Toxoplasma gondii and Neospora caninum antibodies in dogs from Grenada, West Indies

Toxoplasma gondii and Neospora caninum are structurally similar parasites with many common hosts. The prevalence of antibodies to T. gondii and N. caninum was determined in sera from dogs in Grenada, West Indies. Using a modified agglutination test, antibodies to T. gondii were found in 52 (48.5%) of the 107 dogs, with titers of 1:25 in 17, 1:50 in 19, 1:100 in 7, 1:1,600 in 5, and 1:3,200 or higher in 4. Seroprevalence increased with age from 2.2% in dogs <6 mo old to 18.9% in dogs older than 2 yr, indicating postnatal transmission of T. gondii in this population of canines. There was no correlation between the health of the dogs and the seroprevalence or magnitude of the T. gondii titer. Antibodies to N. caninum were determined by the indirect immunofluorescent antibody test (IFAT). Two of the 107 dogs had N. caninum antibodies (IFAT titers 1:100 and 1:400); these dogs had T. gondii titers of 1:1,600 and 1:50, respectively. Results indicate that these 2 structurally similar protozoa are antigenically different.     

Seroprevalence of Toxoplasma gondii in cats from St. Kitts, West Indies

The prevalence of antibodies to Toxoplasma gondii was determined in sera from 106 domestic cats from St. Kitts, West Indies. Using a modified agglutination test, antibodies to this parasite were found in 90 (84.9%) of the cats, with titers of 1:20 in 23 cats, 1:40 in 34 cats, 1:80 in 18 cats, 1:160 in 2 cats, 1:320 in 1 cat, and 1:1,280 or higher in 11 cats. This is the first report of the prevalence of T. gondii infections in cats on St. Kitts and suggests widespread contamination of the environment with oocysts.     

Neospora caninum and Toxoplasma gondii antibodies in dogs from Durango City, Mexico

Toxoplasma gondii and Neospora caninum are structurally similar parasites, with many hosts in common. The prevalence of antibodies to T. gondii and N. caninum was determined in sera from dogs from Durango City, Mexico. Using a modified agglutination test, antibodies to T. gondii were found in 52 (51.5%) of the 101 dogs with titers of 1:25 in 27, 1:50 in 11, 1:100 in 5, 1:200 in 4, 1:400 in 2, 1:800 in 2, and 1:3,200 or higher in 1. Antibodies to N. caninum were determined by the indirect immunofluorescent antibody test (IFAT) and the Neospora sp. agglutination test (NAT). Two of the 101 dogs had N. caninum antibodies; these dogs did not have T. gondii antibodies, supporting the specificity of the tests used. The N. caninum antibody titers of the 2 dogs were: 1:400 by IFAT and 1:200 by NAT in 1, and 1:25 by NAT and IFAT in the other. Results indicate that these 2 structurally similar protozoans are antigenically different.     

Tissue distribution and molecular characterization of chicken isolates of Toxoplasma gondii from Peru

The prevalence of Toxoplasma gondii in free-ranging chickens is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. The prevalence of T. gondii antibodies in sera of 50 free-range chickens (Gallus domesticus) from Peru was 26% on the basis of the modified agglutination test (MAT). Hearts, pectoral muscles, and brains of seropositive (MAT > or =1:5) chickens were bioassayed individually in mice. Tissues from the remaining 37 seronegative chickens were pooled and fed to 2 T. gondii-free cats. Feces of cats were examined for oocysts; they did not shed oocysts. Toxoplasma gondii was isolated from the hearts of 10 seropositive chickens but not from their brains and pectoral muscles. Genotyping of these isolates using the SAG2 locus indicated that 7 isolates were type I and 3 were type III. Six of the 7 type-I isolates were avirulent for mice, which was unusual because type-I isolates are considered virulent for mice. The T. gondii isolates were from chickens from different properties that were at least 200 m apart. Thus, each isolate is likely to be different. This is the first report of isolation of T. gondii from chickens from Peru.     

