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Apicomplexan parasites of the genus Eimeria are widespread in poultry flocks and can cause the intestinal disease coccidiosis. Early studies, concerned with intraspecific variation in oocyst morphology, indicated that phenotypic changes may be induced by selection experiments conducted in vivo . Genetic selection driven by targeted selection for specific phenotypes has contributed to our understanding of the phenomenon of drug resistance and the development of live attenuated vaccines. Our present knowledge regarding genetics of Eimeria is largely based upon the utilization of such selected strains as genetic markers. Practical advantages of working with Eimeria spp. in the chicken are discussed. The selection of drug-resistant strains by serial propagation has provided useful information regarding the mechanisms of drug resistance and likely longevity of anticoccidial drugs when introduced in the field. Selection experiments to develop precocious strains of Eimeria and growth in chicken embryos have contributed to the development of safe and effective live attenuated vaccines for the control of coccidiosis. Establishment of protocols for genetic complementation by transient or stable transfection of Eimeria is now supporting direct manipulation of parasite genotypes, creating opportunities to expand the range and value of live parasite vaccines. Procedures for developing drug-resistant and precocious lines of Eimeria and/or genetic markers described here are likely to prove useful for researchers investigating the propensity for resistance development to novel compounds and the development of new attenuated vaccines. Such investigations can be helpful in providing a better understanding of biochemical and molecular aspects of the biology of these parasites.

A survey of drug combinations employed by the poultry industry indicates that they have played an important role in the control of coccidiosis in chickens. The mode of action of their component drugs is described. Advantages that accrue from their use may include a reduction in potential toxicity, a broadening of their spectrum of activity against different species of Eimeria, activity against different stages of the life cycle, and improved efficacy due to synergism between component drugs. Integration of management procedures involving rotation of drug combinations with vaccination is desirable because this has been shown to result in a restoration of drug sensitivity where drug resistance is present and could contribute to the sustainable control of coccidiosis. Threats to the future use of the most widely used combinations, those that include ionophores, stem from the recent desire to eliminate antibiotics from poultry feeds., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The efficacy of the anticoccidial drugs amprolium, clopidol, diclazuril, monensin, monensin + nicarbazin, narasin, narasin + nicarbazin, and salinomycin against field isolates of Eimeria acervulina obtained from a commercial broiler enterprise before and after immunization with a coccidiosis vaccine was investigated. Evaluated by weight gain, feed conversion, and lesion score following challenge, the isolate obtained before vaccination was resistant to all the drugs tested. By contrast, after vaccination the isolate was sensitive to all drugs evaluated by weight gain, and to most drugs judged by feed conversion and lesion score. It is concluded that vaccination had resulted in the restoration of sensitivity to these drugs., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Three-week-old turkey poults were infected with pure lines of three species of Eimeria ( E. adenoeides , E. gallopavonis , and E. meleagrimitis ) recently isolated from commercial turkey farms. The lines had been propagated from a single oocyst and identified by species-specific PCR amplification of the mitochondrial cytochrome c oxidase subunit I (COI) gene. Five to six days after infection their intestines were removed and examined for the presence of intestinal lesions. A description and review of the pathology caused by these parasites is provided, and a scoring system developed by which the severity of the lesions can be evaluated. The system is similar to that described by Johnson, J. and Reid, W. M. [1970. Anticoccidial drugs: lesion scoring techniques in battery and floor-pen experiments with chickens. Experimental Parasitology , 28, 30-36] for chickens in which a score of zero to four is assigned to lesions of increasing severity. The intestinal lesions observed here, and their assigned scores, are supported by representative illustrations. It is hoped that they may prove a useful tool for evaluating the pathology caused by E. adenoeides , E. gallopavonis , and E. meleagrimitis in the turkey. RESEARCH HIGHLIGHTS A scoring system has been developed for intestinal lesions caused by three species of Eimeria that infect the turkey.The lesions attributable to these species are illustrated.

Coccidiosis is a parasitic disease of a wide variety of animals caused by coccidian protozoa. The coccidia are responsible for major economic losses of the livestock industry. For example, the annual cost due to coccidiosis to the global poultry industry has been estimated to exceed US$ 3 billion annually. Currently available drugs for the control of this disease are either polyether ionophorous antibiotics that are derived from fermentation products, or synthetic compounds, produced by chemical synthesis. Unfortunately, no new drugs in either category have been approved for use for decades. Resistance has been documented for all those of the drugs currently employed and therefore the discovery of novel drugs with unique modes of action is imperative if chemotherapy is to remain the principal means to control this disease. This chapter aims to give an overview of the efficacy and mode of action of the current compounds used to control coccidiosis in livestock and provides a brief outlook of research needs for the future.

