Search

Sarcocystis bertrami (synonym: Sarcocystis fayeri) is a coccidian parasite that infects horses and donkeys in several countries. Dogs are known as definitive hosts of the parasite, however, the patent period is not well defined, and S. bertrami shed by dogs has never been confirmed by molecular methods. Here we investigated the shedding of S. bertrami by experimentally infected dogs and examined the excreted parasites by morphological and molecular tools. Three dogs of small breeds (one Yorkshire terrier and two miniature Pinschers) were acquired with ages of 30 and 60 days and were exclusively fed commercial dog food. Two dogs consumed equine muscle tissues containing cysts of S. bertrami. The third dog served as negative control and was simultaneously fed commercial dog food. The two animals that received equine tissues shed sporocysts and/or oocysts in their feces after prepatent periods of 13 and 23 days. The patent periods were 47 and 14 days. Sporocysts showed average dimensions of 14.19 µm (± 0.53) x 10.06 µm (± 0.44). The control dog did not shed sporocysts or oocysts of the parasite. Interestingly, patent periods had never been reported, and for one dog, the patent period (47 days) was longer than that reported for other Sarcocystidae parasites. PCRs to the gene 18S and to the internal transcribed spacer 1 (ITS1) of the rDNA were successfully performed with DNA extracted from sporocysts. ITS1 sequences were also obtained from the equine tissue cysts used to infect the dogs. Nucleotide sequences of cloned fragments of 18S from sporocysts, and ITS1 from both stages (tissue cysts and sporocysts) matched with S. bertrami (18S: 97.50-99.88 %; ITS1: 88.76-95.21 %), although high molecular diversity was observed with data from these loci. PCR to cox1 using sporocysts' DNA failed to amplify any product. The possibility of the existence of an additional and undescribed Sarcocystis species in the excreted sporocysts, besides S. bertrami, cannot be excluded from this experiment. To our knowledge, this is the first molecular confirmation of S. bertrami in canine feces. Sporocyst dimensions and prepatent periods observed in this study were similar to those previously described for S. bertrami and S. fayeri. In conclusion, the molecular, morphological and biological data generated here fit in previous descriptions for both S. bertrami and S. fayeri., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Vicuñas (Vicugna vicugna) are wild South American camelids (SACs) protected by law in Argentina, and information on pathogens that infect them is scarce. In this study, an adult vicuña found dead in the province of Salta was examined, and evidence of infection by Sarcocystis sp. protozoans was sought. Infection of skeletal muscles by S. aucheniae, with the production of macroscopic sarcocysts, a disease known as SAC sarcocystosis, has been described in the other three SACs - llamas, alpacas, and guanacos - but its occurrence in vicuñas has so far remained unknown. In the analyzed individual, many macroscopic cysts compatible with S. aucheniae were found upon necropsy in the muscular tissue of the neck and diaphragm. Analysis of 18 S rRNA and cytochrome c oxidase subunit 1 (cox-1) gene sequences by BLAST searches and construction of phylogenetic trees demonstrated that the etiological agent was S. aucheniae. Our results show for the first time that vicuñas act as intermediate hosts in the life cycle of this parasite. In addition, this study provides the first cox-1 sequences for S. aucheniae isolates from the four SAC species acting as intermediate hosts and suggests that this marker could be useful for genotypification of this parasite species. The impact of SAC sarcocystosis on the health, well-being, and fitness of vicuñas, and the relevance of vicuña infections in the epidemiology of S. auchaniae, remain to be elucidated., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Sarcocystis spp. are cyst-forming coccidia characterized by a two-host predator-prey life cycle. Sarcocysts are formed in muscles or nervous system of the intermediate host, while sporocysts develop in the small intestine of the definitive host. The intermediate hosts of Sarcocystis falcatula are wild birds. Colombia is one of the countries with the greatest biodiversity of birds, however, there are few studies related to this parasite in wild birds. This study presents the morphological and molecular detection of Sarcocystis falcatula collected from the emerald toucanet (Aulacorhynchus albivitta), a wild bird species endemic to South America. Pectoral muscle samples were obtained, and microscopic and molecular detection was performed by light microscopy, transmission electron microscopy, and amplifying of the first internal transcribed spacer (ITS-1) and surface antigen-encoding genes (SAGs). Sarcocystis measured an average of 161  × 42 μm, with a cyst wall ∼0.4 μm thick. Ultrastructurally, the sarcocyst wall type 11b-like consisted of numerous villar protrusions of 850 nm wide on average. The ITS-1 sequence showed 97.0-99.7% identity to S. falcatula previously described from birds in the United States and Brazil, respectively. Concatenated phylogenetic analysis based on SAG2, SAG3 and SAG4 confirmed that the new isolate is grouped with other sequences of Sarcocystis from South America, but divergent from those isolates obtained in North America. The results of this study demonstrate for the first time the presence of S. falcatula in a wild bird from Colombia., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Sarcocystis infection in sheep has caused significant economic losses in the livestock industry, and the genetic similarity among Sarcocystis species highlights the need for precise diagnostic methods in sheep. This study developed a loop-mediated isothermal amplification (LAMP) method targeting COX-1 and 28S rRNA genes to detect Sarcocystis tenella and Sarcocystis gigantea, respectively. The LAMP method exhibited high specificity, selectively amplifying target DNA sequences without cross-reactivity with closely related protozoa, such as Toxoplasma gondii and Neospora caninum. Detection limits were determined as 3 × 10 5 copies/L for S. tenella and 6 × 10 4 copies/L for S. gigantea, enabling sensitive identification of low-level infections. Comparative analysis with conventional PCR on sheep cardiac tissues demonstrated a higher LAMP detection rate (80.0% vs 66.7%). In conclusion, the LAMP method offers superior sensitivity to conventional PCR, allows visual confirmation of results, and provides a rapid diagnostic tool for identifying S. tenella and S. gigantea infection in sheep. However, due to the limitation of sample availability, we were unable to assess all Sarcocystis species that use sheep as intermediate hosts, which warrants further research., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Abstract: Sarcocystis infections were found for the first time in the muscles of 3 of 3 gray wolves (Canis lupus) from Minnesota. Two kinds (thin-walled and thick-walled) of sarcocysts were detected, based on the appearance of the sarcocyst wall. In wolf 1, sarcocysts were thin-walled (<0.5 μm), and without any visible protrusions. Ultrastructurally, the sarcocyst wall was type 1a and identical to Sarcocystis svanai of the domestic dog (Canis familiaris). The second kind of sarcocyst, with a relatively thicker (>1 μm) sarcocyst wall, was detected in wolves 2 and 3. Ultrastructurally, the sarcocyst wall had undulating, pleomorphic villar protrusion of type 9c; these sarcocysts were identical to Sarcocystis caninum from the domestic dog. Molecularly, the 2 Sarcocystis species were characterized using 18S, 28S, COI, ITS-1, and rpoB genetic markers. All these markers showed 100% identity to either of the 2 species previously described from the domestic dog. The thick-walled sarococyst corresponded to Sarcocystis caninum, whereas the thin-walled sarcocyst corresponded to Sarcocystis svanai., (© American Society of Parasitologists 2024.)

Several wild game meat species, including deer and feral pigs are hunted and consumed in Australia. Feral pigs and deer are not indigenous to Australia, but they have proliferated extensively and established their presence in every state and territory. Following the report of a sambar deer displaying Sarcocystis like white cysts in its rump muscles, the present study was conducted to explore the prevalence of Sarcocystis infections in wild deer and feral pigs in the southeastern regions of Australia. Oesophagus, diaphragm, and heart tissue from 90 deer and eight feral pigs were examined visually for sarcocysts. All results were negative. PCR testing of randomly selected deer and feral pigs yielded positive results, which were subsequently supported by histopathology. This is the first study to report the presence of Sarcocystis spp. in deer and feral pigs in Australia. As no visual cysts were found on the heart or oesophagus that came back positive with PCR, infected animals, particularly those reared free-range, could be passing through meat quality checks unidentified. If people consume this meat without cooking it properly, it may lead to a human infection of Sarcocystis. However, a more targeted study focused on determining the parasite's prevalence and assessing its risks is necessary to determine if it constitutes a food safety issue. As this species has been found not only in feral pigs but also in domestic pigs, the potential for infection spreading between feral pigs and pigs in free-range livestock systems is high, potentially posing a large problem for the Australian pork industry, particularly with the increased emphasis on free-range pig husbandry. Future studies should concentrate on determining the species of Sarcocystis in feral animals commonly consumed as game meat to determine potential zoonotic risks. This could also include a more in-depth look at the prevalence of Sarcocystis infections in other game animals. Identifying where these parasites are present and to what extent, are important areas for future studies., Competing Interests: Declaration of competing interest The authors have no affiliation with any organization with a direct or indirect financial interest in the subject matter discussed in the manuscript., (Copyright © 2024. Published by Elsevier B.V.)

