MX2007012784A

MX2007012784A - Coccidial vaccine and methods of making and using same.

Info

Publication number: MX2007012784A
Application number: MX2007012784A
Authority: MX
Inventors: Larry R Mcdougald; Alberta Lorraine Fuller; Joyce Anita Pritchard
Original assignee: Univ Georgia Res Found
Priority date: 2005-04-15
Filing date: 2006-04-14
Publication date: 2008-01-11
Also published as: BRPI0607550A2; JP2008538364A; SI1877085T1; RU2007142203A; HK1112199A1; DK1877085T3

Abstract

The present invention relates to a vaccine for coccidiosis in chickens prepared from three attenuated Eimeria species: E. acervulina, E. maxima and E. tenella. The vaccine was similar to or superior to other anticoccidial drugs in stimulating protective immunity against coccidiosis.

Description

COCCIDAL VACCINE AND METHODS FOR PREPARING AND USING THE FIELD OF THE INVENTION The present invention relates to the preparation of immunogenic compositions and vaccines against diseases caused by coccidia. The present invention also provides attenuated vaccines against coccidiosis. BACKGROUND OF THE INVENTION [0002] Coccidiosis is a disease caused by infection with one or more of the many species of coccidia which is a subdivision of the phylum Protozoa, intracellular protozoan parasites of the subphylum Apicomplexa and the genus Eimeria. The genus Eimepa contains the most economically important species in domestic birds, such as chickens, ducks, geese, guinea fowl, peacock, pheasants, pigeons and turkeys. While coccidiosis occurs in virtually all classes of birds, parasites are host-specific and each species occurs in one or a limited group of related hosts. On the other hand, bird hosts are known to harbor more than one species of cocci. The Eimeria species that cause coccidiosis in chickens include E. a cervulma, E. brunet ti, E Hagam, E. maximum, E. my tis, E miva ti, E. neca tpx, E. praecox and E. tenella. E. A cervida ina is one of the most common species found in the stretcher of houses of tender chickens. This has a great reproductive potential and is considered to be pathogenic because it produces a marked depression in the body's weight gain, higher feed conversion and produces thick lesions in the upper small intestine. Among domesticated birds, chickens are more susceptible to significant economic losses from coccidiosis, although losses can also occur within turkeys, geese, ducks and guinea fowl. Coccidiosis has also caused serious losses in pheasants and quail bred in captivity. The effects of a coccidiosis infection can take the highly visible form of mortality of the devastating flock, but another undesirable effect of morbidity and / or weight loss resulting from the infection. During the life cycle, the Eimeria parasite passes through a number of stages, (see, for example, U.S. Patent No. 6,100,241 for a review). The life cycle begins when the chicken ingests the infectious stage, known as the sporulated oocyst, during ground feeding or by inhaling dust. The sporulated oocyst wall is broken by a combination of mechanical grinding action and chemical action in the gizzard and intestinal tract, resulting in the release of four sporocysts. Sporocysts pass into the duodenum where they are exposed to bile and digestive enzymes that result in the release of two sporocytes per sporocyst. The sporozoites are mobile and look for the appropriate host epithelial cells in order to penetrate and reproduce in these. After infection of an epithelial cell, the parasite enters the schizont phase of its life cycle, which produces 8 to 16 a > 200 merozoites per schizont. Once released from the schizont, the merozoites are free to further infect the epithelial cells. After two to five of these asexual reproduction cycles, the intracellular merozoites grow in known sexual forms such as the female or macrogametocyte and the male or microgametocyte. After fertilization of the macrogametocyte by the microgametes released from the microgametocyte, a zygote is formed that creates a cyst wall around it. The newly formed oocyst is passed out of the chicken infected with the feces. With the correct environmental conditions of temperature and humidity and sufficient oxygen in the air, the oocyst will sporulate in the infectious stage, ready to infect a new host and in this way spread the disease. Thus, an intermediary host is not required for the transfer of the parasite from bird to bird. The result of the Eimeria parasite that infects the digestive tract of a chicken can be a reduction in weight gain, increased feed conversion, stopping egg production and, in some cases, death. The increase in the intensive production of poultry has been accompanied by severe losses due to this parasite; in fact, coccidiosis has become an economically important parasitic disease. In the past, several methods have been used in attempts to control coccidiosis. Before the arrival of the chemotherapeutic agents, the improved sanitation using disinfectants, together with the mechanical removal of the litter from the animals, was mainly employed; Enough oocysts, however, usually remained to allow disease. The introduction of coccidiostatic agents in food or drinking water, in addition to good management, resulted in some success in controlling the disease. Such agents have been found to suffer from a drop in effectiveness over the years, partially due to the development of drug-resistant coccidial strains. In addition, several chemotherapeutic agents have been found to leave residues in the meat, rendering it unsuitable for consumption. U.S. Patent Nos. 4,438,097; 4,639,372; 4,808,404; 5,055,292; 5,068,104; 5,387,414; 5,602,033; 5,614,195; 5,635,181; 5,637,487; 5,674,484; 5,677,438; 5,709,862; 5,780,289; 5,795,741; 5,814,320; ,843,722; 5,846,527; 5,885,568; 5,932,225; 6,001,363 and 6,100,241 relate to coccidiosis vaccines, including vaccines and recombinants. However, there are problems with existing coccidiosis vaccines, such as reduced efficacy, cross-infection with other parasites (eg, Clos tpdi um spp.) And poor bird performance. Thus, there is still a need for effective coccidiosis vaccines with reduced non-existent cross-infection that do not adversely affect bird performance. The citation or identification of any document in this application is not an admission that such document is available with the prior art for the present invention. BRIEF DESCRIPTION OF THE INVENTION This invention is based in part on an attenuated coccidiosis vaccine which is effective in the face of virulent stimulation, reduced cross-infection with Clostridium um spp. and has better bird performance as defined by feed conversion ratios when compared to other coccidiosis vaccines. The invention relates to a mixture of sporulated oocysts of early E strains. a cervul ma, E. maximum and E. tenel The sporulated oocysts were isolated from a seed culture harvested from one or more chickens plated with a culture of a precocious strain of E. a cervul ma, E. maximum or E. tenel la, that is, one or more chickens are sown with either a precocious strain of E. a cervul ma, E. maximum or E. It has resulted in three groups of chickens, each planted with a different Eimepa strain. The isolated sporulated oocysts were combined to formulate a mixture of sporulated oocysts of early E strains. a cervulma, E. maximum and E. tenella. In an advantageous embodiment, the chickens are SPF chickens from 2 to 8 weeks of age. In another advantageous embodiment, about 100 to about 15,000 oocysts are sown per chicken to generate the seed culture. In another advantageous embodiment, sporulated oocysts of the seed culture are isolated by centrifugation. The present invention also provides verification of the sporulated oocysts that are characteristic of the early E strain. a cervulma, E. maximum or E. tenella. The invention relates to a mixture of sporulated oocysts of early E strains. a cervul ma, E. maximum and E. tenella, where the mixture is from about 10 to about 1000 oocysts of E. to cervix, approximately 10 to approximately 100 E oocysts. maximum and approximately 10 to approximately 1000 E. tenella oocysts. Advantageously, the mixture is about 500 E oocysts. to cervuloma, approximately 50 to approximately 100 oocysts of E. maximum and approximately 100 to approximately 500 E. tenella oocysts. More advantageously, the mixture is about 500 E. acervulma oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts that are combined. The invention also relates to specific relationships of sporulated oocysts isolated from early strains of E. acervulina, E. maxim and E. tenella, where the ratio of E. acervulma, E. maxim, E. tenella is approximately 10: 1. at 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulina, there are approximately 1 to 2 sporocysts of E. maximus and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the relation of E. acervulma: E. maximum E. teneJla is approximately 5: 1: 1 (that is, 10: 2: 2). The invention also relates to the efficacy test of the mixture of sporulated oocysts of the early strains of E. acervulma, E. maxim and E. tenella. Advantageously, the test relates to the administration of a stimulation dose of approximately 100,000 to approximately 500,000 E. acervulina oocysts and approximately 10,000 to approximately 1,000,000 maximum E. oocysts or approximately 10,000 to approximately 100,000 E. tenella oocysts. to the animal In a more advantageous modality, the dose of stimulation is from approximately 200,000 oocysts of E. acervulma and approximately 20,000 to approximately 500,000 oocysts of E. maximus, or approximately 20,000 to approximately 50,000 oocysts of E. tenella. The present invention relates to the immunization of a chicken, advantageously a broiler chicken. However, the methods for making the vaccine described herein can be extrapolated to other animals infected by Eimepa, in particular birds, such as, but not limited to, a chicken, duck, goose, guinea fowl, peacock, pheasant , pigeon, quail or turkey. The invention encompasses an immunogenic or vaccine composition comprising a mixture of sporulated oocysts isolated from the early strains of E. acervulma, E. maximus and E. tenella. In an advantageous embodiment, the mixture is about 10 to about 1000 oocysts of -. Acervulin, about 10 to about 100 E. maximum oocysts and about 10 to about 1000 oocysts of E. tenella. Advantageously, the mixture is about 500 oocysts of E. acervulma, about 50 to about 100 oocysts of E. maximus and about 100 to about 500 oocysts of E. tenella. More advantageously, the mixture is about 500 oocysts of E. acervulma, about 100 oocysts of E. maximus and about 100 oocysts of E. tenella which are combined. The invention also relates to an immunogenic or vaccine composition comprising specific ratios of sporulated oocysts isolated from early E strains. acervulma, E maximum and E. tenel la, where the relation of E. to cervulina: E. maximum E It is approximately 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts from E to cervical, there are approximately 1 to 2 sporocysts of maximum E and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the relationship of E. a cervulma: E. maximum: E. tenella is approximately 5: 1: 1 (that is, 10: 2: 2). The invention also provides for the induction of an immune response or the induction of a protective immune response comprising administering an effective amount of the immunogenic or vaccine composition comprising a mixture of sporulated oocysts isolated from the early strains of E. a cervul ma, E. maximum and E. have it to provoke or induce the response in an animal. Advantageously, the animal is a bird, such as, but not limated to, a chicken, duck, goose, guinea fowl, peacock, pheasant, pigeon, quail or turkey. In the most advantageous mode, the bird is a chicken, advantageously a roast chicken. The method for inducing an immune response or inducing an immune response may also include the administration of an adjuvant, a cytokine, or both.

