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Nasopharyngeal carriage of Streptococcus pneumoniae in unimmunised adults and older children in three remote Australian Aboriginal communities was compared in 2002 and 2004. Universal childhood pneumococcal vaccination with a catch-up program was introduced in late 2001. Carriage prevalence across all age groups of pneumococcal serotypes included in the 7-valent vaccine was 10% in both 2002 and 2004 (12 and 30 months after introduction of vaccination). This carriage prevalence was lower than anticipated. It is likely that indirect effects of childhood vaccination occurred before the 2002 survey. To further assess indirect effects on carriage of childhood pneumococcal vaccination, data prior to 2002 are required. Between 12 and 30 months following introduction of conjugate pneumococcal vaccination, indirect effects on carriage were unchanged.

The dependence of metabolic processes on temperature constrains the behavior, physiology and ecology of many ectothermic animals. The evolution of nocturnality in lizards, especially in temperate regions, requires adaptations for activity at low temperatures when optimal body temperatures are unlikely to be obtained. We examined whether nocturnal lizards have cold-adapted lactate dehydrogenase (LDH). LDH was chosen as a representative metabolic enzyme. We measured LDH activity of tail muscle in six lizard species (n=123: three nocturnal, two diurnal and one crepuscular) between 5 and 35 degrees C and found no differences in LDH-specific activity or thermal sensitivity among the species. Similarly, the specific activity and thermal sensitivity of LDH were similar between skinks and geckos. Similar enzyme activities among nocturnal and diurnal lizards indicate that there is no selection of temperature specific LDH enzyme activity at any temperature. As many nocturnal lizards actively thermoregulate during the day, LDH may be adapted for a broad range of temperatures rather than adapted specifically for the low temperatures encountered when the animals are active. The total activity of LDH in tropical and temperate lizards is not cold-adapted. More data are required on biochemical adaptations and whole animal thermal preferences before trends can be established.

Background: Australian Indigenous children living in remote areas have rates of tympanic membrane perforation as high as 60%, almost 100 times the prevalence in urban child care settings (<1%). Relative rates of pneumococcal nasal carriage do not reflect this difference in disease risk., Methods: Cross-sectional comparison of nasal carriage and hand contamination in children younger than 4 years of age from urban child-care centers and Indigenous children 3-7 years of age from a remote community. Almost identical methods of nasal swab collection, transport and culture were used. Data on pneumococcal antimicrobial susceptibility patterns and serotypes are also reported., Results: For Indigenous children compared with children in child care, the relative risk of nasal carriage of either pneumococcus or noncapsular Haemophilus influenzae was <2-fold [relative risk, 1.7; 95% confidence interval (CI), 1.5, 1.9], the risk of simultaneous nasal carriage was almost 3-fold (78% versus 28%; relative risk, 2.9; 95% CI 2.3, 3.5), and the risk of pneumococcal hand contamination was 8-fold higher (37% versus 4%; relative risk, 8.4; 95% CI 4.6, 15.2). For simultaneous hand contamination, the risk was 23-fold (8% versus 0.3%; relative risk, 23.1; 95% CI 2.9, 185.4). Remote Indigenous children also had a more diverse serotype distribution (25 versus 14 serotypes identified)., Conclusions: Simultaneous nasal carriage of Streptococcus pneumoniae and H. influenzae and hand contamination are simple indicators of risk for use in studies of otitis media in populations at risk for tympanic membrane perforation.