Sarcocystis spp. cause pigeon protozoan encephalitis, a neuronal disease. A female pigeon exhibiting torticollis had a necrotic area in the cerebral hemisphere surrounded by lesions with perivascular cuffing, gliosis, granulomatous foci, and meningitis. Non-necrotic lesions were also observed in the brainstem. Intact and degenerative schizonts were observed within the neuropils and neurons in the lesions. Deoxyribonucleic acid (DNA) was extracted from paraffin-embedded brain tissues and genetically analyzed after gel electrophoresis to determine Sarcocystis spp. using specific primer sets for 28S ribosomal ribonucleic acid and internal transcribed spacer region-1. DNA sequencing confirmed a significant homology with S. calchasi. This is the first report of meningoencephalitis with malacia caused by S. calchasi in a rock pigeon in Japan.

Sarcocystis miescheriana infection is an important cause of carcass condemnation during meat inspection. The infection can cause morbidity and mortality in domestic pigs. In this study, an 8-month-old finisher pig was presented to a local abattoir for slaughter. Multiple white nodular lesions affecting the meat were observed, resulting in the condemnation of the carcass. Consequently, half of the carcass was submitted to the necropsy diagnostic laboratory in the School of Veterinary Medicine for further evaluation. Grossly, all superficial and deep muscle groups had severe multifocal macrocysts (3 mm × 2 mm × 1 mm) on the surface and extending deep into the skeletal musculature. Histopathology revealed moderate multifocal granulomatous and eosinophilic myositis with intralesional degenerated and intact parasites. Sample genomic DNA sequence analysis of the 18S RNA gene showed 100% identity to S. miescheriana in the GenBank. This is the first report of S. miescheriana in Grenada, West Indies., (© 2024 The Author(s). Veterinary Medicine and Science published by John Wiley & Sons Ltd.)

Muscles of 25 bobcats (Lynx rufus) from remote areas of Mississippi in 2017 were tested for parasites. Testing for Sarcocystis infections included microscopic examination of fresh unstained muscle squashes, pepsin digestion of hearts and tongues, and histological sections of paraffin-embedded tissues. Sarcocystis spp. infections were detected in the muscles of 21 (84%) by a combination of methods. Sarcocysts were detected in the unstained tongue squashes of 2 bobcats. Sarcocystis sp. bradyzoites were detected in the pepsin digests of 3 of 19 hearts, and 12 of 19 tongues. In paraffin-embedded histological sections, sarcocysts were detected in 7 of 25 hearts, 17 of 25 tongues, and 5 of 23 limb muscles. Based on the character of the cyst wall, at least 3 morphologic types of sarcocysts were detected: those with small spikes on the cyst wall, corresponding to Sarcocystis felis, those with long villar protrusions, corresponding to Sarcocystis neurona, and those lacking visible cyst wall protrusions, representing an unidentified type of sarcocyst. Myositis associated with sarcocysts was seen in the tongues of 3, and in the limb muscles of 1 bobcat. Multilocus genotyping of the DNA extracted from paraffin-embedded sections from 2 bobcats, employing 18S, 28S, COI, ITS-1, and 5.8S and rpoB genes, diagnosed Sarcocystis caninum, S. felis, Sarcocystis lutrae, and S. neurona. An encapsulated species of Trichinella was identified in the tongue of 1; it represents the first documented occurrences in bobcats from Mississippi. Taken together, these observations suggest intensive exposure of these wild carnivores to Trichinella tissue cysts, implies predation or scavenging on these tissues promotes parasite transmission, and raises caution concerning zoonotic risk when such meat is rendered for human consumption., (© American Society of Parasitologists 2024.)

Asexual replication in the apicomplexan Sarcocystis neurona involves two main developmental stages: the motile extracellular merozoite and the sessile intracellular schizont. Merozoites invade host cells and transform into schizonts that undergo replication via endopolygeny to form multiple (64) daughter merozoites that are invasive to new host cells. Given that the capabilities of the merozoite vary significantly from the schizont, the patterns of transcript levels throughout the asexual lifecycle were determined and compared in this study. RNA-Seq data were generated from extracellular merozoites and four intracellular schizont development time points. Of the 6,938 genes annotated in the S. neurona genome, 6,784 were identified in the transcriptome. Of these, 4,111 genes exhibited significant differential expression between the merozoite and at least one schizont development time point. Transcript levels were significantly higher for 2,338 genes in the merozoite and 1,773 genes in the schizont stages. Included in this list were genes encoding the secretory pathogenesis determinants (SPDs), which encompass the surface antigen and SAG-related sequence (SAG/SRS) and the secretory organelle proteins of the invasive zoite stage (micronemes, rhoptries, and dense granules). As anticipated, many of the S. neurona SPD gene transcripts were abundant in merozoites. However, several SPD transcripts were elevated in intracellular schizonts, suggesting roles unrelated to host cell invasion and the initial establishment of the intracellular niche. The hypothetical genes that are potentially unique to the genus Sarcocystis are of particular interest. Their conserved expression patterns are instructive for future investigations into the possible functions of these putative Sarcocystis -unique genes., Importance: The genus Sarcocystis is an expansive clade within the Apicomplexa, with the species S. neurona being an important cause of neurological disease in horses. Research to decipher the biology of S. neurona and its host-pathogen interactions can be enhanced by gene expression data. This study has identified conserved apicomplexan orthologs in S. neurona , putative Sarcocystis -unique genes, and gene transcripts abundant in the merozoite and schizont stages. Importantly, we have identified distinct clusters of genes with transcript levels peaking during different intracellular schizont development time points, reflecting active gene expression changes across endopolygeny. Each cluster also has subsets of transcripts with unknown functions, and investigation of these seemingly Sarcocystis -unique transcripts will provide insights into the interesting biology of this parasite genus., Competing Interests: The authors declare no conflict of interest.

The genus Sarcocystis includes protozoan parasites with an indirect life cycle. Sarcocystis spp. can infect various animal species and humans, causing sarcocystosis, a parasitosis of economic importance and zoonotic concern. Wild boars can act as intermediate hosts for Sarcocystis miescheriana and the zoonotic Sarcocystis suihominis that infects humans by consumption of raw or undercooked infected swine meat. In the present study, the diaphragmatic muscle tissue of 123 wild boars hunted in Greece was examined to determine the frequency of Sarcocystis spp. The samples were examined by tissue compression and molecular techniques. Under light microscopy, 34 out of 123 (27.6%) wild boars tested positive for Sarcocystis spp., while a higher infection prevalence (75%) was revealed by multiplex PCR performed in 100 of the samples. The partial mtDNA cox1 gene (~ 1100 bp) of 20 samples tested positive for S. miescheriana by multiplex PCR was amplified and sequenced. Sarcocystis miescheriana was identified as the only species involved in these infections. This is the first study on the prevalence of Sarcocystis spp. in wild animals in Greece. Further, large-scale surveys are needed to assess the prevalence and species of this parasite in Greece and to design efficient control and preventive measures in a One Health perspective., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

In the present study, tissue samples (tongue, esophagus and heart) were investigated from dromedary camels of India for identification and characterization of Sarcocystis spp. using histopathology, PCR and gene sequencing. Genomic DNA extracted from these tissue samples was used for PCR amplification of the cytochrome c oxidase subunit I gene (cox1) of Sarcocystis spp. and the partial sequence of small subunit ribosomal RNA (18S rRNA) gene of the S. cameli. The PCR products were purified, sequenced and analyzed using bioinformatics tools. Based on phylogenetic analysis of the cox1 gene, the sequences of the present study clustered with those of S. cameli, hosted by dromedary camels of Iraq and a close association was observed with S. masoni hosted by dogs and alpacas of China. Until now, there are no 18S rRNA sequences of S. cameli available in GenBank and this is the first study recording 18S rRNA sequences of S. cameli which were grouped with S. masoni from alpaca of China and guanaco and llama of Argentina in phylogenetic analysis. These findings could be useful for further studies on the characterization through molecular epidemiology, genetic diversity and host specificity of S. cameli., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

