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PubMed2001

Seroprevalence of Toxoplasma gondii infection in goats by the indirect haemagglutination, immunofluorescence and immunoenzymatic tests in the region of Uberlândia, Brazil.

A comparative study of the indirect haemagglutination (IHA), immunofluorescence (IFAT) and immunoenzymatic (ELISA) tests was carried out to determine the prevalence of Toxoplasma gondii antibodies in goats. One hundred seventy-four serum samples were obtained from four goat herds from the region of Uberlândia, State of Minas Gerais. The distribution of the animals, according to their origin, was as follow: 71 from herd I; 39 from herd II; 37 from herd III; and 27 from herd IV. Serum samples were analyzed by IHA, IFAT and ELISA, considering the reactivity of the serum samples at dilution > or = 1:64 as cut off titer for the three tests. A global seroprevalence of 18.4% was observed, with significantly higher positivity rate in the herd II (66.7%) and older animals (> 36 months). A high and significant positive correlation was found between the titers obtained by the IHA versus IFAT, IHA versus ELISA, and ELISA versus IFAT. Therefore, it can be concluded that the three analyzed tests have shown to be highly concordant and appropriate for epidemiological surveys of Toxoplasma infection in goats. Although the seroprevalence of T. gondii infection in goats is relatively low in this region as compared to other regions of the country, adequate management might be useful and essential to control the infection in the goat herds.

Animals↗
PubMed2002

Anionic sites, fucose residues and class I human leukocyte antigen fate during interaction of Toxoplasma gondii with endothelial cells.

Toxoplasma gondii invades and proliferates in human umbilical vein endothelial cells where it resides in a parasitophorous vacuole. In order to analyze which components of the endothelial cell plasma membrane are internalized and become part of the parasitophorous vacuole membrane, the culture of endothelial cells was labeled with cationized ferritin or UEA I lectin or anti Class I human leukocyte antigen (HLA) before or after infection with T. gondii. The results showed no cationized ferritin and UEA I lectin in any parasitophorous vacuole membrane, however, the Class I HLA molecule labeling was observed in some endocytic vacuoles containing parasite until 1 h of interaction with T. gondii. After 24 h parasite-host cell interaction, the labeling was absent on the vacuolar membrane, but presents only in small vesicles near parasitophorous vacuole. These results suggest the anionic site and fucose residues are excluded at the time of parasitophorous vacuole formation while Class I HLA molecules are present only on a minority of Toxoplasma-containing vacuoles.

Animals↗
PubMed1985

One point dilution enzyme-linked immunosorbent assay (ELISA) for Toxoplasma gondii seroepidemiological surveys.

A one point dilution enzyme-linked immunosorbent assay (ELISA) procedure suitable for determining immunoglobulin G (IgG) antibody levels to Toxoplasma gondii (T. gondii) in community seroepidemiological surveys is described. A two-fold serial dilution ELISA procedure was first used to determine the IgG titers in 56 and 83 sera earlier screened by the Sabin-Feldman dye test (DT) and the indirect hemagglutination test (IHA), respectively. The regression rate of the results by the DT and ELISA was 0.92. Comparison of the results by the IHA and the two-fold serial dilution ELISA gave regression coefficient of 0.92. Using the absorbance values for the test sera at dilutions of 1:20, standard curves made by plotting the optical density versus the corresponding dilution factor of a control sera were used to estimate the antibody levels. The regression coefficient of the results by the two-fold serial dilution method and those by the curves for sera with titers of up to 1:320 was 0.97. The curves could not, however, estimate accurately the antibody level in sera with titers above 1:320. The one point dilution ELISA described is a useful epidemiological tool for the screening of IgG antibody to Toxoplasma gondii in the community. However, larger series are required to confirm our observations.

Antibodies↗
PubMed2003

Detection of Toxoplasma gondii IgG antibodies in juvenile great grey kangaroos, Macropus giganteus giganteus.

