Oocyst Morphology Research Articles

Further characterisation of two Eimeria species (Eimeria quokka and Eimeria setonicis) in quokkas (Setonix brachyurus)

The identification and characterisation of novel Eimeria species has largely been based on sporulated oocyst and sporocyst morphology, the host species and the geographical range. Variation in the size and shape of Eimeria oocysts across their host range however, make the identification and characterisation of novel species using traditional methodologies alone problematic. The use of molecular markers and phylogenetic analysis has greatly advanced our ability to characterise Eimeria species and has recently been applied to understand evolutionary relationships among Eimeria species from Australian marsupials.In the present study, Eimeria species isolated from quokkas (Setonix brachyurus) captured from Two Peoples Bay, Bald Island and Rottnest Island, Western Australia, were morphologically identified as Eimeria quokka and Eimeria setonicis. Both Eimeria species were identified as being polymorphic in nature with regards to sporulated oocyst and sporocyst morphometrics. Phylogenetic analysis using 18S rRNA and COI (cytochrome c oxidase subunit 1) genes, grouped E. quokka and E. setonicis within the Eimeria marsupial clade together with Eimeria trichosuri from brushtail possums, Eimeria macropodis from tammar wallabies (Macropus eugenii) and several unidentified macropod Eimeria species from western grey kangaroos (Macropus fuliginosus). This study is the first to characterise E. quokka and E. setonicis by molecular analysis, enabling more extensive resolution of evolutionary relationships among marsupial-derived Eimeria species.

Prevalence and molecular characterisation of Eimeria species in Ethiopian village chickens.

BackgroundCoccidiosis, caused by species of the apicomplexan parasite Eimeria, is a major disease of chickens. Eimeria species are present world-wide, and are ubiquitous under intensive farming methods. However, prevalence of Eimeria species is not uniform across production systems. In developing countries such as Ethiopia, a high proportion of chicken production occurs on rural smallholdings (i.e. 'village chicken production’) where infectious diseases constrain productivity and surveillance is low. Coccidiosis is reported to be prevalent in these areas. However, a reliance on oocyst morphology to determine the infecting species may impede accurate diagnosis. Here, we used cross-sectional and longitudinal studies to investigate the prevalence of Eimeria oocyst shedding at two rural sites in the Ethiopian highlands.ResultsFaecal samples were collected from 767 randomly selected chickens in May or October 2011. In addition, 110 chickens were sampled in both May and October. Eimeria oocysts were detected microscopically in 427 (56%, 95% confidence interval (95% CI) 52-59%) of the 767 faecal samples tested. Moderate clustering of positive birds was detected within households, perhaps suggesting common risk factors or exposure pathways. Seven species of Eimeria were detected by real time PCR in a subset of samples further analysed, with the prevalence of some species varying by region. Co-infections were common; 64% (23/36, 95% CI 46-79%) of positive samples contained more than one Eimeria spp. Despite frequent infection and co-infection overt clinical disease was not reported. Eimeria oocysts were detected significantly more frequently in October (248/384, 65%, 95% CI 60-69%), following the main rainy season, compared to May (179/383, 47%, 95% CI 42-52%, p < 0.001). Eimeria oocyst positivity in May did not significantly affect the likelihood of detecting Eimeria oocyst five months later perhaps suggesting infection with different species or immunologically distinct strains.ConclusionsEimeria spp oocysts may be frequently detected in faecal samples from village chickens in Ethiopia. Co-infection with multiple Eimeria spp was common and almost half of Eimeria positive birds had at least one highly pathogenic species detected. Despite this, all sampled birds were free of overt disease. Although there was no evidence of a difference in the prevalence of oocysts in faecal samples between study regions, there was evidence of variation in the prevalence of some species, perhaps suggesting regional differences in exposure to risk factors associated with the birds, their management and/or location-specific environmental and ecological factors.