The phylum Apicomplexa includes parasites of medical, zoonotic and veterinary significance. Understanding the global distribution and genetic diversity of these protozoa is of fundamental importance for efficient, robust and long-lasting methods of control. Eimeria spp. cause intestinal coccidiosis in all major livestock animals and are the most important parasites of domestic chickens in terms of both economic impact and animal welfare. Despite having significant negative impacts on the efficiency of food production, many fundamental questions relating to the global distribution and genetic variation of Eimeria spp. remain largely unanswered. Here, we provide the broadest map yet of Eimeria occurrence for domestic chickens, confirming that all the known species (Eimeria acervulina, Eimeria brunetti, Eimeria maxima, Eimeria mitis, Eimeria necatrix, Eimeria praecox, Eimeria tenella) are present in all six continents where chickens are found (including 21 countries). Analysis of 248 internal transcribed spacer sequences derived from 17 countries provided evidence of possible allopatric diversity for species such as E. tenella (FST values ⩽0.34) but not E. acervulina and E. mitis, and highlighted a trend towards widespread genetic variance. We found that three genetic variants described previously only in Australia and southern Africa (operational taxonomic units x, y and z) have a wide distribution across the southern, but not the northern hemisphere. While the drivers for such a polarised distribution of these operational taxonomic unit genotypes remains unclear, the occurrence of genetically variant Eimeria may pose a risk to food security and animal welfare in Europe and North America should these parasites spread to the northern hemisphere., (Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The course of natural Eimeria infections in 6 successive broiler flocks at a commercial farm comprising 4 houses, where different anticoccidial drug programs were employed, was studied by counting the number of oocysts in the litter at weekly intervals. The course of infection in all flocks followed a bell shaped curve in which oocyst numbers, initially low, increased to a peak ranging from 36 × 10(3) to 74 × 10(3) oocysts/g (OPG) of litter around 3 to 4 wk of age. Numbers subsequently declined to 3 × 10(3) to 15 × 10(3) OPG. Oocysts could be detected between flocks when birds were not present. Species of Eimeria identified included E. acervulina, E. maxima, and E. tenella Despite the presence of large numbers of oocysts in the litter, coccidial lesions were not observed in the intestines of the birds. The performance of broilers at the study site was comparable to that of other farms in the area where birds from the same settlement were reared to a similar age using the same drug programs. The results indicate the ubiquitous nature of Eimeria spp. infections in commercial broilers despite prophylactic medication., (© 2016 Poultry Science Association Inc.)

An attenuated line of Eimeria meleagrimitis was established by repeated propagation of the parasite in 9-day old turkey poults and subsequent selection for precocious development. Following 20 passages, the prepatent period decreased from 120 to 104h. A series of experiments were conducted to evaluate the pathogenicity, immunogenicity and fecundity of the newly selected line. Judged by body weight gain, feed consumption and feed efficiency following infection, the attenuated line had appreciably reduced pathogenicity. Immunogenicity of the attenuated line was examined by infecting poults successively with incremental doses of 10(2), 10(3) and 10(4) oocysts at 0, 7, and 14 days of age respectively. No oocysts were detected following challenge with 5×10(2) oocysts, indicating that the attenuated line had retained immunogenicity. Fecundity was assessed by infecting two-week old birds with 5×10(2) oocysts of either parent or attenuated line. Oocyst production from 96 to 240h post-infection showed that the patent period of the attenuated line commenced earlier and was of shorter duration than the parent line., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Oocysts of Eimeria spp. were isolated from litter samples obtained from 30 commercial turkey farms. Genomic DNA was extracted from clean oocysts, and polymerase chain amplification of the species-specific cytochrome c oxidase subunit I (COI) gene was performed for five species of turkey Eimeria. The species tested were Eimeria adenoeides, Eimeria meleagrimitis, Eimeria meleagridis, Eimeria dispersa, and Eimeria gallopavonis. All DNA samples were positive for E. meleagrimitis, nine were positive for E. adenoeides, two were positive for E. dispersa, and none for E. meleagridis and E. gallopavonis. E. meleagrimitis occurred as a single species in 21 (70 %) of the farms while 9 (30 %) farms had a mixed species with E. meleagrimitis and E. adenoeides and 2 (7 %) were triple positive with E. meleagrimitis, E. adenoeides, and E. dispersa. This is the first account of the field prevalence of turkey Eimeria species using molecular methods.

Five successive flocks of broilers were reared in floor-pens and given different drug programs or were vaccinated against coccidiosis. Oocysts of Eimeria were isolated from the litter of pens during the fifth flock and their sensitivity to salinomycin (Sal) investigated by measuring new oocyst production following infection of medicated and unmedicated birds. Parasites obtained following 5 flocks given Sal were not well-controlled and it was concluded that they were partially resistant to the drug. Parasites obtained following 4 unmedicated flocks and one medicated flock were better controlled by Sal and it was concluded that in the absence of continuous medication there had been an improvement in drug efficacy. Sal almost completely suppressed oocyst production of isolates from treatments in which medication was followed by vaccination, indicating that when a drug program is followed by vaccination, restoration of sensitivity to Sal had occurred., (© 2015 Poultry Science Association Inc.)

Anticoccidial effects of Sericea Lespedeza (SL) included in the diet at different levels were evaluated in chickens following oral infection with sporulated oocysts of either Eimeria acervulina, Eimeria maxima or Eimeria tenella. A series of experiments were conducted to determine the effect of SL upon the ability of the parasites to multiply in the intestine, and the effect on bodyweight gain, feed intake, and feed conversion following infection. Chicks infected with a low dose of oocysts (500 oocysts/bird) of E. acervulina or E. maxima did not show differences in the numbers of oocysts produced in the feces whether they were given 0, 1, 2, or 4% SL in the diet. There was no significant difference in the weight gain, feed intake, or FCR of birds infected with high doses of E. acervulina or E. maxima (200,000 or 100,000 oocysts/bird respectively) whether 0, 1, 2, or 4% SL was included in the feed. No significant difference in the numbers of oocysts in the feces, weight gain, feed intake, and FCR of birds infected with E. tenella (low dose of 500 oocysts; high dose of 50,000 oocysts per bird) whether 4% SL was included in the feed. The results of this study indicate that SL has no anticoccidial activity against Eimeria species in the chicken., (Copyright © 2014 Elsevier B.V. All rights reserved.)

