CN110694058B - Rabbit coccidiosis live vaccine and preparation method and application thereof - Google Patents
Rabbit coccidiosis live vaccine and preparation method and application thereof Download PDFInfo
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Abstract
The invention relates to the technical field of animal coccidiosis vaccines, in particular to a rabbit coccidiosis live vaccine and a preparation method and application thereof. The invention provides a rabbit coccidiosis live vaccine composition, which comprises Eimeria macrotype, Eimeria intestinalis and Eimeria intermedia precocious attenuated strains and Eimeria perforated precocious attenuated strains. The rabbit coccidiosis live vaccine provided by the invention has higher immune protection efficacy and safety, can effectively prevent and treat rabbit coccidiosis, reduces or eliminates the use of anti-coccidiosis drugs, and simultaneously obviously reduces the adverse effects of vaccination on growth and production performances of rabbits such as weight gain and the like, thereby having good economic benefits and wide application prospects. The invention also provides a breeding method of the coccidian prematurity attenuated strain, the breeding method can effectively shorten the breeding cycle of the coccidian prematurity attenuated strain, improve the breeding efficiency of the coccidian prematurity attenuated strain, and the obtained prematurity insect strain has stable genetic characteristic, obviously reduced pathogenicity and good immunogenicity.
Description
Technical Field
The invention relates to the technical field of animal coccidiosis vaccines, in particular to a rabbit coccidiosis live vaccine and a preparation method and application thereof.
Background
Antibiotics and anticoccidial drugs have a long history as livestock and poultry feed additives, and are main means for guaranteeing the health of livestock and poultry in an intensive breeding mode. With the frequent occurrence of drug residues and drug-resistant strains (worms) and the threat to the environment and human health, the strategy of preventing and controlling animal epidemic diseases by using drugs gradually exposes the disadvantages. Outbreaks of coccidiosis in rabbits can cause massive rabbit death; chronic and subclinical coccidial infections lead to reduced animal productivity and reduced economic benefits. The continuous occurrence of coccidiosis is the main reason for the poor economic benefit and even loss of many rabbit farms. Therefore, under the dilemma that the use of anticoccidial drugs is limited and the drug resistance of coccidia frequently occurs, so that the prevention and control effects of the drugs are poor, the development of highly effective and safe rabbit coccidiosis vaccines is urgent.
The pathogeny of rabbit coccidiosis is 11 kinds of coccidiosis in Eimeria, which parasitizes on different parts of intestinal tract or liver, and the pathogenicity and prevalence rate of different kinds of coccidiosis are different, thus the harm to rabbit industry is different. Among them, Eimeria macrotype, Eimeria mesotype, Eimeria enterobacter and Eimeria punctata are the main species of infection and prevalence in rabbit farm, and the first three species have strong pathogenicity, cause severe diarrhea, and seriously affect productivity and economic benefits in rabbit industry. Therefore, the development of a rabbit coccidiosis vaccine must achieve two main goals: firstly, the pathogenic coccidian species can be well protected; secondly, the safety is high. The vaccine taking the live oocysts as the components can effectively resist the re-infection of the same coccidia and has great advantages in the prevention of coccidiosis of chickens and turkeys. Therefore, the live vaccine aiming at the epidemic coccidian species is developed for preventing the rabbit coccidiosis, the use of the anti-coccidian drug can be obviously reduced, the anti-coccidian drug is not used, and the food safety, the environment and the economic benefit are effectively improved, so that the application prospect is very wide.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide a rabbit coccidiosis attenuated live vaccine with excellent safety and immune protection efficacy and a method for efficiently and quickly breeding a coccidiosis praecox attenuated strain.
In order to achieve the purpose, the technical scheme of the invention is as follows:
although the live oocysts of the rabbit coccidia are used as the immunogen of the vaccine, the immune protective efficacy of the vaccine is more favorably improved, but the risk of adverse effects on the growth and production of animals is higher, so that the technical problem in the development process of the rabbit coccidia vaccine is how to improve the safety of the live vaccine while ensuring the immune protective efficacy of the rabbit coccidia vaccine. In the research of rabbit coccidian vaccines, highly pathogenic species are mostly utilized for inoculation to obtain the immunity of corresponding species, and low-pathogenic or basically nonpathogenic rabbit coccidian is usually ignored. Furthermore, in order to improve the preparation efficiency of the vaccine, the invention develops a rapid breeding method of the early-maturing attenuated insect strain on the basis of the traditional early-maturing pressure breeding method, effectively shortens the passage and screening period of the early-maturing attenuated insect strain, and obtains the early-maturing attenuated insect strain with stable genetic characteristic, high safety and good immunogenicity.