Advantageously, the amount effective to induce an immune response or induce an immunological or protective response is about 10 to about 1000 E oocysts. to cervulina, approximately 10 to approximately 100 oocysts of E. maximum and approximately 10 to approximately 1000 oocysts of - .. tenella. Advantageously, the effective amount is approximately 500 oocysts of E. to cervulina, approximately 50 to approximately 100 oocysts of E. maximum and approximately 100 to approximately 500 E oocysts. tenella. Most advantageously, the effective amount is about 500 oocysts of E. to cervulina, approximately 100 oocysts of E. maximum and approximately 100 oocysts of E. tenella that are combined. In another embodiment, the effective amount is sufficient to withstand a dose of stimulation of 'approximately 100,000 to approximately 500,000 oocysts of E. to cervulina and approximately 10,000 to approximately 1,000,000 maximum E. oocysts or approximately 10,000 to approximately 100,000 E oocysts. hold the animal. More advantageously, the dose of stimulation is approximately 200,000 oocysts of E. to cervix and approximately 20,000 to approximately 500,000 E oocysts. maximum or approximately 20,000 to approximately 50,000 E. tenella oocysts. The effective amount to induce a response II Immune or induce a protective or immunological response can also be expressed as sporulated oocyst relationships of early strains of E. acervulma, E. maxim and E. tenella, where the ratio of E. acervulma: E. maxim: E. tenella is approximately 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulina, there are approximately 1 to 2 sporocysts of E. maximus and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervullma: E. maximum: E. tenella is approximately 5: 1: 1 (ie, 10: 2: 2). In consideration of the prevalence and pathogenicity of several Eimepa species, a successful attenuated coccidiosis vaccine must contain at least a number of Eimepa strains sufficient to induce an immune response or induce an immune or protective response that is nonpathogenic to the recipient. the vaccine. The present relationship is related to a combination of oocysts of four Eimepa-specific early strains, ie E. acervulma, E. maximus and E. tenella which results in an effective and non-pathogenic vaccine. The addition of other strains of Eimeria, such as E. brunetti, E. necatrix and 17. praecox may be disadvantageous with respect to the efficacy, cross-ampheccato or pathogenicity of the vaccine. Since E. brunetti, E. necatrix and E. praecox are not necessary for the efficacy of the coccidiosis vaccine disclosed herein, it would be advantageous to exclude these strains from the vaccine of the present invention. It is noted that this description and particularly in the claims and / or paragraphs, the terms such as "comprises", "understood", "comprising" and the like can have the meaning attributed to it in the US patent law; for example, they can mean "include", "included", "including" and the like; and those terms such as "consisting essentially of" and "consisting essentially of" have the meaning ascribed to them in the American patent law, for example, they allow elements not explicitly mentioned, but exclude elements that are found in the prior art or that affect a basic or novel feature of the invention. These and other embodiments are disclosed and are obvious from and understood by, the following Detailed Description. DETAILED DESCRIPTION The invention is based, in part, on an attenuated coccidiosis vaccine which is effective in the face of virulent stimulation, reduced cross-infection with Clos tridium spp. and has better bird performance as defined by the feed conversion ratios when compared to other coccidiosis vaccines. The invention relates to a mixture of sporulated oocysts of early E strains. to cervuloma, E. maximum and E. tenel The sporulated oocysts were isolated from a seed culture harvested from one or more chickens plated with a culture of a precocious strain of E. acervulina, E. maxim or E. tenella, that is, one or more chickens are sown with either a precocious strain of E. a cervulma, E. maximum or E. tenella that results in three groups of chickens, each sown with a different Eimeria strain. Optionally, the sporulated oocysts of E strains. My tis can also be added to the mixture of sporulated oocysts. Advantageously, the Eimeria strain is a precocious strain. Early strains are derived from species in the field that are not pathogenic when administered at the correct dose. In an advantageous embodiment, the Eimeria strain is a precocious strain of the respective microorganism as described in Avian Pathology, 17: 305-314, 1988 entitled "Eimeria of American Chickens: Characteristics of S x Attenuated Strains Produced by Selection for Precocious Development, PL Long and Joyce K. Johnson, the description of which is incorporated by reference in its entirety.The microorganisms may be attenuated by their selection for early development as described in Avian Pathology, 17: 305-314, 1988 entitled "Eimeria of American Chickens: Charactepsta cs of Six Attenuated Strains Produced by Selection for Precocious Development, PL Long and Joyce K. Johnson, the description of which is incorporated by reference in its entirety. Briefly, microorganisms are attenuated by selecting an earlier pre-patent period, that is, when oocysts first show up in the stool. Such methods are well known to one of skill in the art and constitute routine experimentation. The extract cultures of Eimepa strains for seed crops include, but are not limited to, the following. The main strain of the Eimepa to cervino, obtained from T. K. Jeffers at Hess and Clark Laboratories 1969, is believed to have been isolated by Dr. M. M. Farr at USDA, Beltsville, MD, which was derived from a single oocyst. The maximum Eimepa culture was derived from an interproduction mixture of 10 purified isolates obtained from Georgia, Delaware, Maryland, Virginia and Texas. The main strain of Eimeria my tis culture was isolated from Gainesville, Georgia in Julao, 1978 and purified by a single oocyst isolate. The main strain of the Eimeria tenel culture was obtained from a crop maintained at Pennsylvania State University by Dr. Patten since the early 1960s and was acquired by the University of Georgia in 1982. Other strains of Eimeri at early ancluyen the isolate 809-13 of E. a precocious cervix LS 100, and E. mitis, precocious LS, obtained from Merck Research Laboratories, which were obtained from Dr. Peter Long. Alternatively, early attenuated Eimeria lines that have been deposited as sporocysts in the European Collection of Animal Cell Cultures ("ECACC") as patent deposits (see, for example, U.S. Patent No. 5,055,292, the description of which it is incorporated by reference in its entirety) are useful extract cultures for generating the Eimepa seed cultures described herein. Specifically, the deposits of E. to the cervix (deposit No. ECACC 86072203), E. maximum (deposit Nos. ECACC 86112011 and 86112012), E. mi ti s (deposit No. ECACC 86072206) and E. tenella (deposit No. ECACC 86072201) as described in U.S. Patent No. 5,055,292 are useful extract cultures for the seed crops of the present invention. Advantageously, the microorganisms are attenuated by their selection for early development as described in the foregoing. In another advantageous embodiment, the culture is pathogen-free. The extract cultures described above are advantageously maintained in the liquid or vapor phase of liquid nitrogen. Such methods are known to one of skill in the art. In an advantageous modality, chickens are two to eight weeks old. The sporulated oocysts are passed successively, without limitations to the passage, in chickens until the number of oocysts are sufficient to be used as seed for production. Advantageously, the crops should not be maintained for more than 12 months in order to maintain viability / infectivity. In an advantageous embodiment, the dedicated facils are maintained for each Eimeria species. Advantageously, a sufficient volume of sporulated oocysts (seeds) are mixed with the feed or alternatively, it is administered orally to provide each chicken with a minimum dose. In an advantageous embodiment, about 5000 to about 15,000 oocytes are planted by chicken to generate the seed culture. The sporulated oocysts of the seed culture are isolated from the feces of the birds. Sell by centrifugation. In one embodiment, the collection is as follows. The defecations are homogenized in an approximate ratio of 10% (w / v) in water. The large particles are removed by passing the homogenate through screens. The solids are separated, either by centrifugation, screening or by holding at 5 ± 3 ° C for up to 24 hours. If the solids separate when holding at 5 ± 3 ° C, they are further concentrated by centrifugation. The supernatant is discarded, and the solids are resuspended in a saturated NaC L solution (80% w / v) in water. The resulting solution is centrifuged. The oocysts are collected (removed) from the upper part of the liquid and resuspended in water. Optionally, the remaining liquid is diluted to 20-40% in NaCl with water and centrifuged. The pellet is then resuspended in a saturated NaCl solution and recentrifuged, until no addnal oocysts are recovered. The oocysts are washed no more than twice. The oocysts are washed free of salt by repeated centrifugation cycles of resuspension, followed by resuspension in a 0.5% solution of sodium hypochlo- prate for 10 to 15 minutes. The oocysts are then washed free of the sodium hypochlo- kick solution by repeated centrifugation and resuspension steps (3X). The final resuspension is done in a 2.5% aqueous solution of potassium dichromate (K2Cr07). The oocysts are then transferred to sporulation vessels. Sporulation is facilitated by dispersing the suspensions with air for a period not exceeding 72 hours at 27 ± 3 ° C. After sporulation the oocysts are maintained at 5 ± 3 ° C until the final product is produced. In another embodiment, the oocysts to be used according to the present method of vaccination can be prepared by any of several methods known to those skilled in the art. Such methods include those described in JF Ryley et al., Parasitology 73: 311-326, 1976, PL Long et al., Folia Veterinaria Lata na VI # 3, 201-217, 1976, and U.S. Patent No. 6,627,205, descriptions of which are incorporated by reference in their entirety. According to one method, commercial roasting chickens, about 2 weeks old, are infected with the Eimeria species of interest by orally priming an appropriate dose of sporulated oocysts. Well-known procedures for the collection and purification of oocysts from infected birds are then followed. For most Eimepa species, faeces are collected from infected birds 5-7 post-mfecca days, mixed and filtered to remove cell debris, then centrifuged at a sufficient rate to form the fecal material pellet. remaining. The pellet is resuspended in a saturated salt solution, in which the oocysts float and most of the contaminating cellular debris can be removed by centrifugation. The oocyst suspension is then diluted to a lower salt concentration. The oocysts are repeatedly washed to remove the salt and resuspended in potassium dichromate solution (2.5% w / v). The oocyst suspension is incubated at 29 ° C with shaking (for example, 140 rpm) for approximately 72 hours to induce sporulation of the oocysts. Alternatively, the oocysts can be treated with sodium hypochlorite and then sporulated. The number of sporulated oocysts / ml is determined by direct counting using either hemocytometer or McMaster plate, and the culture is stored or refrigerated until needed. To prepare the sporocysts, potassium dichromate is removed from the oocyst suspension described above by repeated washing of the oocysts, which involves collection of oocysts by centrifugation and resuspension in deionized or distilled water. When the dichromate has been removed as it is marked by the lack of yellowish-orange coloration, the oocyst suspension is sterilized. Advantageously, the sterilant beta-propiolactone (BPL). In an advantageous embodiment, 97% BPL is diluted 1:10 with sterile water, then 10-20 ml of BPL is added per liter of sporulated oocysts. In an alternate embodiment, the oocyst suspension is mixed with an equal volume of sodium hypochlorite (bleach) and incubated at room temperature for 15 minutes. The bleach is then removed by repeated washes, and the oocysts are resuspended in physiological saline or deionized water. Oocysts can be disrupted to release sporocysts using a variety of known techniques. For example, oocysts can be disrupted to release sporocysts by mixing the oocysts with glass beads of 1-4 mm in diameter and manual shaker, vortex mixer or agitacason incubator, or by using a portable homogenate. Unbroken oocysts and oocyst walls can be separated from the released sporocysts by differential centrifugation in 50% PERCOLL, a colloidal suspension of silica particles coated with polyvinylpyrrolidone (sold by Pharmacia Biotech) or 1 M sucrose as described in Dulski and collaborators, Avian Diseases, 32: 235-239, 1988. Sporocysts can be used in the present method of vaccination either mixed with or separated from the unbroken oocysts and the oocyst walls. Advantageously, the dose of sporocysts is separated from the oocysts and the oocyst walls. In an advantageous embodiment, the specifications for an acceptable collection of the seed culture are known. First, the ratio of sporulated oocysts to total oocysts was determined. Only collections that meet or exceed >40% sporulation is considered acceptable. Second, the size, shape and appearance of each collection of oocysts must be characteristic for the species proposed to be produced. For example, the parameters that are considered in the characterization of the Eimeria species include but are not limited to, DNA-based technologies, floating density of DNA, enzyme variation, host and site specificity, immunological specificity, pathogenicity, pre -patent and sporulation time (see, for example, Long &Joyner, J Protozool, 1984 Nov; 31 (4): 535-41 and Shirley, Acta Vet Hung., 1997; 45 (3): 331-47, descriptions of which are incorporated by reference). The sporulated oocysts staked from the seed cultures described herein combine to formulate a mixture of sporulated oocysts of early strains of E. acervulma, E. maximus and E. tenella. Generally, the mixture is about 10 to about 1000 oocysts of E. acervulina, about 10 to about 100 oocysts of E. maximus and about 10 to about 1000 oocysts of E. tenella. Advantageously, the range of sporulated oocysts in the mixture is from about 125 to about 500 E. acervulma oocysts, about 25 to about 100 E. maximum oocysts and about 25 to about 500 E. tenella oocysts. In one embodiment, a low dose is approximately 125 oocysts of E. acervulma, approximately 25 oocysts of E. maximus and approximately 25 oocysts of E. tenella. In another embodiment, an average dose is approximately 250 oocysts of E. acervulma, approximately 50 oocysts of E. maximus and approximately 50 oocysts of E. tenella. In yet another embodiment, a high dose is approximately 500 E. acervulin oocysts, approximately 100 E. maximum oocysts and approximately 100 oocysts of E. tenella. The mixture may optionally comprise about 10 to about 1000 E. mitis oocysts, advantageously about 125 to about 500 E. mitis oocysts, about 125 E. mitis oocysts at a low dose, about 250 E. mitis oocysts at an average dose and approximately 500 oocysts of E. mjtis in a high dose. The mixture may optionally comprise about 10 to about 1000 E. mitis oocysts, advantageously about 125 to about 500 E. mitis oocysts, about 125 E. mitis oocysts at a low dose, about 250 E. mitis oocysts at an average dose and approximately 500 oocysts of E. mitis in a high dose. The invention also relates to specific relationships of sporulated oocysts isolated from early strains of E. acervulma, E. maximus and E. tenella, where the ratio of E. acervulmaiE. maximum E. tenella is approximately 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulina, there are approximately 1 to 2 sporocysts of E. maximum and approximately 2 to 10 sporocastos of E. tenella). Advantageously, the ratio of E. acervuJma: E. maximum, E. tenella is approximately 5: 1: 1 (ie, 10: 2: 2). In a modality containing E. mitis, the ratio of E. acervul ina: E. maxim: E. mitis: E. tenella is approximately 10: 1 to 2: 10: 2 to 10 (ie, for every 10 sporocysts of E. acervulma, and approximately 1 to 2 sporocysts of E. maximus, approximately 10 sporocysts of E. mitis and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: E. maximum: E. itis: E. tenella is approximately 5: 1: 5: 1 (ie, 10: 2: 10: 2). Advantageously, the mixture is about 500 oocysts of E. acervulina, about 50 to about 100 oocysts of E. maximus and about 100 to about 500 oocysts of E. tenella. In a more advantageous embodiment, the mixture is about 500 E. acervulin oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts. The mixture can optionally contain about 500 oocysts of E. mitis, advantageously about 500 oocysts of E. mitis. Advantageously, the oocysts are suspended in a preservative consisting of a saline solution regulated in 0.01 M phosphate containing gentamicin. In another embodiment, the oocysts are suspended in any variety of preservatives or organic acids such as, but not limited to, acetic acid, citric acid, potassium dichromate or propionic acid. For example, but not by limitation, sufficient sterile 0.01 M phosphate buffered saline containing no more than 30 mcg / ml gentamicin is used to produce 2 ml per bottle for a 2,000-dose presentation, 5 ml per bottle for a presentation of 5,000 doses and 10 ml per bottle for a presentation of 10,000 doses. Advantageously, the oocysts are stored in sterile borosilicate glass vials. For example, but not by limitation, the oocysts are filled aseptically in vaccine bottles with a semi-automatic or automatic supplier, the caps are inserted mechanically or manually and the aluminum cans are placed and plated. In another embodiment, the oocysts are suspended in sterile distilled water containing a suspending agent, for example a polysaccharide suspension agent, such as a gum, for example, xanthan gum or acacia gum, as a cellulose derivative, by example, carboxymethyl cellulose, hydroxypropyl methyl cellulose or microcrystalline cellulose, carrageenan, sodium alginate, pectin or starch; a polypeptide suspension agent such as gelatin; a synthetic polymer suspension agent such as polyacrylic acid, a silicate suspension agent such as aluminum magnesium silicate (see, for example, U.S. Patent No. 5,055,292, the disclosure of which is incorporated by reference in its entirety) . The present invention also provides that the verification of the sporulated oocysts are characteristic of the early strain of E. acervulma, E. maximum or E. tenella. Advantageously, all oocysts are attenuated, in that they are early. In another advantageous embodiment, the size, shape and appearance of each collection of oocysts should be characteristic of the species proposed to be produced. In yet another advantageous embodiment, the mixture of sporulated oocysts is tested for purity, foreign pathogens and / or death or severe lesions of the test animals, eg, chickens. The characteristics of the various species of Eimepa are fully exposed by Long PL and Reid WM (1982: A Guide for the Diagnosis of Coccidiosis m Chickens, University of Georgia Research Report 404) and Joyner LP (1978: Identification and Diagnosis, Avian Coccidiosis, Poultry Science Symposium No. 13, Bptish Poultry Science Ltd), the descriptions of which are incorporated by reference in their totalities. The invention also relates to the efficacy test of a mixture of sporulated oocysts of the early E strains. acervulma, E maximum and E. tenella. Advantageously, the test is related to the administration of a dose of approximately 100,000 to approximately 500,000 E oocysts. acervulma and approximately 10,000 to approximately 1,000,000 E oocysts. maximum or approximately 10,000 to approximately 100,000 E. tenella oocysts to the animal. In a more advantageous embodiment, the stimulation dose is approximately 200,000 E oocysts. to the cervix and approximately 20,000 to approximately 500,000 E oocysts. maximum or approximately 20,000 to approximately 50,000 E oocysts. tenel In a modality comprising E. my tis, as a stimulation dose of approximately 100,000 to approximately 500,000 E oocysts. my tis, advantageously approximately 200,000 oocysts of E. My tis can be used. The invention further provides the determination of bird performance as deduced by the feed conversion ratios as a result of the administration of the sporulated oocysts mixture of early E strains. acervul ina, E max ima and E. Tenel her to the birds. The feed conversion efficiency is defined as the pounds of feed to produce one pound of meat. A common result is approximately 1.90 or 2.00. A point in feed conversion is common that is .01, equal to approximately 0.5% (half a percent). If the metric system is used, the feed conversion is Kg of feed per Kg of meat, and is still proportional to the previous one. The present invention relates to the immunization of a chicken, advantageously a tender chicken for roasting. However, methods for making the vaccine described herein can be extrapolated to other animals infected by Eimepa, in particular birds, such as, but not limited to, a chicken, duck, goose, guinea fowl, peacock, pheasant , pigeon, quail or turkey, or a less advantageous modality such as a rabbit. The invention encompasses an immunogenic or vaccine composition comprising a mixture of sporulated oocysts isolated from the early strains of E. acervulma, E. maximus and E. tenella. The isolated sporulated oocysts are combined to formulate the sporulated oocysts composition of the early strains of E. acervulma, E. maxim and E. tenella. Generally, the mixture is from about 10 to about 1000 oocysts of E. acervulma, about 10 to about 100 oocysts of E. maximus and about 10 to about 1000 oocysts of E. tenella. Advantageously, the range of sporulated oocysts in the composition is from about 125 to about 500 E. acervulin oocysts, about 25 to about 100 E. maximum oocysts and about 25 to about 250 E. tenella oocysts. In one embodiment, a dosa ba is approximately 125 oocysts of E. acervulma, approximately 25 oocysts of E. maximus and approximately 25 oocysts of E. tenella. In another embodiment, an average dose is approximately 250 oocysts of E. acervulma, approximately 50 oocysts of E. maximus and approximately 50 oocysts of E. tenella. In still another embodiment, a high dose is about 500 E. acervulin oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts. Advantageously, the composition is about 500 E. acervulma oocysts, about 50 to about 100 E. maximum oocysts and about 100 to about 500 E. tenella oocysts. In a more advantageous embodiment, the composition is about 500 oocysts of E. acervulma, about 100 oocysts of E. maximus and about 100 oocysts of E. tenella. The mixture may optionally comprise about 10 to about 1000 oocysts of E. mitis, advantageously about 125 to about 500 oocysts of E. miLis, approximately 125 oocysts of E. mitis at a low dose, about 250 oocysts of E. mitis in an average dose and approximately 500 oocysts of E. my tis in a high dosas. The invention also relates to specific relationships of sporulated oocysts isolated from early strains of E. acervulma, E. maxim and E. tenella, where the ratio of E. acervulma: E. maximum E. tenella is approximately 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulma, there are approximately 1 to 2 sporocysts of E. maximus and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: E. maximum: E. tenella is approximately 5: 1: 1 (ie, 10: 2: 2). In a modality containing E. milis, the ratio of E. acervulma: E. maximum: E. mitis: E. tenella is approximately 10: 1 to 2: 10: 2 to 10 (ie, for every 10 E sporocysts). acervulma, there are approximately 1 to 2 sporocysts of E. maximus, approximately 10 sporocysts of E. mitis and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: E. maximum: E. mitis: E. tenella is approximately 5: 1: 5: 1 (ie, 10: 2: 10: 2). The term "immunogenic composition" herein covers any composition capable, once it has been administered to an animal, eg, a bird, of inducing a protective immune response against the parasite or antigen or immunogen or epitope. The term "vaccine" herein covers any composition capable, once it has been administered to the animal, eg, a bird, to address a protective immune response against the virus, or to effectively protect the animal against the parasitic invention. Immunogenic compositions or vaccines according to the invention may also include the pathogen or immunogen, antigen or epitope of the pathogen and at least one immunogen, antigen or epitope of another pathogen, parasite or virus, ie, the coccidiosis vaccine is combined with another bird vaccine. Such an immunogen, antigen or epitope can be, for example, of bacterial, or parasitic or viral origin or a mactivated or attenuated form of the pathogen, parasite or virus. The invention also comprises equipment for preparing these combination compositions, as well as methods for making these combination compositions and the use of the components of these combination compositions for preparing the combination compositions. Accordingly, the invention involves equipment for preparing the immunogenic or combination vaccine compositions of the invention; for example, such equipment comprising (a) an organism, pathogen or virus or antigen or epitope thereof (sale of a pathogen as mentioned herein) and (b) an organism, pathogen or virus or immunogen, antigen or epitope thereof (advantageously to virus or immunogen, antigen or epitope thereof, but other pathogens as mentioned herein are also contemplated) which is different from (a) in separate containers, optionally in the same package, and optionally with instructions for mixing and / or administration. Immunogenic compositions and / or vaccines according to the invention can include Eimeria culture or preparation (e.g., inactivated or attenuated Eimeria, or an immunogen or antigen or epitope thereof) and at least one immunogen, antigen or epitope of another. bird pathogen (including without limitation the pathogen in maimed or attenuated form). For multivalent avian immunogenic compositions and multivalent vaccines, e (the) additional bird pathogen (s) in that the additional bird antigen (s) or an epitope (s) for epitope (s) are included in and / or are expressed by the multivalent immunogenic compositions and multivalent vaccines are viruses, diseases or pathogens of the Marek's disease virus (MDV) (eg, serotypes 1 and 2, advantageously 1), viruses of Newcastle disease (NDV), paramyxoviruses other than Newcastle disease (PMV2 to PMV7), infectious bronchitis virus (IBV), infectious anemia virus or chicken anemia virus (CAV), infectious laryngotracheitis virus (ILTV) , infectious bursal disease virus (IBDV), encephalomyelitis virus or bird encephalomyelitis virus (AEV or ALV bird leukosi virus), turkey haemorrhagic enteritis virus (HEV), pneumovirosa virus (TRTV), virus of bird plague (avian influenza), hid virus chicken ropepcarditis, bird reovirus, coccidiosis, egg drop syndrome (EDS76), fowlpox, inclusion of hepatitis of the body (adenovirus), life-threatening lymphopro disease of turkeys, chicken reticuloendotheliosis, reticuloendotheliosis in turkeys, rotavirus enteritis and turkey rmotraqueitis , Clostridium um spp. , Eschepchia coli, Mycopla sma gallmarum, Mycoplasma ga llisepticum, Haemophilus avi um, Pa s teurella gallmarum, Pasteurella multilife gallicida, and mixtures thereof. Advantageously, for MDV the immunogen is advantageously gB and / or gD, for example, gB and gD, for NDV the immunogen is advantageously HN and / or F, for example, HN and F; for IBDV the immunogen is advantageously VP2; for IBV the immunogen is advantageously S (more advantageously Sl) and / or M and / or N, for example, S (or Sl) and M and / or N; for CAV the immunogen is advantageously VP1 and / or VP2; for ILTV the immunogen is advantageously gB and / or gD; for AEV, the immunogen sold is env and / or gag / pro, for example, env and gag / pro or gag / pro; for HEV the immunogen is advantageously the 100K protein and / or exon; for TRTV the immunogen is advantageously F and / or G, and for bird pest the immunogen is advantageously HA and / or N and / or NP, for example, HA and N and / or NP. Thus, the invention also involves methods for making these compositions, as well as equipment for them. An immunogenic composition or vaccine according to the invention which also comprises such additional immunogenic component (immunogen, antigen or additional epitope) has the advantage that it induces an immune response or protection against various infections or diseases or causative agents thereof at the same time . This additional immunogenic component can be an attenuated or inactivated microorganism, a recombinant construct or subunits, eg, proteins, glycoproteins, polappeptides or epitopes). Methods of epitope determination, such as the generation of superimposed peptide libraries (Hemmer et al., Immunology Today, 1998, 19 (4), 163-168), Pepscan (Geysen HM et al., Proc. Nat. Acad. Sci. USA, 1984, 81 (13), 3998-4002; Geysen H. M. et al., Proc. Nat. Acad. Sci. USA, 1985, 82 (1), 178-182; Van der Zee R. et al., Eur. J. Immunol. , 1989, 19 (1), 43-47; Geysen H.M., Southeast Asia J. Trop. Med. Public Health, 1990, 21 (4), 523-533; Multipm Peptide Synthesis Kits of Chiron) and algorithms (De Groot A. et al., Nature Biotechnology, 1999, 17, 533-561), can be used in the practice of the invention, to determine epitopes of immunogens, antigens, polypeptides, glycoproteins and the like, without undue experimentation. From that information, nucleic acid molecules encoding such epitope can be constructed, and from that knowledge and knowledge in the art, vectors or constructs can be constructed, for example viruses or recombinant plasmids or vectors that express immunogens, epitopes or antigens; all without undue experimentation. The pharmaceutically or veterinarily acceptable carriers or carriers or excipients are well known to one skilled in the art. For example, a carrier or excipient or pharmaceutically acceptable carrier can be a 0.9% NaCl solution (e.g., salt solution) or a phosphate buffer solution. The carrier or vehicle excipient is pharmaceutically or veterinarily acceptable can be any compound or a combination of compounds that facilitates the administration of the vector (or expressed protein of an inventive vector m vi tro); Advantageously, the carrier, vehicle or excipient can facilitate transfection and / or improve the preservation of the vector (or protein). Dosages and dose volumes are discussed herein in the general description of immunization and vaccination methods, and may also be determined by the skilled person from this description read in conjunction with knowledge in the art, without any experimentation. undue The immunogenic compositions and vaccines according to the invention may comprise or consist essentially of one or more adjuvants. Adjuvants suitable for use in the practice of the present invention are (1) polymers of acrylic or methacrylic acid, maleic anhydride or polymers derived from alkenyl, (2) immunostimulatory sequences (ISS), such as oligodeoxyribonucleotide sequences having one or more unmethylated CpG units (Klinman et al., Proc. Nat. Acad. Sd., USA, 1996, 93, 2879-2883; 098/16247), (3) an oil in water emulsion, such as the SPT emulsion described in p 147 of "Vaccine Design, The Subunit and Adjuvant Approach" published by M. Powell, M. Newman, Plenum Press 1995, and the emulsion MF59 described in p 183 of the same work, (4) cation lipids containing a salt of quaternary ammonium, (5) cytokines, (6) aluminum hydroxide or aluminum phosphate or (7) other adjuvants discussed in any of the cited and incorporated by reference in the present application, or (8) any of the combinations or mixtures thereof. The oil-in-water emulsion (3), which is especially suitable for viral vectors, can be based on: light liquid paraffin oil (European pharmacopoeia type), isoprenoid oil such as squalane, squalene, oil resulting from the oligomerization of alkenes, for example, isobutene or decene, acid esters or alcohols having a straight chain alkyl group, such as vegetable oils, ethyl oleate, propylene glycol, di (capplate / caprate), tri (capril ato / caprate) glycerol and propylene glycol dioleate or esters of alcohols or branched fatty acids, especially esters of isostearic acid. The oil is used in combination with emulsifiers to form an emulsion. The emulsifiers can be nonionic surfactants, such as: esters of on the one hand sorbitan, manuro (for example, anhydromanitol oleate), glycerol, polyglycerol or propylene glycol and on the other hand oleic, isostearic, picololic or hydroxystearic acids, esters that are optionally ethoxylated, or polyoxypropylene-polyoxyethylene copolymer blocks, such as Pluronic, e.g., L121. Among the polymers of adjuvants of type (1), preference is given to crosslinked acrylic or methacrylic acid polymers, especially crosslinked by polyalkenyl or sugar ethers or polyalcohols. These compounds are known under the name carbomer (Pharmeuropa, vol.8, No. 2, June 1996). One skilled in the art can also refer to US Pat. No. 2,909,462, which provides such acrylic polymers crosslinked by a polyhydroxyl compound having at least three hydroxyl groups, preferably not more than eight such groups, the atoms of hydrogen of at least three hydroxyl groups which are replaced by aliphatic, unsaturated radicals having at least two carbon atoms. Preferred radicals are those containing 2 to 4 carbon atoms, for example, rings, rings and other ethylenically unsaturated groups. The unsaturated radicals may also contain other substituents, such as methyl. Products sold under the name Carbopol (BF Goodrich, Ohio, USA) are especially suitable. They are crosslinked by allyl sucrose or by allyl pentaeptptol. Among these, the reference is made to Carbopol 974P, 934P and 971P. As for the polymers of maleic anhydride-alkene derivative, preference is given to EMA (Monsanto), which are straight-chain or cross-linked ethylene-maleic anhydride copolymers and are, for example, cross-linked by divmyl ether. Reference is also made to J. Fields et al., Nature 186: 778-780, June 4, 1960. With respect to structure, acrylic, methacrylic acid and EMA polymers are preferably formed by basic units having the following formula: in which; Ri and R2, which may be the same or different, represent H or Cll3 - x = 0 or 1, preferably x = 1 y = 1 or 2, with x + y = 2. For EMA, x = 0 and y = 2 and for carbomers x = y = 1. These polymers are soluble in water or physiological salt solution (20 g / 1 NaCl) and the pH can be adjusted to 7.3 to 7.4, for example, by soda (NaOH), to provide the adjuvant solution in which the expression vector (s) can be incorporated. The polymer concentration in the final vaccine composition can vary between 0.01 and 1.5% w / v, advantageously 0.05 to 1% w / v and preferably 0.1 to 0.4% w / v. The cationic lipids (4) containing a quaternary ammonium salt which are advantageously but not exclusively suitable for plasmids, are preferably those having the following formula: wherein Rx is an unsaturated saturated straight chain aliphatic radical having 12 to 18 carbon atoms, R2 is another aliphatic radical containing 2 or 3 carbon atoms and X is an amine or hydroxyl group. Among these cationic lipids, preference is given to DMRIE (N- (2-hydroxyethanol) -N, Nd? Met? L-2, 3-b? S (tetradecyloxy) -1-propane ammonium; WO96 / 34109), preferably associated with a neutral lipid, preferably DOPC (dioleoyl-phosphatidyl-ethanol amine; Behr JP, 1994, Bioconjugate Chemistry, 5, 382-389), to form DMRIE-DOPE. Advantageously, the mixture with the adjuvant is formed extemporaneously and preferably contemporaneously with the administration of the preparation or shortly before the administration of the preparation; for example, shortly before or prior to administration, the plasmid mixture of an adjuvant is formed, advantageously to give sufficient time before administration for the mixture to form a complex, for example between about 10 and about 60 minutes before administration, such as approximately 30 minutes before administration. When DOPE is present, the moral relationship DMRIE: DOPE is preferably from about 95: about 5 to about 5: about 95, more advantageously about 1: about 1, for example, 1: 1. The weight ratio of DMRIE adjuvant or DMRIE-DOPE: plasmid can be between about 50: about 1 and about 1: about 10, such as about 10: about 1 and about 1: about 5, and preferably about 1: about 1 and about 1: about 2, for example, 1: 1 and 1: 2. The cytokine or cytokines (5) may be in a protein form in the immunogenic or vaccine composition, or may be co-expressed in the host or in the immunogen or immunogens or epitope (s) thereof. The preference is given to the co-expression of the cytochem or cytokines, either by the same vector as that expressing the immunogen or immunogens or epitope (s) thereof, or by a separate vector therefor. The invention comprises the preparation of such combination compositions; for example, administering the active components, advantageously jointly and with an adjuvant, carrier, cytokine and / or diluent. Cytokines that can be used in the present invention, include, but are not limited to, granulocyte colony stimulation factor (G-CSF), granulocyte / macrophage colony stimulation factor (GM-CSF), interferon. a (IFN o1), interferon ß (IFN ß), interferon,, (IFN)), interleukin-l (IL-1 a), n nterleuc na na-1 ß (IL-I ß), interleukin- 2 (IL-2), inter! eucine-3 (1L-3), etherleucma-4 (EL-4), metleucine-5 (IL-5), etherleucine-6 (IL-6), etherleucine-7 (IL-7), methylleukin-8 (IL-8), nterleucin-9 (IL-9), methylleukin-10 (IL-10), nterleucine-11 (IL-1), in ter eucine-12 (IL-12), tumor necrosis factor a (TNL "a), tumor necrosis factor ß (TNF ß), and transforming growth factor ß (TGF ß). can be co-administered and / or sequentially administered with the immunogenic and vaccine composition of the present invention., for example, the vaccine of the present invention may also contain an exogenous nucleic acid molecule that expresses a suitable cytokine m vivo, for example, cytokine matched to this host to be vaccinated in which a molecular response will be induced (by example, a bird cytokine for preparations that are administered to birds). The invention also provides for the induction of an immune response or induction of an immune or protective response comprising administering an effective amount of the immunogenic or vaccine composition comprising a mixture of sporulated oocysts isolated from early strains of E. a cervulma, E. maximum and E. have it to induce or elicit the response in an animal. Advantageously, the animal is a bird such as, but not limited to, a chicken, duck, goose, guinea fowl, peacock, pheasant, pigeon, quail or turkey. In the much more advantageous mode, the bird is a chicken, advantageously a tender chicken for roasting. The method for inducing an immune response or inducing an immune response may also include the administration of an adjuvant, a cytokine or both. The amount effective to induce an immune response or induce an immunological or protective response is from about 10 to about 1000 E oocysts. to cervix, approximately 10 to approximately 100 E oocysts. maximum and approximately 10 to approximately 1000 E oocysts. tenel Advantageously, the amount effective to induce an immune response or induce a protective or immunological response is about 500 E. acervulin oocysts, about 50 to about 100 E. maximum oocysts and about 100 to about 500 E. tenella oocysts. More advantageously, the effective amount is about 500 E. acervulma oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts. In another embodiment, the effective amount is sufficient to withstand a stimulation dose of about 100,000 to about 500,000 E. acervulma oocysts and about 10,000 to about 1,000,000 maximum E. oocytes or about 10,000 to about 100,000 E. tenella oocysts of the animal. . More advantageously, the dose of stimulation is about 200,000 E. acervulma oocysts and about 20,000 to about 500,000 maximum E. oocytes or about 20,000 to about 50,000 E. tenella oocysts. The mixture may optionally comprise about 10 to about 1000 E. mitis oocysts, advantageously about 125 to about 500 E. mitis oocysts, about 125 E. mitis oocysts at a low dose, about 250 E. mitis oocysts at an average dose and approximately 500 oocysts of ---. mitis in a high dose.