Background: Peptides of the trefoil factor family (TFF1, TFF2 and TFF3) are cosecreted with mucus from mucus-producing cells in most organ systems and are believed to interact with mucus to form high-viscosity stable gel complexes. In the gastrointestinal tract, they sustain the mucosal barrier, and both injected and orally administered TFF peptide have protective and healing functions in the gastric mucosa., Aim: To investigate the possible treatment effect of luminally and parenterally administered TFF peptides in experimental colitis in rats., Methods: Colitis was induced by administration of 5% dextran sodium sulphate in the drinking water or by one intraperitoneal injection of mitomycin C, 3.75 mg/kg. TFF peptides were administered as subcutaneous injections or directly into the lumen via a catheter placed in the proximal colon. Treatments were saline, TFF2, TFF3 monomer or TFF3 dimer 5 mg/kg twice per day throughout the study [dextran sulphate sodium (DSS)] or from day 4 to 7 (mitomycin C). Colitis severity was scored in a stereomicroscope and histologically., Results: Luminal treatment with TFF3 in its dimeric form significantly improved the colitis score in both colitis models, whereas TFF2 had positive effect only in DSS-induced colitis. The TFF3 monomer was without any effects in both models. Treatment effect was most pronounced in the middle part of the colon, closest to the tip of the catheter. Injected TFF peptides, especially the TFF3 monomer, aggravated the colitis score in both colitis models., Conclusions: Intracolonic administration of TFF3 dimer and TFF2 improves experimentally induced colitis in rats. The TFF3 monomer has no effect. Parenteral administration of TFF peptides aggravates the colitis especially the TFF3 monomer.

The rate of oxygen consumption (VO2) is commonly used as a measure of whole organism metabolic rate, but requires the animal to be motionless and at rest. Few studies have measured whether animals that appear motionless are truly at rest, or whether being in a novel environment elevates metabolic rate. We investigated whether conditioning of the gecko Naultinus manukanus to experimental procedures influenced the VO2 and probability of achieving a constant rate of oxygen consumption. Metabolic rate was measured at 24 degrees C in 22 individuals until a steady-state was achieved, or for 80 min if no steady-state was reached, once a day on 5 consecutive days (five trials). Geckos in the first trial, when compared with subsequent trials, had a significantly higher mass-adjusted VO2 (0.89+/-0.06 vs. 0.67+/-0.05 ml O2 h(-1), respectively), and time to reach a steady-state VO2 (66+/-8 vs. 47+/-3 min, respectively), as well as a significantly lower probability of reaching a steady-state VO2 (24% vs. 74%, respectively). We conclude that there may be hidden inaccuracies in studies that do not condition animals and that at least one conditioning trial should be used to obtain a metabolic rate at rest for small lizards.

Interactions with thymic stromal cells are known to be critical for the development of T cells from progenitors entering the thymus, yet the molecular mechanisms of stromal cell function remain poorly understood. Accumulating evidence has highlighted the importance of beta-catenin-mediated activation of T cell factor (TCF)/lymphoid enhancer factor (LEF) transcription during thymocyte development. As regulation of this signaling pathway is controlled by binding of soluble Wnt proteins to cell surface Frizzled (Fz) receptors, we studied components of Wnt/Fz-mediated signaling in thecontext of stromal cell regulation of thymocyte development. We show that mRNA for a variety of Wnt family members, notably Wnt-4, Wnt-7a and 7b, and Wnt-10a and 10b, are expressed by thymic epithelium rather then by thymocytes, while thymocytes demonstrate a developmentally regulated pattern of Fz receptor expression. Collectively these findings suggest (1) a functional role for Wnt-producing thymic epithelium in determining TCF/LEF-mediated transcriptional regulation in Fz-bearing thymocytes, and (2) a role for defined Wnt-Fz interactions at successive stages of thymocyte maturation. In support of this we show that separation of thymocytes from Wnt-producing epithelial cells and the thymic microenvironment, triggers beta-catenin phosphorylation and degradation in thymocytes. Thus, sustained exposure to Wnt in the context of an intact stromal microenvironment is necessary for stabilization of beta-catenin-mediated signaling in thymocytes.

Following their migration into the thymus, hemopoeitic stem cell precursors enter a complex developmental pathway involving proliferation, differentiation and alphabetaT-cell receptor (alphabetaTCR)-mediated selection procedures, in order to generate mature T-cell populations ready for export to the periphery. Thus, a critical stage during intrathymic T-cell development involves the generation of functionally mature CD4+8- and CD4-8+ cells from immature CD4+8- precursor thymocytes, a poorly understood process referred to as positive selection. While interactions between the alphabetaTCR and MHC-peptide complexes are known to be essential for the initiation of positive selection, additional unknown signals are also required. Using an in vitro reaggregate thymic organ culture system which allows comparison of the abilities of various cell types to induce maturation of CD4+8+ precursors, we provide evidence that both MHC-peptide complexes and specialised accessory molecules must be provided by thymic epithelium for efficient mediation of positive selection. Moreover, analysis of positive selection in the presence of thymic and non-thymic stromal cells expressing MHC class II molecules with the same limited peptide array suggests that this unique ability of thymic epithelium to mediate positive selection of CD4+8- cells is not solely due to presentation of a specialised peptide repertoire, but is dependent upon provision of specialised accessory interactions.