Evolutionary Plasticity in Coccidia – Striking Morphological Similarity of Unrelated Coccidia (Apicomplexa) from Related Hosts: Eimeria spp. from African and Asian Pangolins (Mammalia: Pholidota)

Two morphologically similar, but phylogenetically unrelated Eimeria species from ancient mammals, African Tree Pangolin Phataginus tricuspis and Sunda Pangolin Manis javanica (Pholidota: Manidae), from two distant biogeographic realms (Afrotropical and Oriental), are characterized and compared morphologically and molecularly. Phylogenetic analyses produced an unstable topology. However, while precise position of the two Eimeria species from pangolins could not be firmly established due to the lack of related taxa, it is evident that they are not closely related and do not fall into any of the so far recognized eimerian lineages. Moreover, an eimerian found in P. tricuspis is described as a new species Eimeria nkaka n. sp., based on morphology of oocysts, endogenous developmental stages and sequence data.

Determination of Phylogenetic Relationships among &lt;i&gt;Eimeria&lt;/i&gt; species, which Parasitize Cattle, on the Basis of Nuclear 18S rDNA Sequence

ABSTRACTWe analyzed almost complete 18S rDNA sequences of 10 bovine Eimeria species, namely Eimeria alabamensis, E. auburnensis, E. bovis, E. bukidnonensis, E. canadensis, E. cylindrica, E. ellipsoidalis, E. subspherica, E. wyomingensis and E. zuernii. Although these sequences showed intraspecific variation in 8 species, the sequences of each species were clustered in monophyletic groups in all species, except E. auburnensis. The sequences constituted 3 distinct clusters in a phylogenetic tree with relatively high bootstrap values; however, the members including each cluster shared no similarities in oocyst morphology.

Enteric coccidiosis in the brown kiwi (Apteryx mantelli)

Enteric coccidiosis may cause significant morbidity and mortality in juvenile brown kiwi (Apteryx mantelli). Morphology of sporulated oocysts indicates that at least two Eimeria species are able to infect the brown kiwi. A histological study of the endogenous stages of coccidia was undertaken in the intestinal tracts of ten naturally infected young kiwi. Sequential sectioning of the entire intestinal tract allowed identification and recording of the distribution of the various coccidial life stages. Macromeronts measuring 268 × 162 μm when mature were found mainly within the lamina propria of the proximal one third of the small intestine. A smaller form of lamina propria meront was also identified (8.7 × 6.4 μm) with a similar distribution to the macromeronts. Small meronts (4.4 × 3.8 μm) were also identified in mucosal epithelial cells, with the overall peak in distribution within the intestinal tract being distal to the lamina propria meronts. Three morphologically distinctive gametocytes were identified. Type A gametocytes contained within epithelial cells shared the same distribution as the epithelial meronts. Polyps containing large numbers of type B gametocytes within the distal intestinal tract were found in two cases, and type C gametocytes were identified throughout the entire intestinal tract in one case only. The observational nature of this study precludes complete knowledge of the parasite life cycles using histology alone. However, it is likely that each of the three morphologically distinct gametocytes represents a separate species of enteric coccidia.

Redescription of Protomagalhaensia blaberae Peregrine, 1970 (Apicomplexa: Eugregarinida: Blabericolidae) Parasitizing the Bolivian Cockroach, Blaberus boliviensis (Dictyoptera: Blaberidae)

Protomagalhaensia blaberae Peregrine, 1970 is redescribed from the type host, the Bolivian cockroach Blaberus boliviensis. Complete morphometric data on all life cycle stages is presented, the taxon is stabilized with the deposition of new voucher specimen material, and P. blaberae is distinguished from all other species in the genus (Protomagalhaensia cerastes, Protomagalhaensia granulosae, and Protomagalhaensia wolfi). Species of Protomagalhaensia are distinguished by differences in relative metric ratios, morphology of oocysts, and by relative metric ratios of mature gamonts in association. The status of Protomagalhaensia serpentula is considered and the taxon is determined species inquirenda.