This article describes some of the milestones in research concerned with protozoan parasites of the genus Eimeria that infect birds and cause the disease coccidiosis. The time period covered is from 1891, when oocysts were first found in the ceca of diseased chickens, to the present. Progress in our understanding has lagged behind that of other protozoan parasites such as Toxoplasma and Plasmodium despite the enormous importance of Eimeria to animal livestock production. Nevertheless, applied research by universities, government agencies, and private industry has resulted in the successful development of methods of control, research that continues today. The topics covered and the references provided are selective and include life cycles and biology, pathology, ultrastructure, biochemistry, immunity, genetics, host cell invasion, species identification, taxonomy, chemotherapy, vaccination, and literature concerned with avian coccidiosis. This review is primarily concerned with the avian species of Eimeria that infect poultry, but some important advances, principally in immunology, have been made using species that infect rodents and rabbits. These are included where appropriate.

1. An experiment was designed to evaluate the effect of different doses of oocysts of Eimeria acervulina on intestinal absorption and skin deposition of xanthophylls (XAs) in broilers. 2. A total of 192 broiler chickens were randomly assigned to 4 groups: an uninfected control group and three groups inoculated with either 1 × 10(2), 1 × 10(4) or 1 × 10(5) sporulated oocysts of E. acervulina by gavaging at 21 d. There were 4 replicate pens (2 male and 2 female) per group. 3. Plasma xanthophyll (PX) and skin yellowness (SY) were measured in live birds weekly. At 42 d of age, SY was measured in the breast and abdomen after chilling and in the breast 24 h post-processing on refrigerated carcasses. 4. In general, in all challenged treatments, and for the duration of the study, the average PX decreased by 0.02 μg/ml (R(2) = 61.6%) for every 1000 inoculated oocysts, whereas PX increased by 1.26 μg/ml/d in uninfected birds. 5. The average SY in live birds from 21 to 42 d of age decreased by 0.019 b*/every 1000 oocysts administered, while SY of uninfected controls increased by 0.57 b*/d. It was also noted that in all treatments females had a greater SY (6.17 b*) than males for the duration of the study. The SY of the breast and abdomen was correlated (r = 0.76) in chilled carcasses. Breast SY in 24 h refrigerated carcasses was greater in the control group and for female birds. 6. Oocyst excretion was different between inoculated treatments only on 7 d post-inoculation (PI). Coccidia lesion scores in the duodenum averaged 1+ in infected birds and 2+ in birds given the highest oocyst dose.

Newly hatched turkey poults were infected with 10(2) oocysts of Eimeria adenoeides and subsequently reinfected with 10(3) and 10(4) oocysts at 6 and 12 d of age, respectively. Three peaks in oocyst production were observed in the feces of poults following this series of infections. A second group of poults given the same dosing regimen was challenged with 5 × 10(4) oocysts/poult at different times to evaluate the acquisition of immunity. Judging by weight gain and mortality, no protection had been acquired at 6 d of age, but partial protection was observed by 12 and 18 d of age. A third group of poults were also infected with 10(2) oocysts and subsequently reinfected with 10(3) and 10(4) oocysts at 6 and 12 d of age to evaluate cellular immune responses to infection. Sections of ceca from infected poults showed a significantly higher leukocyte infiltration on d 6, 10, 12, 16, and 18 after infection than uninfected controls. The percent area occupied by CD4+ and CD8+ lymphocytes in the ceca, as assessed by immunohistochemistry, was significantly elevated in infected poults on d 12, 16, and 18. The relative expression of chemokine CXCLi2, and cytokines IL1β, IFNγ, IL10, IL13, IL2, IL12b, and IL18 was measured by real-time reverse-transcription PCR. The expression of CXCLi2 and IL10 was found to be elevated on d 12, and IFNγ on d 10, 12, and 16. Expression of IL13 and IL18 was increased on d 10 and IL2 on d 10 and 16, and that of IL12b on d 16 in infected poults. Increase in the infiltration of leukocytes, percent area occupied by CD4+ and CD8+ lymphocytes, and changes in the relative expression of cytokines in the ceca characterize the dynamics of immune responses in turkey poults infected with E. adenoeides early in life.

Coccidiosis is a widespread and economically significant disease of livestock caused by protozoan parasites of the genus Eimeria. This disease is worldwide in occurrence and costs the animal agricultural industry many millions of dollars to control. In recent years, the modern tools of molecular biology, biochemistry, cell biology and immunology have been used to expand greatly our knowledge of these parasites and the disease they cause. Such studies are essential if we are to develop new means for the control of coccidiosis. In this chapter, selective aspects of the biology of these organisms, with emphasis on recent research in poultry, are reviewed. Topics considered include taxonomy, systematics, genetics, genomics, transcriptomics, proteomics, transfection, oocyst biogenesis, host cell invasion, immunobiology, diagnostics and control., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

In July 1971, the polyether ionophorous antibiotic monensin was introduced in the United States for the control of coccidiosis in poultry. At that time, prospects for new anticoccidial agents were not good. Amprolium had enjoyed several years of use, but many other compounds had been abandoned as resistance to them developed. After the introduction of monensin, most commercial broilers were medicated with the drug and it is still widely used for this purpose today. Apart from in poultry, monensin is also used to control coccidiosis in game birds, sheep, and cattle. Indeed, more animals have been medicated with ionophores, such as monensin, for control of disease than any other medicinal agents in the history of veterinary medicine. In this review, we discuss the discovery, mode of action, and efficacy of monensin, together with matters of importance to the poultry industry such as commercial use, drug resistance, toxicity, pharmacology and residues, host immunity to coccidiosis, and effects in other avian species.

