Stable isotope probing: Technical considerations when resolving ¹⁵N-labeled RNA in gradients

RNA based stable isotope probing (SIP) facilitates the detection and identification of active members of microbial populations that are involved in the assimilation of an isotopically labeled compound. ¹⁵N-RNA-SIP is a new method that has been discussed in recent literature but has not yet been tested. Herein, we define the limitations to using ¹⁵N-labeled substrates for SIP and propose modifications to compensate for some of these shortcomings. We have used ¹⁵N-RNA-SIP as a tool for analysing mixed bacterial populations that use nitrogen substrates. After incubating mixed microbial communities with ¹⁵N-ammonium chloride or ¹⁵N₂ we assessed the fractionation resolution of ¹⁵N-RNA by isopycnic centrifugation in caesium trifluoroacetate (CsTFA) gradients. We found that the more isotopic label incorporated, the further the buoyant density (BD) separation between ¹⁵N- and ¹⁴N-RNA, however it was not possible to resolve the labeled from unlabeled RNA definitively through gradient fractionation. Terminal-restriction fragment length polymorphism (T-RFLP) analysis of the extracted RNA and fluorescent in situ hybridisation (FISH) analysis of the enrichment cultures provided some insight into the organisms involved in nitrogen fixation. This approach is not without its limitations and will require further developments to assess its applicability to other nitrogen-fixing environments

Association of nutrition in early childhood with body composition and leptin in later childhood and early adulthood

Objectives: Using data from the Avon Longitudinal Study of Parents and Children (ALSPAC), this study aimed to replicate the finding of the Etude Longitudinale Alimentation Nutrition Croissance des Enfants (ELANCE) that low fat intake in early childhood was associated with increased adiposity in adulthood. Methods: Diet was assessed at 8 and 18 months using 3-day food records. Body composition variables were measured at 9 and 17 years, and serum leptin at 9 years. Associations were modelled using adjusted linear regression. Results: In replication analyses, in contrast to ELANCE, there was a positive association between fat intake (% energy) at 18 months and fat mass (FM) at 9 years (B coefficient 0.10 (95% CI 0.03, 0.20) kg, p = 0.005). There was no association with serum leptin. In extended analyses fat intake at 18 months was positively associated with FM in boys (0.2 (0.00, 0.30), p = 0.008) at 9 years but not in girls. Fat intake was positively associated with serum leptin concentration in boys (0.2 (0.1, 0.4) ng/mL, p = 0.011) but not in girls. Conclusions: Our results did not corroborate the findings from the ELANCE study. A high fat diet in early life may have implications for later childhood and adolescent obesity

Covalent attachment of active enzymes to upconversion phosphors allows ratiometric detection of substrates

Upconverting phosphors (UCPs) convert multiple low energy photons into higher energy emission via the process of photon upconversion and offer an attractive alternative to organic fluorophores for use as luminescent probes. Here, UCPs were capped with functionalized silica in order to provide a surface to covalently conjugate proteins with surface?accessible cysteines. Variants of green fluorescent protein (GFP) and the flavoenzyme pentaerythritol tetranitrate reductase (PETNR) were then attached via maleimide?thiol coupling in order to allow energy transfer from the UCP to the GFP or flavin cofactor of PETNR, respectively. PETNR retains its activity when coupled to the UCPs, which allows reversible detection of enzyme substrates via ratiometric sensing of the enzyme redox state

Antibacterial Gene Transfer Across the Tree of Life

Though horizontal gene transfer (HGT) is widespread, genes and taxa experience biased rates of transferability. Curiously, independent transmission of homologous DNA to archaea, bacteria, eukaryotes, and viruses is extremely rare and often defies ecological and functional explanations. Here, we demonstrate that a bacterial lysozyme family integrated independently in all domains of life across diverse environments, generating the only glycosyl hydrolase 25 muramidases in plants and archaea. During coculture of a hydrothermal vent archaeon with a bacterial competitor, muramidase transcription is upregulated. Moreover, recombinant lysozyme exhibits broad-spectrum antibacterial action in a dose-dependent manner. Similar to bacterial transfer of antibiotic resistance genes, transfer of a potent antibacterial gene across the universal tree seemingly bestows a niche-transcending adaptation that trumps the barriers against parallel HGT to all domains. The discoveries also comprise the first characterization of an antibacterial gene in archaea and support the pursuit of antibiotics in this underexplored group

Henoch Schonlein Purpura – A 5-Year Review and Proposed Pathway

Henoch Schonlein Purpura (HSP) is the commonest systemic vasculitis of childhood typically presenting with a palpable purpuric rash and frequently involving the renal system. We are the first group to clinically assess, critically analyse and subsequently revise a nurse led monitoring pathway for this condition

A direct comparison of errorless and errorful therapy for object name relearning in Alzheimer's disease

Developing rehabilitation techniques to combat cognitive decline is a key goal of healthcare strategies aimed at promoting increased longevity and better quality of life for individuals with Alzheimer's disease (AD). In AD, problems with episodic memory and word-finding greatly affect everyday life and, as such, these symptoms provide a clear clinical target for therapeutic interventions. Errorless learning (EL) has been proposed as a particularly effective technique for relearning in individuals with memory dysfunction, including AD. However, EL learning has rarely been directly contrasted with other more traditional trial-and-error techniques (errorful learning or EF) in individuals with AD, especially in the context of alleviating word-finding problems. In the current study, we directly contrasted the therapeutic gains of an EL learning paradigm (consisting of reading/repetition of object names) with an EF learning technique (comprised of phonemic/orthographic cueing) in eight mild to moderate AD patients with pronounced anomia. Both techniques were administered concurrently in sessions run twice a week over a five-week period. Therapeutic gains were assessed at one week and five weeks post-intervention using confrontation naming. Our results suggest that, both at the group and individual patient level, EL and EF techniques were equally effective. Correlational analyses of overall therapy gains and background assessments of patient neuropsychology revealed that individuals with better scores on measures of semantic memory, pre-intervention naming, and recognition memory demonstrated larger therapy gains. No individual patient showed a significant advantage for EL over EF learning, however, for patients that showed a numerical advantage in this direction. These results suggest that either EL or EF therapy can be used to alleviate word-finding problems in AD. © 2012 Copyright Psychology Press

Proficiency deficiency: mastery of fundamental movement skills and skill components in overweight and obese children

The purpose of this observational study was to compare the mastery of 12 fundamental movement skills (FMS) and skill components between a treatment-seeking sample of overweight/obese children and a reference sample from the United States. Mastery of six locomotor and six object-control skills (24 components in each subdomain) were video-assessed by one assessor using the test of gross motor development-2 (TGMD-2). The 153 overweight/obese children (mean ± s.d. age = 8.3 ± 1.1 years, BMI z-score = 2.78 ± 0.69, 58% girls, 77% obese) were categorized into age groups (for the underhand roll and strike: 7–8 years and 9–10 years; all other FMS: 6–7 years and 8–10 years) and mastery prevalence rates were compared with representative US data (N = 876) using χ2 analysis. For all 12 skills in all age groups, the prevalence of mastery was lower among overweight/obese children compared with the reference sample (all P < 0.05). This was consistent for 18 locomotor and upto 21 object-control skill components (all P < 0.05). Differences were largest for the run, slide, hop, dribble, and kick. Specific movement patterns that could be targeted for improvement include positioning of the body and feet, the control or release of an object at an optimal position, and better use of the arms to maintain effective force production during the performance of FMS. Physical activity programs designed for overweight and obese children may need to address deficiencies in FMS proficiency to foster the movement capabilities required for participation in health-enhancing physical activity