Specifically, the technical scheme of the invention is as follows:
the invention provides a rabbit coccidiosis live vaccine composition, which comprises at least 3 precocious attenuated strains of highly pathogenic rabbit Eimeria and low pathogenic rabbit Eimeria; the low-pathogenicity rabbit coccidia is a wild insect strain and/or a prematurity attenuated insect strain.
Preferably, in the rabbit coccidiosis live vaccine composition, the ratio of the number of oocysts of the precocious attenuated strain of the highly pathogenic rabbit eimeria and the low pathogenic rabbit eimeria is 1: (2-20). Preferably 1: (5-20). The content proportion in the range is compounded for use, so that the safety and the immune protection efficacy of the vaccine can be better ensured at the same time.
Preferably, the low-pathogenicity rabbit coccidia adopts a precocious attenuated strain.
Preferably, the low pathogenic rabbit eimeria comprises one or more of eimeria minutissima, eimeria perforata, eimeria vimentis.
Among the numerous highly pathogenic and less pathogenic rabbit coccidia, the present inventors have found that the perforated Eimeria is most effective in improving the safety of the large Eimeria, intestinal Eimeria and intermediate Eimeria component vaccines. Thus, in the rabbit coccidiosis live vaccine composition, the highly pathogenic prawned attenuated strains of rabbit eimeria coccidia preferably include prawned attenuated strains of eimeria macrolepia, eimeria intestinalis, and eimeria mellifera; the low pathogenic E.rabbit preferably comprises perforated E..
On the basis of meeting the content ratio of the components, in order to better ensure the safety and the immune protection efficacy of the vaccine at the same time, the rabbit coccidiosis live vaccine composition preferably comprises the following components: 100-3000 oocysts per head part of precocious attenuated strain of large Eimeria, 500-5000 oocysts per head part of precocious attenuated strain of intestinal Eimeria, 500-6000 oocysts per head part of precocious attenuated strain of medium Eimeria, and 1500-40000 oocysts per head part of wild Eimeria strain or precocious attenuated strain of perforated Eimeria.
More preferably, the rabbit coccidiosis live vaccine composition comprises the following components: 500-3000 oocysts per head part of precocious attenuated strain of large Eimeria, 1000-5000 oocysts per head part of precocious attenuated strain of intestinal Eimeria, 1000-5000 oocysts per head part of precocious attenuated strain of medium Eimeria, and 3000-30000 oocysts per head part of wild Eimeria strain or precocious attenuated strain of perforated Eimeria.
In order to expand the immune protection range of the vaccine, the rabbit coccidiosis live vaccine composition can comprise a plurality of high-pathogenic coccidia and low-pathogenic coccidia, thereby forming a multivalent coccidia vaccine.
As a preferred embodiment of the present invention, the present invention provides a tetravalent rabbit coccidia live vaccine composition consisting of a Prenylated strain of Eimeria macrotype, Eimeria intestinalis and Eimeria intermedia and a Periplaneta praecox strain of Eimeria punctata. The immunogen composition of the rabbit coccidiosis live vaccine composition is as follows: 1000-3000 oocysts per head part of precocious attenuated strain of large Eimeria, 1000-4000 oocysts per head part of precocious attenuated strain of intestinal Eimeria, 2000-5000 oocysts per head part of precocious attenuated strain of medium Eimeria, and 3000-30000 oocysts per head part of wild Eimeria strain or precocious attenuated strain of perforated Eimeria. Wherein, the ratio of the number of oocysts of the E.perforatum to the number of oocysts of E.macrogol, E.enterobacteriaceae and E.intermedia is preferably (5-20): 1.
the rabbit coccidiosis live vaccine has the following immunization modes: 20-50 day old rabbits were inoculated orally.