Serial IgG antibody titers to Toxoplasma gondii from three juvenile great grey kangaroos (Macropus giganteus giganteus) with clinical signs consistent with toxoplasmosis were compared with titers from three clinically normal juveniles of similar age. The modified agglutination test was used to measure antibodies to T. gondii. Results demonstrated declining antibody titers to T. gondii in five of six animals. Antibody titers were similar in animals with and without clinical signs. This was interpreted as an indication that maternal IgG antibodies to T. gondii were transferred to juveniles, presumably in milk, rather than an indication of clinical disease. Therefore, clinicians should interpret Toxoplasma serology results in juvenile macropods cautiously.

Agglutination Tests↗
PubMed2000

Is stage conversion the initiating event for reactivation of Toxoplasma gondii in brain tissue of AIDS patients?

Reactivation of chronic toxoplasmosis resulting in Toxoplasma encephalitis (TE) is a common event in acquired immune deficiency syndrome (AIDS) patients. Conversion from Toxoplasma gondii bradyzoites to tachyzoites is a prerequisite for reactivation. Until recently, the study of stage conversion in human tissue was not possible due to the lack of antibodies that recognize stage-specific epitopes after long-term formaldehyde fixation. Using the combination of a polyclonal anti-T. gondii antibody, the cyst-stage-specific monoclonal antibody CC2, and a tachyzoite-specific polyclonal antibody (anti-SAG1, recombinant), we tried to demonstrate parasite differentiation in the brain tissue of 10 AIDS patients with clinically suspected TE. Double labeling of the stage-specific antibodies enabled us to demonstrate interconversion between tachyzoites and bradyzoites for the first time in human tissue. The study confirmed that the transformation process is nonsynchronous and that the manifestation of TE depends on the degree and site of tissue destruction caused by invading tachyzoites. The original source of tachyzoites could never be located, but a few samples suggested that tachyzoites may invade by dissemination across the blood-brain barrier. Cyst rupture as the first event in the process of reactivation was not seen. We conclude that the initial site(s) of reactivation will be destroyed by tissue-destructive tachyzoites long before clinical symptoms occur.

AIDS-Related Opportunistic Infections↗
PubMed2000

Prevalence of antibodies to Toxoplasma gondii in ostriches (Struthio camelus).

Serum samples from 973 ostriches (Struthio camelus) in Canada were examined for antibodies to Toxoplasma gondii by the modified agglutination test incorporating mercaptoethanol and formalin-fixed whole tachyzoites. Twenty-eight (2.9%) of the 973 birds were found to be seropositive for antibodies to T. gondii at titers of 1:25 in 15 birds, 1:50 in 12 birds, and 1:500 in 1 bird. This is the first record of T. gondii exposure in ostriches, and it supports the hypothesis that all avian species are susceptible to Toxoplasma infection. Nevertheless, the results of this study suggest that the risk of acquiring toxoplasmosis from ostriches as a food source is low.

Agglutination Tests↗
PubMed2000

Intergenic spacer (IGS) polymorphism: a new genetic marker for differentiation of Toxoplasma gondii strains and Neospora caninum.

The region between the 28S and 18S rRNA genes, including the intergenic spacer (IGS) region and the 5S rRNA gene, from 32 strains of Toxoplasma gondii and the NC1 strain of Neospora caninum was amplified and used for DNA sequencing and/or restriction fragment length polymorphism (RFLP) analysis. The 5S rDNA sequences from 20 strains of T. gondii were identical. The IGS region between the 5S and 18S rRNA genes (nontranscribed spacer 2 or NTS 2) showed 10 nucleotide variations. Six of the 10 variant positions correlated with the murine virulence of the strains. Intraspecific polymorphisms distinguished the virulent strains of zymodemes 5, 6, and 8 from other virulent strains (in zymodeme 1). RFLP methods (IGS-RFLP) were developed and used to characterize the virulent and avirulent patterns among 29 T. gondii strains. Sequence diversity of 19.8% was found between T. gondii and N. caninum when comparing a region of 919 bp at the 3' end of NTS 2. The sequence variation in ribosomal IGS could therefore be a useful marker for Toxoplasma strain identification and for distinguishing N. caninum from T. gondii.

