This study aimed to evaluate and document the excystation process of Cryptosporidium muris oocysts in various incubation media, and to monitor the behaviour of excysting and freshly excysted sporozoites. A test of oocyst viability, using fluorescent double staining with fluorescein diacetate and propidium iodide, was performed prior to each experimental assay. Light microscope observations confirmed that relatively often only three sporozoites were released; the fourth one either left the oocyst later together with a residual body or remained trapped within the oocyst wall. These results suggest that successful oocyst excystation is not limited by the viability of all four sporozoites. Darkening of oocysts to opaque and their specific movement (the so-called "oocyst dancing") preceded the final excystation and liberation of sporozoites, while the dormant oocysts appeared refractive. The process of excystation in C. muris is not gradual as generally described in cryptosporidia but very rapid in an eruptive manner. Experiments were performed using oocysts stored at 4 °C for various time periods, as well as oocysts freshly shed from host rodents (Mastomys coucha) of different ages. The most suitable medium supporting high excystation rate (76 %) and prolonged motility of sporozoites was RPMI 1640, enriched with 5 % bovine serum albumin (BSA). Our results emphasize that to reliably evaluate the success of in vitro excystation of cryptosporidia, not only the number of released sporozoites in a set time period should be taken into consideration but also their subsequent activity (motility), as it is expected to be essential for the invasion of host cells.

• Klíčová slova
• Cryptosporidium muris, Excystation rate, Motility, Oocyst, Sporozoite, Viability test,
• MeSH
• Cryptosporidium účinky léků   fyziologie   MeSH
• krysa rodu Rattus MeSH
• kultivační média farmakologie   MeSH
• mikrobiální viabilita účinky léků   MeSH
• oocysty fyziologie   MeSH
• propidium MeSH
• sporozoiti účinky léků   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kultivační média MeSH
• propidium MeSH

Urosporid eugregarines (Apicomplexa: Urosporidae) are unicellular eukaryotic parasites inhabiting the coelom or the intestine of marine invertebrates such as annelids, molluscs, nemerteans, and echinoderms. Despite the availability of published morphological and phylogenetical analyses of coelomic gregarines, their long-term survival in the host body cavity and dispersal routes into the marine environment remain unclear. Here, we focus on Urospora gametocysts and oocysts with sporozoites, which were found viable inside the so-called brown bodies floating in the body cavity of the polychaete Travisia forbesii. Brown bodies form as a result of host defence where coelomocytes encapsulate dead host cells and foreign objects including potential pathogens. We hypothesise the long-term persistence of Urospora eugregarines in brown bodies through evasion of the host immunity and outline possible pathways for their egress into the marine environment, applicable as dispersal routes for other parasites as well. Unique features revealed by detailed ultrastructural analysis of detected eugregarine stages include asynchronous sporogony, a massive sporozoite secretion apparatus, as well as the presence of free (possibly autoinfective) sporozoites within the gametocyst. The assignment to the genus Urospora and the complete identity with U. ovalis and U. travisiae were confirmed by analysing 18S rDNA sequences obtained from isolated gametocysts. The 18S rDNA phylogeny confirmed the affiliation of Urosporidae to Lecudinoidea and the grouping of all Urospora sequences with Difficilina from nemerteans and environmental sequences from the Artic region. We also enriched the Apicomplexa set by partial 28S rDNA sequences of two Urospora species enabling more complex phylogenetic analyses prospectively.