Further, the invention provides a preparation method of the rabbit coccidiosis live vaccine composition, which comprises the breeding of prematurity attenuated strains; the breeding of the early-maturing attenuated insect plants adopts a classical early-maturing pressure breeding method or a breeding method comprising the following steps: and (3) orally inoculating the rabbit with the to-be-attenuated coccidia strain, and orally inoculating the rabbit with the to-be-attenuated coccidia strain at a high dose once or more within 12-72 hours after inoculation, and harvesting oocysts discharged earliest for passage amplification.
Preferably, the first inoculation is followed by the second inoculation, and the time interval between the second inoculations is 12-60 hours; the high dose is 5-10 times of the immunization dose of the first inoculation.
Specifically, the high-dose coccidian strain to be attenuated is inoculated orally once or more times twice respectively at 12-72 hours and 24-84 hours after the first inoculation.
Preferably, the breeding method of the early-maturing attenuated insect strain comprises the following steps: the coccidia to be attenuated is added at 1.0X 105～2.0×105Inoculating dose of sporulated oocysts/rabbits to the rabbits orally, and inoculating the to-be-attenuated coccidian strains at 5.0 × 10 hours after the first inoculation5～1.0×106The sporulated oocysts/rabbits were inoculated once more orally; then the first inoculation is carried out 24-84 hours later, the coccidia strain to be attenuated is treated at the ratio of 1.0 multiplied by 106～2.0×106The sporulated oocysts/rabbit were inoculated once orally at the inoculation dose and the earliest shed oocysts were harvested for passage expansion.
The invention also provides application of the rabbit coccidiosis live vaccine composition or the rabbit coccidiosis live vaccine composition obtained by the preparation method in preparation of a preparation for preventing or treating rabbit coccidiosis.
The invention provides a rabbit coccidiosis live vaccine which comprises the rabbit coccidiosis live vaccine composition.
Preferably, the rabbit coccidiosis live vaccine further comprises an adjuvant and an auxiliary material which are allowed in the field of animal vaccines.
On the other hand, the invention provides a breeding method of a premature and weak coccidia strain of the coccidia, which comprises the following steps: and (3) orally inoculating the rabbit to be attenuated coccidian strain, and orally inoculating the rabbit again with the high-dose coccidian strain to be attenuated for one time or more 12-72 hours after the first inoculation, and harvesting oocysts discharged earliest for passage amplification.
Preferably, the first inoculation is followed by the second inoculation, and the time interval between the second inoculations is 12-60 hours; the high dose is 5-10 times of the immunization dose of the first inoculation. More preferably, the time interval between the first inoculation and the second inoculation is 12-48 hours, and the time interval between the first inoculation and the second inoculation is 12-24 hours.
The rapid weakening breeding method of the rabbit coccidia is different from the most commonly adopted classical early maturing pressure breeding method at present, the cycle of the early maturing weakening insect strain obtained by increasing one or more times of high dose inoculation and collecting oocysts discharged at the earliest stage for passage and amplification is shortened by 2-5 generations compared with that of the classical early maturing pressure breeding method, and the obtained early maturing insect strain is stable in genetic characteristic, remarkably reduced in pathogenicity and kept good in immunogenicity.
Preferably, the oral inoculation once or more times of the high-dose coccidia strain to be attenuated is twice the inoculation 12-72 hours and 24-84 hours after the first inoculation respectively. More preferably, the secondary inoculation is performed twice at 12 to 48 hours and 24 to 72 hours after the primary inoculation, respectively.
More preferably, the breeding method comprises: the coccidia to be attenuated is added at 1.0X 105～2.0×105Inoculating dose of sporulated oocysts/rabbits to the rabbits orally, and inoculating the to-be-attenuated coccidian strains at 5.0 × 10 hours after the first inoculation5～1.0×106The sporulated oocysts/rabbits were inoculated once more orally; then the first inoculation is carried out 24-84 hours later, the coccidia strain to be attenuated is treated at the ratio of 1.0 multiplied by 106～2.0×106The sporulated oocysts/rabbit were inoculated once orally at the inoculation dose and the earliest shed oocysts were harvested for passage expansion.
The invention further provides application of the breeding method of the coccidian presenile attenuated strain in breeding of the coccidian attenuated strain or preparation of a coccidiosis vaccine.
