Molecular detection of cattle Sarcocystis spp. in North-West Italy highlights their association with bovine eosinophilic myositis.

BACKGROUND: Cattle are intermediate hosts of six Sarcocystis species, among which Sarcocystis hominis and Sarcocystis heydorni can infect humans through the consumption of raw or undercooked meat. In addition to the zoonotic potential, there is increasing interest in these protozoa because of the evidence supporting the role of Sarcocystis spp. in the occurrence of bovine eosinophilic myositis (BEM), a specific inflammatory myopathy which leads to carcass condemnation and considerable economic losses. Actually, all the prevalence studies carried out on cattle in Italy have been based on either morphological or 18S rDNA-based molecular techniques, most likely leading to misidentification of closely related species. Therefore, there is a strong need for new data on the prevalence of the different Sarcocystis spp. in cattle in Italy and their association with bovine eosinophilic myositis. METHODS: To reach our aim, individual striated muscle samples from BEM condemned carcasses (N = 54) and diaphragm muscle samples from randomly sampled carcasses (N = 59) were obtained from Northwest Italy slaughterhouses. Genomic DNA was extracted and analyzed by multiplex-PCR targeting 18S rDNA and cox1 genes. PCR products amplified using the genus-specific primer set in absence of the specific fragment for S. hirsuta, S. cruzi, S. hominis or S. bovifelis were sequenced to achieve species identification. RESULTS: Sarcocystis DNA was detected in 67.8% of the samples from slaughter cattle and in 90.7% of the samples from BEM condemned carcasses. S. cruzi was identified as the most prevalent species in slaughter cattle (61%), followed by S. bovifelis (10.2%), S. hominis (8.5%) and S. hirsuta (1.7%). Notably, among the different Sarcocystis spp. detected, the presence of S. bovifelis and S. hominis was significantly higher in samples isolated from BEM condemned carcasses (46.3% and 40.7% respectively), while there was no statistically significant difference between the presence of S. cruzi or S. hirsuta in BEM condemned carcasses (42.6% and 1.8%, respectively) and randomly sampled carcasses. Furthermore, DNA sequence analysis revealed the presence of a putative new species in two carcasses. CONCLUSIONS: Our study contributes to updating the data on the prevalence of the different Sarcocystis spp. in cattle in Italy, highlighting the presence of three Sarcocystis spp., S. cruzi, S. hominis and S. bovifelis, in BEM lesions and allowing us to speculate on the possible role of S. hominis and S. bovifelis as the major sarcosporidian species involved in bovine eosinophilic myositis.

Schistosoma mansoni granulomas in the skeletal striated muscles in the murine model of neuroschistosomiasis: histological findings.

Schistosomiasis mansoni presents many clinical manifestations during migration of schistosomes in their hosts, including diarrhea, hepatomegaly, splenomegaly, liver abscesses, skinlesions, brain tumors and myeloradiculopathy. No lesions have been reported in skeletal striated muscles due to schistosomiasis mansoni in the literature. This short communication reports the histopathological findings on skeletal musculature in a murine model of neuroeschistosomiasis mansoni. Lesions were found in the tongue, masseter muscle, buccinator muscle, digastric muscle and temporalis muscle. Worm recovery was carried out to confirm the infection. We describe here, for the first time in the literature, injuries in the skeletal musculature due to Schistosoma mansoni nfection.

Schistosoma mansoni granulomas in the skeletal striated muscles in the murine model of neuroschistosomiasis: histological findings

Schistosomiasis mansoni presents many clinical manifestations during migration of schistosomes in their hosts, including diarrhea, hepatomegaly, splenomegaly, liver abscesses, skinlesions, brain tumors and myeloradiculopathy. No lesions have been reported in skeletal striated muscles due to schistosomiasis mansoni in the literature. This short communication reports the histopathological findings on skeletal musculature in a murine model of neuroeschistosomiasis mansoni. Lesions were found in the tongue, masseter muscle, buccinator muscle, digastric muscle and temporalis muscle. Worm recovery was carried out to confirm the infection. We describe here, for the first time in the literature, injuries in the skeletal musculature due to Schistosoma mansoni nfection.

