Effects of diclazuril on the expression of enolase in second-generation merozoites of Eimeria tenella

Abstract

Eimeria tenella is an obligate intracellular parasite of the chicken cecum; it brings huge economic loss to the chicken industry. Enolase is a multifunctional glycolytic enzyme involved in many processes of parasites, such as infection and migration. In this study, the effect of diclazuril on the expression of enolase in second-generation merozoites of E. tenella ( Et ENO ) was reported. The prokaryotic expression plasmid pET-28a- Et ENO was constructed and transformed into Escherichia coli BL21 (DE3). Then, it was subjected to expression under the induction of isopropyl-β-D-1-thiogalactopyranoside. The expressed products were identified and purified. The purified Et ENO protein was used for antibody preparation. The Et ENO mRNA and protein expression levels were analyzed via real-time PCR and Western blotting. Localization of Et ENO on the merozoites was examined by immunofluorescence technique. The mRNA and protein expression levels of Et ENO were decreased by 36.3 and 40.36%, respectively, by diclazuril treatment. Et ENO distributed in the surface, cytoplasm, and nucleus of the infected/control group. With diclazuril treatment, it was significantly reduced in the surface and cytoplasm and even disappeared in the nucleus of the infected/diclazuril group. These observations suggested that Et ENO may play an important role in mechanism of diclazuril anticoccidial action and be a potential drug target for the intervention with E. tenella infection.

Diclazuril-induced expression of CDK-related kinase 2 in the second-generation merozoites of Eimeria tenella

“…Diclazuril, a classic anticoccidial drug, signi cantly induces merozoite apoptosis, effectively decreases the number of merozoites, and alleviates cecum damage induced by E. tenella [19][20][21][22][23][24]. Here, we reported changes in EtCRK2 in response to diclazuril treatment against E. tenella infection and observed the expression levels of EtCRK2 mRNA and protein and the spatial location of EtCRK2.…”

“…At 120 h postinoculation, chickens were killed through CO 2 asphyxiation, and the pooled caecal tissues from 10 randomly selected chickens in each replicate were used for the preparation of the second-generation merozoites in accordance with a previously described method [19][20][21]. Brie y, caecal tissues were cut into pieces, then incubated with enzyme digestion solution (0.12 mol/L NaCl, 3.0 mmol/L K 2 HPO 4 •3H 2 O, 9 mmol/L CaCl 2 , 1 mg/mL hyaluronidase, 1 mg/mL BSA and 0.02 mol/L Tris; pH = 7.4) at 37 °C for 60 min.…”

Nano-encapsulation of halofuginone hydrobromide enhances anticoccidial activity against Eimeria tenella in chickens

“…Second-generation merozoites (SGM) of E. tenella developed deep inside the lamina propria and musculature, and the merozoite stage caused the most deadly harm to intestinal mucosa and the overall physiological function of infected chickens. 40 In this study, changes in merozoite morphology and numbers after HTPM and HF treatments were investigated. As shown in Fig.…”

Influence of Qingchang Oral Liquid on Second Generation Merozoite of the Chicken Eimeria tenella

“…According to the life history of E. tenella, the damage of E. tenella mainly occurs in the fission stage where the schizont expand repeatedly and the mature schizont releases a large number of merozoites which invade and damage the intestinal epithelial cells. It resulted in the excretion of bloody flux, destruction of intestinal mucosa tissue, digestive disorder, anemia, emaciation and other clinical symptoms (Matsubayashi et al, 2019;Zhou et al, 2020). The fission stage is the most serious hazard developmental stage, which mainly occurs 3-5 days after infection.…”

“…The cecum was taken according to the literature method (Zhou et al, 2020) and the cecum contents were washed with PBS solution containing 2000U penicillin streptomycin double antibody. The small pieces of cecum were placed in a 1L beaker, and 10 times of the volume of hyaluronidase digestion solution was added.…”