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Molecular diagnostics of Sarcocystis spp. infections

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Protozoa of the genus Sarcocystis (phylum Apicomplexa, family Sarcocystidae) is one of the most common parasites affecting animals. Interspecies diagnostic of Sarcocystis genus was based on electron microscopy for many years. Because of absence of visible differences between species with reachable magnifications, light microscopy is useless. In many cases serological diagnostic method have lack of sensitivity. A variety of molecular methods have been developed and used to detect and identify Sarcocystis spp. and to assess the genetic diversity among this protozoan from different population/hosts. Nowadays, molecular diagnostic is the common, time/cost effective method used all over the world to interspecies differentiation.
Three different forms of sarcocysts from the zoo animals Pseudois nayaur, Capricomis crispus and Ovibos moschatus (Bovidae, Caprinae) were investigated by light and transmission electron microscopy, in special consideration of the cyst wall. Two of these forms were not distinguishable from Sarcocystis capracanis or the pair of sibling species S. hircicanis/ arieticanis by their morphology. They were, therefore, designated as S. cf. capracanis (in Pseudois) and S. cf. hircicanis/arieticanis (in Pseudois, Capricornis and Ovibos) in this paper. The third form (from Capricornis) was not identifiable as one of the species hitherto known. Its cyst wall, provided with stumpy nail-like, in the outline T-shaped villar protrusions, could not be attributed to one of the types established by Dubey et al. (1989). It was, therefore, described as Sarcocystis capricornis sp. n. The Sarcocystis species described in Caprinae so far were discussed.
Sarcocysts were found during the examination of muscle samples from four different Equidae from the zoo in Berlin-Friedrichsfelde (Przewalski’s feral horse, Chapman’s plain zebra, kulan, kiang), a Damara plain zebra from the Cologne zoo, and a domestic horse from Mongolia. Light and electron microscopic features of these sarcocysts correspond largely. The cyst walls of all the six forms represent type 11 from the classification system by Dubey et al. 1989. On this ultrastructural basis, one could postulate the existence of two or only one species, depending on the emphasis that is laid on the presence or absence of large osmiophilic granules within the cyst wall protrusions. Equus caballus przewalskii, E. burchellii chapmani, E. burchellii antiquorum, E. kiang holdereri, and E. onager kulan have not been previously described as hosts for sarcocysts belonging to ТЕМ wall type 11. In addition to the ТЕМ wall type 11 forms, a type 7 form is known in horses. It is not possible for the time being to list Sarcocystis asinus Gadaev, 1978 as a synonym of a Sarcocystis species found in equids. Hence S. asinus is declared a species inquirenda.
This work presents the results of investigation of cardiac muscle in 137 bisons shot in the Białowieża Forest during breeding selection in the years 1990-1995. The investigation focused upon the occurrence of protozoan Sarcocystis sp. and the lesions caused by it. Macroscopic inspection did not show any lesions characteristic of Sarcocystis sp. Histopathological examination was carried out of sections of the left heart ventricle, the right heart ventricle and interventricular septum. For 137 bisons examined in 117 animals the presence of Sarcocystis sp. in the heart muscle was confirmed, which means 85,4%. Moreover, the cardiac muscle showed inflammatory cell infiltration, hyperemia, degeneration and less frequently necrosis. The animals in which the presence of Sarcocystis was not confirmed belonged to the age group under 1 year. Differences in structure of cysts walls indicate the occurrence of various species of Sarcocystis in bisons.
Acalculous cholecystitis and cholangitis are increasingly being recognized as complications of AIDS. The opportunistic parasites that have been most commonly associated with these disorders are Cryptosporidium species, Isospora belli, Cyclospora cayetanensis and Enterocytozoon bieneusi. The authors performed a parasitological survey on the gallbladder tissue sections of patients underwent cholecystectomy due to chronic acalculous cholecystitis at the Shiraz University of Medical Sciences, Iran. Light microscopic investigation in more than three hundred archived histopathological slides revealed the presence of sexual stages (i.e., mature sporocysts) of a coccidial protozoan in a patient with AIDS who developed acalculous cholecystitis as confirmed by histological, parasitological and molecular tests in which Sarcocystis species was the only identifiable pathogen in gallbladder sections. In the best of our knowledge it’s the first documented case of chronic non-calculous cholecystitis due to Sarcocystis parasite in an Iranian AIDS patient from worldwide.
To investigate the occurrence of sarcocystosis in water buffalo (Bubalus bubalis) in Ahvaz, the Khuzestan province, Iran, and to evaluate an ELISA for the diagnosis of sarcocystosis, serum and oesophagus samples were collected from the 300 water buffaloes, aged 0.5-7 years, and then slaughtered at the Ahvaz abattoir. The oesophagus samples were examined for sarcocysts under microscopic examination, using the digestion method. One hundred and seventy-one (57%) animals were found to be positive for Sarcocystis bradyzoites. One hundred and sixty-three (54.3%) serum samples were positive for sarcocystis antibodies in the ELISA. The prevalence of sarcocystosis was statistically age related, with significantly higher rates in adult buffaloes than young animals (P<0.05). The prevalence did not differ significantly in relation to the gender (P>0.05). The Mc Nemar test revealed a high correlation (94%) between the digestion method and ELISA. The ELISA, with the use of antigens from S. fusiformis bradyzoites, as presented in this study, can be adapted to detect antibodies to Sarcocystis sp., with an acceptable specificity and sensitivity.
A species of Sarcocystis is reported from a naturally infected African grey parrot, Psittacus erithacus, from Costa Rica. Only mature sarcocysts, measuring up to 2 mm in length and up to 750 μm in width, were observed. The sarcocyst wall was smooth. The villar protrusions on the sarcocyst wall were up to 5 μm long and up to 1.1 μm wide; they were folded over the sarcocyst wall giving a thin-walled appearance. The microtubules in villar protrusions were smooth and confined to villar protrusions. Bradyzoites in sections were 5.4–6.6 × 1.3–2.0 μm in size. Sequencing the small subunit and first internal transcribed spacer portions of ribosomal DNA related this parasite to, but distinguished it from, previously characterized species of Sarcocystis that encyst in the musculature of birds and complete their sexual development in New World opossums of the genus Didelphis. This evidence suggests that the parrot may have acquired its infection from an opossum from which it suffered a debilitating attack a year prior to the onset of depression, anorexia, and ultimately death.
A species of Sarcocystis is reported from two naturally infected Buffon’s macaws (Ara ambigua) from Costa Rica. Only mature sarcocysts, measuring up to 950 μm in length and up to 75 μm in width, were observed. By light microscopy the sarcocyst wall was thin (< 1 μm thick) and smooth. The villar protrusions on the sarcocyst wall were up to 4.0 μm long and up to 0.6 μm wide; they were folded over the sarcocyst wall giving a thin-walled appearance. The microtubules in villar protrusions were smooth and confined to villar protrusions. Bradyzoites in sections were 4.0–5.9 × 0.8–1.8 μm in size. Structurally, sarcocysts from the macaw appeared different from sarcocysts of other avian species. This is the first report of Sarcocystis infection in this host.
Sarcocystosis of wild ducks, relatively common in North America, has hitherto been recorded only three times in Europe. The present study yielded a single case of macroscopically detected sarcocysts in the skeletal muscles of a mallard (Anas platyrhynchos). This finding constitutes the first such record in Poland. A PCR technique was used for identification of the parasite from sarcocysts. The results obtained suggest that the sarcocysts were produced by protozoans, which were the most closely related to Frenkelia glareoli and Sarcocystis neurona (homology of the nucleotide sequences was 96.6% in both cases).
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