Electrochemical detection of gene expression in tumor samples: overexpression of Rak nuclear tyrosine kinase

Absolute quantification of Rak nuclear tyrosine kinase mRNA in breast tissue samples was determined by competitive RT-PCR. The total RNA from the same samples was also chemically amplified through conventional RT-PCR, and the relative amounts of these amplified RT-PCR products were determined by adsorption onto an indium tin oxide (ITO) electrode followed by electrochemical detection. The electrochemical detection was performed using the inorganic metal complex Ru(bpy)(3)(2+) (bpy = 2,2' bipyridine) to catalyze the oxidation of the guanine residues of the immobilized RT-PCR products. Using the competitive RT-PCR values as standards, it was found that an optimized conventional RT-PCR coupled with electrochemical detection provides a simple method for measuring relative gene expression among a series of mRNA samples from breast tumors. The use of electrochemical detection potentially eliminates the need for gel electrophoresis and fluorescent or radioactive labels in detecting the target genes.     

Expression patterns of chondrocyte genes cloned by differential display in tibial dyschondroplasia

Tibial dyschondroplasia (TD) appears to involve a failure of the growth plate chondrocytes within growing long bones to differentiate fully to the hypertrophic stage, resulting in a mass of prehypertrophic chondrocytes which form the avascular TD lesion. Many biochemical and molecular markers of chondrocyte hypertrophy are absent from the lesion, or show reduced expression, but the cause of the disorder remains to be identified. As differentiation to the hypertrophic state is impaired in TD, we hypothesised that chondrocyte genes that are differentially expressed in the growth plate should show altered expression in TD. Using differential display, four genes, B-cadherin, EF2, HT7 and Ex-FABP were cloned from chondrocytes stimulated to differentiate to the hypertrophic stage in vitro, and their differential expression confirmed in vivo. Using semi-quantitative RT-PCR, the expression patterns of these genes were compared in chondrocytes from normal and TD growth plates. Surprisingly, none of these genes showed the pattern of expression that might be expected in TD lesion chondrocytes, and two of them, B-cadherin and Ex-FABP, were upregulated in the lesion. This indicates that the TD phenotype does not merely reflect the absence of hypertrophic marker genes, but may be influenced by more complex developmental mechanisms/defects than previously thought.     

Cannibalism or congeneric predation? The African clawed frog,Xenopus laevis (Daudin), preferentially predates on larvae of Cape platannas,Xenopus gilli Rose & Hewitt

Predators are not limited to prey from other species as they can cannibalise vulnerable individuals within their own population. The African clawed frog, Xenopus laevis (Daudin), is a predator with a broad diet, known to consume multiple prey species, including congeners and conspecifics. African clawed frogs occur in sympatry with the Endangered Cape platanna, Xenopus gilli Rose & Hewitt, which are under threat through competition and predation from X. laevis. We investigated the threat of X. laevis predation on X. gilli using choice and no‐choice experiments to evaluate the relative vulnerability of X. laevis and X. gilli larvae. Results showed that large X. gilli larvae had a significantly higher vulnerability to X. laevis predation compared to small X. gilli larvae. However, the same discrimination was not discerned when offered large and small X. laevis larvae, or mixed larvae of the same size. We report ontogenic shifts in behaviour of X. gilli larvae that may be a factor in contributing to the vulnerability of large X. gilli larvae to adult X. leavis predation. Congeneric predation likely has negative implications for the population structure of the Endangered X. gilli. Our study underlines the call for the removal of X. laevis to conserve populations of X. gilli.     

How repeatable is the Environmental Impact Classification of Alien Taxa (EICAT)? Comparing independent global impact assessments of amphibians

The magnitude of impacts some alien species cause to native environments makes them targets for regulation and management. However, which species to target is not always clear, and comparisons of a wide variety of impacts are necessary. Impact scoring systems can aid management prioritization of alien species. For such tools to be objective, they need to be robust to assessor bias. Here, we assess the newly proposed Environmental Impact Classification for Alien Taxa (EICAT) used for amphibians and test how outcomes differ between assessors. Two independent assessments were made by Kraus (Annual Review of Ecology Evolution and Systematics, 46, 2015, 75‐97) and Kumschick et al. (Neobiota, 33, 2017, 53‐66), including independent literature searches for impact records. Most of the differences between these two classifications can be attributed to different literature search strategies used with only one‐third of the combined number of references shared between both studies. For the commonly assessed species, the classification of maximum impacts for most species is similar between assessors, but there are differences in the more detailed assessments. We clarify one specific issue resulting from different interpretations of EICAT, namely the practical interpretation and assigning of disease impacts in the absence of direct evidence of transmission from alien to native species. The differences between assessments outlined here cannot be attributed to features of the scheme. Reporting bias should be avoided by assessing all alien species rather than only the seemingly high‐impacting ones, which also improves the utility of the data for management and prioritization for future research. Furthermore, assessments of the same taxon by various assessors and a structured review process for assessments, as proposed by Hawkins et al. (Diversity and Distributions, 21, 2015, 1360), can ensure that biases can be avoided and all important literature is included.     

