Ultrastructure of sarcocysts from water buffalo in India

The ultrastructure of sarcocysts of macro- and microscopic species of Sarcocystis was compared from naturally infected water buffalo from India. Grossly visible sarcocysts had walls consisting of cauliflower-like villar protrusions, typical of S. fusiformis. The sarcocyst wall of the microscopic species of Sarcocystis was 6.4 microns thick and consisted of tightly packed conical villar protrusions that were 9.6 microns long and 3.7 microns wide at the base. At approximately 3 microns above the base, the distal two-thirds of the villar protrusion became conical shaped and was bent laterally at an angle of 45 degrees to the sarcocyst surface. The granular layer beneath the villar protrusions was 0.9 microns thick. In S. levinei the granular layer was 1.9 microns thick, the villar protrusions were narrow and it had a highly undulating primary cyst wall. Whether the microscopic S. levinei-like sarcocysts of Indian and Malaysian water buffalo are distinct species of Sarcocystis will require further investigation.     

Association of eosinophilic myositis with an unusual species of Sarcocystis in a beef cow.

The carcass of a mature cow had numerous, disseminated lesions typical of eosinophilic myositis. To elucidate the nature and possible cause of the lesions, histological sections were examined by light microscopy and selected areas were removed and processed for electron microscopy. The lesions were granulomatous in nature. Each granuloma contained at its centre an intact or ruptured sarcocyst associated with degenerate muscle fibers. Surrounding this was a layer of epithelioid cells and an intense accumulation of inflammatory cells, most of which were eosinophils. The primary cyst wall of the sarcocysts in these granulomas consisted of hair-like protrusions that featured many unusual electron-dense bodies. Sarcocysts with ultrastructures characteristic of Sarcocystis cruzi and Sarcocystis hirsuta were also present in muscle from the same animal, but these sarcocysts lacked any associated cellular responses. The eosinophilic myositis in this case appeared to be associated with sarcocystosis of an unknown species. Possibly, the inflammatory reaction was due to the host-parasite interaction in an unusual host.     

Prevalence of Sarcocystis spp. in Argentinean cattle

Sarcocystis cruzi, S. hirsuta and S. hominis are apicomplexan parasites that affect cattle worldwide with variable prevalence. The aim of the present study was to evaluate the prevalence of Sarcocystis spp. in Argentinean cattle comparing microscopic fresh examination and molecular methods. Blood, myocardium and loin samples were collected in five slaughterhouses from a total of 380 bovines. Origin of animals was representative of the major beef cattle production area of Argentina. Samples were analyzed by fresh microscopical examination, transmission electron microscopy (TEM), IFAT and PCR-RFLP. Thin walled sarcocysts corresponding with S. cruzi were found in 99.5% of heart samples. Sarcocysts were detected in 73.1% of loin samples; 71.5% had S. cruzi cysts and 23.1% had thick walled sarcocysts (S. hirsuta or S. hominis). TEM observation revealed the presence of characteristic S. hominis and S. hirsuta cyst walls in 7 and 1 loin samples respectively. Using IFAT, 379/380 animals had titers 25 or higher, showing a full agreement with fresh examination. Amplification products were detected in 35.5% (135/380) of loin samples; however Sarcocystis species could only be determined by RFLP in 29 samples. Agreement between fresh examination and PCR was low (Kappa value=0.262). This is the first report of S. hominis and S. hirsuta in Argentina. Further studies are needed to improve the sensitivity of molecular methods for species identification, especially for differentiation of S. cruzi and S. hirsuta from the zoonotic species S. hominis. The results of the present study and others focusing on sensitivity and specificity of Sarcocystis spp. diagnostic methods should contribute to improve food safety.     

Prevalence of Sarcocystis in domestic pigs in India.

