Monitoring the prevalence and abundance of parasites over time is important for addressing their potential impact on host life histories, immunological profiles and their influence as a selective force. Only long-term ecological studies have the potential to shed light on both the temporal trends in infection prevalence and abundance and the drivers of such trends, because of their ability to dissect drivers that may be confounded over shorter time scales. Despite this, only a relatively small number of such studies exist. Here, we analysed changes in the prevalence and abundance of gastrointestinal parasites in the wild Soay sheep population of St. Kilda across 31 years. The host population density (PD) has increased across the study, and PD is known to increase parasite transmission, but we found that PD and year explained temporal variation in parasite prevalence and abundance independently. Prevalence of both strongyle nematodes and coccidian microparasites increased during the study, and this effect varied between lambs, yearlings and adults. Meanwhile, abundance of strongyles was more strongly linked to host PD than to temporal (yearly) dynamics, while abundance of coccidia showed a strong temporal trend without any influence of PD. Strikingly, coccidian abundance increased 3-fold across the course of the study in lambs, while increases in yearlings and adults were negligible. Our decades-long, intensive, individual-based study will enable the role of environmental change and selection pressures in driving these dynamics to be determined, potentially providing unparalleled insight into the drivers of temporal variation in parasite dynamics in the wild.

Within-year variation in infection is a ubiquitous feature of natural populations, but is determined by a complex interplay of environmental, parasitological and host factors. At the same time, co-infection is the norm in the wild. Longitudinal dynamics of co-infecting parasites may therefore be further complicated by covariation across multiple parasites. Here, we used fecal parasite egg and oocyst counts collected repeatedly from individually marked wild Soay sheep to investigate seasonal dynamics of six gastrointestinal parasite groups. Prevalence and abundance tended to be higher in spring and summer, and abundance was higher in lambs compared to adults. We found that within-year variation in highly prevalent strongyle nematode counts was dependent on adult reproductive status, where reproductive ewes had distinct dynamics compared to males and barren ewes. For similarly prevalent coccidia we found an overall peak in oocyst counts in spring but no differences among males, barren and pregnant ewes. Using multivariate mixed-effects models, we further show that apparent positive correlation between strongyle and coccidia counts was driven by short-term within-individual changes in both counts rather than long-term among-individual covariation. Overall, these results demonstrate that seasonality varies across demographic and parasite groups and highlight the value of investigating co-infection dynamics over time.

Many parasites spend part of their life cycle as infectious forms released from an infected host in the external environment, where they may encounter and infect new hosts. The emergence of infectious life stages often occurs once a day to minimize mortality in adverse environments. In bird hosts, intestinal parasites such as coccidia are generally released with feces in the late afternoon. This dynamic is adaptive since it allows avoiding desiccation and ultraviolet (UV) radiation, thus reducing mortality of oocysts in the environment until transmission to the next host. If this circadian rhythm is the result of natural selection to increase oocyst survival, we may hypothesize that oocysts will appear in feces at different times depending on the environment where hosts live. Particularly, in an environment where UV radiation and desiccation are very low, we may expect oocyst circadian release to disappear since the main selective pressure would be relaxed. We sampled different species of birds in tropical and temperate forests in spring and investigated coccidian oocyst output. A strong circadian variation in the prevalence of hosts shedding coccidian oocyst was detected for species caught in the temperate forest with an increase in prevalence in the late afternoon, whereas prevalence of birds shedding oocysts was constant over the course of the day for most species sampled in the tropical rain forest. These results are consistent with the hypothesis that oocysts’ circadian output is maintained by natural selection to increase oocyst survival. We discuss the adaptive significance of diurnal periodicity in parasite output.