Identification and genetic characterization of Sarcocystis arctica and Sarcocystis lutrae in red foxes (Vulpes vulpes) from Baltic States and Spain.

BACKGROUND: Typically, carnivores serve as definitive hosts for Sarcocystis spp. parasites; currently, their role as intermediate hosts is being elucidated. The present study aimed to identify and molecularly characterize Sarcocystis cysts detected in striated muscle of red foxes from different populations in Latvia, Lithuania and Spain. METHODS: Muscle samples from 411 red foxes (Vulpes vulpes) and 269 racoon dogs (Nyctereutes procyonoides) from Latvia, 41 red foxes from Lithuania and 22 red foxes from Spain were examined for the presence of Sarcocystis sarcocysts by light microscopy (LM). Sarcocystis spp. were identified by transmission electron microscopy (TEM) and molecular biology techniques. RESULTS: Sarcocystis cysts were detected in 11/411 (2.7%) Latvian, 3/41 (7.3%) Lithuanian, and 6/22 (27.3%) Spanish red foxes, however, cysts were not observed in the muscles of racoon dogs. Based on LM, TEM, 18S rDNA, 28S rDNA, ITS1, cox1 and rpoB sequences, Sarcocystis arctica and Sarcocystis lutrae cysts were identified in red fox muscles from Latvia and Lithuania, whereas only S. arctica was detected in Spain. The 18S rDNA, 28S rDNA and ITS1 sequences from the 21 isolates of S. arctica from Latvia, Lithuania and Spain were identical. By contrast, two and four haplotypes were determined based on mtDNA cox1 and apicoplast rpoB sequences, respectively. Polymorphisms were not detected between the two isolates of S. lutrae from Latvia and Lithuania. Based on phylogenetic results, S. arctica and S. lutrae were most closely related to Sarcocystis spp. using predatory mammals as intermediate hosts and to Sarcocystis species with a bird-bird life-cycle. CONCLUSIONS: Based on current knowledge, the red fox and Arctic fox (Vulpes lagopus) could act as intermediate host for the same two Sarcocystis species. Molecular results suggest the existence of two genetic lineages of S. arctica, and such divergence relies on its geographical distribution but not on their intermediate host species.

Antiprotozoal drug nitazoxanide enhances parasitemia, tissue lesions and mortality caused by Trypanosoma cruzi in murine model.

Chagas' disease is caused by unicellular parasite Trypanosoma cruzi (T. cruzi). It is endemic throughout Latin America, but nowadays has become a global challenge due to tourism and migration. Non-treated infection may result in health-threatening complications and lead to death. Current medications for this infection are nifurtimox (NFT) and benznidazol. Both drugs may cause side effects and are ineffective in the chronic phase. Therefore, new antichagasic compounds are urgently required. Nitazoxanide (NTZ) is a broad spectrum antiparasitic drug, proposed recently as a potential candidate to be added to the list of essential medicines for integrated neglected tropical disease control and elimination. Although the effect of NTZ against T. cruzi epimastigotes in vitro was reported, the corresponding experiments in animal models of T. cruzi infection have never been undertaken. The present work was designed to fill this gap and evaluate the effect of NTZ on experimental murine trypanosomiasis, in comparison with classical antichagasic agent NFT. Highly sensitive to T. cruzi BALB/c mice were infected using Albarrada T. cruzi strain, recently isolated in Mexico. Experimental groups were either left untreated, or otherwise treated with NFT, NTZ (100 and 1000 mg/kg), or with both drugs simultaneously. The severity of the infection was estimated based on criteria such as parasitemia, lesions in target tissues (heart, muscles and lungs) and mortality. Despite the expected protective effect, NTZ drastically aggravates the course of T. cruzi infection. Namely, parasitemia, tissue lesions and mortality caused by T. cruzi infection were significantly higher in NTZ-treated mice groups, even in comparison with untreated infected animals. NTZ by itself no produced mortality o tissue damage, and NFT showed an expected protective effect. Our results indicate that NTZ cannot be considered for Chagas' disease treatment. Moreover, NTZ should be used with caution in patients positive for T. cruzi infection.

