Serologic responses of cats against experimental Sarcocystis neurona infections

Sarcocystis neurona is the most important cause of a neurologic disease of horses, equine protozoal myeloencephalitis (EPM). Cats and other carnivores can act as its intermediate hosts and horses are aberrant hosts. Little is known of the sero-epidemiology of S. neurona infections in cats. In the present study, antibodies to S. neurona were evaluated by the S. neurona agglutination test (SAT). Cats fed sporocysts from the feces of naturally infected opossums or inoculated intramuscularly with S. neurona merozoites developed high levels (> or =1:4000) of SAT antibodies. Antibodies to S. neurona were not found in a cat inoculated with merozoites of the closely related parasite, Sarcocystis falcatula. These results should be useful in studying sero-epidemiology of S. neurona infections in cats.     

Experimental inoculation of domestic cats (Felis domesticus) with Sarcocystis neurona or S. neurona-like merozoites

Sarcocystis neurona is the parasite most commonly associated with equine protozoal myeloencephalitis (EPM). Recently, cats (Felis domesticus) have been demonstrated to be an experimental intermediate host in the life cycle of S. neurona. This study was performed to determine if cats experimentally inoculated with culture-derived S. neurona merozoites develop tissue sarcocysts infectious to opossums (Didelphis virginiana), the definitive host of S. neurona. Four cats were inoculated with S. neurona or S. neurona-like merozoites and all developed antibodies reacting to S. neurona merozoite antigens, but tissue sarcocysts were detected in only two cats. Muscle tissues from the experimentally inoculated cats with and without detectable sarcocysts were fed to laboratory-reared opossums. Sporocysts were detected in gastrointestinal (GI) scrapings of one opossum fed experimentally infected feline tissues. The study results suggest that cats can develop tissue cysts following inoculation with culture-derived Sarcocystis sp. merozoites in which the particular isolate was originally derived from a naturally infected cat with tissue sarcocysts. This is in contrast to cats which did not develop tissue cysts when inoculated with S. neurona merozoites originally derived from a horse with EPM. These results indicate present biological differences between the culture-derived merozoites of two Sarcocystis isolates, Sn-UCD 1 and Sn-Mucat 2.     

Utilization of stress in the development of an equine model for equine protozoal myeloencephalitis

Neurologic disease in horses caused by Sarcocystis neurona is difficult to diagnose, treat, or prevent, due to the lack of knowledge about the pathogenesis of the disease. This in turn is confounded by the lack of a reliable equine model of equine protozoal myeloencephalitis (EPM). Epidemiologic studies have implicated stress as a risk factor for this disease, thus, the role of transport stress was evaluated for incorporation into an equine model for EPM. Sporocysts from feral opossums were bioassayed in interferon-gamma gene knockout (KO) mice to determine minimum number of viable S. neurona sporocysts in the inoculum. A minimum of 80,000 viable S. neurona sporocysts were fed to each of the nine horses. A total of 12 S. neurona antibody negative horses were divided into four groups (1-4). Three horses (group 1) were fed sporocysts on the day of arrival at the study site, three horses were fed sporocysts 14 days after acclimatization (group 2), three horses were given sporocysts and dexamethasone 14 days after acclimatization (group 3) and three horses were controls (group 4). All horses fed sporocysts in the study developed antibodies to S. neurona in serum and cerebrospinal fluid (CSF) and developed clinical signs of neurologic disease. The most severe clinical signs were in horses in group 1 subjected to transport stress. The least severe neurologic signs were in horses treated with dexamethasone (group 3). Clinical signs improved in four horses from two treatment groups by the time of euthanasia (group 1, day 44; group 3, day 47). Post-mortem examinations, and tissues that were collected for light microscopy, immunohistochemistry, tissue cultures, and bioassay in KO mice, revealed no direct evidence of S. neurona infection. However, there were lesions compatible with S. neurona infection in horses. The results of this investigation suggest that stress can play a role in the pathogenesis of EPM. There is also evidence to suggest that horses in nature may clear the organism routinely, which may explain the relatively high number of normal horses with CSF antibodies to S. neurona compared to the prevalence of EPM.     

