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Proceedings of the
55th Annual Convention of
the American Association of
Equine Practitioners
December 5–9, 2009, Las Vegas, Nevada
Program Chair : Nathaniel A. White

ACKNOWLEDGMENTS
Dr. David D. Frisbie, Educational Programs Committee Chair
Carey M. Ross, Scientific Publications Coordinator

Published by the American Association

of Equine Practitioners
www.aaep.org
ISSN 0065–7182
© American Association of Equine Practitioners, 2009
Published in IVIS with the permission of the AAEP Close this window to return to IVIS

MEDICINE—NEUROLOGY

Evidence-Based Review of Diagnosis and

Treatment of Sarcocystis neurona Infection
(Equine Protozoal Myeloencephalitis)

Amy L. Johnson, DVM, Diplomate ACVIM-LAIM

Equine protozoal myeloencephalitis (EPM) should only be diagnosed when compatible clinical signs
and seropositivity are present and other likely diseases have been excluded. Available evidence
suggests that the Western blot test is highly sensitive but not as specific as the indirect fluorescent
antibody test, which is also highly sensitive. New data indicate that the surface antigen-1 (SAG-1)
enzyme-linked immunosorbent assay (ELISA) may be insensitive, likely because of the Sarcocystis
neurona strain variation. None of the available/approved treatments are obviously superior; all have
shown an ⬃60% success rate in clinical trials. Author’s address: New Bolton Center, 382 West
Street Road, Kennett Square, Pennsylvania 19348; e-mail: aljdvm03@gmail.com. © 2009 AAEP.

1. Introduction peripheral neuropathy to diffuse CNS dysfunction.

Sarcocystis neurona infection is the most common
cause of EPM (equine protozoal myeloencephalitis).
This neurologic disease presents a diagnostic chal-
lenge to practitioners, because many horses are ex-
posed to the protozoa and clinical signs can mimic
many other conditions. Treatment is also challeng-
ing, because several medications are available and
response to treatment is not consistent among
horses. This review summarizes current commer-
cially available diagnostic and therapeutic options
and the evidence supporting each option.
2. Diagnosis
A complete neurologic exam should always be the
mainstay of diagnosis. If clinical signs cannot be at-
tributed to lesions in one or more regions of the central
nervous system (CNS), EPM should not be considered.
However, EPM can mimic almost any neurologic dis-
ease and cause signs ranging from a single cranial or
Certain patterns, such as multifocal signs, mixed
lower motor neuron and upper motor neuron signs
(e.g., both muscle atrophy and spinal proprioceptive
ataxia), and asymmetric signs are more commonly ob-
served with EPM than other neurologic diseases.
Additional support for a diagnosis of EPM should
include exclusion of other common neurologic dis-
eases and confirmation of exposure to S. neurona
through serology and/or cerebrospinal fluid (CSF)
analysis. For example, negative results of survey
cervical radiography and/or myelography can ex-
clude cervical vertebral malformation or cervical
stenotic myelopathy. Absence of fever and respira-
tory disease, as well as negative buffy coat and nasal
swab polymerase chain reaction (PCR), can exclude
Equine Herpes Virus-1 myeloencephalopathy. Nega-
tive IgM capture enzyme-linked immunosorbent assay
(ELISA) or absence of mosquito vectors can exclude
West Nile virus from consideration.

