In this study three Toxoplasma strains were used. The transgenic T. gondii β-Gal strain derived from T. gondii RH (Type I) constitutively expresses β-galactosidase was kindly provided by David Sibley, Washington University, St. Louis, MO, USA, and the T. gondii Me49 (a type II strain, kindly provided by Furio Spano, Istituto Superiore di Sanità, Rome) were both maintained by serial passaging in human foreskin fibroblast monolayer cultures in Dulbecco's modified Eagles's medium (DMEM) with 10% fetal calf serum (FCS) and penicillin/streptomycin as previously described (Winzer et al., 2015 (link)). Oocysts of the T. gondii ShSp1 strain (TgShSp1 (type II) were kindly provided by Complutense University of Madrid (Imhof et al., 2021 (link); Sánchez-Sánchez et al., 2018 (link)) and stored at 4 °C until used for experimental infection of CD1 outbred mice.
Compounds used in this study are shown in Fig. 1 and include (A) Sulfadoxine (SDX); (B) the di-ruthenium complex (RU); (C) the hybrid RU-SDX conjugate (RU-SDX); (D) the hybrid RU complex-9-(2-hydroxyethyl)-adenine (RU-adenine). All compounds were synthesized as previously described (Desiatkina et al., 2021 (link); Păunescu et al., 2021 (link)). Pyrimethamine (PYR) and the two mitochondrial uncouplers carbonylcyanide-trifluoromethoxyphenylhydrazone (FCCP) and carbonylcyanide-3-chlorophenylhydrazone (CCCP) were purchased from Merck (Darmstadt, Germany). For RU-based drugs and PYR, stock solutions were prepared at 1 mM in dimethylsulfoxide (DMSO) and were stored at −20 °C.
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