The amount effective to induce an immune response and induce an immune or protective response is also related to specific relationships of sporulated oocysts isolated from the early strains of E. acervulina, E. maximus and E. tenella, where the ratio of E. acervulma , E. maximal and E. tenella, is approximately 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulma, there are approximately 1 to 2 sporocysts of E. maximus and approximately 2 to 10 of E. tenella). Advantageously, the ratio of E. acervulma, E. maximal and E. tenella is approximately 5: 1: 1 (ie, 10: 2: 2). In a modality containing E. mitis, the ratio of E. acervulina: E. maxim, E. milis: E. tenella is approximately 10: 1 to 2: 10: 2 to 10 (ie, for every 10 E sporocysts). acervulma, there are approximately 1 to 2 sporocysts of E. maximus, approximately 10 sporocysts of E. mitis and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: E. maximum: E. mitis: E. tenella is approximately 5: 1: 5: 1 (ie, 10: 2: 10: 2). Another aspect of the present invention is an immunization method or a method of vaccination using the immunogenic compositions or vaccine compositions according to the invention, respectively. The method includes at least one administration of an animal of an efficient amount of the immunogenic composition or vaccine according to the invention. The animal can be male or female. This administration can be not simply by intramuscular (IM), intradermal (and) (ID) or subcutaneous (SC) injection or by the intranasal or oral administration route, wherein oral administration includes but is not limited to administration over food or water to drink, gels or sprays. The immunogenic composition of the vaccine according to the invention can be administered by means of a syringe or a needle-free device (similar to for example Pigjet or Biojector (Bioject, Oregon, USA)). In an advantageous embodiment, the administration is oral. The compositions according to the invention can also be administered to other mammals, for example, mice or laboratory animals, for example to generate polyclonal antibodies or to prepare hybridomas for monoclonal antibodies. The present invention provides the immunization of animals, advantageously birds. Methods for administering coccidiosis vaccines are described in U.S. Patent Nos. 4,438,097; 4,639,372; 4,808,404; 5,055,292; 5,068,104; 5,387,414; 5,602,033; 5,614,195; 5,635,181; 5,637,487; 5,674,484; 5,677,438; 5,709,862; 5,780,289; 5,795,741; 5,814,320; 5,843,722; ,846,527; 5,885,568; 5,932,225; 6,001,363 and 6,100,241, the descriptions of which are incorporated by reference in their totalities. The method comprises administering the animal an effective immunizing dose of the vaccine of the present invention. The vaccine can be administered by any of the methods well known to those skilled in the art, for example, by intramuscular, subcutaneous, intrapeptoneal, intravenous, intranasally, orally, mdermic, mtrabursal injection, just above chick ventilation), m ovo, o ocularly. The methods of administration are known to those skilled in the art. For example, U.S. Patent Nos. 5,693,622; 5,589,466; 5,580,859; and 5,566,064 are hereby incorporated by reference in their entirety. Birds can also be administered with vaccines in a spray cabinet. The birds may also be administered in the m ovo vaccine, as described in U.S. Patent Nos. 4,458,630 and 6,627,205, the descriptions of which are incorporated by reference. Advantageously, the birds are administered with vaccines in a spray cabinet, that is, a cabinet in which the birds are placed and exposed to a vapor containing vaccine, or by spray in progress. In another advantageous modality, the immunogenic or vaccine composition is administered orally. Alternatively, the immunogenic or vaccine composition can be administered in the drinking water or in the food. The invention encompasses an immunogenic composition or vaccine comprising a mixture of sporulated oocysts of early strains of E. acervulma, E. maximus and E. tenella. The sporulated oocysts of early strains of E. acervulina, E. maxim and E. tenella are isolated from the seed cultures described herein. Generally, the dose range of sporulated oocysts in the composition is from about 10 to about 1000 E. acervulma oocysts, about 10 to about 100 E. maximum oocysts and about 10 to about 1000 E. tenella oocysts. Advantageously, the dose range of sporulated oocysts in the composition is from about 125 to about 500 E. acervulma oocysts, about 25 to about 100 E. maximum oocysts and about 25 to about 500 E. tenella oocysts. In one embodiment, a low dose is approximately 125 oocysts of E. acervulma, or approximately 25 oocysts of E. maximus and approximately 25 oocysts of E. tenella. In another embodiment, an average dose is approximately 250 oocysts of E. acervulma, approximately 50 maximum oocysts and about 50 E. tenella oocysts. In yet another embodiment, a high dose is about 500 E. acervulin oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts. Advantageously, the dose of the immunogenic or vaccine composition is about 500 E. acervulin oocysts, about 50 to about 100 E. maximum oocysts and about 100 to about 500 E. tenella oocysts. In a more advantageous embodiment, the dose is about 500 E. acervulin oocysts, about 100 E. maximum oocysts and about 100 E. tenella oocysts. The mixture may optionally comprise about 10 to about 1000 E. mitis oocysts, advantageously about 125 to about 500 E. milis oocysts, about 125 E. mitis oocysts in a low dose, about 250 E. mitis oocysts. in a medium dose and approximately 500 oocysts of E. mitis in a high dose. The invention also relates to specific ratios of sporulated oocysts isolated from early strains of E. acervulma, E. maximus and E. tenella in a dose of the immunogenic or vaccine composition, wherein the ratio of E. acervulina E. maximal E. it is about 10: 1 to 2: 2 to 10 (ie, for every 10 sporocysts of E. acervulma, there are approximately 1 to 2 sporocysts of E. maximus and about 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: E. maxim, E. tenella is approximately 5: 1: 1 (ie, 10: 2: 2). In a modality containing E. mitis, the ratio of E. acervulma: E. maxim: E. mitis: E. tenella is from about 10: 1 to 2: 10: 2 to 10 (ie, for every 10 sporocysts of E. acervulma, there are approximately 1 to 2 sporocysts of E. maximus, approximately 10 sporocysts of E. mitis and approximately 2 to 10 sporocysts of E. tenella). Advantageously, the ratio of E. acervulma: - .. maximum: E. milis: E. tenella is approximately 5: 1: 5: 1 (ie, 10: 2: 10: 2). By sale, the oocysts are suspended in a preservative consisting of a 0.1 M phosphate buffered saline solution containing gentamicin. In another embodiment, the oocysts are suspended in any of a variety of preservatives or organic acids such as, but not limited to, acetic acid, citric acid, potassium dichromate or propionic acid. For example, but not by limitation, sufficient saline regulated 0.01 M sterile phosphate solution containing no more than 30 mcg / ml of gentamicin, is used to produce 2 ml per bottle for a presentation of 2, 000 doses, 5 ml per bottle for a presentation of 5,000 doses and 10 ml per bottle for a presentation of 10,000 doses. Advantageously, the oocysts are stored in sterile borosilicate glass vials.