Trichosomoides nasalis (Nematoda: Trichinelloidea) in the murid host Arvicanthis niloticus: migration to the epithelium of the nasal mucosa after intramuscular development.

Knowledge of the biology of the trichinelloid subfamily Trichosomoidinae is poor. Trichosomoides nasalis is a common parasite of Arvicanthis niloticus (Muridae) in Senegal, and a procedure for experimental infections has been established. It has been demonstrated that larvae develop in striated muscle fibres, similar to Trichinella spp., but they are not arrested in the first stage, and they reach the adult stage within three weeks. In the present histological study it is shown that T. nasalis females and dwarf males migrate from the abdomen and thorax to the host's muzzle, moving through connective tissues and between muscles. A few migrating specimens were also found in the blood vessels of the nasal mucosa. While sexes were still separated in the lamina propria of the mucosa, females recovered from the epithelium contained intra-uterine males. Worms were found between the incisors in the mucosa of the anterior and median conchae which are rich in mucous cells. Only the pseudostratified epithelium was parasitized. Under natural conditions, the inflammation of the nasal mucosa that is induced by the parasites might reduce the competitiveness of infected rodents when foraging or looking for potential mates.

Development of Trichosomoides nasalis (Nematoda: Trichinelloidea) in the murid host: evidence for larval growth in striated muscle fibres.

Trichosomoides nasalis (Trichinelloidea) is a parasite of Arvicanthis niloticus (Muridae) in Senegal. Female worms that harbour dwarf males in their uteri, occur in the epithelium of the nasal mucosa. Young laboratory-bred A. niloticus were either fed females containing larvated eggs or intraperitoneally injected with motile first-stage larvae recovered from female uteri. Both resulted in successful infection. Organs examined during rodent necropsy were blood and lymphatic circulatory systems (heart, large vessels, lymphnodes), lungs, liver, kidneys, thoracic and abdominal cavities, thoracic and abdominal muscular walls, diaphragm, tongue, and nasal mucosa. Development to adult nasal stages took three weeks. Recovery of newly hatched larvae from the peritoneal fluid at four-eight hours after oral infection suggests a direct passage from the stomach or intestinal wall to the musculature. However, dissemination through the blood, as observed with Trichinella spiralis, cannot be excluded even though newly hatched larvae of T. nasalis are twice as thick (15 µm). Developing larvae were found in histological sections of the striated muscle of the abdominal and thoracic walls, and larvae in fourth moult were dissected from these sites. Adult females were found in the deep nasal mucosa where mating occurred prior to worms settling in the nasal epithelium. The present study shows a remarkable similarity between T. nasalis and Trichinella species regarding muscle tropism, but the development of T. nasalis is not arrested at the late first-larval stage and does not induce transformation of infected fibres into nurse cells. T. nasalis seems a potential model to study molecular relations between trichinelloid larvae and infected muscle fibres.

Description of a new species of myxozoan from Notropis hudsonius in the Great Lakes region of Canada.

Myxobolus burti n. sp. is described from striated muscle of Notropis hudsonius (Cyprinidae) collected from localities in Lake Superior, Lake Michigan, Lake Huron, Lake St. Claire, Lake Erie, Lake Ontario, and the St. Lawrence River. Myxobolus burti is intracellular, forming thin-walled, cigar-shaped plasmodia within striated muscle cells of the body. It exhibits disporoblastic, oval spores, approximately 10 µm long, 8 µm wide, and 6 µm thick, with a length-to-width ratio of 1.3 ∶ 1. They resemble most closely those of Myxobolus bellus Kudo, 1934, and Myxobolus mutabilis Kudo, 1934, but have polar capsules unequal in length (5.3 vs. 4.7 µm) and oblique filament coils. The new parasite was not encountered in routine examination of other small-bodied fishes at collection locations and thus looks to be specific to N. hudsonius. A comparative plate of similar species encountered during surveys of these fishes in the northeast is presented to contrast the uniqueness of the new species.

Detection of Leishmania spp. and associated inflammation in ocular-associated smooth and striated muscles in dogs with patent leishmaniosis.