NOTES

172 2009 Ⲑ Vol. 55 Ⲑ AAEP PROCEEDINGS

Proceedings of the Annual Convention of the AAEP - Las Vegas, NV, USA, 2009
Published in IVIS with the permission of the AAEP
3. Diagnostic Tests
The three most commonly used ante-mortem diag-
nostic tests (Western blot [WB], indirect fluorescent
antibody test [IFAT], and surface antigen-1 (SAG-1)
ELISA) are all based on anti-S. neurona antibodies
and can be performed on serum or CSF. These
tests provide supportive evidence only; none is con-
sidered a gold standard. Necropsy remains the
only definitive test. In a previous review, the evi-
dence to support the use of these tests was discussed
in detail.1 This review summarizes the evidence
and sensitivity/specificity estimates previously dis-
cussed as well as summarizes new data available
regarding the SAG-1 ELISA.
WB
Initial estimates of the sensitivity (Se) and specific-
ity (Sp) of the WB test were high (serum Se ⫽ 89%;
serum Sp ⫽ 71%; CSF Se ⫽ 89%; CSF Sp ⫽ 89%).2
Shortly thereafter, other data indicated that the WB
Sp likely was lower for detecting EPM, because sim-
ilar proportions of normal horses and abnormal
horses (with neurologic signs, gait abnormalities, or
performance problems) were positive on CSF WB.3
A more recent prospective study estimated WB
Se/Sp for both neurologic and normal horses; results
on CSF yielded Se ⫽ 87% and Sp ⫽ 44% for neuro-
logic horses and Se ⫽ 88% and Sp ⫽ 60% for normal
horses (serum results were similar).4 If weak pos-
itive results were considered negative, Sp improved
but Se declined. Limitations of this study included
questionable accuracy of classification of horses, a
small number of confirmed positive cases, and blood
contamination of CSF samples. A modified WB
(with a blocking step) reported improved Se/Sp
(100% and 98%, respectively); however, it was only
tested on a small number of confirmed positive
horses and was never tested on horses that had
potentially been exposed to S. neurona but did not
have clinical disease.5
One of the difficulties in interpreting WB results
is the high number of seropositive horses. For this
reason, testing CSF has been advocated over serum
and will likely result in a reduction in false-positive
results. When a large number of paired serum and
CSF results were analyzed, 29% of CSF samples
from seropositive horses were negative.6 Unfor-
tunately, only a small amount of blood contami-
nation may affect CSF results; if the blood is
moderately or strongly reactive, as few as 8 red
blood cells (RBCs)/␮l in the CSF may cause false-
positive results.7
IFAT
Initial comparative analysis of the IFAT using a
small subset of serum samples from the Daft et al.4
study yielded comparable Se values (88.9% for IFAT,
WB, and modified WB) but differing Sp values
(IFAT ⫽ 100%; WB ⫽ 87.2%; and modified WB ⫽
69.2%).8 The IFAT was subsequently analyzed
with a larger sample set of naturally infected horses
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MEDICINE—NEUROLOGY
(including samples used in previous studies4,8),
yielding a Se value of 83.3% and a Sp value of 96.9%
on serum samples and a Se value of 100% and a Sp
value of 99% on CSF samples.9 As before, limita-
tions included the small number of positive cases
and questionable classification of horses.
One caveat to the IFAT is that crossreaction with
Sarcocystis fayeri could lead to false-positive re-
sults10; it is currently unclear whether or not this is
a clinically relevant problem. One benefit of the
IFAT compared with the WB is that CSF blood con-
tamination has a far less significant effect; there
was no effect on IFAT results at any serological titer
when the CSF was contaminated with ⬍104
RBCs/␮l.11
SAG-1 ELISA
The newest commercially available test is the SAG-1
ELISA, which is based on an immunodominant sur-
face antigen of S. neurona (SAG-1). Its first evalu-
ation in the literature using naturally occurring
cases lacks detail and does not include Se and Sp
estimates.12 Three hundred thirty samples (serum
and CSF) from horses with “a presumptive diagnosis
of EPM” were tested, and 85.4% were positive.
Unfortunately, no inclusion criteria were specified.
One hundred forty-seven samples from presump-
tively normal horses were also analyzed, and 24%
were positive. Again, these cases were not defined.
Data from the author of this review indicate that
the SAG-1 ELISA shows poor Se and fair Sp for
naturally occurring cases in the mid-Atlantic region.
In this patient population, preliminary results
showed that the overall Se of the SAG-1 ELISA was
17% and Sp was 79% with a cutoff value of 1:16.13
The low sensitivity severely limited the usefulness
of this test in this population.
Hoane et al.14 independently evaluated the use of
ELISAs for EPM diagnosis. They determined that
Se (90.9 –95.5%) and Sp (78.6 –92.9%) values were
high for ELISAs based on other surface antigens
(SAGs 2, 3, and 4), but their SAG-1 ELISA had a Se
of only 68.2% and Sp of 71.4%.14 They proposed
that the reduced sensitivity of the SAG-1 ELISA
may be explained by the fact that this surface anti-
gen may be absent in certain S. neurona isolates in
nature.14 Later work supported S. neurona surface
antigen diversity and showed that SAG-5 is an al-
ternative surface antigen of S. neurona strains,
which is mutually exclusive to SAG-1.15 Therefore,
the SAG-1 ELISA is only likely to be useful in areas