Pinning down a polymorphic parasite: New genetic and morphological descriptions of Eimeria macropodis from the Tammar wallaby (Macropus eugenii)

Identification of the protozoan parasite, Eimeria has traditionally relied on oocyst morphology, host range and life-cycle attributes. However, it is increasingly recognized that Eimeria species can vary in size and shape across their host range, an attribute known as ‘polymorphism’ that presents a unique challenge for identification. Advances in molecular tools hold promise for characterising Eimeria that may otherwise be misclassified based on morphology. Our study used morphologic and molecular traits of the oocyst life stage to identify a polymorphic parasite, Eimeria macropodis in a captive Tammar wallaby (Macropus eugenii) population in Australia. Molecular characterization highlighted the need to use multiple genetic markers (18S SSU and cytochrome c oxidase subunit I) to accurately identify E. macropodis owing to heterozygous alleles at the 18S SSU locus. This study provided an opportunity to assess the utility and shortcomings of morphologic and molecular techniques for ‘pinning down’ a polymorphic species. Moreover, our study was able to place E. macropodis in an evolutionary context and enhance resolution of the under-studied marsupial clade.

Synoptic Revision of Blabericola (Apicomplexa: Eugregarinida: Blabericolidae) Parasitizing Blaberid Cockroaches (Dictyoptera: Blaberidae), with Comments On Delineating Gregarine Species Boundaries

Complete synoptic redescriptions, including complete morphometric data for all life cycle stages, species recognition characters, and differential comparisons are presented for the 4 gregarine species comprising Blabericola . Blabericola cubensis ( Peregrine, 1970 ), Blabericola haasi (Geus, 1969), Blabericola migrator ( Clopton, 1995 ), and Blabericola princisi ( Peregrine, 1970 ) are redescribed from their type hosts, i.e., the discoid cockroach Blaberus discoidalis , the lobster cockroach Nauphoeta cinerea , the Madagascar hissing cockroach Gromphadorhina portentosa , and the Bolivian cockroach Blaberus boliviensis , respectively. These gregarine species descriptions are stabilized through deposition of extensive new voucher collections. Species of Blabericola are distinguished by differences in relative metric ratios, morphology of oocysts, and by relative metric ratios of mature gamonts in association. This work is discussed as a model for morphological species descriptions in the Eugregarinorida including the 6 principles for morphological gregarine species descriptions, i.e., a centroid and population variation approach, adequate sample size, partitioning developmental variation and sexual dimorphism, recognition and minimization of fixation and physiological artifacts to eliminate false morphotypes, and comparative data sets across multiple life cycle stages.

Use of Artemisia annua as a natural coccidiostat in free-range broilers and its effects on infection dynamics and performance

This work investigated the preventive effect of Artemisia annua L. dried leaves supplied as a botanical coccidiostat to two broiler genotypes reared in a Danish free-range system in a factorial experiment (two genotypes and ± supplement of dried A. annua leaves). The genotypes White Bresse L40, a pure slow-growing line, and Kosmos 8 Ross, a hybrid genotype with medium growing characteristics, were used. Broilers were raised indoor until 29-days-old and kept free of parasites. Twelve groups of 30 randomly selected broilers were placed in the range forming three replicates for each treatment combination. The paddocks were cultivated with a mix of grass and clover. A separate group of broilers was naturally infected with Eimeria spp. oocysts and five animals nominated as “seeders” were introduced to the above mentioned 12 groups, 10days after its formation, with each group consisting of 35 animals per plot. This infection strategy was meant to imitate the transmission pathway observed at farm level. Ten individual birds from each of the 12 groups, in total 120 animals of mixed sex, were monitored twice weekly for 30days for oocysts excretion. PCR of pooled faecal samples, oocyst morphology and localization upon necropsy were used to identify the Eimeria species involved in the infection. In general, broilers from both genotypes in the range coped well with a coccidia infection caused by Eimeria acervulina and Eimeria maxima as no clinical symptoms, or deaths, were reported during the experiment. In general, broilers supplemented with A. annua dried leaves showed a significantly (p<0.05) reduced number of excreted oocysts during the infection with no interaction to genotype. Females generally had a significantly higher shedding of oocysts than males (p<0.05). The overall body weight gain and the daily weight gain when infection was subdued showed a three-way interaction among genotype, sex and treatment – accounted mainly for the fact that Kosmos females responded positively to the Artemisia treatment while Kosmos males responded negatively, and only minor differences were found between sexes for the White Bresse genotype. In conclusion, supply of A. annua dried leaves as a botanical coccidiostat significantly reduced oocyst output in free ranged broilers and thus may form part of a strategy to prevent commercial losses.