Muscle samples from 890 slaughtered pigs were examined for the presence of sarcocysts. A high prevalence rate of 67.98% was observed. Two types of microsarcocysts were recorded. The sarcocyst wall of one type had redial striations and the other possessed hair-like villar protrusions. The species were identified as Sarcocystis miescheriana and Sarcocystis suihominis; there was a slightly higher incidence of the latter species (47.11%) than of the former (43.14%). S. suihominis has been identified for the first time from pigs in India.     

The structure and identity of macroscopically visible Sarcocystis cysts in cattle

Macroscopically visible Sarcocystis spp. cysts isolated from the skeletal muscle of slaughtered cattle were examined by light- and electronmicroscopy. Transmission experiments involving cats, dogs and a human volunteer were also carried out. The cysts could only be transmitted to cats which establishes them with a high degree of certainty as Sarcocystis hirsuta. The cyst wall (including protrusions) ranged from 3.3 to 7.0 micron in thickness and the individual cyst wall protrusions from 1.2 to 2.6 micron in width. Transmission and scanning electronmicroscopy revealed previously undescribed features of the cyst wall. It appears that, with increasing age, the cyst wall protrusions become larger and develop a highly irregular surface. Their attachments to the cyst wall are slender and widely spaced indicating that growth of the cyst continues without the formation of new protrusions. Within the protrusions the fibrils become disorganised and numerous osmiophilic granules appear. It is evident that major changes in the structure of sarcocysts can occur with age.     

Sarcocystis infections in mule deer (Odocoileus hemionus) in Montana and the descriptions of three new species

Four structural types of sarcocysts of Sarcocystis were found in skeletal muscles of mule deer in Montana. Type I sarcocysts were thin walled (1 to 3 micron) and belonged to S hemionilatrantis. Types II to IV sarcocysts were thick walled (2 to 10 microns) and new names were proposed for them. Type II sarcocysts with long villar projections were named S hemioni. Type III sarcocysts with club-shaped villi of uneven thickness were named S youngi. Type IV sarcocysts with walls of uneven thickness and containing hair-like protrusions were named S americana.     

First isolation of Sarcocystis hominis from cattle in Japan.

Sarcocystis hominis was first isolated from slaughtered cattle raised in Japan. Cysts were 1,220-4,460 x 80-384 microns in size and their wall was 3 to 6 microns thick and appeared radially striated in the histopathological sections because of the presence of palisade-like villar protrusions on the surface. The protrusions were 3.1-4.3 x 0.7-1.1 microns in size and had many microtubules in the core. Two cynomolgus monkeys, Macaca fascicularis, fed with the Sarcocystis cysts began to pass sporocysts, which measured a size of 14.3-15 x 9.5-10 microns, in the feces 10 days after ingestion.     

The prevalence and identity of Sarcocystis in beef cattle in New Zealand

Muscle tissue from the oesophagus and diaphragm of 500 beef cattle slaughtered in New Zealand was examined for Sarcocystis infection by microscopic examination of cysts isolated from muscle samples. All cattle were infected with Sarcocystis; based on light microscopy of cysts, 98% had thin-walled Sarcocystis cruzi cysts and 79.8% had thick-walled (Sarcocystis hirsuta/Sarcocystis hominis) cysts. Cysts were also collected for electron microscopy and transmission experiments. Thick-walled cysts could not be distinguished as S. hirsuta or S. hominis by light or electron microscopy. Thick-walled cysts were fed to three cats and one human volunteer; one cat shed sporocysts but not the human volunteer. Electron microscopy of the cysts revealed many features that have not been described previously.     

The prevalence and identity of Sarcocystis cysts in cattle in Belgium

Muscle tissue from the oesophagus, diaphragm and heart of 100 cattle slaughtered in Belgium was examined for Sarcocystis infection by microscopic examination of tissue and artificial digestion. Intact sarcocysts or cystozoietes were recovered from 97% of the cattle examined. There was a difference in sensitivity between the method (digestion or histology) used and the muscle processed. The digestion of the oesophagus muscle resulted in the highest number of positive animals whereas the heart muscles contained most cysts during histological examination. Thin-walled cysts were recovered from all positive animals especially in the heart and they were indistinguishable from those of S. cruzi. Thick-walled cysts were recovered from 56% of animals but these could not be identified as S. hirsuta and/or S. hominis on morphological grounds. A correlation between pathological changes and the infection grade could not be proved.     