For example, but not by limitation, the oocysts are filled aseptically in vaccine bottles with a semi-automatic or automatic supplier, the caps are inserted mechanically or manually and aluminum seals are placed and folded. The vaccine is marketed as a multiple dose containing 2,000-dose vials, 5,000-dose vials, 10,000-dose vials, or 20,000-dose vials. The expiration date of the product shall not exceed 13 months from the date of initiation of the power test. The animals, advantageously birds, can be vaccinated at any suitable age, and usually are approximately one to three days of age before the first vaccination. Advantageously, the animals are vaccinated once. Optionally, when two doses of vaccine are used, the first is usually given when the animals are 3 days to a week old and subsequently after an additional 1-10 weeks depending on the type of animal being vaccinated. The use, dosage and route of administration for each species of animal in an advantageous embodiment is as follows. The immunogenic or vaccine composition of the present invention was used for the vaccination of healthy day-old or older chickens, an aid in the prevention of the disease due to coccidiosis. Advantageously, the dosage was one dose per chicken by the running water spray of 20 ml per 100 chickens. The invention will now be described further by means of the following non-limiting examples. EXAMPLES Example 1: Production of coccids Composition of the product. The microorganisms that can be used are Eimepa a cervulma, Eimepa maximus, Eimeria mi ti s and Eimeria tenella. The origins of the extract cultures of the microorganisms are as follows. The origin strain of the Eimeria a cervul ma was obtained from T. K. Jeffers and collaborators Hess and Clark Laboratories in 1969 and is believed to have been isolated by Dr. M. Farr at USDA, Beltsville, MD, which was derived from a single oocyst. The maximum Eimepa culture was derived from an interproduction mixture of 10 purified isolates obtained from Georgia, Delaware, Maryland, Virginia and Texas. The strain of origin of the Eimepa mi tis culture was isolated from Gainesville, Georgia in July 1978 and purified by a single oocyst isolate. The strain of origin of the Eimeria tenella culture was obtained from a crop maintained at Pennsylvama State University by Dr. Patten from the early 1960s, and was acquired by the University of Georgia in 1982. Each of the Eimeria strains used was a precocious strain of the respective microorganism as described in Avian Pathology, 17: 305-314, 1988 entitled " Eimepa by American Chickens: Charactepstics of Six Attenuated Strains Produced by Selection for Precocious Development, PL Long and Joyce K. Johnson, the description of which is incorporated by reference in its entirety.The mechanisms were attenuated by their selection for the early development of According to this method, each Eimepa species used in the product was identified by its unique microscopic appearance, size and shape, and the area of the intestine or cecal area of infected chicken, as described in Long, PL and M. Reid, A Guide for the Diagnosis of coccidiosis in chickens Re. Report 404 (Report 355 revised), Athens, GA: College of Agriculture Experiment Station, Univ. Of Georgia; ription of which is incorporated by reference in its entirety. The microorganisms were attenuated by their selection for early development as described above. Each culture was pathogen free by testing using the procedures described in 9 CFR 113.37. The extract cultures described in the above were maintained in the liquid or vapor phase of liquid nitrogen. The chickens, from 2 to 8 weeks of age, were used for the production of seed or seed crops. The administration route was per os (ie, orally). All seeds were sporulated oocysts and were produced in SPF chickens, from 2 to 8 weeks of age. The dedicated facilities were maintained for each Eimeria species. A sufficient volume of sporulated oocysts (seed) was mixed with the food, free of anticoccidials, or administered by oral priming to provide each chicken with a minimum dose listed in Table 1. The sporulated oocysts were passed successively, without limitations to the passage. , in SPF chickens from 2 to 8 weeks of age until the number of oocysts was sufficient to be used as seed for production. Crops can not be maintained for longer than 12 months in order to maintain viability / infectivity. Table 1: Minimum dosage of sporulated oocysts (seed) The defecations were collected daily from the third to the eighth day after the inoculation. The random chickens were sacrificed and observed for the characteristic infection of each species other than my tis. No collection was done if there was any evidence of a strange disease.

