Fig. 5: Restoration of stumpy formation in T. b. gambiense.
a In vivo infectivity of T. b. gambiense LiTat 1.3 (T. b. gambiense; green line) and T. b. gambiense expressing T. b. b. HpHbR (T. b. g. + b18S; red line) and T.b. g. HpHbR (T. b. g. + g18S; black line) from the 18S rRNA locus was evaluated by infecting mice with 3 × 106 cells (n = 3). The parasitemia was counted daily till day 6 when the experiment was terminated. b Indirect immunofluorescence with PAD1 antibody (red), specific for BS-ST in cell lines described in (a). DNA in the nucleus (N) and kinetoplast (k) was stained with DAPI (blue). Scale bar, 5 μm. c Quantification of PAD1-positive and PAD1-negative cells (ex vivo), day 4 p.i.) in cell lines described in (a) (n = 3). In the columns, the stumpy form is shown in red. d Morphological characterization of T. b. gambiense and T. b. g. + b18S cell lines containing the PAD1-negative cells (black dots) or PAD1-positive cells (red dots). Left panel: the distance between the nucleus (N) and the kinetoplast (k). T. b. gambiense (n = 30 cells), T. b. g. + b18S (n = 25 cells); Right panel: cell area. T. b. gambiense (n = 30 cells), T. b. g. + b18S (n = 25 cells). Results were analyzed using the Mann–Whitney test (ns, p < 0.05; ****, p < 0.0001). e In vivo infectivity of T. b. gambiense Bosendja strain. The parasitemia was counted daily till day 4, when the experiment was terminated (n = 3). f Immunofluorescence using PAD1 antibody, revealing a fraction of the PAD1-positive cells. DNA in the nucleus (N) and kinetoplast (k) was stained with DAPI (blue). Scale bar, 5 μm. g Quantification of PAD1-positive and PAD1-negative cells in the T. b. gambiense Bosendja strain ex vivo days 2-4 p.i. h Western blot analysis of WT T. b. brucei and ABbin situ, as well as T. b. gambiense Bosendja strain with PAD1 antibody. Enolase antibody was used as a loading control. Error bars indicate ±SD. Source data are provided as a Source Data file.
