toxoplasmosis in farm animals

1. Invited Review
Toxoplasma gondii prevalence in farm animals in the United States
D.E. Hill ⇑
, J.P. Dubey
United States Department of Agriculture, Agricultural Research Service, Beltsville Agricultural Research Center Animal Parasitic Diseases Laboratory, BARC-East,
Beltsville, MD 20705, USA
a r t i c l e i n f o
Article history:
Received 20 July 2012
Received in revised form 21 September 2012
Accepted 30 September 2012
Available online 28 November 2012
Keywords:
Toxoplasma gondii
Food safety
Pigs
Lambs
Goats
Chickens
a b s t r a c t
Toxoplasmosis, caused by Toxoplasma gondii, is one of the most common parasitic infections of humans
and other warmblooded animals. It has been found worldwide and nearly one-third of humans have been
exposed to the parasite. Congenital infection occurs when a woman becomes infected during pregnancy
and transmits the parasite to the foetus. Besides congenital infection, humans become infected by ingest-
ing food or water contaminated with sporulated oocysts from infected cat faeces or through ingestion of
tissue cysts in undercooked or uncooked meat. Food animals (pigs, chickens, lambs and goats) become
infected by the same routes, resulting in meat products containing tissue cysts, which can then infect
consumers. Toxoplasma infection is common in food animals in the United States. Implementation of
management factors such as biosecure confinement housing are important in reducing the levels of infec-
tion in animals destined for human consumption.
Published by Elsevier Ltd. on behalf of Australian Society for Parasitology Inc.
1. Introduction
Toxoplasmosis continues to be a significant public health prob-
lem in the United States (US). It is estimated that 1,075,242 per-
sons are infected with Toxoplasma gondii and approximately
2,839 persons develop symptomatic ocular disease annually (Jones
and Holland, 2010). In the US, various surveys have found that 10–
50% of the adult population has been exposed to T. gondii (Dubey
and Beattie, 1988; Jones et al., 2001, 2003, 2007; Dubey and Jones,
2008). The cost of illness in the US caused by Toxoplasma has been
estimated to be nearly US $3 billion and an annual loss of 11,000
quality-adjusted life year (QALY) (Batz et al., 2012; Hoffmann
et al., 2012). Recent publications have linked suicide and schizo-
phrenia to Toxoplasma infection (Pedersen et al., 2012; Torrey
et al., 2012).
The relative contribution of foodborne (meat) sources of Toxo-
plasma infection versus oocyst transmission of Toxoplasma to hu-
man infection is unknown, and various studies have suggested
widely disparate estimates of foodborne transmission. Mead
et al. (1999) suggested that T. gondii is one of three pathogens (to-
gether with Salmonella and Listeria) which account for >75% of all
deaths due to foodborne disease in the US Roghmann et al.
(1999) suggested that 50% of Toxoplasma infections in the US could
be ascribed to foodborne transmission. Scallan et al. (2011) esti-
mated that Toxoplasma caused 8% of hospitalizations and 24% of
deaths resulting from foodborne illnesses. In contrast, Dubey
et al. (2005), in a nationwide survey of retail meats (beef, chicken
and pork) found no viable Toxoplasma in any beef or chicken sam-
ples, and seven positive pork samples out of 2,094 samples assayed
from each type of meat, concluding that there was not enough via-
ble Toxoplasma present in retail meats to account for the level of
Toxoplasma infection found in the US population. Recent studies
(Boyer et al., 2011; Hill et al., 2011) have suggested that oocyst
exposure is the predominate route of Toxoplasma transmission in
the US.
Despite the uncertainty of human infection sources, Toxoplasma
is recognised as a foodborne risk and infection is common in many
domesticated animals used for food in the US, including pigs,
chickens, lambs and goats. Animal infections with Toxoplasma,
especially infections in non-meat eating ruminants, birds and pigs
raised in confinement, likely result from environmental exposure
to T. gondii oocysts. Oocyst contamination of the environment is
widespread as a result of faecal contamination of soil and ground-
water by the estimated 140 million domestic and feral cats in the
US, each of which can deposit hundreds of millions of oocysts in
faeces during infection (www.avma.org; Levy and Crawford,
2004; Dubey, 2010a). Oocyst contaminated runoff surface water
entering the marine environment has resulted in devastating dis-
ease in endangered sea otters off the west coast of the US (Miller
et al., 2002; Conrad et al., 2005), and even wild herbivores have
been shown to have very high seroprevalence as a result of expo-
sure to the environmentally resistant oocysts (Hill et al., 2005).
Most T. gondii isolates from human and animal sources in North
0020-7519/$36.00 Published by Elsevier Ltd. on behalf of Australian Society for Parasitology Inc.
http://dx.doi.org/10.1016/j.ijpara.2012.09.012
⇑ Corresponding author. Tel.: +1 301 504 8770; fax: +1 301 504 5306.