T cell development in the thymus occurs through a series of events beginning with thymic colonization by migrant precursors and ending with the emigration of functionally competent CD4+ and CD8+ T cells to the periphery. It is well accepted that signals through the pre-T cell receptor (pre-TCR) and alpha-beta TCR (alphabetaTCR) complex play pivotal roles in the maturation of CD4-8- and CD4+8+ thymocytes, respectively. It is clear that stromal cells constituting the thymic microenvironment provide non-TCR-mediated interactions which are essential for several developmental events. Examples of such will be discussed here in relation to early and late events in T cell development.

Intrathymic T cell development is a multistage process involving discrete phases of proliferation as well as differentiation. From studies on IL-7 or IL-7Ralpha-deficient mice, it is clear that the IL-7 receptor (IL-7R) plays a critical role during the initial stages of intrathymic CD4-8- precursor development. In contrast, the role of IL-7R in later stages of thymocyte development are unclear. Here, we have used various approaches to investigate directly the role of the IL-7R in thymocyte positive selection and the recently described phase of postselection proliferation. First, we show that positive selection involves selective up-regulation of IL-7Ralpha- and IL-7Rgamma-chains, with the majority of CD4+ and CD8+ cells being IL-7R+. Second, MHC class II+ thymic epithelium-which drives postselection proliferation-expresses IL-7 mRNA. Finally, analysis of positive selection and postselection proliferation in thymocytes from IL-7Ralpha-/- neonates shows that positive selection occurs normally, whereas postselection expansion is drastically reduced. Thus, our data provide the first evidence that, as well as playing a role during early phases of thymic development, IL-7R mediates intrathymic expansion of positively selected thymocytes, which may aid in establishment of the neonatal peripheral T cell pool.

Positive selection is a crucial stage in T-cell development because it is here that CD4+CD8+ cells bearing T-cell receptors that interact with self-major histocompatibility complex molecules are rescued from cell death, resulting in the generation of mature T cells. Here, Graham Anderson and colleagues review recent studies indicating that positive selection is a multistage process involving interactions with thymic epithelium.

In the thymus, phenotypically and functionally mature single positive cells are generated from immature CD4+8+ precursors by a process known as positive selection. Although this event is known to involve alphabetaTCR ligation by peptide/MHC complexes expressed on thymic stromal cells, it is clear that positive selection is a multistage process involving transition through an intermediate CD4+8+69+ phase as well as subsequent postselection phases. By analyzing the development of preselection CD4+8+69- and intermediate CD4+8+69+ thymocytes in the presence of MHC class I-deficient, MHC class II-deficient, and MHC double-deficient thymic stromal cells, we investigated the role of MHC molecules at three distinct points during positive selection. Although the initiation of positive selection is critically dependent upon MHC interactions, we find the that later stages of maturation, involving the differentiation of CD4+8- and CD4-8+ cells from CD4+8+69+ thymocytes, occur in the absence of MHC molecules. Moreover, an analysis of the postselection proliferation of newly generated CD4+8- and CD4-8+ thymocytes shows that this also occurs independently of MHC molecules. Thus, our data provide direct evidence that, although positive selection is a multistage process initiated by TCR-MHC interactions, continuation of this process and subsequent postselection events are independent of ongoing engagement of the TCR.

