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SRY is the Y-chromosomal gene that determines male sex development in humans and most other mammals. After three decades of study, we still lack a detailed understanding of which domains of the SRY protein are required to engage the pathway of gene activity leading to testis development. Some insight has been gained from the study of genetic variations underlying differences/disorders of sex determination (DSD), but the lack of a system of experimentally generating SRY mutations and studying their consequences in vivo has limited progress in the field. To address this issue, we generated a mouse model carrying a human SRY transgene able to drive testis determination in XX mice. Using CRISPR-Cas9 gene editing, we generated novel genetic modifications in each of SRY's three domains (N-terminal, HMG box, and C-terminal) and performed a detailed analysis of their molecular and cellular effects on embryonic testis development. Our results provide new functional insights unique to human SRY and present a versatile and powerful system in which to functionally analyze variations of SRY including known and novel pathogenic variants found in DSD.

De novo germline mutations in GNB1 have been associated with a neurodevelopmental phenotype. To date, 28 patients with variants classified as pathogenic have been reported. We add 18 patients with de novo mutations to this cohort, including a patient with mosaicism for a GNB1 mutation who presented with a milder phenotype. Consistent with previous reports, developmental delay in these patients was moderate to severe, and more than half of the patients were non-ambulatory and nonverbal. The most observed substitution affects the p.Ile80 residue encoded in exon 6, with 28% of patients carrying a variant at this residue. Dystonia and growth delay were observed more frequently in patients carrying variants in this residue, suggesting a potential genotype-phenotype correlation. In the new cohort of 18 patients, 50% of males had genitourinary anomalies and 61% of patients had gastrointestinal anomalies, suggesting a possible association of these findings with variants in GNB1. In addition, cutaneous mastocytosis, reported once before in a patient with a GNB1 variant, was observed in three additional patients, providing further evidence for an association to GNB1. We will review clinical and molecular data of these new cases and all previously reported cases to further define the phenotype and establish possible genotype-phenotype correlations.

The aim of this study was to analyse whether altered cytokine production by cord blood (CB) T cells is of relevance regarding the development of allergic diseases during the first 2 yr of life independent from known or suspected risk factors for allergy. Within an ongoing birth cohort study (Life style - Immune System - Allergy; LISA) the cytokine production of PMA/ionomycin-stimulated CB cells was measured by intracellular cytokine staining. Data of 98 children from Leipzig and Munich with complete information on cytokine production at birth and allergic outcomes during the first 2 yr were analysed. Statistical analysis was performed using a regression model adjusted for gender, month of birth, parental history of atopy, parental education, exposure to environmental tobacco smoke, maternal smoking during pregnancy, renovation activities during pregnancy, pet ownership and study centre. During the first 2 yr of life, 17.3% of the children developed a physician-diagnosed atopic dermatitis. Children with reduced frequencies of interferon-g (IFN-g)-producing CD4+ T cells in the CB (1st quartile) had a higher risk to develop atopic dermatitis (adjusted OR 5.16, 95% CI: 1.04-25.6). Furthermore, a high percentage of interleukin (IL)-4-producing T cells in CB in children from the Leipzig cohort were associated with an increased risk for atopic dermatitis (adjusted OR 8.92, 95% CI: 1.40-56.93 for the 90th percentile). CD8+ cytokine-producing CB T cells had no relation to increased risk for atopic dermatitis. Low amounts of IFN-g and high amounts of IL-4-producing T cells at birth may enhance the risk of subsequent development of atopic dermatitis.

Biological treatment of 2,4,6-trinitrotoluene (TNT) in soil rarely results in complete mineralization of the parent compound. More often, the largest proportion of the TNT carbon is incorporated into the soil organic matrix. Therefore, we evaluated the stability of nonextractable residues from various bioremediation processes of 14C-TNT in soils. The extractable amounts of the residual radioactivity varied between 7 and 33% and thus the nonextractable amount between 93 and 67% (3–15% in fulvic acids, 26–46% in humic acids, and 27–44% in the humin fraction). The residue-containing soils were analyzed for the release of radioactivity after treatment by physical (freeze and thaw, grinding of soil, and steam extraction), chemical (acid rain and addition of metal complexing agent), and biological methods (addition of compost, white rot fungi, radical-generating enzymes, and germination of plants). Freeze and thaw treatment and grinding of the soil did not alter the partitioning of the label significantly. Steam extraction and acid rain extraction increased the water extractability to 11 to 29% and to 51.6% in the native TNT-contaminated soil. The addition of ethylenediamine-tetraacetate (EDTA) increased the extractability from 7 to 12%. After biological treatment, only slightly increased extractability (<<10%) was observed. No increase of extractable TNT or known metabolites was observed with any of the treatments. Thus, under the treatment conditions applied in this study, the residues formed during microbial transformation of TNT may be biogenic residues with low mobilization potential and low hazardous impact.