E-mail address: dolores.hill@ars.usda.gov (D.E. Hill).
International Journal for Parasitology 43 (2013) 107–113
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2. America have been grouped into one of three clonal lineages
including Types I, II and III (Dardé et al., 1992; Howe and Sibley,
1995; Ajzenberg et al., 2002a,b), and are biologically and geneti-
cally different from isolates from Brazil and Columbia, but similar
to isolates from Europe (Dubey et al., 2002a, 2007a; Lehmann et al.,
2006; Dubey and Su, 2009). Recent genotyping studies of isolates
from pigs, lambs and goats demonstrate that the Type II lineage
predominates in food animals in the US, followed by Type III iso-
lates and atypical genotypes; Type I isolates have rarely been found
in farm animals (Dubey et al., 2008a,b, 2011; Velmurugan et al.,
2009).
2. Surveillance in foods
Transmission of Toxoplasma from consumption of infected meat
products is difficult to quantify, since meat from infected animals
may undergo post-harvest treatments such as heating, freezing,
salting or pumping (injection of water and salt-based solutions to
retard microbial growth) that can render the parasite non-viable
(Hill et al., 2004, 2006), and few comprehensive assessments have
been completed in meat available for retail purchase. Complicating
matters is the fact that the number of T. gondii organisms in meat
from naturally infected food animals is very low, making the para-
site difficult and expensive to detect by direct methods. It is esti-
mated that as few as one tissue cyst may be present in 100 g of
meat (Dubey, 2010a). In addition, there is no predilection site for
Toxoplasma in meat animals; virtually all edible portions of an ani-
mal can harbour viable T. gondii tissue cysts (Dubey et al., 1986),
and tissue cysts can remain viable in food animals for years.
Beef, chicken and pork are the main meat types consumed in
the US In a case control study of 148 recently (<6 months) infected
individuals, Jones et al. (2009) identified elevated risks of infection
associated with eating raw ground beef, rare lamb, locally pro-
duced cured, dried or smoked meat, raw oysters, clams or mussels,
working with meat and drinking unpasteurized goat’s milk.
The relative risk to US consumers of acquiring T. gondii infec-
tion from undercooked meat was recently determined in a nation-
wide survey of retail chicken, beef and pork. The survey of 698
retail outlets in 28 metropolitan statistical areas (MSAs as defined
by the US Census Bureau) covered 80% of the US population. Only
pork was found to harbour viable T. gondii tissue cysts, which
were isolated from 0.38% of samples (7/2,094) by cat bioassay,
and 0.57% of pork samples were suspected to be infected based
on positive ELISA results (Dubey et al., 2005). No beef samples
were positive by bioassay or by ELISA, while 1.4% of chickens were
positive by ELISA only. The northeastern United States had a high-
er number of positive pork samples than other regions of the
country, reflecting the higher risk of pig infection due to regional
management practises (outdoor versus confinement rearing;
Dubey et al., 2005). Thus, while the extent of human infection
resulting from meat sources remains undetermined, the lack of
viable organisms in beef and chicken and the low prevalence of
T. gondii infection in market pigs found in this comprehensive
study would not account for the estimated incidence and mea-
sured seroprevalence in humans in the US.
3. Pigs
Serological surveys for Toxoplasma prevalence have been con-
ducted frequently in the US over the last 30 years. Of the major
meat animal species investigated thus far, pigs (and chickens) are
the only species shown to frequently harbour the parasite (Dubey
et al., 1991; (http://www.pork.org/FileLibrary/ResearchDocu-
ments/00-130%20-PATTONUofTenn.pdf); Dubey and Jones, 2008;
Hill et al., 2010). Dubey et al. (1991), in a nationwide survey con-
ducted in 1984, found 23% of market pigs and 42% of sows were
seropositive for Toxoplasma. Viable T. gondii tissue cysts were iso-
lated from 17% of 1,000 adult pigs (sows) from a slaughter plant
in Iowa, US (Dubey et al., 1995a), and from 51 of 55 (92.7%) market
weight pigs from the New England region of the US (Dubey et al.,
2002b). Serological surveys of pigs from Illinois (US) pig farms re-
vealed an infection rate of 3% in market weight animals, but higher
in breeding pigs (20%), suggesting that age is a factor for pigs
acquiring Toxoplasma infection (Weigel et al., 1995a,b). Serological
surveys on New England farms of pigs raised predominantly in out-
door conditions revealed an overall infection rate of 47% (Gamble
et al., 1999). The United States Department of Agriculture (USDA)
initiated the National Animal Health Monitoring System (NAHMS)
in 1983 to collect, analyse and disseminate data on animal health,
management and productivity in US domestic livestock popula-
tions, including swine (http://nahms.aphis.usda.gov; Losinger
et al., 1998). The program includes an on-farm serological sam-
pling component which can be used to monitor seroprevalence of
various pathogens including, since 1990, T. gondii. Sera and data on
management practises for this study are collected from breeding/
grower/finisher swine production sites located in 17 states
accounting for >90% of US swine production (Arkansas, Colorado,
Iowa, Illinois, Indiana, Kansas, Michigan, Minnesota, Missouri, Ne-
braska, North Carolina, Ohio, Pennsylvania, South Dakota, Texas,
Oklahoma and Wisconsin). Only sows were sampled in 1990.
