Localization of transforming growth factor-beta at the human fetal-maternal interface: role in trophoblast growth and differentiation.

We examined the localization of transforming growth factor (TGF)-beta in first-trimester and term human decidua and chorionic villi and explored the role of this factor on the proliferation and differentiation of cultured trophoblast cells. Two antibodies, 1D11.16.8, a mouse monoclonal neutralizing antibody capable of recognizing both TGF-beta 1 and TGF-beta 2 and CL-B1/29, a rabbit polyclonal antibody capable of recognizing TGF-beta 2, were used to immunolocalize TGF-beta in fixed, paraffin-embedded, or fixed, frozen sections of placenta and decidua, providing similar results. Intense labeling was observed in the extracellular matrix (ECM) of the first-trimester decidua and cytoplasm of term decidual cells. Syncytiotrophoblast cell cytoplasm as well as the ECM in the core of the chorionic villi of both first-trimester and term placentas exhibited a moderate degree of labeling. Strong cytoplasmic labeling was observed in the cytotrophoblastic shell of the term placenta. To examine the role of TGF-beta on trophoblast proliferation and differentiation, early passage cultures of first-trimester and primary cultures of term trophoblast cells were established and characterized on the basis of numerous immunocytochemical and functional markers. These cells expressed cytokeratin, placental alkaline phosphatase, urokinase-type plasminogen activator, and pregnancy-specific beta glycoprotein, but not factor VIII or 63D3; they also produced hCG and collagenase type IV. Exposure of first-trimester trophoblast cultures to TGF-beta 1 significantly inhibited proliferation in a dose-dependent manner. An antiproliferative effect was also noted in the presence of TGF-beta 2. These effects were abrogated in the presence of the neutralizing anti-TGF-beta antibody (1D11.16.8) in a concentration-dependent manner. In a 3-day culture, exogenous TGF-beta 1 stimulated formation of multinucleated cells by the first trimester as well as term trophoblast cells. Addition of neutralizing anti-TGF-beta antibody to first-trimester trophoblast cells stimulated proliferation beyond control levels in a 24-h culture and reduced formation of multinucleated cells in a 3-day culture, indicating the presence of endogenous TGF-beta activity. These results indicate that TGF-beta produced at the human fetal-maternal interface plays a major regulatory role in the proliferation and differentiation of the trophoblast.     

Regulated temporal and spatial expression of the calcium-binding proteins calcyclin and OPN (osteopontin) in mouse tissues during pregnancy.

In situ hybridization and northern/slot blot analyses were used to quantify the expression of calcyclin (2A9, 5B10), osteopontin (opn, secreted phosphoprotein, 2ar) and calmodulin mRNAs in mouse tissues that support pregnancy. High-to-moderate levels of the mRNAs of all three genes were detected at discrete locations in the uterus, decidua and placenta as a function of gestation time. Calmodulin expression was constant in these tissues; calcyclin mRNA was high during early pregnancy and declined after day 8-9 of gestation; and opn mRNA was undetectable before day 7, with maximal levels on days 9-12 in each of these tissues. Calcyclin, but not opn, expression was also observed in the chorioamnion after day 12. Calcyclin was expressed throughout the decidua on day 8 but became restricted to the primary (antimesometrial) decidual zone and decidua lateralis on day 9, and the decidua capsularis after day 9. By contrast, opn mRNA was localized on day 9 to the mesometrial triangle, which contains a large population of granulated metrial gland cells, and to the decidua basalis. These two genes may serve as markers for the two types of decidual tissue. We suggest that one function of OPN, which may be an indicator of cells in the decidua that have a bone marrow genealogy, is to mediate the flux of calcium from the maternal circulation to the developing embryo.     

Light and electron microscopical study of Nosema eurytremae

A nuclear-polyhedrosis virus (NPV) of the silkworm, Bombyx mori, which forms an icosahedral inclusion body, was transmitted to larvae of the rice stem borer, Chilo suppressalis. Serial passages of Bombyx NPV in the alternate host by injecting the supernatant of diseased hemolymph produced inclusion bodies with cuboidal and other shapes that differed from the original shape formed in Bombyx. These different shapes increased with times of passages, and after the twelfth passage, only cuboidal inclusion bodies were formed. The icosahedral inclusion bodies in B. mori and the cuboidal inclusion bodies in C. suppressalis occluded singly enveloped virions of the same size (350 × 75 nm), but the cuboidal inclusion bodies contained only a few virions and a large number of membraneous spherical structures. The formation process of the cuboidal inclusion body differed from that of the icosahedral. At first, irregularly branched inclusion bodies containing “vacant” spaces appeared in the infected nuclei. The bodies grew larger with the deposition of protein in the spaces between the branches, and this was accompanied with the occlusion of a large number of membraneous structures formed in the vicinity of the inclusion bodies, which became cuboidal in shape.     