Description of the two strains of turkey coccidia Eimeria adenoeides with remarkable morphological variability

Although oocyst morphology was always considered as a reliable parameter for coccidian species discrimination we describe strain variation of turkey coccidia, Eimeria adenoeides, which remarkably exceeds the variation observed in any other Eimeria species. Two strains have been isolated - the first strain maintains the typical oocyst morphology attributed to this species - large and ellipsoidal - while the second strain has small and ovoid oocysts, never described before for this species. Other biological parameters including pathogenicity were found to be similar. Cross-protection between these 2 strains in 2 immunization and challenge experiments was confirmed. Sequencing and analysis of 18S and ITS1 ribosomal DNA revealed a close relationship according to 18S and a relatively distant relationship according to ITS1. Analysis of 18S and ITS1 sequences from commercial turkey coccidiosis vaccines Immucox®-T and Coccivac®-T revealed that each vaccine contains a different strain of E. adenoeides and that these strains have 18S and ITS1 sequences homologous to the sequences of the strains we have isolated and described. These findings show that diagnostics of turkey coccidia according to oocyst morphology have to be carried out with caution or abolished entirely. Novel PCR-based molecular tools will be necessary for fast and reliable species discrimination.

Construction of PCR Primers to Detect and Distinguish Eimeria spp. in Northern Bobwhites and a Survey of Eimeria on Gamebird Farms in the United States

Coccidiosis is an important disease in captive gamebirds, including northern bobwhites (Colinusvirginianus). Three Eimeria species, Eimeria lettyae, Eimeria dispersa, and Eimeria colini, have been described in bobwhites. Distinguishing the various Eimeria spp. is often problematic because of similarity in oocyst morphology and site of infection and thus requires live bird infections to distinguish between the coccidian species. To aid in identification and diagnosis, PCR specific primers were generated against the internal transcribed spacer region 1 (ITS-1) of the ribosomal RNA gene using sequences obtained from coccidian-positive samples collected from diagnostic cases or litter from captive bobwhites. Three distinct Eimeria spp. were detected. Species-specific primers were constructed and used to survey the prevalence of the species in 31 samples collected from 13 states. The primers survey results identified E. lettyae, E. dispersa, and Eimeria sp. in 20 (64.5%), 22 (71%), and 29 (93.5%) of the samples, respectively. Mixed infections were common: 13 (41.9%) samples had 3 Eimeria spp., 14 (45.2%) had 2 spp., and 4 (12.9%) samples had only 1 species. The species were widely distributed over the area sampled and were not associated with the age of the flock.

The molecular characterization of an Eimeria and Cryptosporidium detected in Asian seabass (Lates calcarifer) cultured in Vietnam

An intestinal Eimeria was previously reported as a significant pathogen of Asian seabass (Lates calcarifer) in nurseries in Vietnam. In the present study, both Eimeria and Cryptosporidium were detected by sequence analyses of fragments of the 18S rRNA gene amplified from these Vietnamese L. calcarifer tissues. Based on these analyses, the Eimeria from the Vietnamese L. calcarifer formed clades with the Eimeria detected in L. calcarifer tissues from Australia, but clustered separately from other known Eimeria and Goussia species. The Cryptosporidium detected in L. calcarifer from Vietnam clustered closest with C. parvum and C. hominis. In situ hybridization using DIG-labeled DNA probes generated from 18S PCR products on the Vietnamese L. calcarifer wax block tissues showed that this method could not be used to distinguish between Eimeria and Cryptosporidium, due to the conserved nature of the 18S locus. A previously published study on the morphology of parasite developmental stages and oocysts in the Vietnamese L. calcarifer tissues showed only an intestinal Eimeria infection. The Cryptosporidium could be present at very low levels undetectable by microscopy in intestines, or being ubiquitous, was a possible contaminant from feed or water. While molecular analysis is a very useful tool in the study of disease and identification of aetiological agents, this study reiterates the importance of demonstrating organisms in situ in tissues.

