Occurrence of antibodies against Neospora caninum in water buffaloes (Bubalus bubalis) on four ranches in Corrientes province, Argentina.

Abstract

The aim of the present work was to describe the occurrence of antibodies to Neospora caninum in water buffaloes on four ranches located in Corrientes province in the northeast of Argentina. Antibodies against N. caninum were determined in sera of 449 water buffaloes by using an indirect fluorescent antibody test (IFAT). A Bayesian logistic regression mixed model was used to quantify the strength of association between positive serological results to N. caninum and gender, age and category (calf, steer, heifer, cow) as risk factors. Antibody titers were found in 287 (64%) buffaloes. All ranches had seropositive animals. Age was more strongly associated with positive results to N. caninum (OR: 1.4; CI 95%: 0.86-2.22) than gender (OR: 1.02, CI 95%: 0.40-2.59) and category (OR: 0.88, CI 95%: 0.57-0.88). Results suggest a high exposure of water buffaloes to N. caninum by postnatal transmission in these four ranches located in Corrientes province, Argentina. Further studies are needed to quantify the consequences of Neospora-infections in the water buffalo industry.

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Short communication
Occurrence of antibodies against Neospora caninum in water
buffaloes (Bubalus bubalis) on four ranches in
Corrientes province, Argentina
C.M. Camperoa,*, A. Pe ´rezb, D.P. Mooreb, G. Crudelic, D. Benitezd,
M.G. Draghid, D. Canoa, J.L. Konradc, A.C. Odeo ´na
aInstituto Nacional de Tecnologı ´a Agropecuaria (INTA), 7620 Balcarce, Argentina
bConsejo Nacional de Investigaciones Cientı ´ficas y Te ´cnicas (CONICET), Argentina
cFacultad de Ciencias Veterinarias, Universidad Nacional del Nordeste (UNNE), Corrientes, Argentina
dInstituto Nacional de Tecnologı ´a Agropecuaria (INTA), Mercedes, Corrientes, Argentina
Received 9 April 2007; received in revised form 5 September 2007; accepted 5 September 2007
Abstract
The aim of the present work was to describe the occurrence of antibodies to Neospora caninum in water buffaloes on four
ranches located in Corrientes province in the northeast of Argentina. Antibodies against N. caninum were determined in sera of 449
water buffaloes by using an indirect fluorescent antibody test (IFAT). A Bayesian logistic regression mixed model was used to
quantify the strength of association between positive serological results to N. caninum and gender, age and category (calf, steer,
heifer, cow) as risk factors. Antibody titers were found in 287 (64%) buffaloes. All ranches had seropositiveanimals. Agewas more
strongly associated with positive results to N. caninum (OR: 1.4; CI 95%: 0.86–2.22) than gender (OR: 1.02, CI 95%: 0.40–2.59)
and category (OR: 0.88, CI 95%: 0.57–0.88). Results suggest a high exposure of water buffaloes to N. caninum by postnatal
transmission in these four ranches located in Corrientes province, Argentina. Further studies are needed to quantify the
consequences of Neospora-infections in the water buffalo industry.
# 2007 Elsevier B.V. All rights reserved.
Keywords: Neospora caninum; Bubalus bubalis; Argentina
1. Introduction
Neospora caninum, a coccidian parasite that has
domestic dogs and coyotes as its definitive hosts
(McAllister et al., 1998; Gondim et al., 2004), causes
serious disease in cattle and dogs worldwide (Dubey,
2003). Evidence of Neospora-infections has been
reported in many other domestic and wild species
including water buffaloes (Bubalus bubalis) (Dubey
etal., 1998; Huong et al., 1998). Detection of antibodies
to N. caninum in buffaloes has been reported in Brazil
(Fujii et al., 2001; Gennari et al., 2005); Egypt (Dubey
et al., 1998); Italy (Guarino et al., 2000) and Vietnam
(Huong et al., 1998). Because different cut-off points
were used in these studies, it is difficult to compare the
seroprevalence of antibodies against N. caninum in
these countries. The reported percentage of exposed
animals varied from 0 (0/40) in China (Yu et al., 2007)
to 70.9% (139/196) in Brazil (Gennari et al., 2005).
According tonational records, approximately80,000
head of water buffalo are raised under extensive
conditions in wet areas of the northeast of Argentina
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Veterinary Parasitology 150 (2007) 155–158
* Corresponding author at: Veterinary Pathology, INTA Balcarce,
CC 276, 7620 Balcarce, Argentina. Tel.: +54 2266 439100x20;
fax: +54 2266 439101.
E-mail address: ccampero@balcarce.inta.gov.ar (C.M. Campero).
0304-4017/$ – see front matter # 2007 Elsevier B.V. All rights reserved.
doi:10.1016/j.vetpar.2007.09.002