Direct agglutination test for the detection of antibodies to Sarcocystis neurona in experimentally infected animals

Equine protozoal myeloencephalitis (EPM) is a serious neurological disease of horses in the Americas. The apicomplexan protozoan most commonly associated with EPM is Sarcocystis neurona. A direct agglutination test (SAT) was developed to detect antibodies to S. neurona in experimentally infected animals. Merozoites of the SN6 strain of S. neurona collected from cell culture were used as antigen and 2-mercaptoethanol was added to the antigen suspension to destroy IgM antibodies when mixed with test sera. Mice fed sporocysts of S. speeri or S. falcatula-like sporocysts from opossums did not seroconvert in the SAT. The sensitivity of the SAT was 100% and the specificity was 90% in mice.     

Prevalence of Sarcocystis neurona sporocysts in opossums (Didelphis virginiana) from rural Mississippi

Sarcocystis species sporocysts were found in intestinal scrapings from 24 of 72 opossums (Didelphis virginiana) from rural Mississippi. The number of sporocysts in each opossum varied from a few ( < 100000) to 187 million. Sporocysts from 24 opossums were bioassayed for Sarcocystis neurona infections by feeding to gamma-interferon knockout (KO) mice. S. neurona was detected in the brains of KO mice fed sporocysts from 19 opossums by immunohistochemical staining with anti-S. neurona specific polyclonal rabbit serum, and by in vitro culture from the brains of KO mice fed sporocysts. The isolates of S. neurona from opossums were designated SN16-OP to SN34-OP. Merozoites from 17 of 19 isolates tested at the 25/396 locus were identical to previously described S. neurona isolates from horses. The high prevalence of S. neurona sparocysts in D. virginiana suggests that this opossum constitutes an ample reservoir of infection in the southern United States.     

Sporocyst size of isolates of Sarcocystis shed by the Virginia opossum (Didelphis virginiana)

The Virginia opossum (Didelphis virginiana) is a definitive host for multiple Sarcocystis species including Sarcocystis neurona, one of the causative agents of equine protozoal myeloencephalitis (EPM), a severe, neuromuscular disease of horses. Size and morphologic characteristics of isolates of Sarcocystis shed by the opossum were examined to determine if differences were useful in discriminating between the isolates and/or species. Collections of sporocysts from 17 opossums were molecularly characterized and measured using an ocular micrometer. The mean sporocyst size of isolates of S. neurona was 10.7 microm x 7.0 microm, Sarcocystis falcatula 11.0 microm x 7.1 microm, Sarcocystis speeri 12.2 microm x 8.8 microm, 1085-like isolate 10.9 microm x 6.8 microm, and 3344-like isolate 19.4 microm x 10.5 microm. The length and width of S. speeri were statistically different (p < 0.05) from the sporocysts of other types. The length of S. neurona and S. falcatula sporocysts were statistically different (p < 0.05) from each other and the width of S. falcatula and 1085 differed (p < 0.05). The fifth sporocyst type (3344) was observed, but due to pronounced morphological characteristics, statistical analysis was not performed. There was no consistent difference between the taxa based on internal structure of the sporocyst.     

