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Fine structure of the oocyst walls of Isospora serini and Isospora canaria and excystation of Isospora serini from the canary, Serinus canarius L.

Oocysts of Isospora serini and Isospora canaria, from the canary Serinus canarius, were broken, added to a cell suspension, fixed in Karnovsky's fluid, and studied in the electron microscope. The oocyst wall of each species had an electron-lucent inner layer, a more osmiophilic middle layer and an outer layer of electron-lucent (I. serini) or electron-dense material interspersed with some electron-lucent material (I. canaria). A few, relatively large lipid-like bodies were present in the outer or middle layer of the oocyst wall of I. canaria. As many as 9 membranes were present in the oocyst wall of I. canaria and 3 in that of I. serini. When exposed to a trypsin-sodium taurocholate fluid, sporozoites of I. serini excysted from 5-month-old sporocysts in vitro, but not from sporocysts stored for more than 6 months. No excystation occurred in 15-month-old I. canaria sporocysts. Similarities and differences in excystation between I. serini and other Isospora, Eimeria, and Sarcocystis species are discussed.

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The genus Atoxoplasma (Garnham 1950) as a junior objective synonym of the genus Isospora (Schneider 1881) species infecting birds and resurrection of Cystoisospora (Frenkel 1977) as the correct genus for Isospora species infecting mammals.

Molecular and morphological data permit a rational subdivision of the paraphyletic Isospora into 2 apparently monophyletic groups of parasites, i.e., Isospora and Cystoisospora. Atoxoplasma was determined to be a junior objective synonym for Isospora. Tetrasporozoic, diplosporocystic oocysts possessing Stieda bodies in their sporocysts belong to Isospora (Eimeriidae) and have been described principally from the feces of birds. Tetrasporozoic, diplosporocystic oocysts without Stieda bodies in their sporocysts belong to Cystoisospora (Sarcocystidae).

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Excystation of Isospora arctopitheci Rodhain, 1933 with notes on a similiar process in Isospora begemina (Stiles, 1891) Lühe, 1906.

The in vitro excystation process of sporozoites of Isospora arctophitheci Rodhain, 1933 from the titi marmoset Saguinus geoffroyi and of Isospora bigemina (Stiles, 1891) Lühe, 1906 from the bobcat, Lynx rufus is presented. Sporocysts of both species lack a Stieda body and when exposed to a trypsin-sodium taurocholate (pH 7.4) excysting fluid the walls of both collapse in a similar fashion, along apparently predetermined lines. Similarities and differences on excystation between. I. arctopitheci, I. bigemina, and other Isospora, Eimeria, and Sarcocytis species are summarized. Such studies show that 2 distinct patterns of sporozoite excystation have been described to date, and both appear to be related to the structure of the sporocyst.

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Exogenous stages of Isospora serini (Aragão)and Isospora canaria sp. n. in the canary (Serinus canarius Linnaeus).

Exogenous stages of Isospora serini (Aragao) and Isospora canaria sp. n. from the canary (Serinus canarius Linnaeus) are described. Oocytes of I. serini are spheroid and average 19.2 times 20.1 mum, while those of I. canaria are larger, more ellipsoid, and average 21.8 times 24.6 mum. No oocyst residuum is present and the oocyst walls of both species are colorless, transparent, and single layered. Sporocysts average 9.4 times 15.2 mum for I. serini and 11.5 times 18.1 mum for I. canaria. The I. canaria sporocyst has a substiedal body, but none was found in I. serini sporocysts. Both species have a spherical sporocyst residuum; this was obscured in the I. serini sporocyst by scattered granules. Living sporozoites of I. canaria average 3.6 times 16.9 mum and have 1 to 3 refractile globules; those of I. serini have 2 globules and average 2.8 times 12.6 mum. A disseminated infection of the mononuclear phagocytes results from administration of I. serini while I. canaria oocysts give rise to a typical coccidian infection restricted to the intestinal epithelium. Asexual stages of I. serini in macrophages are indistinguishable from parasites previously called avian Toxoplasma, Atoxoplasma, and Lankesterella.

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Re-descriptions of Isospora ameivae Carini, 1932 in the teiid lizard Ameiva ameiva and isospora hemidactyli Carini, 1936 in the gecko Hemidactylus mabouia, with particular reference to their endogenous stages.

Redescriptions are given of the mature oocysts of Isospora ameivae Carini, 1932, from the teiid lizard Ameiva ameiva, and Isospora hemidactyli Carini,1936 from the gecko Hemidactylus mabouia, in north Brazil. The endogenous stages of the two parasites in the small intestine are described. Those of I. ameivae are intracytoplasmic, whereas those of I. hemidactyli are intranuclear.