The extract cultures for reference were kept in liquid nitrogen. The production cultures were maintained at 5 ± 3CC and passed into SPF chickens for two to eight we The preparation of suspensions for the seed or inoculation involved the serial passage of seed cultures in SPF chickens, from two to eight weof age, until enough oocysts were produced for the manufacture. A sufficient volume of sporulated oocysts (seed) was mixed with the food, free of anticocciodials, and was administered to provide each chicken with the minimum dose listed in Table 1. The defecations were collected as a bath from the third to the eighth day after of the inoculation. Random chickens were sacrificed and observed for the characteristic infection for each species with the exception of my ti s. No collection was done if there was any evidence of a strange disease. The microorganisms are attenuated by their selection for early development as described in the above. Harvest . Before the removal of the microorganisms, for production purposes, the chickens were housed in a room designated for the production of individual species. The chickens were kept on a diet free of anticocciodiales and were left with free access to water. The defecations were collected daily from the third to the eighth day after the inoculation, and were maintained at 5 ± 3 ° C until they were processed. The technique of collecting microorganisms for production purposes were collected as follows. The defecations were homogenized in an approximate ratio of 10% (p / v) in water. The long particles were removed by passing the homogenate through screens. The solids were separated either by centrifugation, screening or by holding 5 ± 3 ° C for up to 24 hours. If the solids were separated by holding at 5 ± 3 ° C, they were further concentrated by centrifugation. The supernatant was discarded, and the solids were resuspended in saturated NaCl solution (80% w / v) in water. The resulting solution was centrifuged. The oocysts were collected (removed) from the upper part of the liquid, and resuspended in water. Optimally, the remaining liquid was diluted to 20-40% NaCl with water and centrifuged. The pellet was then resuspended in a saturated NaCl solution and centrifuged, until no additional oocysts were recovered. The oocysts were washed no more than twice. The oocysts were washed free of salt by centrifugation cycles, repeated resuspension followed by resuspension of a 0.5% sodium hypochlorite solution from 10 to 15 minutes. The oocysts were then washed free of the sodium hypochlorite solution by repeated centrifugation (3X) and the resuspension steps. The final resuspension was done in a 2.5% aqueous solution of potassium dichromate (K2Cr07). The oocysts were then transfected into sporulation vessels. Sporulation was facilitated by dispersing the suspensions with air for a period not exceeding 72 hours at 27 ± 3 ° C. After sporulation the oocysts were maintained at 5 ± 3 ° C until the final product was produced. In an advantageous embodiment, the specifications for an acceptable collection were as follows. First, the ratio of sporulated oocysts to total oocysts is determined. Only collections that meet or exceed sporulation > 40% were considered acceptable. Second, the size, shape and appearance of each oocyst collection must be characteristic of the species proposed to be produced. Discarded material not used in production was disposed of in a manner consistent with 9 CFR 114.15. Preparation of the product. Series or subseries of final product were combined with the collection fluids in volume. All oocysts were attenuated, in that they are precocious, as it is described in the previous. The oocysts were resuspended in a preservative consisting of a 0.01 M phosphate buffered saline solution containing no more than 30.0 mcg of gentamicin, the product was standardized to produce at least final numbers of sporulated oocysts of each species as indicated in Table 2. Table 2: Standardized dosage of sporulated oocysts A series was assembled as follows. The bulk suspension lots of each of the four Eimeria species are removed from storage at 5 ± 3 ° C and allowed to warm to room temperature while stirring. A sufficient volume of each lot (s) was removed aseptically and removed and combined, to produce no less than the oocysts per doses listed in Table 2 as determined by calculations based on counts of each of the suspensions by volume. If necessary, multiple batches of each Eimeria species were combined to provide the required number of sporulated oocysts. Sufficient regulated saline solution of 0.01 M sterile phosphate containing more than 30 mcg / ml of Gentamicin, to produce 2 ml per bottle for a presentation of 2,000 doses, 5 ml per bottle was used for a presentation of 5 ml dose and 10 ml per bottle for a presentation of 10,000 doses. The series in volume was kept under constant agitation.