where SAG-1 expressing strains of S. neurona pre-
dominate; in areas where SAG-5 expressing strains
predominate, the test will fail to identify infected
horses. S. neurona strains lacking SAG-1 have
been identified in a variety of geographic locations
(including the states of California, Oregon, Missouri,
and Ohio), but insufficient information is available
to predict the likely utility of the SAG-1 ELISA for
specific geographic regions.
AAEP PROCEEDINGS Ⲑ Vol. 55 Ⲑ 2009 173
Published in IVIS with the permission of the AAEP
MEDICINE—NEUROLOGY
4. Treatment
There are four treatments currently approved by the
Food and Drug Administration (FDA) for EPM, but
only three have been commercially available: a sul-
fadiazine and pyrimethamine combination, pona-
zuril, and nitazoxanide. Diclazuril has been
approved but not yet marketed. Recently (spring of
2009), the commercially available form of nitazox-
anidea has been discontinued. Although multiple
investigators have examined in vitro activity of var-
ious drugs, their pharmacokinetics, and their use in
prevention of experimental infection, this review
will focus only on the efficacy and side effects of the
four approved treatments as documented in clinical
cases.
Sulfadiazine and Pyrimethamine
The FDA-approved combination is sulfadiazine and
pyrimethamineb dosed at 20 mg/kg sulfadiazine and
1 mg/kg pyrimethamine daily for a minimum of 90
days. If the FDA-approved combination is unavail-
able, some practitioners opt to use trimethoprim-
sulfa tablets (20 –30 mg/kg, q 12–24 h, PO) with
pyrimethamine tablets (1 mg/kg, q 24 h, PO) be-
cause of availability or ease of administration.
However, these drugs are not FDA-approved for
EPM treatment, and therefore, this regimen consti-
tutes extra-label/unapproved use. All three of
the drugs (trimethoprim, sulfadiazine, and py-
rimethamine) inhibit enzymes in the folic acid
pathway and thereby, inhibit thymidine synthe-
sis; trimethoprim and pyrimethamine inhibit di-
hydrofolate reductase, whereas sulfadiazine
blocks the conversion of para-aminobenzoic acid to
dihydrofolic acid.
A field study was performed during the approval
process at multiple sites to evaluate two dose levels
of sulfadiazine and pyrimethamine (1⫻ and 2⫻ the
aforementioned dose for 90 –270 days).16 Inclusion
criteria included neurologic signs and positive CSF
WB; treatment success was defined by negative CSF
WB and/or marked clinical improvement (two or
more grades of improvement). Of the horses com-
pleting the study, 16 of 26 (61.5%) treated at the 1⫻
dose had successful outcomes. The most common
adverse reaction was bone marrow suppression
(anemia, leucopenia, neutropenia, and/or thrombo-
cytopenia) in 12% of the 1⫻ group and 21% of the 2⫻
group.
An epizootic of EPM at a farm was described dur-
ing which 12 horses were diagnosed with EPM based
on neurologic signs and positive CSF WB.17 All
horses were treated with pyrimethamine and tri-
methoprim-sulfamethoxazole until they were nega-
tive on CSF WB (45–211 days). Eleven of twelve
horses became negative on CSF WB in this time
period. Adverse effects attributed to treatment in-
cluded fever, leucopenia, anorexia, depression, acute
worsening of ataxia, mild anemia, and abortions.
In addition to blood dyscrasias, folic acid defi-
ciency may lead to gastrointestinal disturbances
174 2009 Ⲑ Vol. 55 Ⲑ AAEP PROCEEDINGS
Proceedings of the Annual Convention of the AAEP - Las Vegas, NV, USA, 2009
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such as glossitis.18 Stallions treated for 90 days
with trimethoprim-sulfamethoxazole and py-
rimethamine may have changes in copulatory form
and agility along with altered pattern and strength
of ejaculation.19 Three mares receiving sulfon-
amides, pyrimethamine (with or without tri-
methoprim), and folic acid delivered foals with
congenital defects that died or were euthanized.20
Ponazuril
Ponazuril is a triazinetrione antiprotozoal drug that
targets the “apicoplast” organelle and inhibits en-
ergy metabolism (respiratory chains). The label
dosage regimen for ponazurilc is 5 mg/kg PO daily
for 28 days.
A multi-center field study was performed during
the approval process for ponazuril (Marquis) and
subsequently published.21 Inclusion criteria man-
dated that horses have neurologic gait abnormali-
ties, normal cervical radiographs, and positive CSF
WB results. Horses were divided into two treat-
ment groups: 5 mg/kg daily for 28 days or 10 mg/kg
daily for 28 days. One hundred one horses were
included, and 63 (62%) had a favorable outcome
(improving by at least one neurologic grade 90 days
after stopping treatment or becoming negative on
CSF WB). Sixty percent (28 of 47) in the low-dose
group improved and 65% (35 of 54) in the high-dose
group improved. No adverse effects were noted in
either group as reported in the efficacy study.
However, information provided by the manufacturer
reports “unusual daily observations” in eight ani-
mals that may have been related to treatment in-
cluding blisters on nose and mouth, skin rash or
hives, loose stools, mild colic, and a seizure.c This
information also mentioned slightly different results
for the high-dose group in that 32 of 55 (58%) horses
improved.
Nitazoxanide
As previously mentioned, nitazoxanide pastea is no
longer commercially available. This drug is a 5-ni-