Evidence to support horses as natural intermediate hosts for Sarcocystis neurona

Opossums (Didelphis spp.) are the definitive host for the protozoan parasite Sarcocystis neurona, the causative agent of equine protozoal myeloencephalitis (EPM). Opossums shed sporocysts in feces that can be ingested by true intermediate hosts (cats, raccoons, skunks, armadillos and sea otters). Horses acquire the parasite by ingestion of feed or water contaminated by opossum feces. However, horses have been classified as aberrant intermediate hosts because the terminal asexual sarcocyst stage that is required for transmission to the definitive host has not been found in their tissues despite extensive efforts to search for them [Dubey, J.P., Lindsay, D.S., Saville, W.J., Reed, S.M., Granstrom, D.E., Speer, C.A., 2001b. A review of Sarcocystis neurona and equine protozoal myeloencephalitis (EPM). Vet. Parasitol. 95, 89-131]. In a 4-month-old filly with neurological disease consistent with EPM, we demonstrate schizonts in the brain and spinal cord and mature sarcocysts in the tongue and skeletal muscle, both with genetic and morphological characteristics of S. neurona. The histological and electron microscopic morphology of the schizonts and sarcocysts were identical to published features of S. neurona [Stanek, J.F., Dubey, J.P., Oglesbee, M.J., Reed, S.M., Lindsay, D.S., Capitini, L.A., Njoku, C.J., Vittitow, K.L., Saville, W.J., 2002. Life cycle of Sarcocystis neurona in its natural intermediate host, the raccoon, Procyon lotor. J. Parasitol. 88, 1151-1158]. DNA from schizonts and sarcocysts from this horse produced Sarcocystis specific 334bp PCR products [Tanhauser, S.M., Yowell, C.A., Cutler, T.J., Greiner, E.C., MacKay, R.J., Dame, J.B., 1999. Multiple DNA markers differentiate Sarcocystis neurona and Sarcocystis falcatula. J. Parasitol. 85, 221-228]. Restriction fragment length polymorphism (RFLP) analysis of these PCR products showed banding patterns characteristic of S. neurona. Sequencing, alignment and comparison of both schizont and sarcocyst DNA amplicons showed 100% identity. Although Koch's postulates have not been demonstrated in this case study, the finding of mature, intact S. neurona schizonts and sarcocysts in the tissues of this single horse strongly suggests that horses have the potential to act as intermediate hosts. Further studies are needed to demonstrate Koch's postulates with repeated transfer of S. neurona between opossums and horses.     

Risk factors associated with the presence of Sarcocystis neurona sporocysts in opossums (Didelphis virginiana).

Sarcocystis neurona is the most important cause of equine protozoal myeloencephalitis (EPM) in horse in the Americas. The only known definitive host for this parasite in the United States is the opossum (Didelphis virginiana); however, despite the importance of the disease, the epidemiology of the parasite in the definitive host is poorly understood. To begin addressing these data gaps, potential risk factors were evaluated for their association with the presence of sporocysts of S. neurona in opossums live-trapped in March 1999 and November 1999 to May 2000. Sporocysts of S. neurona were found in 19 of the 72 animals examined. Potential risk factors evaluated were locality, trap date, age, gender, the presence of young in the pouch of females, and body condition score. Variables that were associated with the presence of S. neurona sporocysts were used in logistic regression analysis. Of the factors examined, season and body condition score were associated with increased odds of an animal harboring sporocysts.     

Immunoconversion against Sarcocystis neurona in normal and dexamethasone-treated horses challenged with S. neurona sporocysts