Grower/finisher and sow/breeder populations were surveyed con-
currently in 1995 and 2000. In 2006 the swine biological sampling
targeted grower/finisher swine, which is the source of most fresh
pork consumed in the US The initial 1990 survey documented
nearly 20% seroprevalence of Toxoplasma in the US sow population.
Subsequent NAHMS surveys have shown a decline in Toxoplasma
seroprevalence in sows from 20% in 1990 to 15% in 1995, to 6%
in 2000 (Patton et al., 1996, 2002 (see URL above)). In contrast,
seroprevalence in grower/finisher swine has remained somewhat
stable over that period. The measured seroprevalence in 1995
and 2000 was 3.2% and 0.9%, respectively (http://www.pork.org/
FileLibrary/ResearchDocuments/00-130%20-PATTONUofTenn.pdf),
and 2.6% in 2006 (herd prevalence of 21.6%; Hill et al., 2010). Data
from the 2006 NAHMS for swine indicated that 145 of 185 farms
had no Toxoplasma-positive samples. A sampling of grower/finisher
populations in Tennessee, North Carolina and Illinois during this
same period revealed a seroprevalence of 1–3% (Assadi-Rad et al.,
1995; Dubey et al., 1995b; Patton et al., 1996). Risk factors associ-
ated with Toxoplasma infection identified during these surveys in-
cluded swine raised outdoors and exposed to organic material
potentially contaminated with cat faeces and oocysts, hogs raised
on small farms, and cats and other sylvatic reservoirs on the pre-
mises. The steep decline in seroprevalence in grower/finisher hogs
from 23% in 1984 (Dubey et al., 1991), to 3.2% in 1995 (NAHMS
Swine 1995), to 0.9% in 2000 (NAHMS Swine 2000; (http://
www.pork.org/FileLibrary/ResearchDocuments/00-130%20-PAT-
TONUofTenn.pdf), to 2.6% in 2006 (NAHMS Swine 2006; Hill et al.,
2010) likely reflects the increased use of confinement housing sys-
tems in the US pork industry; these studies indicated that pigs
raised indoors in biosecure environments have a lower level of
infection (Gamble et al., 1999; Dubey et al., 2002b). NAHMS 2006
data indicated that the odds of a farm being positive for Toxoplasma
were 7.7 times higher when grower/finisher pigs were not housed
in total confinement (Hill et al., 2010). While the reduced seroprev-
alence in sow populations probably resulted from the large-scale
movement of the swine industry towards total confinement rear-
ing (80% of sows are kept in confinement) and an emphasis on
facility biosecurity, the stable seroprevalence of 2.0% in grower/
finisher pigs may reflect gaps in adherence to good production
practises known to prevent exposure to Toxoplasma in confine-
ment-reared pigs. Boot hygiene has been shown to be essential
108 D.E. Hill, J.P. Dubey / International Journal for Parasitology 43 (2013) 107–113

3. in eliminating Toxoplasma infection in pigs raised in total confine-
ment, since infectious oocysts can be tracked into barns on con-
taminated footwear (Gamble et al., 2008, unpublished data). The
seroprevalence reported in live animals from the NAHMS 2006 sur-
vey is 4.5–6.8 times higher than the prevalence (0.38–0.57%) of
viable Toxoplasma in pork reported by the retail meat study of
Dubey et al. (2005), likely a result of post harvest treatments such
as heating, freezing, salting and pumping which are commonly
applied to fresh pork prior to sale and which reduce Toxoplasma
viability (Hill et al., 2004, 2006).
Toxoplasma gondii isolates (n = 168; designated TgPgUs15-182)
from pigs from several states in the US (Pennsylvania, Iowa, Mas-
sachusetts and New Jersey) were genotyped using 10 PCR-restric-
tion fragment length polymorphism (PCR-RFLP) markers (SAG1,
SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico)
(Velmurugan et al., 2009). Genotyping data from an additional 14
isolates collected from T. gondii-infected pigs in Maryland, US were
included for analysis. Nine genotypes (1–9) were recognised from
the 182 T. gondii isolates. Most (56%, 102) isolates were clonal Type
II (genotypes 1 and 2) and 27% (49) were clonal Type III (genotype
3) strains. Genotype 4 had Type II alleles, with the exception of
Type I alleles at loci Apico and L358. Eight isolates (genotype 5)
from Iowa had a combination of alleles I, II and III at different loci.