• Klíčová slova
• 18S and 28S rDNAs, Gametocyst, Invertebrate immunity, Oocyst, Sporozoite, Urosporidae,
• MeSH
• Apicomplexa * MeSH
• fylogeneze MeSH
• oocysty ultrastruktura   MeSH
• Polychaeta * parazitologie   MeSH
• ribozomální DNA genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ribozomální DNA MeSH

Cryptosporidium parvum contains a unique fusion protein pyruvate:NADP+ oxidoreductase (CpPNO) that is composed of two distinct, conserved domains, an N-terminal pyruvate:ferredoxin oxidoreductase (PFO) and a C-terminal cytochrome P450 reductase (CPR). Unlike a similar fusion protein that localizes to the mitochondrion of the photosynthetic protist Euglena gracilis, CpPNO lacks an N-terminal mitochondrial targeting sequence. Using two distinct polyclonal antibodies raised against CpPFO and one polyclonal antibody against CpCPR, Western blot analysis has shown that sporozoites of C. parvum express the entire CpPNO fusion protein. Furthermore, confocal immunofluorescence and transmission electron microscopy confirm that CpPNO is localized within the cytosol rather than the relict mitochondrion of C. parvum. The distribution of this protein is not, however, strictly confined to the cytosol. CpPNO also appears to localize posteriorly within the crystalloid body.

• MeSH
• Cryptosporidium parvum cytologie   enzymologie   genetika   růst a vývoj   MeSH
• cytosol enzymologie   MeSH
• Euglena gracilis cytologie   enzymologie   MeSH
• fluorescenční mikroskopie MeSH
• ketonoxidoreduktasy analýza   genetika   imunologie   MeSH
• konfokální mikroskopie MeSH
• NADPH-cytochrom c-reduktasa analýza   genetika   imunologie   MeSH
• organely enzymologie   MeSH
• protozoální proteiny analýza   MeSH
• pyruvátsynthasa analýza   genetika   imunologie   MeSH
• sporozoiti cytologie   enzymologie   genetika   MeSH
• transmisní elektronová mikroskopie MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• ketonoxidoreduktasy MeSH
• NADPH-cytochrom c-reduktasa MeSH
• protozoální proteiny MeSH
• pyruvate dehydrogenase (NADP+) MeSH Prohlížeč
• pyruvátsynthasa MeSH

Naive and immune specific-pathogen-free rabbits were inoculated in the duodenum with sporocysts of Eimeria coecicola or Eimeria intestinalis. Samples were taken from the following tissues: duodenum (site of penetration of sporozoites), ileum (specific target site of the endogenous development of E. intestinalis), vermiform appendix (target site of E. coecicola) and two extraintestinal sites, mesenteric lymph nodes (MLNs), and spleen. The presence of sporozoites was checked by immunohistochemistry. In rabbits primary-infected with E. coecicola, large numbers of sporozoites were detected in the duodenum, extraintestinal sites, and vermiform appendix. The abundance of sporozoites in the spleen, MLN, and appendix was significantly reduced in the immune rabbits, and the migration seemed impeded. In the rabbits infected with E. intestinalis, sporozoites were absent in the spleen and MLN, indicating that the route of migration is different from that of E. coecicola. The number of sporozoites in the crypts of the ileum was markedly reduced in the immune animals.

• MeSH
• antigeny protozoální metabolismus   MeSH
• Eimeria růst a vývoj   imunologie   patogenita   MeSH
• interakce hostitele a parazita MeSH
• kokcidióza imunologie   parazitologie   MeSH
• králíci MeSH
• lymfatické uzliny parazitologie   MeSH
• organismy bez specifických patogenů MeSH
• parazitární nemoci střev * MeSH
• Peyerovy pláty metabolismus   parazitologie   MeSH
• protilátky protozoální imunologie   MeSH
• slezina parazitologie   MeSH
• sporozoiti růst a vývoj   imunologie   patologie   MeSH
• tenké střevo metabolismus   parazitologie   patologie   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny protozoální MeSH
• protilátky protozoální MeSH

In all the studies performed on the invasion of the rabbit by sporozoites of 4 Eimeria species it was shown that the sporozoites first penetrate the duodenal epithelium; and then very quickly appear within the IELs of the duodenal epithelium. A few hours later they are found in IELs of their specific site of multiplication. The relationship between an "extraintestinal" route for sporozoites and the IELs homing phenomenon is therefore an exciting hypothesis.

