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. 2020 Mar 23;88(4):e00027-20. doi: 10.1128/IAI.00027-20

FIG 7.

FIG 7

IFN-γ production and in vivo bioluminescence imaging of transgenic T. gondii ME49 expressing either T. gondii or N. caninum profilin. (A) Alignment of the T. gondii profilin predicted amino acid sequence with those from the near relatives N. caninum and Hammondia hammondi. Putative substrate-binding motifs identified previously by Kucera and colleagues (59) are outlined as follows: the red box indicates an acidic loop (AL), and the green box indicates a β-hairpin. Sequences were obtained from ToxoDB (H. hammondi HHA_293690, T. gondii TGME49_293690, and N. caninum NCLIV_000610). (B) Anti-HA immunofluorescence assay (IFA) of ME49:TgProfilinHA (Tg:TgProHA) (top row) or ME49:NcProfilinHA (Tg:NcProHA) (bottom row) showing the expected cytoplasmic localization. (C) Western blotting using anti-HA (top blot) or anti-SAG1 (bottom blot) as a loading control. Arrows indicate sizes in kilodaltons. (D to F) Six- to eight-week-old BALB/c mice were infected with 106 tachyzoites (n = 3 per parasite strain) or injected with soluble tachyzoite antigen (STAg) preparations equivalent to 106 tachyzoites of the indicated parasite strains (n = 3 per parasite strain). (D) Quantification of bioluminescence imaging (total flux [photons per second]) over the time course of infection of mice with Tg:TgProHA or Tg:NcProHA. Data were log transformed, and two-way repeated-measures ANOVA (alpha value of 0.05) with Sidak’s multiple-comparison test was performed (analyses were performed on days 0 to 5). (E) IFN-γ ELISA of samples collected over the course of infection with Tg:TgProHA or Tg:NcProHA. Two-way repeated-measures ANOVA (alpha value of 0.05) and Sidak’s multiple-comparison test were performed. (F) IFN-γ ELISA of samples collected after injection of Tg:TgProHA STAg or Tg:NcProHA STAg. Two-way repeated-measures ANOVA (alpha value of 0.05) with Sidak’s multiple-comparison test was performed (analysis for 0 to 48 h). No statistically significant differences were identified between mice injected with Tg:TgProHA STAg and those injected with Tg:NcProHA STAg. *, P < 0.05; **, P <0.01.