The remaining six isolates were divided into genotypes 6–9 and
had a combination of different alleles. Eight of the nine genotypes
were previously reported in different animal species and geo-
graphic regions. These data demonstrated that there is a predomi-
nance of clonal Type II and III strains in pigs in the US as well as a
few diverse, previously unrecognised T. gondii lineages circulating
in pigs used for human consumption.
Transmission of T. gondii to pigs on the farm occurs in a similar
manner to human infection: ingestion of oocysts from the environ-
ment; consumption of infected animals such as mice, birds and
other wildlife; consumption of undercooked meat scraps; and
through in utero transmission. In several studies of management
factors in the US, outside housing of swine, access of cats to swine,
infection in local cats and mice, and small herd size were positively
correlated with T. gondii infection (Assadi-Rad et al., 1995; Weigel
et al., 1995a,b; Zimmerman et al., 1990). Further, the use of traps,
bait, poison, an exterminator or some other method besides cats or
dogs for rodent control on the farm reduced the prevalence of Tox-
oplasma-positive results by 63% (Hill et al., 2010). Gamble et al.
(unpublished data) audited 58 pork production sites in the US
and documented persistent Toxoplasma infection in confinement-
reared grower/finisher swine on six of the sites. Increased risk of
infection was associated with the presence of domestic cats, feral
cats and wildlife, as well as poor practises for disposal of swine car-
casses. Increased compliance with production practises which re-
duced the risk of exposure of pigs, including the introduction and
stringent use of barn-only boots in infected production sites, re-
sulted in a reduction in Toxoplasma prevalence on infected farms,
with all but one non-compliant site becoming negative for T. gondii
infection after three production cycles.
The low tissue and seroprevalence reported by the 2005 retail
meat study (Dubey et al., 2005) does not support the contention
that pork contributes significantly to human infection in the US
However, over 110 million hogs are slaughtered for food in the
US each year (USDA, National Agricultural Statistics Service,
2011, http://usda01.library.cornell.edu/usda/nass/LiveSlau//
2010s/2011/LiveSlau-12-23-2011.txt). Based on the prevalence re-
ported from NAHMS 2006, it is possible that 2.8 million Toxo-
plasma-infected hogs could enter the US food chain each year. A
single market weight hog (113 kg) yields approximately 70 kg of
meat or 620 individual 113 g servings (http://www.pork.org/new-
sandinformation/quickfacts); even if one half of the harvested
meat is processed (salting, cooking, freezing, etc.) by methods
which are known to kill Toxoplasma, nearly 870,000,000 individual
113 g servings of Toxoplasma-infected pork could be available for
consumption in the US each year. As the annual per capita con-
sumption of pork in the US is 22.4 kg (49.4 lbs), ample opportunity
exists for exposure to Toxoplasma from infected pork and pork
products. Further, recent changes in USDA recommended cooking
temperatures for pork in the US from 160 °F (71.1 °C) to 145 °F
(62.7 °C) make exposure to viable, infectious bradyzoites more
likely (www.fsis.usda.gov/news/NR_052411_01/index.asp). Dubey
et al. (1990) demonstrated that although bradyzoites in tissue
cysts in ground pork were generally rendered non-viable by heat-
ing to 142 °F (61 °C) or higher temperature for 3.6 min, bradyzoites
did survive in one instance at 147 °F (64 °C) for 3 min. Home cook-
ing temperatures could also impact on risk, as one recent study
showed that 24% of pork samples prepared by consumers partici-
pating in the study were cooked at temperatures that did not reach
145 °F (http://foodrisk.org/default/assets/File/EcoSure%202007%20
Cold%20Temperature%20Report.pdf). Lethality studies on cooking
temperatures for killing of Toxoplasma in unground prime cuts of
pork have not been completed.
