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Evolutionary relationships among cyst-forming coccidia Sarcocystis spp. (Alveolata: Apicomplexa: Coccidea) in endemic African tree vipers and perspective for evolution of heteroxenous life cycle.

Cyst-forming coccidia of the genus Sarcocystis (Alveolata: Apicomplexa: Coccidea) parasitize vertebrates worldwide. Data from the small subunit rRNA genes (SSU) and the D2 domain of the large subunit rRNA genes were used to reconstruct phylogeny for all species in the Sarcocystidae for which sequences are currently available. We have focused on the evolutionary history of species that circulate between snakes as definitive hosts and rodents as intermediate hosts. Trees were reconstructed using maximum parsimony, minimum evolution, maximum likelihood and the bayesian phylogenetics. Our reconstructions support monophyly of Sarcocystidae but fail to robustly resolve the relationship within clades. Using a concatenated dataset of available rDNAs, the "isosporoid" coccidia Neospora, Toxoplasma, Besnoitia, Isospora and Hyaloklossia form a sister group to the monophyletic Sarcocystis. Moreover, we show that Sarcocystis from arboreal vipers of the genus Atheris, which are endemic to the mountain rain forests of the Equatorial Africa, are monophyletic, with sister species parasitizing the desert viper Pseudocerastes persicus from the Near East. We report the co-evolution of Sarcocystis spp. with their final snake hosts. The geological history of the African continent, mountain ranges, forests and general SSU rDNA rates were used to construct a linearized tree. Possible origin of the heteroxenous life cycle of Sarcocystis is discussed.

Animals↗

Fine structure of zoosporulation in Perkinsus atlanticus (Apicomplexa: Perkinsea).

Light and transmission electron microscopy were used to study different stages of Perkinsus atlanticus (Apicomplexa) during induced zoosporulation, with fluid thioglycollate medium and seawater. Cytokinesis and nucleokinesis of different developmental stages were studied every 12 h during the incubation period of 72 h, at which time the zoospores became free. Uninucleated and flagellated zoospores present the apical complex formed by conoid, polar ring, micronemes, rhoptries and subpellicular microtubules observed at different sections. Ultrastructural details were compared with the other two species of the genus Perkinsus.

Animals↗

Two new species of Caryospora Léger, 1904 (Apicomplexa, Eimeriidae) from accipitrid raptors.

Two new species of Coccidia (Apicomplexa: Eimeriidae) are described from European accipitrid raptors (Falconiformes: Accipitridae). Oöcysts of Carvospora aquilae n. sp. found in faeces of the gold eagle Aquila chrysaetos are subspherical to broad ellipsoidal and measure 43 (40-49) x 37.5 (34-39) microm. Polar granule, oöcyst residuum and micropyle are absent. Each oöcyst contains one spherical to subspherical slightly polygonal sporocyst measuring 23.8 (23-25) x 23.3 (22-25) microm. Stieda and substieda bodies are absent. The sporocyst residuum is composed of numerous small granules less than 0.5 microm in diameter dispersed randomly among the sporozoites. Sporulated oöcysts of Carvospora circi n. sp. from faeces of the marsh harrier Circus aeruginosus are widely oval, measuring 24.5 (23-25) x 21.8 (21-24) microm. A polar granule, oöcyst residuum and micropyle are absent. Each oöcyst contains one spherical to subspherical sporocyst measuring 16.2 (15-17) x 15.6 (15-17) microm. A compact granular, spherical to subspherical sporocyst residuum, 10.4 (10-11) x 8.5 (7-9), was present in 76% of measured sporocysts. In 24% of sporocysts the granules of sporocyst residuum were scattered among the sporozoites.

Animals↗

Aggregata sagittata n. sp. (Apicomplexa: Aggregatidae), a coccidian parasite from the European flying squid Todarodes sagittatus (Mollusca: Cephalopoda).

A new species of the coccidian genus Aggregata Frenzel, 1885 (Apicomplexa, Aggregatidae) is described from the digestive tract of Todarodes sagittatus (Lamarck), an ommastrephid squid that lives in temperate waters in the northeastern Atlantic Ocean. Using light microscopy. oöcysts were 225-333 microm in length and 220-323 microm in width. Each oöcyst contained 600-1,000 sporocysts. Sporocysts were 16-18 microm in length and 15-17 microm in width, with a smooth surface and thick wall. Each sporocyst contained 4-8 sporozoites each with a length of 11-13 microm. This paper presents the description of a new Agregata species in a nerito-oceanic flying squid. Moreover. it extends the distribution of Aggregata to oceanic cephalopod host species. A. sagittata represents the third named species in the genus Aggregata in European waters and the seventh worldwide.