A senile male black capuchin monkey (Sapajus nigritus) kept under human care in a Zoo was found dead after 2 weeks presenting signals of weight loss and hyporexia. Histopathological revealed a necrotizing encephalitis. Although it was not observed microscopically, Sarcocystis sp infection was detected in brain tissue from molecular assays. These infections have been rarely described in neotropical primates, particularly associated with tissue lesions., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Currently, research on apicomplexan Sarcocystis parasites is mainly carried out by analyzing animal carcasses. However, environmental studies would not only allow faster detection of possible sources of infection but also avoid the use of animals for investigations. Therefore, in the current study, we aimed to identify tested Sarcocystis species in sediment collected from water bodies located in the southeastern Baltic countries. A total of 99 sediment samples were collected during the summer from different types of water bodies in Estonia, Latvia, Lithuania, and Poland. Species-specific nested PCR targeting cox1 gene was used for the detection of selected Sarcocystis species (S. cruzi, S. bovifelis, S. hirsuta, S. arieticanis, S. tenella, S. capracanis, S. miescheriana, and S. bertrami) infecting livestock. The results showed a statistically lower (p < 0.05) occurrence of Sarcocystis parasites in Estonia (50%) compared to three countries, where the detection rate of Sarcocystis spp. DNA was remarkably higher, ranging from 88 to 100%. Among Sarcocystis species tested, S. cruzi (83.8%) and S. arieticanis (55.6%) using cattle and sheep as their intermediate hosts were most commonly identified. The detection rates of some of the analyzed Sarcocystis species were significantly different in southeastern Baltic countries. It is discussed that the detection rates of certain Sarcocystis species depend not only on the number of animals per 1 km 2 but also on various ecological factors and farming practices that differ in the amount of contact domestic animals have with predators and the potential for animals to become infected through natural water or food sources., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Sarcocystis spp. are protozoan parasites that form cysts in the organs and musculature of various animal species. The species Sarcocystis miescheriana and Sarcocystis suihominis are pathogenic to pigs and wild boars (Sus scrofa), acting as intermediate hosts, while humans are the definitive host for S. suihominis. To date, there have been no reports of the identification of these coccidian species in Sus scrofa in Brazil. Therefore, in this study, we conducted the first molecular identification of Sarcocystis species using PCR-RFLP and sequencing. A total of 210 samples were analyzed, of this total, 67 tested positive for Sarcocystis spp., representing 31.9% of the total samples assessed. Out of the total positive samples, 55 (82.1%) were identified as S. miescheriana and 8 (11.9%) as S. suihominis, a zoonotic species. Additionally, other species related to bovines, such as S. cruzi and zoonotic S. hominis, were detected in 3.0% of the samples, serving as contaminants in the pork products. The presence of S. suihominis in swine and wild boar samples is concerning due to the zoonotic risk and potential environmental contamination, as humans act as definitive hosts, also for the presence of S. hominis as a bovine contaminant in pork sausages. Furthermore, we confirmed the efficacy of the PCR-RFLP technique as a reliable tool for the identification of Sarcocystis species, demonstrating its potential use in laboratories for molecular diagnosis and rapid identification of these parasites, aiming to protect public health and ensure food safety., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Sarcocystis spp. are apicomplexan cyst-forming parasites that can infect numerous vertebrates, including birds. Sarcosporidiosis infection was investigated in three muscles (breast, right and left thigh muscle) and one organ (heart) of four Razorbill auks (Alca torda) stranded between November and December 2022 on the shores of the Mediterranean Sea in Nabeul and Bizerte governorates, Northern Tunisia. Two of the four tested A. torda were PCR positive for 18S rRNA Sarcocystis spp. gene. Among the examined 16 muscles/organs, only one breast and one right thigh were Sarcocystis spp. PCR-positive (12.5% ± 8.3, 2/16). Our results showed a relatively high molecular prevalence of Sarcocystis spp. in Razorbill auks (A. torda). Sarcocystis spp. sequence described in the present study (GenBank number: OR516818) showed 99.56-100% identity to Sarcocystis falcatula. In conclusion, our results confirmed the infection of Razorbill auks (A. torda) by S. falcatula. Further research is needed on different migratory seabirds' species in order to identify other Sarcocystis species., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

A case of suspected food poisoning related to the consumption of raw meat from a common minke whale (Balaenoptera acutorostrata) was reported in Tokyo, Japan, in June 2020. Microscopic analysis revealed tissue cysts of Toxoplasma gondii and sarcocysts of Sarcocystis sp. in whale meat. The SAG2 and ITS1 region sequences of T. gondii were detected in the DNA extracted from the meat. Genotyping of the multilocus nested PCR-RFLP using the genetic markers SAG1, SAG2 (5'- SAG2, 3'-SAG2, and alt. SAG2), SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico revealed that the genotype of T. gondii was type II, with a type I pattern for the L358 locus. In the phylogenetic analyses of the six loci (GRA6, GRA7, SAG1, HP2, UPRT1, and UPRT7), these sequences clustered into haplogroup 2. Moreover, the sequences of the virulence-related genes ROP5 and ROP18 of T. gondii isolated from whale meat were similar to those of the type II ME49 reference strain. Sequence analyses of the mtDNA cox1 gene, 18S rRNA gene, and ITS1 region indicated the highest similarity of sarcocyst isolated from whale meat to Sarcocystis species that infect birds or carnivores as intermediate hosts; however, the species could not be identified. To our knowledge, this is the first report of T. gondii and Sarcocystis spp. being detected in same whale meat ingested by patients involved in a suspected food poisoning case in Japan., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023. Published by Elsevier B.V.)

Background: members of the genus Sarcocystis are intracellular obligate protozoan parasites classified within the phylum Apicomplexa and have an obligate heteroxenous life cycle involving two hosts. A more comprehensive understanding of the prevalence and geographic range of different Sarcocystis species in marine ecosystems is needed globally and nationally. Hence, the objective of this study was to document the incidence of Sarcocystis infection in sharks within the aquarium ecosystem of Egypt and to identify the species through the characterization of the SSU rDNA gene., Methods: All organs of the mako shark specimen underwent macroscopic screening to detect the existence of a Sarcocystis cyst. Ten cysts were collected from the intestine and processed separately to extract the genomic DNA. The polymerase chain reaction (PCR) was accomplished by amplifying a specific 18S ribosomal RNA (rRNA) gene fragment. Subsequently, the resulting amplicons were subjected to purification and sequencing processes., Results: Macroscopic examination of the mako shark intestinal wall sample revealed the presence of Sarcocystis cysts of various sizes and shapes, and sequencing of the amplicons from Sarcocystis DNA revealed a 100% nucleotide identity with the sequence of Sarcocystis tenella recorded from sheep in Iran; The mako shark sequence has been deposited in the GeneBank with the accession number OQ721979. This study presents the first scientific evidence demonstrating the presence of the Sarcocystis parasite in sharks, thereby documenting this specific marine species as a novel intermediate host in the Sarcocystis life cycle., Conclusions: This is the first identification of Sarcocystis infection in sharks, and we anticipate it will be an essential study for future screenings and establishing effective management measures for this disease in aquatic ecosystems., (© 2024. The Author(s).)

