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The cathepsin-D expression in the cyto-invasive stages of coccidia in intestinal mucosa of chickens, and the relevance of this proteinase for intracellular penetration and spreading of coccidia in host tissues.

The Cathepsin-D proteinase was demonstrated immunohistochemically in cases of intestinal coccidiosis in chickens; its expression concerned almost the cyto-invasive stages of reproductive cycle of several strains of coccidia (merozoites of the II-nd generation and also microgametes). The authors consider Cathepsin-D as a potentional "penetration/spreading factor", being significant for intracellular invasion and spreading of coccidia into host cells. In diagnostic practice, an immunohistochemical demonstration of Cathepsin-D is a relatively simple and sensitive method, which brings information on the extent of parasitic lesions, and on participation of particular evolutive forms of coccidial life cycle; it may also be useful in proving isolated coccidia in the intestinal epithelium, poorly recognized in routine HE preparation. Some Cathepsin-D protease reactivity in intestinal Cryptosporidiosis is also briefly mentioned. The examined cases of cerebral toxoplasmosis and primary acanthamoebic meningoencephalitis (PAME) showed in above mentioned methods negative results.

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[Eight new species of coccidia (Sporozoa, Coccidia) in fishes from the continental waters of Russia].

Description of coccidian species infecting freshwater fishes is given. Localities: Neman river, Goussia peleci sp. n.--oocyst 37.5-45 x 35-42.5 microns, oocyst residuum (OR) absent, sporocysts 20-22.5 x 15-17.5 microns, Stieda body (SB) absent, sporocyst residuum (SR) present, host (h)--Pelecus cultratus; G. cultrati sp. n.--oocyst 22.5-30 microns, OR absent, sporocysts 12.5 x 15 microns, SR, SB absent, wall 2-2.5 microns, h--P. cultratus; G. arinae sp. n.--oocyst 12.5 x 17.5 microns, OR present, sporocysts 5-7.5 microns, SR, SB absent, h--P. cultratus; G. vimbae sp. n.--oocyst 15-22.5 microns, OR absent, wall 2.5 microns, sporocysts 5-7.5 microns, SR, SB absent, h--Vimba vimba vimba; G. gymnocephali sp. n.--oocyst 25 x 25 microns, OR absent, sporocysts 10 x 12.5 microns, SR, SB absent, h--Gymnocephalus cernuus; G. cernui sp. n.--oocyst 15-22.5 microns, OR present, sporocysts 5-12.5 microns, SR, SB absent, h--Gym. cernuus; G. luciopercae sp. n.--oocyst 30-35 x 30 microns, OR absent, sporocysts 12.5-15 x 10-12.5 microns, SB absent, SR present, h--Stizostedion lucioperca; Eimeria fluviatili sp. n.--oocyst 20-22.5 microns, OR present, sporocysts 7.5 x 12.5 microns, SB present, SR absent, h--Perca fluviatilis.

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Concurrent infections with reoviruses and coccidia in broilers.

These experiments investigated the interaction among two species of coccidia (Eimeria acervulina and E. mitis) and three strains of reovirus (virus 2035, a weak to moderate pathogen; and viruses 2408 and 1733, severe pathogens). When reoviruses were not present, high inoculation dosages (10(6) sporulated oocysts/bird) of both E. acervulina and E. mitis depressed weight gain, plasma pigment, and plasma protein. Low doses of coccidia (10(4) oocysts) in the absence of virus had no such effect on weight gain. When high doses of coccidia were present at the same time as virus 2035 or 2408, they resulted in a significantly greater depression of weight gain than when either virus or coccidia were present alone. With virus 2035, this greater depression was seen even when low doses of coccidia were used. Lesion scores due to coccidiosis and the number of oocysts produced were not affected by previous exposure to reovirus. Both coccidiosis and reovirus infections increased the frequency of some leg problems and other abnormal conditions. The most obvious interaction between coccidia and reovirus was the marked increase in swollen hocks seen when coccidia and virus 2035 were present together (20-27%) compared with either the virus or coccidia alone (0-10%). Virus 2408 interfered slightly with the development of immunity to coccidia. There was some indication that early coccidiosis could increase the ability of some virus isolates to infect various tissues of the host.

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The conceptual basis for a new classification of the coccidia.