Sarcocystis sinensis is an ultrastructurally distinct parasite of water buffalo that can cause foodborne illness but cannot complete its life-cycle in human beings

In this study, we compared the morphology of Sarcocystis sinensis and Sarcocystis hominis, and assessed the infectiousness of S. sinensis for human volunteers. The cysts of S. sinensis were from water buffalo (Bubalus bubalis) and those of S. hominis were from cattle (Bos taurus). Transmission electron microscopy of S. sinensis cysts revealed that the cyst wall had leaning, finger-like protrusions measuring 1.44-5.08 μm in length and without invaginations on the tip surface of the protrusions. In contrast, the cyst wall of S. hominis had upright, finger-like protrusions measuring 9.43 μm×2.42 μm and with vesicle-like invaginations on the tip surface of the protrusions. Scanning electron microscopy revealed that surface of the protrusions was arranged as rectangles in S. sinensis, as compared to tongue-shaped in S. hominis. Other distinguishing features of S. sinensis include a thin ground substrate (GS) zone with microtubules and small, circle-like structures located at the base of the protrusions. Human volunteers, after consuming S. sinensis cysts, produced no sporocysts or oocysts in feces, suggesting that humans could not serve as definitive hosts for S. sinensis. By contrast, many sporocysts and oocysts were passed in feces of a human volunteer 11-29 days after ingestion of S. hominis cysts. These results showed that S. sinensis and S. hominis are separate species and S. sinensis cannot use human being as the definitive host.     

Naturally occurring Sarcocystis neurona-like infection in a dog with myositis

Tissue stages similar to those of Sarcocystis neurona, the causative agent of equine protozoal myeloencephalitis, were identified in skeletal muscles of a dog. The dog, a 6-year-old Labrador retriever, was seropositive for Toxoplasma gondii infection and euthanized due to a history of polymyositis and progressive muscular atrophy. Histologically, 30, variably sized, microscopic, intracellular sarcocysts were observed in 60 sections of skeletal muscles taken from the neck, fore limbs and hind limbs. The cysts were only observed in inflamed skeletal muscles, but were mostly in myocytes at the periphery of areas infiltrated with leukocytes. Ultrastructurally, the cyst wall had villar protrusions consistent with sarcocysts. Immunohistochemistry with monoclonal S. neurona antibodies demonstrated positive labeling of zoites in merozoites or schizonts in the skeletal muscle interstitium, but no labeling of the sarcocysts. Initial PCR analysis with primers amplifying a genetic sequence encoding Apicomplexan 18s rRNA, and subsequent PCR analysis with differentiating primers indicated that the genetic sequences had 100% identity with sequences reported for S. neurona.     

Prevalence of Sarcocystis sarcocysts in nine-banded armadillos (Dasypus novemcinctus) from Florida.

The prevalence and identity of Sarcocystis spp. sarcocysts in the skeletal muscles of nine-banded armadillos (Dasypus novemcinctus) collected from Alachua County, FL, were determined. H & E stained sections of skeletal muscle from tongue and thigh were examined. Thirty nine of 63 (61.9%) armadillos examined contained Sarcocystis sarcocysts. Two species were identified, Sarcocystis dasypi and Sarcocystis diminuta. Sarcocystis dasypi sarcocysts were found in 38 of 63 (60.3%) and S. diminuta sarcocysts were found in 6 of 63 (9.5%). Sarcocysts of S. dasypi were larger, more densely packed with bradyzoites, and bradyzoites contained within the sarcocyst were smaller than those of S. diminuta. Mixed infections occurred in 5 of 63 (7.9%) armadillos examined.     