Animals↗

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.

Animals↗

Intestinal infections by Eimeria (s. l.) vanasi n. sp. (Eimeriidae, Apicomplexa, Protozoa) in cichlid fish.

Eimeria (s. l.) vanasi n. sp. is described from the intestine of a variety of cichlids in Israel and South Africa. Merogony and gamogony stages are both intra- and epiepithelial, Sporogony is exogenous with young zygotes being released from the fish in the faeces. Fully developed sporocysts are ellipsoid and apparently lacking both a Stieda body (characteristic of Eimeria s. st. and Epieimeria species) and a suture line (characteristic of Goussia species). In view of the present controversial generic status of piscine coccidia, the species is tentatively designated as Eimeria (s. l.) vanasi n. sp. (Eimeriidae, Apicomplexa, Protozoa).

Animals↗

Proteomics analysis and protein expression during sporozoite excystation of Cryptosporidium parvum (Coccidia, Apicomplexa).

Cryptosporidiosis, caused by coccidian parasites of the genus Cryptosporidium, is a major cause of human gastrointestinal infections and poses a significant health risk especially to immunocompromised patients. Despite intensive efforts for more than 20 years, there is currently no effective drug treatment against these protozoa. This study examined the zoonotic species Cryptosporidium parvum at two important stages of its life cycle: the non-excysted (transmissive) and excysted (infective) forms. To increase our understanding of the molecular basis of sporozoite excystation, LC-MS/MS coupling with a stable isotope N-terminal labeling strategy using iTRAQ reagents was used on soluble fractions of both non-excysted and excysted sporozoites, i.e. sporozoites both inside and outside oocysts were examined. Sporozoites are the infective stage that penetrates small intestinal enterocytes. Also to increase our knowledge of the C. parvum proteome, shotgun sequencing was performed on insoluble fractions from both non-excysted and excysted sporozoites. In total 303 C. parvum proteins were identified, 56 of which, hitherto described as being only hypothetical proteins, are expressed in both excysted and non-excysted sporozoites. Importantly we demonstrated that the expression of 26 proteins increases significantly during excystation. These excystation-induced proteins included ribosomal proteins, metabolic enzymes, and heat shock proteins. Interestingly three Apicomplexa-specific proteins and five Cryptosporidium-specific proteins augmented in excysted invasive sporozoites. These eight proteins represent promising targets for developing vaccines or chemotherapies that could block parasite entry into host cells.

Animals↗

Experimental transmission of Caryospora simplex (Apicomplexa: Eimeriidae) to Palestine vipers, Vipera xanthina palestinae (Serpentes: Viperidae).

Four littermate, laboratory-reared Palestine vipers, Vipera xanthina palestinae (#149, #150, #151, #152) (Serpentes: Viperidae) were used to determine modes by which Caryospora simplex (Apicomplexa: Eimeriidae) could be transmitted to snakes. Viper #149 was inoculated orally by stomach tube with 5.0 X 10(4) sporulated oocysts of C. simplex obtained from the feces of an Ottoman viper, V. x. xanthina and began passing unsporulated oocysts of C. simplex 121 days post-inoculation (DPI). Viper #150 was fed five mice that had been inoculated orally greater than or equal to 25 days previously with 5.0 X 10(4) sporulated oocysts of C. simplex and it began passing unsporulated oocysts of C. simplex 33 days after being fed the first two of five mice. Viper #151 was inoculated orally with sporulated oocysts of C. simplex obtained from viper #150 and began passing oocysts 52 DPI. Viper #152 served as an uninoculated control and did not pass oocysts of any species of coccidian. This study demonstrates that snake/snake and mouse/snake transmission of C. simplex readily occurs.

Animals↗

Two new species of coccidia (Apicomplexa: Eimeriidae) from Madagascar Gekkonids.