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Isospora mcquistioni and Isospora bioccai (Apicomplexa, Eimeriidae): two new coccidian parasites from Carduelis sinica (Passeriformes, Fringillidae).

The following two species are described from Carduelis sinica (Greenfinch) from Italy. The oocysts of Isospora mcquistioni n. sp. were 26.0 x 22.6 (24.0-28.5 x 20.0-24.2) microns and ovoid with a smooth bilayered wall. Neither micropyle nor oocyst residuum were observed. One polar granule was found. Sporocysts were oval, 18.1 x 11.4 (16.0-19.8 x 11.0-12.0) microns, and with a symmetrical Stieda complex. The residuum was compact and spherical. Isospora bioccai n. sp. oocysts were spherical to subspherical and 24.0 x 23.6 (22.0-26.0 x 21.0-25.8) microns. The oocyst wall was smooth and bilayered. A micropyle and oocyst residuum were absent; 4 to 10 elongate polar granules were present. Sporocysts were 19.5 x 11.6 (18.0-20.0 x 10.0-12.4) microns, ellipsoidal, and with a symmetrical Stieda complex. The sporocyst residuum was diffuse and composed of a few granules.

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A new species of Isospora Schneider, 1881 (Apicomplexa: Eimeriidae) from the Bali (Rothschild's) mynah Leucopsar rothschildi (Passeriformes: Sturnidae), and comments concerning the genera Atoxoplasma Garnham, 1950 and Isospora.

A new species of isosporan (Apicomplexa: Eimeriidae) is reported from the Bali mynah Leucopsar rothschildi (Passeriformes: Sturnidae). Oöcysts of Isospora rothschildi n. sp. are spherical to subspherical, 22.3 x 21.6 (20-26 x 19-23) microm, with a shape index (length/width) of 1.03 (1.00-1.15). A micropyle and oöcyst residuum are absent, but one to many polar granules are present. Sporocysts are ovoidal, 15.9 x 10.6 (15-18 x 9.5-11) microm, with Stieda and substieda bodies and a shape index of 1.50 (1.39-1.65). Each sporocyst contains an ellipsoidal sporocyst residuum, 8.0 x 5.8 (6-11 x 5-8), and each sporozoite contains two refractile bodies. No correlation was found between the presence of coccidian oöcysts in the faeces of some of the birds and Atoxoplasma in blood smears.

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Isospora araponga sp. n. (Apicomplexa: Eimeriidae), a new species of Isospora Schneider from a bare-throated bellbird, Procnias nudicollis (Vieillot, 1817) (Passeriformes: Cotingidae) from Brazil.

Faecal samples from a couple of bare-throated bellbirds Procnias nudicollis imported from Brazil to Barcelona Zoo contained oocysts of Isospora araponga n. sp. Sporulated oocysts were subspherical to broadly ellipsoidal, 19.5 (17-22) x 15.5 (14-16.5) microm, shape index (length:width ratio) 1.26 (1.13-1.38) with smooth and colourless bilayered wall, about 1 microm thick, and with varying number (1-3) of polar granule, but without a micropyle or residuum. The sporocysts were ellipsoidal, slightly asymmetric, 12.5 (12-13) x 8.5 (7.5-9) microm with barely visible Stieda body and indistinguishable substieda body. Sporozoites were elongated, possessing smooth surface and two distinct refractile bodies.

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Activation and excystation of Isospora felis and Isospora rivolta sporozoites.

Studies on the activation and excystation of I. felis and I. rivolta sporozoites revealed that, although some differences were apparent between the two, both were capable of activation under a wide range of conditions. Activation of both species took place in trypsin and bile between 21 and 43 C (the range tested) occurring rapidly at 39 C. Although the presence of bile appeared to be essential for this process that of trypsin did not. Neither the concentration of bile (above 5%), the type of bile, nor the hydrogen ion concentration had a marked effect on the level of activation. Pretreatment of oocysts in aqueous cysteine hydrochloride under CO2 or with sodium hypochlorite before exposure to trypsin and bile was not an essential prerequisite for the activation of I. felis and I. rivolta. However, higher levels of activation were attained when pretreatment was used. But for I. rivolta, the level of activation appeared to be less dependent on pretreatment for oocysts stored in sulfuric acid than for those stored in potassium dichromate. The process of activation and excystation of both species was essentially similar to that described for other species of disporocystic coccidia with heteroxenous life cycles. Sporozoites escaped following the collapse of the sporocyst wall and completed excystation through indentations and fractures at one or both ends of the oocyst.