In the US, organically raised, ‘humanely raised’ and free range
(FR) pork products have become increasingly popular in recent
years, leading to the recruitment of pork producers to switch to
non-confinement rearing of hogs (Honeyman et al., 2006; Dimitri
and Greene, 2002). The US National Organic Program (NOP) stan-
dards (http://www.ams.usda.gov/nop/) require that all organically
raised animals must have access to the outdoors. Although ‘huma-
nely raised’ and ‘FR’ products have standards that are less strin-
gently defined, outdoor access is also considered a requirement
for labelling. These practises substantially increase the risk of
exposure of pigs to Toxoplasma. Some grocery retailers in the US
have developed specific animal welfare guidelines for producers
of animals, including pigs, whose meat is sold in their stores
(http://www.wholefoodsmarket.com/about-our-products/quality-
standards/animal-welfare-standards). Although not certified or-
ganic, animals raised using these guidelines are given access to
pasture. Recent studies have demonstrated the increased risk asso-
ciated with pork produced in FR/organic/outdoor management sys-
tems. Dubey et al., 2002b isolated Toxoplasma from 51 of 55 FR pigs
from a New England farm, and from 14 of 48 FR pigs from a farm in
Maryland (Dubey et al., 2008a). In one study, 22 of 324 (6.8%) FR
pigs in North Carolina were seropositive for T. gondii, while three
of 292 conventionally raised pigs (1.1%) were seropositive
(Gebreyes et al., 2008). Dubey et al. (2012) determined seropreva-
lence to T. gondii in organically raised pigs on two farms in the
upper midwest US Serum and tissue samples were collected from
pigs at slaughter, and ELISA and the modified agglutination test
(MAT) were positive in 30 of 33 animals (91%). Toxoplasma was iso-
lated from 17 pigs by mouse bioassay including one from a sero-
negative (both ELISA and MAT) pig. Genetic typing of 16 of the
17 T. gondii isolates revealed clonal Type II and Type III; no Type
I was found. These studies revealed a very high prevalence of T.
gondii in FR/organic pigs in the US, indicating a potentially in-
creased health risk of consuming pork products from FR/organic
systems.
The US feral pig population is estimated at five million animals
and is growing rapidly. Feral pigs are now found in at least 39
states due to natural range expansion and illegal movement of ani-
mals for hunting opportunities (USDA, APHIS Bulletin #799 (2005);
#2086 (2011), http://www.aphis.usda.gov/publications/wild-
life_damage/content/printable_version/feral%20pigs.pdf; http://
www.aphis.usda.gov/publications/wildlife_damage/content/print-
able_version/feral_swine.pdf; www.feralswinemap.org). Feral pigs
are frequently hunted for food and meat from these animals are of-
ten used to make cured products which would not involve cooking
or freezing to inactivate T. gondii tissue cysts. A cross-sectional
D.E. Hill, J.P. Dubey / International Journal for Parasitology 43 (2013) 107–113 109

4. serological survey was conducted to estimate the prevalence of T.
gondii and risk factors associated with infection in feral pigs in
the US (Hill et al., in press). Serum samples were tested from
3,247 feral pigs from 32 states. Maximum entropy ecological niche
modeling and spatial scan statistics were underutilised to predict
the geographic range and to investigate clusters of infection of T.
gondii in feral pigs. The overall seroprevalence of antibodies to T.
gondii, indicating infection, was 17.7%, although in some states
(Hawaii) seroprevalence was as high as 50%. Toxoplasma seroposi-
tive feral pigs were widespread across the south and midwest, and
more restricted in the arid west. Dubey et al. (1997) found similar
prevalence (18%) in 170 feral pigs from Georgia, while Diderrich
et al. (1996) found 34% of surveyed feral hogs from South Carolina
were seropositive for Toxoplasma. Thus, the potential for Toxo-
plasma infections in humans from eating undercooked meat from
feral pigs appears to be high.
Currently, there are no inspection programs for the testing and
detection of Toxoplasma in pigs at slaughter in the US Infected pigs
enter the food chain but cannot be tracked because there is no na-
tional system for identifying individual pigs slaughtered in the US
other than the identification system which has been established for
animals enrolled in the Trichinella Certification Program (Pyburn
et al., 2005). In addition, no regulations requiring that pork under-
go further processing are mandated to inactivate the parasite.
However, many of the methods that are in place for processing
pork for inactivation of Trichinella spiralis (US Code of Federal Reg-
ulations, Title 9, CFR318.10, http://ecfr.gpoaccess.gov; US Depart-
ment of Agriculture, Food Safety and Inspection Service, 2003)
are also effective for the inactivation of T. gondii. While these reg-
ulations remain in effect, processed pork products should be safe
for human consumption without further treatment. Meat from
other animals which are not required to undergo inactivation pro-
cedures as described for processed pork may still harbour viable
parasites. Unlike bacterial pathogens, Toxoplasma does not undergo
logarithmic growth in meats during prolonged storage at subopti-
mal temperatures. However, it is possible that ground meat con-
taining viable parasites could contaminate grinding equipment,
or become mixed with meat from several animals to produce large
quantities of infective meat product.