Purpose: Using molecular techniques, we have previously shown that carnivorous mammals of the family Mustelidae might be common definitive hosts for various protozoan Sarcocystis species. In the present study we aimed to unravel whether Sarcocystis species using ungulates as intermediate hosts and canids or felids as definitive hosts can be found in intestine of mustelids., Methods: Small intestine samples of 93 individual mustelids of five different species from Lithuania were examined. Sarcocystis species were identified based on species-specific PCR and subsequent cox1 sequencing., Results: Six Sarcocystis species (S. arieticanis, S. bertrami, S. capracanis, S. capreolicanis, S. linearis and S. morae) defined by ungulate-canid life cycle were detected for the first time in small intestines of mustelids. By contrast, the prevalence of Sarcocystis characterised by ungulate-felid life cycle was low (3.2%). Overall, 76% of the examined animals were positive for at least one of the studied Sarcocystis species. Four species, S. arieticanis, S. bertrami, S. capracanis and S. morae were most commonly found, with the detection rate of about 40%., Conclusions: The current finding, in addition to our previous studies, suggests that mustelids play an important role in the spread of various Sarcocystis species., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Sarcocystis are Apicomplexan protozoa with a dixenous life cycle that includes a predator and a prey as definitive and intermediate hosts, respectively. Domestic and wild pigs are intermediate hosts of S. suihominis, with formation of sarcocysts in their muscles, while humans and non-human primates act as final hosts. After ingesting raw or undercooked sarcocyst-infested pork, signs of gastroenteritis including inappetence, nausea, vomiting, and diarrhea may develop in humans. Moreover, excretion of infective forms with human feces leads to dissemination of the parasite in the environment. In this study, macroscopic sarcocysts of white color, oval shape, and a diameter of approximately 3-8 mm were found in the skeletal muscle of a slaughtered domestic pig (Sus scrofa domesticus) destined for human consumption in an abattoir of Makurdi, Benue State, Nigeria. Sarcocyst DNA was used as template to PCR amplify the near-complete length of the 18S rRNA gene and a fragment of the cytochrome c oxidase subunit 1 (cox-1) gene. Amplicons were sequenced and used to construct phylogenetic trees with selected available Sarcocystis spp. sequences. In both cases, the placement of the analyzed sequences with S. suihominis was strongly supported, confirming the species identity of this macroscopic sarcocyst-forming parasite. This constitutes the first molecular identification of S. suihominis in Nigeria and the African continent. Proximity between pigs and humans, and poor sanitary conditions frequently encountered in pig farms of Nigeria might favor the dissemination of this zoonotic parasite, posing a threat to public health., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The epidemiological picture of Taenia saginata infections in Kenya is fragmented with limited available data. Although Sarcocystis species are significant meat-borne parasites, few studies have explored their occurrence in Kenya. This study aimed to estimate the occurrence of bovine cysticercosis and screen for the presence of Sarcocystis spp. A meat inspection-based survey was conducted in ten abattoirs in Narok County, Kenya, and inspection for T. saginata cysticerci was limited to the Triceps brachii muscle. The apparent occurrence of the parasite was 5.4% (95% CI, 3.8, 7.6, n =573). Molecular confirmation of T. saginata was done via nested polymerase chain reaction targeting the mitochondrial 12S ribosomal RNA gene and restricted fragment length polymorphism. Sarcocystis species were identified using a multiplex polymerase chain reaction method targeting the 18S ribosomal RNA gene sequences and the mitochondrial cytochrome c oxidase subunit I gene. Of the 31 cystic lesions tested, 26/31 (83.9%) were confirmed to be T. saginata. Sarcocystis cruzi and S. hominis were detected in 8/31 (25.8%) and 1/31 (3.2%) of the cystic lesions, respectively. Co-infections of S. cruzi and T. saginata were found in 6/31 lesions (19.4%). The confirmation of bovine cysticercosis and S. hominis is suggestive of the presence of risky culinary and sanitation practices that facilitate transmission. This is the first report and molecular confirmation of Sarcocystis spp. in cattle in the country. The presence of both zoonotic S. hominis and pathogenic S. cruzi highlights an underexplored concern of veterinary and human health significance, warranting further epidemiological investigation.

Recently, the wild deer population has been increasing in Japan, causing serious feeding-related damage to the agricultural and forestry industries. In conjunction with the government's promotion of hunting for population control, the effective utilization of resources and promotion of the game meat industry as a sixth sector of industrialization are desired by local governments. However, several cases in which patients showed intestinal symptoms such as diarrhea due to the consumption of sika deer meat infected with protozoan Sarcocystis spp. have been reported, and the pathogenic microorganisms found in wild deer should be investigated. In this study, Sarcocystis sp. parasitized Kyushu sika deer ( Cervus nippon nippon ) in Nagasaki Prefecture, Japan, was examined for its enterotoxicity. A phylogenetic analysis based on the sequence of the 18S rRNA gene and cox1 showed that the species was highly homologous to Sarcocystis japonica and/or Sarcocystis sp. HM050622. We attempted to confirm the diarrhea-evoking toxicity of Sarcocystis sp. in sika deer meat, which has been previously reported in human case reports. A mouse ileal loop assay showed that Sarcocystis sp. in sika deer meat induced significant fluid accumulation in the loop at doses of ∼5 × 10 6 bradyzoites. Western blotting showed that these Sarcocystis parasites possess actin-depolymerizing factor, a diarrhea-evoking factor, similar to Sarcocystis fayeri , which exists in horsemeat. However, the pathogenic conditions of the ileal loop were different from those of similar experiments with S. fayeri . This study suggests that S. japonica parasitizing C. n. nippon may cause diarrhea via a different mechanism from that of S. fayeri .

At least three Sarcocystis species (S. falcatula, S. halieti and S. wobeseri-like) have been detected infecting raptorial birds. By histopathology and PCR-sequencing of the ITS1 marker, S. halieti was detected in a bearded vulture (Gypaetus barbatus) and a black kite (Milvus migrans) from the Catalonia region in North Spain. The 241 bp-long sequences obtained from the Sarcocystis organisms detected in both raptors showed 97.5-99.6% and 97.9-100% similarity with those of previously identified S. halieti; also, the phylogenetic trees generated placed the identified sequences together with other sequences of S. halieti available in GenBank. In sum, the description of the bearded vulture as a new intermediate host for S. halieti adds new insights on the complex epidemiology of the genus involving avian hosts., (© 2023. The Author(s).)

Horses are intermediate hosts of Sarcocystis spp. capable of forming cysts in their musculature. This study aimed to detect sarcocysts and investigate the presence of nucleic acids from Sarcocystis spp. in samples of striated muscles from horses in the State of Rio Grande do Sul, Brazil, necropsied at the Veterinary Pathology Laboratory of the Federal University of Santa Maria. A total of 108 samples were collected from 24 horses and examined through direct examination. Microscopic tissue cysts were observed in three samples: tongue (2) and esophagus (1) from two animals. Extractions were performed on the found cysts and tissues, even though sarcocystosis detection was not present. DNA samples were subjected to Nested-PCR using Tg18s primers, and the amplified products were subjected to Restriction Fragment Length Polymorphism (RFLP) using DdeI and HpaII enzymes. DNA belonging to Sarcocystis spp. was amplified in tissues from 91.7% (22/24) of the equines, and 67.6% (73/108) of the samples tested positive in the Nested-PCR reaction. The tissues with the highest detection frequency were: diaphragm 92.3% (12/13), gluteal muscle 77.2% (17/22), and esophagus 66.7% (4/6). In RFLP, Sarcocystis spp. was detected in 21 tissues from 11/22 equines, and cysts, identified through nucleotide sequencing, were determined to be S. bertrami. S. neurona was identified in 11 samples from 7/22 animals, with co-infection detected in 5/22 cases. The high detection rate indicates a concerning circulation of the protozoan, particularly the zoonotic S. bertrami found in all tissues, which are commonly exported for human consumption., Competing Interests: Declaration of Competing Interest The authors have no financial or proprietary interests in any material discussed in this article., (Copyright © 2023. Published by Elsevier B.V.)

Background: Sarcocystis species are obligatorily heteroxenous protozoan parasites with predator-prey life cycles. Global Knowledge about the epidemiology and the distribution pattern of different Sarcocystis species in dog feces are very scarce. Therefore, the current investigation was conducted to declare the occurrence of Sarcocystis in the fecal specimens of the most common canids in Egypt, the domestic dogs, and to identify the species present using various parasitological and molecular approaches., Methods: A total of 100 dog fecal samples were collected and screened using fecal sugar flotation test for the presence of Sarcocystis oocysts/sporocysts. Additionally, thirty samples were used for genomic DNA extraction. The 18S rRNA gene fragment was the target of primers for a PCR, followed by purification and sequencing of the amplicons., Results: Currently, the results obtained reviewed that 4% of fecal samples were positive for Sarcocystis spp. using LM. Additionally, Sarcocystis spp. were verified in sixteen dogs (53.3%, 16/30) using PCR and subsequent sequencing protocols. Statistically, insignificant difference in prevalence of sarcocystosis relative to age and gender was noticed. Morphologically, the detected sporocysts measured 13.2-16.0 × 9.4-11 μm. Based on the 18S rRNA gene, sequencing analysis of amplicons from sporocysts DNA revealed 99.82% nucleotide homology with published S. tenella partial nucleotide sequences from sheep in Iraq and Iran., Conclusions: This is the first molecular evidence in support of the final host role of domestic dogs in the life cycle of S. tenella in Egypt, which provides a precious diagnostic tool for further epidemiological studies and for the assessment of the effectiveness of control measures for this disease., (© 2023. The Author(s).)

