRs in GLIPH2 groups estimated to be Mtb-specific from C. TCRs grouped by GLIPH2 and TCRdist3 were compared using a chi-square analysis and Cramer’s V post hoc test.
Figure 4.

TCRs from additional LTBI cohorts enhance the ability of GLIPH2 to distinguish Mtb-specific clonotypes. (A and B) Pie charts comparing percentage (and number) of (A) GLIPH2 groups or (B) unique TCRβs linked to a response to Mtb-infected macrophages (green), MTB300 only (blue), or lysate only (yellow). (C) Bar graph of GLIPH2 groups estimated to be Mtb-specific (x axis) containing TCRs contributed by ≥3 participants, rank-ordered by the sum of the highest number of TCR copies recovered in an experiment (y axis). Responses to infected macrophages (green), MTB300 only (blue), both (blue stripes), or lysate only (yellow) are indicated. GLIPH2 groups containing TCRs previously annotated as Mtb antigen–specific in IEDB are in red font; % indicates any amino acid substitution. (D and E) Pie charts comparing (D) total or (E) percentage of total CD4+ T cell TCRβs sequenced from 10 × 106 unstimulated PBMCs after cross-referencing GLIPH2 groups from Cleveland participants’ responses to infected macrophages (± MTB300 or lysate) to determine natural circulating frequency. (F) Bar graph of the mean number of each TCRβ clonotypes (different symbols) sequenced from unstimulated PBMCs (for Cleveland participants only), corresponding to each GLIPH2 group. (G) Bar graphs of the number (and percentage) of unique TCRs within GLIPH2 groups estimated to be Mtb-specific or non–Mtb-specific bystander GLIPH2 groups that contain either 100% (left) or 97–99% (right) CDR3α homology with MAIT cells. (H) Comparison of TCRdist3 meta-clonotypes generated from all TCRs in respective GLIPH2 groups estimated to be Mtb-specific in C. Thickness of each line corresponds to the number of TCR sequences across all samples. For meta-clonotypes, “.” indicates any amino acid substitution (analogous to % in GLIPH2), “?” indicates an optional amino acid, and brackets indicate an either–or substitution. (I) Chi-square statistics comparing membership for all TCRs (All) or only the GLIPH2 groups in C (GLIPH2 [Mtb]). (J) Proportional Venn diagram showing the number of unique TCRs clustered by GLIPH2 or TCRdist3 within groups (or meta-clonotypes) with TCRs from ≥2 participants. The green circle represents TCRs in GLIPH2 groups estimated to be Mtb-specific from C. TCRs grouped by GLIPH2 and TCRdist3 were compared using a chi-square analysis and Cramer’s V post hoc test.

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