4. Chickens
Although chickens are naturally resistant to clinical toxoplas-
mosis, they are considered one of the most important hosts in
the epidemiology of Toxoplasma infection in many countries be-
cause they are an efficient source of infection for cats that excrete
the environmentally resistant oocysts, and because humans may
become infected with this parasite after eating undercooked in-
fected chicken meat. Toxoplasma was considered a neurotropic par-
asite based on studies in rodents, however findings from recent
studies indicate that T. gondii encysts in muscle more efficiently
than in the brain (Dubey and Beattie, 1988), making chicken meat,
the most commonly consumed meat in the US, a potentially signif-
icant source of infection. However, most chicken in US is cooled to
near freezing or is completely frozen at the packing plant (Chan
et al., 2001), which should kill organisms in tissue cysts (Kotula
et al., 1991). In the US, the per capita yearly consumption of poul-
try is estimated as 37.2 kg, and approximately 8.5 billion chickens
are killed for human consumption. The risk factors associated with
infection in chickens raised indoors is likely similar to the risk fac-
tors associated with confinement raised pigs, i.e., transport of oo-
cysts into animal facilities on contaminated footwear or
equipment; however, no studies have been completed which could
answer these questions. The prevalence of Toxoplasma infection in
chickens raised indoors in the US has been investigated in only a
few studies, which found low levels (3%) of infection (Jacobs
and Melton, 1966). In a recent survey of retail meats, viable T. gon-
dii was not isolated from any of the 2,094 chicken meat samples
obtained from grocery stores in the US (Dubey et al., 2005). Despite
the results of this study, infected chickens may still pose a risk of
infection for humans for several reasons. First, in this study, chick-
en breasts were selected for sampling due to the experimental de-
sign that called for testing 1 kg of boneless meat for each sample,
although the prevalence of T. gondii in chicken breast is lower than
in other tissues. In a previous study, Toxoplasma was isolated from
breast meat of only 18.6% of infected chickens (Dubey, 2010b). Sec-
ond, many of the chicken breasts had been injected with enhancing
solutions that have a deleterious effect on T. gondii viability (Hill
et al., 2004, 2006). Third, antibodies to T. gondii were found in
1.3% of the tissue fluids extracted from the breast meat using an
ELISA (Dubey et al., 2005). FR chickens play an important role in
the epidemiology of T. gondii in the rural environment, perhaps
more than rodents, because they are clinically resistant to T. gondii
and live longer than rodents. FR chickens are one of the best indi-
cators for soil contamination with T. gondii oocysts because they
feed from the ground, exposing them to infection with oocysts.
Cats fed naturally infected chicken tissues can shed millions of oo-
cysts (Dubey et al., 2002a). FR chickens are known to be infected,
often at high rates. Isolation of Toxoplasma by bioassay from small
numbers (11–20) of chickens from across the US (backyard opera-
tions on small farms in Mississippi, Montana, Texas, Ohio, Illinois,
Georgia and Louisiana (Gibson and Eyles, 1957; Eyles et al., 1959;
Foster et al., 1969; Dubey, 1981; Dubey et al., 2003, 2007b) re-
vealed high levels (27–100%) of infection in FR chickens. Free range
chickens have not yet been identified as a source of human infec-
tion in the US Although T. gondii has been isolated from ovaries
and oviducts of naturally infected hens (Jacobs and Melton,
1966; McCulloch, 1968. Toxoplasmosis review and assessment.
Proceedings, 72nd Annual Meeting of the US Livestock Sanitary
Association and the 11th Annual Meeting of the American Associ-
ation of Veterinary Laboratory Diagnosticians, New Orleans, Louisi-
ana. October 6–11, 1968, 503–516; Foster et al., 1969; Peixoto and
Lopes, 1990), raw hens’ eggs are extremely unlikely to transmit T.
gondii infection. Shelled eggs have not been found to be infected
with T. gondii (Jacobs and Melton, 1966) and are unlikely to be a
source of infection for humans.
5. Beef
Serological evidence of infection in cattle is equivocal, since
most serological tests perform poorly with cattle sera. Cattle are
resistant to infection and are not considered to be important hosts
for Toxoplasma (Dubey, 2010a). The ingestion of beef is not consid-
ered important in the epidemiology of T. gondii due to the observed
innate resistance of cattle to the parasite. In one study, an attempt
was made to isolate T. gondii from a single naturally-exposed beef
cow in the US (Dubey, 1992). This 500 kg cow was killed and 100–
500 g portions of its tissues were bioassayed in cats (500 g of each
tissue) and mice (100 g of each tissue). None of the 12 cats, fed
approximately 6 kg of beef, shed oocysts. Viable T. gondii was not
isolated from any of the edible tissues of the cow by bioassays in
mice but was isolated from a homogenate of intestine of the
cow. Additionally, there is little, if any, danger of T. gondii infection
by consumption of dairy products or from drinking cows’ milk and,
in any case, cows’ milk is generally pasteurised or even boiled prior
to sale (Dubey and Beattie, 1988). Although cattle can be infected
with the parasite, tissue stages are eliminated or reduced to unde-
tectable levels within a few weeks or months. In a recent nation-
wide survey of pork, chicken and beef available for purchase in
retail stores in the US, viable Toxoplasma was not found in 2,049
samples of beef assayed for infection, and all samples were nega-
tive for antibody against the parasite (Dubey et al., 2005). Recent
110 D.E. Hill, J.P. Dubey / International Journal for Parasitology 43 (2013) 107–113

5. reports have suggested that consumption of ground beef is a risk
factor for infection with Toxoplasma (Jones et al., 2009), however
the potential for cross contamination of ground beef with meat
from other species more likely to carry the parasite cannot be ruled
out since meat grinding and mixing equipment may not be cleaned
sufficiently between commodities.
