Toxoplasmosis: beyond animals tohumansYaowalark Sukthana

Faculty of Tropical Medicine, Mahidol University, 420/6 Rajvithi Road, Bangkok 10400, Thailand

The parasitic zoonosis toxoplasmosis, which was poorly

understood before the advent of the HIV epidemic, has

become a major clinical problem worldwide. Humans

acquire toxoplasmosis from cats, from consuming raw or

undercooked meat and from vertical transmission to the

foetus through the placenta during pregnancy. Studies of

the unique environmental factors in various commu-

nities indicate the important roles that eating habits and

culture have on the transmission of this infection. The

socioepidemiological aspects of toxoplasmosis are

thought to be important contributing factors for the

spread of this disease. Preventative measures should

consider the cultures and beliefs of people in various

communities more than solving poverty and giving

orthodox health education.

Toxoplasma gondii

Toxoplasma gondii can infect humans, and warm-bloodeddomestic and wild animals such as birds and rodents. In1908, Nicolle and Manceaux (from the Pasteur Institute inTunisia) isolated a new parasite from the African rodentCtenodactylus gundi, differentiated it from Leishmaniaand named it T. gondii a year later [1]. The first congenitalcase of toxoplasmosis was described in 1923 and the firstadult case was diagnosed in 1940 [2]. However, its lifecycle was not known until 1969 [1]. The development ofthe dye test by Sabin and Feldman in 1948 [2] was animportant milestone and it is now the ‘gold standard’serological method for diagnosing toxoplasmosis [3].

Cats and other felids are the only definitive hosts ofToxoplasma in which sexual reproduction occurs toproduce infective oocysts. Warm-blooded animals,including humans, are intermediate hosts that harbourtissue cysts in their bodies. Although asymptomatic innormal hosts, T. gondii can cause severe disease inimmunodeficient subjects. Since the HIV–AIDS pandemic,concurrent Toxoplasma infection has become an import-ant health problem, with its frequency increasingworldwide during the 1980s. Patients with toxoplasmicencephalitis were first documented in Thailand in 1992,and the number of cases has been increasing annually,particularly in the northern part of the country [4].

Early diagnosis is, therefore, crucial. However, diagnosisof toxoplasmosis is not straightforward because ofpleomorphic clinical manifestations. Laboratory diagnosisis important in primary infections, in pregnant women and

Corresponding author: Sukthana, Y. (tmymv@mahidol.ac.th).Available online 30 January 2006

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in congenital toxoplasmosis, whereas clinical manifestationsare more likely to be obvious in late reactivated cases (e.g.ocular toxoplasmosis) and in immunocompromised individ-uals. Practical strategies for investigating and interpretingresults should be focused for each of the clinical groups.

Diagnosis and management of congenital infection

The diagnosis of congenital toxoplasmosis relies mainly onfinding specific antibodies in patient serum and on theisolation of T. gondii DNA from amniotic fluid. Manyserological screening methods can detect IgG and IgM thatarespecific forT.gondii.Whenconfirmationof initial serologyis required, ranges of secondary tests are available,including the Sabin–Feldman dye test and a test for specificIgM or other immunoglobulin such as IgA, IgE and IgGavidity [5,6]. The sensitivity and specificity of screeningmethods range from 95.6% to 100% and from 94.8% to 99.8%,respectively [7–10], and they are commercially available andeasy to perform. However, they should be used with cautionfor screening pregnant women living in an area with a lowprevalence of Toxoplasma infection, such as in Thailand.Sukthana et al. [10] found that the commercial latexagglutination test showed 100% sensitivity and 94.8%specificity compared with the Sabin–Feldman dye test butits positive predictive value was only 71.3%. Therefore, thenumber of false seropositive cases would be higher than itought to be and clinicians would miss a certain number ofseronegative women who should be given preventativemeasures to protect them from T. gondii infection duringpregnancy. Therefore, confirmation of seropositivityis needed.