Equine protozoal myeloencephalitis is a common neurologic disease of horses in the Americas usually caused by Sarcocystis neurona. To date, the disease has not been induced in horses using characterized sporocysts from Didelphis virginiana, the definitive host. S. neurona sporocysts from 15 naturally infected opossums were fed to horses seronegative for antibodies against S. neurona. Eight horses were given 5x10(5) sporocysts daily for 7 days. Horses were examined for abnormal clinical signs, and blood and cerebrospinal fluid were harvested at intervals for 90 days after the first day of challenge and analyzed both qualitatively (western blot) and quantitatively (anti-17kDa) for anti-S. neurona IgG. Four of the challenged horses were given dexamethasone (0.1mg/kg orally once daily) for the duration of the experiment. All challenged horses immunoconverted against S. neurona in blood within 32 days of challenge and in CSF within 61 days. There was a trend (P = 0.057) for horses given dexamethasone to immunoconvert earlier than horses that were not immunosuppressed. Anti-17kDa was detected in the CSF of all challenged horses by day 61. This response was statistically greater at day 32 in horses given dexamethasone. Control horses remained seronegative throughout the period in which all challenged horses converted. One control horse immunoconverted in blood at day 75 and in CSF at day 89. Signs of neurologic disease were mild to equivocal in challenged horses. Horses given dexamethasone had more severe signs of limb weakness than did horses not given dexamethasone; however, we could not determine whether these signs were due to spinal cord disease or to effects of systemic illness. At necropsy, mild-moderate multifocal gliosis and neurophagia were found histologically in the spinal cords of 7/8 challenged horses. No organisms were seen either in routinely processed sections or by immunohistochemistry. Although neurologic disease comparable to naturally occurring equine protozoal myeloencephalitis (EPM) was not produced, we had clear evidence of an immune response to challenge both systemically and in the CNS. Broad immunosuppression with dexamethasone did not increase the severity of histologic changes in the CNS of challenged horses. Future work must focus on defining the factors that govern progression of inapparent S. neurona infection to EPM.     

Comparison of Sarcocystis neurona isolates derived from horse neural tissue

Sarcocystis neurona is a protozoan parasite that can cause neurological deficits in infected horses. The route of transmission is by fecal-oral transfer of sporocysts from opossums. However, the species identity and the lifecycle are not completely known. In this study, Sarcocystis merozoites from eight isolates obtained from Michigan horses were compared to S. neurona from a California horse (UCD1), Sarcocystis from a grackle (Cornell), and five Sarcocystis isolates from feral opossums from Michigan.Comparisons were made using several techniques. SDS-PAGE analysis with silver staining showed that Sarcocystis spp. from the eight horses appeared the same, but different from the grackle isolate. One Michigan horse isolate (MIH6) had two bands at 72 and 25kDa that were more prominent than the UCD1 isolate and other Michigan horse isolates. Western blot analysis showed that merozoites of eight of eight equine-derived isolates, and the UCD1 S. neurona isolate had similar bands when developed with serum or CSF of an infected horse. Major bands were seen at 60, 44, 30, and 16kDa. In the grackle (Cornell) isolate, bands were seen at 60, 44, 29, and 16kDa. DNA from merozoites of each of the eight equine-derived isolates and the grackle-derived isolate produced a 334bp PCR product (Tanhauser et al., 1999). Restriction fragment length polymorphism (RFLP) analysis of these horse isolates showed banding patterns characteristic for S. neurona. The grackle (Cornell) isolate had an RFLP banding pattern characteristic of other S. falcatula species. Finally, electron microscopy examining multiple merozoites of each of these eight horse isolates showed similar morphology, which differed from the grackle (Cornell) isolate. We conclude that the eight Michigan horse isolates are S. neurona species and the grackle isolate is an S. falcatula species.     

A review of Sarcocystis neurona and equine protozoal myeloencephalitis (EPM)

Equine protozoal myeloencephalitis (EPM) is a serious neurological disease of horses in the Americas. The protozoan most commonly associated with EPM is Sarcocystis neurona. The complete life cycle of S. neurona is unknown, including its natural intermediate host that harbors its sarcocyst. Opossums (Didelphis virginiana, Didelphis albiventris) are its definitive hosts. Horses are considered its aberrant hosts because only schizonts and merozoites (no sarcocysts) are found in horses. EPM-like disease occurs in a variety of mammals including cats, mink, raccoons, skunks, Pacific harbor seals, ponies, and Southern sea otters. Cats can act as an experimental intermediate host harboring the sarcocyst stage after ingesting sporocysts. This paper reviews information on the history, structure, life cycle, biology, pathogenesis, induction of disease in animals, clinical signs, diagnosis, pathology, epidemiology, and treatment of EPM caused by S. neurona.     