Table 3: Example of the volume in series (based on a batch of 10,000,000 doses of the product of 10,000 doses) An average of the 10,000 dose series varies from 20L to 40L. The filling volume is 1.0 ml per 1000 doses. Vaccine bottles are borosilicate glass bottles, sterile. The method and technique of filling and sealing the final containers were as follows. The vaccine was aseptically taken in vaccine bottles with a semiautomatic or automatic supplier. The plugs were inserted mechanically or manually. The aluminum seals are placed and folded. Each dose contained no less than the dose listed in Table 2 of sporulated oocysts, as calculated from the volume counts. A serial number was assigned to each quantity per batch and presented in a single operation. Proof . Each series was tested for purity using the test methods described in 9CFR 113.27 (e). The average count in the incubation condition should not exceed 1 colony per dose. If the average count in any incubation exceeded one colony per dose in the initial test, a test again was optionally conducted using 20 unopened final containers. If the average count of any incubation condition of the final test of one colony per dose, the series was unsatisfactory. Sa lmonel la was shown to be effectively exterminated by both sodium hypochlo- prate and potassium dichromate, therefore, no additional test was conducted. Mycoplasma was shown to be effectively killed by sodium hypochlorite, therefore, no additional test was conducted. Each series was tested for foreign pathogens in accordance with 9 CFR 113.37. To test the safety of the vaccine, each of the 25-day-old specific pathogen-free chickens was vaccinated by spray vaccination with 10 doses of vaccine. The chicks were observed every day for 21 days. The chickens that died during the period were examined and the cause of death was determined. If at least 20 chickens did not survive the observation period, the test was inconclusive. If any disease or death was directly attributable to the vaccine, the series is unsatisfactory. The average intestinal lesions characteristic of the vaccine were considered normal, and are not considered a safety assessment. If less than 20 chickens survived the observation period there were no deaths or severe injuries attributable to the vaccine, the test was optionally repeated once. If the test is not repeated, the series was declared unsatisfactory. The first series of final product produced from each new seed batch of production will be tested for potency. The product will be approved in SPF chickens or broiler chickens from 1 to 14 days of age. Subsequent series produced from the seed of production will be evaluated for potency using preformulation counts specified in the above and in the recovery of oocysts from inoculated birds. No more than 70 chickens were used. No more than 35 were vaccinated per os with a dose in the field of vaccine. Twenty-six to thirty days after the initial vaccination all the chicks were individually identified and the weights recorded. No more than 10 vaccinates and 10 controls of each of the groups listed below, were stimulated per os, with 1.0 ml of each of the doses of stimulation as shown in the Table. Table 4. Stimulation dose Group 1 oocysts E. maximum 10,000 to 1,000,000 oocysts Stimulation of E. my ti s 100,000 to 500,000 Group 2 oocysts Stimulation of E. tenel the 10,000 to 100, 000 Group 3 oocysts The stimulation doses were also selected on the basis of pathogenicity in which the selected level will give a minimum record of at least two. Six days after the stimulation, the vaccinates and controls of each of the Groups in Table 4 were sacrificed, weighed and the intestines and cecal area were examined and recorded for the lesions. The record was in accordance with the coccidial registration system of Johnson and Reid as described in Experimental Papsitology, Vol. 28, p 30-36, 1970, the description of which is incorporated by reference in its entirety, as shown in Table 5. Table 5: Injury record Post-preparatory stages. The vaccine is marketed as a multiple dose containing, vials of 2000 doses, vials of 5000 doses, vials of 010,000 doses or vials of 20,000 doses. The collection, storage and presentation of representative samples were in accordance with 9 CFR 113.3. The expiration date of the product did not exceed 13 months from the date of the power test initiation, and was confirmed in accordance with 9 CFR 114.13. The use, dosage and route of administration for each species of animal was as follows. This product was used for the vaccination of healthy chickens 1 day of age or older, as an aid in the prevention of disease due to coccidiosis. The dosage was one dose per chicken by coarse water spray; 20 ml per 100 chickens. Example 2: Efficacy of the Experimental Vaccine Containing Attenuated Coccidial Strains of Birds Compared to a Vaccine of Commercial Live Coccidiosis and Salmomycin, an Anticoccidial Drug in Commercial Poultry. The objective of this Example was to determine the efficacy of the attenuated Eimeri oocysts vaccine, compared with a life-saving oocysts vaccine approved by the USDA (Coccivac-B) for the protection of roast-type chickens from stimulation with virulent Eimepa species and to compare the use of the attenuated vaccine with a conventional anticoccidial drug, Salmomycin. The objective variables were as follows. The primary variable was the intestinal lesion records. The acceptance criteria were lower records of post-stimulation coccidial bowel choice when compared to the stimulated, unvaccinated control birds. Secondary variables were (1) weight gain and feed conversion during a 7-week growth phase and (2) mortality. Acceptance criteria included equal or greater weight gain and feed conversion in birds vaccinated against birds vaccinated with CoccivacB or medicated with the anticoccidial drug and low mortality in vaccinated birds against unvaccinated control birds, birds vaccinated with CoccivacB and birds treated with anticoccidial. Materials . The vaccines are described in Table 6. Table 6: Vaccines Table 7: Virulent Organisms / Stimulation Table 8: Animals The animals were handled in a similar way and with due respect to their welfare. The animals were handled in accordance with all applicable regulations. Experimental design. The experimental design was as follows. Table 9: Experimental treatment groups * 20 ml supplied by 100 chickens The timeline was as follows. On Day 0, the birds were weighed per pen. Birds in Groups 1 and 2 were vaccinated as described in Table 9, with 1300 birds per vaccine regimen, divided between 2 pens, one in each half of the housing. Birds in Group 3 were started on Salmomycin, 60g / ton, divided as described by Groups 1 and 2. The pens were closed. Extra hatchmates were retained in wire cages as unvaccinated chickens. On Day 21, the feed of the starter to the grower was changed for all the birds. Salinomycin was continued in Group 3. On Day 28, the birds to be stimulated were moved to wire cages. Salmomycin was discontinued in Group 3 to birds that are stimulated. All the birds and the food were weighed. Fresh fecal samples were collected from the bed of each group in the large pens, 20 samples per pen. The oocysts were quantified by species. On Day 29, the birds in the cages were stimulated using the number of oocysts described in Table 7. On Day 35, the stimulation birds were terminated, the lesions were recorded and the weight gains were taken. On Day 42 all groups were switched to the non-medicated terminator feed and the feed was weighed again. On Day 49, all the birds and feed were weighed and the experiment was finished. Experimental procedures Vaccination / Medication. Three treatments of birds identified as Groups 1-3 in Table 9 were replicated in half of houses, three pens per house, 650 birds per pen, for a total of 1300 birds per treatment. Birds in Groups 1 through 3 were vaccinated as described in Table 9. An additional treatment group (4) was kept separately in clean cages as untreated, unvaccinated controls for use in the stimulation test only. Treatment Groups 1, 2 and 4 were given non-medicated food. Treatment Group 3 was given Salinomycin (60 g / ton) starting on Day 1 until the day 42. The birds in Group 3 to be stimulated were given unmedicated feed after moving to the stimulation cages. The starter food was given until 21 days, the grower up to 42 days and the terminator up to 49 days. The food issued was recorded, and any remaining uneaten food was passed again at the time of feeding changes and termination.

