| Chicken coccidiosis is an important animal parasitic disease with global distribution with Eimeria tenella being the most pathogenic species.Global economic loss caused by chicken coccidiosis was up to 10.04 billion pound(about 89.8 billion RMB)in 2020.Currently,prevention and control of coccidiosis still relies on drugs and traditional live oocyst vaccines,but there are obvious problems such as serious resistance to commonly used anticoccidial drugs and the risk of potential outbreaks of chicken coccidiosis after vaccination with traditional chicken coccoidal vaccines.Eimeria tenella secretes a series of proteins when invading host cells,They play an important role in parasite invasion and survival and may be potential vaccine candidates.In this study,we investigated whether Et ROP21 has secretion properties and whether it has potential values as a candidate vaccine antigen protein.The details of the study are as follows.In this study,we amplified and cloned the Et ROP21 gene from the cotyledon c DNA of Eimeria tenella and further constructed the prokaryotic expression vector PET32a-Et ROP21,and expressed Et ROP21 protein using E.tenella.In order to evaluate the immunoprotective effect of ROP21 protein,three groups were set up in this study for chicken attack test: immunized group(immune attack group),positive group(non-immune attack group)and negative group(non-immune non-attack group).One week after the three immunizations,all chicks in the immunized and positive groups were given 20 million E.tenella sporulated oocysts orally.The data of weight gain,mortality,and oocyst discharge were monitored during the attack period,and postattack dissection was performed to observe the cecum lesions.Finally,the comprehensive data analysis was performed to evaluate the immunoprotective effect of ROP21 recombinant protein.The results showed that the survival rate after challenge was 100% after immunization with p ET32a(+)-Et ROP21 subunit vaccine.Weight gain after immune challenge was comparable to the non-immune/non-challenge group and slightly higher in the non-immune/non-challenge group(P<0.05);The oocyst discharge in the immune group was smaller than that of the positive group with no significant difference(P>0.05);The cecum lesion count score of the immune group was less severe than that of the positive group(P<0.05);Finally,the ACI(anticoccidial index)of the immune group was 163.23.The Et ROP21 protein subunit vaccine was able to provide partial immune protection against E.tenella.Western-blot results on chicken sera showed that ROP21 recombinant protein immunization resulted in specific antibodies in chickens,which were immunogenic at dilutions in the range of 1:100-12800 as measured by ELISA. |