The immunological response to T. gondii is unique in thatIgM appears first, approximately one or two weeks afterinfection, closely followed by IgA and IgE [11]. In most cases,these acute-phase immunoglobulins peak after approxi-mately two months. The time at which IgM can no longerbe detected varies depending on the sensitivity of the assayemployed but it is usually between six and nine months afterinfection. IgG appears after IgM and reaches maximal levelsafter four months, then declines to lower levels over the next12–24months,whereasIgGpersistsata low-titre level for theremainder of the subject’s life [11]. Based on all recent dataand antibody dynamics, pregnant women with high positivetitres for Toxoplasma IgG and IgM, or seroconverted motherswill be offered additional testing using IgA, IgE or IgG avidityto try to determine the time of primary infection moreaccurately [5,6,12,13]. Low avidity is indicative of recentinfection. However, the current consensus is that high IgG

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. doi:10.1016/j.pt.2006.01.007

* J.M. Miro, et al., abstract 796, 10th World AIDS Conference on Retroviruses andOpportunistic Infection, Boston, February 2003.

Review TRENDS in Parasitology Vol.22 No.3 March 2006138

avidity can be used to rule out recent infection. An IgG avidityindex higher than 0.300 indicates an infection that wasacquired more than four months before testing [14,15].

In 2001, a multicentre evaluation of primary T. gondiiinfection in pregnant women from 20 European referencecentres recommended that a combination of two assays,each 95% specific, would reach a net specificity of 99.75%.A pair of positive results would be regarded as a positivediagnosis. Furthermore, the best assay combination wasthe measurement of IgM and IgG avidity, which gave adiagnostic result with specificities of w99% while main-taining diagnostic sensitivities at 95% or higher [6].

In the past decade, the use of PCR has facilitated majorimprovement in the diagnosis of many parasitic infections,including toxoplasmosis. T. gondii DNA can be detected inamniotic fluid, foetal tissue, blood, cerebrospinal fluid(CSF) and other clinical specimens. Primers are selectedfrom either the P30 or the B1 gene [16–18]. The sensitivityof amniotic fluid PCR can be increased from 81% to 91%when combined with mouse inoculation [17].

The routine prenatal screening of all pregnant womenremains controversial and the cost-effectiveness must betaken into account, especially in areas with a low prevalenceof toxoplasmosis. Although the screening of all pregnantwomen would decrease the incidence of congenitaltoxoplasmosis compared with targeted screening only whenfoetal anomalies are noted, it would cause 18.5 pregnancylosses from invasive prenatal testing for each child withcongenital toxoplasmosis prevented. Even if all foetusesidentified as being affected were terminated, there would stillbe 12.1 pregnancy losses for each case of congenitaltoxoplasmosis avoided [19]. Consequently, apart form Franceand Austria, postnatal or neonatal screening is preferred incountries with lower prevalences [20–22].

Recently, the management of congenital toxoplasmosishas improved because the early diagnosis of Toxoplasmainfection in mother, foetus and newborn can be made by acombination of serology and PCR methods [23]. Therefore,the treatment of congenital infection is now initiated beforebirth. However, the prevention of infection in pregnantwomen should be emphasized by using education abouthygiene and avoidance of the risks of infection. The more timethat elapses after maternal seroconversion and before thestart of treatment, the greater the risk of congenital infection.In 2001, Gilbert et al. [24] suggested that treatment startingwithin four weeks after maternal seroconversion did notincrease the risk of congenital infection, whereas it was 1.29and 1.44 times higher if treatment commenced between fourand seven weeks, and more than eight weeks, respectively.When foetal infection is proven, treatment by pyrimethamineand sulfadiazine is recommended until birth. Folinic acidshould always be administered to prevent bone marrowsuppression. Therapy using pyrimethamine and sulfona-mides is continued immediately after birth in infants who areknown, from the prenatal diagnosis, to be infected. Alterna-tive drugs include spiramycin and clindamycin for patientswith early pregnancy. A clinical profile of the infant iscompleted by ophthalmological examination and transfonta-nelle ultrasound scan.