The molecular diversity of arbuscular mycorrhizal (AM) fungi colonizing roots of Prunus africana and of AM fungal spores obtained from baiting cultures of indigenous soils from two dry afromontane forests of Ethiopia was investigated. The internal transcribed spacer (ITS) rDNA region from colonized roots and single spores of three AM fungal spore types was amplified, cloned and sequenced using AM fungal specific primers.Phylogenetic analysis using the 5.8S rDNA data set revealed that 109 of the sequences obtained belong to members of the Glomeromycota. Subsequent 5.8S/ITS2 rDNA sequence analysis indicated high AM fungal diversity and dominance of Glomus types. Twenty sequence types belonged to the Glomeraceae and one each to the Diversisporaceae and Archaeosporaceae. Two of the three spore types were identified as Glomus etunicatum and Glomus mosseae.Twenty of the AM fungal types identified are new to Ethiopia and to science. The AM fungal community differed between the two sites studied.

The fates of the labels from [14C] and [15N] trinitrotoluene were analyzed in bioreactors under aerobic conditions in soil treated by a fungal bioremediation process with Stropharia rugosoannulata and in control soil. Up to 17.5% of the 15N label had a different fate than the 14C label. Three N-mineralization processes were identified in detailed experiments with [15N]TNT. About 2% of the 15N label was found as NO3− and NH4−, showing simultaneous processes of direct TNT denitration (I) and reduction with cleavage of the amino groups (II). The enrichment of NO2−/NO3− (up to 7.5 atom% 15N abundance) indicates the formation of Meisenheimer complexes with a denitration of [15N]TNT. A 1.4% of the label was found distributed between N2O and N2. However, the 15N enrichment of the N2O (up to 38 atom%) demonstrated that both N atoms were generated from the labeled TNT and clearly indicates a novel formation process (III). We propose, as an explanation, the generation of N2O by cleavage from condensed azoxy metabolites. In addition, 1.7% of the 15N label was detected as biogenic amino acids in the wheat straw containing the fungus. Overall, 60 to 85% of the applied [15N]TNT was degraded and 52 to 64% was found as nonextractable residues in the soil matrix. Three percent was detected as 2-amino-4,6-dinitrotoluene and 4-amino-2,6-dinitrotoluene.

Background: Within an ongoing birth cohort study (LISA) the cytokine production of cord blood T cells was compared between neonates from Leipzig (East Germany) and Munich (West Germany). The aim of this study was to analyse regional differences and influencing factors of the immune status.Methods: Cytokine production was measured in a randomly selected subgroup of 158 children from the LISA (Life style - Immune system - Allergy) cohort by intracellular cytokine staining. Information on family "atopy" history (FAH) and home characteristics was obtained from questionnaires.Results: Reduced numbers of interferon-? (IFN-?) and tumor necrosis factor-a (TNF-a) producing T cells were found in association with biparental FAH and housing renovation during pregnancy. In addition, cytokine production was influenced by season. In Munich, the frequency of biparental FAH and of renovation measures during pregnancy was significantly higher as compared to Leipzig. Neonates from Munich showed significantly decreased amounts of IFN-? and TNF-a and elevated levels of interleukin-4 (IL-4) producing T cells. Differences in cytokine production between Munich and Leipzig were influenced by season (IL-4) and housing renovation (IFN-?, TNF-a).Conclusions: Since differences in the T cell cytokine production of neonates in Munich and Leipzig are independent from FAH our findings may provide evidence for the impact of environmental factors upon the fetal immune system.