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PubMed1990

Detection of specific immunoglobulin E in patients with toxoplasmosis.

An immunocapture assay was developed to detect Toxoplasma gondii-specific immunoglobulin E (IgE) in sera from adults with acute acquired infection or reactivation and from babies with congenital toxoplasmosis. The components of this assay were monoclonal antibody to human IgE, samples from patients, and T. gondii tachyzoites treated with Formalin. When T. gondii-specific IgE antibodies were present, visually detectable agglutination occurred. Sera, umbilical cord blood, fetal blood, cerebrospinal fluid, and amniotic fluid were tested by this method. Specific IgE antibodies were detected in sera from 25 (86%) of 29 adults who developed specific IgG antibody during pregnancy or had specific IgA and IgM antibodies. Specific IgE was present early during infection, at the time that IgM antibodies were present, and slightly preceding the presence of specific IgA antibodies. In 23 patients tested serially, IgE antibodies never persisted for longer than 4 months. No nonspecific anti-T. gondii IgE was detected in sera from uninfected individuals. Maternal IgE antibodies did not cross the placenta. In sera of patients with congenital toxoplasmosis, specific IgE antibodies were found at birth, during the first year of life, and during immunologic recrudescence following discontinuation of pyrimethamine-sulfonamide therapy. The IgE immunocapture assay is simple to perform. It is especially useful for determining when T. gondii was acquired by recently infected pregnant women.

Acquired Immunodeficiency Syndrome↗
PubMed1993

Serodiagnosis of toxoplasmosis by using a recombinant form of the 54-kilodalton rhoptry antigen expressed in Escherichia coli.

A 330-residue carboxy-terminal antigenic fragment of the Toxoplasma gondii 54-kDa rhoptry protein (ROP2) was expressed in Escherichia coli as a fusion polypeptide containing a 48-amino-acid sequence derived from phage lambda protein Cro and E. coli protein LacI followed by six consecutive histidyl residues. Metal chelate affinity chromatography provided an easy way to isolate the recombinant product in a highly purified form (> 95%). When this material was used to develop an immunoglobulin G (IgG) enzyme-linked immunosorbent assay for diagnosis of toxoplasmosis, the test reached a sensitivity of 89%. The sensitivity of the assay was similar whether the sera contained T. gondii-specific IgM or were devoid of such IgM. It was also found that ROP2 is a conserved antigen since antibodies against the recombinant antigen could be detected in mice experimentally infected with 11 independent T. gondii isolates originating from infected human tissues tested. Thus, the 54-kDa rhoptry antigen could advantageously complement other previously described T. gondii antigens for the development of more-sensitive and more-informative recombinant antigen-based tests for toxoplasmosis diagnosis.

Animals↗
PubMed1995

Diagnosis of congenital toxoplasmosis by immunoblotting and relationship with other methods.

Immunoblot has been evaluated as a diagnostic method for congenital toxoplasmosis. Like enzyme-linked immunofiltration assay (ELIFA), immunoblot can be used to compare antibody patterns and to determine if the antibodies are transmitted by the mother or synthesized by the fetus or infant. Among the 48 infants tested, 27 had congenital toxoplasmosis and 21 were suspected but had none. Reproducibility, sensitivity, specificity, and positive predictive values in immunoblot for immunoglobulins (Igs) G+M+A and/or G+M were 90, 92.6, 89.1, and 92.4%, respectively. G+M immunoblot and G+M ELIFA have better sensitivities than the conventional IgM immunosorbent agglutination assay, IgM enzyme-linked immunosorbent assay (ELISA), IgM immunofluorescence antibody test, in vitro culture, and mouse inoculation. The novel antibodies, i.e., those synthesized by infants against Toxoplasma gondii, were of the IgG class in most cases, although a confident diagnosis could be related to the number of observed Ig classes (G+M, G+A, and G+M+A). Immunoblot has a better resolution than ELIFA. In prenatal diagnosis, immunoblot could be complementary to in vitro culture and mouse inoculation. In the other cases, early detection by immunoblot appears to give the best results when compared with the other serological methods.

Animals↗
PubMed1996

Urine sample used for congenital toxoplasmosis diagnosis by PCR.