At the joint meeting of the 8th International Coccidiosis Conference and the Annual Scientific Meeting of the Australian Society for Parasitology in Palm Cove, Australia, in July 2001, a Controversial Roundtable was held on 'New classification of coccidia'. The aim of this Roundtable was to stimulate and encourage discussion and debate on current classification schemes for the group of parasitic protozoa known as the eimeriid coccidia. In the past, such classifications have been based only on phenotypic characters such as morphology, ultrastructure, life cycles, and host specificity. However, over the past 10-15 years, molecular phylogenetic studies on taxa of the eimeriid coccidia have revealed that several of the families, subfamilies, and genera that have been erected based on non-molecular characters are paraphyletic. Therefore, this Roundtable was an important forum for initial discussions on how a new and more comprehensive classification of the eimeriid coccidia, which takes into consideration both phenotypic and molecular characters, can be devised. The stimulus came from invited speakers who gave introductions into selected areas of taxonomy and classification. Following these introductions, a more general discussion with the audience addressed potential steps that may be taken in future work. This review is the immediate outcome of the Roundtable. It describes advantages and disadvantages of the use of phenotypic or molecular characters as the base for taxonomic schemes for eimeriid coccidia. It gives specific examples for drawbacks of current classifications based only on phenotypic characters as well as potential pitfalls associated with the use of only molecular phylogenies. It addresses current controversies as well as rules of taxonomy and nomenclature relevant for the eimeriid coccidia. Finally, it recommends the establishment of an international group of scientists to meet on a regular basis, stimulate further discussions, and give direction on how the final goal, i.e. a proposal for a revised, and widely accepted, classification of the eimeriid coccidia, may be achieved.

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Evaluation of interspecific hybrids of the chicken, guinea fowl, and Japanese quail for innate resistance to coccidia.

Experimental chicken/guinea fowl hybrids, guinea fowl, and chickens were orally inoculated with Eimeria acervulina or E. tenella, which are specific for chickens, or with E. grenieri, which is specific for guinea fowl. No intact oocysts were found in feces within 24 hr of inoculation, suggesting that excystation occurred in the normal and abnormal hosts. No oocysts were found in the feces of hybrids during a 9-day postinoculation period. The guinea fowl passed oocysts of guinea fowl coccidia (E. grenieri) but not those of chicken coccidia, and the chickens passed oocysts of chicken coccidia (E. acervulina and E. tenella) but not those of guinea fowl coccidia. Some asexual development (schizogony) occurred in hybrids inoculated with E. tenella, but sexual development (gametogony) did not. In contrast, quail/chicken hybrids became infected with oocysts of chicken coccidia (E. acervulina, E. tenella, and E. maxima) and quail coccidia (E. bateri) and passed a few oocysts during the normal patent period; control chickens and quails became heavily infected with oocysts of chicken and quail coccidia, respectively.

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[Paul-Louis Simond and coccidia].

In 1880, during a sojourn at Constantine, Alphonse Laveran discovered the etiological agent of human malaria. During his microscopic observations of the parasite in freshly collected blood, Laveran's attention was attracted to the movement of flagellar bodies in the preparations. For Laveran these flagellar bodies corresponded to living organisms, in fact he considered them the most characteristic stage of the parasite; perhaps the sexual form of the parasite. In 1884, back in Paris at the military hospital Val de Grâce, A. Laveran showed these flagellated bodies to Pasteur, Roux and Chamberland who all thought that is was impossible not to recognize a living body in this mass of protoplasm displacing the surrounding red blood cells with its protruding flagella. As early as 1890, Elie Metchnikoff established a link between the flagellar bodies of Laveran and the stealthy stage of parasitic Coccidia infecting the intestinal epithelium of salamanders. It was the Pasteuriens and a few others scientists like Danilewski and Pfeiffer who firmly believed that the flagellar forms were indeed a normal stage during the hematozoan life cycle. On the contrary, Grassi and the Italian school, as well as the French protozoologist Labbé were convinced that the flagellar bodies corresponded to a degenerative form of the parasite provoked by the exposure of the parasites to air during slide preparation. In early 1896, P-L Simond joined Metchnikoff's laboratory and was assigned to study salamander coccidia in order to clarify the nature of the flagellar bodies. In a very short period of time, Simond clearly demonstrated that the coccidia had two types of life cycles; one of which resulted in the formation of flagellar bodies. Simond called the flagellar bodies "chromatozoites" due to the important quantities of chromatin twisted around the flagellum. From these observations and the sperm-like movement of the chromatozoites, Simond put forth the hypothesis that the chromatozoites were the male sexual forms of the parasite. He noted that they were found in all species of coccidia, as well as in the different species of malaria causing hematozoans of man and birds. Simond went even further in his interpretation to suggest that malaria parasites undergo sexual reproduction. This was indeed true, but Simond thought erroneously that sexual reproduction in hematozoans would result in the formation of resistant spores like in the Coccidia. The sexual reproduction hypothesis of malaria parasites was also formulated independently a year later by W. G. MacCallum (1898) working in the United States on Halteridium, a hematozoan infecting crows. Initially MacCallum was not aware of Simond's work, but later gave Simond full credit for his work on Coccidia. Furthermore, MacCallum observed for the first time the chromatozoite (microgamete) enter a female element (macrogamete) to form a mobile worm-like stage of the parasite know today as the ookinete. In his writings Simond is very modest, giving most of the credit to his mentors Metchnikoff and Laveran, but Laveran in an article which appeared in 1899 recognizes P.-L. Simond as the first person having put forth the hypothesis that the flagellar bodies of hematozoans causing human and bird malaria were the male forms of the parasite destined to fertilize the female elements.

