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Contents lists available at SciVerse ScienceDirect

Veterinary Parasitology

jo urnal homepage: www.elsevier.com/locate/vetpar

Short communication

Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine

a,∗ b c c

B. Moreno , E. Collantes-Fernández , A. Villa , A. Navarro ,

b b

J. Regidor-Cerrillo , L.M. Ortega-Mora

a

Centro de Investigación en Encefalopatías y Enfermedades Transmisibles Emergentes, Pathology Department, Faculty of Veterinary Sciences, Zaragoza

University, Miguel Servet, 177, 50013 Zaragoza,

b

SALUVET, Animal Health Department, Faculty of Veterinary Sciences, Complutense University of , Ciudad Universitaria s/n, 28040 Madrid, Spain

c

Laboratorio Exopol. Polígono del Río Gállego D/8, 50840 San Mateo, Zaragoza, Spain

a r t i c l e i n f o a b s t r a c t

Article history: Neospora caninum and Toxoplasma gondii are closely related cyst-forming apicomplexan

Received 10 November 2011

parasites identified as important causes of reproductive failure in cattle and small rumi-

Received in revised form

nants, respectively. Protozoan in small ruminants is traditionally associated with

15 December 2011

T. gondii, but the importance of N. caninum remains uncertain. The aim of this study was

Accepted 29 December 2011

to investigate the presence of N. caninum and T. gondii infections in abortion cases in small

ruminants submitted for diagnosis. For this purpose, 74 ovine and 26 caprine aborted foe-

Keywords:

tuses were recovered from different areas in Spain. Foetal histopathology was used to

Neospora caninum

detect the presence of protozoal-associated lesions in brain. The presence of N. caninum

Toxoplasma gondii

Abortion and T. gondii was confirmed by PCR. Protozoal infection was detected in 17 out of 100 (17%)

Ovine foetuses examined by at least one of the diagnostic techniques used. Lesions suggestive

Caprine of protozoal infection were observed in 10.8% (8/74) and 15.4% (4/26) of the ovine and

caprine abortions respectively. N. caninum and T. gondii infection was detected by PCR in

6.8% (5/74) and 5.4% (4/74) of sheep foetuses, respectively, of which five showed protozoal-

associated lesions. N. caninum DNA was detected in 11.5% (3/26) of goat foetuses, of which

two showed protozoal-associated lesions, whereas T. gondii DNA was detected in one goat

foetus with no lesions. The simultaneous presence of N. caninum and T. gondii DNA was

detected in one sheep foetus with severe lesions. This study demonstrates that N. caninum

plays a significant role in abortion in small ruminants in the studied population. In addi-

tion, our results highlight the importance of differentiating between protozoa whenever

characteristic lesions are observed.

© 2012 Elsevier B.V. All rights reserved.

1. Introduction range of factors must be considered. Although other causes

may exist, infectious aetiologies seem to predominate in

The productivity of small ruminant flocks depends sheep and goats. The diagnosis of an infection cause

greatly on their reproductive efficiency. Abortion is a of abortion is often difficult and should be made in a

major cause of economic losses in the livestock industry specialised veterinary laboratory; thus, a high percentage

due to its high prevalence. In a large number of abortions, of cases remain undiagnosed (Kirkbride, 1993; Moeller,

the precise cause cannot be determined because a broad 2001). Definitive diagnosis rate of between 40% and 62%

are achieved in good diagnostic laboratories (Oporto et al.,

2006; Edmondson et al., 2012). A variety of pathogens

∗ have been classically reported as being abortive in these

Corresponding author. Tel.: +34 876554185; fax: +34 976762508.

E-mail address: [email protected] (B. Moreno). species, some of which also have zoonotic potential,

0304-4017/$ – see front matter © 2012 Elsevier B.V. All rights reserved.

doi:10.1016/j.vetpar.2011.12.034

Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034

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including Brucella melitensis, Chlamydophila abortus, Toxo- Spain. A complete necropsy was performed on all foetuses

plasma gondii, Coxiella burnetii and the emerging pathogen submitted, and the presence of macroscopic lesions was

Campylobacter jejuni. In addition, due to the high number of also evaluated. Samples of brain, lung, heart, liver, spleen

cases that remain undiagnosed (Edmondson et al., 2012), and kidney were obtained and fixed in 10% neutral buffered

it is important to search for new pathogens or agents not formalin at room temperature for histopathology. In addi-

typically associated with abortions in these species. tion, samples of anterior brain, mesencephalon, cerebellum

Neospora caninum and T. gondii are closely related cyst- and pons were subjected to PCR for the detection of N.

forming apicomplexan parasites identified as important caninum and T. gondii.

causes of reproductive failure in cattle (Dubey et al., 2007;

Dubey and Schares, 2011) and small ruminants, respec-

2.2. Histopathological analysis

tively (Buxton, 1998; Hurtado et al., 2001; Pereira-Bueno

et al., 2004; Masala et al., 2007; Dubey and Schares, 2011).