Phylogenetic Exploration of Nosocomial Transmission Chains of 2009 Influenza A/H1N1 among Children Admitted at Red Cross War Memorial Children’s Hospital,Cape Town,South Africa in 2011

Traditional modes of investigating influenza nosocomial transmission have entailed a combination of confirmatory molecular diagnostic testing and epidemiological investigation. Common hospital-acquired infections like influenza require a discerning ability to distinguish between viral isolates to accurately identify patient transmission chains. We assessed whether influenza hemagglutinin sequence phylogenies can be used to enrich epidemiological data when investigating the extent of nosocomial transmission over a four-month period within a paediatric Hospital in Cape Town South Africa. Possible transmission chains/channels were initially determined through basic patient admission data combined with Maximum likelihood and time-scaled Bayesian phylogenetic analyses. These analyses suggested that most instances of potential hospital-acquired infections resulted from multiple introductions of Influenza A into the hospital, which included instances where virus hemagglutinin sequences were identical between different patients. Furthermore, a general inability to establish epidemiological transmission linkage of patients/viral isolates implied that identified isolates could have originated from asymptomatic hospital patients, visitors or hospital staff. In contrast, a traditional epidemiological investigation that used no viral phylogenetic analyses, based on patient co-admission into specific wards during a particular time-frame, suggested that multiple hospital acquired infection instances may have stemmed from a limited number of identifiable index viral isolates/patients. This traditional epidemiological analysis by itself could incorrectly suggest linkage between unrelated cases, underestimate the number of unique infections and may overlook the possible diffuse nature of hospital transmission, which was suggested by sequencing data to be caused by multiple unique introductions of influenza A isolates into individual hospital wards. We have demonstrated a functional role for viral sequence data in nosocomial transmission investigation through its ability to enrich traditional, non-molecular observational epidemiological investigation by teasing out possible transmission pathways and working toward more accurately enumerating the number of possible transmission events.     

Evaluating the Potential for the Environmentally Sustainable Control of Foot and Mouth Disease in Sub-Saharan Africa

Strategies to control transboundary diseases have in the past generated unintended negative consequences for both the environment and local human populations. Integrating perspectives from across disciplines, including livestock, veterinary and conservation sectors, is necessary for identifying disease control strategies that optimise environmental goods and services at the wildlife-livestock interface. Prompted by the recent development of a global strategy for the control and elimination of foot-and-mouth disease (FMD), this paper seeks insight into the consequences of, and rational options for potential FMD control measures in relation to environmental, conservation and human poverty considerations in Africa. We suggest a more environmentally nuanced process of FMD control that safe-guards the integrity of wild populations and the ecosystem dynamics on which human livelihoods depend while simultaneously improving socio-economic conditions of rural people. In particular, we outline five major issues that need to be considered: 1) improved understanding of the different FMD viral strains and how they circulate between domestic and wildlife populations; 2) an appreciation for the economic value of wildlife for many African countries whose presence might preclude the country from ever achieving an FMD-free status; 3) exploring ways in which livestock production can be improved without compromising wildlife such as implementing commodity-based trading schemes; 4) introducing a participatory approach involving local farmers and the national veterinary services in the control of FMD; and 5) finally the possibility that transfrontier conservation might offer new hope of integrating decision-making at the wildlife-livestock interface.     