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[Formation and diversity of parasitophorous vacuoles in parasitic protozoa. The Coccidia (Sporozoa, Apicomplexa)].

Data on parasitophorous vacuole (PV) formation in host cells (HC) harbouring different intracellular protozoan parasites have been reviewed and critically analysed, with special reference to the main representatives of the Coccidia. The vacuole membrane (PVM) is the interface between host and parasite, playing a role in nutrient acquisition by the parasite from the HC. The PV phenomenon is regarded as a generalized HC response to the introduction of alien bodies (microorganisms), which eventually reflects the evolutionary established host-parasite relationships at cellular, subcellular and molecular levels. Special attention has been paid to the existing morpho-functional diversity of the PVs within the same genera and species of parasites, and even at different stages of the parasite life cycle. The PVM is generally considered to derive from the HC plasmalemma, whose biochemical composition undergoes significant changes as the intravacuolar parasite grows. The original HC proteins are selectively excluded from the PVM, while those of the parasite are incorporated. As the result, the changed PVM becomes not fusigenic for HC lysosomes. For Toxoplasma gondii and other cyst-forming coccidia (Isospora, Sarcocystis), a definite correlation has been noticed between the extent of rhoptry and dense granule secrets released by a zoite during HC internalization, on the one hand, and the pattern of the PV that forms, on the other one. In T. gondii, tachyzoites, known to discharge abundant secrets, commonly force the development of PVs limited with a single unit membrane and equipped with a tubulovesicular network in the lumen. Unlike, bradyzoites known to be deficient in secretory materials trigger the formation of PVs with a three-membrane lining composed of the changed invaginated plasmalemma in addition to two membranes of endoplasmic reticulum. The two different types of PV harbour, respectively, exoenteric and enteric stages of T. gondii, the latter being confined to the cat intestine only. Unlike, all endogenous stages of the classic intestinal coccidia (Eimeria spp.) develop within PVs limited with a single membrane, with some invaginations extending into the PV lumen. Unusual PV patterns are characteristic of the extracytoplasmic eimerian coccidia (Cryptosporidium, Epieimeria) and adeleid haemogreagarines (Karyolysus). In cyst-forming coccidia, the PVM is actively involved in tissue cyst wall formation, thus protecting the encysted parasites from recognition by the host immune system. All this strongly suggests that the PV is far from being an indifferent membraneous vesicle containing a parasite, but represents a metabolically active compartment in infected cells. Since all the coccidia are obligate intracellular parasites, the mode of their intimate interaction with the HC, largely accomplished via the PV and its membrane, is vital for their survival as biological species.

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Drug sensitivity of coccidia from broiler breeder pullets and from broilers in the same integrated company.

Coccidia isolated from 12 breeder pullet farms and 18 broiler farms were tested for sensitivity to anticoccidial drugs. The species Eimeria acervulina, E. maxima, and E. tenella were common in the isolates and were fully pathogenic. Broiler coccidia were generally resistant to Zoalene but sensitive to nicarbazin. About half of the isolates had reduced sensitivity to amprolium and monensin. Most of the breeder pullet coccidia were resistant to Zoalene and amprolium, but almost all were highly sensitive to nicarbazin and monensin. There was some difference in responses of broiler isolates to various ionophores. Isolates responded best to salinomycin and lasalocid, which had not been previously used on the farms. Monensin controlled coccidia slightly better when given at 120 ppm than when given at 100 ppm. These results suggest that broiler coccidia have lost some responsiveness to ionophores as a result of long-term exposure, but that breeder pullet coccidia, which have never been exposed to those drugs, have retained their sensitivity. Resistance or sensitivity to other drugs was consistent with history of use: more isolates were sensitive to older drugs that had not been used for several years than was reported in previous surveys.