For the histopathological study, fixed tissues were

Protozoan abortion in sheep and goats is traditionally asso-

dehydrated through graded alcohols before being embed-

ciated with T. gondii, where characteristic necrotic lesions

ded in paraffin wax using routine procedures. Blocks

in the central nervous system are usually observed (Buxton,

were cut in 4 ␮m sections, deparaffinised, rehydrated,

1998; Hurtado et al., 2001; Dubey, 2003). However, similar

stained with hematoxylin and eosin (H/E) and exam-

lesions can be also observed in Neospora-associated abor-

ined by light microscopy. Organs were examined for

tion; therefore, discrimination between these infections

protozoal-associated lesions. For brain tissues, microscopic

is difficult from a pathological point of view (McAllister

examination was done on the mesencephalon, cerebellum

et al., 1996; Moreno, personal communication; Conraths

and pons. Identified lesions were classified as previously

and Gottstein, 2007). Nevertheless, in small ruminants, N.

reported (Pereira-Bueno et al., 2004). Briefly, in brain tis-

caninum seems to be much less relevant and its impor-

sues, “characteristic” lesions contained multifocal necrotic

tance remains unclear (Otter et al., 1997; Buxton, 1998;

foci surrounded by inflammatory cells and/or multiple foci

Moeller, 2001; Dubey, 2003; Moore, 2005; Givens and

of non-suppurative infiltrates, while “compatible” lesions

Marley, 2008). Serological surveys in both sheep (Helmick

were less severe, with scarce glial foci, diffuse gliosis and

et al., 2002; Reichel et al., 2008; Bártová et al., 2009;

no necrotic areas. Additionally, based on the number of

Panadero et al., 2010) and goats (Abo-Shehada and Abu-

necrotic foci, “characteristic” lesions were classified as mild

Halaweh, 2010; Dubey and Schares, 2011) have shown a

(≤2 necrotic foci), moderate (2–5 necrotic foci) or severe

variable prevalence ranging from 0.6% to 30.8%. Moreover,

(>5 necrotic lesions). “Compatible” lesions were classi-

abortion cases attributed to N. caninum infection have been

fied as mild. Heart, lung, liver, kidney and spleen tissues

occasionally reported in small ruminants (Barr et al., 1992;

were also examined for the presence of mononuclear infil-

Dubey et al., 1996; Corbellini et al., 2001; Kobayashi et al.,

tration and/or necrotic foci. These lesions were classified

2001; Dubey, 2003; Eleni et al., 2004). Much less frequently,

as mild (+), moderate (++) or severe (+++) as described

N. caninum has been associated with epizootic abortion

above.

(Moreno, personal communication; Moeller, 2001; Hässig

et al., 2003). In addition, Neospora infection has occasion-

ally been detected in neonates, adult animals with nervous 2.3. DNA extraction and nested PCR for the detection of

signs (Dubey et al., 1992; Corbellini et al., 2001; Bishop N. caninum and T. gondii

et al., 2010) and asymptomatic animals (Koyama et al.,

2001). These studies indicate the presence of N. caninum Genomic DNA extraction was performed on three dif-

infection and the potential for transplacental transmission ferent samples of all brain sections recovered at necropsy.

in small ruminants; however, the frequency and aetiology DNA was extracted from 20 mg of brain tissue using a

of abortion due to this parasite remain unclear. commercial kit (Real Pure, Durviz, Spain) according to the

The aim of this article is to provide data about the occur- manufacturer’s protocol. DNA concentrations were deter-

rence of T. gondii and N. caninum in cases of small ruminant mined by spectrophotometric analysis at A260/280, and all

abortion in Spain by means of some of the most widely used samples were diluted to a final concentration of 60 ng/␮l.