A number of in vitro systems are currently being used to study both thymocyte development and thymic stromal cell function. However, the usefulness of dispersed culture systems is limited since they often involve disruption of interactions within the normal three-dimensional architecture of the thymus in vivo which are critical for normal development to proceed. In contrast, Fetal Thymus Organ Culture (FTOC) provides an experimental system where such interactions are maintained, thereby allowing in vitro access to key aspects of thymocyte development. More recently, Reaggregate Thymus Organ Cultures (RTOCs) have allowed detailed analysis of thymic stromal cell function, while retroviral transfection of thymocyte subsets under FTOC conditions provides a rapid means to investigate thymocyte development at the molecular level. Current use of the FTOC approach is summarised here, and where appropriate is compared to the use of dispersed culture systems., (Copyright 1999 Academic Press.)

To investigate events following the initiation of positive selection, we have used reaggregate organ cultures to follow the maturation of purified CD4+8+69+ thymocytes; these thymocytes represent a subpopulation of thymocytes which have already received positive selection signals. Using a dilution analysis of an FITC-based membrane-binding dye, 5-(and -6)-carboxyfluorescein diacetate succinimidyl ester, to allow a quantitative measure of proliferation, we show that while newly selected CD4+ and CD8+ cells are nondividing, both subsets subsequently undergo a wave of postpositive selection proliferation involving multiple cell divisions. Moreover, in the presence of fetal stromal cells, postselection expansion is more extensive in newborn thymocytes compared with adult thymocytes, suggesting that this phase of expansion is developmentally regulated. We also show that proliferation of CD4+ and CD8+ cells is seen in reaggregates of purified MHC class II+ thymic epithelial cells, while CD4+ and CD8+ cells generated from bcl-2 transgenic CD4+8+69+ thymocytes in the absence of stromal cell support survive but do not proliferate; this observation indicates that MHC class II+ thymic epithelial cells are both necessary and sufficient to mediate this wave of cell division. Finally, the maturation of CD4+8+69+ thymocytes and the subsequent proliferation of CD4+ and CD8+ cells occur in the presence of MHC-mismatched thymic stromal cells, suggesting that the later stages of positive selection and the associated postselection events do not depend on interactions with the same peptide/MHC complexes responsible for initiation.

As well as signaling through the alphabeta T cell receptor complex, positive selection of immature CD4+ 8+ thymocytes involves additional ill-defined accessory interactions provided by thymic epithelial cells. Here, we have isolated CD4+ 8+ thymocytes at a pre-positive selection stage of development (TCR- CD69- 4+ 8+ cells), or after initiation of positive selection (CD69+ 4+ 8+ cells), from mice where the normal lifespan of thymocytes is extended by the presence of a bcl-2 transgene, to allow us to discriminate between requirements for maintenance and differentiation signals during positive selection. We find that MHC class II+ thymic epithelial cells drive positive selection of TCR- CD69- 4+ 8+ bcl-2 tg thymocytes to the CD4+ and CD8+ stage, while no such mature subsets are observed when thymocytes are cultured alone or with major histocompatibility complex (MHC) class II+ salivary epithelial cells. However, CD4+ 8+ cells remain in such cultures in considerable numbers, and retain the potential for positive selection if re-cultured with thymic epithelium, suggesting that thymic epithelial cells provide specific differentiation-inducing signals for positive selection. In contrast, intermediate CD69+ 4+ 8+ thymocytes show some capacity for phenotypic conversion in the absence of thymic stromal cells although strikingly the single-positive CD4+ and CD8+ cells generated are not functionally competent. Finally, we show that prior culture of thymic epithelial cells under monolayer conditions abrogates their ability to support the initiation of positive selection, suggesting that the epithelial cell molecules necessary for the provision of differentiation signals during positive selection are down-regulated under such conditions.

A case is described of a patient with an oat cell carcinoma of the bronchus with moderately elevated levels of plasma corticotrophin (ACTH) and antidiuretic hormone (ADH). Ectopic secretion of ACTH induced severe hypokalaemia and concealed the effects of concomitant ADH secretion on renal function. Normal renal responsiveness was restored following correction of hypokalaemia. The hypokalaemia was associated with evidence of a marked increase in corticosteroid secretion but plasma ACTH concentrations did not show a proportionate elevation. Chromatographic studies on tumour extracts suggest that the presence of a large fraction of high molecular weight ACTH in plasma could explain this discrepancy.