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A plea for improved presentation of type material for coccidia.

The "true" coccidia (phylum Apicomplexa, suborder Eimeriina) constitute a large and heterogeneous group of parasitic protozoa. Despite the large number of described species (ca. 1,650) and the medical and veterinary importance of some (e.g., Toxoplasma), 2 facts are clear: (1) the majority of coccidia species are probably yet undescribed, and (2) the phylogenetic relationships of those described species are poorly known. Contributing to the latter dilemma is the lack of a tradition to provide type specimens by those who describe new species, even though the International Code of Zoological Nomenclature specifically recommends the designation of a type specimen with the description of a new species. With the publication of a new edition of the Code (1985), explicit provisions are made for the unique concerns of taxonomists working with Protozoa. Here we remind those interested in the taxonomy of coccidia of an already established method for preserving oocysts in resin and, as an alternative, suggest the standardization of a photographic procedure through which type specimens of coccidia oocysts might also be submitted to and maintained in accredited museums. Thus, coccidia taxonomists should no longer have an excuse for their failure to designate types.

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Porcine coccidia in Papua New Guinea.

Faecal samples from 232 domestic pigs raised on concrete, 98 free-ranging village pigs, and five wild boar showed 46.6 (108/232), 54 (53/98) and 80% (4/5) prevalence of coccidian oocysts, respectively. Eight species of Eimeria, and Isospora suis, were recovered. In their descending order of predominance in the pigs raised on concrete, the species of coccidia were E. debliecki (26.7%), E. scabra (22.4%), E. neodebliecki (19.8%), E. porci (15.5%), E. suis (11.6%), E. polita (8.6%), E. perminuta (7%), E. spinosa (5.6%) and I. suis (3.9%). The first five species listed above predominated in the village pigs as well. E. polita, E. spinosa and I. suis were not found in the wild boar. I. suis oocysts prevailed in 8.3% of the 36 sows on concrete, and in 11.1% (3/27) of those which were positive for coccidia. Isosporoid oocysts were absent in the village sows. Of the 125 less than 24-day-old piglets, 29.6% were diarrhoeic, and of these, 43.2% were positive for coccidia. Four of the 16 (25%) coccidia-positive, diarrhoeic piglets, and four of the 37 (10.8%) coccidia-positive non-diarrhoeic piglets shed I. suis oocysts, an observation which seems to weaken the present contention that I. suis is the primary causative agent of neonatal porcine coccidiosis. The highest mean number of oocysts per gram faeces (23,550) was recorded from the diarrhoeic farm piglets on concrete, and the lowest of 6,100 from the gestating farm sows. Mean opg data revealed very little significant quantitative variation between the corresponding age groups of the free-ranging village pigs and the commercially-farmed ones. One of the most interesting findings in the study was that the sows were more frequently infected than all other age groups.

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Prevalence and identity of coccidia in pen-raised wild turkeys.

One hundred nineteen pen-raised wild turkeys (Meleagris gallopavo) from 12 locations in nine states in the United States were examined for coccidia by sugar flotation of intestinal contents and mucosa or by subinoculating the contents into uninfected domestic turkeys. Seventy-eight (66%) of the turkeys were positive for coccidia. There were no differences in the frequency of coccidia among adult, sub-adult or juvenile turkeys. More females (75%) were infected than males (48%). The species of coccidia from 30 of the turkeys were identified based on microscopic examination of oocysts, fresh scrapings, stained sections and inoculations of bobwhites (Colinus virginianus). The frequency of each species was Eimeria meleagrimitis (97%), E. gallopavonis (47%), E. meleagridis (27%), E. dispersa (17%), E. innocua-E. subrotunda (13%), E. adenoeides (7%) and an undescribed species (3%). Of the 30 turkeys in which the species of coccidia was determined, 30% had a single species infection, 40% had two species, 20% had three species and 10% had four species.

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The current status of the small subunit rRNA phylogeny of the coccidia (Sporozoa).