◦

techniques for diagnosis of infections in foetuses. DNA samples were stored at −20 C prior to PCR analysis.

Nested PCR was performed on all samples to detect

2. Materials and methods N. caninum and T. gondii DNA. For N. caninum, four

oligonucleotide primers were used to amplify the internal

2.1. Samples transcribed spacer (ITS1) region, as described by Buxton

et al. (1998). For T. gondii, a single-tube nested-PCR assay

Seventy-four sheep foetuses and twenty-six goat foe- was performed as previously described (Hurtado et al.,

tuses were sent to the Diagnostic Service of Exopol in 2008 2001). Positive controls (purified N. caninum or T. gondii

and 2009 for general aetiological diagnosis. In addition to tachyzoite DNA) and negative controls (double-distilled

T. gondii and N. caninum, typical pathogens associated with water) were included in each set of PCR reactions. To

abortion in sheep and goats, including C. abortus, Coxiella avoid false-positive reactions, DNA extraction, PCR sam-

burnetti, Leptospira spp., B. melitensis, Border disease virus, ple preparation and electrophoresis were performed in

Campylobacter spp., Salmonella abortus ovis and Listeria separate rooms with different sets of instruments, and

spp., were also analysed. All foetuses were obtained from aerosol barrier tips and disposable gloves were employed.

abortion cases that took place on farms in different areas of Amplification products were separated by 1.8% agarose gel

Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034

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Fig. 1. Brain lesions in a sheep foetus with N. caninum infection. Necrotic

focus is observed in the white matter of the mesencephalon. H/E.

electrophoresis, stained with ethidium bromide and visu-

alised under UV light.

2.4. N. caninum microsatellite analysis

Microsatellite analysis was performed on N. caninum

PCR-positive DNA samples extracted from aborted foetuses

as previously reported (Pedraza-Díaz et al., 2009). Briefly,

MS4, MS5, MS6A, MS6B, MS7, MS8, MS10, MS12 and MS21

markers were amplified using specific primers and nested-

PCR conditions as described by Pedraza-Díaz et al. (2009).

The sizes of the PCR products for all microsatellites were

also determined using a 48-capillary 3730 DNA Analyser

(Applied Biosystems, Foster City, CA) with GeneScan-500 Fig. 2. Lesions in goat foetuses with N. caninum infection. (a) Brain.

Glial focus in the white matter of the mesencephalon. (b) Liver. Necrotic

(LIZ) Size Standards (Applied Biosystems,) at the Unidad

focus in the parenchyma. Several degrees of interstitial hepatitis and

Genómica del Parque Científico de Madrid, and the results

haematopoiesis as well as autolysis can be observed. H/E.

were analysed with GeneMapper1 v3.5 software. Addition-

ally, the markers MS7 and MS10 were sequenced using

a Big Dye Terminator v3.1 Cycle Sequencing Kit (Applied

N. caninum DNA was detected in 11.5% (3/26) of goat

Biosystems) and a 3730 DNA Analyser (Applied Biosys-

foetuses, of which two also showed protozoal-associated

tems) at the Unidad Genómica del Parque Científico de

lesions (Fig. 2a). T. gondii DNA was detected in one foetus

Madrid. Sequences were analysed using BioEdit Sequence

with no lesions. The simultaneous presence of N. caninum

Alignment Editor v.7.0.1 software (Copyright 1997–2004

and T. gondii DNA was detected in only one sheep foetus,

Tom Hall, Ibis Therapeutics, Carlsbad, CA 92008, USA).

which also showed the most severe lesions. In two sheep

foetuses with severe lesions, cyst-like structures were

3. Results observed close to inflammatory foci in the nervous tissue

of one foetus and in the myocardium of the other.

Protozoal infections were detected by at least one of the According to our classification of severity based in the

diagnostic techniques in 17 out of 100 (17%) foetuses exam- presence and number of necrotic foci in the brain, the most

ined. Of the infected foetuses, 14.9% (11/74) and 23% (6/26) severe lesions were found in sheep. Characteristic necrotic

were ovine and caprine foetuses, respectively (Table 1). In lesions were observed in six sheep foetuses and in zero goat

ovine foetuses, “characteristic” lesions of protozoal abor- foetuses (Table 1). In goats, lesions mainly corresponded

tion were observed in the brain tissues of seven foetuses with glial proliferation in the brain. Histological lesions