6. Lambs
Although sheep and goats are important hosts of T. gondii in
some countries and pose a major risk for human exposure, they
are minor food commodities in the US However, goat meat is very
popular with many ethnic groups in the US Since sheep and goats
are typically raised outdoors, exposure to contaminated soil con-
taining oocysts is constant. It is thought that only a small percent-
age of lambs are congenitally infected (Dubey, 2010a). Serological
evidence of infection in ruminants is widespread, with sheep and
goats demonstrating high levels of infection and associated clinical
disease, especially abortion. However, of the ruminant species
commonly consumed in the US, lamb poses a much greater risk
of infection to humans than does beef. According to US Department
of Agriculture regulations, sheep 1 year old (without permanent
teeth) are classified as lambs and slaughtered for human consump-
tion, while older animals are classified as sheep and their meat
(mutton) is sold for pet food and export. In the US, lambs and sheep
are slaughtered in separate commercial slaughter facilities. Be-
tween 3 and 3.6 million lambs are slaughtered in the US for food
each year, and the per capita consumption of lamb meat in the
US is approximately 0.5 kg per year (NASS Agricultural Statistics,
2011, http://www.nass.usda.gov/Publications/Ag_Statistics/2011/
Chapter07.pdf; p. 25, Table 7–43).
Toxoplasma gondii was isolated four decades ago from two of 50
lamb chops from retail meat stores in California (Remington, 1968)
and from the diaphragms of eight of 86 (9.3%) sheep from a slaugh-
terhouse in Baltimore, Maryland (Jacobs et al., 1960). In one survey,
indirect haemagglutination antibodies to T. gondii were found in 8%
of 1,056 lambs in a California slaughterhouse (Riemann et al.,
1977). In another study, MAT antibodies were found in 65.5% of
1,564 ewes from 33 farms in the northwest US (Dubey and Kirk-
bride, 1989a). Malik et al. (1990) reported ELISA antibodies to T.
gondii in 42% of 345 lambs and 80% of adult sheep from five slaugh-
terhouses in the northeastern US Dubey and Kirkbride (1989b) iso-
lated T. gondii from eight of eight naturally-infected lambs from a
flock in South Dakota, US The lambs were from a flock that had
aborted due to toxoplasmosis. Toxoplasma gondii was found histo-
logically in 11 of 30 lambs that were born dead. Lambs that sur-
vived the first week after birth remained asymptomatic and were
bled when 3–4 months old; antibodies (MAT 1:1,024 or higher)
were found in 67 of 112 lambs. Eight of the lambs with MAT titres
of 1:4,096 or higher were slaughtered when they were 7 months
old. Toxoplasma gondii was isolated from the hearts of three, ton-
gues of seven, legs of eight, and lamb chops of seven. Lamb hearts
are edible and are available for sale in the US, and the habit of con-
suming lamb meat that is medium to rare in preparation assures
that viable organisms contained in the tissues are not killed during
cooking. A recent survey of hearts collected from 383 lambs raised
in the mid-Atlantic states in the US and butchered at a commercial
abattoir demonstrated that 104 (27%) of the lambs were infected
with viable Toxoplasma organisms, and this meat was offered for
sale to consumers at retail outlets (Dubey et al., 2008b). In total,
53 isolates of T. gondii were obtained from 68 of the seropositive
lambs (77.9%). Fifty-seven T. gondii strains representing 15 geno-
types were identified using 10 PCR-RFLP markers. Four lambs
had infections with two T. gondii genotypes. Twenty-six (45.6%)
strains belong to the clonal Type II lineage. Eight (15.7%) strains be-
long to the Type III lineage. The remaining 22 strains were divided
into 11 atypical genotypes. Phylogeneic analyses indicated that the
clonal Type II lineage and its closely related genotypes accounted
for 68% (39/57) of the isolates. The Type III lineage accounted for
14% (8/57) of the strains and was the second most prevalent geno-
type. No Type I isolates were found. These results indicated high
parasite prevalence and high genetic diversity of T. gondii in lambs
from Maryland, Virginia and West Virginia. This data is in agree-
ment with previous finding that the Type II and III lineages pre-
dominate in food animals in North America (Howe and Sibley,
1995). Recently, a mouse virulent atypical T. gondii genotype was
isolated from the tissues of an aborted lamb from a chronically in-
fected ewe during an abortion storm in Texas (Edwards and Dubey,
2012).