A primary infection of 12.5 x 10(3) oocysts of Eimeria adenoeides , given to 20-d-old turkey poults, resulted in depression of weight gain, and the production of large numbers of oocysts in the feces, compared with uninfected controls. Poults were raised under conditions to prevent possible reinfection to determine the ability of the primary infection to confer protective immunity against a challenge infection of 5 x 10(4) oocysts given at 34 d of age. Using weight gain and oocyst production after challenge as criteria for protection, the results indicated that immunity had developed. The concentration and proportions among white blood cell (WBC) populations in peripheral blood were determined at different times after the primary infection. The WBC concentration of infected poults was elevated on d 7 and 11, primarily due to elevated levels of lymphocytes and monocytes on d 7 and eosinophils on d 11. There were no differences in heterophil and basophil concentrations between infected and uninfected poults at any of the time points examined. With the exception of increased percentages of eosinophils on d 11, infection was not associated with alterations in the proportions among WBC populations. Comparison of CD4+ and CD8+ defined lymphocyte subpopulations in the blood of infected versus uninfected poults revealed higher concentrations of CD4+ lymphocytes on d 11, lower concentrations of CD8+ cells on d 4, and higher concentrations of CD8+ cells on d 11 of infection, as well as elevated ratios of CD4+:CD8+ lymphocytes in infected birds on d 4 and 11. These alterations in WBC profiles after primary E. adenoeides infection in turkey poults suggest initiation of both innate and adaptive cellular immune activities designed to effectively cope with a parasitic, intracellular pathogen.

Infectious and metabolic disorders are common in poultry and cause stress, poor performance, and mortality that results in considerable economic loss. Identifying the nature of stress in chickens will assist the development of appropriate measures to improve health and welfare. Acute phase proteins are hepatic proteins, the blood concentrations of which change significantly in the event of many health problems including inflammation and physical injuries. Thus, acute phase proteins are used as nonspecific diagnostic markers for various health disorders. Our previous studies showed that serum ovotransferrin (OVT) is an acute phase protein in chickens. Therefore, in the present study, we investigated whether OVT concentration can be a marker of physiological stress using sera from chickens with different infectious and metabolic disorders. A competitive enzyme immunoassay was developed to measure serum OVT concentrations. The results show that with experimentally induced pulmonary hypertension syndrome and tibial dyschondroplasia, there were no significant changes in OVT levels compared with matched controls. In contrast, when chickens were infected with microbes such as the bacterium Escherichia coli, or protozoan parasites such as Eimeria maxima and Eimeria tenella, there was a significant increase in the levels of OVT in the serum. Chickens with spontaneous autoimmune vitiligo also showed a significant increase in blood OVT levels. These studies suggest that blood OVT concentration is modulated under inflammatory and microbial stress and can therefore be used as a diagnostic marker of infection and inflammation in chickens.

The sensitivity of field isolates of turkey coccidia from the United States to the anticoccidial drugs amprolium, clopidol, diclazuril, and monensin was investigated. Clopidol and diclazuril were the most effective, followed by monensin and amprolium. Thirty-one isolates were classified as resistant to amprolium, 23 resistant to monensin, 10 resistant to diclazuril, and 6 resistant to clopidol. Six isolates were partially resistant to monensin, 10 partially resistant to clopidol, and 11 partially resistant to diclazuril. Four isolates were sensitive to monensin, 12 sensitive to diclazuril, and 17 sensitive to clopidol.

"Continuous feeding of low concentrations of sulfaquinoxaline for the control of coccidiosis in poultry" by L. C. Grumbles, J. P. Delaplane, and T. C. Higgins [Poult. Sci. (1948) 27:605-608] was the first paper to demonstrate that it was possible to control coccidiosis by the continuous inclusion of a low level of a drug in the feed of chickens. The principle involved (prevention or prophylaxis) has had a profound impact on our ability to grow chickens and turkeys under intensive conditions. Indeed, it is possible that the modern poultry industry could never have developed to its present extent without the advent of drugs used prophylactically to control coccidiosis. One particular insight was that use of a compound in this manner did not necessarily prevent the acquisition of immunity, an important principle that helps explain the continued efficacy of ionophorous antibiotics used today. The significance of this work to the poultry industry and individuals involved in research, whether employed by government, academia, or pharmaceutical companies, cannot be overstated. Economic benefits, in terms of improved productivity, have been demonstrated in numerous studies published in Poultry Science. In addition, the livelihoods of many poultry farmers have been helped by the control of a disease that in the past caused substantial morbidity and mortality in their flocks. The paper is brief and contains no critical science involving novel procedures but has had a profound influence on the health of poultry for the last 6 decades. For this reason, it is nominated as a landmark contribution from the first 100 yr of Poultry Science.

Coccidiosis is a widespread disease of considerable economic importance to the turkey industry, as is evident by the fact that many millions of dollars are spent annually on its control. Despite this, in recent years little research has been conducted on the causative organisms--protozoan parasites of the genus Eimeria. This is unfortunate because current methods of control are not without problems. It is possible, therefore, that in the future optimal productivity in turkey production will be difficult to achieve. There has been no recent comprehensive literature review of coccidiosis in the turkey. In the present article our knowledge of the biology of the species of Eimeria that infect the turkey is critically reviewed. Other topics considered include host specificity, host resistance, acquired immunity, in vitro cultivation, pathology, pathogenicity, clinical signs, diagnosis, epidemiology, prevalence, significance, chemotherapy, drug resistance, drug toxicity, and vaccination. The review is intended to provide a baseline for future studies of this important disease.

