David Modrý

Redescription of Neospora caninum and its differentiation from related coccidia

Neospora caninum is a protozoan parasite of animals, which before 1984 was misidentified as Toxoplasma gondii. Infection by this parasite is a major cause of abortion in cattle and causes paralysis in dogs. Since the original description of N. caninum in 1988, considerable progress has been made in the understanding of its life cycle, biology, genetics and diagnosis. In this article, the authors redescribe the parasite, distinguish it from related coccidia, and provide accession numbers to its type specimens deposited in museums.

Genetic diversity of Cryptosporidium spp. in captive reptiles.

ABSTRACT The genetic diversity of Cryptosporidium in reptiles was analyzed by PCR-restriction fragment length polymorphism and sequence analysis of the small subunit rRNA gene. A total of 123 samples were analyzed, of which 48 snake samples, 24 lizard samples, and 3 tortoise samples were positive for Cryptosporidium. Nine different types of Cryptosporidium were found, including Cryptosporidium serpentis, Cryptosporidium desert monitor genotype, Cryptosporidium muris, Cryptosporidium parvum bovine and mouse genotypes, one C. serpentis-like parasite in a lizard, two new Cryptosporidium spp. in snakes, and one new Cryptosporidium sp. in tortoises. C. serpentis and the desert monitor genotype were the most common parasites and were found in both snakes and lizards, whereas the C. muris and C. parvum parasites detected were probably the result of ingestion of infected rodents. Sequence and biologic characterizations indicated that the desert monitor genotype was Cryptosporidium saurophilum. Two host-adapted C. serpentis genotypes were found in snakes and lizards.

Morphology, Ultrastructure and Life Cycle of Vitrella brassicaformis n. sp., n. gen., a Novel Chromerid from the Great Barrier Reef

Chromerida are photoautotrophic alveolates so far only isolated from corals in Australia. It has been shown that these secondary plastid-containing algae are closely related to apicomplexan parasites and share various morphological and molecular characters with both Apicomplexa and Dinophyta. So far, the only known representative of the phylum was Chromera velia. Here we provide a formal description of another chromerid, Vitrella brassicaformis gen. et sp. nov., complemented with a detailed study on its ultrastructure, allowing insight into its life cycle. The novel alga differs significantly from the related chromerid C. velia in life cycle, morphology as well as the plastid genome. Analysis of photosynthetic pigments on the other hand demonstrate that both chromerids lack chlorophyll c, the hallmark of phototrophic chromalveolates. Based on the relatively high divergence between C. velia and V. brassicaformis, we propose their classification into distinct families Chromeraceae and Vitrellaceae. Moreover, we predict a hidden and unexplored diversity of the chromerid algae.

Dog shedding oocysts of Neospora caninum: PCR diagnosis and molecular phylogenetic approach

Results of molecular determination of a dog isolate of Neospora caninum in the Czech Republic are presented. Colorless bisporocystic oocysts measuring 10-13 micro m x 10-11 micro m were recovered from feces and used for DNA isolation. A diagnostic PCR procedure using previously described molecular methods was performed to determine the species. The N. caninum species-specific primers based on the Nc 5 region produced a positive result, while primers specific for Hammondia heydorni rDNA internal transcribed spacer 1 (ITS1) was negative. Sequencing and phylogenetic comparison of ITS1 rDNA and the D2 domain of the large subunit rDNA (D2 LSU) determined our isolate to be N. caninum. Phylogenetic analysis of closely related genera Toxoplasma, Neospora and Hammondia based on ITS1 and D2 LSU robustly distinguished three clades: (i). Toxoplasma gondii + Hammondia hammondi, (ii). N. caninum + Neospora hughesi, and (iii). H. heydorni. Based on phylogenetic relationships we propose three acceptable suggestions to solve the problem of taxonomy of these genera.