The invention also provides a coccidiosis live vaccine which comprises the coccidiosis prematurity attenuated strain obtained by breeding by using the breeding method of the coccidiosis prematurity attenuated strain.
The invention has the beneficial effects that:
the rabbit coccidiosis live vaccine provided by the invention has higher immune protection effect and safety, can effectively prevent and treat rabbit coccidiosis, has long immune duration, can effectively reduce or eliminate the use of anti-coccidiosis drugs, reduces the breeding cost, obviously reduces the adverse effects of vaccination on growth and production performance of rabbits such as weight gain and the like, improves the breeding benefit, is easy to produce, has no pollution to animal products and environment, and has good economic benefit and wide application prospect.
The breeding method of the coccidian prematurity attenuated strain provided by the invention can effectively shorten the breeding cycle of the coccidian prematurity attenuated strain, effectively improve the breeding efficiency of the coccidian prematurity attenuated strain, and the obtained prematurity insect strain has stable genetic property, obviously reduces pathogenicity, and simultaneously keeps good immunogenicity, thereby providing an efficient method for breeding the coccidian prematurity attenuated strain and preparing a coccidian vaccine.
Detailed Description
Preferred embodiments of the present invention will be described in detail with reference to the following examples. It is to be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the present invention. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit and scope of this invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example 1 Breeding method of coccidia precocious attenuated insect strain (1)
This example provides a method for breeding a premature attenuated coccidian strain of coccidian, comprising the following steps:
(1) primary inoculation: the coccidia strain to be attenuated is treated at 1.0X 105Sporulated oocyst/rabbit vaccination dose clean-grade (coccidiosis free) new zealand white rabbits were inoculated orally;
(2) and (3) repeating inoculation: the coccidia strains to be attenuated are inoculated at 5.0X 10 hours after the first inoculation5The sporulated oocysts/rabbits were inoculated once more orally; then the coccidian strain to be attenuated is inoculated at 1.0X 10 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once orally at an inoculation dose;
(3) the oocysts that are expelled earliest are harvested and subcultured after sporogenesis under appropriate conditions.
Example 2 Breeding method of coccidian precocious attenuated insect strain (2)
This example provides a method for breeding a premature attenuated coccidian strain of coccidian, comprising the following steps:
(1) primary inoculation: the coccidia strain to be attenuated is treated at 1.0X 105Sporulated oocyst/rabbit vaccination dose clean-grade (coccidiosis free) new zealand white rabbits were inoculated orally;
(2) and (3) repeating inoculation: the coccidia strains to be attenuated are inoculated at 5.0X 10 hours 24 hours after the first inoculation5The sporulated oocysts/rabbits were inoculated once more orally; then the coccidia strain to be attenuated is treated at 1.0X 10 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once orally at an inoculation dose;
(3) the oocysts that are expelled earliest are harvested and subcultured after sporogenesis under appropriate conditions.
Example 3 Breeding method of coccidia precocious attenuated insect strain (3)
This example provides a method for breeding a premature attenuated coccidian strain of coccidian, comprising the following steps:
(1) primary inoculation: the coccidia strain to be attenuated is treated at 2.0X 105Sporulated oocyst/rabbit vaccination dose clean-grade (coccidiosis free) new zealand white rabbits were inoculated orally;
(2) and (3) repeating inoculation: the coccidia strains to be attenuated are inoculated at 1.0X 10 hours 12 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once more orally; then the coccidian strain to be attenuated is inoculated at 2.0X 10 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once orally at an inoculation dose;
(3) the oocysts that are expelled earliest are harvested and subcultured after sporogenesis under appropriate conditions.
Example 4 Breeding method of coccidian precocious attenuated insect strain (4)
This example provides a method for breeding a premature attenuated coccidian strain of coccidian, comprising the following steps:
(1) primary inoculation: the coccidia strain to be attenuated is treated at 2.0X 105Sporulated oocyst/rabbit vaccination dose clean-grade (coccidiosis free) new zealand white rabbits were inoculated orally;
(2) and (3) repeating inoculation: the coccidia strains to be attenuated are inoculated at 1.0X 10 hours 36 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once more orally; then the coccidian strain to be weakened is inoculated at 2.0X 10 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once orally at an inoculation dose;
(3) the oocysts that are expelled earliest are harvested and subcultured after sporogenesis under appropriate conditions.