Antigenic similarity between the coccidian parasites Toxoplasma gondii and Hammondia hammondi

The antigens that are present in the coccidian parasites Toxoplasma gondii and Hammondia hammondi were demonstrated and defined by using SDS-PAGE and immunoenzymatic techniques with 125I-labeled and unlabeled antigens of T. gondii and sera of mice infected orally or intraperitoneally with H. hammondi . All cell surface antigens of T. gondii that were labeled with 125I were recognized by antibodies in the sera of the mice infected with H. hammondi except the antigen of approximate molecular weight of 21.5 Kd. This suggests that this antigen is specific for T. gondii. Various antigens in the T. gondii-lysed antigen preparations were recognized by antibodies to H. hammondi . The number of recognized antigens increased as the infection of the mice with H. hammondi progressed. Oral infection with H. hammondi appeared to induce the formation of antibodies that recognized more T. gondii antigens than infection by intraperitoneal inoculation.     

First biologic and genetic characterization of Toxoplasma gondii isolates from chickens from Africa (Democratic Republic of Congo, Mali, Burkina Faso, and Kenya)

The prevalence of Toxoplasma gondii in free-ranging chickens (Gallus domesticus) is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. In the present study, prevalence of T. gondii in chickens from Democratic Republic of Congo, Mali, Burkina Faso, and Kenya is reported. The prevalence of T. gondii antibodies in sera of 50 free-range chickens from Congo was 50% based on the modified agglutination test (MAT); antibody titers were 1:5 in 7, 1:10 in 7, 1:20 in 6, 1:40 in 1, and 1:160 or more in 4 chickens. Hearts, pectoral muscles, and brains of 11 chickens with titers of 1:20 or more were bioassayed individually in mice; T. gondii was isolated from 9, from the hearts of 9, brains of 3, and muscles of 3 chickens. Tissues of each of the 14 chickens with titers of 1:5 or 1:10 were pooled and bioassayed in mice; T. gondii was isolated from 1 chicken with a titer of 1:10. Tissues from the remaining 25 seronegative chickens were pooled and fed to 1 T. gondii-free cat. Feces of the cat were examined for oocysts, but none was seen. The results indicate that T. gondii localizes in the hearts more often than in other tissues of naturally infected chickens. Genotyping of these 10 isolates using the SAG2 locus indicated that 8 were isolates were type III, 1 was type II, and 1 was type I. Two isolates (1 type I and 1 type III) were virulent for mice. Toxoplasma gondii was isolated by mouse bioassay from a pool of brains and hearts of 5 of 48 chickens from Mali and 1 of 40 chickens from Burkina Faso; all 6 isolates were avirulent for mice. Genetically, 4 isolates were type III and 2 were type II. Sera were not available from chickens from Mali and Burkina Faso. Toxoplasma gondii antibodies (MAT 100 or more) were found in 4 of 30 chickens from Kenya, and T. gondii was isolated from the brain of 1 of 4 seropositive chickens; this strain was avirulent for mice and was type II. This is the first report on isolation and genotyping of T. gondii from any source from these 4 countries in Africa.     

Serological survey of Toxoplasma gondii infection in free-ranging Eurasian lynx (Lynx lynx) from Sweden

To investigate the prevalence of Toxoplasma gondii infection in free-ranging Eurasian lynx (Lynx lynx) in Sweden, serosanguinous fluids and feces were collected from 207 carcasses of lynx killed or found dead from 1996 to 1998. Sera were tested for antibodies against T. gondii by the direct agglutination test, and 156 (75.4%) of the sera tested positive at antibody titers>or=40. Antibody prevalence was significantly lower in lynx originating from the northern parts of Sweden than in lynx from the more southern regions that are more densely populated by humans. Age-related differences also were found, with a significantly lower prevalence (55%) in juvenile (<1-yr-old) than in subadult and adult animals (82%). There was no significant difference in seroprevalence between males and females. Oocysts typical of T. gondii were not detected in any of the fecal samples.     