New Insights into the Excystation Process and Oocyst Morphology of Rodent Eimeria Species

In this study, the mechanism of excystation of the rodent parasites Eimeria nieschulzi, from rats, and Eimeria falciformis, from mice, was investigated. In vitro, oocysts of both species are susceptible to the protease pepsin, and sporocysts and sporozoites can be excysted in a similar way. Scanning electron microscopy (SEM) revealed a collapse of the oocysts wall at both polar ends after pepsin treatment. This occurs without any visible damage of the outer wall. Using fluorescence and transmission electron microscopy (TEM) we observed that pepsin enters sporulated oocysts at both polar ends and causes degradation of the inner oocyst wall. Using scanning electron microscopy we could identify two polar caps in both investigated rodent Eimeria species, but only one is harbouring the micropyle. Thus the polar caps are the entry site for the pepsin. Furthermore, we provide evidence that the oocyst cap and micropyle are functionally different structures. This study complements the morphological description of both Eimeria species and is of relevance for other coccidian species.

Disseminated visceral coccidiosis in Eurasian cranes (Grus grus) in the UK

Clinical disease and mortalities due to disseminated visceral coccidiosis were identified for the first time in a group of captive juvenile Eurasian cranes (Grus grus) in the UK during 2008....

Redescription of Protomagalhaensia granulosae Peregrine, 1970 (Apicomplexa: Eugregarinida: Blabericolidae) Parasitizing the Discoid Cockroach, Blaberus discoidalis (Dictyoptera: Blaberidae)

Protomagalhaensia granulosae Peregrine, 1970, is redescribed from the type host, the discoid cockroach, Blaberus discoidalis. Complete morphometric data on all life cycle stages is presented, and P. granulosae is distinguished from other species in the genus and stabilized with the deposition of new voucher specimen material. Species of Protomagalhaensia are distinguished by differences in relative metric ratios, morphology of oocysts, and by relative metric ratios of mature gamonts in association.

New species ofEimeriaandIsospora(Protozoa: Eimeriidae) inGeochelonespp. (Chelonia: Testudinidae) from Amazonian Brazil

Tetrasporocystic, dizoic oocysts of reptiles have been separated by some authors into the genera Eimeria, Choleoeimeria and Acroeimeria (Protozoa: Eimeriidae), based on the site and mode of development of their endogenous stages. The majority of Eimeria species have been, and still are, however, described on oocyst morphology alone. Four different oocysts with this basic morphology were encountered in the faeces of Brazilian tortoises, Geochelone carbonaria Spix, 1824 and are assigned to the genus Eimeria, with the view that they can readily be transferred to the genus Choleoeimeria or Acroeimeria if this is indicated by a future examination of their endogenous development. A morphological comparison distinguishes the oocysts from those of Eimeria spp., previously described in chelonids of the family Testudinidae, and the names E. amazonensis, E. carbonaria, E. carajasensis and E. wellcomei n. spp. are proposed. Coccidial infection appears to be common in G. carbonaria, with three of seven animals examined passing oocysts. Oocysts of Isospora rodriguesae n. sp. (Protozoa: Eimeriidae) are described in the faeces of Geochelone denticulata Linnaeus, 1766. They are morphologically very different from those of Isospora testudae, Davronov, 1985 in Testudo horsfieldi. Eimeria motelo Hůrková et al., 2000, previously described in Geochelone denticulata from Peru, is here recorded in the some chelonid from Amazonian Brazil.