(9.5%; 7/74) and “compatible” lesions were observed in one associated with protozoal infection were also observed in

brain (1.4%; 1/74). N. caninum was detected by specific PCR other organs in most of the foetuses, and they were mainly

in 5 out of 74 foetuses (6.8%), three of which showed “char- characterised by several degrees of mononuclear inflam-

acteristic” lesions of protozoal infection in the brain (Fig. 1). matory infiltrates (Table 1). However, in one goat foetus

T. gondii DNA was detected in four out of 74 foetuses (5.4%), infected by N. caninum, an atypical pattern of moderate

three of which showed protozoal-associated lesions. Of the diffuse gliosis in the brain, with no necrotic lesions and

goat foetuses, three (11.5%; 3/26) showed characteristic numerous necrotic foci in the lung, liver and spleen, was

lesions and one (3.8%; 1/26) showed compatible lesions. observed (Fig. 2b).

Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034

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Table 1

Summary of lesions and PCR results obtained in samples corresponding to protozoal-infected foetuses.

a b

Species Sample no. Results of histology in Results of

Brain Lung Liver Kidney Heart Nc PCR Tg PCR

1 ++(a) − A A − + −

2 ++(c) − + A + + +

3 + + + + + − −

4 ++(a) + + + + − +

5 ++(a) A + A − − −

Sheep 6 ++(a) A A A A − −

−

7 ++(b) + A − + −

8 ++(b) + − + + − +

9 − − + − − + −

10 −A A A A A + −

11 − − − − − − +

12 + − + + + − −

13 ++ + + + + − −

14 ++ + + + − + −

Goat

15 ++ − + − − + −

16 −A A A A A + −

17 − − − − − − +

A, autolysis.

a

Pathological features: absence (−) and presence (+) of protozoal-associated lesion. In brain tissues, “+” indicates samples showing “compatible” lesions

of protozoal infection and “++” “characteristic” lesions. In addition (a) means mild lesions with ≤2 necrotic foci, (b) means moderate lesions with 2–5

necrotic foci and (c) means severe lesions with >5 necrotic foci.

b

Nc, N. caninum; Tg, T. gondii.

Amplification of N. caninum microsatellite markers was caninum could be an important in small rumi-

only achieved using DNA from brain samples of an ovine nants, as N. caninum DNA associated with characteristic

foetus that also tested positive by PCR (Table 2). New alleles lesions was found as frequently as T. gondii in the stud-

were not identified after satellite marker analysis. How- ied animals. N. caninum has been sporadically associated

ever, comparison of this multilocus profile with others with abortions in small ruminants (Conraths and Gottstein,

previously described for N. caninum isolates and infected 2007; Dubey and Schares, 2011). N. caninum was detected

clinical samples showed a unique multilocus pattern for in the brains of 3 out of 18 aborted ovine foetuses in New

this sample. Zealand (Howe et al., 2008), in 18.9% of 74 aborted ovine

foetuses in England (Hughes et al., 2006) and in 2% of 292

4. Discussion aborted sheep foetuses in (Masala et al., 2007). In goat

abortions, N. caninum was found in 8.6% of 23 aborted foe-

T. gondii is considered to be the primary parasite tuses in Italy (Masala et al., 2007) and in 8.3% of 12 aborted

responsible for both sheep and goat abortion. However, foetuses in Spain (Moreno, personal communication). On

the economic, clinical and epidemiological importance of the other hand, T. gondii DNA was only detected in four

N. caninum infection in these species remains uncertain (5.4%) ovine foetuses and one (3.8%) caprine foetus in the

(Dubey and Schares, 2011). To date, this work is the largest present study. These figures are lower than those previ-

study that has been performed to investigate the involve- ously reported in Spain, in which T. gondii was detected

ment of N. caninum and T. gondii in cases of small ruminant in 15–23% of aborted sheep foetuses (Hurtado et al., 2001;

abortion, both in terms of the number of analysed foe- Pereira-Bueno et al., 2004; Oporto et al., 2006) and in 8.3% of

tuses and the range of diagnostic techniques applied. In this goat abortions (Moreno, personal communication). These

study, we used foetal histopathology combined with PCR to differences may be influenced by geographical distribution

detect the presence of protozoal infection in aborted foe- of infection because those studies were mainly focused on

tuses from sheep and goats. Confirmation of the presence of abortions recovered from Northern Spain, whereas in this

the etiologic agent using specific techniques such as PCR is study T. gondii was detected in abortions from all Spanish

needed because N. caninum and T. gondii may cause similar regions. Furthermore, they may be also influenced by the

lesions. different methodologies employed, as Pereira-Bueno et al.