Example 5 Breeding method of coccidian precocious attenuated insect strain (5)
This example provides a method for breeding a premature attenuated coccidian strain of coccidian, comprising the following steps:
(1) primary inoculation: the coccidia strain to be attenuated is treated at 2.0X 105Sporulated oocyst/rabbit vaccination dose clean-grade (coccidiosis free) new zealand white rabbits were inoculated orally;
(2) and (3) repeating inoculation: the coccidia strains to be attenuated are inoculated at 1.0X 10 hours 48 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once more orally; then the coccidian strain to be attenuated is inoculated at 2.0X 10 hours 72 hours after the first inoculation6The sporulated oocysts/rabbits were inoculated once orally at an inoculation dose;
(3) the oocysts that are expelled earliest are harvested and subcultured after sporogenesis under appropriate conditions.
Experimental example 6 Breeding of early maturing attenuated insect plants
By using the breeding method of the coccidia precocious attenuated strain of the embodiment 1-5, the precocious attenuated strain of the Eimeria macrotype, the Eimeria mesotype, the Eimeria intestinalis and the Eimeria perforated by the weak pathogenic species with high prevalence rate and strong pathogenicity is bred respectively. The breeding method of coccidia premature attenuated strains provided in examples 1-5 is compared with the breeding times and the latent period of the traditional breeding method reported in the literature (the traditional early breeding method is specifically that oral inoculation is carried out on coccidia strains to be attenuated at high dose, and oocysts which are discharged earliest are harvested, and the specific method and the experimental results are shown in the literature, and the results are shown in Table 1. The results show that the breeding generations required by the breeding method for the coccidia prematurity attenuated insect strains provided by the embodiments 1-5 are obviously reduced, the prematurity insect strains with weakened virulence can be obtained more quickly, and the obtained insect strains still keep the shortened prepatent period (Table 1) after 10 generations of pressureless continuous propagation, which shows that the genetic characteristics of the attenuated insect strains obtained by the method of the present embodiment are stable.
TABLE 1 comparison of efficiency of obtaining precocious attenuated strains of rabbit coccidia by different breeding methods
Example 7 development of trivalent Rabbit Coccidiosis live vaccine Using Rabbit Coccidia precocious attenuated Strain
In this example, the precocious attenuated strains of Eimeria maxima, Eimeria intermedia and Eimeria intestinalis obtained by the breeding method of example 1 were used for the preparation of rabbit coccidiosis live vaccines, and the development of trivalent live vaccines against rabbit coccidiosis was carried out.
First, to effectively resist 5.0 × 104The optimal immunization dose of each precocious attenuated strain was determined using a virulent strain of the wild insect infected with the sporulated oocysts as a standard (table 2). According to the experimental results shown in table 2, the optimal immune dose of each early-maturing attenuated insect strain is determined as follows: e.macrotype 1500/rabbit, E.mesoscale 3000/rabbit and E.enteron 2000/rabbit.
TABLE 2 optimal immunization dose screening of Eimeria maxima, Eimeria intermedia and Eimeria intestinalis precocious Weak strains
And further performing multi-prescription proportioning based on the determined optimal immune dose of the single insect strain, determining the appropriate immune dose of each insect strain in the trivalent vaccine, namely 1500/rabbit of large Eimeria, 3000/rabbit of medium Eimeria and 2000/rabbit of Eimeria intestinalis, and preparing the trivalent rabbit coccidia live vaccine according to the determined immune dose.
The safety and immunoprotection efficacy analysis is carried out on the prepared trivalent rabbit coccidium live vaccine by the following specific method: after proportioning the vaccines according to the determined optimal immune dose, new zealand white rabbits (10 rabbits each group) with the age of 35 days are inoculated orally, and an immune group, a repeated immune group, a high-dose immune group, an immune control group and an immune attack group are arranged. Wherein the repeated immunization group is used for carrying out secondary immunization according to the same dose 14 days after the immunization according to the optimal immunization dose; the high dose immunization group was immunized 10 times the recommended dose. After immunization, all rabbits were normally raised to ensure sufficient feed and drinking water. Evaluation of safety: weighing is carried out 14 days after immunization, and the change of the weight gain of the rabbits before and after each group of immunization is calculated. Evaluation of the immunopotency: at 14 days after immunization, mixed wild venom strains (each containing 1.0X 10) were administered at high doses5Sporulated oocysts) are subjected to toxin attack, and oocyst discharge and body weight gain after toxin attack are counted.