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(NEA) (SAGPYA, 2007). Water buffalo are better
adapted to wet tropical and subtropical environments
than cattle. The buffalo industry is increasing in the
NEA area, with an impact on the regional economy.
Bovine neosporosis has been reported as a cause of
abortions in dairy and beef cattle in Argentina
(Campero et al., 1998, 2003; Moore et al., 2002,
2003). In contrast, N. caninum has not been investigated
as a cause of abortion in water buffaloes in this country.
The aim of this study was to determine the
occurrence of antibodies to N. caninum in buffaloes
from herds in the province of Corrientes, Argentina.
2. Materials and methods
2.1. Experimental animals and blood samples
Blood samples were collected from the jugular vein
from 449 Mediterranean and Murrah crossbreed water
buffaloes (B. bubalis), from 1 to 20 years of age, from
four ranches located in Corrientes province, Argentina.
Approximately 40% of the buffalo population from
each ranch was bled. The animals were divided into
three age groups: <3 years, 3–5 years and >5 years.
Data regarding gender and category (calf, steer, heifer,
and cow) were also recorded when available. Repro-
ductiveparameters in the four buffalo herds werewithin
the normal range suggested by SAGPYA (2007) having
pregnancy rates of about 80%, calving rates of about
75% and weaning rates of >70% (data not shown). Sera
were kept at ?20 8C until analysis.
2.2. Serological analysis
N. caninum antibody levels were measured by an
indirect fluorescent antibody test (IFAT) (Fujii et al.,
2001) on tachyzoite slides made with the NC-1 N.
caninum strain (kindly provided by Dr. MC Venturini,
La Plata Veterinary College, Argentina). Rabbit anti-
bovine IgG labeled with fluorescein isothiocyanate
(Sigma, St. Louis, MO) was used as conjugate. The
samples were tested at a dilution of 1:100 (Rodrigues
et al., 2004). Positive and negative control sera from
buffalo were kindly provided by Dr. S.M. Gennari,
Veterinary College, Sa ˜o Paulo University, Brazil. The
IFAT was considered to be positive when peripheral
staining patterns typical of tachyzoites were observed.
2.3. Statistical analysis
A Bayesian logistic regression mixed model was
used to quantify the strength of the association between
positive serological results for N. caninum and gender
(male, female), age (years), and category (calf, steer,
heifer, cow) of the sampled animals. A hierarchical
model was necessary because animals within the same
herd are more likely to have a similar probability of
being positive to N. caninum than animals in different
herds. Risk factors (gender, age and category) were
included into the model as fixed effects. A random
effect was included to account for herd level factors not
assessed in this study. We used diffuse prior distribu-
tions oftheforms
Gamma(0.5,0.0005)] for the regression coefficients
andthe randomeffects, respectively.Themodelwasrun
using the Win-BUGS software (Spiegelhalter et al.,
1996), with 20,000 iterations after 500 iterations were
discarded. We used a stepwise procedure to succes-
sively include variables into the model. The odds ratio
(OR), calculated as the exponent of the posterior
distribution of the regression coefficients, was used to
estimate the strength of the association between each
factor and positive status to N. caninum. The combina-
tion of variables that resulted in the lowest value of the
deviance inference criterion (DIC) was considered the
model that best fit the data (Spiegelhalter et al., 2002).
N(0,4)andN[0,d?
3. Results
The distribution of age groups was as follows: <3
years—56 animals, 3–5 years—227 animals, >5
years—166 animals. There were 437 females (209
cows, 228 heifers) and 12 males (10 steers and 2 calves
with more than 6 months of age).
Specific antibodies were found in 287 (64%)
buffaloes. All farms had seropositive animals. The
proportion of seropositive animals on each farm is
shown in Table 1. The distribution of N. caninum
seropositive animals according to age groups is shown
in Fig. 1.
Age was more strongly associated with positive
resultstoN.caninum(OR:1.4;CI95%:0.86–2.22)than
gender (OR: 1.02, CI 95%: 0.40–2.59) and category
(OR: 0.88, CI 95%: 0.57–0.88). Moreover, the model
C.M. Campero et al./Veterinary Parasitology 150 (2007) 155–158 156
Table 1
Occurrence of antibodies by IFAT against N. caninum in water
buffaloes on four ranches in Corrientes province, Argentina
Farm%Seropositive animalsTotal
A
B
C
D
56.5
63.7
75.5
60.6
115
138
102
94