Apicomplexans, including species of Eimeria, pose a real threat to the health and wellbeing of animals and humans. Eimeria parasites do not infect humans but cause an important economic impact on livestock, in particular on the poultry industry. Despite its high prevalence and financial costs, little is known about the cell biology of these ‘cosmopolitan’ parasites found all over the world. In this review, we discuss different aspects of the life cycle and stages of Eimeria species, focusing on cellular structures and organelles typical of the coccidian family as well as genus-specific features, complementing some ‘unknowns’ with what is described in the closely related coccidian Toxoplasma gondii.

Cystoisospora (Isospora) belli is a coccidian parasite of humans. It can cause serious digestive disorders involving infection of intestines, biliary tract and gallbladder, especially in those with depressed immunity. It has a direct fecal–oral transmission cycle. After ingestion of sporulated oocysts, the parasite multiplies asexually and sexually within host epithelial cells, resulting in unsporulated oocysts that are excreted in feces. The details of asexual and sexual stages are not known and certain inclusions in epithelial cells in biopsy samples have been erroneously identified recently as C. belli. Here, we provide details of developmental stages of C. belli in two patients, in duodenal biopsy of one and biliary epithelium of the other. Immature and mature asexual stages (schizonts/meronts) were seen in epithelial cells. The merozoites were seen singly, in pairs and in groups in single parasitophorous vacuole (pv) in host cytoplasm. Immature and mature meronts were seen together in the same pv; up to eight nuclei were seen in meronts that retained elongated crescent shape; round multinucleated schizonts, seen in other coccidians, were not found. Meronts were up to 25 µm long and contained up to ten merozoites that were 8–11 µm long. The merozoites and meronts contained PAS-positive granules. Microgamonts (male) contained up to 30 nuclei that were arranged at the periphery and had condensed chromatin; 1–3 PAS-positive, eosinophilic, residual bodies were left when microgametes were formed. The microgametes were 4 µm long and PAS-negative. All stages of macrogamonts, including oocysts were PAS-positive. The detailed description of the life cycle stages of C. belli reported here should facilitate in histopathologic diagnosis of this parasite.

Cyclospora cayetanensis is a coccidian parasite associated with diarrheal illness. In the USA, foodborne outbreaks of cyclosporiasis have been documented almost every year since the mid-1990s. The typical approach used to identify this parasite in human stools is an examination of acid-fast-stained smears under bright-field microscopy. UV fluorescence microscopy of wet mounts is more sensitive and specific than acid-fast staining but requires a fluorescence microscope with a special filter not commonly available in diagnostic laboratories. In this study, we evaluated a new DNA extraction method based on the Universal Nucleic Acid Extraction (UNEX) buffer and compared the performances of four published real-time polymerase chain reaction (PCR) assays for the specific detection of C. cayetanensis in stool. The UNEX-based method had an improved capability to recover DNA from oocysts compared with the FastDNA stool extraction method. The best-performing real-time PCR assay was a C. cayetanensis-specific TaqMan PCR that targets the 18S ribosomal RNA gene. This new testing algorithm should be useful for detection of C. cayetanensis in human stool samples.

Coccidia are protozoal parasites which compromise mucosal integrity of the intestine, potentiating poultry morbidity. The host's Zn status influences the course of infection. Therefore, two experiments were designed to determine how supplemental Zn regimens impacted jejunal and caecal immune status and Zn transporter expression. Coccivac®-B was administered weekly at ten times the recommended dose as a mild coccidial challenge (10CV). Zn was provided through a basal diet, supplemental zinc sulfate (ZnSO4), or a supplemental 1:1 blend of ZnSO4 and Availa®-Zn (Blend). Mucosal jejunum (Expt 1) and caecal tonsils (Expt 2) were evaluated for intracellular Zn concentrations and phagocytic capacity. Messenger expression of Zn transporters ZnT5, ZnT7, Zip9 and Zip13 were investigated to determine Zn trafficking. With 10CV, phagocytic capacity was decreased in jejunal cells by 2 %. In the caecal tonsils, however, phagocytic capacity increased with challenge, with the magnitude of increase being more pronounced with higher dietary Zn (10CV × Zn interaction; P= 0·04). Intracellular Zn within caecal tonsils was found significantly reduced with 10CV (27 %, P= 0·0001). 10CV also resulted in an overall increase in the ratio of Zip:ZnT transporters. With the exception of Zip13 transporter expression, dietary Zn source had little impact on any of the measured cellular parameters. Thus, intestinal mucosal tissues had reductions in intracellular free Zn during coccidial challenge, which was coupled with an upregulation of measured Zip transporters. This suggests that under coccidial challenge, intestinal cells attempt to compensate for the drop in intracellular Zn.