The diagnosis of toxoplasmosis in congenitally infected infants can be difficult; serology is unreliable, and diagnosis must be based on the combination of symptomatology and direct demonstration of the parasite. Four infants suspected of having Toxoplasma gondii infection were studied by serological analysis, tissue culture, and PCR determination. T. gondii was isolated from the urine of one patient. The parasite was detected by PCR in the blood and cerebrospinal fluid of three infants and in the urine in all patients. Because nested PCR proved to be a sensitive, relatively rapid, and specific method and because it can be applied to a variety of different clinical samples, PCR can be a valuable technique for the identification of T. gondii infections in children. The present study indicates that PCR examination of urine, a fluid never before used for diagnosis in this age group, may be valuable in diagnosing cases of congenital toxoplasmosis.

Animals↗
PubMed1999

Value of prenatal diagnosis and early postnatal diagnosis of congenital toxoplasmosis: retrospective study of 110 cases.

We reviewed the files of 110 women with Toxoplasma seroconversion during pregnancy. Prenatal diagnosis was attempted for 94 women by amniotic fluid sampling. Toxoplasma gondii was detected by PCR, with or without tissue culture and mouse inoculation. The early neonatal diagnostic procedure included placental testing by PCR and/or mouse inoculation, cord blood serological testing, and comparison of maternal and newborn antibodies by Western blotting (WB). Serological follow-up of the infants was conducted during the first year of life or until the diagnosis of congenital toxoplasmosis (CT) could be ruled out. Congenital infection was diagnosed in 27 individuals (20 live births) in the prenatal and/or neonatal period. The sensitivity and specificity of prenatal diagnosis were 81 and 100%, respectively. Placental examination was positive for 66.7% of individuals with CT and was always negative for neonates without CT. Cord blood serology detected immunoglobulin M (IgM) and/or IgA in 80% of infected newborns, with respective specificities of 91.2 and 87.7%. By WB we detected bands on IgG and IgM blots recognized by the newborn serum but not by the maternal serum (neosynthesized IgG and/or IgM) for 88.2% of infected infants within the first 2 months of life with a specificity of 100%. Early postnatal diagnosis was negative for 2 of the 20 neonates with CT. Both of these newborns had a negative prenatal diagnosis and were asymptomatic, suggesting a very low parasite load. In conclusion, despite the use of advanced methods, some cases of congenital toxoplasmosis cannot be detected early, which underlines the importance of careful follow-up of newborns who are at risk.

Animals↗
PubMed2000

Early aqueous humor analysis in patients with human ocular toxoplasmosis.

To evaluate the diagnostic sensitivity of a panel of laboratory tests for ocular toxoplasmosis performed at the time of presentation, paired samples of aqueous humor and serum were collected from 49 consecutive episodes of ocular toxoplasmosis with a clinical course of less than 3 weeks. Total immunoglobulin G (IgG) and Toxoplasma gondii-specific IgG, IgM, and IgA were quantified by enzyme-linked immunosorbent assay. The avidity of T. gondii-specific IgG was determined, and DNA extracted from aqueous humor was amplified for detection of a glycoprotein B gene sequence of T. gondii. The diagnosis was confirmed for 73% (36 of 49) of the patients; this rate rose to 79.5% if data from a later analysis of aqueous humor derived from five of the negative patients were included. The analysis of serum (detection of T. gondii-specific IgM and analysis of consecutive serum samples) alone did not contribute to the diagnosis. Calculation of local antibody production lacked diagnostic sensitivity when it was determined less than 3 weeks after the manifestation of clinical symptoms (28 of 49 patients [57%]), but this rose to 70% after an analysis of a second aqueous humor sample. The antibody avidity index attained diagnostic significance in only 8 of 43 instances (19%), and T. gondii DNA was amplified from no more than 6 of 39 (16%) aqueous humor samples. However, T. gondii-specific IgA was found within the aqueous humors of 11 of 43 patients (26%); measurement of the T. gondii-specific IgA level thus contributed substantially to the diagnostic sensitivity of the laboratory tests.

Animals↗
PubMed2001

Genotypic characterization of Toxoplasma gondii strains associated with human toxoplasmosis in Spain: direct analysis from clinical samples.

Genetic analysis of the SAG2 locus was performed to determine the prevalence of the different genotypes of Toxoplasma gondii (strain types I, II, and III) associated with human toxoplasmosis in Spain. This determination was made directly from primary clinical samples, obviating the previous process of isolation in mice or cell culture. A total of 34 isolates of T. gondii, collected from immunocompromised patients and congenital infection cases, were analyzed. Restriction fragment length polymorphism in PCR-amplified SAG2 products was used to group strains into one of the three genotypes of T. gondii. Complete characterization of the SAG2 gene was successful in 76.5% of the cases, demonstrating the feasibility of direct genotype analysis from clinical samples of different origins. Strains of T. gondii type II were the most prevalent in immunocompromised patients, with 52% of cases, while strains of type I were present in 75% of the congenital infection cases. These data differ from previous reports that show type II strains to be mostly associated with all kinds of human toxoplasmosis. These differences might be an effect of selection in the process of culture and isolation of the samples performed by other researchers prior to strain characterization.