• MeSH
• buněčný cyklus MeSH
• duodenum mikrobiologie   MeSH
• Eimeria cytologie   MeSH
• králíci MeSH
• lymfocyty mikrobiologie   MeSH
• střevní sliznice mikrobiologie   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The invasive phase of Eimeria coecicola was studied during the first 80 h postinoculation (p.i.). Using a method that synchronized the life cycle, sporozoites were observed in the duodenum and the jejunum until 32 h p.i. They were seen first in the villous epithelial cells or in host cells resembling intraepithelial lymphocytes (IEL). Later they were observed in IEL in the lamina propria. After 48 h p.i., no coccidian stage was identifiable in the mucosa of the small intestine but sporozoites appeared in the lymphoid cells of lymphatic follicles of the gut-associated lymphoid tissue (vermiform appendix, sacculus rotundus, and Peyer's patches). The first merogony was observed 64 h p.i. in these lymphoid cells and in membranous epithelial cells (M-cells) but was never seen in the epithelium itself. Morphologically there were two types of meronts, depending on the host cell type, but in both cases the merozoites contained a refractile body and resembled sporozoites. The first meronts of the second generation were observed 80 h p.i. in the villous epithelial cells of the domes of the follicles of the gut-associated lymphoid tissue, where the further development of this Eimeria takes place. This pattern of invasion strongly suggests that sporozoites take an exclusively extraintestinal route to reach the target cells. Moreover, to our knowledge this is the first description of an eimerian merogony that does not take place in epithelial cells.

• MeSH
• apendix parazitologie   MeSH
• biologická proměna MeSH
• Eimeria fyziologie   ultrastruktura   MeSH
• kokcidióza parazitologie   MeSH
• lymfoidní tkáň parazitologie   ultrastruktura   MeSH
• Peyerovy pláty parazitologie   MeSH
• pohyb buněk MeSH
• střeva parazitologie   ultrastruktura   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Archigregarines, an early branching lineage within Apicomplexa, are a poorly-known group of invertebrate parasites. By their phylogenetic position, archigregarines are an important lineage to understand the functional transition that occurred between free-living flagellated predators to obligatory parasites in Apicomplexa. In this study, we provide new ultrastructural data and phylogenies based on SSU rDNA sequences using the type species of archigregarines, the Selenidiidae Selenidium pendulaGiard, 1884. We describe for the first time the syzygy and early gamogony at the ultrastructural level, revealing a characteristic nuclear multiplication with centrocones, cryptomitosis, filamentous network of chromatin, a cyst wall secretion and a 9+0 flagellar axoneme of the male gamete. S. pendula belongs to a monophyletic lineage that includes several other related species, all infecting Sedentaria Polychaeta (Spionidae, Sabellaridae, Sabellidae and Cirratulidae). All of these Selenidium species exhibit similar biological characters: a cell cortex with the plasma membrane - inner membrane complex - subpellicular microtubule sets, an apical complex with the conoid, numerous rhoptries and micronemes, a myzocytosis with large food vacuoles, a nuclear multiplication during syzygy and young gamonts. Two other distantly related Selenidium-like lineages infect Terebellidae and Sipunculida, underlying the ability of archigregarines to parasite a wide range of marine hosts.