Observations. The birds were weighed by pen in 0 and 28 days and by pen and sex in 49 days. The dead birds were collected twice a day and the necropsy was performed to determine the cause of death. Fecal samples collected on Day 28 were observed for viable oocysts and quantified by species. Stimulation On Day 28, 3 reps of 10 birds from each pen (total of 60 per treatment) were weighed and moved to the battery cages for stimulation with Eimeria field strains (corresponding to the species used in the vaccine). All the birds were given non-medicated feed in the cages. On Day 29, the birds of group 1-4 were stimulated and then finished on Day 35. The records of injury and weights were recorded. The animals were randomly assigned to each treatment group by randomly collecting them from the chicken boxes before vaccination. The scientists were not blinded to the treatment groups since strict safety was maintained between the groups to prevent cross-contamination. Concurrent Medications and Management of Adverse Events. The birds vaccinated with Coccivac became ill 14 days in the experiment. The diagnosis made was necrotic enteritis. The responsible agent was identified as Clos tridi um sordell i. All the birds in the experiment were administered with Penicillin G to reduce the contamination, and the effects of the same, between the pens and to eliminate any difference in the treatments between the groups that could present the results of coccidiosis. Data Analysis Criteria for Measurement. The primary variables for the floor corral phase were live weights in 28 and 49 days and the feed conversion to 49 days. For the stimulation phase, the primary variables were weighed again six days after the stimulation, and the injury records of the upper, middle and cecal sections, also at six days after the stimulation. Statistic analysis. All statistical analyzes were conducted using SAS, Gary, NC (Version 8.2). The statistical significance was based on tests of two extremes of the null hypothesis that were examined in this study, resulting in p-values of 0.05 or less. Intestinal Lesion Registry. The incidence and severity (registers of categorical lesion) of post-stimulation intestinal coccidial lesions were analyzed using a logit model with factors of the treatment group and the analysis model of blockade and / or survival with factors of the treatment group and blockade (depending on of the nature of the data).

Mortality. If the mortality related to the stimulation of the virulent Eimeria species occurred, an ANOVA (ie, an analysis of variance) with factors from the treatment and blockage group was conducted to determine whether significant differences existed between the means of fetal mortality (without consider the day of occurrence) for each treatment group. Weight Gain and Food Conversion Phase of the Floor Corral. Within the treatment groups, paired t-tests (by blockage and as a whole group, with no blocking effect considered) were conducted to determine if there are significant differences in average live weights within each treatment group. Paired comparisons included weights from day 0 against Day 28 (pre-stimulation) weights from Day 28 against Day 49 (post-stimulation). Paired t tests (by blockage as a whole group, without considering blocking effect) were conducted to determine if there are significant differences in the weights of the average feed conversion within each treatment group. Paired comparisons included the feed conversion weights of Day 28 versus Day 49 (post-stimulation) and feed conversion weights of Day 42 versus Day 49 (effect of the unmedicated terminator feed).