In this study, for the first time, Sarcocystis species were identified molecularly in sika deer (Cervus nippon) that form free-ranging populations in several countries of Europe. Diaphragm muscle samples from 151 deer from 10 populations in Germany and Austria were examined for sarcocysts. By one-gram methylene-blue staining, sarcocysts were recorded in samples of 114 animals (75.5%) which originated from all populations. Sarcocysts were more often (p < 0.0001) recorded in yearling and adult deer than in calves. Infection intensity was generally low with ~ 70% of the sarcocyst positive deer harbouring ≤ 10 sarcocysts per 1-gram diaphragm muscle. Based on cox1 sequence comparison, 10 species of Sarcocystis, all previously reported parasitizing cervids, were identified: S. elongata, S. entzerothi, S. hjorti, S. iberica, S. japonica, S. linearis, S. morae, S. pilosa, S. silva and S. truncata. The prevailing S. hjorti was detected in sika deer of all 10 populations. The identification in sika deer of S. hjorti, S. iberica, S. elongata, S. linearis, S. morae and S. silva constitutes new host records. With the additional species records of this study, the highest number of Sarcocystis species, at least 16, was identified in this host., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Different food-safety institutions, including the European Food Safety Authority, encourage monitoring and characterising Sarcocystis spp. in animals and foodstuffs; among meat-producing animals, domestic pigs (Sus scrofa domesticus) can host two different Sarcocystis spp., that is Sarcocystis miescheriana and the zoonotic Sarcocystis suihominis. Herein, we report for the first time the presence of macrocysts of Sarcocystis miescheriana in a domestic pig resulting in carcass condemnation. In North-West Italy, in June 2022 the carcass of a clinically healthy sow was condemned due to the detection of multifocal macroscopic whitish fusiform lesions. Affected muscle samples were submitted to histological and molecular analyses targeting the mtDNA cox1 and 18S rRNA genes. At gross examination and histology, well demarcated, oval or elongated macrocysts up to 8 mm in length characterized by a calcified central core surrounded by fibrosis were detected. The molecular amplification and sequencing of the cox1 mtDNA and 18S rRNA genes revealed the presence of Sarcocystis miescheriana DNA in all sampled macrocysts. Our study provides the first molecularly confirmed case of Sarcocystis miescheriana infection in a domestic pig in Italy. The present report highlights the need to increase data related to the occurrence and the prevalence of Sarcocystis spp. in meat-producing animals, and in wild and domestic pigs in particular, taking into account the zoonotic potential of Sarcocystis suihominis and the possible financial losses related to carcass discard due to macroscopic Sarcocystis spp. cysts., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

The increase of wild boar populations density and their meat consumption across Europe could expose humans to a plethora of foodborne diseases as sarcocystosis, caused by the zoonotic protozoan Sarcocystis suihominis. Humans become infected by eating raw or undercooked pig (Sus scrofa domesticus) containing S. suihominis sarcocysts. Despite this, to date very few data are available on the risk of infection by this parasite to wild boar (Sus scrofa) meat consumers. Thus, the present study aimed to assess the occurrence of Sarcocystis spp. in wild boars from southern Italy, applying both histology and a new multiplex PCR assay targeting the cox1 gene. Between 2019 and 2020, 997 muscle tissues (i.e., n = 269 oesophagus, n = 277 diaphragms, n = 298 hearts, n = 153 tongues) from 311 wild boars were collected and screened by a combined histological and molecular approach. Overall, 251 (80.7%) animals tested were positive for Sarcocystis spp., and S. miescheriana whose definitive hosts are canids, was the only molecularly identified species. A statistically significant difference (p < 0.05) in the prevalence of Sarcocystis infection was found according to the wild boar age and muscle tissue. Findings outlined the low zoonotic potential of infection to humans via wild boar meat consumption in Italy and the importance of the application of new molecular methods in distinguishing different Sarcocystis species., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023. Published by Elsevier Ltd.)

Sarcocystis spp. and Toxoplasma gondii are two apicomplexan protozoa that infect a broad range of vertebrates, however, little is known about the infection of equids with these parasites. A total of 184 slaughtered equids from slaughterhouses of Bizerte and Tunis located in Northern Tunisia, were examined for meat infections with Sarcocystis spp. and T. gondii by PCR. The prevalence of infections with Sarcocystis spp. and T. gondii were 38% (95% CI: 31-45) and 39.7% (95% CI: 32.6-46.7), respectively. The highest prevalence of infection with Sarcocystis spp. was observed in donkeys (48.6%; 95% CI: 37.3-60) followed by mules (32.8%; 95%CI: 21.3-44.3), and horses (28.3%; 95% CI: 15.2-41.2) (P = .04). Similarly, the highest prevalence of infection with T. gondii was also observed in donkeys (66.2%; 95% CI: 55.4-77), followed by mules (18.7%; 95%CI: 9.2-28.3), and horses (26.1%; 95%CI: 13.4-38.8) (P < .001). The coinfection prevalence was estimated to be 17.4% (95%CI: 11.9-22.9). Taking into consideration that humans can be infected following consumption of infected equid meat with T. gondii and/or some Sarcocystis species, it is important to assess the risk of human infection. Thus, further studies are needed to better understand the epidemiology of these zoonoses., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Sarcocystis spp. infects water buffaloes (Bubalus bubalis) causing sarcocystosis. In the present study, Sarcocystis fusiformis was recognized in Egyptian water buffaloes based on histological observation and molecular analysis of internal transcribed spacer 1 (ITS1), 18S ribosomal RNA (18S rRNA) and cytochrome c oxidase subunit I (COX-1) gene fragments. Chemotherapy and vaccines against Sarcocystis spp. could potentially target proteases because they may play a crucial role in the infection. Cysteine proteases are multifunctional enzymes involved in vital metabolic processes. However, the involvement of proteases in S. fusiform infection has not yet been characterized. Here, the purification and study on some biochemical properties of protease isolated from cysts of S. fusiform were carried out. Protease with a molecular weight of 100 kDa was purified. LC-MS/MS analyzed the protein sequence of purified protease and the data suggested that the enzyme might be related to the cysteine protease. The purified protease exhibited maximum activity at pH 6 and a temperature of 50 °C. The Michaelis-Menten constant (K m ), the maximum velocity (V max ), and the turnover number (K cat ) were determined. The complete inhibition effect of cysteine inhibitors indicated that the purified enzyme is a cysteine protease. The results suggested that S. fusiform proteolytic enzyme may be necessary for parasite survival in water buffaloes by digesting host tissues. Therefore, cysteine protease could be a suitable target for vaccinations., (© 2023. The Author(s).)

Background: Equine protozoal myeloencephalitis (EPM) caused by Sarcocystis neurona remains an antemortem diagnostic challenge in some horses. Recent work suggested the use of real-time PCR (rtPCR) on cerebrospinal fluid (CSF) as a promising diagnostic tool., Objective: To evaluate the sensitivity and specificity of S. neurona rtPCR on CSF for EPM diagnosis using horses with EPM and S. neurona-seropositive horses with other neurologic conditions., Animals: Ninety-nine horses with neurologic disease that underwent complete neurologic examination, CSF collection, and, if euthanized, necropsy including the central nervous system (CNS)., Methods: Retrospective case-control study using banked CSF samples. Samples from horses with neurologic abnormalities and necropsy-confirmed EPM diagnosis, presumptive EPM diagnosis using strict criteria (SnSAG2/4/3 ELISA serum:CSF titer ratios <50) and horses diagnosed with other neurologic diseases were used., Results: Fifty-two horses had EPM; 23 were confirmed on necropsy, and 29 were presumptive clinical diagnoses. The other 47 horses all had necropsy-confirmed diagnoses. Four of the 47 horses had normal neurologic findings on necropsy and the remaining 43 horses had neurologic diseases including equine degenerative myeloencephalopathy (EDM), cervical vertebral stenotic myelopathy, trauma, and other miscellaneous conditions. One CSF sample was weakly positive for S. neurona by rtPCR, this sample was obtained from a horse with confirmed EDM. Samples from the other 98 horses were negative for S. neurona by rtPCR., Conclusions and Clinical Importance: Our study contradicts previous conclusions that S. neurona rtPCR is potentially useful for EPM diagnosis, because our results indicate that the assay has a low sensitivity (0%) for EPM., (© 2023 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals LLC on behalf of American College of Veterinary Internal Medicine.)