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Diagnosis and management of toxoplasmic reactivation

in HIV–AIDS

For clinically suspected central nervous system (CNS)toxoplasmosis in AIDS patients, the Centers for DiseaseControl and Prevention (http://www.cdc.gov/) criteria shouldbe applied for presumptive diagnosis. These criteria consistof: (i) the recent onset of a focal neurological abnormality thatis consistent with intracranial disease or reduced conscious-ness; (ii) evidence from brain imaging of a lesion with masseffect or a lesion that appears on a radiograph after injectionof a contrast medium; and (iii) positive serum antibody to T.gondii or successful response to treatment for toxoplasmosis[25]. Computerized tomography and magnetic resonanceimaging are preliminary diagnostic tools for clinicians,although neuroimaging often cannot differentiate cerebraltoxoplasmosis from tumours such as lymphoma. Single-photon emission computerized tomography could provide amore precisediagnosis but it is costlyand not widelyavailableat present [26,27].

Serum and intrathecal levels of T. gondii antibody arealways low [28] and parasite isolation from blood and CSFis successful in !40% of CNS toxoplasmosis cases.However, in suspected cases, even a low titre of T. gondiiantibody aids the diagnosis. By contrast, when T. gondiiantibody is negative, toxoplasmic encephalitis is unlikelyto have occurred. Brain biopsy is often impracticable,although direct tissue staining with haematoxylin–eosinand enhanced by immunocytochemistry could beapplied [29].

DNA-amplification-based techniques greatly contrib-ute to the diagnostic improvement. Blood PCR as a singletest is not sensitive; CSF PCR has a higher sensitivity(50–100%) and specificity (97–100%). Repeated testingand combining both CSF and blood PCR enhancesensitivity [30–32]. Stage-specific oligonucleotide primerscould provide a more precise laboratory diagnosis ofreactivated toxoplasmic encephalitis, especially in recur-rent cases [33,34].

The treatment of suspected toxoplasmic encephalitisusing a combination of pyrimethamine and sulfadiazinewith folinic acid is effective. Unfortunately, up to 40–50%of patients treated develop adverse effects that require achange of therapy. Clindamycin is an alternative drug inthe case of intolerance to sulfa- compounds [35]. Mainten-ance therapy using half the dose of therapeutic drugs toprevent recurrent toxoplasmic encephalitis is necessarybecause the available drugs are ineffective against thetissue cyst that could later be reactivated [35]. The use ofhighly active antiretroviral therapy (HAART) suppressesthe HIV viral load and improves the CD4CT-cell count,followed by a strong reduction of opportunistic infections,including toxoplasmic encephalitis. The influence ofHAART in reducing toxoplasmic encephalitis has beenconfirmed in a randomized, controlled clinical trial andthere is no increase in risk of developing this disorder evenif drug prophylaxis is discontinued* [36].

Table 1. Seropositivity rates in Europe, the Americas and

Southeast Asia

Continents and

countries

Year Seropositivity (%) Refs

Western Europe

Austria 1998 43 [46]

Belgium 1997 50 [47]

France 2001 Up to 75 [39]

Germany 2004 26–54 [48]

Italy 2001 18–60 [39]

The Netherlands 2004 40.5 [49]

Spain 2004 28.6 [50]

Switzerland 1995 46 [51]

Scandinavia

Denmark 1999 27.8 [52]

Finland 1995 20.3 [53]

Norway 1998 10.9 [54]

Sweden 2001 14.0–29.4 [22]

Central and Eastern

Europe

Croatia 2000 38.1 [55]

Poland 2001 46.4–58.5 [56]

Slovenia 2002 34 [57]

UK 1998 57–93 [58]

Yugoslavia 1992 23–33 [59]

The Americas

USA 2004 16–40 [60]

Central America

Costa Rica 1996 76 [61]

Cuba 1993 60 [62]

Mexico 2001 35 [39]

Panama 1988 90 (at 60 years of age) [63]

South America

Argentina 2001 72 [39]

Brazil 2001 59 [39]

West Indies 1991 29.7 [64]

Southeast Asia

Indonesia 2000 58a [65]

Malaysia 2004 44.8 [66]

Thailand 1992, 1997,

2000, 2001

2.3–21.9 [67–70]

aMale:female ratioZ63:52.