Protozoal infection was present based on at least one (2004) used serological techniques and Moreno (personal

of the diagnostic techniques in 17% (11 sheep and 6 goats) communication) histological and immunohistochemical

of the 100 foetuses analysed. In 12 foetuses (8 sheep and techniques.

4 goat foetuses), lesions suggestive of protozoal infection In this study, we observed characteristic lesions of pro-

were observed, and in the remaining five, only proto- tozoal infection in 9.5% and 11.5% of ovine and caprine

zoal DNA was detected. Our results indicate that protozoal foetuses, respectively. These data are similar to those

infection is an important cause of abortion in sheep and reported in previous studies in Spain (Moreno, personal

goats, as is often reported in the literature (Dubey, 2003; communication; Hurtado et al., 2001; Pereira-Bueno et al.,

Masala et al., 2007; Pereira-Bueno et al., 2004; Dubey, 2004) and elsewhere (Moeller, 2001; Dubey, 2003). In

2009). We also provide presumptive evidence that N. these studies, protozoal-associated lesions associated with

Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034

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T. gondii infection were found in 9.4% and 11.1% of aborted

-TT

ovine foetuses (Hurtado et al., 2001; Pereira-Bueno et al.,

6

2004) and in 16.7% of caprine abortion cases (Moreno,

personal communication). However, N. caninum detection

was not performed in these studies. Interestingly, Hurtado

et al. (2001) found a 100% correlation between lesions -TACC-(TACA)

3

and T. gondii DNA detection; however, this is not the rule,

as very often PCR and lesions present a fair correspon-

dence (Pereira-Bueno et al., 2004). This poor agreement

MS21 TG-(TACA)

is thought to be related to the uneven localisation and

low levels of the parasite (Pereira-Bueno et al., 2004). In -GC

15

our study, abortion cases with characteristic lesions were

mainly associated with the presence of N. caninum (62.5%)

and less often with T. gondii (37.5%). It has been suggested

MS12 GC-(GT)

8 that N. caninum may have been misidentified as T. gondii in

previous studies (Dubey, 2003). Lesions induced by N. can-

-(TGA) inum and T. gondii in abortion cases are similar, so specific

14

techniques must be used to detect the pathogen involved.

Consequently, N. caninum infection in small ruminant abor- -(AGA) b

6

tions may be underdiagnosed, and its importance remains

).

largely unknown (Dubey, 2003; Conraths and Gottstein, MS10 (ACT) 2006

2007). al.,

Another aim of this study was to compare lesions -GG et

12

between N. caninum and T. gondii. However, in both sheep

and goats, the small number of foetuses with lesions

MS8 AC-(AT) precluded a comparison between ruminant species and

9

between parasites. In sheep, characteristic necrotic foci

were typically seen, while in goats only glial reactions Regidor-Cerrillo (

b

were observed. Interestingly, in goats, similar to previ-

MS7 ATTA-(TA) ous observations (Moreno, personal communication), N.

analysis caninum-associated lesions mainly corresponded with glial

-GT

proliferation and very rare necrotic foci. While our study 12

failed to detect necrotic lesions, in a previous investiga-

fragment

tion only a small necrotic focus in the mesencephalon was MS6B CC-(AT)

size found (Moreno, personal communication). Some of the few

-AC pathologic descriptions of N. caninum-associated abortions 15

.

in goats have also described only glial reactions (Corbellini

et al., 2001) and less often the presence of necrotic foci according

MS6A GC-(TA)

(Eleni et al., 2004). More studies with an increased number caninum

N. of foetuses will be needed to determine whether patho-

expected

logic differences exist between N. caninum and T. gondii with

infections in sheep and goats.