Brown-headed cowbirds (Molothrus ater) harbor Sarcocystis neurona and act as intermediate hosts

We tested the hypothesis that brown-headed cowbirds (Molothrus ater) harbor Sarcocystis neurona, the agent of equine protozoal myeloencephalitis (EPM), and act as intermediate hosts for this parasite. In summer 1999, wild caught brown-headed cowbirds were collected and necropsied to determine infection rate with Sarcocystis spp. by macroscopic inspection. Seven of 381 (1.8%) birds had grossly visible sarcocysts in leg muscles with none in breast muscles. Histopathology revealed two classes of sarcocysts in leg muscles, thin-walled and thick-walled suggesting two species. Electron microscopy showed that thick-walled cysts had characteristics of S. falcatula and thin-walled cysts had characteristics of S. neurona. Thereafter, several experiments were conducted to confirm that cowbirds had viable S. neurona that could be transmitted to an intermediate host and cause disease. Specific-pathogen-free opossums fed cowbird leg muscle that was enriched for muscle either with or without visible sarcocysts all shed high numbers of sporocysts by 4 weeks after infection, while the control opossum fed cowbird breast muscle was negative. These sporocysts were apparently of two size classes, 11.4+/-0.7 microm by 7.6+/-0.4 microm (n=25) and 12.6+/-0.6 microm by 8.0+/-0 microm (n=25). When these sporocysts were excysted and introduced into equine dermal cell tissue culture, schizogony occurred, most merozoites survived and replicated long term and merozoites sampled from the cultures with long-term growth were indistinguishable from known S. neurona isolates. A cowbird Sarcocystis isolate, Michigan Cowbird 1 (MICB1), derived from thin-walled sarcocysts from cowbirds that was passaged in SPF opossums and tissue culture went on to produce neurological disease in IFNgamma knockout mice indistinguishable from that of the positive control inoculated with S. neurona. This, together with the knowledge that S. falcatula does not cause lesions in IFNgamma knockout mice, showed that cowbird leg muscles had a Sarcocystis that fulfills the first aim of Koch's postulates to produce disease similar to S. neurona. Two molecular assays provided further support that both S. neurona and S. falcatula were present in cowbird leg muscles. In a blinded study, PCR-RFLP of RAPD-derived DNA designed to discriminate between S. neurona and S. falcatula showed that fresh sporocysts from the opossum feeding trial had both Sarcocystis species. Visible, thick-walled sarcocysts from cowbird leg muscle were positive for S. falcatula but not S. neurona; thin-walled sarcocysts typed as S. neurona. In 1999, DNA was extracted from leg muscles of 100 wild caught cowbirds and subjected to a PCR targeting an S. neurona specific sequence of the small subunit ribosomal RNA (SSU rRNA) gene. In control spiking experiments, this assay detected DNA from 10 S. neurona merozoites in 0.5g of muscle. In the 1999 experiment, 23 of 79 (29.1%) individual cowbird leg muscle samples were positive by this S. neurona-specific PCR. Finally, in June of 2000, 265 cowbird leg muscle samples were tested by histopathology for the presence of thick- and thin-walled sarcocysts. Seven percent (18/265) had only thick-walled sarcocysts, 0.8% (2/265) had only thin-walled sarcocysts and 1.9% (5/265) had both. The other half of these leg muscles when tested by PCR-RFLP of RAPD-derived DNA and SSU rRNA PCR showed a good correlation with histopathological results and the two molecular typing methods concurred; 9.8% (26/265) of cowbirds had sarcocysts in muscle, 7.9% (21/265) had S. falcatula sarcocysts, 1.1% (3/265) had S. neurona sarcocysts, and 0.8% (2/265) had both. These results show that some cowbirds have S. neurona as well as S. falcatula in their leg muscles and can act as intermediate hosts for both parasites.     

