Natural Variant of Collagen-Like Protein A in Serotype M3 Group A Streptococcus Increases Adherence and Decreases Invasive Potential

Group A Streptococcus (GAS) predominantly exists as a colonizer of the human oropharynx that occasionally breaches epithelial barriers to cause invasive diseases. Despite the frequency of GAS carriage, few investigations into the contributory molecular mechanisms exist. To this end, we identified a naturally occurring polymorphism in the gene encoding the streptococcal collagen-like protein A (SclA) in GAS carrier strains. All previously sequenced invasive serotype M3 GAS possess a premature stop codon in the sclA gene truncating the protein. The carrier polymorphism is predicted to restore SclA function and was infrequently identified by targeted DNA sequencing in invasive strains of the same serotype. We demonstrate that a strain with the carrier sclA allele expressed a full-length SclA protein, while the strain with the invasive sclA allele expressed a truncated variant. An isoallelic mutant invasive strain with the carrier sclA allele exhibited decreased virulence in a mouse model of invasive disease and decreased multiplication in human blood. Further, the isoallelic invasive strain with the carrier sclA allele persisted in the mouse nasopharynx and had increased adherence to cultured epithelial cells. Repair of the premature stop codon in the invasive sclA allele restored the ability to bind the extracellular matrix proteins laminin and cellular fibronectin. These data demonstrate that a mutation in GAS carrier strains increases adherence and decreases virulence and suggest selection against increased adherence in GAS invasive isolates.     

Diabetes im Kindes- und Jugendalter

Type 1 diabetes is one of the commonest chronic disorders encountered in children and adolescents. When it first becomes apparent in children, approximately 20% of them have clinical and biochemical signs of ketoacidosis (DKA). In the presence of unusual clinical symptoms it is necessary to consider the possibility of associated conditions, such as coeliac disease, immunothyroiditis and Addison’s disease. Children with diabetes must be treated by a multidisciplinary team made up of paediatrician, paediatric diabetes specialist, psychologist, social worker, ophthalmologist, dietitian, nurse and diabetes counsellor, making it essential for them to be treated in regional specialised centres. They are treated in their own psychosocial environment and their families are involved in the therapy. Comprehensive, multidisciplinary treatment strategies have now made it possible for these patients to enjoy normal physical wellbeing and near-normal psychosocial development. Prevention and early treatment of of disturbances associated with diabetes remain an important concern. The fact that type 2 diabetes must now be looked for in overweight children and adolescents is a new aspect of diabetes medicine.     

Influence of Short‐term Confinement and Exercise on Tibia Development in Growing Pigs

The influence of short‐term confinement and moderate exercise on tibia development was investigated in growing pigs (36–41 kg, 10 weeks at the beginning of the study). Animals were kept for 2 weeks either in individual crates of 0.8 m² (‘confinement’, n = 4) or kept in groups of four animals in large crates of 5.6 m² without (‘control’) or with additional exercise (30 min walking at 5 days/week; ‘exercise’). Bone density and morphological parameters were evaluated by computer tomography (CT). Periosteal apposition and longitudinal growth were determined after polychrome sequential labelling in weekly intervals. Cortical areas of the cross sections at the mid‐shaft and at 75% of the length of the bone measured distal to the carpus by CT were significantly correlated to each other (r = 0.70) and revealed a significant reduction in confined animals. This difference was explained by lower periosteal apposition rates in these animals compared with exercised and group‐housed pigs. Similarly longitudinal growth and the formation and calcification of spongiosa of the tibiae were inhibited by confinement. Thus, 2 weeks of confinement led to significant differences in bone growth and metabolism in young growing piglets when compared with animals with moderate activity.     

Liver transplantation in children with biliary atresia and polysplenia syndrome

Biliary atresia is the most common indication for orthotopic liver transplantation (OLT) in children. The polysplenia syndrome anomalies, which occur in approximately 10% of children with biliary atresia, may represent special difficulties at liver transplantation. We have reviewed our experience with this syndrome in 116 children with biliary atresia who underwent liver transplantation between March 1984 and December 1989. The main features of the polysplenia syndrome, which included absence of the inferior vena cava, preduodenal portal vein, midgut malrotation, aberrant hepatic artery, and situs inversus, were encountered in 12 of the 116 children (10.3%). Severe portal vein hypoplasia (3.5 mm or smaller) was also present in 7 of these children. Eight patients received a complete and four received a reduced liver graft. The vascular anomalies increased the technical difficulty of OLT but could be surmounted, although they did contribute to the peroperative death of one child. The 1-month survival rate was 83% for the 12 children with features of the polysplenia syndrome and 88% for the other 92 children with biliary atresia alone.     

Detergent fractionation with subsequent subtractive suppression hybridization as a tool for identifying genes coding for plasma membrane proteins

Abstract:  The identification of tumor-specific proteins located at the plasma membrane is hampered by numerous methodological pitfalls many of which are associated with the post-translational modification of such proteins. Here, we present a new combination of detergent fractionation of cells and of subtractive suppression hybridization (SSH) to gain overexpressed genes coding for membrane-associated or secreted proteins. Fractionation of subcellular components by digitonin allowed sequestering mRNA of the rough Endoplasmatic reticulum and thereby increasing the percentage of sequences coding for membrane-bound proteins. Fractionated mRNAs from the cutaneous T-cell lymphoma (CTCL) cell line HuT78 and from normal peripheral blood monocytes were used for SSH leading to the enrichment of sequences overexpressed in the tumor cells. We identified some 21 overexpressed genes, among them are GPR137B, FAM62A, NOMO1, HSP90, SLIT1, IBP2, CLIF, IRAK and ARC. mRNA expression was tested for selected genes in CTCL cell lines, skin specimens and peripheral blood samples from CTCL patients and healthy donors. Several of the detected sequences are clearly related to cancer, but have not yet been associated with CTCL. qPCR confirmed an enrichment of these mRNAs in the rough endoplasmic reticulum fraction. RT-PCR confirmed the expression of these genes in skin specimens and peripheral blood of CTCL patients. Western blotting verified protein expression of HSP90 and IBP2 in HuT78. GPR137B could be detected by immunohistology in HuT78 and in keratinocytes of dysplastic epidermis, but also in sweat glands of healthy skin. In summary, we developed a new technique, which allows identifying overexpressed genes coding preferentially for membrane-associated proteins.