The acquisition of immunity by chickens infected 18 h post-hatch with 100 oocysts of Eimeria maxima and reared in floor-pens in contact with their droppings was investigated. In the first experiment, birds were placed on new litter and immunity was measured at 2, 3, 4, and 5 weeks by calculation of weight gain from days 0 to 7 following challenge with 100000 oocysts or by oocyst production in the faeces from days 5 to 8 following challenge with 500 oocysts. In the second experiment, birds were placed on new litter or reused litter from the first experiment (1 week after birds from the first experiment had been removed when 6 weeks of age), and were challenged at 1, 2, and 3 weeks of age. In the first experiment, immunity had developed in birds challenged at 3, 4, and 5 weeks, judged by weight gain and oocyst production, but immunity was not complete at 2 weeks. In the second experiment, immunity had developed in birds challenged at 1, 2, and 3 weeks measured by either criterion. In both experiments, birds produced small numbers of oocysts in their faeces following challenge. Judged by the weight gain following challenge, no significant difference in the acquisition of immunity was observed whether birds were reared on new or reused litter.

A precocious line of Eimeria meleagridis was developed by repeated propagation of the parasite in young turkeys and collection of the very first oocysts produced following infection. After 20 generations of selection, the prepatent period was reduced by 4-8 hr. Weight gain of poults given this line, from days 0-3 after infection, was significantly greater than that of poults given the parental strain, indicating that selection resulted in a loss of pathogenicity. Poults immunized with the precocious line produced no oocysts following challenge with the parental strain, indicating that immunogenicity was retained following selection. This is the first published report of the selection of a precocious line of Eimeria in the turkey.

An isolate of Eimeria meleagridis Tyzzer, 1927 was obtained by harvesting oocysts from the ceca of a turkey from northwest Arkansas and a pure line was established by infecting birds with a single oocyst. Oocysts were first produced in the ceca of infected birds from 102 to 108 hr after inoculation and were of similar size (mean length X width, 24.9 X 17.0 microm) to those of Eimeria adenoeides Moore and Brown, 1951 and Eimeria gallopavonis Hawkins, 1952. The line was identified as E. meleagridis based upon the development of large schizonts in the midintestine, and small schizonts in the ceca. Two generations of large schizonts were found 48 and 72 hr after infection, and at least two generations of small schizonts were found from 60 to 108 hr after infection. An inoculum of 2 X 10(5) oocysts was found to cause a significant reduction in weight gain from days 0-3 and 0-6 after infection, suggesting that the significance of this species of Eimeria as a pathogen of turkeys should be reassessed.

These guidelines are intended to aid those engaged in poultry research in the design, implementation and interpretation of laboratory, floor-pen and field studies for the assessment of the efficacy and safety of live anticoccidial vaccines for immunization of chickens and turkeys against Eimeria species. In addition to efficacy and safety requirements, manufacture, quality control and licensing considerations are discussed. The guidelines do not address subunit vaccines comprising non-viable material, but many of the principles described will be relevant to such vaccines if they are developed in the future. Guidelines are available in some countries for avian vaccines of bacterial or viral origin but specific standards for anticoccidial vaccines in poultry have not, as far as we know, been produced. Information is provided on general requirements of registration authorities (based upon regulations applicable in the European Union and the USA) for obtaining marketing authorizations for vaccines. These guidelines may assist poultry specialists in providing specific information for administrators involved in the decision-making process leading to registration of new vaccines, and are intended to facilitate the worldwide adoption of consistent, standard procedures.

The effects of diclazuril and monensin, when included in the feed of turkeys from 0 to 10 wk, upon performance and development of immunity to Eimeria species was investigated. Birds were initially inoculated with a low dose of oocysts of three species of Eimeria at 3, 5, 7, and 9 days of age in order to simulate a natural infection. Weight gain and feed intake from 0 to 6 wk of age was significantly greater in medicated birds compared with those that received no anticoccidial medication. Weight gain and feed intake from 6 to 10 wk was greater in birds that received diclazuril than in unmedicated birds. No differences in performance were evident after drug withdrawal from 10 to 16 wk. Immunity to Eimeria species developed by 10 wk in birds that received no anticoccidial medication but did not develop in those given diclazuril or monensin.

These guidelines have been written to aid in the design, implementation and interpretation of studies for the assessment of drug efficacy against Eimeria species in chickens and turkeys. The information provided deals with many aspects of how to conduct controlled studies in battery cages (dose determination), floor pens (dose confirmation), and commercial facilities (field effectiveness studies), the selection of birds, housing, feeding, preparation of medicated rations, record keeping, diagnostic techniques, and methods for the preparation, maintenance and use of parasites. These guidelines are also intended to assist investigators in conducting specific studies, provide specific information for registration authorities involved in the decision-making process, assist in the approval and registration of new anticoccidial drugs, and facilitate the world-wide adoption of standard procedures.

The timing and magnitude of a coccidian infection, judged by the numbers of oocysts of Eimeria present in the litter, were affected by the duration of medication. In birds medicated for 6 wk and infected at 35 d of age, fewer oocysts were produced than in birds medicated for 4 or 5 wk whether infected at 18 or 35 d of age. Feed conversion at 6, 7, and 8 wk of birds infected at 18 d and medicated for 6 wk was less than that of birds medicated for 4 or 5 wk. Birds infected at 35 d and medicated for 6 wk had a lower feed conversion than birds medicated for 5 wk. Immunity to Eimeria tenella had developed by 8 wk in birds medicated for 4, 5, or 6 wk if infected at 18 d of age. Immunity did not develop in those birds medicated for 6 wk when infected at 35 d.