Parasitemia in an immunocompetent horse experimentally challenged with Sarcocystis neurona sporocysts

Equine protozoal myeloencephalitis (EPM) is a serious neurological disease of horses in Americans. Most cases are attributed to infection of the central nervous system with Sarcocystis neurona. Parasitemia has not been demonstrated in immunocompetent horses, but has been documented in one immunocompromised foal. The objective of this study was to isolate viable S. neurona from the blood of immunocompetent horses. Horses used in this study received orally administered S. neurona sporocysts (strain SN 37-R) daily for 112 days at the following doses: 100/day for 28 days, followed by 500/day for 28 days, followed by 1000/day for 56 days. On day 98 of the study, six yearling colts were selected for attempted culture of S. neurona from blood, two testing positive, two testing suspect and two testing negative for antibodies against S. neurona on day 84 of the study. Two 10 ml tubes with EDTA were filled from each horse by jugular venipuncture and the plasma fraction rich in mononuclear cells was pipetted onto confluent equine dermal cell cultures. The cultures were monitored weekly for parasite growth for 12 weeks. Merozoites grown from cultures were harvested and tested using S. neurona-specific PCR with RFLP to confirm species identity. PCR products were sequenced and compared to known strains of S. neurona. After 38 days of in vitro incubation, one cell culture from a horse testing positive for antibodies against S. neurona was positive for parasite growth while the five remaining cultures remained negative for parasite growth for all 12 weeks. The Sarcocystis isolate recovered from cell culture was confirmed to be S. neurona by PCR with RFLP. Gene sequence analysis revealed that the isolate was identical to the challenge strain SN-37R and differed from two known strains UCD1 and MIH1. To our knowledge this is the first report of parasitemia with S. neurona in an immunocompetent horse.     

Inoculation of Sarcocystis neurona merozoites into the central nervous system of horses

Equine protozoal myeloencephalitis (EPM) is a neurologic syndrome in horses from the Americas and is usually caused by infection with the apicomplexan parasite, Sarcocystis neurona. A horse model of EPM is needed to test the efficacy of chemotherapeutic agents and potential vaccines. Five horses that were negative for antibodies to S. neurona in their serum and cerebrospinal fluid (CSF) were injected in the subarachnoid space with living merozoites of the SN2 isolate of S. neurona. None of the horses developed clinical disease or died over a 132-day observation period. All five horses developed antibodies to S. neurona in their CSF and serum 3-4 weeks after injection. Two of the horses were examined at necropsy and no parasite induced lesions were observed in their tissues and no parasites were recovered from portions of their spinal cords inoculated on to cell cultures. Results of this study demonstrate that merozoites of the SN2 isolate of S. neurona will induce seroconversion but not clinical disease when inoculated directly into the CSF of nonimmune horses.     

Development and evaluation of a Sarcocystis neurona-specific IgM capture enzyme-linked immunosorbent assay

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease of horses caused primarily by the protozoal parasite Sarcocystis neurona. Currently available antemortem diagnostic testing has low specificity. The hypothesis of this study was that serum and cerebrospinal fluid (CSF) of horses experimentally challenged with S neurona would have an increased S neurona-specific IgM (Sn-IgM) concentration after infection, as determined by an IgM capture enzyme linked immunoassay (ELISA). The ELISA was based on the S neurona low molecular weight protein SNUCD-1 antigen and the monoclonal antibody 2G5 labeled with horseradish peroxidase. The test was evaluated using serum and CSF from 12 horses experimentally infected with 1.5 million S neurona sporocysts and 16 horses experimentally infected with varying doses (100 to 100,000) of S neurona sporocysts, for which results of histopathologic examination of the central nervous system were available. For horses challenged with 1.5 million sporocysts, there was a significant increase in serum Sn-IgM concentrations compared with values before infection at weeks 2-6 after inoculation (P < .0001). For horses inoculated with lower doses of S neurona, there were significant increases in serum Sn-IgM concentration at various points in time after inoculation, depending on the challenge dose (P < .01). In addition, there was a significant increase between the CSF Sn-IgM concentrations before and after inoculation (P < .0001). These results support further evaluation of the assay as a diagnostic test during the acute phase of EPM.     