Coccidiosis, a serious disease resulting from infection with parasitic protozoa of the genus Eimeria, causes significant economic losses to the poultry industry, where intensive rearing facilitates transmission of infectious oocysts via the fecal/oral route. Current control relies primarily on prophylactic drugs in feed but, whilst cost effective, the rise of drug resistance and public demands for residue-free meat has encouraged development of alternative control strategies. Chickens that recover from infection with Eimeria develop solid immunity that is directed against the early asexual stages of the parasite life cycle. This has allowed development of a number of vaccines that utilize deliberate infection with controlled doses of virulent oocysts or reproductively attenuated lines of Eimeria. The latter are immunogenic but non-pathogenic. The realization that both prophylactic drugs and attenuated vaccines control but do not eradicate infection with Eimeria encouraged development of a vaccine based upon maternal immunity. Laying hens exposed to Eimeria are able to transfer protective antibodies to hatchlings via egg yolks and these antibodies have been used to identify parasite proteins that are conserved across the genus. When delivered maternally, these provide an economical means of preventing coccidiosis, offering immediate protection to newly hatched chicks.

Eimeria is a common genus of apicomplexan parasites that infect diverse vertebrates, most notably poultry, causing serious disease and economic loss. Like all apicomplexans, eimerians have a complex life cycle characterized by asexual divisions that amplify the parasite population in preparation for sexual reproduction. This can be divided into three events: gametocytogenesis, producing gametocytes from merozoites; gametogenesis, producing microgametes and macrogametes from gametocytes; and fertilization of macrogametes by microgametes, producing diploid zygotes with ensuing meiosis completing the sexual phase. Sexual development in Eimeria depends on the differential expression of stage-specific genes, rather than presence or absence of sex chromosomes. Thus, it involves the generation of specific structures and, implicitly, storage of proteins and regulation of protein expression in macrogametes, in preparation for fertilization. In Eimeria, the formation of a unique, resilient structure, the oocyst wall, is essential for completion of the sexual phase and parasite transmission. In this review, we piece together the molecular events that underpin sexual reproduction in Eimeria and use additional details from analogous events in Plasmodium to fill current knowledge gaps. The mechanisms governing sexual stage formation and subsequent fertilization may represent targets for counteracting parasite transmission.

Coccidian oocysts from feces of 46 individuals of the garden dormouse, Eliomys quercinus (Rodentia: Gliridae), were morphologically and molecularly characterized. Both morphological and sequence data (18S rDNA and ORF 470) showed low variability, indicating that all samples represent a single species. By comparison with published morphological descriptions of coccidia from glirid rodents, we determined that the samples represent Eimeria myoxi. Molecular data suggest that this species does not fall within the 2 known rodent-specific groups but branches as a third independent lineage. However, its exact position in respect to other eimerian clusters could not be established due to the lack of phylogenetic information at this taxonomic level for the 18S rRNA and ORF 470 genes. Based on these results, we provide a re-description of Eimeria myoxi, which contains morphological and molecular characteristics sufficient for its further unequivocal identification.