These guidelines have been prepared to assist in the planning, operation and interpretation of studies designed to assess the effectiveness of drugs against helminth parasites of chickens and turkeys. They are the first to be compiled under the auspices of the World Association for the Advancement of Veterinary Parasitology (WAAVP) for these parasites. The advantages and disadvantages of the widely used critical and controlled tests are discussed. Information is provided on the selection of animals for experiments, animal housing, feed, dose determination studies, confirmatory and field trials, record keeping and necropsy procedures. This document should help investigators and those involved in product approval and registration in conducting and evaluating studies concerned with determining the effectiveness and safety of anthelmintic drugs.

In 1910, H. B. Fantham described the life cycle of a coccidian parasite in birds. Fantham was a parasitologist at Cambridge University in the United Kingdom working for an enquiry into diseases affecting the red grouse. Despite the growing importance of the poultry industry and the realization that coccidiosis was an important disease of the fowl, little further work was carried out in the United Kingdom until coccidiosis research was initiated at the Veterinary Laboratory, Weybridge almost 30 yr later. Further progress depended upon research carried out at academic and agricultural institutions in the United States. E. E. Tyzzer at Harvard University provided the solid foundation upon which our present knowledge of coccidiosis, and the species of Eimeria involved in the disease, is based. Agricultural experiment stations (AESs) throughout the nation played an important role in communicating advances to the agricultural community. W. T. Johnson at Western Washington and, subsequently, Oregon AES made significant contributions to our understanding of the disease, as did C. A. Herrick and coworkers at Wisconsin AES, J. P. Delaplane and coworkers at Rhode Island AES, and P. P. Levine at Cornell University.

Under simulated natural conditions of bird production and parasite challenge, the effects of ascaridiasis and the effectiveness of fenbendazole treatment (6-day regimes in the feed at 16 ppm) were documented. Birds were artificially challenged with ascarid larvae on a daily basis from day 35 to 112, with bird grow out ending on day 119. Experimental groups, on a per pen basis, were infected control, treated with fenbendazole at days 63-69, treated with fenbendazole at days 63-69 and days 91-97, and uninfected control. In the same order as above, and on an experimental group mean bird basis, final weights were 13.34, 13.47, 13.59, and 13.78 kg, average daily gains from day 7 to day 119 were 117.8, 118.9, 120.1, and 121.8 g, and units gained per unit of feed consumed from day 7 to day 119 were 0.337, 0.341, 0.347, and 0.362. Infected control bird mean Ascaridia dissimilis burdens, with all stages combined, ranged from 351.1 on day 63 to 117.2 on day 91, levels seen commonly with naturally infected commercial turkeys. Trial data dearly indicated that moderate A. dissimilis burdens negatively impacted animal performance (average daily gains and feed efficiencies) and that these parasite burdens are effectively removed by fenbendazole treatment.

The development of new methods of administering coccidiosis vaccines has facilitated their use in the hatchery and thereby improved prospects for the economic vaccination of broilers. The acquisition of protective immunity to Eimeria species is boosted by further exposure to infection after vaccination. Factors that affect the reproductive efficiency of non-attenuated and attenuated vaccines are considered and the key role that oocyst production plays in establishing and maintaining uniform immunity in a flock of chickens is discussed. In addition to immunisation, a possible advantage to the application of certain vaccines is that their use could repopulate poultry houses with drug-sensitive organisms. Theoretical rotation programmes in which the use of drugs is alternated with that of vaccines are described. Variability of the cross-protective immune response between strains of the same species should be considered during vaccine development and subsequent use. The significance of less common species of Eimeria, not included in all vaccines, also needs to be assessed. An important consideration is the occurrence of pathogens other than Eimeria (such as the bacterium Clostridium) in flocks given coccidiosis vaccines and the methods by which they might be controlled. More research is required into the relationship between bacterial and viral infections of poultry and coccidiosis vaccination. Vaccines need to be developed that are simple to apply and cost effective for use in areas of the world where small-scale poultry production is commonplace. In the near future it is likely that more live vaccines based upon oocysts derived from attenuated strains of Eimeria will be developed but in the longer term vaccines will be based on the selective presentation to the host of specific molecules that can induce protective immunity. This achievement will require significant investment from the private and public sectors, and, if successful, will facilitate the sustainable control of coccidiosis in poultry production.

Data collected by Agri Stats Inc. in the US for the years 1995 to 1999 was evaluated to determine the types of anticoccidial drug programs used by broiler plants, their frequency and duration, and whether there was any correlation with performance of the birds. Information was available for five feed types (prestarter, starter, grower, first withdrawal, and final withdrawal). The most popular program was an ionophore (ION; principally salinomycin) in starter and grower feeds. A chemical (principally nicarbazin) was also used in the starter feed followed by an ION in the grower feed, or two different ION in the starter and grower feeds. Seasonal differences were apparent in the type of program and in the frequency of program changes. There was no consistent pattern in the sequence with which different programs were employed. There were no significant differences in calorie conversion or the number of days to produce a 2.27 kg bird, whether a single ION or a chemical followed by an ION was used, but mortality was significantly higher for the latter. For 1999, calorie conversion and mortality were higher in plants where chemical-ION programs had been used for more than 40% of the time during the previous 4 yr. The duration and frequency with which different programs were employed were similar whether birds were raised to final weights of 1.5 to 2.0 kg, 2.0 to 2.5 kg, or more than 2.5 kg. There were significant differences in the use of salinomycin and nicarbazin in different regions of the US.