Animals↗
PubMed2001

Strategy for diagnosis of congenital toxoplasmosis: evaluation of methods comparing mothers and newborns and standard methods for postnatal detection of immunoglobulin G, M, and A antibodies.

In a study involving 14 laboratories supported by the European Community Biomed 2 program, we evaluated immunologic methods for the postnatal diagnosis of congenital toxoplasmosis (CT). Among babies born to mothers who seroconverted to positivity for toxoplasmosis during pregnancy, we analyzed 55 babies with CT on the basis of persistent anti-Toxoplasma immunoglobulin G (IgG) at 1 year of life and 50 control babies without anti-Toxoplasma IgG at 1 year of life in the absence of curative treatment with pyrimethamine-sulfonamides. We tested in-house methods such as the enzyme-linked immunofiltration assay (ELIFA) or Immunoblotting (IB) for the detection of IgG or IgM; these methods allowed comparison of the immunologic profiles of the mothers and the infants. We compared ELIFA and IB with a commercial enzyme immunoassay (EIA) or in-house immunosorbent agglutination assay (ISAGA) for the detection of IgM or IgA. The performances of combinations of methods were also assessed. A cumulative sensitivity of 98% during a 1-year follow-up was obtained with the ELIFA plus ISAGA combination. Only one case of CT was missed by the ELIFA plus ISAGA combination, whereas three cases were missed by the IB plus ISAGA combination, even though 48% of patients with CT were treated with pyrimethamine-sulfonamides, which are known to inhibit antibody neosynthesis. A similar performance was obtained with either ELIFA or IB in combination with EIA. The difference in performance between ELIFA plus ISAGA and IB plus ISAGA was not statistically significant (P = 0.31), and we conclude that both combinations of tests can be used for the diagnosis of CT in newborns.

Adult↗
PubMed2006

Timely diagnosis of disseminated toxoplasmosis by sputum examination.

The diagnosis of disseminated toxoplasmosis in a 14-year-old allogeneic bone marrow recipient with graft-versus-host disease was determined by the detection of Toxoplasma gondii tachyzoites in sputum smears. Sputum analysis is a valuable alternative in the clinical assessment of pulmonary toxoplasmosis, especially when conventional invasive techniques are not practicable.

Adolescent↗
PubMed1978

Premarket evaluation of commercial toxoplasmosis indirect fluorescent-antibody reagents.

The quality of commercial toxoplasmosis indirect fluorescent-antibody reagents was evaluated over a 6-year period. Seven manufacturers voluntarily submitted their products for evaluation in the Center for Disease Control Premarket Evaluation Program. Each product was tested in accordance with the Center for Disease Control performance and labeling specifications and evaluation methods. Only 49% of all of the products tested met the Center for Disease Control requirements. Performance criteria are outlined, and suggestions are offered to assist laboratorians in obtaining toxoplasmosis indirect fluorescent-antibody reagents of high quality.

Antibodies↗
PubMed1991

Serological investigation of ocular toxoplasmosis.

The limitations of serological assessment in toxoplasma infection of the eye are well recognised, but the predictive value of clinical examination is not defined. We undertook a prospective investigation into the role of clinical examination and of serological findings in cases of suspected toxoplasma infection of the eye by means of the dye test and multiple IgM assays. Seventy-four cases of retinal disease and 202 control patients were studied. Patients with retinal disease had a significantly higher incidence of toxoplasma seropositivity than the control group. This was because some patients with retinal disease had acquired the infection congenitally. Half the patients investigated for toxoplasmosis were seronegative. Possible explanations for these findings included misdiagnosis, clinical uncertainty, or, the use of serology testing in the confirmation of other diseases. An excess of IgM reactivity among the retinal disease group may indicate low level immunoglobulin-M production associated with an acute exacerbation of ocular toxoplasmosis. There is a need to consider invasive procedures in cases of ocular infection and for novel techniques to aid the diagnosis of toxoplasma retinochoroiditis.

Adolescent↗
PubMed1980

Demonstration of lesions of cerebral toxoplasmosis by computerized tomography.