The intake of food and water containing the Sarcocystis parasite has been linked to a number of outbreaks worldwide, including Malaysia. Nevertheless, the lack of surveys and epidemiological data on Sarcocystis infections in Malaysia makes it difficult to estimate its occurrence in humans and animals. A cross-sectional study was conducted to determine the prevalence of Sarcocystis and the risk factors associated with infection among village chickens and pigs reared under different farm managements in Peninsular Malaysia. Phylogenetic trees were constructed using partial fragments of the 18S rRNA gene and ITS1 sequences. In the present study, 680 sera samples were collected from village chickens (n=250) and commercial pigs (n=433) and anti-Sarcocystis antibodies were screened using the enzymelinked immunosorbent assays (ELISA) kit. At the animal level, the prevalence of Sarcocystis was 9.2% (95% CI: 5.92-13.48) and at the farm level, it was 64.0% (95% CI: 42.52-82.03) in village chickens. The animal-level seroprevalence of Sarcocystis for pigs was 3.7% (95% CI: 2.13-5.93) and 36.8% (95% CI: 16.29-61.64) at the farm-level. Polymerase Chain Reaction (PCR) was conducted on meat samples from various parts of village chickens (n=250) consisting of brain, heart, lung, and pectoralis muscle tissues, and pork (n=121) consisting of intercostal muscle, diaphragm, and tongue. Sarcocystis DNA was detected in 6.4% (95% CI: 4.60-11.60) of village chicken samples but zero in pork samples. A total of 11 unique Sarcocystis haplotypes were isolated from these tissue samples. Multivariable logistic regression analysis of the putative risk factors showed a statistically significant association between Sarcocystis infection in pigs and uncovered storage of feed. Although no zoonotic Sarcocystis was isolated in this study, we reported the first discovery of S. wenzeli in Malaysia.

Background: Sarcocystis is an intracellular parasite of particular importance as it infects many domestic animals as camels that play the role of intermediate host for the parasite., Aim: This study aimed to identify Sarcocystis species in camels by molecular assay with confirmation of local isolates by phylogenetic analysis., Methods: A total of 200 slaughtered camels ( Camelus dromedarius ) that were slaughtered in Al-Najaf province (Iraq) abattoirs from October (2021) to July (2022) were subjected to collect the fresh tissues from four organs (esophagus, diaphragm, skeletal muscle, and heart), to be tested later by the conventional polymerase chain reaction (PCR). Then, a total of 20 positive genomic DNA samples were sequenced, named, got specific access numbers (OP785703.1 to OP785722.1), and compared with the NCBI-GenBank isolates., Results: Targeting Cox1 gene, 80% of collected tissues were found positive by the conventional PCR assay. Phylogenetic tree analysis revealed that the local Sarcocystis isolates were identical to Indian S. cameli isolates at 99.70%-99.90%. Significantly, an increase in Sarcocystis infection was seen in the esophagus compared to the diaphragm, skeletal muscle, and heart; older (>4 years) than younger (≤4 years) camels, and in females more than males., Conclusion: To the best of our knowledge, this represents the first molecular study in Iraq that identifies Sarcocystis cameli in camels. However, additional epidemiological and molecular studies in camel populations as well as in other domestic and wild animals appeared to be necessary., Competing Interests: The authors declare that there is no conflict of interest.

The aim of this study was to determine the presence of deoxyribonucleic acid (DNA) from Toxoplasma gondii, Sarcocystis spp. and Neospora caninum, in tissues of wild boars slaughtered in southern Brazil. A total of 156 samples were collected from different organs of 25 wild boars, and DNA from at least one of the protozoa investigated was detected in 79 samples. To differentiate between infectious agents, restriction fragment length polymorphism was performed using the restriction enzymes DdeI and HpaII. For N. caninum, conventional PCR was performed with specific primers. The DNA of at least one of the studied pathogens was detected in each animal: 26.58% for T. gondii, 68.36% for Sarcocystis spp. and 5.06% for N. caninum. Coinfection between T. gondii and Sarcocystis spp. occurred in 14 animals, between T. gondii and N. caninum in only one male animal, between Sarcocystis spp. and N. caninum in a female, while co-infection with the three agents was equally observed in only one male animal. Considering the high frequency of detection and its zoonotic risk, especially T. gondii, it appears that wild boars can be potential sources of transmission of infectious agents and the adoption of monitoring measures in these populations should be prioritized.

Infections by Sarcocystis in cattle are ubiquitous worldwide. There is considerable debate concerning the identity of Sarcocystis spp. in cattle. Proper diagnosis of Sarcocystis spp. is important to assess their economic and public health importance. Currently there are seven named species: Sarcocystis hirsuta, Sarcocystis cruzi, Sarcocystis hominis, Sarcocystis bovifelis, arcocystis heydorni, Sarcocystis bovini and Sarcocystis rommeli. Additionally, there are unnamed Sarcocystis spp. Two species, S. hominis and S. heydorni, are zoonotic. One out of seven species (S. hirsuta, contracted from cats) forms macroscopic cysts which can be visible during carcass inspection. Current molecular characterization is based on DNA extracted from sarcocysts from naturally infected cattle because DNA was not characterized from tissues of experimentally infected cattle or feces of experimentally infected definitive hosts. Sarcocystis cruzi (transmitted via canids) is recognized as the most pathogenic species and it causes abortion, low milk yield, poor body growth, and outbreaks of clinical sarcocystosis and death. Additionally, Sarcocystis infections have been linked to an inflammatory condition of striated muscles termed bovine eosinophilic myositis (BEM). Cattle affected by BEM appear clinically normal. Diagnosis of BEM at slaughter occurs when inspecting the carcass surface, or once the carcass has been divided into prime cuts or quarters. Sex and breed have no apparent influence on prevalence of BEM. The condition evidently occurs with equal frequency in steers, cows, and heifers. Virtually all striated muscles can be affected including skeletal muscles, the muscles of the eye, larynx, and the heart. In the USA, regulations require condemnation of BEM-affected parts, or (in severe cases) the entire carcass. These aesthetic considerations result in economic losses. Cattle experimentally infected with Sarcocystis did not have BEM at slaughter. Here, we review the status of Sarcocystis spp. and BEM in cattle including prevalence, lesions, epidemiology, and association of BEM with different species of Sarcocystis., (Published by Elsevier Ltd.)

South American opossums (Didelphis spp.) are definitive hosts of Sarcocystis neurona, Sarcocystis speeri, Sarcocystis lindsayi and Sarcocystis falcatula. In Brazil, diverse studies have demonstrated a high frequency of Sarcocystis falcatula-like in sporocysts derived from opossums, and high genetic diversity has been observed in surface antigen-encoding genes (SAGs). In this study, genetic diversity of Sarcocystis spp. derived from Didelphis albiventris and Didelphis aurita from the cities of Campo Grande and São Paulo, was accessed by sequencing SAG2, SAG3, SAG4, the first internal transcribed spacer (ITS-1) and cytochrome c oxidase subunit I (cox1). Molecular identification was performed for 16 DNA samples obtained from sporocyst or culture-derived merozoites. The ITS-1, cox1, and SAG3 fragments were cloned, whereas SAG2 and SAG4 were sequenced directly from PCR products. Four alleles variants were found for SAG2, 13 for SAG3 and seven for SAG4, from which four, 13 and four, respectively, were novel. Twenty-seven allele variants were found for ITS-1, all phylogenetically related to S. falcatula-like previously described in Brazil. Sarcocystis sp. phylogenetically related to Sarcocystis rileyi was evidenced by cox1 in three opossums. Further studies are needed to clarify the role of Didelphis spp. as definitive hosts of Sarcocystis spp. other than that previous described.