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that best fit (DIC: 586.2) the serological status to N.
caninum only included age as a covariate (OR: 1.3, CI
95%: 0.96–1.7), suggesting that age was the most
important factor contributing to the serological status to
N. caninum in the populations of buffaloes assessed in
this study.
4. Discussion
This report demonstrates a high exposure of water
buffaloes to N. caninum in four ranches located in
Corrientes province, Argentina. These results agree
with those reported by Fujii et al. (2001) and Gennari
et al. (2005) from Brazil, who found a prevalence of
antibodies to N. caninum >60% in 222 and 196 water
buffaloes, respectively using IFATwith a serum dilution
of 1.25. When a serum dilution of 1:200 was used in
IFAT, the prevalence of antibodies was 34.6% in 1377
animals tested in Italy (Guarino et al., 2000). Only 1.5%
of 200 animals tested seropositive in Vietnam (Huong
et al., 1998); however, a serum dilution of 1:640 was
usedinthe IFAT.BecauseNeosporawas isolated from5
of 6 water buffalo testing seropositive by IFAT using a
serum dilution of 1:100 (Rodrigues et al., 2004), this
dilution factor was used in the present work.
The probability of being seropositive to N. caninum
increased 30% per year in the populations of buffaloes
assessed in this study. Older animals are expected to
have a higher probability of cumulative exposure to
infectious agents compared with younger animals. No
association between gender or category and serological
status to N. caninum was detected, suggesting that the
age association estimated in this study may simply be a
reflection of the yearly increment in the probability of
exposure to the parasitic oocysts. A similar association
between seroprevalence of antibodies against N.
caninum and age was found in buffalo populations
from southern Italy (Guarino et al., 2000) and south-
easternBrazil(Fujiietal., 2001).Ontheotherhand,this
association was not found in water buffaloes from
northernBrazil(Gennari et al., 2005). Although vertical
transmission occurs in buffaloes (Rodrigues et al.,
2005) and it is a frequent and efficient route of infection
in cattle (Dubey, 2003), it is unclear which route of
infection is most important in buffaloes.
Finally, although two aborted buffalo foetuses
showed non-suppurative encephalitis and non-suppura-
tivemyocarditis with Neospora-like cystsintheir brains
(Guarino etal., 2000),itisnotclear whetherN.caninum
isa frequent agent ofabortion inbuffaloes. In this study,
a high occurrence of antibodies against N. caninum was
associated with normal reproductive records in all four
ranches (data not shown). We have reported and
identified N. caninum as a cause of bovine spontaneous
abortions (Campero et al., 2003); however, no
information exists concerning reproductive losses in
buffaloes. Although more studies should be done to
clarify the consequences of Neospora-infections in
water buffalo, the epidemiological role of infected
buffaloes and the relationship with bovine neosporosis
should be also investigated.
Acknowledgments
Authors express their appreciation to Miss Marı ´a R.
Leunda and Mrs. Susana Mendez for their technical
assistance. The authors also thank Dr. Solange M.
Gennari from the Veterinary College, Sa ˜o Paulo
University, Brazil for kindly providing positive and
negative control buffalo sera and Dr. Douglas C.
Hodgins from the Department of Pathobiology, Ontario
Veterinary College, University of Guelph, Canada for
help in preparation of this paper.
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