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Phylogenetic analysis of coccidia based on 18S rDNA sequence comparison indicates that Isospora is most closely related to Toxoplasma and Neospora.

The phylogenetic relationships and taxonomic affinities of coccidia with isosporan-type oocysts have been unclear as overlapping characters, recently discovered life cycle features, and even recently discovered taxa, continue to be incorporated into biological classifications of the group. We determined the full or partial 18S ribosomal RNA gene sequences of three mammalian Isospora spp., Isospora felis, Isospora ohioensis and Isospora suis, and a Sarcocystis sp. of a rattlesnake, and used these sequences for a phylogenetic analysis of the genus Isospora and the cyst-forming coccidia. Various alveolate 18S rDNA sequences were aligned and analyzed using maximum parsimony to obtain a phylogenetic hypothesis for the group. The three Isospora spp. were found to be most closely related to Toxoplasma gondii and Neospora caninum. This clade in turn formed the sister group to the Sarcocystis spp. included in the analysis. The results confirm that the genus Isospora does not belong to the family Eimeriidae, but should be classified together with the cyst-forming coccidia in the family Sarcocystidae. Furthermore, there appear to be two lineages within the Sarcocystidae. One lineage comprises Isospora and the Toxoplasma/Neospora clade which share the characters of having a proliferative phase of development preceding gamogony in the definitive host and an exogenous phase of sporogony. The other lineage comprises the Sarcocystis spp. which have no proliferative phase in the definitive host and an endogenous phase of sporogony.

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Taxonomic position of the human intestinal protozoan parasite Isospora belli as based on ribosomal RNA sequences.

The taxonomic positions of Isospora belli and other members of the genus Isospora are controversial. We determined the small-subunit ribosomal RNA of I. belli and used this sequence in combination with other coccidian RNA sequences for analysis of the taxonomic position of I. belli. The phylogenetic trees we obtained provide molecular evidence for three clades within a monophyletic group that represents the suborder Eimeriina. The clade containing I. belli consists of tissue-cyst-forming coccidia (Toxoplasma and Neospora) and members of the genus Isospora (I. ohioensis, I. suis, I. belli). The second clade, representing a sister clade of that containing the Isospora species, contains members of the genus Sarcocystis. The third one consists of members of the family Eimeriidae, including Eimeria and Cyclospora species. This shows that although I. belli as well as other members of the genus Isospora belong to the suborder Eimeriina, the family to which they belong is not Eimeriidae but rather Sarcocystidae. We suggest that the genus Isospora should be removed from the family Eimeriidae and placed into the family Sarcocystidae within the suborder Eimeriina.

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Detection of Isospora belli by polymerase chain reaction using primers based on small-subunit ribosomal RNA sequences.

The aim of the present study was to use small-subunit (SSU)-rRNA sequences of Isospora belli to design specific primer pairs and a hybridization probe for the detection of Isospora belli in human samples by PCR and Southern blot hybridization. PCR amplification with the primer pairs produced correct DNA fragments with target DNA from samples of Isospora belli-infected patients and from cloned SSU-rRNA of Isospora belli. The nature of the PCR products was confirmed by Southern blot hybridization. No amplification was seen with template DNA extracted from other parasites. Although Isospora belli infections can be easily diagnosed using light microscopy, molecular-based techniques may prove useful as an additional diagnostic tool.

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Five new species of Isospora from Hawaiian birds.

The following species are described from Hawaiian birds: Isospora brayi sp. n., with oocysts 27 X 26 micron and sporocysts 19 X 12 micron, from the Japanese white-eye, Zosterops japonicus Temminck & Schlegel; Isospora cardinalis sp. n., with oocysts 24 X 23 micron, and sporocysts 16 X 10 micron, from the cardinal, Cardinalis cardinalis (Linnaeus); Isospora ivensae sp. n., with oocysts 26 X 25 micron, and sporocysts 18 X 12 micron, from the spotted or white-throated munia, Lonchura punctulata (Linnaeus); Isospora loxopis sp. n., with oocysts 26 X 23 micron, and sporocysts 16 X 13 micron, from the amakihi or honeycreeper, Loxops virens (Gmelin); and Isospora phaeornis sp. n., with oocysts 27 X 19 micron, and sporocysts 16 X 11 micron, from the omao or Hawaiian thrush, Phaeornis obscurus (Gmelin). All the host birds belong to the order Passerorida.

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