Prevalence of thin-walled <Emphasis Type="Italic">Sarcocystis cruzi</Emphasis> and thick-walled <Emphasis Type="Italic">Sarcocystis hirsuta</Emphasis> or <Emphasis Type="Italic">Sarcocystis hominis</Emphasis> from cattle in Iran

Bovine sarcocystosis is caused by Sarcocystis cruzi and is known to cause considerable morbidity and mortality in cattle. This species is distributed worldwide in cattle and is the most prevalent of the Sarcocystis species infecting cattle. There is high infection rate of sarcocyst in cattle in Iran, but to our knowledge, there is no study about identification of Sarcocystis species. This work aimed to survey prevalence of S. cruzi cyst in slaughtered cattle of Isfahan, Iran. In this study, esophageal and diaphragmatic muscles of 100 cattle were collected from Fesaran abattoir of Isfahan and examined for the presence of Sarcocystis spp. cysts macroscopically and microscopically. No macroscopic sarcocysts were found in any of the samples. In light microscopy, 89 out of 100 cattle (89%) had thin-walled cysts of S. cruzi, while 21 out of them (21%) had thick-walled sarcocysts. In addition to light microscopy, ultrastructural features of the thin-walled cyst confirmed the presence of S. cruzi.     

Sarcocystis sp. from cattle slaughtered in Japan

Sarcocystis sp. was detected from cattle slaughtered in Saitama Prefecture, Japan. The cysts were 3,400-4,400 x 198-238 microm in size and had the thick cyst wall which was 7 to 10 microm thick and provided with finger-like villar protrusions. The protrusions were 8-9.5 x 2-2.5 microm in size and had microtubules in the core.     

Sarcocystis infection in the white-tailed deer (Odocoileus virginianus) in Montana: intensity and description of Sarcocystis odoi n sp

Sarcocystis infection was diagnosed in 27 of 36 (75%) samples of meat from white-tailed deer (Odocoileus virginianus) in Montana. Two structurally distinct thin- and thick-walled sarcocysts were found in the white-tailed deer; the thin-walled sarcocysts were those of S odocoileocanis. A new name, S odoi, was proposed for the thick-walled sarcocysts. Sarcocysts of S odoi were up to 1,050 microns long and 260 microns wide and contained a 6.5- to 12-microns thick wall. The ultrastructure of sarcocysts of S odocoileocanis was compared with that of S odoi. A cat fed meat containing thin- and thick-walled sarcocysts shed oocysts and sporocysts 24 days later. The sporocysts in the cat's feces were 13.3 X 9.6 microns and probably belonged to S odoi.     

Experimental infection with Sarcocystis medusiformis in sheep

The development of the parasite was studied in 48 sheep killed between 188 and 1132 days after experimental inoculation with Sarcocystis medusiformis sporocysts from cats. Immature sarcocysts were present at 188 days post inoculation (d.p.i.). At 331 d.p.i. macroscopic sarcocysts with an elongate fusiform appearance were seen in the laryngeal, abdominal and diaphragm musculature. The largest cyst measured 2 mm in length by 0.5 mm in width at 331 d.p.i.; histologically they contained metrocytes at the periphery of the cyst with more densely staining merozoites in the central region. By 443 d.p.i. typical 'thin' cysts 2-3.5 mm in length were seen in the flank and external thoracic muscles. By 765 d.p.i. sarcocysts were 5 mm in length. The ultrastructure of the cyst wall of these cysts resembled that of S. medusiformis. At 1132 d.p.i. sarcocysts measured 4 mm X 0.5 mm.     