7. Goats
The 2011 inventory of meat goats in the US recorded more than
2.5 million head (http://www.nass.usda.gov/Publications/Ag_Sta-
tistics/2011/Chapter07.pdf, p. 38, Table 7–64). More than 850,000
goats are slaughtered in the US each year for food, and this number
is expected to grow as the immigrant population from areas where
goat meat is a diet staple continues to increase. In a recent study,
hearts of 234 goats obtained from a grocery store were examined
serologically for T. gondii infection using the MAT (Dubey et al.,
2011). Like lamb hearts, goat hearts are edible and are sold com-
mercially. The goats originated from farms in Maryland, Virginia
and Pennsylvania, US Antibodies to T. gondii were found in 125
(53.4%) of 234 goats. Toxoplasma was isolated from 29 of 112 goat
hearts bioassayed in mice. Genotyping of the 29 T. gondii isolates
using 10 PCR-RFLP markers from DNA obtained from cell culture
grown tachyzoites revealed 12 genotypes. Nine isolates were clo-
nal Type II lineage, four isolates had Type II alleles at all loci except
a Type I allele at the Apico locus, and four isolates were clonal Type
III. The remaining 12 strains were divided into nine atypical geno-
types, including five new and four previously identified genotypes.
There were no Type I strains isolated. This study showed the dom-
inance of atypical strains (55%) infecting goats in the US Only 31%
(9/29) of goats were infected with clonal Type II strains and this is
far below the percentage recovered from other domestic animals
studied in the US These results indicate high parasite prevalence
and moderate genetic diversity of T. gondii in goats. The dominance
of atypical, Type II and Type III lineages in goats in the US (and the
absence of Type I strains) is in agreement with findings in lambs
and pigs in the US (Dubey et al., 2008a,b; Velmurugan et al.,
2009). These studies suggest that goat meat can be an important
source for T. gondii transmission to humans.
Little is known of the excretion of T. gondii in goat milk, how-
ever Toxoplasma tachyzoites have been recovered from goat’s milk
(Dubey, 2010a,b) and milk from other mammals (Pettersen, 1984;
Powell et al., 2001). Tachyzoites survive exposure to pepsin and
trypsin, and are orally infectious (Dubey, 1998), suggesting that
consuming milk containing tachyzoites can result in clinical infec-
tion. Fatal toxoplasmosis in humans has been linked to the drink-
ing of unboiled, unpasteurized goat’s milk (Riemann et al., 1975;
Sacks et al., 1982; Chiari and Neves, 1984; Skinner et al., 1990),
and recent studies identified drinking unpasteurized goat milk as
a risk factor for recently-acquired toxoplasmosis in pregnant wo-
men in the US (Patton et al., 1990; Jones et al., 2009). Although pas-
teurisation will kill T. gondii in milk, unpasteurized, raw milk is
sold by small goat farmers, and goat cheeses made from raw milk
could be a source of T. gondii infection.
8. Future directions and trends
Toxoplasmosis continues to be a significant public health prob-
lem in the United States; more than 1 million people are newly
D.E. Hill, J.P. Dubey / International Journal for Parasitology 43 (2013) 107–113 111

6. infected each year. Seroprevalence in pigs and chickens from non-
biosecure management systems is high and clinical disease, espe-
cially abortion, and high seroprevalence occurs in sheep and goats.
Beef cattle do not appear to be important in the epidemiology of
Toxoplasma or in transmission to humans. Type II and Type III clo-
nal lineages and atypical genotypes predominate in food animals in
the United States. Although toxoplasmosis is estimated to have a
disease burden and economic impact comparable to that of campy-
lobacteriosis and salmonellosis, there are presently no explicit
monitoring programs to screen animals entering the food chain
and no standardized reporting of human toxoplasmosis between
different countries. Dubey et al. (2005) have shown a minimal risk
of purchasing beef or chicken containing viable Toxoplasma tissue
cysts in the US The risk of purchasing infected pork was minimal
in most of the US (southern, western and midwestern regions),
while the risk in the northeastern states was 78% over a 10 year
period. The increasing demand for food safety, together with the
potential economic impact of legislation aimed at risk reduction,
has brought attention to the need for development and standardi-
zation of diagnostic tests for Toxoplasma infection. Such tests will
need to provide an accurate estimate of risks of transmission of
Toxoplasma to humans and must perform with comparable speci-
ficity and sensitivity across a range of animal species.
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