There is no current comprehensive assessment of the molecular phylogeny of the coccidia, as all recently published papers either deal with subsets of the taxa or sequence data, or provide non-robust analyses. Here, we present a comprehensive and consistent phylogenetic analysis of the available data for the small-subunit ribosomal RNA gene sequence, including a number of taxa not previously studied, based on a Bayesian tree-building analysis and the covariotide model of evolution. The assumptions of the analysis have been rigorously tested, and the benefits and limitations highlighted. Our results provide support for a number of prior conclusions, including the monophyly of the families Sarcocystidae (cyst-forming coccidia) and Eimeriidae (oocyst-forming coccidia), but with bird-host Isospora species in the Eimeriidae and mammal-host species in the Sarcocystidae. However, it is clear that a number of previously reported relationships are dependent on the evolutionary model chosen, such as the placements of Goussia janae, Lankesterella minimia and Caryospora bigenetica. Our results also confirm the monophyly of the subfamilies Toxoplasmatinae and Sarcocystinae, but only some of the previously reported groups within these subfamilies are supported by our analysis. Similarly, only some of the previously reported groups within the Eimeriidae are supported by our analysis, and the genus Eimeria is clearly paraphyletic. There are unambiguous patterns of host-parasite relationship within the coccidia, as most of the well-supported groups have a consistent and restricted range of hosts, with the exception of the Toxoplasmatinae. Furthermore, the previously reported groups for which we found no support all have a diverse range of unrelated hosts, confirming that these are unlikely to be natural groups. The most interesting unaddressed questions may relate to Isospora, which has the fewest available sequences and host-parasite relationships apparently not as straightforward as elsewhere within the suborder.

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Prevalence and risk factors for Giardia and coccidia species of pet cats in 2003-2004.

Prevalence and risk factors for feline coccidia and Giardia species infections were estimated for cats visiting 434 Banfield hospitals in 40 states in 2003-2004. Evaluated were 631,021 cats making 1,456,712 office visits (encounters) and having 211,105 fecal examinations. The overall fecal prevalences of coccidia and Giardia species were 1.4% and 0.58%, respectively. Cats at increased risk of coccidia infection were under 4 years of age, intact, and seen during the summer, fall, and spring months compared to winter. Cats at increased risk of Giardia species infection were under 4 years of age. Those at decreased risk were mixed breed and seen during the summer, fall, and spring. The highest regional risk of coccidia and Giardia species infection was for cats in the East South Central region and Mountain region, respectively, compared to the South Pacific region.

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The seasonal incidence of coccidia infections in trade cattle, sheep, and goats in Nigeria.

A survey of coccidia infections in trade cattle, sheep, and goats was undertaken in Nigeria between April 1978 and March 1979. Faecal examinations showed coccidia oocysts in 1,456 (56 per cent) of 2,600 calves, 832 (80 per cent) out of 1,040 sheep, and 468 (45 per cent) out of 1,040 goats. There were relatively few coccidia oocysts between October and March and peaks occurred in August and September. Nine species of Eimeria were identified in bovine faeces, and seven species in both sheep and goats. Eimeria bovis and E. zurnii are predominant in cattle, whereas E. faurei and E. ninakohlyakimovae have the highest percentage occurrence in sheep and goats in Nigeria. The effects of seasonal influences on the abundance of coccidia oocysts among ruminants in Nigeria are stressed.

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Attempted cross-transmission of coccidia between sheep and goats and description of Eimeria ovinoidalis sp. n.

Attempted infection of 2 young lambs with oocysts of Eimeria christenseni from a goat was unsuccessful. Negative results were obtained also when young kids were fed oocysts of Eimeria ninakohlyakimovae from sheep. There was no difficulty in infecting lambs with the sheep coccidium resembling E. ninakohlyakimovae nor goats with the goat coccidium E. christenseni. Oocysts from the goat measured 38.4 X 26.7 microns, but were easily distinguished from Eimeria ahsata from the sheep by sporocyst size and shape, and from Eimeria ovina by oocyst size. Eimeria ninakohlyakimovae-like oocysts from sheep averaged 23.0 X 18.2 microns and were morphologically indistinguishable from previously reported goat coccidia. Since no cross infections of sheep and goats could be accomplished with oocysts of Eimeria sp. characteristic of one or the other host, I concluded that sheep coccidia previously known as E. ninakohlykimovae are distinct from morphologically similar goat coccidia and therefore constitute a separate species. Since the name E. ninakohlyakimovae was first used for coccidia from the goat, the sheep coccidium is renamed Eimeria ovinoidalis with oocyst structure and endogenous stages similar to those previously described from the sheep.

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