• Klíčová slova
• Apicomplexa, Archigregarines, Selenidium pendula, phylogeny, sporozoite., ultrastructure,
• MeSH
• Apicomplexa klasifikace   genetika   růst a vývoj   ultrastruktura   MeSH
• fylogeneze * MeSH
• mikroskopie elektronová rastrovací MeSH
• protozoální DNA genetika   MeSH
• ribozomální DNA genetika   MeSH
• transmisní elektronová mikroskopie MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• protozoální DNA MeSH
• ribozomální DNA MeSH

One of three (33%) captive specimens of Oustalet's chameleon, Furcifer oustaleti (Mocquard) originally from Madagascar and housed at the Oklahoma City Zoological Park Herpetarium, Oklahoma County, Oklahoma, USA, was found to be passing an undescribed species of Choleoeimeria in its faeces. Oocysts of Choleoeimeria fischeri sp. n. were cylindroidal, 30.3 x 16.8 (28-34 x 15-18) microm, with a smooth, bilayered wall and a length/width ratio (L/W) of 1.8. A micropyle and oocyst residuum was absent but a fragmented polar granule was often present. Sporocysts were ovoidal, 9.6 x 8.0 (9-10 x 7-9) jm, with an L/W of 1.2. Stieda, sub-Stieda, and para-Stieda bodies were absent. The sporocyst residuum consists of large globules dispersed between sporozoites. Sporozoites were elongate, 8.6 x 2.9 (8-10 x 2-3) microm, with an elongate posterior refractile body. The new species represents the second coccidian described from this lizard.

• MeSH
• Eimeriidae klasifikace   růst a vývoj   izolace a purifikace   MeSH
• feces parazitologie   MeSH
• ještěři parazitologie   MeSH
• oocysty klasifikace   růst a vývoj   MeSH
• sporozoiti klasifikace   růst a vývoj   MeSH
• zvířata v ZOO klasifikace   parazitologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Oklahoma MeSH

The fine structure of sporocysts and sporozoites of parent strains and precocious lines of rabbit coccidia Eimeria intestinalis, E. magna and E. media was studied. The parent strains and precocious lines differ only in the shape and size of refractile bodies (RB). In the sporocysts of precocious lines of E. magna and E. media, one extremely large RB was seen, either inside one of the sporozoites, or free in the sporocyst. In the oocysts of the precocious strain of E. intestinalis, two sporocysts resembled those of the precocious lines of E. magna and E. media, whereas the other two sporocysts did not harbour any RB. Sporozoites of all the precocious lines contained no, or very small, RB after in vitro excystation.

• MeSH
• Eimeria růst a vývoj   ultrastruktura   MeSH
• elektronová mikroskopie metody   MeSH
• kokcidióza parazitologie   MeSH
• králíci MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

In the present study, we have investigated the role of antimalarial drug halofantrine (HF) in inducing the sterile protection against challenges with sporozoites of the live infectious Plasmodium yoelii (Killick-Kendrick, 1967) in Swiss mice malaria model. We observed that during the first to third sequential sporozoite inoculation cycles, blood-stage patency remains the same in the control and chemoprophylaxis under HF drug cover (CPS-HF) groups. However, a delayed blood-stage infection was observed during the fourth and fifth sporozoite challenges and complete sterile protection was produced following the sixth sporozoite challenge in CPS-HF mice. We also noticed a steady decline in liver stage parasite load after 3th to 6th sporozoite challenge cycle in CPS-HF mice. CPS-HF immunisation results in a significant up-regulation of pro-inflammatory cytokines (IFN-γ, TNF-α, IL-12 and iNOS) and down-regulation of anti-inflammatory cytokines (IL-10 and TGF-β) mRNA expression in hepatic mononuclear cells (HMNC) and spleen cells in the immunised CPS-HF mice (after 6th sporozoite challenge) compared to control. Overall, our study suggests that the repetitive sporozoite inoculation under HF drug treatment develops a strong immune response that confers protection against subsequent challenges with sporozoites of P. yoelii.

• Klíčová slova
• CPS-immunisation, Real Time PCR, cytokines, mRNA expression, sporozoites,
• MeSH
• chemoprofylaxe MeSH
• fenantreny MeSH
• imunizace MeSH
• léčivé přípravky * MeSH
• malárie * prevence a kontrola   MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• Plasmodium yoelii * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fenantreny MeSH
• halofantrine MeSH Prohlížeč
• léčivé přípravky * MeSH