Among the treatment groups, an ANOVA of repeated measurements with factors of treatment group, day, block and group-day interaction was used to determine if there were different signifiers in the average live weights between the groups, especially the weights of the groups. days were recorded. Stimulation phase. Within the treatment groups, paired t tests (by blockage and as a whole group, without considering the blocking effect) were conducted to determine if there were significant differences in the average live weights within each treatment group. Paired comparisons included the weights of Day 28 versus Day 35 (6 days post-stimulation). Among the treatment groups, the analysis of the live weights between the treatment groups, post-stimulation, were conducted using the results of the analysis of the treatment group for the floor corral phase, focusing the comparisons on Day 28 and Day 35. Results * Means statistical differences by grouping Table 11: Summary of Day 49 Table 12. Stimulation Results 60 birds of stimulation per treatment. * It means statistical differences by grouping. a, b, c and d are Duncan's separation from the means, (Duncan's test is a statistical hoc test to separate the means). 1 is the total injury record 2 is the average total injury record Discussion. There was no significant difference in injury records for the experimental vaccine and Coccivac for E stimulations. maximum, E. to cervulina and E. my tis Coccivac had records of lower lesions of E. tenella as compared to the experimental strains. It is important to note that birds vaccinated with Coccivac became infected with Cl ostridi um sordell i. Clostridial disease has been linked to an infection with coccidiosis in the field. Although the infection was dispersed to the other treatment groups, birds vaccinated with Coccivac were the most affected, showing 11.77% mortality. Conclusions Experimental Eimeria strains were shown to be effective in the face of virulent stimulation. The birds vaccinated with the experimental strain have better bird performance as defined by the feed conversion ratios when compared to the birds treated with Coccivac and Salinomycin in 49 days which corresponds to an economic advantage to the poultry producer. Example 3: Live m potency test for the Coccidiosis Vaccine, Live Oocysts, Chicken Origin. This example presents a procedure to demonstrate the potency of the collection material of each new Eimeria production seed, as formulated in the final product. Materials for this Example included chicken feed, free of anticoccidials, SPF or roast type chickens 1-14 days of age and chicken housing. The first series of final product produced and each subsequent series produced using a new batch of seed or production seeds are tested for potency. The first series produced, which contains all the new seeds of production are stimulated with each species. For subsequent series, which contain one or more new seeds of production, only those antigens representing the new seeds are tested for potency as described below. A final product is tested on SPF-type chickens or for broiler 1-14 days of age. Subsequent series produced from the production seed are evaluated for power using preformulation counts specified below. The power of E. acervulina, E maximum and E. They are evaluated singularly or in combination, in the same group of animals. For each strain tested, no more than 24 chickens are used. At least 10 and no more than 12 are individually vaccinated but with a dose of vaccine in the field. Ten to 12 chickens will remain as controls. Twenty-six to 30 days after the initial vaccination all the chicks are identified individually. Each stimulation strain used in the potency test must have a control group of at least 10 chickens. At least 10 vaccinates and 10 controls of the applicable group listed below are individually stimulated per os, with 1.0 ml of stimulation or homologous stimulations in the dosage listed in Table 13 Table 13. Stimulation The dose of stimulation is also selected on the basis of the pathogenicity of Eimeria, in which the selected level will cause at least 50% of the unvaccinated stimulation controls to have an injury record of two or more high. No group of vaccinates or controls can have less than eight animals. The record is in accordance with the coccidial registration system of Johnson and Reid as described in Experimental Parisitology, Vol. 28, p 30-36, 1970. Table 14. Registration Six days after the stimulation, the vaccinates and controls are sacrificed, and the intestine and cecal area is examined and recorded for individual lesions according to Eimeria species and the section corresponding to the intestine that correlates with an infection, ie , E. a cervulina affects the upper intestine, E. maximum infects the middle section of the intestine and E. tenella infects the cecal area. The data is analyzed so that a significant difference between the vaccinated and controls can be determined at the 0.05 level of significance using the Mann-Whitney U-Test. For each species of Eimeria stimulated for potency, the total of the respective individual injury record for each of the vaccinated and controls. Each bird receives a record for E. maximum, a record for E. to cervulina and a record for E. tenella. The classification records of all animals from the smallest to the largest. For identical classifications, the allocation to the average of each animal, for example, if the 5th and 6th animals have an identical record, assigning a rating of 5.5 to each. The retention of the identity of the classification as soon as it is a vaccinated or control. The total of the classification numbers of the control group. Called this total "SRc.dd" The total of the classification numbers of the vaccinated group. Called this total "SRV." The determination of the sum of the classifications for the control group, the Mann-Whitney U Statistics of the U (Uc) controls is calculated using the following equation: Where nc represents the number of animals in the control group and nv represents the number of vaccinated animals. In determining the sum of classifications for the vaccinated group, the Mann-Whitney U Statistic of the controls (Uv) is calculated using the following equation: + nv (n, -f- l) _ ^, R u "= nvnc Where nc represents the number of animals in the control group and nv represents the number of animals vaccinated. The significant test of the level 0.05 if ÜJ is less than Uv, and ÜJ is less than or equal to the critical value listed in Table 15 for the appropriate group sizes. Table 15. Table of Critical values for the Mann-Whitney U statistic (Values 0.05 of Two Extremes) Validity criteria. For a valid test, at least 50% of unvaccinated stimulation controls must have the injury record of two or more high. No group of vaccinates or controls can have less than eight animals. If the animals meet the validity requirements and the calculated Mann-Whitney U statistic is significantly different from the 0.05 level, the series is satisfactory. Re-test requirements.- If a vaccinated group fails to show a significant difference (the statistics Uc calculated is greater than the critical values listed in Table 15) of the control group, the results of the test are inconclusive and the series can be rejected. To determine if the series was not satisfactory due to overstimulation, the injury records of the vaccinates from the first test session and the second test session are analyzed to determine if a significant difference between the sessions can be ascertained at the level 0.05 of meaning using the Mann-Whitney U test. Individual injury records for strains of the vaccinated are totaled from both test sessions. The classification records of all animals from the smallest to the largest. For identical classifications, the average is assigned to each animal, for example, if the 5th or 6th animal has an identical record, a classification of 5.5 is assigned to each one. Detention of the identity of the classification as to whether it is from the first or second test session. The total of the classification numbers of the first test session. Called this total "S RJ 'The total classification number of the second session of the test, called this total" SR2. "In the determination of the sum of the classifications for the vaccinated of the first test session, the U statistic The Mann-Whitney of the first test session (Ui) is calculated using the following equation: In determining the sum of the classifications for the vaccinated group from the second test session, the Mann-Whitney U statistic of the controls (Uv) is calculated using the following equation: Where neither represents the number of animals of the vaccinated in the first test section and n2 represents the number of vaccinated in the second test session. Interpretation of the results of the test. The test is significant at the 0.05 level if Ui is less than U2, and U2 is less than or equal to the critical value listed in Table 16 for the appropriate group sizes. Table 16. Table of Critical values for the Mann-Whitney U statistic (Values 0.05 of Two Extremes).

If the results of the comparison of the first and second test sessions is significant at a level of 0.05, the initial result should be considered a "No test" due to overstimulation, and the vaccinated and controls will be compared using the data generated from the second test session. The data analysis will be conducted as it is previously summarized. If the comparison of the two test sessions fails to show significance at the 0.05 level, the data from both test sessions will be combined, and the data will be analyzed as summarized previously. The test is significant at the 0.05 level if Uc is less than Uv, and Uc is less than or equal to the critical value listed in Table 17 for the appropriate group sizes. Table 17. Table of Critical Values for Mann-Whitney U Statistics (Series Retest) Reference: Sheskin, David J. Handbook of Parametric and Nonparametric Statistical Procedures, 3rd ed. 2003; 423-428, 1151. * * * Having described in this manner in detail the advantageous embodiments of the present invention, it is to be understood that the invention will not be limited to the particular details set forth in the foregoing description since many obvious variations thereof are possible without departing from the spirit. or scope of the present invention.

Claims

CLAIMS 1. An immunogenic or vaccine composition for protection against E. acervulma, E. maxim and E. tenella, characterized in that it comprises a pharmaceutically acceptable excipient and a mixture of sporulated oocysts isolated from early strains of E. acervulma, E. maxim and E. tenella, wherein the mixture consists essentially of about 500 E. acervulma oocysts, about 50 to about 100 E. maximum oocysts and about 100 to about 250 E. tenella oocysts.
2. The composition according to claim 1, characterized in that the mixture consists essentially of about 500 oocysts of E. acervulma, about 100 oocysts of E. maximus and about 100 oocysts of E. tenella.
3. The composition according to claim 1, characterized in that the mixture consists essentially of about 500 oocysts of E. acervulma, about 50 oocysts of E. maximus and about 250 oocysts of E. tenella.
4. An immunogenic or vaccine composition for protection against E. acervulma, E. maxim and E. tenella, characterized in that it comprises a pharmaceutically acceptable excipient and a mixture of sporulated oocysts, wherein the mixture of sporulated oocysts consists of sporulated oocysts isolated of early strains of E. acervulina, E. maximum and E. tenella, where for every 10 sporocysts of E. to cervulina, there are approximately 1 to 2 sporocysts of E. maximum and approximately 2 to 10 sporocysts of E. tenella.
5. The composition according to claim 4, characterized in that for every 10 sporocysts of E. to cervulina, there are approximately 2 sporocysts of E. maximum and approximately 2 sporocysts of i-., tenella.
6. A method for inducing an immune response, characterized in that it comprises administering an effective amount of the immunogenic or vaccine composition according to claim 1 to induce the response in a chicken.
7. A method for inducing an immune or protective response, characterized in that it comprises administering an effective amount of the immunogenic or vaccine composition according to claim 1 to induce the response in a chicken.
8. A method for inducing an immune response, characterized in that it comprises administering an effective amount of the immunogenic or vaccine composition according to claim 4 for inducing the response in a chicken.
9. A method for inducing an immune or protective response, characterized in that it comprises administering an effective amount of the immunogenic or vaccine composition according to claim 4 for inducing the response in a chicken.
10. The method of compliance with the claim 6, characterized in that the effective amount is about 500 oocysts of E. acervulin, approximately 50 to approximately 100 E oocysts. maximum and approximately 100 to approximately 250 E oocysts. tenella.
11. The method according to the claim 7, characterized in that the effective amount is about 500 oocysts of E. acervulina, about 50 to about 100 oocysts of E. maximum and approximately 100 to approximately 250 E oocysts. tenella.
12. The method according to claim 6, characterized in that the effective amount is about 500 oocysts of E. to cervulina, approximately 100 oocysts of E. maximum and approximately 100 oocysts of E. tenella. The method according to claim 7, characterized in that the effective amount is about 500 E oocysts. to cervulina, approximately 100 oocysts of E. maximum and approximately 100 oocysts of E. tenel The method according to claim 8, characterized in that the effective amount of the sporulated oocysts mixture, wherein for every 10 sporocysts of E. to cervus, there are approximately 2 sporocysts of E. maximum and approximately 2 sporocysts of E. tenella in the mixture. 15. The method according to claim 9, characterized in that the effective amount of the mixture of sporulated oocysts, wherein for every 10 sporocysts of E. to cervulin, there are approximately 2 sporocysts of E. maximum and approximately 2 sporocysts of E. hold it in the mix. 16. The method according to claim 6, characterized in that the effective amount is sufficient to withstand a stimulation dose of about 100,000 to about 500,000 E oocysts. to cervix and approximately 10,000 to approximately 1,000,000 E oocysts. maximum or approximately 10,000 to approximately 100,000 E oocysts. hold the animal. The method according to claim 7, characterized in that the effective amount is sufficient to withstand a stimulation dose of about 100,000 to about 500,000 E oocysts. to cervix and approximately 10,000 to approximately 100,000 E oocysts. maximum or approximately 10,000 to approximately 100,000 E oocysts. hold the animal. 18. The method of compliance with the claim 8, characterized in that the effective amount is sufficient to withstand a stimulation dose of approximately 100,000 to approximately 500,000 E oocysts. to cervulina and approximately 10,000 to approximately 1,000,000 of E oocysts. maximum or approximately 10,000 to approximately 100,000 E oocysts. Tenel the animal. 19. The method according to the claim 9, characterized in that the effective amount is sufficient to withstand a stimulation dose of about 100,000 to about 500,000 oocysts of the cervix and about 10,000 to about 1,000,000 of E oocysts. maximum or approximately 10,000 to approximately 100,000 E oocysts. hold the animal.