The efficacy of different concentrations of monensin in turkeys exposed to field isolates of Eimeria and the effect of the withdrawal of monensin from their diet were investigated. Results of a battery study indicated that 66 ppm monensin was effective against three species of Eimeria in the turkey. In floor pens in which poults were exposed to infection, use of 59.5, 79.4, and 99.2 ppm monensin resulted in reduced mortality and improved feed conversion at 3 wk of age compared with birds that had received no medication. Poults given 59.5 ppm monensin weighed more and had a better feed conversion at 3 wk than poults given 79.4 or 99.2 ppm monensin, but at 10 wk no differences in the body weight or feed conversion of poults given different concentrations of drug were apparent. At 14 wk (and from 10 to 14 wk), poults that had been given 99.2 ppm monensin had a lower feed intake and weighed less than birds that had been given 59.5 ppm of the drug but there were no significant differences in feed conversion. There was no significant difference in the weight gain or feed conversion of poults from 10 to 14 wk of age whether monensin was present in the feed or had been withdrawn. No evidence of compensatory growth was found in the present study.

Anticoccidial drugs undergo a vigorous series of test during development in order to demonstrate their efficacy and safety. Evaluation of efficacy may also be necessary after a drug is utilised in the field in order to establish whether resistance has developed. The principles and procedures used to evaluate anticoccidial efficacy were reviewed in a series of papers presented at a symposium held at the University of Georgia in May 1969. It is a testimony to the quality of this symposium that much of the information presented remains valid today. In the USA, general procedures that should be followed in new drug investigations are given in various guidelines and memoranda issued by the Food and Drug Administration. Specific guidelines relate to anticoccidial investigations. Although not a legal requirement, they provide a standard for those investigators seeking approval of a new anticoccidial drug. These guidelines are periodically revised, and investigators should consult the FDA for advice on specific protocols. Many of these procedures are also appropriate for the determination of drug resistance.

Day-old turkeys poults were given a live coccidiosis vaccine via the eye at the hatchery, and all poults produced oocysts in their droppings 7 d later. No oocytes were present in droppings of unvaccinated controls. The weight gain of vaccinated poults challenged at 3 wk of age with a mixture of three species of Eimeria was lower than that of unchallenged poults, indicating that they were susceptible to infection. No significant difference in weight gain was found when poults were challenged at 6 wk, however, indicating that they had acquired immunity to the species used for challenge.

No significant differences in the BW, feed intake, feed conversion, or mortality of broilers that had been vaccinated at 3 d of age with a live coccidiosis vaccine were observed irrespective of whether they had been given roxarsone, bacitracin methylene disalicylate, or both drugs in the feed. The weight gain of birds challenged with a high dose of the vaccine at 51 d was not significantly different from that of unchallenged birds and no lesions of Eimeria species were found in their intestines irrespective of the medication given. It is concluded that roxarsone and bacitracin methylene disalicylate do not interfere with the acquisition of immunity to Eimeria species in broilers.

The performance of broilers reared in floor pens and given monensin in the feed at 121 ppm was compared with that of birds given no drug. Feed intake and BW gain of medicated birds was significantly lower than that of unmedicated birds from 0 to 22 d of age. Feed intake and feed conversion of medicated birds was significantly reduced, compared with unmedicated birds, from 22 to 53 and 0 to 60 d of age. Total mortality, and mortality due to leg abnormalities from 22 to 53 and 0 to 60 d, was significantly lower in birds given monensin. There was no difference in the incidence of tibial dyschondroplasia (TD) by 60 d. No differences in mortality due to pulmonary hypertension syndrome (PHS) were observed for any age period. Birds removed from pens at 28 d that had received monensin had lower hematocrit and percentage saturation of hemoglobin with oxygen in the blood than unmedicated birds. No differences in these variables were found at 54 d. There were no differences in the right ventricle weight: total ventricular weight ratios or electrocardiogram lead II values at 28 or 54 d. The results indicate that PHS does not occur more frequently in broilers medicated with monensin.

The sensitivity of field isolates of Eimeria to monensin was investigated following the use of a commercial coccidiosis vaccine. Monensin was not effective against isolates from five commercial farms where ionophores had been used for many flocks. Monensin was more effective, however, against isolates obtained from the same farms following the use of a coccidiosis vaccine. No further improvement in drug efficacy was noted after five successive flocks in which the vaccine was used.

A floor-pen trial was conducted to investigate the effects of different shuttle programs upon the growth of broilers to 8 wk of age. Nicarbazin, halofuginone, and robenidine, when included in the starter feed for 3 wk, were effective in preventing lesions due to Eimeria. The effects of medication upon performance were apparent, medicated groups gaining more weight by 6 wk and having a lower feed conversion at 6, 7, and 8 wk than the unmedicated controls. There were no significant differences in body weight at 6, 7, or 8 wk or feed conversion at 6 or 7 wk among the medicated groups, whether medication was withdrawn for 7 or 14 days. A decrease in the number of small and medium oocysts in the litter was observed as the trial progressed. Few large oocysts (Eimeria maxima) were seen in the medicated groups. Numbers of oocysts did not increase following withdrawal of medication. Birds from all medicated groups were challenged at 6 wk with oocysts of Eimeria acervulina, Eimeria maxima, or Eimeria tenella. Weight gains were similar to that of the unchallenged controls, indicating that they had acquired immunity to these species of Eimeria.