Seroprevalence of Toxoplasma gondii in wild toque macaques (Macaca sinica) at Polonnaruwa, Sri Lanka

From a natural population that inhabits the dry evergreen forest at Polonnaruwa, serum samples of 170 toque macaques were examined for antibodies to Toxoplasma gondii by the modified agglutination test. Of these, 21 (12%) were found with titers of 1:16 in 9, 1:32 in 9, 1:256 in 1, 1:1,024 in 1, and 1:4,096 in 1. There was no evidence of maternal transmission of antibodies or congenital toxoplasmosis. None of the infected macaques died within 1 yr after sampling. Toxoplasma gondii infection was closely linked to human environments where domestic cats were common. Macaques having frequent contact with human settlements showed a significantly greater (P < 0.0001) prevalence (19% infected) than macaques restricted to forest habitat, none of which was infected. Although infection with T. gondii has been noted in several species of Asian primates, this is the first report of T. gondii antibodies in toque macaques (Macaca sinica) that are endemic to the island of Sri Lanka.     

Serologic survey for Toxoplasma gondii and Neospora caninum in the common brushtail possum (Trichosurus vulpecula) from urban Sydney, Australia

The common brushtail possum (Trichosurus vulpecula) has well adapted to increasing urbanization, resulting in greater interaction with humans and their domestic pets. Wildlife species in urban areas face a higher risk of exposure to zoonotic pathogens and may be affected by parasites hosted by cats (Toxoplasma gondii) or dogs (Neospora caninum), yet it is unknown to what extent urban T. vulpecula are exposed to these parasites. Antibodies to T. gondii and N. caninum were assayed in sera of 142 adult possums from the city of Sydney, Australia. Using the modified agglutination test, antibodies to T. gondii were found in 9 (6.3%) of the 142 animals in titers of 1:25 (4), 1:50 (1), 1:100 (1), 1:800 (1), 1:3,200 (1), 1:6,400 (1), and 1:12,800 (1). Of some T. vulpecula multiple sera samples within a 2-yr frame could be collected, but seropositive animals in general were not recaptured after initial seroconversion. One possum had a high T. gondii titer on 2 consecutive bleedings, 14 mo apart, and seropositive possums appeared normal when captured. Sex seemed not to have an affect on antibody prevalence, but age and location may play a role. Antibodies to N. caninum were not detected in 1:25 dilution of sera in the N. caninum agglutination test, indicating that T. vulpecula may not have been exposed to this parasite. This is the first serological survey for T. gondii and N. caninum infections in urban T. vulpecula.     

The use of commercial immunoenzyme kits for the diagnosis of toxoplasmosis

The results of comparative study of sera obtained from donors and from several groups of patients with suspected toxoplasmosis are presented. The study has been carried out with the use of commercial enzyme immunoassay (EIA) kits: Toxoplasma gondii IgG EIA and Toxoplasma gondii IgM EIA manufactured by Labsystems (Finland), Sevatest ELISA IgG Toxo Micro I manufactured by Sevac (Czechoslovakia). Statistical processing of the results has confirmed the identity of these kits. The necessity of using evaluation criteria (the separation point, the scale for the interpretation of results) when working with the Sevac kits is emphasized. Comparative evaluation of antibody profiles in the sera under test suggests that the titer less than 1:1600 should be regarded as the separation point for these kits. IgM antibodies to T. gondii have been found only in 22% of patients with high titers of IgG antibodies.     