Background: Limited data are currently available on protozoan parasites of the genus Sarcocystis that infect their avian hosts within the order Anseriformes (waterfowl). To date, no Sarcocystis species has been recorded in ducks in China., Methods: Leg muscles were sampled from 26 domestic ducks (Anas platyrhynchos) in China in 2021. Morphological characteristics of sarcocysts detected in the muscle tissue were described using light microscopy (LM) and transmission electron microscopy (TEM). Genomic DNA was extracted from single sarcocysts obtained from different ducks, and three genetic markers, 18S ribosomal DNA (18S rDNA), 28S ribosomal DNA (28S rDNA) and mitochondrial (mt) cytochrome oxidase subunit 1 (cox1), were amplified and cloned for sequence analyses., Results: Sarcocysts were observed by LM in only three of the 28 samples (10.7%). These sarcocysts had a thick cyst wall with numerous brush-like villar protrusions (vps) of 3.8-4.3 μm in length (n = 30) on the cyst surface. TEM observation showed that the sarcocysts had lanceolated vps. Each vps narrowed in the stalk and contained a bundle of microtubules that extended into the ground substance. Comparisons of the new sequences with those deposited in GenBank showed that the most similar sequences were those of Sarcocystis halieti in the great cormorant Phalacrocorax carbo and European starling Sturnus vulgaris, and Sarcocystis calchasi in the domestic pigeon (Columba livia) at the 18S rDNA (99.1% identity); Sarcocystis wenzeli from the domestic chicken Gallus gallus at the 28S rDNA (95.9-96.0% identity); and Sarcocystis speeri from the opossum at the mtcox1 (98.2% identity). The new 18S rDNA, 28S rDNA and mitochondrial cox1 sequences shared up to 99.0%, 95.6% and 97.7% identity, respectively, with those of Sarcocystis spp. obtained from Anseriformes avian hosts. Phylogenetic analysis inferred from the sequences of the three genetic markers placed the organism within a group of Sarcocystis spp. obtained from avian or carnivorous intermediate hosts and avian, marsupial or carnivorous definitive hosts. Based on the morphological observation and molecular analyses, the organism found in the Chinese domestic ducks was regarded as a new species and named Sarcocystis platyrhynchosi n. sp., Conclusions: Based on morphology and sequence analyses, the microcysts diagnosed in the domestic ducks examined in this study were named as a new species. This is the first record of Sarcocystis spp. from waterfowl in China. Sarcocysts of similar morphology occur frequently in different Anseriformes birds, and the relationships among these species need to be further clarified in future studies using more molecular markers., (© 2023. The Author(s).)

Pigeons are common birds around the world and may act as intermediate hosts of the tissue cyst-forming apicomplexan parasites Toxoplasma gondii, Neospora caninum and Sacrocystis spp. This study aimed to provide an overview on the prevalence of and exposure to these parasites in Iranian domestic rock pigeon (Columba livia domestica) through molecular, serological and histopathological examination. Blood and tissue samples (i.e., brain, heart, gizzard, neck, thigh, and pectoral muscles) were taken from 100 pigeons. Sera were screened by agglutination tests for detection of anti- T. gondii and N. caninum antibodies, genomic DNA from tissue samples were assessed by respective species-specific PCRs, and histopathological examination of tissues was carried out. A seroprevalence of 45 % to anti-T. gondii and 35 % to anti-N. caninum IgG was recorded. PCR detected T. gondii DNA in 28 pigeons. Sacrocystis spp. was detected in one animal, but sequencing of the 28 S rRNA gene product did not reveal the identity of the species. Histopathology revealed myocarditis, myositis, and gliosis in the heart, skeletal muscles, and brain, respectively. No Sarcocystis tissue-cysts were detected, but T. gondii tissue cyst-like structures in the brain (i.e., 4 %) and heart (i.e., 3 %) were found by histology. Data reported herein demonstrate that pigeons from Iran are infected with tissue cyst-forming apicomplexans, particularly T. gondii. Since domestic pigeons are in close contact with human populations, and consumption of their meat and egg is popular in different societies, control strategies for minimizing the risk of infection in both pigeons and humans are suggested., Competing Interests: Conflict of interest The authors declare that they have no competing interests., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Sarcocystis cruzi is a member of the genus Sarcocystis, infecting bovine animals such as cattle and bison as intermediate hosts, and canids such as dogs and raccoon dogs as definitive hosts. Acute sarcocystosis of S. cruzi causes occasional symptoms in cattle, including weight loss, reduced milk production, abortions, and death, and similar to other Sarcocystis species can potentially cause food poisoning in humans when raw or undercooked infected cattle meat is consumed. Despite these issues, genetic information on S. cruzi is scarce, and there is no specific quantitative method for the detection and quantification of the parasite in infected cattle. In this study, we aimed to develop a method based on high-throughput sequencing of S. cruzi genome and transcriptome that specifically and quantitatively detects the S. cruzi acetyl-CoA synthetase gene (ScACS). Cardiac muscles were collected from slaughterhouses in Saitama Prefecture to obtain sarcocysts from which DNA and RNA were extracted for the high-throughput sequencing. Using the sequences, we developed a specific quantitative PCR assay which could distinguish S. cruzi ACS from that of Toxoplasma gondii by taking advantage of the differences in their exon/intron organizations and validated the assay with the microscopic counting of the S. cruzi bradyzoites. Thus, this assay will be useful for future studies of S. cruzi pathogenesis in cattle and for the surveillance of infected animals, thereby easing public health concerns.

Despite its frequent presence in buffaloes, Sarcocystis buffalonis remains as one of the most under studied parasite. In the present study, isolates of S. buffalonis from,Mathura, Uttar Pradesh India were characterized for 18S rRNA (MF595842-MF595844), cox 1 (MG792800-MG792802), 28S rRNA (MH793418-MH793420) and ITS 1 (MH793421-MH793423) genes. Analysis revealed multiple haplotypes for each individual gene viz., 18S rRNA (three haplotypes), cox 1 (two haplotypes), 28S rRNA (two haplotypes) and ITS 1 (single haplotype). The studied Indian sequences showed variable homologies for individual gene loci viz., 18S rRNA (99.3-99.9%); cox 1 (99.8-100.0%); 28S rRNA (99.9-100.0%) and ITS 1 (100.0%) The phylogenetic association between S. buffalonis and closely related Sarcocystis spp. infecting buffaloes and cattle was delineated. All these gene loci placed S. buffalonis close to S. hirsuta. The study has generated A vital phylogenetic data about this erstwhile neglected parasite., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Several cases of gastrointestinal symptoms including diarrhea and vomiting due to the consumption of Sarcocystis -infected venison have been reported in Japan. However, the control of case incidence is difficult, as epidemiological information concerning Sarcocystis in venison in Japan is insufficient. We examined the prevalence and parasite load of Sarcocystis in 89 samples of Yezo-deer ( Cervus nippon yesoensis ) venison in Hokkaido by quantifying the copy numbers of the 18S rRNA gene of Sarcocystis , followed by a statistical analysis that considered the sampling area, age, and sex to clarify the parameters related to the parasite load. The copy numbers per gram of venison in samples ranged from 4.8 to 8.8 log. Wilcoxon rank-sum test, the one-way factorial analysis of variance (ANOVA), Steel-Dwass test, and a two-way factorial ANOVA showed significant differences in the copy numbers among sampling areas, not by age or sex, suggesting that the load of Sarcocystis in wild deer depended on the sampling area in Hokkaido. Notably, more than 80% of Hokkaido venison has a higher gene copy number than the meat that caused Sarcocystis fayeri -food poisoning. This information is expected to contribute to the establishment of hygiene standards for safe venison consumption and the control of gastrointestinal symptom cases due to consumption of Sarcocystis -infected venison.

The seroprevalence of Sarcocystis spp. and Toxoplasma gondii was researched in swine raised in Santa Maria, RS, Brazil. Serum samples from 84 pigs from 31 farms were tested using indirect immunofluorescence assay (IFA) for both agents. Additionally, 53 samples of pork sausages and tissues destined for human consumption, including: salami, sausage, black pudding, heart, tongue, brain, and rib muscle, were submitted to PCR to detect DNA for each agent. The frequency of anti-Sarcocystis spp. antibodies was 36.9% (31/84), with titers ranging from 32 to 1024, and 25% (21/84) for anti-T. gondii antibodies, with titers ranging from 64 to 2048. Sarcocystis spp. and T. gondii DNA were detected in 67.9% (36/53) and 13.2% (7/53) of samples, respectively. The presence of antibodies and the detection of DNA from Sarcocystis spp., and T. gondii suggests that the pigs were infected and may serve as an important reservoir for both parasites. The infection by these protozoa in the swine population is relevant to public health due to their zoonotic potential.