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Epidemiology and transmission

Geographic distribution

T. gondii is found worldwide but its prevalence is unevenamong people of different countries in the same continent,such as those in Western and Central Europe and inSoutheast Asia (Table 1).

Transmission to humans

Humans become infected with toxoplasmosis mainly byeating uncooked meat that contains viable tissue cysts orby ingesting food or water contaminated with oocysts fromthe faeces of infected cats. A high prevalence of infection inFrance has been related to a preference for eating raw orundercooked meat, whereas it has been related in CentralAmerica to large numbers of stray cats in a climate thatfavours the survival of oocysts. A major concern is whetheracquired toxoplasmosis is mainly the result of consuming

Table 2. Percentage of Toxoplasma infection in humans associated

Country Beef (%) Pork (%) La

Belgium 6 2 10

Denmark 27 2 8

France ORZ5.5; (95% CI: 1.1–27.0) – OR

Italy 12.5 3 0

Norway 19 3 21

Switzerland 8 13 10

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infected meat or consuming food contaminated withoocysts from cat excreta [37].

A European multicentre study that included selectedcities in Belgium, Denmark, Italy, Norway, Switzerlandand the UK identified the consumption of undercookedmeat as the strong risk factor for acquiring a T. gondiiinfection [38]. Consuming raw pork and tasting raw meatduring meal preparation were the principal risk factors inPoland [39]. These risk factors have also been associatedwith seroconversion for T. gondii in case-control studies ofhealthy adults in France [40], Yugoslavia and the USA[39]. However, whereas the consumption of raw orundercooked meat was consistently identified as a riskfactor, the relative importance of the risk factor and thetype of meat associated with it varied among differentcountries (Table 2). These findings might reflect differ-ences in the eating habits of consumers or differentprevalences of infection in meat-producing animals inthe affected regions [39].

Domestic cats have a key role in the epidemiology ofT. gondii infection. Cats and other feline species canbecome infected with T. gondii either by ingestinginfectious oocysts from the environment or by ingestingtissue cysts from an intermediate host through feeding onfood scraps that contain meat or viscera of livestock orgame animals. Depending on the host species, thegeographic area and the season of the year, up to 73% ofsmall rodents and up to 71% of wild birds might beinfected with T. gondii [39]. The prepatent period in cats isshort and, after a primary infection with T. gondii,domestic cats can shed large numbers of oocysts into thehousehold, thereby putting their owners at risk ofinfection. Stray or domestic cats that are allowed toroam about could contaminate the environment withoocysts; this might affect livestock that will later beslaughtered for human consumption.

Antibody to T. gondii can be detected in up to 74% of theadult cat population, depending on the type of feeding andwhether the cats are kept indoors or outdoors [41].Seroprevalences are usually higher in stray or feral catsthan in cats living in an urban or suburban environment.Between 9% and 46% of pet cats in Europe, South Americaand the USA showed serological evidence of past exposureto T. gondii, whereas seroprevalences of T. gondii infectionhave been estimated to be 6–9% in Asia [39,41].

In Central and South America, where levels of T. gondiiinfection are high, transmission by consumption of tissuecyst can be excluded because meat is usually well cooked[41]. Access for cats to the outdoor environment, andfeeding cats with leftovers or with raw viscera and rawmeat were the risk factors for human infection in Mexicoand Brazil [42,43]. A study of residents and workers on

with types of meat consumed

mb (%) Salami (%) Refs

10 [38]

4 [38]

Z3.1; (95% CI: 0.85–14.00) – [40]

.5 12.5 [38]

3 [38,71]

5 [38]

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swine farms in Illinois (USA) showed that canine infectionwith T. gondii increases the risk of human infection andthat contact with soil is a probable mechanism oftransmission [44]. The increased risk of seropositivity inhuman males is attributed to less attention being paid tocleanliness in food preparation and consumption thanin females.

In Thailand, cats are kept as pets but they are allowedto roam freely outdoors. They are rarely trained todefecate in litter boxes and, thus, do so anywhere, fromthe backyard to the roof of the house. Buddhism is themajor religion in Thailand and it has a vital role in Thaisociety. Belief in the five precepts of Buddhism, one ofwhich states that killing is sinful, and practices such asnever killing, or allowing the killing of, unwanted petslead to unwanted pet dogs and cats being abandoned intemples, where monks are obliged to care for them(Figure 1). It is estimated that w20 000 dogs andw10 000 cats are left in 500 Buddhist temples in Bangkok(Thailand).