Prevalence of agglutinating antibodies to Toxoplasma gondii in adult and fetal mule deer (Odocoileus hemionus) from Nebraska

Toxoplasma gondii is an apicomplexan parasite of mammals and birds. Herbivores acquire postnatal infection by ingesting oocysts from contaminated food or water. Toxoplasma gondii infection is common in white-tailed deer, Odocoileus virginianus, but little is known about the prevalence of infection in mule deer, O. hemionus. We examined sera from 89 mule deer from Nebraska for agglutinating antibodies to T. gondii using the modified direct agglutination test (MAT) with formalin-fixed tachyzoites as antigen. Thirty-one (35%) of the samples were positive at dilutions of > or = 1:25. Samples were examined from 29 fetuses from these mule deer and none were positive in the MAT. Sera from 14 white-tailed deer from Nebraska were also examined and 6 (43%) were positive for T. gondii. Samples were examined from 5 fetuses from these white-tailed deer and none was positive in the MAT. Our results in both deer species from Nebraska are similar to studies conducted in white-tailed deer from other regions of the United States. Our findings indicate that mule deer are frequently infected with T. gondii and that mule-deer meat may be a source of human infection.     

Seroprevalence of Toxoplasma gondii antibodies among Atayal aboriginal people and their hunting dogs in northeastern Taiwan

Atayal aborigines, living at an altitude of 1,500-1,600 m in northeastern Taiwan, still hunt for wild animals with the help of hunting dogs. In this study, the latex agglutination test (LAT) was used to detect sera anti-toxoplasma antibodies in this community as a measure of their exposure to Toxoplasma gondii. The positive rates for sera anti-toxoplasma antibodies were 21.8% and 19.6%, respectively, in 422 Atayal and 51 hunting dogs tested. Neither of the positive rates were found to be significantly different between male (22.1%) and female Atayal (21.4%), or between humans (21.8%) and dogs (19.6%) (P > 0.05) when compared by the Chi-Squared test (chi 2-test). A significant difference was observed between the positive rates in adults (28.3%) and children (18.7%) (P < 0.05), and the age pattern of prevalence is consistent with an increasing duration of exposure to Toxoplasma gondii with age. The consumption of raw liver of wild animals or insufficiently cooked meat may be the major mode of transmission of toxoplasmosis in Atayal.     

Murine immune responses to recombinant Toxoplasma gondii antigens

Recombinant proteins of the RH strain of Toxoplasma gondii were produced by expression in Escherichia coli as glutathione S-transferase (GST) fusion proteins. Enzyme-linked immunosorbent assays were established using 2 of these fusion proteins termed H4/GST and H11/GST. The assays were able to detect antibodies in the sera of mice orally infected with either the cyst or oocyst stage of a pork isolate of T. gondii. In addition, the sera from mice infected with 1 of 4 different T. gondii isolates were investigated for their binding to these fusion proteins. Antibodies in the sera of all mice bound to H11/GST, but not all sera recognized H4/GST. Delayed-type hypersensitivity (DTH) responses to the fusion proteins were found when the mice were sensitized intradermally with H4/GST and H11/GST and challenged with the homologous fusion protein. However, no DTH response was recorded when mice were challenged with homologous fusion proteins after infection with T. gondii, or after immunization with a sonicate of the RH strain of the parasite. In addition, cellular responses were not stimulated against either of the fusion proteins in in vitro assays. These 2 fusion proteins were recognized by anti-T. gondii antibodies in experimental murine infections, and they are therefore potential candidates as antigens in assays for the diagnosis of human toxoplasmosis.     

Prevalence of antibodies to Toxoplasma gondii in the sera of rare wildlife in the Shanghai Zoological Garden, People's Republic of China

The prevalence of antibodies to Toxoplasma gondii in the sera of rare wildlife in the Shanghai Zoological Garden, PR China, was examined using a modified agglutination test (MAT) and an enzyme-linked immunosorbent assay (ELISA). Forty-one (35%) of 117 animals belonging to two classes, 10 orders, 18 families, 37 genera and 52 species (including sub-species) were sero-positive for MAT. By MAT, T. gondii antibodies were found in 11.1% (4/36) of birds, in 25% (4/16) of primates, in 69.4% (25/36) of carnivores and in 27.6% (8/29) of herbivores. Thirty-three (33.7%) of 98 animals tested by protein A ELISA were sero-positive. By ELISA, T. gondii antibodies were found in none of 36 birds, in 33.3% (4/12) of primates, in 87.1% (27/31) of carnivores and in 10.5% (2/19) of herbivores.     