Two new species of coccidia (Apicomplexa: Eimeriidae) are described from the Madagascar giant day gecko, Phelsuma madagascariensis grandis, and the Golddust day gecko, P. laticauda. Both species of coccidia were found to infect the anterior one-half of the small intestine. Oocysts of Eimeria brygooi n. sp. are spherical or subspherical, 23.0 X 21.3 (18.8-25.2 X 16.4-23.2)micron; shape index (L/W) 1.1 (1.0-1.2). A micropyle, oocyst residuum, and polar granule are absent. Sporocysts are ovoid, 9.2 X 7.9 (8.0-10.0 X 7.2-8.8) micron; shape index 1.2 (1.0-1.3), with a Goussia-type suture; Stieda and substieda bodies are absent. A sporocyst residuum is present, 4.2 X 3.0 (3.2-6.4 X 2.4-4.0) micron. Sporozoites are elongate, with anterior and posterior refractile bodies. This coccidian was found to infect five of six (83%) P. m. grandis and one of five (20%) P. laticauda examined. Oocysts of Isospora gekkonis n. sp. are spherical or subspherical, 24.2 X 22.0 (21.6-26.4 X 20.0-23.6) micron; shape index 1.1 (1.0-1.2). A micropyle and oocyst residuum are absent; polar granule present. Sporocysts are ovoid, 12.2 X 9.4 (11.2-12.8 X 8.4-10.0) micron, with Stieda and substieda bodies; shape index 1.3 (1.2-1.4). A sporocyst residuum is present, either compact, 5.1 X 4.2 (4.0-7.2 X 3.2-5.6) micron or diffuse. Sporozoites are elongate, with anterior and posterior refractile bodies. Isospora gekkonis was found in two of six (33%) P. m. grandis and one of five (20%) P. laticauda. In addition, oocysts of Cryptosporidium sp. were found in the cloacas of two of six (33%) necropsied P. m. grandis.

Animals↗

Eimeria saudiensis n. sp. (Apicomplexa: Eimeriidae) from the Arabian oryx (Oryx leucoryx) in Saudi Arabia.

Oocysts of Eimeria saudiensis n. sp. (Apicomplexa: Eimeriidae) are described from the feces of the Arabian oryx, Oryx leucoryx, from the Riyadh Zoo, Saudi Arabia. The oocysts were ellipsoidal or slightly ovoid, 31.2 x 24.5 (24.3-36.5 x 20.0-27.6) micron with a bilayered wall about 1.7 micron thick. The micropyle was covered by a dome-shaped cap. The oocyst residuum was absent, but tiny polar granules were present. The sporocysts were elongate ovoid, 14.3 x 7.2 (11.5-18.5 x 6.0-9.0) micron, had a Stieda body, but lacked a substiedal body. The sporocyst residuum was present, composed of numerous small granules. The sporozoites were elongate club-shaped, and contained two prominent refractile bodies.

Animals↗

Isospora peromysci Davis, 1967 (Apicomplexa: Eimeriidae) in Peromyscus leucopus and P. maniculatus (Rodentia: Cricetidae) from Texas.

Oocysts of Isospora peromysci (Davis, 1967) (Apicomplexa: Eimeriidae) were recovered from the feces of 1/30 (3.3%) white-footed mice, Peromyscus leucopus, in Johnson County, Texas. This report represents a new host and geographic record for the parasite. The coccidium was also found in 1/20 (5.0%) deer mice, P. maniculatus, from the same locale. Morphological data are provided on the sporulated oocyst of I. peromysci and comparisons are made with previously published information on the species from other geographic localities.

Animals↗

Early development of Eimeria papillata (Apicomplexa: Eimeriidae) in the mouse.

Early development of Eimeria papillata (Apicomplexa) in the mouse was evaluated using Nomarski interference-contrast and brightfield microscopy. Sporozoite-shaped meronts, which were motile and contained a large posterior refractile body and a smaller anterior refractile body, were observed entering and leaving host cells in the jejunum of an experimentally infected mouse at 26 h post inoculation (HPI). However, early developmental stages were not observed in tissue of the duodenum, ileum, cecum and colon. The mean length and width of these meronts (n = 20) were 12.0 microns and 3.7 microns, respectively. Spherical or subspherical meronts containing crescent-shaped merozoites were observed at 36 HPI.

Animals↗

Lymphokine-induced inhibition of growth of Eimeria bovis and Eimeria papillata (Apicomplexa) in cultured bovine monocytes.