These animals can roam everywhere in temple groundsat any time, even when religious functions are beingperformed. They are fed with leftover food that, althoughplentiful, might not be hygienic. There is no regularveterinarian visit and no antiparasitic drugs are pre-scribed. The overcrowded conditions make Buddhisttemples a place of risk for acquiring zoonotic infection.

A seroepidemiological study of T. gondii in cats andtheir owners in the metropolitan area of Bangkok wasconducted involving community households and those inBuddhist temples, covering an area of 106.6 km2

containing 494 931 inhabitants [37]. Serum sampleswere collected from 327 household members, monks,novices and nuns living in the temples and from 315stray cats in the temple boundary. It was found that7.3% of the cats studied and 6.4% of the humansstudied were seropositive for Toxoplasma antibody. Therisk of Toxoplasma seropositivity in the exposed humangroup was five times that in the non-exposed group [OR(95% CI) Z5.43 (1.28–23.04): pZ0.01]. Of the studiedcats, up to 80% defecated anywhere, and in most cases(up to 75%) the excreta of the cats were not buried orremoved. Infected cats with unrestricted defecation

Figure 1. Abandoned cat and dog in a Buddhist temple in Bangkok (Thailand) being

looked after by an obliging monk. Photograph courtesy of Bangklao Nok Temple.

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would contaminate their immediate environment and,therefore, provided a potential source of humaninfection. Thus, it seems that cat ownership is a riskfactor for Toxoplasma infection in Thailand. Risk wasincreased in and around temples, particularly ifcourtyards were made of earth or grass, suggestingthat ground temperature is an important determinantof oocyst survival [37].

As mentioned, access for cats to the outdoor environ-ment, and feeding cats with leftovers or with raw visceraand raw meat are risk factors for human Toxoplasmainfection in Mexico and Brazil [42,43] but the situation isdifferent in Thailand. Thai cats eat rice and well-cookedfish but not raw meat. They also catch rodents in responseto hunting instincts but usually not for eating. This couldexplain why the prevalences of Toxoplasma infection incats and humans are low in Thailand, even though manyfactors seem to promote transmission. Moreover, Dubey[45] demonstrated that oocysts remain infective for onlyone minute at 608C compared with 54 months at 48C.Thus, oocysts from cat excreta deposited on hot roofs andstone or concrete courtyards are unlikely to be successfulin disease transmission [37].

Prevention

Although consumption of infected meat and closeassociation with infected cats are the two main sourcesof Toxoplasma infection in humans, details as to howthese are brought about differ in different countries oreven in different ethnic communities that belong to thesame country. Eating habits vary even among inhabi-tants of the West. For example, the distribution of beef,pork and lamb consumption in Europe is uneven(Table 2) and there are religious differences amongSoutheast Asian people (e.g. Buddhist or Islamic faith).Even the manner in which domestic cats are kept andfed in various communities and countries is not similar.Cats in Thailand are not kept indoors; they roam aboutand are fed on rice and cooked fish but not meat orlivestock viscera as in Mexico. Although the climate andground temperature in both countries are similar, theprevalence of T. gondii in Mexico is higher than inThailand. In Southeast Asia, the Toxoplasma seroposi-tivity rate is much higher in Surabaya (Indonesia), apredominantly Islamic community in which cats arebetter looked after than in Bangkok.

Addressing the problem of disease prevention should,therefore, take into account ethnic and cultural differ-ences. Proper hygienic measures when taking food andwhen keeping pets will reduce the chance of Toxoplasmatransmission to humans but ignorance and poverty are notthe only important factors that contribute to the highprevalence of zoonotic diseases. Increasing both the publicawareness of these diseases and the sense of responsibilityfor looking after pet animals is important. However, socialscience research strategies regarding the importance ofculture, tradition and beliefs in the transmission ofdisease are also necessary.

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