Plasmodium falciparum parasitaemia in the first half of pregnancy, uterine and umbilical artery blood flow, and foetal growth: a longitudinal Doppler ultrasound study

ABSTRACT: BACKGROUND: During early pregnancy, the placenta develops to meet the metabolic demands of the foetus. The objective of this analysis was to examine the effect of malaria parasitaemia prior to 20 weeks' gestation on subsequent changes in uterine and umbilical artery blood flow and intrauterine growth restriction. METHODS: Data were analysed from 548 antenatal visits after 20 weeks' gestation of 128 women, which included foetal biometric measures and interrogation of uterine and umbilical artery blood flow. Linear mixed effect models estimated the effect of early pregnancy malaria parasitaemia on uterine and umbilical artery resistance indices. Log-binomial models with generalized estimating equations estimated the effect of early pregnancy malaria parasitaemia on the risk of intrauterine growth restriction. RESULTS: There were differential effects of early pregnancy malaria parasitaemia on uterine artery resistance by nutritional status, with decreased uterine artery resistance among nourished women with early pregnancy malaria and increased uterine artery resistance among undernourished women with early pregnancy malaria. Among primigravidae, early pregnancy malaria parasitaemia decreased umbilical artery resistance in the late third trimester, likely reflecting adaptive villous angiogenesis. In fully adjusted models, primigravidae with early pregnancy malaria parasitaemia had 3.6 times the risk of subsequent intrauterine growth restriction (95% CI: 2.1, 6.2) compared to the referent group of multigravidae with no early pregnancy malaria parasitaemia. CONCLUSIONS: Early pregnancy malaria parasitaemia affects uterine and umbilical artery blood flow, possibly due to alterations in placentation and angiogenesis, respectively. Among primigravidae, early pregnancy malaria parasitaemia increases the risk of intrauterine growth restriction. The findings support the initiation of malaria parasitaemia prevention and control efforts earlier in pregnancy.     

Valley‐scale hydrogeomorphology drives river fish assemblage variation in Mongolia

River hydrogeomorphology is a major driver shaping biodiversity and community composition. Here, we examine how hydrogeomorphic heterogeneity expressed by Functional Process Zones (FPZs) in river networks is associated with fish assemblage variation. We examined this association in two distinct ecoregions in Mongolia expected to display different gradients of river network hydrogeomorphic heterogeneity. We delineated FPZs by extracting valley‐scale hydrogeomorphic variables at 10 km sample intervals in forest steppe (FS) and in grassland (G) river networks. We sampled fish assemblages and examined variation associated with changes in gradients of hydrogeomorphology as expressed by the FPZs. Thus, we examined assemblage variation as patterns of occurrence‐ and abundance‐based beta diversities for the taxonomic composition of assemblages and as functional beta diversity. Overall, we delineated 5 and 6 FPZs in river networks of the FS and G, respectively. Eight fish species were found in the FS river network and seventeen in the G, four of them common to both ecoregions. Functional richness was correspondingly higher in the G river network. Variation in the taxonomic composition of assemblages was driven by species turnover and was only significant in the G river network. Abundance‐based taxonomic variation was significant in river networks of both ecoregions, while the functional beta diversity results were inconclusive. We show that valley‐scale hydrogeomorphology is a significant driver of variation in fish assemblages at a macrosystem scale. Both changes in the composition of fish assemblages and the carrying capacity of the river network were driven by valley‐scale hydrogeomorphic variables. River network hydrogeomorphology as accounted for in the study has, therefore, the potential to inform macrosystem scale community ecology research and conservation efforts.     