Sporozoites of Eimeria bovis penetrated and developed normally to first-generation meronts in bovine monocytes (BM) and Madin-Darby bovine kidney (MDBK) cells that had been pretreated with culture medium (CM) or supernatant (NS) from nonstimulated bovine T cells. At 240 h after sporozoite inoculation (ASI), the mean percent development (meronts/[sporozoites + meronts]) in CM- and NS-pretreated BM was 52 and 28%, respectively; values for MDBK cells were 36 and 35%, respectively. Pretreatment of BM and MDBK cells with supernatant (ConAS) from concanavalin A-stimulated bovine T cells had no effect on the ability of sporozoites to penetrate cells; however, at 240 h ASI, only 1% of the sporozoites in ConAS-pretreated BM cultures had developed to meronts. In contrast, ConAS had no adverse effect on the ability of E. bovis sporozoites to develop to first-generation meronts in MDBK cells. At 240 h ASI, E. bovis meronts in ConAS-pretreated BM were abnormal in appearance and retarded in development, whereas sporozoites appeared structurally normal by light microscopy. Pretreatment of BM with ConAS had no effect on the ability of sporozoites of Eimeria papillata (Apicomplexa) to penetrate cells. Sporozoites of E. papillata did not develop to meronts in ConAS-pretreated BM and, in contrast to E. bovis, most sporozoites were destroyed intracellularly.

Animals↗

Kinetics of murine delayed-type hypersensitivity response to Eimeria falciformis (Apicomplexa: Eimeriidae).

Mice recovering from a primary infection with an intestinal protozoan parasite, Eimeria falciformis (Apicomplexa: Eimeriidae), showed a classic delayed-type hypersensitivity (DTH) reaction to oocyst antigen challenge. This reaction was characterized by a biphasic pattern of footpad swelling. The first swelling peaked at 2 h after antigen challenge, whereas the second swelling peaked at 24 to 48 h after challenge. The DTH reaction was transferable with a T-cell-enriched spleen cell population from mice that had recovered from E. falciformis infection. Cytotoxic depletion of immune T cells with anti-L3T4 antibody and complement abrogated DTH transfer, indicating that L3T4-positive T cells were required. A T-cell-enriched spleen cell population from acutely infected mice suppressed the transfer of DTH with immune cells from recovered animals, implicating the existence of infection-induced immunoregulatory cells controlling the parasite-specific immune response during infection. Immune spleen cells also transferred resistance to infection as measured by oocyst production and death rate of recipients. Together, these results indicate that the DTH reaction, induced by infection with E. falciformis, is mediated by L3T4-positive T cells and is associated with resistance to infection.

Acute Disease↗

Occurrence of Cryptosporidium (Apicomplexa, Cryptosporidiidae) in Crotalus durissus terrificus (Serpentes, Viperidae) in Brazil.

The objective of the present study was to investigate the prevalence of Cryptosporidium (Apicomplexa, Cryptosporidiidae) in the snake Crotalus durissus terrificus (Serpentes, Viperidae). Fifty animals were evaluated for the presence of oocysts of Cryptosporidium sp. at the time of arrival and 30 and 60 days later. Intestinal washings with saline solution (1% body weight), fecal samples, and organ scrapings were collected during the study. Oocysts were concentrated by an ether-phosphate-buffered saline sedimentation technique and then separated by a density gradient centrifugation technique. Smears were made with the sediment and submitted to modified acid-fast and auramine-rhodamine staining. Cryptosporidium-positive smears were used as controls for the experimental findings. The overall prevalence of Cryptosporidium sp. oocysts was 14%. Among the positive snakes, oocysts were detected only in the intestinal washing in two specimens, only in the feces in four specimens, and in both materials at least once in one specimen. The positive snakes were predominantly from Santa Maria da Serra city State of São Paulo (57.1%). We also observed that all of the examinations that presented positive results were obtained at least 27 days after the capture of the animals.

Animals↗

Three new species of Isospora Schneider, 1881 (Apicomplexa: Eimeriidae) from the lesser seed-finch, Oryzoborus angolensis (Passeriformes: Emberizidae) from Brazil.

