Sulfadiazine resistance in Toxoplasma gondii: transcriptomic approach

Alexandre Mzabi a,b, Sandie Escotte-Binet a,b, Dominique Aubert a,b, Isabelle Villena a,b a.Laboratory of Parasitology-Mycology, EA 3800, SFR CAP-Santé - University of Reims Champagne-Ardenne - France b.Laboratory of Parasitology-Mycology, National Reference Centre for Toxoplasmosis, Biological Resource Centre Toxoplasma - Reims University Hospital - France

often prescribed active associations are Background pyrimethamine-(sulfadiazine or sulfadoxine), and trimethoprim-sulfamethoxazole. These Toxoplasma gondii, the causative agent of associations (except the pyrimethamine- toxoplasmosis, is a protozoan parasite that is sulfadoxine association) are used for the estimated to infect more than one-third of the treatment of severe toxoplasmosis in world's human population. Its life cycle immunocompromised patients. The comprises a series of five developmental pyrimethamine-(sulfadiazine or sulfadoxine) stages in the intestinal epithelium of the feline association is used in congenital definitive host and three stages relevant to toxoplasmosis (prenatal and postnatal Fig. 1. Multiple alignment of folate-biopterin transporters (FBT) relevant to this work infection in intermediate hosts which include treatment). As indicated above, in order to The depth of shading represents degrees of similarity. A (*) symbol indicates residues identical in all all other species of mammals. These stages limit adverse haematological events, these sequences. The signature sequence for the FBT family (black box) has been identified in all sequences. are sporozoites (contained in oocysts), and treatments are administered with folinic acid. the obligate intracellular stages tachyzoites, In clinical practice, therapeutic failures and and bradyzoites. relapses have been observed. The The principal drugs known as active understanding of the failure mechanisms against T. gondii are divided into two major against the main active drugs on T. gondii is groups: the inhibitors of folic acid synthesis essential because there are currently few and the macrolides. These drugs are effective and validated therapeutic primarily active against the tachyzoites and alternatives. Considering a drug’s action have no effect on cysts. mechanism, and in analogy to the other The inhibitors of folic acid synthesis include protozoa, the existence of a sensitivity dihydrofolate reductase (DHFR) inhibitors change for drugs and/or the development of (pyrimethamine and trimethoprim), and drug resistance could be feared. dihydropteroate synthase (DHPS) inhibitors (sulfonamides). These two anti-Toxoplasma Objectives Fig. 2. Expression levels of FBT genes in Toxoplasma gondii drugs are used in combination because of a synergistic effect on two key enzymes of folic In order to better understand the Results - Discussion parasite, by the internalization and the acid metabolism, DHPS and DHFR. These mechanisms of resistance to sulfadiazine, we sequestration of folate, and the efflux of drugs act together by inhibiting the folic acid have analysed the expression levels of 7 The transcriptomic analysis of various T. sulfadiazine. Furthermore, the potential synthesis in T. gondii, but also that of its host. folate transporter genes [1] (Fig. 1), 11 ABC gondii strains has highlighted in resistant involvement of folate transporters in drug As a consequence, they have a powerful transporter genes [2], and 14 enzyme genes strains an overexpression of folate resistance could, in the long term, raise the anti-parasitic effect (changes of purine which are involved in folic acid synthesis. transporter genes (Fig. 2), one ABC question about the systematic administration synthesis and parasitic division), which is This study by real time RT-PCR focused on transporter gene (data not shown), and 2 of folinic acid, which could potentially associated with adverse haematological sensitive strains (RH and ME-49), natural enzyme genes which are involved in folic acid enhance the parasite’s survival. events (neutropenia, thrombocytopenia). resistant strains (TgH 32006 and TgA synthesis (data not shown). Overexpression These drugs diffuse well into the organism 103001) [3], and induced resistant strains References of these genes would allow the survival of and cross the placental barrier. The most (RH-RSDZ and ME-49-RSDZ) [4]. [1] Massimine (2005) Mol. Biochem. Parasitol. 144, 44-54. [2] Sauvage (2004) Mol. Biochem. Parasitol. 134, 89-95. [3] Meneceur (2008) Antimicrob. Agents Chemother. 52, 1269-77. [4] Doliwa (2013) Experimental Parasitol. 133, 131-36.