Potential effects on zooplankton from species shifts in planktivorous mussels: a field experiment in the St Lawrence River

1. Suspension feeding by bivalves exceeds that by other planktivores in many North American rivers, and food webs may be altered substantially by differences in feeding patterns between native unionid mussels and invading dreissenid mussels. 2. We conducted an experiment comparing zooplanktivory by one unionid and two dreissenid species that addressed several primary questions. Is benthic planktivory important in this river? Has this linkage been altered substantially by dreissenids? Do the two dreissenid species differ in planktivory, and is this ecologically important if quagga mussels extend their geographical range? 3. Our 12‐day experiment consisted of controls (no mussels) and treatments with unionid (Elliptio complanata), quagga (Dreissena bugensis) or zebra (D. polymorpha) mussels in 3500‐L, 80‐μm mesh enclosures placed in a slackwater area of the St Lawrence River. 4. The density of the most abundant calanoid copepod Eurytemora affinis increased in the presence of dreissenids, probably as an indirect food web response. By day 12, a cumulative effect was shown by the most overwhelmingly abundant rotifer, Polyarthra, whose density declined dramatically in dreissenid enclosures compared with control and unionid enclosures. Rotifer densities in unionid enclosures were not different from controls, nor were dreissenid treatments different from each other. The effects on rotifers were probably from predation, as Chl‐a did not vary among treatments. 5. We conclude that benthic‐pelagic coupling via planktivory is important in slackwater areas. Dreissenids have strengthened this linkage, but range extension of quaggas should not appreciably alter effects produced by a similar biomass of zebra mussels.     

Temporal dynamics influenced by global change: bee community phenology in urban,agricultural, and natural landscapes

Urbanization and agricultural intensification of landscapes are important drivers of global change, which in turn have direct impacts on local ecological communities leading to shifts in species distributions and interactions. Here, we illustrate how human‐altered landscapes, with novel ornamental and crop plant communities, result not only in changes to local community diversity of floral‐dependent species, but also in shifts in seasonal abundance of bee pollinators. Three years of data on the spatio‐temporal distributions of 91 bee species show that seasonal patterns of abundance and species richness in human‐altered landscapes varied significantly less compared to natural habitats in which floral resources are relatively scarce in the dry summer months. These findings demonstrate that anthropogenic environmental changes in urban and agricultural systems, here mediated through changes in plant resources and water inputs, can alter the temporal dynamics of pollinators that depend on them. Changes in phenology of interactions can be an important, though frequently overlooked, mechanism of global change.     

Maternal investment and size-number trade-off in a bee, Megachile apicalis, in seasonal environments

Maternal investment in offspring size and number differed between spring- and summer-emerging individual females of Megachile apicalis, a solitary multivoltine bee. Data from experimentally initiated female populations indicated that spring-emerging females produced a relatively large number of progeny but allocated a small amount of food to each, resulting in small progeny. Adult females of larger body sizes provisioned food at a greater rate than did smaller females, and this body-size effect was significant in spring-emerging females. The large body size of these females allowed them to increase the number of progeny produced under the abundant floral resources that occurred during the spring. Conversely, summer-emerging females produced fewer progeny under the diminishing resources for brood production, but allocated each with more food, producing larger progeny, most of which emerged in the spring of the following year. Field data using trap-nests also indicated the same pattern of seasonal offspring size allocation found in the experimental populations. This maternal investment strategy entails a trade-off between the size and number of progeny, so that the daughters upon emergence can best perform in their brood production under the seasonally variable environments where they reproduce.     

Tumour necrosis factor induction of ELAM-1 and ICAM-1 on human umbilical vein endothelial cells--analysis of tumour necrosis factor-receptor interactions.

Induction of the adhesion molecules ELAM-1 and ICAM-1 on endothelial cells is a key pro-inflammatory effect of tumour necrosis factor (TNF). Earlier work in non-human systems has suggested that unlike other cell types, endothelial cells interact with the N-terminus of the TNF molecule, thereby implying novel TNF receptors on endothelial cells. This is also supported by 125I-TNF cross-linking studies on bovine endothelial cells. The present study aimed to see whether TNF induction of ELAM-1 and ICAM-1 on human umbilical vein endothelial cells (HUVECs) involved novel TNF-receptor interactions. Three approaches were employed. First, antibodies directed at different sites on the TNF molecule were tested for inhibition of TNF-induction of ELAM-1 and ICAM-1 on HUVECs. Inhibition was seen only with antibodies reacting with epitopes outside the N-terminal region. Second, an N-terminal TNF peptide (residues 1-26) failed to induce ELAM-1 and ICAM-1 on HUVECs or antagonise TNF induction of these molecules. Third, HUVEC/125I-TNF cross-linking revealed a major complex characteristic of the known 55 kDa TNF receptor: this was confirmed with receptor-specific monoclonal antibodies. It is concluded that (a) the same part of the TNF molecule interacts with TNF-receptors on HUVECs and other cell types and (b) TNF induction of ELAM-1 and ICAM-1 on HUVECs is mediated via the well-characterized 55 kDa TNF receptor.