Three new coccidian (Apicomplexa: Eimeriidae) species are reported from the lesser seed-finch, Oryzoborus angolensis from Brazil. Sporulated oocysts of Isospora curio n. sp. are spherical to subspherical; 24.6 x 23.6 (22-26 x 22-25) microm, shape-index (SI, length/width) of 1.04 (1.00-1.15). Oocyst wall is bilayerd, approximately 1.5 microm thick, smooth and colourless. Micropyle and oocyst residuum are absent. The sporocysts are ovoid, 13.2 x 10.9 (15-17 x 10-13) microm, SI = 1.56 (1.42-1.71), with a small Stieda body and residuum composed of numerous granules scattered among the sporozoites. Sporozoites are elongated and posses a smooth surface and two distinct refractile bodies. Oocysts of Isospora braziliensis n. sp. are spherical to subspherical, 17.8 x 16.9 (16-19 x 16-18) microm, with a shape-index of 1.06 (1.00-1.12) and a smooth, single-layered wall approximately 1 microm thick. A micropyle, oocyst residuum and polar granules are absent. Sporocysts are ellipsoid and slightly asymmetric, 13.2 x 10.8 (12-14 x 9-12) microm, SI = 1.48 (1.34-1.61). Each sporocyst contains a barely visible Stieda body and a residuum composed numerous of granules. Sporozoites are elongated and each of them contains two distinct refractile bodies. Oocysts of Isospora paranaensis n. sp. are subspherical to broadly ellipsoid 24.3 x 19.8 (22-26 x 18-22) microm, SI = 1.22 (1.15-1.38) with smooth single-layered wall approximately 1.5 microm thick. A micropyle and oocyst residuum are absent, but one distinct ellipsoid polar granule (2.5-3.5 x 1.5-2.5 microm) is present. Sporocyst are ovoid, 15.7 x 10.1 (14-18 x 8-12) microm, SI = 1.46 (1.31-1.72), with distinct Stieda and sub-Stieda bodies. Each sporocyst contains a spherical sporocyst residuum, 4 microm in diameter All described isosporan species represent a possible cause of acute coccidiosis for O. angolensis in captivity.

Animals↗

Phylogenetic relationships of Hepatozoon (Apicomplexa: Adeleorina) based on molecular, morphologic, and life-cycle characters.

To evaluate higher-level affinities of Hepatozoon species within Apicomplexa, we sequenced the 18S rRNA gene from 2 parasites (Hepatozoon americanum and Hepatozoon canis) of dogs and 1 (Hepatozoon catesbianae) of bullfrogs. Sequences from other apicomplexans among the Sarcocystiidae, Eimeriidae, Theileriidae, Plasmodiidae, Cryptosporiidae, and Babesiidae, a Perkinsus species and 2 dinoflagellates were obtained from GenBank. Phylogenetic analysis indicated that Plasmodium, Cryptosporidium, and Hepatozoon form a monophyletic group distinct from representatives of other apicomplexan families. Although equivocal, our analysis indicated that Plasmodium and Cryptosporidium are sister taxa and that Hepatozoon is basal to them. To evaluate phylogenetic affinities among H. americanum, H. canis, and other species of Hepatozoon, we examined 18 morphologic and life-cycle features of 13 species currently assigned to Hepatozoon. This analysis indicates paraphyly of Hepatozoon (as currently arranged) because Hepatozoon lygosomarum was found most closely related to Hemolivia mauritanicum. These results, combined with results of previous studies, support elevating Hepatozoon to familial level (Hepatozoidae) as originally suggested by Wenyon in 1926. Both DNA sequence data and morphologic and life-cycle characters support a sister-group relationship between H. americanum and H. canis.

Animals↗

Eimeria species (Apicomplexa: Eimeriidae) from arctic ground squirrels (Spermophilus parryii) and red squirrels (Tamiasciurus hudsonicus) in Alaska and in Siberia, Russia.

Fecal samples from arctic ground squirrels (Spermophilus parryii) collected in Alaska (n = 90) and Russia (n = 46) and from red squirrels (Tamiasciurus hudsonicus) in Alaska (n = 35) were examined for the presence of Eimeria spp. (Apicomplexa: Eimeriidae). Four species were recovered from arctic ground squirrels, including Eimeria callospermophili (prevalence = 18%), Eimeria cynomysis (23.5%), Eimeria lateralis (19%), and Eimeria morainensis (77%). A single species, Eimeria tamiasciuri (91%), was recovered from red squirrels. Eimerians recovered from arctic ground squirrels represent new host records, and the single species from red squirrels is a new geographic record. Alaskan arctic ground squirrel prevalence was higher for E. callospermophili (Alaska = 22% vs. Russia = 9%), E. cynomysis (34% vs. 2%), and E. lateralis (27% vs. 4%), but not E. morainensis (78% vs. 76%).

Alaska↗