The Phylogeny of Goussia and Choleoeimeria (Apicomplexa; Eimeriorina) and the Evolution of Excystation Structures in Coccidia

The phylogenetic relationships of Goussia janae and Choleoeimeria sp. were analyzed using the small subunit ribosomal RNA gene (SSU rDNA). This is a first attempt to study the molecular phylogeny of coccidian genera parasitizing strictly poikilotherm hosts. The biliary Eimeria-like coccidia of reptiles classified into the genus Choleoeimeria form a sister clade to the family Eimeriidae, which confirms the separate generic status of the genus Choleoeimeria. The position of Goussia is less robustly resolved, since it forms a trichotomy with the Eimeriidae and Sarcocystidae, or alternatively constitutes the earliest branch of the coccidian lineage. Morphological similarities, namely the extracytoplasmic location of the endogenous stages, and the presence of sutures in the sporocyst wall are discussed in the context of the traditional classification of eimeriids. In contrast to the morphology-based systematics, the monophyly of Goussia and Choleoeimeria is not supported by the SSU rDNA data.

Phylogenetic analysis of Sarcocystis spp. of mammals and reptiles supports the coevolution of Sarcocystis spp. with their final hosts

Sequences of the small subunit rRNA genes were obtained for two coccidians, Sarcocystis dispersa and an unnamed Sarcocystis sp. which parasitise the European barn owl and an African viperid snake as their final host, respectively, and share mouse as their intermediate host. Phylogenetic analysis of the sequence data showed that Sarcocystis sp. from the viperid snake is most closely related to another Sarcocystis sp. isolated from an American crotalid snake, while S. dispersa grouped with other bird-transmitted species. The available dataset failed to resolve the evolutionary relationships among four major branches into which all Sarcocystidae and Isospora spp. were split. However, within these branches, the phylogenetic relationships of the majority of analysed members of the genus Sarcocystis reflected coevolution with their final, rather than intermediate hosts.

A NEW SPECIES OF HEPATOZOON (APICOMPLEXA: ADELEORINA) FROM PYTHON REGIUS (SERPENTES: PYTHONIDAE) AND ITS EXPERIMENTAL TRANSMISSION BY A MOSQUITO VECTOR

Hepatozoon ayorgbor n. sp. is described from specimens of Python regius imported from Ghana. Gametocytes were found in the peripheral blood of 43 of 55 snakes examined. Localization of gametocytes was mainly inside the erythrocytes; free gametocytes were found in 15 (34.9%) positive specimens. Infections of laboratory-reared Culex quinquefasciatus feeding on infected snakes, as well as experimental infection of juvenile Python regius by ingestion of infected mosquitoes, were performed to complete the life cycle. Similarly, transmission to different snake species (Boa constrictor and Lamprophis fuliginosus) and lizards (Lepidodactylus lugubris) was performed to assess the host specificity. Isolates were compared with Hepatozoon species from sub-Saharan reptiles and described as a new species based on the morphology, phylogenetic analysis, and a complete life cycle.

A NEW SPECIES OF MYXIDIUM (MYXOSPOREA: MYXIDIIDAE), FROM THE WESTERN CHORUS FROG, PSEUDACRIS TRISERIATA TRISERIATA, AND BLANCHARD'S CRICKET FROG, ACRIS CREPITANS BLANCHARDI (HYLIDAE), FROM EASTERN NEBRASKA: MORPHOLOGY, PHYLOGENY, AND CRITICAL COMMENTS ON AMPHIBIAN MYXIDIUM TAXONOMY

