Evaluation of a novel rapid one-step immunochromatographic assay for detection of monoclonal Helicobacter pylori antigen in stool samples from children.

A new rapid one-step immunochromatographic test using monoclonal antibodies for detection of Helicobacter pylori antigen in stool in children was evaluated on coded stool samples from 159 children (mean age, 9.7 +/- 5.0 years; 118 from Munich, 41 from Vienna): 86 children were H. pylori infected defined by positive culture and/or > or =2 other positive tests ([13C]urea breath test, histology, rapid urease test), and 73 children showed concordant negative results. Seventy-nine patients (12.1 +/- 3.8 years; 42 from Munich; 37 from Vienna) were tested 6 to 8 weeks after anti-Helicobacter pylori therapy with urea breath test and stool test. In Munich, all 160 tests (118 pre- and 42 posttreatment) were independently read by two observers. Equivocal results were excluded for calculation of sensitivity and specificity but were considered as false to assess accuracy. The two observers in Munich agreed in 63 out of 65 positive and 89 out of 95 negative results, while eight times (5.0%) they judged the test as equivocal. Pretreatment and posttreatment results for sensitivity were 88.1% (79.2 to 94.1) and 88.9% (51.8 to 99.7), specificity 88.1% (77.8 to 94.1) and 93.9% (85.2 to 98.3), and accuracy 83.5% and 81.5%, respectively. We conclude that the new monoclonal immunochromatographic quick test shows a good interobserver agreement, but equivocal results occur in 5%. Performance is comparable before and after therapy. The test may become a good alternative in children in settings where a [13C]urea breath test or a reliable enzyme immunoassay stool test are not available.

Pulmonary chemokines and their receptors differentiate children with asthma and chronic cough.

BACKGROUND: Cough is a frequent symptom in children, but the differentiation of asthmatic cough from cough of other origins can be difficult. Chemokines recruit T lymphocytes to inflamed tissues, and the corresponding chemokine receptors are differentially expressed on T H 1 and T H 2 cells. OBJECTIVE: We sought to determine whether levels of T H 1/T H 2-related chemokines and their receptors differ in bronchoalveolar lavage fluid (BALF) from 12 children with allergic asthma, 15 nonatopic children with chronic cough, and 10 children without airway disease. METHODS: The T H 1-related (IFN-gamma-inducible protein of 10 kd [IP-10], IFN-gamma-inducible T cell alpha chemoattractant [ITAC], monokine induced by IFN-gamma [Mig], and IFN-gamma) and T H 2-related (thymus- and activation-regulated chemokine [TARC], macrophage-derived chemokine [MDC], IL-5, and IL-4) chemokines and cytokines were quantified in BALF by ELISA and a particle-based multiplex array. Percentages of pulmonary lymphocytes expressing CXCR3 + and CCR5 + (T H 1) and CCR4 + and CCR3 + (T H 2) chemokine receptors were determined in BALF by flow cytometry. RESULTS: Pulmonary CCR4 + CD4 + cells and levels of TARC and MDC were significantly increased in asthmatic children versus children with chronic cough or without airway disease. In asthmatic children CCR4 + CD4 + cells correlated positively with levels of TARC, MDC, and serum IgE levels and negatively with FEV 1 . In contrast, CXCR3 + CD8 + cells and levels of ITAC were significantly increased in children with non-atopic chronic cough compared with the other groups. In children with chronic cough, CXCR3 + CD8 + cells correlated with levels of ITAC and IFN-gamma. CONCLUSION: Pulmonary CCR4 + CD4 + and CXCR3 + CD8 + cells and their ligands TARC, MDC, and ITAC clearly differentiate asthmatic children from nonatopic children with chronic cough. The analysis of these markers could facilitate the diagnostic discrimination of asthma versus other reasons for chronic cough in children.

FISH analysis of native smears from bone marrow and blood for the monitoring of chimerism and clonal markers after stem cell transplantation in children.

After stem cell transplantation (SCT) close follow-up of chimerism and/or clonal disease markers is essential for early treatment of graft failure or relapse. We wanted to assess the sensitivity, clinical reliability and practicability of inter-phase FISH on untreated, native smears of BM or PB for this purpose. We investigated 23 children after SCT with sex mismatch (MM) and/or clone specific markers (monosomy 7, trisomy 8, MLL rearrangement, bcr-abl, TEL-AML-1). Diagnoses were ALL (8), AML (6), MDS (2), CML (2), large cell anaplastic lymphoma (1) and SAA (4). Eighteen children were transplanted from sex-mismatched donors, seven among them had shown a clonal marker at diagnosis. The remaining five patients with sex matched donors also had a clonal marker. For FISH, we used commercial probes on fresh or stored unmanipulated smears of PB or BM. Cut-off levels for clonal markers were established on control probands without hematologic disease, for host sex on probands of the opposite sex, respectively (mean +3 SD). The presence of host cells and/or clonal markers established at diagnosis by conventional karyotyping was followed up after SCT at regular intervals by FISH. Nineteen of the 23 patients studied achieved and maintained complete continuous hematologic remission with corresponding absence of host and/or disease markers. In one of them, a fatal extramedullary relapse occurred. The associated mixed chimerism was confirmed by FISH. In all four cases with hematological relapse, the respective marker (MLL, bcr-abl, Mo 7) reappeared and was successfully monitored during DLI and repeat SCT in two as well as parallelled by simultaneous demonstration of host cells in the two sex mismatched cases among them. We demonstrate the usefulness of FISH on native smears for clinical routine follow-up of SCT patients. FISH allowed identification of cell origin in non-hematologic material (spinal fluid, pericardial effusion). Chimerism analysis in BM was slightly more sensitive than in PB. FISH was feasible on frozen stored smears as well.