For Isospora (Protozoa: Eimeriidae) parasites of passerine birds, diurnal periodicity of oocyst output is a well-described phenomenon. From the temporal zone to the tropics, oocyst production is correlated with the light-dark cycle, peaking in the afternoon hours. However, nothing is known about the existence of diurnal periodicity of these parasites in the birds of High Arctic environments, under permanent light during summer. We sampled free-ranging Snow Bunting (Aves: Passeriformes), on Svalbard in summer and tested oocysts output of Isospora plectrophenaxia. Here we show that under the permanent light conditions of Arctic summer in the wild, Isospora plectrophenaxia, a parasite of the Snow Bunting, still keeps the 24-h rhythm of oocyst output with the peak in the post-meridiem hours, despite the absence of diurnal periodicity in host's activity. Our findings prove the ability of avian Isospora to invoke alternative cues for synchronizing the circadian rhythms. Possible cues and adaptive significance of diurnal periodicity of parasite output in High Arctic are discussed. The maintenance of synchronization and timing of the parasite life-cycle stages is under positive selection pressure even in permanent daylight in the Arctic.

The Regent Honeyeater (Xanthomyza phyrigia) is an endangered Australian bird species. Breeding populations have been established at Australian zoos in support of re-introduction programmes. This species is the host of a new species of Isospora (Apicomplexa). Oocysts are spherical, 25·8 (22·5–28·75) by 23·8 (20–26·25) μm with a colourless to pale yellow smooth wall undergoing rapid exogenous sporulation, 90% sporulated oocysts in 8 h at 20°C. Each oocyst contains 1 polar granule. Sporocysts are ovoid, 18·67 (17–19) by 9·49 (9–10) μm with a flat Stieda body and spherical substieda body devoid of a hyaline body. The asexual stages and sexual phase is within the enterocytes of the duodenum and jejunum. Faeces collected in the morning (AM, n=84) and in the afternoon (PM, n=90) revealed significant diurnal periodicity in oocyst shedding; 21% (18 of 84) of the AM were positive with the mean of 499 oocysts.g−1 compared to the PM with 91% (82 of 90) bird faeces positive with the mean of 129 723 oocysts.g−1. Therefore, parasite checks for these birds should be carried out in the afternoon to obtain an accurate result. The ecological significance of the high parasite burden in captive birds requires further investigation and comparison to the wild counterparts.

The composition and structure of a community of Eimeria was investigated in a population of Eastern cottontail (Sylvilagus floridanus) introduced into Italy. Eight Eimeria species were found, of which all but 1 had North American origins and were presumably introduced into Italy together with imported cottontails. The success of cottontails in spreading microparasites is probably related to their massive release for hunting purposes. Nearly all cottontails were infected with at least 1 Eimeria species, with bimonthly prevalence ranging from 0–6·3% (E. leporis) to 42·9–89·3% (E. environ). Bayesian model averaging and multivariate techniques were used to investigate the relationships between the occurrence of each parasite and the structure of the relative community. Among the host parameters, only sex was found to be associated with the prevalence of E. honessi, while the rest of the parameters were only weakly correlated with prevalence and species richness. This indicates that individual phenotypic host characteristics are probably less important than environmental factors in determining levels of parasite prevalence and diversity. The community of Eimeria species was probably structured by competition, with less species co-occurrence than expected under a null hypothesis. This was made evident by the low co-occurrence of E. environ and E. neoirresidua with E. poudrei, E. honessi, and E. maior.

Intraerythrocytic parasites (Apicomplexa: Sarcocystidae) of the South American mouse opossum (Thylamys elegans) from Chile, South America, and of the yellow-bellied glider (Petaurus australis) from Australia were found to be monophyletic using SSU rDNA and partial LSU rDNA sequences. Phylogenetic reconstruction placed both species within the family Sarcocystidae. These intraerythrocytic parasites of marsupials represent an as yet unnamed genus predicted to have bisporocystic oocysts and tetrazoic sporocysts, which is a characteristic feature of all members of the family Sarcocystidae. These results show that erythrocytic parasites share a common ancestor and suggest co-evolution with their vertebrate host.