Small geographic range but not panmictic: how forests structure the endangered Point Arena mountain beaver (Aplodontia rufa nigra)

The landscape genetics framework is typically applied to broad regions that occupy only small portions of a species’ range. Rarely is the entire range of a taxon the subject of study. We examined the landscape genetic structure of the endangered Point Arena mountain beaver (Aplodontia rufa nigra), whose isolated geographic range is found in a restricted (85 km²) but heterogenous region in California. Based on its diminutive range we may predict widespread gene flow and a relatively weak role for landscape variation in defining genetic structure. We used skin, bone, tissue and noninvasively collected hair samples to describe genetic substructure and model gene flow. We examined spatial partitioning of multilocus DNA genotypes and mitochondrial DNA haplotypes. We identified 3 groups from microsatellite data, all of which had low estimates of effective population size consistent with significant tests for historical bottlenecks. We used least-cost-path analysis with the microsatellites to examine how vegetation type affects gene flow in a landscape genetics framework. Gene flow was best predicted by a model with “Forest” as the most permeable, followed by “Riparian”. Agricultural lands demonstrated the highest resistance. MtDNA data revealed only two haplotypes: one was represented in all 57 individuals that occurred north of the east–west flowing Garcia River. South of the river, however, both haplotypes occurred, often at the same site suggesting that the river may have affected historical patterns of genetic divergence.     

Dengue Outbreak in Mombasa City,Kenya, 2013–2014: Entomologic Investigations

Dengue outbreaks were first reported in East Africa in the late 1970s to early 1980s including the 1982 outbreak on the Kenyan coast. In 2011, dengue outbreaks occurred in Mandera in northern Kenya and subsequently in Mombasa city along the Kenyan coast in 2013–2014. Following laboratory confirmation of dengue fever cases, an entomologic investigation was conducted to establish the mosquito species, and densities, causing the outbreak. Affected parts of the city were identified with the help of public health officials. Adult Ae. aegypti mosquitoes were collected using various tools, processed and screened for dengue virus (DENV) by cell culture and RT-PCR. All containers in every accessible house and compound within affected suburbs were inspected for immatures. A total of 2,065 Ae. aegypti adults were collected and 192 houses and 1,676 containers inspected. An overall house index of 22%, container index, 31.0% (indoor = 19; outdoor = 43) and Breteau index, 270.1, were observed, suggesting that the risk of dengue transmission was high. Overall, jerry cans were the most productive containers (18%), followed by drums (17%), buckets (16%), tires (14%) and tanks (10%). However, each site had specific most-productive container-types such as tanks (17%) in Kizingo; Drums in Nyali (30%) and Changamwe (33%), plastic basins (35%) in Nyali-B and plastic buckets (81%) in Ganjoni. We recommend that for effective control of the dengue vector in Mombasa city, all container types would be targeted. Measures would include proper covering of water storage containers and eliminating discarded containers outdoors through a public participatory environmental clean-up exercise. Providing reliable piped water to all households would minimize the need for water storage and reduce aquatic habitats. Isolation of DENV from male Ae. aegypti mosquitoes is a first observation in Kenya and provides further evidence that transovarial transmission may have a role in DENV circulation and/or maintenance in the environment.     

Functional effects of enhancing or silencing adenosine A2b receptors in cardiac fibroblasts

Cardiac fibroblasts (CF) express adenosine (ADO) receptors, and pharmacological evidence suggests the possible involvement of the A2 (A2a and A2b) receptor (A2aR and A2bR) subtypes in inhibiting cell functions involved in fibrosis. The main objective of this study was to define the contributions of A2a and/or A2b receptors in modulating ADO-induced decreases in CF functions. For this purpose, CF were either treated pharmacologically or had the A2aR or A2bR levels modified through the use of recombinant adenovirus or siRNA. The assessment of mRNA expression in adult rat CF yielded evidence for A1R, A2bR, A2a), and A3R. Endogenously or exogenously enhanced ADO significantly inhibits CF proliferation, collagen, and protein synthesis. A2R and A2aR agonists, although capable of inhibiting CF protein and collagen synthesis, were unable to define the contributions derived from A2aR or A2bR. Overexpression of A2bR in CF yielded significant decreases in basal levels of collagen and protein synthesis and correlated with increases in cAMP levels. However, at higher doses of ADO receptor agonists, significant increases in protein and collagen synthesis were observed. CF with underexpression of A2bR yielded increases in protein and collagen synthesis. In contrast, A2aR underexpression did not modify ADO-induced decreases in CF protein or collagen synthesis. In conclusion, results derived from the molecular manipulation of receptor levels indicate that A2bR are critically involved in ADO-mediated inhibition of CF functions.