-TGTA-GG

13 In addition, it is not clear whether an epidemiologi-

sequence

infected cal association exists between small ruminant and cattle

Neospora infections. Taking into account that N. caninum MS5 motif

sequencing.

foetus is one of the most important causes of abortion in cat-

by -AC CG-(TA)

tle in Spain, and that some farms have sheep, goats and 2

ovine

cattle in the same fields, an epidemiological relationship repetitive an

may be involved. Indeed, a previous study investigated this the for

possibility, although no relationship was found (Spilovská determined as

et al., 2009). A molecular characterisation and compari- -ACATTT-(AT) also 13

son of small ruminant and bovine isolates of N. caninum obtained shown

would improve our knowledge of the epidemiology of this were

are

pathogen. Microsatellite markers have been demonstrated MS4 GC-(AT) profile

to be a suitable tool for genotyping N. caninum from clin-

alleles

alleles

ical samples (Pedraza-Díaz et al., 2009). Thus, with the loci

genetic

a

MS10 aim of investigating N. caninum isolates disseminated in motif

ovine and caprine abortions, microsatellite genotyping was and

performed in the present study. However, we were able 2

sequence

Microsatellite MS7

to determine only a single microsatellite profile, from an a Microsatellite Repetitive b

Table

Microsatellite ovine sample, and were therefore unable to perform any

Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034

G Model

VETPAR-6207; No. of Pages 7 ARTICLE IN PRESS

6 B. Moreno et al. / Veterinary Parasitology xxx (2012) xxx–xxx

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trol of neosporosis and Neospora caninum. Clin. Microbiol. Rev. 20,

from those previously found in bovine samples (Regidor-

323–367.

Cerrillo et al., 2006, 2008; Rojo-Montejo et al., 2009; Basso

Dubey, J.P., Schares, G., 2011. Neosporosis in animals – the last five years.

et al., 2009a,b; Pedraza-Díaz et al., 2009). Vet. Parasitol. 180, 90–108.

Edmondson, M.A., Roberts, J.F., Baird, A.N., Bychawski, S., Pugh, D.G., 2012.

Our results confirm that transplacental transmission of

Theriogeneology of sheep and goats. In: Pugh, D.G., Baird, A.N. (Eds.),

N. caninum occurs in sheep and goats and indicate that N.

In Sheep and Goat Medicine. , 2nd ed. Elsevier, Saunders, Maryland

caninum could be a causative agent of abortion in domes- Heights, Missouri, p. 63043.

Eleni, C., Crotti, S., Manuali, E., Costarelli, S., Filippini, G., Moscati, L., Mag-

tic small ruminants. Consequently, neosporosis should be

nito, S., 2004. Detection of Neospora caninum in an aborted goat .

included in a differential diagnosis of causes of abortion

Vet. Parasitol. 123, 271–274.

in these species; however, further studies are necessary to Givens, M.D., Marley, M.S.D., 2008. Infectious causes of embryonic and

investigate the role of N. caninum as a cause of abortion fetal mortality. Theriogenology 70, 270–285.

Hässig, M., Sager, H., Reitt, K., Zeigler, D., Strabel, D., Gottstein, B., 2003.

in sheep and goats and to evaluate the resulting economic

Neospora caninum in sheep: a herd case report. Vet. Parasitol. 117,

losses to the industry. 213–220.

Helmick, B., Otter, A., McGarry, J., Buxton, D., 2002. Serological investiga-

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Acknowledgements

Howe, L., West, D.M., Collet, M.G., Tattersfield, G., Pattison, R.S., Pomroy,

W.E., Kenyon, P.R., Morris, S.T., Williamson, N.B., 2008. The role of

Neospora caninum in three cases of unexplained ewe abortions in the

We thank Carmen Cuevas (Animal Health Department,

southern North Island of New Zealand. Small Rumin. Res. 75, 115–122.

Complutense University of Madrid) for her excellent tech-

Hughes, J.M., Willians, R.H., Morley, E.K., Cook, D.A., Terry, R.S., Mur-

nical assistance. This work was funded by Programa de phy, R.G., Hide, G., 2006. The prevalence of Neospora caninum and

co-infection with Toxoplasma gondii by PCR in naturally occurring

Creación y Consolidación de Grupos de Investigación Uni-

mammal populations. Parasitology 132, 29–36.

versidad Complutense-Comunidad de Madrid (GR35-10).

Hurtado, A., Aduriz, G., Moreno, B., Barandita, J., García-Pérez, A.L., 2001.

Single tube nested PCR for the detection of Toxoplasma gondii in fetal

tissues from naturally aborted ewes. Vet. Parasitol. 102, 17–27.

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Please cite this article in press as: Moreno, B., et al., Occurrence of Neospora caninum and Toxoplasma gondii infections in

ovine and caprine abortions. Vet. Parasitol. (2012), doi:10.1016/j.vetpar.2011.12.034