First record of Eimeria furonis infection in a ferret, Japan, with notes on the usefulness of partial small subunit ribosomal RNA gene sequencing analysis for discriminating among Eimeria species

Eimeria is a genus of apicomplexan parasites found in a variety of vertebrates including the weasel. At present, three species have been reported in members of the weasel family, but the presence of weasel Eimeria in Japan have been quite unclear. The identification of Eimeria species has been performed based on oocyst morphology, host species, and habitat in the host, but sometimes discriminating among morphologically similar species under light microscopy is impossible. The present study detected for the first time the weasel coccidium, E. furonis, in a ferret in Japan. Since molecular information for E. furonis has been quite unclear, this species was compared molecularly with other Eimeria species, and also the usefulness of sequencing analysis of the small subunit ribosomal ribonucleic acid gene (SSUrDNA) for discriminating among Eimeria species was examined. About the 350-bp sequence of SSUrDNA of all species including E. furonis compared differed from each other, and its differences found in Eimeria species were also reflected in the phylogram constructed using the partial nucleotide sequences. The sequencing analysis of partial SSUrDNA examined in the present study thus appears useful for discriminating among morphologically similar Eimeria species.

Occurrence of gastrointestinal protozoa in Didelphis albiventris (opossum) in the central region of Rio Grande do Sul state

The aim of this study was to evaluate the parasitism by gastrointestinal protozoa in Didelphis albiventris (opossum) in the central region of Rio Grande do Sul state. Fecal samples from six free living opossums were collected for research of parasites. Samples were analyzed by the centrifugal-flotation method with zinc sulfate and parasites were identified microscopically based on (oo)cyst size and morphology. Cysts of Giardia sp. and oocysts of Cryptosporidium sp. and Eimeria sp. were observed in four of the six opossums. All four infected marsupials showed mild infection by protozoa. This is the first report of Giardia sp. in D. albiventris.

La cryptosporidiose zoonosique humaine due àCryptosporidium felisdans le monde

Cryptosporidium is an important enteric pathogen worldwide distributed causing diarrhoeal illness in humans and animals. Identifying Cryptosporidium species using conventional criteria, such as oocyst morphology, is inadequate. The advent of molecular techniques has conducted to characterize different species and genotypes of Cryptosporidium infecting humans. The vast majority of human cases of cryptosporidiosis in the world are caused by both species, Cryptosporidium hominis and Cryptosporidium parvum. However other species including Cryptosporidium felis can infect humans too. In this review, we analyse 58 reported cases of human C. felis infection in different parts of the world. To date this emerging protozoan disease is present in humans around the world, except in Australia and Oceania. Adults and children are infected, more often when immunocompromised by HIV infection (83 % of reported cases). Apparently immunocompetent individuals are also infected by C. felis. In developing countries, inhabitants are more likely infected by C. felis probably through the oocyst contamination of drinking or recreational water. The public health importance of C. felis infection in tropical countries remains to be evaluated.

Comparison of the morphology of oocysts and the phylogenetic analysis of four Ascogregarina species (Eugregarinidae: Lecudinidae) as inferred from small subunit ribosomal DNA sequences

This study on the ultrastructure of the oocysts of four isolated species of Ascogregarina ( A. taiwanensis (Lien and Levine) (Eugregarinidae: Lecudinidae) from Aedes albopictus (Skuse), A. culicis (Ross) (Eugregarinidae: Lecudinidae) from Aedes aegypti (L.), A. armigerei (Eugregarinidae: Lecudinidae) from Armigeres subalbatus (Coquillet), and Ascogregarina sp. (Eugregarinidae: Lecudinidae) from Ochlerotatus japonicus japonicus (Theobald)) using a scanning electron microscope revealed significant differences in size and in surface structure. The average length of the oocyst was greatest in A. armigerei (13.2 ± 0.2 μm) (mean ± SD) and least in A. culicis (8.8 ± 0.4 μm). Oocysts were of moderate length in A. taiwanensis (9.9 ± 0.6 μm) and in Ascogregarina sp. (10.7 ± 1.1 μm) isolated from O. j. japonicus. The ultrastructure of the surface of the A. culicis oocyst was rough in texture with numerous dense spots and was easily distinguishable from the oocysts of the other three Ascogregarina spp. The maximum likelihood tree inferred from small subunit ribosomal DNA (SSU rDNA) sequences indicated that the four Ascogregarina spp. form a monophyletic cluster among other gregarine parasites. Within the Ascogregarina clade, A. culicis, A. taiwanensis, and Ascogregarina sp. from O. j. japonicus showed a close relationship, whereas A. armigerei was a distantly related species.