Treatment of toxoplasmosis in pregnancy: concentrations of spiramycin and neospiramycin in maternal serum and amniotic fluid.

Toxoplasma infection during pregnancy is widely treated with oral spiramycin to reduce the risk of congenital toxoplasmosis in the infant. Failures of therapy have been observed, however. In this study, a sensitive high-performance liquid chromatography technique was used to measure concentrations of spiramycin and neospiramycin, one of the major metabolites of spiramycin, in maternal serum and amniotic fluid. Samples were obtained from 18 women who underwent amniocentesis for polymerase chain reaction (PCR) diagnosis of fetal infection 5-109 days following the prescription of spiramycin therapy (3 g/day). Concentrations of spiramycin and neospiramycin in both serum and amniotic fluid were highly variable, ranging from nondetectable values to 1 microg/ml. None of the concentrations measured were within the range reported to inhibit growth of the parasite in vitro. Consistent with previous reports, part of the observed variability in maternal and fetal drug concentrations could be explained by individual differences in several pharmacokinetic parameters: intestinal absorption, tissue distribution, cellular uptake, metabolism, transfer across the placenta, drug accumulation in fetal tissue, and maternal and fetal drug elimination. The heterogeneity of the data could also be related to differences in patient compliance with the medication prescribed. By addressing factors that could impair adequate treatment of toxoplasmosis during pregnancy, the data presented call for a larger-scale controlled study to determine individual and diurnal variations in maternal drug levels, patient compliance, and outcomes of the offspring. The activity of neospiramycin against Toxoplasma gondii should be assessed.

A second European collaborative study on polymerase chain reaction for Toxoplasma gondii, involving 15 teams.

In order to investigate the accuracy and practicability of the polymerase chain reaction (PCR) in the antenatal diagnosis of congenital toxoplasmosis, a collaborative study involving 15 European laboratories was performed under the auspices of the Biomed 2 Programme of the European Community. Each team received 12 aliquots (four negative, eight positive) of 'artificial samples' made of amniotic fluid spiked with tachyzoites of the RH strain of Toxoplasma gondii. Each team performed its own PCR protocol (all were different). Nine of the 15 laboratories were able to detect a single parasite, but two of the 15 found all samples negative. Four of the 15 laboratories found one or more control samples to be falsely positive. This study highlights the lack of homogeneity between PCR protocols and performance and underlines the need for an external quality assurance scheme which could provide 'reference' samples that could be used by any laboratory wanting to establish and maintain an accurate diagnostic test based on PCR.

Follow-up of infants with congenital toxoplasmosis detected by polymerase chain reaction analysis of amniotic fluid.

This study was conducted to assess the validity of performing the polymerase chain reaction (PCR) on amniotic fluid for detecting fetal Toxoplasma infection. The primary endpoint was the outcome of the infant at 1 year of age. A prospective, consecutive study was performed in 49 infants born to mothers with primary Toxoplasma infection during pregnancy. PCR determinations of Toxoplasma gondii DNA in amniotic fluid were carried out as part of their prenatal management. Infants were examined at birth, and at 1, 3, 6, 9, and 12 months of age. Nine of 11 infants from pregnancies with positive PCR results proved to be infected based on follow-up serological investigations conducted during the first year of life. Two fetal deaths occurred. All 38 infants with negative PCR results remained uninfected at 1 year of age, irrespective of whether their mothers had received treatment with sulfadiazine/pyrimethamine or spiramycin alone. Psychomotor development was normal in all infants. This follow-up study confirms that PCR performed on amniotic fluid is a useful method for identification or exclusion of fetal Toxoplasma infection. Treatment of infected pregnant women and - in the event of a positive PCR result subsequent treatment of their infants is associated with a favorable outcome.

Prenatal allergen contact with milk proteins.

BACKGROUND: Cellular proliferation to various allergens (Dermatophagoides pteronyssinus, beta-lactoglobulin, bovine serum albumin, ovalbumin) has been found in cord blood cells. Whether this reflects a sensitization during foetal life is uncertain. OBJECTIVE: We studied the cellular reactivity and cytokine production of cord blood cells in response to cow's milk proteins in a randomly selected group of newborns. The delineation of possible in utero allergen contact was attempted. METHODS: Cord blood mononuclear cells from 39 neonates were incubated with cow's milk proteins (alpha-lactalbumin, beta-lactoglobulin, casein, alpha-casein, beta-casein, kappa-casein, bovine serum albumin) for 7 days, and proliferation was assessed by incorporation of [3H]thymidine. Cord blood cell-derived interferon-gamma (IFN gamma) and interleukin-4 (IL-4) secretion was evaluated in response to allergen or phytohaemagglutinin (PHA) stimulation. RESULTS: A pronounced proliferation of cells stimulated with alpha-lactalbumin (ALA: mean stimulation index 8.0, 95% confidence interval 5.2-10.8), beta-lactoglobulin (BLG: mean stimulation index 5.9, 95% confidence interval 3.2-8.6) and alpha-casein (2.6, 95% confidence interval 2.9-9.1), as opposed to unstimulated cells in medium, was found. No correlation was found between cellular proliferation to milk proteins and parental atopy, maternal total IgE or cord blood IgE. IFN gamma production (but not IL-4) was inducible by PHA (range 429-1810 pg/ml), but only in one individual upon stimulation with BLG. Preferentially, reduced IFN gamma levels were found in individuals with positive parental allergic history. CONCLUSION: The recognition of allergen by cord blood cells indicates that allergen priming must occur prenatally. The relevance for subsequent sensitization is unclear.

[Direct detection of Toxoplasma gondii with polymerase chain reaction in diagnosis of fetal toxoplasma infection]. / Direkter Nachweis von Toxoplasma gondii mit Polymerase-Kettenreaktion zur Diagnostik einer fetalen Toxoplasma-Infektion.