OBJECTIVE: Canine leishmaniosis is a disease characterized by the wide distribution of the parasite throughout the tissues of the host. The purpose of this study was to describe the presence of Leishmania spp. and associated inflammation in ocular-associated muscles of dogs with patent leishmaniosis. PROCEDURES: Smooth muscles (iris dilator muscle, iris sphincter muscle, ciliary muscle, Müller muscle, smooth muscle of the periorbita and smooth muscle of the nictitating membrane) and striated muscles (orbicularis oculi muscle, obliquus dorsalis muscle and dorsal rectus muscle) were evaluated. Routine staining with hematoxylin and eosin and immunohistochemistry to detect Leishmania spp. were performed on tissue sections. RESULTS: Granulomatous inflammation was seen surrounding muscular fibers and was composed mainly of macrophages with scattered lymphocytes and plasma cells. This infiltrate could be seen in 52/473 (10.99%) samples of smooth muscle and 36/142 (25.35%) samples of striated muscle. Parasites were detected in 43/473 (9.09%) samples of smooth muscle and in 28/142 (19.71%) samples of striated muscle. CONCLUSIONS: To the authors' knowledge, this is the first report assessing the presence of Leishmania spp. and associated infiltrate in intraocular, extraocular and adnexal smooth and striated muscles. The inflammation present in those muscles could contribute to clinical signs already described, such as blepharitis, uveitis, and orbital cellulitis.

Taxonomy of Myxobolus ridouti N. Sp. and M. ridgwayi N. Sp. (Myxozoa) from Pimephales notatus and Semotilus atromaculatus (Cypriniformes) in Ontario.

Myxobolus ridouti n. sp. and M. ridgwayi n. sp. are described, respectively, from Pimephales notatus (Rafinesque) and Semotilus atromaculatus (Mitchill) in Algonquin Park, Ontario, Canada. Both are intracellular in striated muscles of the body flank. Spores of M. ridouti n. sp. are subcircular (9.5-10.5 µm long by 9.4-10.9 µm wide); those of M. ridgwayi n. sp. are oval (10.0-12.1 µm long by 9.5-10.5 µm). Both species have spores with a small, distinct swelling (1 µm) on the spore valve near the posterior sutural ridge, similar to that described for Myxobolus insignis Eiras, Malta, Varella, and Pavanelli, 2005. It looks to be non-nuclear in origin, apparently part of the internal face of a spore valve, and could be taxonomically useful. Partial sequence data of the 18S rDNA from both species are unique among those species in the genus that have been sequenced. The sequences have the greatest overall identity with various species of Myxobolus from cyprinids in North America, Europe, and Asia, with a constructed molecular phylogeny having the 2 new species forming a separate clade.

Immunization with an engineered mutant trans-sialidase highly protects mice from experimental Trypanosoma cruzi infection: a vaccine candidate.

Chagas' disease is a major tropical disease for which a cure for chronic phase does not exist yet. Trypanosoma cruzi trans-sialidase (TS) seems to be involved in relevant processes such as infectivity, host survival and, very importantly, disease pathogenesis. In this study, we show that mice vaccinated with an engineered enzymatically deficient mutant TS containing the catalytic domain without the immunodominant SAPA (Shed Acute Phase Antigen) repeats, were highly protected against T. cruzi infection. Adult male BALB/c mice were immunized with mutant protein, purified from Pichia pastoris yeast, using three inoculations in Freund's adjuvant. All immunized mice were protected against challenge with a lethal dose of T. cruzi trypomastigotes. The protected immunized mice developed no clinical or tissue evidence of infection throughout the study. In contrast, 60-90% mortality and 100% occurrence of myocardial lesions were observed in the non-immunized counterparts. Titers of circulating antibody against TS did not correlate with protection, while anti-SAPA antibodies were coincident with disease severity. Further studies indicated that a single inoculation of mutant recombinant protein in Freund's complete adjuvant was not associated with blood or organic alterations, per se. Mutant TS vaccination seems to be a promising tool for immune intervention strategies in Chagas' disease, aimed at preventing T. cruzi-related heart tissue damage.