Seroprevalence of Toxoplasma gondii antibodies in cats from Durango City, Mexico

The prevalence of antibodies to Toxoplasma gondii was determined in sera from 105 domestic cats from Durango City, Mexico. Using a modified agglutination test, antibodies to this parasite were found in 21% of the 105 cats, with titers of 1:25 in 3 cats, 1:50 in 4 cats, 1:200 in 5 cats, 1:400 in 2 cats, 1:800 in 2 cats, 1:1,600 in 4 cats, and 1:3,200 or higher in 2 cats. Cats older than 1 yr had a significantly higher frequency of infection than that found in cats younger than 0.5 yr (41 vs. 13.2%, respectively; odds ratio = 4.55; 95% CI = 1.24-17.18; P = 0.01). Overall, the seroprevalence of T. gondii antibodies in cats in Durango, Mexico, is much lower compared with those reported in other countries.     

Epitope-selected monospecific antibodies to recombinant antigens from Toxoplasma gondii reacted with dense granules of tachyzoites.

Epitope-selected monospecific antibodies were applied to investigate the localization of antigenic molecules in Toxoplasma gondii by immunoelectron microscopy. Eighty cDNA clones encoding antigenic polypeptides were immunoscreened from lambda gt11 expression library with T. gondii infected mouse sera. Twenty different clones with no crossreactivity were selected from eighty clones. Monospecific antibodies to antigens derived from respective cDNA clones extracted from infected mouse sera by the epitope selection method were used in Western blot analysis and immunoelectron microscopy. Eleven antigens were detected with epitope-selected antibodies in lysates of T. gondii tachyzoites. Five of the antigens with molecular weights of 60, 40, 35, 28, and 27 KD were localized in the dense granules. Monospecific antibodies purified by the epitope selection method were useful for investigating the localization of antigens without preparation of a monoclonal antibody from a hybridoma.     

Prevalence of antibodies to Trypanosoma cruzi, Toxoplasma gondii, Encephalitozoon cuniculi, Sarcocystis neurona, Besnoitia darlingi, and Neospora caninum in North American opossums, Didelphis virginiana, from southern Louisiana

We examined the prevalence of antibodies to zoonotic protozoan parasites ( Trypanosoma cruzi, Toxoplasma gondii, and Encephalitozoon cuniculi) and protozoans of veterinary importance ( Neospora caninum, Sarcocystis neurona, and Besnoitia darlingi) in a population of North American opossums ( Didelphis virginiana) from Louisiana. Samples from 30 opossums were collected as part of a survey for T. cruzi in Louisiana. Frozen sera from these 30 opossums were examined using an indirect immunofluorescent antibody test (IFAT) against in vitro-produced antigenic stages of these protozoans. Additionally, 24 of the 30 samples were examined using hemoculture, and all 30 were examined in the modified direct agglutination test (MAT) for antibodies to To. gondii. The prevalences of reactive IFAT samples were as follows: 60% for T. cruzi, 27% for To. gondii, 23% for E. cuniculi, 17% for S. neurona, 47% for B. darlingi, and 0% for N. caninum. Hemoculture revealed that 16 (67%) of 24 samples were positive for T. cruzi, compared to 18 of 30 (60%) by IFAT. The sensitivity and specificity for the IFAT compared to hemoculture was 100% for each. The modified direct agglutination test revealed that 9 (30%) of the 30 samples from opossums had antibodies to To. gondii , compared to 8 (27%) using the IFAT. The sensitivity and specificity of the IFAT compared to the MAT was 100% and 72%, respectively.     