trothiazole antiparasitic drug that inhibits the pyru-
vate:ferredoxin oxidoreductase (PFOR) enzyme-
dependent electron transfer reaction essential for
anaerobic energy metabolism.
Before the research provided by the manufacturer
of nitazoxanide paste, there were two case series
that described the use of nitazoxanide. McClure
and Palma22 described two horses diagnosed with
EPM on the basis of neurologic signs and positive
CSF WB results that were treated with nitazoxanide
tablets and/or paste at 50 mg/kg daily for either 28
or 42 days; both horses improved, and the one with
longer follow-up information had no residual neuro-
logic abnormalities.22 Vatistas et al.23 described
their initial experiences using nitazoxanide (50 or 75
mg/kg daily for 28 days) in seven horses diagnosed
with EPM using the same criteria; five horses be-
came neurologically normal, one horse improved,
Published in IVIS with the permission of the AAEP
and one was unchanged.23 Reported side effects in
these studies included inappetence and depression.
Two field studies for efficacy and safety were con-
ducted during the approval process for nitazoxanide.
When the efficacy data were analyzed, 49 horses
from the first study and 250 from the second study
were included. Inclusion criteria were more strin-
gent for the first study; horses were required to have
asymmetric spinal ataxia or multifocal neurologic
signs along with positive CSF WB results. In the
second study, neither a videotape of the neurologic
exam nor CSF analysis was required. Also, some
horses in the second study had been previously
treated for EPM with other drugs. In the first
study, 57% of horses improved by one grade and/or
became negative on CSF WB as assessed by clinical
investigators. In the second, less stringent study,
81% were considered treatment successes. Side ef-
fects were seen in 22 of 81 (27%) horses in the first
study and 129 of 416 (31%) horses in the second
study. The most common adverse reactions were
fever, anorexia/reduced appetite, and lethargy/de-
pression. Twenty-eight horses died or were eutha-
nized in the second study, and five of these cases
were potentially caused by nitazoxanide use,
prompting the following warning: “administration
of nitazoxanide can disrupt the normal microbial
flora of the gastrointestinal tract leading to entero-
colitis. Deaths due to enterocolitis have been ob-
served while administering the recommended dose
in field studies.”a
Diclazuril
Diclazuril is a triazinetrione antiprotozoal agent
similar to ponazuril with an unknown (but possibly
similar) mechanism of action.
A multi-center clinical field study was performed
using diclazuril pellets at a dose of 1 mg/kg daily for
28 days.24 Criteria for inclusion were similar to
other field trials and consisted of neurologic signs
and positive CSF WB tests. Horses were also
tested for other neurologic diseases and excluded if
abnormal results were obtained. Forty-two horses
completed the study; 67% were considered successes
(negative CSF WB and/or improvement by one
grade) by clinical investigators. Reported adverse
reactions were not clearly linked to drug adminis-
tration and included worsening neurologic status
and laminitis.
5. Conclusions
Ante mortem diagnosis of EPM is always presump-
tive and should be based on the presence of compat-
ible neurologic signs, exclusion of other likely
diseases, and positive serology and/or CSF analysis
for S. neurona infection. Testing CSF samples
(alone or in addition to serum samples) improves
specificity. Three types of serologic tests are com-
mercially available to aid in diagnosis. Available
evidence indicates that both the WB and IFAT have
similar sensitivities (⬃90%). The estimated IFAT
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MEDICINE—NEUROLOGY
specificity is higher than the estimated WB specific-
ity, and the IFAT CSF results are much less affected
by blood contamination than WB CSF results. The
SAG-1 ELISA has the least amount of evidence to
support its use, and preliminary data yield a low
sensitivity that will likely limit its use in some or
most geographic regions.
Results of efficacy studies were surprisingly sim-
ilar for each of the four approved therapies when
similar methodologies and means of assessing im-
provement were used. Regardless of drug, ⬃60% of
treated horses improved by at least one neurologic
grade or became negative on CSF WB. Prospective,
randomized, blinded clinical trials would aid in as-
sessing if one drug shows superior efficacy. Based
on reported side effects, ponazuril and diclazuril
seem to have the fewest reported adverse effects.
References and Footnotes
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MEDICINE—NEUROLOGY
13. Johnson AL, Sweeney RW. Utility of two serologic tests for
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176 2009 Ⲑ Vol. 55 Ⲑ AAEP PROCEEDINGS
Proceedings of the Annual Convention of the AAEP - Las Vegas, NV, USA, 2009
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azole and pyrimethamine. J Am Vet Med Assoc 1999;215:
1317–1319.
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a
Navigator®, Idexx Laboratories, Inc., Westbrook, ME 04092
(no longer available).
b
Rebalance™ Antiprotozoal Oral Suspension, IVX Animal
Health Inc., St. Joseph, MT 64503.
c
Marquis®, Bayer HealthCare LLC, Shawnee Mission, KS
66201.