Mono-sulfonated tetrazolium salt based NAD(P)H detection reagents suitable for dehydrogenase and real-time cell viability assays

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and is important for several biological processes. For GDH inhibitor screening, we developed a novel mono-sulfonated tetrazolium salt (EZMTT), which can be synthesized using H₂O₂ oxidation and purified easily on silica gel in large quantities. The EZMTT detection method showed linear dose responses to NAD(P)H, dehydrogenase concentration and cell numbers. In E. coli GDH assay, the EZMTT method showed excellent assay reproducibility with a Z factor of 0.9 and caused no false positives in the presence of antioxidants (such as BME). Using the EZMTT-formazan-NAD(P)H system, we showed that EGCG is a potent E. coli GDH inhibitor (IC₅₀ 45 nM) and identified that Ebselen, a multifunctional thioredoxin reductase inhibitor, inactivated E. coli GDH (IC₅₀ 213 nM). In cell-based assays at 0.5 mM tetrazolium concentration, EZMTT showed essentially no toxicity after a 3-day incubation, whereas 40% of inhibition was observed for WST-8. In conclusion, EZMTT is a novel tetrazolium salt which provides improved features that are suitable for dehydrogenases and real-time cell-based high-throughput screening (HTS).     

Foraging success under uncertainty: search tradeoffs and optimal space use

Understanding the structural complexity and the main drivers of animal search behaviour is pivotal to foraging ecology. Yet, the role of uncertainty as a generative mechanism of movement patterns is poorly understood. Novel insights from search theory suggest that organisms should collect and assess new information from the environment by producing complex exploratory strategies. Based on an extension of the first passage time theory, and using simple equations and simulations, we unveil the elementary heuristics behind search behaviour. In particular, we show that normal diffusion is not enough for determining optimal exploratory behaviour but anomalous diffusion is required. Searching organisms go through two critical sequential phases (approach and detection) and experience fundamental search tradeoffs that may limit their encounter rates. Using experimental data, we show that biological search includes elements not fully considered in contemporary physical search theory. In particular, the need to consider search movement as a non‐stationary process that brings the organism from one informational state to another. For example, the transition from remaining in an area to departing from it may occur through an exploratory state where cognitive search is challenged. Therefore, a more comprehensive view of foraging ecology requires including current perspectives about movement under uncertainty.     

Effect of polymorphisms in the CD36 and STAT3 genes on different dietary interventions among patients with coronary artery disease: study protocol for a randomized controlled trial

BackgroundCardiovascular disease has become a major health problem, and it has been associated with both environmental and genetic factors. Studies have shown that the Mediterranean Diet (MeDiet), or its components such as nuts and olive oil, may be strongly associated with the improvement of cardiovascular risk factors in specific populations. The purpose of the GENUTRI study is to investigate the interaction of genetics with cardiovascular risk factors in a non-Mediterranean population with coronary artery disease (CAD) according to three different diets: rich in pecan nuts, in extra-virgin olive oil or a control diet.Methods/designThe GENUTRI study is a single-center, randomized, open-label, parallel-group, 12-week pragmatic clinical trial conducted in patients aged 40 to 80 years and diagnosed with CAD. A standardized questionnaire will be applied to data collection and a blood sample will be obtained for lipid, glycemic and inflammatory profile evaluation. Polymorphisms in the CD36 and STAT3 genes will be detected using the TaqMan® SNP Genotyping Assay. Patients will be allocated in three groups: group 1: 30 g/day of pecan nuts; group 2: 30 ml/day of olive oil; and group 3: control diet. The primary outcome will consist of changes in LDL-cholesterol (in mg/dl) after 12 weeks of intervention.DiscussionStudies have shown the beneficial effects of diets rich in nuts and olive oil mainly in the Mediterranean population. GENUTRI is a clinical trial focusing on the effects of nuts or olive oil supplementation in Brazilian individuals. Additionally, we will try to demonstrate that genetic polymorphisms linked to cardiovascular disease may modulate the effects of different diets on biochemical and inflammatory markers among these subjects.

Trial registration

ClinicalTrials.gov Identifier: {"type":"clinical-trial","attrs":{"text":"NCT02202265","term_id":"NCT02202265"}}NCT02202265 (registered on 18 July 2014: first version).     

Diamondback moth performance and preference for leaves of Brassica oleracea of different ages and strata

The preference–performance hypothesis predicts that moth behaviour links plant variations with caterpillar attack and distribution, and the plant‐age hypothesis states that specialist herbivores are more successful in exploring younger plant tissue. We integrated these predictions to investigate underlying mechanisms by which moths and caterpillars of Plutella xylostella L. (Lepidoptera: Plutellidae) track and exploit within‐plant variability of leaf age and stratification. We measured leaf proteins, glucosinolates and fibre, as well as larval choice, developmental performance, and moth oviposition preference with regard to leaf age classes (young, mature and senescent) of three varieties (collard, cauliflower and cabbage) of the main host plant Brassica oleracea L. Larvae consistently fit the prediction that specialist herbivores prefer and perform better on young, upper leaves that have the highest protein level, despite the highest content of defence compounds. Conversely, moths laid more eggs on fibrous and less nutritious leaves from the lower and senescent stratum. We argue that the leaf stratification of host plants imposes conflicting selective pressures concerning offspring feeding and protection on adult females. If egg mortality is catastrophic on the upper nutritious leaves in a particular microclimatic context (e.g. sun, heat, winds, drought or rain‐washing), then oviposition preference will remain for the suboptimal lower and senescent leaves. The ability of larvae to spread upwards over the plant to access the more nutritious leaf stratum is critical when eggs are preferentially laid on the protective low‐quality leaves.     

Plasticity of mycangia in Xylosandrus ambrosia beetles

Insects that depend on microbial mutualists evolved a variety of organs to transport the microsymbionts while dispersing. The ontogeny and variability of such organs is rarely studied, and the microsymbiont*s effects on the animal tissue development remain unknown in most cases. Ambrosia beetles (Coleoptera: Curculionidae: Scolytinae or Platypodinae) and their mutualistic fungi are an ideal system to study the animalfungus interactions. While the interspecific diversity of their fungus transport organ一 mycangia—is well-known, their developmental plasticity has been poorly described. To determine the ontogeny of the mycangium and the influence of the symbiotic fungus on the tissue development, we dissected by hand or scanned with micro-CT the mycangia in various developmental stages in five Xylosandrus ambrosia beetle species that possess a large, mesonotal mycangium: Xylosandrus amputatus. Xylosandrus compactus, Xylosandrus crassiusculus, Xylosandrus discolor, and Xylosandrus germanus. We processed 181 beetle samples from the United States and China. All five species displayed three stages of the mycangium development:(1) young teneral adults had an empty, deflated and cryptic mycangium without fungal mass;(2) in fully mature adults during dispersal, the promesonotal membrane was inflated, and most individuals developed a mycangium mostly filled with the symbiont, though size and symmetry varied;and (3) after successful establishment of their new galleries, most females discharged the bulk of the fun gal inoculum and deflated the mycangium. Experimental aposymbiotic individuals demonstrated that the pronotal membrane invaginated independently of the presence of the fungus, but the fungus was required for inflation. Mycangia are more dynamic than previously thought, and their morphological changes correspond to the phases of the symbiosis. Importantly, studies of the fungal symbionts or plant pathogen transmission in ambrosia beetles need to consider which developmental stage to sample. We provide illustrations of the different stages, including microphotography of dissections and micro-CT scans.