A comparison of several serologic tests to detect antibodies to Toxoplasma gondii in naturally exposed bottlenose dolphins (Tursiops truncatus)

Toxoplasma gondii infection in marine mammals is intriguing and indicative of contamination of the ocean environment and coastal waters with oocysts. In a previous study, 138 of 141 (97.8%) bottlenose dolphins (Tursiops truncatus) from the coasts of Florida and California had antibodies to T. gondii by the modified agglutination test (MAT). Although the MAT has been found to be highly sensitive and specific for T. gondii antibodies from several species of terrestrial animals, it has not yet been validated for T. gondii infections in marine mammals. Furthermore, T. gondii has yet not been isolated from dolphins. In the present study, sera from 146 (60 from the 2004 samples and 86 from the 2003 samples) T. truncatus from the coastal areas of South Carolina and Florida were tested for antibodies to T. gondii. Sera from 2004 were tested by the MAT, the indirect fluorescent antibody test (IFAT), the Sabin-Feldman dye test (DT), an indirect hemagglutination test (IHAT), an enzyme-linked immunosorbent assay (ELISA), and Western blot. All 60 dolphins were seropositive, with MAT titers of 1:20 in 3, 1:40 in 19, 1:80 in 29, 1:160 in 2, 1:1,280 in 3, 1:2,560 in 2, and 1:5,120 or higher in 2, and these results were confirmed in another laboratory. The DT titers of these dolphins were <1:10 in 53, 1:800 in 3, 1:1,600 in 2, and 1:3,200 in 2. The IHAT titers were <1:64 in 52, 1:128 in 1, 1:512 in 2, and 1:2,048 in 5. The IFAT titers were <1:20 in 3, 1:20 in 11, 1:40 in 36, 1:80 in 2, 1:160 in 1, and 1:320 or higher in 7. All 7 DT-positive dolphins had high MAT titers, but 2 were negative by the IHAT. Western blot results closely followed MAT results; ELISA results matched MAT results, which were 1:40 or higher. In sera from the 2003 samples, MAT antibodies were found in 86 of 86 dolphins with titers of 1:25 in 29, 1:50 in 23, 1:100 in 27, 1:200 in 3, 1:1,600 in 1, and 1:3,200 in 3; these sera were not tested by other means. Overall, MAT antibodies were found in all 146 dolphin sera tested. Because marine mammals are considered sentinel animals indicative of contamination of the coastal and marine waters by T. gondii oocysts, serologically positive infections need to be validated by the detection of T. gondii organisms in the tissues of seropositive animals.     

Seroprevalence of Toxoplasma gondii from free-ranging black bears ( Ursus americanus ) from Florida

Toxoplasma gondii is a significant worldwide parasitic protozoan. In the present study, prevalence of antibodies of T. gondii was examined from 29 free-ranging black bears ( Ursus americanus ) from south-central Florida where the host species was listed as state threatened during this project. Overall T. gondii prevalence was found to be 44.8%, specifically 46.2% in male and 43.8% in female U. americanus , using a modified agglutination test (1:25 titer). Seroprevalence differences between sexes were not significant (P > 0.05). Results of the present study add supportive data to the growing body of evidence suggesting that U. americanus has one of the highest T. gondii seroprevalences among all known intermediate hosts. In addition, our data emphasize the importance of understanding parasitic disease dynamics from a conservation perspective.     

Direct evidence for Toxoplasma gondii infection in a wild serow (Capricornis crispus) from mainland Japan

Toxoplasma gondii infection was studied in 41 Japanese serows ( Capricornis crispus ), a goat-antelope in mainland Japan. Blood and muscle specimens were collected from 41 subjects between 2006 and 2010. Presence of antibodies to T. gondii in the sera was examined by using the latex agglutination test (cutoff titer 1:32); 10 of 41 (24.4%) were seropositive. Toxoplasma gondii DNA was detected in muscle tissue of 1 seropositive serow using a semi-nested PCR assay for the B1 gene. A partial nucleotide sequence (220 bp) corresponding to the B1 gene of T. gondii was obtained by direct sequencing; the sequence was 99.1% identical to that of the RH strain. This study is the first report to show direct evidence for the T. gondii infection in Japanese serows.