Primary infection with Toxoplasma gondii during pregnancy may affect the fetus and result in congenital toxoplasmosis. In Austria serological screening for detection of newly acquired infection during pregnancy was introduced in 1975. In this study we used polymerase chain reaction (PCR) for detection of fetal infection with Toxoplasma gondii. Amniotic fluid samples were analyzed from 11 women with serological indication of acute toxoplasmosis infection. Nine of these women had already received treatment prior to amnio-centesis and no evidence of Toxoplasma gondii DNA was detected with PCR in the respective amniotic fluid samples. Isolation of the organism by mouse inoculation was negative in these cases and follow-up serology as well as clinical examination of the infants confirmed these results. In 2 patients investigation of the amniotic fluid samples by means of PCR was positive; both women had not yet been treated at the time of amniocentesis. Our results indicate that identification of Toxoplasma gondii in amniotic fluid is a useful procedure for diagnosing or excluding fetal infection. Moreover, the current recommendations of the screening program appear to be successful in preventing congenital toxoplasmosis.

[Congenital adrenal gland insufficiency and myopathy]. / Kongenitale Nebenniereninsuffizienz und Myopathie.

We report a rare case of a female newborn presenting with muscular hypotonia, pneumonia, and cardiovascular and renal insufficiency. Adrenal insufficiency was diagnosed clinically and proven by extremely low cortisone (0.4-0.8 microgram/dl) and high ACTH plasma levels. Myopathy was diagnosed clinically, as well as by muscular biopsy. DNA analysis of both X chromosomes showed no abnormality in the region of the genes for adrenal hypoplasia and Duchenne muscular dystrophy. After 4 weeks of intensive care therapy the patient died of multiorgan failure. At autopsy she had only microscopically visible fetal adrenal cells and multiple porencephalic lesions.

Altered regulation of apolipoprotein A-IV gene expression in the liver of the genetically obese Zucker rat.

Apolipoprotein (apo) A-IV, a structural component of chylomicrons and high-density lipoproteins, may play a role in the catabolism of triglyceride-rich lipoproteins and in reverse cholesterol transport. To study the regulation of apoA-IV gene expression by genetic and nutritional factors, we determined the effect of a fish oil-rich and a sucrose-rich diet on apoA-IV gene transcription and nuclear and total cellular apoA-IV mRNA abundance in livers of genetically obese, hyperlipoproteinemic (fa/fa) Zucker rats and their lean (Fa/-) littermates. In obese rats fed chow, hepatic apoA-IV gene expression was more than twofold higher than in lean rats because of a post-transcriptional mechanism. apoA-I gene expression and apoC-III mRNA levels, studied as controls, were similar in both groups. The fish oil-rich diet reduced total cellular apoA-IV mRNA abundance transcriptionally to 34 +/- 4% of basal values in lean rats, but did not alter apoA-IV gene expression in obese rats. In contrast, this diet reduced apoA-I gene expression in both lean and obese animals. The sucrose-rich diet increased apoA-IV gene expression twofold in both lean and obese rats. Thus, genetic obesity alters the response of hepatic apoA-IV gene expression to a lipid-lowering diet rich in fish oil by a mechanism affecting transcriptional regulation.

Do heat shock proteins play a role in Graves' disease? Heat shock protein-specific T-cells from Graves' disease thyroids do not recognize thyroid epithelial cells.

Thyroid-derived T-cells from patients with Graves' disease were analyzed for their reactivity to recombinant heat shock proteins (hsp) and autologous thyroid epithelial cells (TEC). Five of six uncloned T-cell lines responded to stimulation with recombinant mycobacterial 71-kilodalton (kDa) hsp and cross-reacted with the corresponding amoebial and human proteins. Only one line reacted with recombinant 65-kDa hsp. Thyroid-derived T-cell lines also showed a proliferative response to TEC, which could be increased in four of the lines, when hsp expression was induced in thyroid cells by heat stress before the initiation of coculture. Clonal specificity analysis of thyroid-derived T-cell clones, however, demonstrated that distinct T-cells were responsible for the recognition of recombinant hsp and TEC. None of the clones responsive to recombinant hsp recognized TEC, whereas TEC-responsive clones did not react with recombinant hsp. Interestingly, the response of the majority of TEC-reactive clones could be dramatically increased when heat-shocked TEC were used as stimulator cells. These results suggest that T-cells specific for hsp of the 70- or 60-kDa families do not recognize TEC in the autoimmune thyroid gland. Heat shock-inducible proteins may, however, still play a role in the autoimmune process by facilitating the presentation of thyroid-specific autoantigen(s) to autoreactive T-cells.

The effect of TSH and TSI on the thyroglobulin expression of cultured human thyroid cells.

Human thyroid cells in culture stimulated by TSH and TSI were used in order to detect thyroglobulin expression. After three days stimulation the cells were incubated with monoclonal thyroglobulin antibody and FITC-conjugated antiglobulin. Fluorescent index (the intensity of fluorescence related to hundred analysed cells) was estimated for each experimental group. The most effective stimulation of the thyroglobulin expression was detected after TSH stimulation at the concentration of 0.1 mU/ml. TSI from active Graves' patients provoked the highest expression of thyroglobulin at concentration of 1.0 mg/ml, but the fluorescence index was lower than after TSH stimulation. The thyroglobulin expression was intracellular, large, partly confluent granules were detectable mainly in the perinuclear area. Antigen expression on the surface of cultured thyroid cells could not be detected. The morphology of thyroglobulin expression as detected by immunofluorescence, was the same after TSH and TSI stimulation. It is concluded, that both stimulating factors, i.e. TSH and TSI, are involved in the thyroglobulin expression of human thyroid cells.