Reactivity against Sarcocystis neurona and Neospora by serum antibodies in healthy French horses from two farms with previous equine protozoal myeloencephalitis-like cases

Sarcocystis neurona is considered a leading cause of equine protozoal myeloencephalitis (EPM), a common infectious neurological disease in horses in the Americas. EPM-like cases associated with S. neurona peptide reactive antibodies in Western blots were recently described in Normandy, France. In this report, antibodies reacting with S. neurona merozoites were detected using an agglutination assay at titers ranging from 50 to 500 in sera from 18/50 healthy horses from two farms with a previous EPM-like case. Higher values were found in older animals. Four out of six horses which traveled or stayed in the US exhibited titers over 50, a higher figure than in the group which did not travel out of France or stayed in an other European country. No correlation was found between anti-S. neurona and anti-Neospora sp. antibody titers. Data prompt further study of significance of anti-S. neurona antibodies in clinically healthy or diseased European horses, and identification of putative immunizing parasite(s) and their host(s).     

Neurologic disease in gamma-interferon gene knockout mice caused by Sarcocystis neurona sporocysts collected from opossums fed armadillo muscle.

Fifteen gamma-interferon gene knockout mice were each orally inoculated with 5 x 10(3) Sarcocystis sporocysts derived from Virginia opossums (Didelphis virginiana) fed nine-banded armadillo (Dasypus novemcinctus) muscle containing sarcocysts. Three mice were inoculated with similarly obtained homogenates, but in which no sporocysts were detected. Mouse M8 was pregnant when inoculated and gave birth during the trial. Fifteen of 15 (100%) mice inoculated with sporocysts developed neurologic signs and/or died by day 30 d.p.i. One of 3 (33.3%) mice inoculated with homogenates in which no sporocysts were detected developed clinical signs and died at 34 d.p.i. All young of mouse M8 had maternally acquired antibodies to Sarcocystis neurona, but none developed clinical neurologic signs or had protozoal parasites in their tissues. All brains from mice that developed clinical signs contained merozoites that reacted positively to S. neurona antibodies using immunohistochemical techniques. Evidence from this study further supports the nine-banded armadillo being an intermediate host of S. neurona.     

First isolation of Sarcocystis neurona from the South American opossum, Didelphis albiventris, from Brazil

Sarcocystis neurona was isolated from sporocysts from two of eight South American opossums, Didelphis albiventris, from Brazil. Interferon gamma gene knock out (KO) mice fed sporocysts from two opossums developed neurologic sarcocystosis. S. neurona was demonstrated in the brains of infected KO mice by immunohistochemical staining with anti-S. neurona antibody. The parasite was cultivated in cell culture and S. neurona DNA was isolated from cultured merozoites. This is the first report of isolation of S. neurona from Brazil and the first report from its new host, D. albiventris.     

A herd-level analysis of risk factors for antibodies to Sarcocystis neurona in Michigan equids

Equine protozoal myeloencephalitis (EPM) is a neurological disease of horses and ponies caused by infection of the central nervous system with the protozoan parasite Sarcocystis neurona. A herd-level analysis of a cross-sectional study of serum antibodies to S. neurona in Michigan equids was conducted, using data collected in 1997 for study that included 1121 equids from 98 Michigan horse farms. Our objective was to identify specific herd-level risk factors associated with seropositivity. We tested associations between herd seroprevalence and various farm-management practices (including feed-storage methods and wildlife control). Multivariable models were developed for three strata based on relative opossum abundance (opossum districts). Herd seroprevalence ranged from 0 to 100% (median=57%). No risk factor was significantly associated with herd seroprevalence at P< or = 0.05 in all opossum districts. Our results suggest that equids living in areas with large opossum populations might be infected with S. neurona from multiple sources.