Frequency of Valpha24+CD161+ natural killer T cells and invariant TCRAV24-AJ18 transcripts in atopic and non-atopic individuals.

Th2 cells play a central role in type I allergies. However, the source of interleukin-4 which may lead to a Th1/Th2 imbalance is unknown. Valpha24+CD161+ Natural killer T (NKT) cells secrete high amounts of interleukin-4 and/or interferon-gamma and are assumed to participate in the initiation of Th1/Th2 immune responses. Their contribution to the development of Th2-dependent type I allergies is controversial. Our objective in this paper was to determine whether Valpha24+CD161+ NKT cells differ in atopic and non-atopic adults. Venous blood was obtained from thirteen atopic and sixteen healthy adult probands. Valpha24+CD161+ NKT cells were determined in CD4+, CD8(bright/dim) and CD4-CD8- lymphocytes by flow cytometry. At the molecular level, the amounts of T cell receptor (TCR) AV24-AJ18 transcripts were quantified with respect to TCRAV24 chain transcripts alone or to all TCR alpha chain transcripts. To detect potential inserted nucleotides in the N-region, a novel real-time PCR-based technology was applied. Both CD4+ and CD4-CD8- NKT cells were present at higher frequencies than CD8+ NKT cells in all probands. CD8(dim) NKT cell levels were lower in healthy individuals, although not statistically significantly different to the patients. Amounts of AV24-AJ18 transcripts in relation to total TCR alpha-chains and to TCRAV24 alone were equal in both proband groups. N-region diversity was detected in four clones from four different individuals, but altered the amino acid sequence in only one clone of an atopic donor. Analysis of Valpha24+CD161+ NKT cell frequencies at both the cellular and molecular levels failed to reveal significant differences in peripheral blood of atopic and non-atopic probands. If NKT cells contribute to development of type I allergies they must do so at earlier times or in other locations.

Reinfection rate in children after successful Helicobacter pylori eradication.

OBJECTIVES: This study was performed to determine the rate of Helicobacter pylori reinfection after its successful eradication in children living in Germany. DESIGN: A total of 102 children (48 boys; 31 German and 71 other nationalities; age 1.8-18 years) with a negative (13)C-urea breath test 8 weeks after triple therapy were followed up by a (13)C-urea breath test every 6 months. The cohort included 11 children aged <6 years, 58 children aged > or =6 to 12 years, and 33 children > or =12 years. RESULTS: The mean duration (+/- standard deviation) of follow-up was 15.5 +/- 11.9 months with a maximum of 4.9 years, representing 132 patient years. Only three children (aged 9.7-14.9 years, one German, two Turkish) tested positive at 6, 12, and 18 months, respectively. The calculated reinfection rate was 2.3% per person per year. CONCLUSION: The risk of reinfection with H. pylori is low in children living in Germany. There is no evidence that the reinfection rate depends on the age, sex, or nationality of the child. The low reinfection rate indicates that it is unnecessary to screen or treat asymptomatic family members in order to prevent reinfection.

Different effects of endotoxin versus mite and cat allergen exposure on T-cell differentiation in infants.

BACKGROUND: Early exposure to bacterial endotoxin has been proposed to protect against allergy development in children. Whether endotoxin is able to direct T-cell differentiation into a predominance of type 1 immunity is still unresolved. OBJECTIVE: We sought to compare the effects of endotoxin and mite and cat allergens on T-cell differentiation in infants. METHODS: In a random population sample of 135 2-year-old children of an ongoing birth-cohort study, peripheral blood CD4+ and CD8+ T-cell subsets were defined by the expression of the chemokine receptors CCR5 and CCR3 as surrogate markers for type 1 and type 2 T cells, respectively. Endotoxin and mite and cat allergens were measured in house dust collected from the mother's mattress at the child's age of 3 months to assess early exposure. RESULTS: In the CD4+ T-cell subset, endotoxin levels were positively associated with high proportions of type 1 CCR5+ cells (odds ratio for fourth exposure quartile [OR(Q4)], 7.68; 95% CI, 1.35-43.75), whereas cat allergen levels were associated with increased proportions of type 2 CCR3+ cells (OR(Q4), 4.07; 95% CI, 1.05-15.85). In contrast to endotoxin, allergen levels were associated with CD8+ T cells, showing an inverse relationship between mite allergen concentrations and high proportions of CCR5+ or CCR3+ cells (CCR5+ cells: OR(Q4), 0.14; 95% CI, 0.03-0.74; CCR3+ cells: OR(Q4), 0.16; 95% CI, 0.03-0.89) and a positive association of cat allergen levels with increased proportions of CCR5+ cells (OR(Q4), 9.24, 95% CI, 1.61-53.10), as well as CCR3+ cells (OR(Q3), 6.64; 95% CI, 1.21-36.51). CONCLUSION: Our results indicate that endotoxin has the potential to promote the development of type 1 CD4+ T cells, whereas mite and cat allergens primarily modify the proportion of CD8+ cells of both types.