Animals↗
PubMed2000

Detection of Toxoplasma gondii by polymerase chain reaction in sera of acutely infected mice.

The presence of Toxoplasma gondii DNA was detected in sera of acutely infected mice by polymerase chain reaction. Adult mice were inoculated intraperitoneally with 5 x 10(3) T. gondii RH strain tachyzoites. Five mice were killed every 3 hr from 3 to 21 hr post infection (PI) and every day from 1 to 7 days PI. Toxoplasma gondii DNA was first detected in 60% of the infected mice 18 hr PI and in 100% of the animals 21 hr PI and from 1 to 7 days PI. No mice survived longer than 7 days.

Acute Disease↗
PubMed2001

Analysis of the humoral responses of Toxoplasma gondii-infected cats using immunofluorescent assays with tachyzoite, bradyzoite, and gametogenic stages.

We investigated levels of Toxoplasma gondii specific antibodies present in sera, intestinal secretions, and fecal extracts obtained from cats following primary and challenge infections. Antibodies specific to T. gondii tachyzoites, bradyzoites, sporozoites, and enteroepithelial stages were detected by indirect immunofluorescence assay. Enteroepithelial stage-specific antibodies were detected in serum as early as 2 wk after infection, whereas antibodies from intestinal secretions did not appear until 3 wk following infection. The T. gondii-specific IgG and IgA antibodies were present in serum, but only specific IgA antibodies were detected in the intestinal secretions. Serum IgG bound to tachyzoites, bradyzoites, sporozoites, and enteroepithelial stages of T. gondii, whereas serum IgA bound strongly to enteroepithelial stages but only weakly to tachyzoites and bradyzoites. IgA from intestinal secretions bound to antigens on all enteroepithelial stages and the distal tips of sporozoites and bradyzoites but did not bind to tachyzoites. IgA present in fecal extracts also bound to enteroepithelial stages of T. gondii. Toxoplasma gondii infection in cats induces the production of antibodies that bind with all forms of the parasite, including the enteroepithelial stages. Comparison of the staining patterns of T. gondii stages for serum and intestinal secretion IgA indicated differences. Thus, the intestinal antibody immune response may be uniquely focused on the intestinal stages relative to the circulating antibodies, resulting in a compartmentalization of the humoral response.

Animals↗
PubMed2001

Organ infectivity of Toxoplasma gondii in interferon-gamma knockout mice.

To determine the influence of interferon (IFN)-gamma on the organ infectivity and on the genetic susceptibility of susceptible (C57BL/6) and resistant (BALB/c) strains after peroral infection with cysts of Toxoplasma gondii. IFN-gamma knockout (KO) mice in C57BL/6 and BALB/c backgrounds were utilized. The kinetics of the changes in T. gondii abundance were evaluated with a quantitative competitive polymerase chain reaction assay in various organs at different times after peroral infection. In IFN-gamma KO mice, a T. gondii-specific gene, SAG1, was detected in all organs examined, and the protozoan proliferated much more actively than in wild-type mice. The abundance of T. gondii was much higher in mesenteric lymph nodes and the heart than in other organs. In contrast, in the nervous system organs and kidneys, only a weakly detectable reaction was observed. Toxoplasma gondii grew at a more rapid rate in the organs of IFN-gamma KO C57BL/6 mice than in the organs of IFN-gamma KO BALB/c mice during the course of infection. Destruction of the IFN-gamma gene showed remarkable effects on the infectivity in both susceptible and resistant mice.

Animals↗
PubMed2002

Tachyzoite-induced life cycle of Toxoplasma gondii in cats.