Forty-six broiler houses were examined for the presence of oocysts of Eimeria in the litter before and after withdrawal of lasalocid or salinomycin from the feed. A decrease in number of small oocysts (Eimeria acervulina or Eimeria mitis) was observed following withdrawal of medication. Numbers of medium-sized oocysts (probably Eimeria tenella) remained the same, but an increase in large oocysts (Eimeria maxima) was recorded. No lesions attributable to Eimeria tenella were found in the ceca of birds after withdrawal of the drug. More small- and medium-sized oocysts were found at sites with new litter than at sites where the litter had been employed for previous flocks. No difference in the number of oocysts was found, whether birds were reared on oak shavings or a mixture of pine shavings and rice hulls. The number of oocysts was positively correlated with the moisture content of the litter, but there was no correlation between oocysts present and the final BW or feed conversion of the birds. Moisture levels were highest (after withdrawal of drug) for new litter or oak shavings. There was no correlation between moisture content of the litter and BW or feed conversion.

The incidence of oocysts was monitored in the litter at two broiler farms where birds were reared to 8 wk of age using a shuttle program involving nicarbazin (from 0 to 21 days of age) and salinomycin (from 21 to 44 days of age). Some birds from these farms were challenged with three species of Eimeria to see whether immunity had developed as a result of prior exposure to infection. Oocysts of at least three species of Eimeria were present in small numbers in the litter. Birds from both farms were immune to Eimeria acervulina and Eimeria maxima (judged by weight gain) although the immunity was incomplete as evidenced by oocyst production. Birds from one farm were immune to Eimeria tenella (judged by weight gain) but birds from the other farm were susceptible to this species.

The development of resistance by the Houghton strain of Eimeria tenella (H) to monensin has been studied by serially passaging the strain in groups of 30 birds, each inoculated with 10(6) oocysts and given 100 p.p.m. of drug in the food. After 16 passages a line partially resistant to this concentration of monensin was produced. Passage of the line in birds medicated with 200 or 300 p.p.m. of monensin did not result in the acquisition of a greater degree of resistance. A further reduction in sensitivity to 100 p.p.m. monensin was obtained, however, when the line was serially passaged in 15 groups of 6 birds each given 3 X 10(6) oocysts. Five passages in the absence of monensin resulted in restoration of sensitivity, suggesting that the resistance developed was not stable. The pathogenic effects of the line that had been passaged in chickens given 300 p.p.m. of monensin were less than those of the parent line.

Twenty-four field isolates of Eimeria spp. which included Eimeria acervulina, E. praecox and E. mivati were tested for sensitivity to arprinocid included in the diet at 60 ppm. Twelve isolates from sites where arprinocid had been use for 5-7 successive flocks were resistant to the drug, whereas 12 isolates from sites where arprinocid had never been used were sensitive. These results indicate that resistance had been acquired as a result of the use of arprinocid in the field.

Resistance to arprinocid was developed in Eimeria tenella after seven serial passages in chickens given progressively greater concentrations of drug. Resistance to the quinolone decoquinate developed after eight passages. These results taken together with the observation that resistance to both drugs developed within a year of their introduction suggest that some indication of the likelihood of the emergence, in the field, of resistance to a given drug, may be obtained by comparing it with a standard reference drug in experimental studies.

Drug resistance is now recognised as a major cause of the failure of drugs to control coccidiosis in the fowl. In this article, biological, biochemical and genetic aspects of resistance in Eimeria are reviewed and some of the problems that may limit progress in understanding the nature of resistance in coccidia are discussed.

The development of resistance by the Houghton strain (H) of E. tenella to robenidine has been studied in the laboratory, by serially passaging the strain in chickens fed increasing concentrations of drug. Resistance to robenidine developed more readily in experiments using larger numbers of birds with higher numbers of oocysts in the inoculum. Both these factors increased the parasite population and increased the chance of selecting parasites resistant to the drug. E. tenella (H) was made resistant to 264 ppm robenidine and showed no cross-resistance to other anticoccidial agents. Resistance arose in a series of 'steps' as the concentration of drug was increased. E. tenella (H) was continuously passaged at concentrations ranging from 2 to 33 ppm of robenidine. After 16 passages, lines passaged at 2,4 and 8 ppm were not resistant to 33 ppm robenidine, suggesting that the degree of resistance developed was dependent upon the drug selection pressure.

The effects of Eimeria maxima or restricted pair-feeding on weight gain, plasma concentrations of protein, glucose, free fatty acids (FFA) and uric acid and liver glycogen were compared in immature fowl. Food intake/kg body weight and weight gain decreased during the acute phase of infection (days 5-7) while weight loss was prolonged for an extra day compared with pair-fed birds. During recovery, food intake/kg body weight of infected birds was greater than that of non-infected controls but there was no evidence for an increase in growth rate compared with controls when body weight was considered. Growth rate of pair-fed birds was greater than that of infected birds during recovery, indicating their better use of ingested food. Liver glycogen and plasma protein concentration were decreased during the acute phase of infection but the concentrations of plasma glucose, free fatty acid (FFA) and uric acid were not affected. In pair-fed birds liver glycogen was depleted, concentrations of plasma glucose and uric acid decreased and FFA increased, and these changes persisted for the remainder of the experiment. The findings are similar to those in birds whose food has been withheld and were probably due to the pattern of food intake imposed by the experimental protocol. It is concluded that the metabolic differences between infected and pair-fed birds are of doubtful significance.