Development of Sarcocystis falcatula in its intermediate host, the Brown-headed Cowbird (Molothrus ater)

Sporocysts of Sarcocystis falcatula obtained from experimentally infected Virginia opossums (Didelphis virginiana) were inoculated orally to 60 wild-caught Brown-headed Cowbirds (Molothrus ater). Another 30 Brown-headed Cowbirds were not challenged and served as uninfected controls. Two inoculated and one control cowbird were necropsied every 2 weeks and the pectoral and thigh muscles were examined grossly for cyst development. Stained histologic sections of pectoral muscle, thigh muscle, and lung were examined by light microscopy and presence, density, and size of sarcocysts were determined. Sarcocysts were present by 6 weeks post-inoculation (PI) and were still growing at 40 weeks PI. The sarcocysts from birds 40 weeks post-infection were infective to an opossum. The morphology of the sarcocyst wall by transmission electron microscopy substantiated the identification as S. falcatula. Lung sections were examined for the presence of schizonts, but were seen only at 2 weeks PI. This evaluation was complicated by the presence of unidentified microfilariae. These birds are migratory and the continued growth and development of muscle cysts would allow them to be a source of infection at both extremes of their geographic range, regardless of which end of the migration at which they were infected.     

Sarcocystis infection and myocardial pathological changes in cattle from south-eastern Norway

The incidence of myocardial sarcocystis infection and of myocardial pathological changes was recorded in samples of 79 healthy cattle obtained from an abattoir. The incidence rate of thin-walled cysts of S. cruzi was 81.0 %, while mixed infection with thick-walled cysts of S. hominis and/or S. hirsuta was found in 5.0 %. Focal interstitial myocarditis was found in 31.6 % of the samples. The sarcocystis infection and the interstitial mononuclear cell infiltrates were positively associated (P < 0.05). Intimai proliferations of musculo-elastic or fibro-elastic tissues in the intramural coronary arteries were found in 75.0 % of the cattle older than 3% years of age, and in 45.7 % of the cattle less than 3½ years old. No association of the arterial lesions and the sarcocystis infection was demonstrated.     

Ultrastructure of the cysts of Sarcocystis tarandivulpes from skeletal muscle of reindeer (Rangifer tarandus tarandus)

The cysts of S. tarandivulpes were found to be limited by a unit membrane which has been called the cyst membrane. The surface of the cysts was covered by closely packed and hexagonally arranged knob-like protrusions. The protrusions were 0.6–1.2 μm long and had an elliptical cross section. At the base of and between the bases of the protrusions the cyst mem brane was raised into low anastomosing folds which delineated shallow compartments. Between the folds the cyst membrane formed small vesicle-like invaginations into the cyst. On the apical part of the protrusions the cyst membrane had a smooth contour and was underlined by 2 layers of electron-dense material. Cyst ground substance divided the interior of the cyst into compartments containing either metrocytes or cystozoites. Cystozoites undergoing endodyogeny were present among the nondividing cystozoites. Some new terms were introduced to denote structures at the border of the cyst. The old terms are reviewed and the structural resemblance between S. tarandivulpes and S. odocoileocanis from Odocoileus virginianus is discussed.     

Ultrastructural and transmission evidence of Sarcocystis cruzi associated with eosinophilic myositis in cattle.

Skeletal muscle, diaphragm, tongue, esophagus and heart of beef carcasses that were condemned for eosinophilic myositis and those that were unaffected were collected at an abattoir in Colorado and studied to determine the involvement of Sarcocystis spp. All affected carcasses contained similar granulomatous lesions with adjacent infiltrations of leukocytes. Intact or fragments of sarcocysts were found within 32 of 363 granulomas, and whole sarcocysts were present in nearby unaffected muscle cells. Light and electron microscopic examinations revealed that sarcocysts, affected or unaffected by cellular response in condemned carcasses, as well as those found in unaffected carcasses, were consistent with those of S. cruzi. Transmission experiments confirmed that S. cruzi were present in all carcasses, and that dogs, but not cats, were the definitive hosts. The results of pepsin-HCl digestion assays showed that unaffected carcasses that were approved for human consumption generally contained more infective parasites than carcasses that were condemned for eosinophilic myositis. This study provides evidence to support the suggestion that dogs, rather than cats, and unaffected rather than eosinophilic myositis-affected carcasses, have greater potential for contributing to the perpetuation of eosinophilic myositis in the cattle industry.