During March 2001–April 2004, 164 adult anurans of 6 species (47 Rana blairi, 35 Rana catesbeiana, 31 Hyla chrysoscelis, 31 Pseudacris triseriata triseriata, 11 Bufo woodhousii, and 9 Acris crepitans blanchardi) from Pawnee Lake, Lancaster County, Nebraska, were surveyed for myxozoan parasites. Of these, 20 of 31 (65%) P. triseriata triseriata and 1 of 9 (11%) A. crepitans blanchardi were infected with a new species of Myxidium. Myxidium melleni n. sp. (Myxosporea) is described from the gallbladder of the western chorus frog, P. triseriata triseriata (Hylidae). This is the second species of Myxidium described from North American amphibians. Mature plasmodia are disc-shaped or elliptical 691 (400–1,375) × 499 (230–1,200) × 23 (16–35) μm, polysporic, producing many disporic pansporoblasts. The mature spores, 12.3 (12.0–13.5) × 7.6 (7.0–9.0) × 6.6 (6.0–8.0) μm, containing a single binucleated sporoplasm, are broadly elliptical, with 2–5 transverse grooves on each valve, and contain 2 equal polar capsules 5.2 (4.8–5.5) × 4.2 (3.8–4.5) μm positioned at opposite ends of the spore. Myxidium melleni n. sp. is morphologically consistent with other members of Myxidium. However, M. melleni n. sp. was phylogenetically distinct from other Myxidium species for which DNA sequences are available. Only with improved morphological analyses, accompanied by molecular data, and the deposit of type specimens, can the ambiguous nature of Myxidium be resolved. Guidelines for descriptions of new species of Myxidium are provided.

Infectivity of Cryptosporidium muris isolated from cattle

The infectivity of a bovine isolate of Cryptosporidium muris for various animals was studied by transmission experiments. Neonatal BALB/c mice, adult BALB/c mice, SCID mice, common voles (Microtus arvalis), bank voles (Clethrionomys glareolus), common field mice (Apodemus sylvaticus), Mongolian gerbils (Meriones unguiculatus), desert gerbils (Gerbilus gerbilus), guinea pigs, rats, rabbits and goats were used to test the infectivity of this isolate. Among these host species, only Mongolian gerbils were susceptible to the infection and discharged C. muris oocysts in their faeces. The prepatent period for 8-week-old Mongolian gerbils was 15-19 days, the patent period varied between 18 and 36 days. More protracted chronic infections have been observed in gerbils immunosuppressed with methylprednisolone. No signs of clinical illness or macroscopic changes were seen in infected gerbils. Cryptosporidial developmental stages were detected in the stomach, histopathological changes were characterized by epithelial hyperplasia and mucosal hypertrophy without inflammatory exudate. In spite of the fact that C. muris was able to infect gerbils, we do not consider gerbils to be a true hosts for C. muris of cattle origin. Based on our results, we suggest that significant differences in host specificity of individual C. muris isolates exist, and that wild rodents are not reservoir for C. muris infection of cattle.

Hyalomma aegyptium as dominant tick in tortoises of the genus Testudo in Balkan countries, with notes on its host preferences

Collection of 1327 ticks sampled throughout Greece, Bulgaria, Romania and Croatia, from 211 tortoises belonging to three species, Testudo marginata Schoepff, T. graeca Linnaeus, and T. hermanni Gmelin, revealed the presence of four species of ixodid ticks, namely Hyalomma aegyptium (Linnaeus), Haemaphysalis sulcata Canestrini and Fanzago, H. inermis Birula and Rhipicephalus sanguineus (Latreille). Study confirmed the strong dominance of all life stages of H. aegyptium among ticks parasitizing west Palaearctic tortoises of genus Testudo Linnaeus. Furthermore, a considerable portion of ticks collected from tortoises in southwestern Bulgaria represent larvae and nymphs of H. sulcata. At the same area we collected as exception one larva and one nymph of H. inermis from a single specimen of T. hermanni. Our findings of four adults of R. sanguineus is the first record of this species from reptilian host. According to our results achieved on localities with syntopic occurrence of two tortoise species, T. marginata and T. graeca represent in the Balkans the principal hosts of H. aegyptium, whereas T. hermanni serves only as an alternative host in the areas close to range of either T. marginata or T. graeca.