Background: Sarcocystis species are diverse apicomplexan parasites, though only two zoonotic species (S. hominis and S. heydorni) circulate between cattle and humans. Due to the importance of cattle in the human food chain and to prevent the consequences of parasitosis in humans, the first global systematic review and meta-analysis on molecular epidemiology, species distribution, and zoonotic significance of Sarcocystis infection in cattle was performed., Methods: For this aim, four international English databases (PubMed, Scopus, Google Scholar, and Web of Science) were systematically searched till 20th September 2021, and random-effect models were drawn to calculate total estimates and their 95% confidence intervals (CIs)., Results: Finally, 44 papers from 21 countries were qualified for this review which examined 8526 cattle regarding Sarcocystis infection, rendering a total prevalence of 62.7% (95% CI 53-71.5%). Globally, 12 Sarcocystis spp. have been reported from cattle, including S. cruzi, S. hominis, S. hirsuta, S. rommeli, S. heydorni, S. bovifelis, S. bovini, S. sinensis, S. gigantea, S. fusiformis, S. hjorti and S. tenella. Among them, S. cruzi (37 studies), S. hominis (22 studies) and S. hirsuta (19 studies) were the 3 most common species, with 76.4% (95% CI 64.8-85%), 30.2% (95% CI 19.3-44%) and 8.7% (95% CI 3.8-18.6%), respectively. However, molecular identification was not performed in 48.4% (95% CI 27.3-70.1%) of the positive samples., Conclusion: Despite the zoonotic significance of Sarcocystis spp., particularly S. hominis, the epidemiology and distribution of Sarcocystis infection in cattle remains unclear and demands more extensive researches around the world., (© 2022. The Author(s) under exclusive licence to Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Eosinophilic myositis in bovine striated muscle thought to be caused by a hypersensitivity reaction to the degradation of Sarcocystis tissue cysts, is a rare reason for carcase condemnation in the United Kingdom. This paper describes the identification of Sarcocystis cruzi associated with lesions of generalised eosinophilic myositis in three English beef carcases, by gross and histopathological examination followed by PCR with subsequent sequencing. Samples from two unaffected animals were also examined. Although sarcocystosis caused by S.cruzi is not considered a public health risk, the clinically affected carcases were deemed unfit for human consumption due to the extensive lesions affecting meat quality. We believe this to be the first report from the UK describing the molecular-based identification of Sarcocystis cruzi in meat affected and unaffected with eosinophilic myositis., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Sika deer (Cervus nippon), which are native to the Japanese islands and the adjacent mainland of eastern Asia, have been introduced into Europe and established free-ranging populations in several countries. Various Sarcocystis species have been identified recently from farmed "mainland sika" deer in Lithuania and native "Japanese sika" deer in Japan. To study the distribution, prevalence and intensity of Sarcocystis infection in free-ranging sika deer outside of their natural range heart and/or diaphragm samples of 311 animals from nine populations in Germany and Austria were examined by histology.Overall, sarcocysts were detected in either heart or diaphragm of 107/311 deer (34.4%) with prevalence ranging roughly from 5 to 50% among the populations. Considering the 263 animals with both heart and diaphragm available, prevalence varied significantly (p < 0.0001) among calves (20.2%), yearlings (40.3%), and adult deer (49.1%) but did not differ between male and female deer (48.3% vs. 43.7%; p = 0.6483). Occurrence of sarcocysts in heart vs. diaphragm indicated a marginal difference (27.8% vs. 20.9%; p = 0.0839). Intensity of infection in all but one heart positive and all diaphragm positive animals was low (< 10 sarcocysts per square centimeter muscle cut). While heart sarcocyst counts of yearlings and adult deer were higher than those of calves and were higher in male compared to female sika deer, diaphragm sarcocyst counts did not differ significantly between age groups and sexes. Sarcocystis infection was demonstrated at variable prevalence in sika deer in all populations but intensity is apparently low. Further studies are needed to identify the species of Sarcocystis infecting sika deer naturalized outside their natural range., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Protozoans of genus Sarcocystis are widespread parasites infecting mammals, birds, and reptiles. Morphology of their sarcocysts is an important criterion for species identification. However, as more and more morphologically similar Sarcocystis species are being found and described, additional methods for their routine diagnostics are needed. We investigated restriction fragment length polymorphism (RFLP) as potential alternative to sequencing data analysis for the identification of Sarcocystis species using birds as an intermediate host. The internal transcribed spacer 1 (ITS1) region sequences of seventeen Sarcocystis species (S. albifronsi, S. anasi, S. calchasi, S. columbae, S. cornixi, S. corvusi, S. cristata, S. halieti, S. falcatula, S. fulicae, S. kutkienae, S. lari, S. lindsayi, S. rileyi, S. turdusi, S.wenzeli, and S. wobeseri) of interest were analysed and five best-fitting endonucleases generating most informative restriction fragments were selected for routine testing. In general, RFLP analyses are always inconclusive as they target very short DNA sequences. However, it can be an irreplaceable technique when fast and cheap identification and discrimination of known species are required, which was our main goal and preliminary results indicate that RFLP could be successfully used when identifying closely related avian Sarcocystis species with just two nucleases., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Two captive vulturine guineafowl ( Acryllium vulturinum ) were presented with lethargy, hyporexia, weight loss, and progressive neurologic signs. One of the guineafowl was seropositive for Sarcocystis falcatula (1:50 dilution). Both guineafowl died within 5 d of presentation. Histologic examination revealed nonsuppurative meningoencephalitis with gliosis, associated with occasional schizonts in the neuropil. Using fresh-frozen brain tissue, PCR was performed to amplify the ITS1 RNA region and portions of the 18S ribosomal RNA gene (18S gene) and the 28S ribosomal RNA gene (28S gene). Analysis of nucleic acid sequences from the resulting amplicons indicated that Sarcocystis calchasi was the likely cause of disease. To our knowledge, S. calchasi -associated disease has not been reported previously in the order Galliformes .

Infections with the Apicomplexa Toxoplasma gondii, Neospora caninum, and Sarcocystis spp. are common causes of reproductive disorders in sheep. However, few epidemiological studies regarding co-infections with these three protozoa are reported in sheep in Italy. For this reason, this study aims to evaluate possible co-infections with T. gondii, N. caninum, and Sarcocystis spp. in sheep slaughtered for human consumption. From April to July 2019, individual blood, brain, heart, and diaphragm samples were collected from 138 sheep after slaughtering. The presence of IgG anti-Toxoplasma in serum samples was evaluated through ELISA. DNA of the three protozoa was investigated using specific PCRs. Co-infection with T. gondii, N. caninum and Sarcocystis spp. was found in 66.7% of the examined sheep. Antibodies against T. gondii were found in the 36.2% of serum samples. The presence of T. gondii DNA was detected in the 67.4%, 77.5%, and 21.7% of the brain, heart, and diaphragm samples, respectively. Neospora caninum DNA was found in 72.5% of the examined brain samples. Sarcocystis spp. DNA was detected in 92% and 52.2% of the heart and diaphragm samples, respectively. Sequence analysis of the Sarcocystis spp. revealed the sole presence of Sarcocystis tenella. The present study demonstrates that sheep have a high risk of infection with the three Apicomplexa investigated, suggesting the need to adopt adequate measures to prevent the spread of these parasitic infections considering their clinical and economic impact on ovine production. Furthermore, the possible role sheep play in the zoonotic transmission of toxoplasmosis to humans was highlighted., (© 2022. The Author(s).)

The closely related apicomplexa protozoa, Toxoplasma gondii, Neospora spp., and Sarcocystis neurona, have all been associated with neurological and reproductive diseases in horses. However, there is limited data regarding the presence of these three parasites in equine placental tissues and amniotic fluid. The aim of the present report was to investigate the presence of the DNA of T. gondii, Neospora spp. and S. neurona in placentas and amniotic fluid in mares. Anti-T. gondii, anti- S. neurona and anti- Neospora spp. antibody titers were evaluated in 31 mares in the final third of pregnancy by indirect fluorescent antibody test (IFAT). The presence of parasite DNA in placentas and amniotic fluid was evaluated by polymerase chain reaction (PCR), using two target loci (ITS1 and Nc5). No antibodies to were identified nor was any T. gondii DNA detected in any mare. Antibodies to Neospora spp. were found in 6 mares (19.35 %) and DNA from this protozoan was detected in four placentas (12.9 %) and in five amniotic fluid samples (16.6 %). Antibodies to Sarcocystis spp. were detected in nine mares and S. neurona DNA was found in only one placenta (3.23 %). Our results suggest that the transplacental route may be a potential source of Neospora caninum infection in mares. Further studies are needed to understand the role of transplacental transmission in the epidemiology of these protozoa., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Data mining animal of genomes has been used before to identify endoparasites, and may be a particularly useful tool to surpass some difficulties faced by studies in the marine environment. We detected a species of Sarcocystis Lankester, 1882, contamination in the sperm whale (Physeter catodon Linnaeus) reference genome available in the GenBank database. We identified and extracted multiple gene fragments and placed the sequences in a phylogenetic framework. Our results indicate that the sequences of Sarcocystis sp. found in the genome do not correspond to any currently described species, despite a few other similar sequences having been identified in fur seals (Pinnipedia) and another sperm whale. Including data from previous studies, we suggest there is enough evidence to support the occurrence of at least four species of Sarcocystis in marine mammals. We also demonstrate that the term "S. canis-like" has been used for samples not closely related to Sarcocystis canis Dubey et Speer, 1991.