The tachyzoite-induced cycle of Toxoplasma gondii was studied in 46 cats. Tachyzoites of the M-7741 or Me-49 strain of T. gondii were administered orally to cats by pouring into the mouth or by stomach tube, or by intraintestinal inoculation. Ten weaned cats that had been inoculated with tachyzoites directly in the intestine were killed 1, 3, 6, 9, 12, 15, 18, or 25 days later, and their tissues were studied histologically and bioassayed in mice. Toxoplasma gondii was demonstrable in the blood of 8 cats and in other tissues of all these 10. Four out of five 1- to 8-day-old cats fed tachyzoites by stomach tube became infected with T. gondii, and 1 became ill because of toxoplasmosis. All 19 weaned cats fed tachyzoites (poured into the mouth) became infected, and 6 died of acute toxoplasmosis 9-15 days after being fed T. gondii. Six out of 12 weaned cats fed tachyzoites by stomach tube became infected but were asymptomatic. Overall, 12 out of 26 cats observed for 19 days or more shed oocysts with a prepatent period (pp) of 19 days or more, with the sole exception of 1 cat that shed oocysts with a pp of 5 days. Enteroepithelial stages of T. gondii were not found in any cat before oocysts were shed. Cats shed up to 360 million oocysts in a day, and oocysts were shed for 4-6 days.

Animals↗
PubMed2003

Toxoplasma gondii isolates from free-ranging chickens from the United States.

Prevalence of Toxoplasma gondii infection in chickens is a good indicator of the strains prevalent in their environment because they feed from ground. The prevalence of T. gondii was determined in 118 free-range chickens from 14 counties in Ohio and in 11 chickens from a pig farm in Massachusetts. Toxoplasma gondii antibodies (> or = 1: 5) were found using the modified agglutination test (MAT) in 20 of 118 chickens from Ohio. Viable T. gondii was recovered from 11 of 20 seropositive chickens by bioassay of their hearts and brains into mice. The parasite was not isolated from tissues of 63 seronegative (< or = 1:5) chickens by bioassay in cats. Hearts, brains, and muscles from legs and breast of the 11 chickens from the pig farm in Massachusetts were fed each to a T. gondii-negative cat. Eight cats fed chicken tissues shed oocysts; the 3 cats that did not shed oocysts were fed tissues of chickens with MAT titers of 1:5 or less. Tachyzoites of 19 isolates of T. gondii from Ohio and Massachusetts were considered avirulent for mice. Of 19 isolates genotyped, 5 isolates were type II and 14 were type III; mixed types and type I isolates were not found.

Agglutination Tests↗
PubMed2003

Isolation and genotyping of Toxoplasma gondii from free-ranging chickens from Argentina.

The prevalence of Toxoplasma gondii in free-ranging chickens can be considered a good indicator of the prevalence of T. gondii oocysts in the environment because chickens feed from the ground. In the present study, prevalence of T. gondii in 29 free-range chickens (Gallus domesticus) from Argentina was investigated. Blood, heart, and brain from each chicken were examined for T. gondii infection. Antibodies to T. gondii, assayed with the modified agglutination test (MAT), were found in 19 of 29 (65.5%) chickens. Hearts and brains of seropositive (MAT > or = 1:5) chickens were bioassayed in mice. Toxoplasma gondii was isolated from 9 of 19 seropositive chickens. Genotyping of chicken isolates of T. gondii using the SAG2 locus indicated that 1 was type I, 1 was type II, and 7 were type III. This is the first report of isolation of T. gondii from chickens from Argentina.

Agglutination Tests↗
PubMed2004

Endogenous polyamine levels in macrophages is sufficient to support growth of Toxoplasma gondii.