A patient who was treated for 11 years with cytotoxic drugs for Hodgkin's disease developed cerebral toxoplasmosis. Discrete lesions including an occipital abscess were visualized in the brain by computerized tomography (CT). This permitted a brain biopsy to be taken but the appearances were non-specific. At post-mortem pseudocysts of Toxoplasma gondii were found in several sites within the CNS. The recognition of these CT appearances as being due to toxoplasmosis should in future prompt serological investigation and urgent treatment of this potentially curable condition, despite negative biopsy material.

Brain Diseases↗
PubMed1977

Toxoplasmosis in coypu.

Serological studies indicate that toxoplasmosis is a common infection in coypus now established in Norfolk. A strain of Toxoplasma gondii was isolated from an animal which had shown a high titre to the dye test for toxoplasma antibodies. This was similar to the many tissue cyst producing strains which are of low virulence for most of the small laboratory animals. Congenital toxoplasmosis was proved in a litter born to one of the coypus during captivity. Superinfection occurred when animals, previously infected with a strain of low virulence, were given a challenge of the highly virulent RH strain, but coypus are so resistant that the latter strain caused no apparent illness even in previously uninfected seronegative animals. Antibodies developed to titres of 1:100,000 or more and, at lower levels, persisted for at least a year.

Animals↗
PubMed1983

Role of enzootic abortion and toxoplasmosis in an outbreak of abortion in a Scottish sheep flock.

During 1978-79 there was an outbreak of abortion in a large sheep flock during which approximately 10 per cent of the breeding ewes aborted. Both Toxoplasma gondii and Chlamydia ovis (the agent of enzootic abortion of ewes) were considered to be involved. In the year following this outbreak (1979-80), 156 ewes (11.4 per cent) aborted and the majority of cases were diagnosed as enzootic abortion: only one case showed gross pathology typical of toxoplasmosis. Serology carried out on sera collected from ewes at the time of abortion and at two post abortion samplings demonstrated that large numbers of animals had high titres against enzootic abortion of ewes while the prevalence of sheep with titres against toxoplasmosis was relatively low. Following the introduction of control measures to reduce the spread of enzootic abortion of ewes, the abortion rate in 1980-81 fell to 2.2 per cent. A small-scale trial was carried out to investigate the prophylactic effect of long acting oxytetracycline against enzootic abortion of ewes when given to pregnant sheep three weeks before lambing. Results indicated that treatment reduced the number of abortions in comparison with untreated controls.

Abortion, Veterinary↗
PubMed2004

Detection of toxoplasmosis in experimentally infected goats by PCR.

PCR was used to diagnose toxoplasmosis in two pairs of Barbari goats infected by oral administration of doses of either 10(4) or 10(5) oocysts of Toxoplasma gondii. Blood and lymph node aspirates were collected from the infected goats and control goat at intervals, and tissues were also collected from a fetus that was aborted and a doe that died during the trial. Both processed and unprocessed samples were used for the PCR, using primers directed to the multicopy B1 gene. None of the blood samples was positive, but a specific signal was obtained from the lymph node aspirates after partial DNA extraction. Direct PCR of the lung, muscle and mesenteric lymph node of the doe and lung tissue of the aborted fetus yielded the target fragment. The simplified PCR protocols, including partial DNA extraction and direct assay of lung tissue, were effective for the diagnosis of toxoplasmosis.

Aborted Fetus↗
PubMed1991

Primary central nervous system lymphoma versus toxoplasmosis in AIDS.

The imaging studies of 16 patients with acquired immunodeficiency syndrome (AIDS) and proved primary central nervous system (CNS) lymphoma were reviewed. All studies included computed tomography (CT); six also included magnetic resonance (MR) imaging. A periventricular lesion was seen in 50% of patients. At least one such lesion exhibited subependymal spread or ventricular encasement in 38%. One-third of lesions in three of five patients who underwent nonenhanced CT were hyperattenuating. Five lesions were at least in part hypointense on T2-weighted MR images. The specificity of these findings was evaluated with a similar review of the imaging studies in 28 patients with AIDS and proved toxoplasmosis. Only 3% of lesions were periventricular. None exhibited subependymal spread or encasement. None were hyperattenuating on nonenhanced CT scans. Similar findings in other CNS lesions in AIDS patients could not be found in the literature. A focal enhancing mass with subependymal spread on CT or MR images and hyperattenuation at nonenhanced CT were the most reliable features in distinguishing between primary CNS lymphoma and toxoplasmosis in AIDS patients.

Acquired Immunodeficiency Syndrome↗