Before the discovery of Toxoplasma gondii as a coccidium of the cat in 1970, cat and dog coccidia were classified in the genus Isospora and considered of little clinical or zoonotic significance. Since 1970, several new (Hammondia sp., Neospora sp.) and previously described species, including Sarcocystis, Besnoitia, and Cryptosporidium have been found as coccidians of cats and dogs with clinical and zoonotic significance. In the present paper I review salient features of the evolution of cat and dog coccidia.

Codon usage of Crytosporidium parvum was compared with those of other Eimeriorina Toxoplasma gondii and Eimeria tenella and revealed a biased use of synonymous codons with a preference for NNU (40·0%) and NNA (33·4%). There was no close resemblance of the codon usage of C. parvum to T. gondii (correlation coefficient, r = 0·14) or E. tenella (r = 0·14) but it was similar to Entamoeba histolytica (r = 0·75) and Plasmodium falciparum (r = 0·5). Analysis of the codon usage in homologous gene sequences (actin, β-tubulin) also failed to reveal a close relationship between C. parvum and T. gondii or E. tenella. The low usage codons in C. parvum were most frequently used codons in T. gondii and E. tenella. These observations are consistent with 18S rRNA sequence analysis which shows no close relationship of Cryptosporidium with other Eimeriorina (Sarcocystis, Toxoplasma and Eimeria) and questions the validity of the current classification of C. parvum.

Codon usage has been analysed in individual gene sequences, derived from a variety of parasitic protozoa in the class Sporozoa of the phylum Apicomplexa using metric multidimensional scaling. The two groups of codon usage patterns detected reflect the two main subgroups of organisms studied (the coccidia and the piroplasms), and it is the pattern usage of synonymous codons that has the largest influence on overall codon usage in the individual genes, rather than being the pattern of amino acid composition of the gene product. The magnitude of the codon usage bias in the sequences was determined using three commonly used indices – NC', GC3s and B. In general, although relatively low levels of codon usage bias were detected in these gene sequences, codon usage bias does explain at least some of the codon usage patterns observed. Codon usage bias was observed to be dependent on the overall base composition of the genes analysed, which in turn was reflected in the types of codons that were either over-or under-represented in the nucleotide sequences. keeping with observations on prokaryotic organisms, it is speculated that the codon usage patterns detected in these parasitic protozoa are the result of directional mutation pressure on the base composition of the genomic DNA.

When sporozoites of Eimeria tenella were inoculated into an isolated horn of the bicornuate caecum of the domestic fowl, replicating infections, culminating in the production of oocysts, were found, almost exclusively, in the inoculated horn. In contrast, sporozoites of E. tenella introduced into the bloodstream via a superficial vein induced infections of equivalent intensity in both horns of the caecum. These findings are discussed with reference to (1) the detection (by tissue transfer) of infective sporozoites in samples of cardiac blood and vascular tissues taken within 3 to 18 h of the oral inoculation of oocysts, and (2) the (previously published) observation that the sporozoites of E. tenella are transported from the superficial epithelium of the caecum (site of invasion) to the crypts (site of initial development) within host intra-epithelial lymphocytes. We suggest that a local inflammatory response to the invasion of sporozoites may be responsible for their re-location in proximity to the site of entry. This could act by directing the homing (or retention) of the parasite-bearing lymphocytes, and/or by providing a stimulus for the sporozoite to leave its transporting cell.

The pattern of oocyst production of 8 inbred lines of chickens was compared for each of the 7 species of Eimeria which infect this host. Both the overall numbers and the pattern of oocyst production differed in the inbred lines, but there was no evidence of prolonged cycling of schizogenic developmental stages. Comparison of the numbers of oocysts produced by the different lines indicates that there may be common genetic factors affecting susceptibility to 6 of the 7 species. Surprisingly there appears to be an inverse relationship between susceptibility to E. tenella and susceptibility to the other species: lines which produced most oocysts of E. tenella produced least oocysts of the other species and vice-versa.