Cytotoxic-activated macrophages control Toxoplasma gondii growth by producing nitric oxide (NO). However, the parasite can partially inhibit NO production. NO is generated from arginine within the polyamine biosynthetic pathway. Two enzymes of this pathway are ornithine, decarboxylase (ODC) and arginine decarboxylase (ADC). The aim of the present work was to investigate whether T. gondii is able to modulate polyamine metabolism in macrophages. Toxoplasma gondii infection did not affect basal ODC or ADC activity. However, lipopolysaccharide induced an increase in ODC activity. Polyamine-treated macrophages exhibited a T. gondii-infection index similar to controls but a higher adhesion index; the parasite did not grow in methyl-ornithine (ODC inhibitor)-treated macrophages. The parasites were able to take up putrescine with a Km of 0.92 microM, indicating the presence of a high-affinity putrescine-transporter system. Putrescine-treated T. gondii actively penetrated macrophages and Vero cells. However, NO production and lysosomal parasitophorous vacuole fusion were not inhibited. Considered together, these results demonstrate that T. gondii requires polyamines for multiplication. However, as opposed to Trypanosoma cruzi and because of a relatively high-affinity putrescine-transporter system in the parasite, constitutive macrophage levels of putrescine seem sufficient to support T. gondii survival and multiplication.

Animals↗
PubMed2004

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.

Agglutination Tests↗
PubMed2004

Survival of Toxoplasma gondii oocysts in Eastern oysters (Crassostrea virginica).

Toxoplasma gondii has recently been recognized to be widely prevalent in the marine environment. It has previously been determined that Eastern oysters (Crassostrea virginica) can remove sporulated T. gondii oocysts from seawater and that oocysts retain their infectivity for mice. This study examined the long-term survival of T. gondii oocysts in oysters and examined how efficient oysters were at removing oocysts from seawater. Oysters in 76-L aquaria (15 oysters per aquarium) were exposed to 1 x 10(6) oocysts for 24 hr and examined at intervals up to 85 days postexposure (PE). Ninety percent (9 of 10) of these oysters were positive on day 1 PE using mouse bioassay. Tissue cysts were observed in 1 of 2 mice fed tissue from oysters exposed 21 days previously. Toxoplasma gondii antibodies were found in 2 of 3 mice fed oysters that had been exposed 85 days previously. In another study, groups of 10 oysters in 76-L aquaria were exposed to 1 x 10(5), 5 x 10(4), or 1 x 10(4) sporulated T. gondii oocysts for 24 hr and then processed for bioassay in mice. All oysters exposed to 1 x 10(5) oocysts were infected, and 60% of oysters exposed to 5 x 10(4) oocysts were positive when fed to mice. The studies with exposure to 1 x 10(4) oocysts were repeated twice, and 10 and 25% of oysters were positive when fed to mice. These studies indicate that T. gondii can survive for several months in oysters and that oysters can readily remove T. gondii oocysts from seawater. Infected filter feeders may serve as a source of T. gondii for marine mammals and possibly humans.

Animals↗
PubMed2004

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.

Agglutination Tests↗
PubMed2004

Biologic and molecular characteristics of Toxoplasma gondii isolates from striped skunk (Mephitis mephitis), Canada goose (Branta canadensis), black-winged lory (Eos cyanogenia), and cats (Felis catus).

Toxoplasma gondii isolates can be grouped into 3 genetic lineages. Type I isolates are considered virulent to outbred mice, whereas Type II and III isolates are not. In the present report, viable T. gondii was isolated for the first time from striped skunk (Mephitis mephitis), Canada goose (Branta canadensis), and black-winged lory (Eos cyanogenia). For the isolation of T. gondii, tissues were bioassayed in mice, and genotyping was based on the SAG2 locus. Toxoplasma gondii was isolated from 3 of 6 skunks, 1 of 4 Canada geese, and 2 of 2 feral cats (Felis catus) from Mississippi. All donor animals were asymptomatic. Viable T. gondii was also isolated from 5 of 5 lories that had died of acute toxoplasmosis in an aviary in South Carolina. Genotypes of T. gondii isolates were Type III (all skunks, lories, and the goose) and Type II (both cats). All 5 Type III isolates from birds and 2 of the 3 isolates from skunks were mouse virulent.

Animals↗