TABLE 3 safety and immunoprotective efficacy of trivalent rabbit coccidia live vaccines
The results are shown in table 3, and the results show that after the three rabbit coccidia precocity attenuated insect strains are immunized according to the optimal proportion, the rabbit coccidia precocity attenuated insect strains have good immunoprotection for infection of a large dose of the virulent insect strains, but the immunization of the trivalent rabbit coccidia live vaccine has certain adverse effects on the weight gain of rabbits.
Example 8 development of tetravalent Rabbit coccidia live vaccine
This example further introduced a high dose (3000 oocysts per head part, 10000 oocysts per head part, or 30000 oocysts per head part) of the perforated Eimeria precocious attenuated strain obtained by the breeding method of example 1 on the basis of the trivalent coccidia live vaccine of example 2 to prepare a tetravalent rabbit coccidiosis live vaccine.
The immunoprotective efficacy and safety of the tetravalent rabbit coccidia live vaccine obtained above were evaluated, and the effect of the introduction of the perforated eimeria on the immunoprotective efficacy and safety of eimeria maxima, eimeria intermedia and eimeria intestinalis was analyzed by the following specific method:
after proportioning the vaccines according to the optimal immunization dose, the vaccine components are inoculated with 35-day-old New Zealand white rabbits (10 in each group) by mouth according to different high doses of the E.perforatum (3000 oocysts/head part, 10000 oocysts/head part or 30000 oocysts/head part) in the vaccine components, and the groups are shown in Table 4. After immunization, all rabbits were normally raised to ensure sufficient feed and drinking water. Evaluation of safety: weighing is carried out 14 days after immunization, and the change of the weight gain of the rabbits before and after each group of immunization is calculated. Evaluation of the immunopotency: at 14 days after immunization, mixed wild venom strains (each containing 1.0X 10) were administered at high doses5Sporulated oocysts) are subjected to toxin attack, and oocyst discharge and body weight gain after toxin attack are counted.
The results are shown in table 4, and the results indicate that the addition of a high dose of perforated eimeria has no significant effect on the immunoprotective efficacy of the trivalent coccidia vaccine consisting of large eimeria, medium eimeria and eimeria enterica, but the adverse effect on the weight gain of rabbits is significantly reduced, demonstrating that the tetravalent rabbit coccidia live vaccine consisting of large eimeria, medium eimeria, eimeria enterica and perforated eimeria has both high immunoprotective efficacy and high safety.
TABLE 4 immunopotentiation and safety analysis of tetravalent Rabbit coccidia live vaccine
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (3)
1. A rabbit coccidiosis live vaccine composition is characterized by comprising the following components: 1500 oocysts per head of large Eimeria praecox attenuated strain, 2000 oocysts per head of Eimeria enteric attenuated strain, 3000 oocysts per head of medium Eimeria praecox attenuated strain, and 10000-30000 oocysts per head of Eimeria per puncture attenuated strain.
2. The method for preparing the rabbit coccidiosis live vaccine composition of claim 1, wherein the breeding method of the prematurity attenuated strain comprises the following steps: the coccidia to be attenuated is added at 1.0X 105~2.0×105Inoculating dose of sporulated oocysts/rabbits to the rabbits orally, and inoculating the to-be-attenuated coccidian strains at 5.0 × 10 hours after the first inoculation5~1.0×106The sporulated oocysts/rabbits were inoculated once more orally; then the first inoculation is carried out 24-84 hours later, the coccidia strain to be attenuated is treated at the ratio of 1.0 multiplied by 106~2.0×106The sporulated oocysts/rabbit were inoculated once orally at the inoculation dose and the earliest shed oocysts were harvested for passage expansion.
3. Use of the live rabbit coccidiosis vaccine composition of claim 1 for the preparation of a formulation for the prevention of rabbit coccidiosis.
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