| Rabbit coccidiosis,caused by infection with parasites of the genus Eimeria has greatly affected the rabbit industry.For control of coccidiosis,immunoprophylaxis with live oocyst vaccines,as those being extensively and successfully used in poultry industry,is the most promising strategy.Unfortunately,there is no live oocyst vaccine available for rabbit coccidiosis.Thus,in our present work,five original strains and 2 precocious lines of rabbit coccidia were thoroughly investigated for their vaccine-related properties.Moreover,laboratory trials of a triple-combination vaccine against rabbit coccidiosis was completed.First,the original strains of E.magna,E.media,E.flavescens,E.intestinalis and E.irresidua,and the precocious lines of E.media and E.intestinalis were investigated for their identification,pathogenicity,immunogenicity and anticoccidial drug sensitivity.We confirmed the identification criteria for morphological identification and single oocyst PCR for the 5 original strains.All 5 original strains possess various pathogenicity and can be applied as testing strains.All original stains and precocious lines except for E.flavescens were of great immunogenicity.All three candidate strains did not have resistance to common anticoccidial drugs.Above all,we demonstrated that the precocious lines of E.intestinalis and E.media and the original strain of E.irresidua can be further studied as the components of the rabbit coccidiosis vaccine.Second,we conducted laboratory trials of a triple-combination vaccine of rabbit coccidiosis.Based on obtained data on optimal immunization dosages of the 3 components,we conducted a co-immunization assay to verify any cross-influences in establishing immunity.Results showed that immunization with precocious lines didn’t cause interference while the original strains affected the establishment of immunity of the precocious lines.Thus,we confirmed the optimal dosage of each component:1 × 103 oocysts of E.intestinalis precocious line,1 × 103 oocysts of E.media precocious line,and 3×102 oocysts of E.irresidua original strain.In subsequent experiments,we determined the optimal vaccination days of age and immunity duration.Results showed that vaccination performed on 30-35 days old weaning rabbits provided efficient and long-lasting immunity till 60 days post immunization.On this basis,we conducted field trials on meat and rex rabbits.It was well demonstrated that vaccination with the formulated live oocyst vaccine protected the rabbits against massive re-infection in terms of excellent growth and feed conversion.In addition,we exploited to find immunodominat antigens of rabbit coccidia.By using degenerated primers and genome walking analysis,we acquired the E.magna profilin gene.EmagPRF was a sectretory protein and was continuously expressed in sporozoites and schizogony stages.Immunization with recombinant EmagPRF provided partial immunity against challenge with E.magna oocysts.More importantly,ex vivo stimulation using EmagPRF elevated relative transcriptional levels of IFN-γ and IL-2 of spleen and mesenteric lymph node of rabbits irmmunized with E.magna oocysts,strongly suggesting that the very antigen had grest potential to be applied as a testing molecule of cellular immunity induced by rabbit coccidia.In conclusion,in this study,we systematically studied the vaccine-related characteristics of 5 original strains and 2 precocious lines of rabbit coccidia.We,for the first time,conducted laboratory trials of a triple-combination vaccine of rabbit coccidiosis which was proved to be safe and efficient in subsequent field trials.Our study provided ground knowledge regarding rabbit coccidia and encourages further development of live oocyst vaccine of rabbit coccidiosis. |
