Toltrazuril and diclazuril: comparative evaluation of anti-coccidial drugs using a murine model

Abstract

Intestinal coccidiosis caused by Eimeria protozoan species is an economically important disease, especially in poultry and cattle. Anti-coccidial drugs commonly used for controlling coccidiosis are toltrazuril (TTZ) and diclazuril (DCZ). In this study, the efficacies of TTZ and DCZ were compared using a murine model, and the effect of these treatments on the induction of acquired resistance was evaluated. Male C57BL/6J mice were inoculated with 1,000 sporulated E. vermiformis oocytes and treated with TTZ or DCZ. The recommended TTZ dose for cattle (15 mg/kg) completely prevented oocyte excretion. But, mice required 5 mg/kg of DCZ, which is five times the recommended dose for cattle, to reduce oocyte excretion. In E. vermiformis re-infection, TTZ (15 mg/kg) and DCZ (5 mg/kg) treatments did not interfere with the development of acquired resistance. Bodyweight gain was significantly higher in the TTZ-treated group than in the control (untreated/infected) group and the DCZ-treated group, and no significant difference in bodyweight gain was observed between the TTZ-treated group and the healthy (uninfected/untreated) group. Analysis of T lymphocyte subsets in the spleen and mesenteric lymph nodes indicated that the relative populations of CD4⁺ and CD8⁺ T cells were reduced in the DCZ-treated and control (untreated/infected) groups, suggesting there was immunosuppression during the infection. However, no reductions in T cell populations were observed in the TTZ-treated group. The results indicated that an optimal anti-coccidial drug is one that can completely break the parasite life cycle in the host animal.

Keywords: Eimeria vermiformis; body-weight gain; diclazuril; host immunity; toltrazuril.

Fig. 1.

Effect of toltrazuril (TTZ) and diclazuril (DCZ) treatment on oocytes excreted per day (OPD) in Eimeria vermiformis-infected mice. Mice were orally inoculated with 1,000 sporulated oocytes of E. vermiformis on day 0 and orally treated with several doses of TTZ (a) and DCZ (b) on day 3. Oocyte count and identification were performed using a modified McMaster’s method. Results are shown as mean ± SEM, *P<0.05, n=5/group.

Fig. 2.

Effect of toltrazuril (TTZ) and diclazuril (DCZ) treatment against Eimeria vermiformis re-infection. Mice were orally inoculated twice with 1,000 sporulated oocytes of E. vermiformis on days 0 and 21 and orally treated with TTZ at 15 mg/kg and DCZ at 5 mg/kg on day 3. The single-infection group was infected once with 1,000 sporulated E. vermiformis oocytes. Oocyte count and identification were performed using a modified McMaster’s method. Results are shown as mean ± SEM, *P<0.05, n=5/group.

Fig. 3.

Effect of toltrazuril (TTZ) and diclazuril (DCZ) treatment on body weight (BW) gain in Eimeria vermiformis-infected mice. Mice were orally inoculated twice with 1,000 sporulated oocytes of E. vermiformis on days 0 and 21 and orally treated with TTZ at 15 mg/kg and DCZ at 5 mg/kg on day 3. BW gain in each group was assessed during the whole period of infection. The mean percentage BW gain in the healthy (uninfected/untreated) control was considered to be 100%. Results are shown as mean ± SEM, *P<0.05, n=5/group.

Fig. 4.

Effect of toltrazuril (TTZ) and diclazuril (DCZ) treatment on splenic and mesenteric lymph node T cell subsets in Eimeria vermiformis-infected mice. Mice were inoculated twice with 1,000 sporulated E. vermiformis oocytes on days 0 and 21 and orally treated with TTZ at 15 mg/kg and DCZ at 5 mg/kg on day 3. Mice were sacrificed on days 10, 21, 24, and 28, and spleens and MLNs were harvested. The populations of CD4⁺ (a) and CD8⁺ cells (b) in the spleen and the populations of CD4⁺ (c) and CD8⁺ cells (d) in MLNs were analyzed by flow cytometry. Results are shown as mean ± SEM, *P<0.05, n=4/group.