Escherichia coli viability determination using dynamic light scattering: a comparison with standard methods

To determine the concentration of bacteria in a sample is important in the food industry, medicine and biotechnology. A disadvantage of the plate-counting method is that a microorganism colony could arise from one cell or from many cells. The other standard methodology, known as optical density determination, is based on the turbidity of a suspension and registers all bacteria, dead and alive. In this article, dynamic light scattering is proposed as a fast and reliable method to determine bacterial viability and, consequently, time evolution. Escherichia coli was selected because this microorganism is well known and easy to handle. A correlation between the data from these three techniques was obtained. We were able to calculate the growth rate, usually determined by plate counting or optical density measurement, using dynamic light scattering and to predict bacterial behavior. An analytical relationship between the colony forming units and the light scattered intensity was also deduced.     

“Plastic” and “static” behavior of vessel-anatomical features in Oriental beech (Fagus orientalis Lipsky) in view of xylem hydraulic conductivity

Key message

Xylem anatomical traits can be categorized into two groups: plastic properties which show a high inter-annual variability, and static characteristics which vary in a more conservative range.

Abstract

Water conduction in broad-leaved trees depends mainly on the size, number, and arrangement of vessels, which vary from year to year in response to varying exogenous factors, thus contributing to a safe and/or efficient water transport. However, the nature of such compensation is not clear; in particular, it is not obvious which traits act independently and which ones coincidentally. To better understand these inter-relations, tree-ring width (TRW), vessel-related anatomical traits, and the theoretical hydraulic conductivity were measured or modeled in the last 50 growth rings of mature Oriental beech trees growing at different altitudes in northern Iran. The study trees followed similar strategies compensating the effects of external factors by modifying their vessel-anatomical features. TRW and the number of vessels per unit of area were highly but negatively correlated and both were affected by exogenous factors. However, a decrease in vessel frequency (VF) is not a mirror effect of wider tree rings, but trees actively control the number of vessels produced. Principal component analysis revealed that the features VF, TRW and relative total conductivity were more plastic, whereas average vessel-lumen area, tree-ring porosity, and relative specific conductivity behaved more static. Moreover, we suggest that in theoretical approaches, total hydraulic conductivity rather than the specific hydraulic conductivity is a better indicator of a tree’s hydraulic behavior in a given growing season.     

Infrared Spectroscopy of Pollen Identifies Plant Species and Genus as Well as Environmental Conditions

Background

It is imperative to have reliable and timely methodologies for analysis and monitoring of seed plants in order to determine climate-related plant processes. Moreover, impact of environment on plant fitness is predominantly based on studies of female functions, while the contribution of male gametophytes is mostly ignored due to missing data on pollen quality. We explored the use of infrared spectroscopy of pollen for an inexpensive and rapid characterization of plants.

Methodology

The study was based on measurement of pollen samples by two Fourier transform infrared techniques: single reflectance attenuated total reflectance and transmission measurement of sample pellets. The experimental set, with a total of 813 samples, included five pollination seasons and 300 different plant species belonging to all principal spermatophyte clades (conifers, monocotyledons, eudicots, and magnoliids).

Results

The spectroscopic-based methodology enables detection of phylogenetic variations, including the separation of confamiliar and congeneric species. Furthermore, the methodology enables measurement of phenotypic plasticity by the detection of inter-annual variations within the populations. The spectral differences related to environment and taxonomy are interpreted biochemically, specifically variations of pollen lipids, proteins, carbohydrates, and sporopollenins. The study shows large variations of absolute content of nutrients for congenital species pollinating in the same environmental conditions. Moreover, clear correlation between carbohydrate-to-protein ratio and pollination strategy has been detected. Infrared spectral database with respect to biochemical variation among the range of species, climate and biogeography will significantly improve comprehension of plant-environment interactions, including impact of global climate change on plant communities.     

Experimental selection of long‐term intracellular mycobacteria

Some intracellular bacteria are known to cause long‐term infections that last decades without compromising the viability of the host. Although of critical importance, the adaptations that intracellular bacteria undergo during this long process of residence in a host cell environment remain obscure. Here, we report a novel experimental approach to study the adaptations of mycobacteria imposed by a long‐term intracellular lifestyle. Selected Mycobacterium bovis BCG through continuous culture in macrophages underwent an adaptation process leading to impaired phenolic glycolipids (PGL) synthesis, improved usage of glucose as a carbon source and accumulation of neutral lipids. These changes correlated with increased survival of mycobacteria in macrophages and mice during re‐infection and also with the specific expression of stress‐ and survival‐related genes. Our findings identify bacterial traits implicated in the establishment of long‐term cellular infections and represent a tool for understanding the physiological states and the environment that bacteria face living in fluctuating intracellular environments.     

Radial stem variations of Tabebuia chrysantha (Bignoniaceae) in different tropical forest ecosystems of southern Ecuador

Stem diameter increments of the broadleaved deciduous tree species Tabebuia chrysantha were measured with high-resolution dendrometers in a tropical lower montane forest and in a dry forest in southern Ecuador, the latter showing a distinct dry season. Those analyses were complemented by wood anatomical studies on regularly collected microcores to determine the season of active cambial growth and the time of formation of annual growth boundaries. The length of the cambial active period varied between 3 and 7 months at the tropical lower montane forest and 2 and 4 months in the dry forest, respectively. During dry days, amplitudes of daily stem diameter variations correlated with vapour pressure deficit. During October and November, inter-annual climate variations may lead to dry and sunny conditions in the tropical lower montane forest, causing water deficit and stem diameter shrinkage in T. chrysantha. The results of the climate–growth analysis show a positive relationship between tree growth and rainfall as well as vapour pressure deficit in certain periods of the year, indicating that rainfall plays a major role for tree growth.     

Effects of land-use change on functional and taxonomic diversity of Neotropical bats

Human land-use changes are particularly extensive in tropical regions, representing one of the greatest threats to terrestrial biodiversity and a key research topic in conservation. However, studies considering the effects of different types of anthropogenic disturbance on the functional dimension of biodiversity in human-modified landscapes are rare. Here, we obtained data through an extensive review of peer-reviewed articles and compared 30 Neotropical bat assemblages in well-preserved primary forest and four different human-disturbed habitats in terms of their functional and taxonomic diversity. We found that disturbed habitats that are structurally less similar to primary forest (pasture, cropland, and early-stage secondary forest) were characterized by a lower functional and taxonomic diversity, as well as community-level functional uniqueness. These habitats generally retained fewer species that perform different ecological functions compared to higher-quality landscape matrices, such as agroforestry. According to functional trait composition, different bat ensembles respond differently to landscape change, negatively affecting mainly gleaning insectivorous bats in pasture, narrow-range species in cropland, and heavier animalivorous bats in secondary forest. Although our results highlight the importance of higher-quality matrix habitats to support elevated functional and taxonomic bat diversity, the conservation of bat species that perform different ecological functions in the mosaic of human-modified habitats also depends on the irreplaceable conservation value of well-preserved primary forests. Our study based on a pooled analysis of individual studies provides novel insights into the effects of different human-modified habitats on Neotropical bat assemblages.     

Time-resolved fluorescence resonance energy transfer and surface plasmon resonance-based assays for retinoid and transthyretin binding to retinol-binding protein 4

Retinol-binding protein-4 (RBP4) is an emerging candidate drug target for type 2 diabetes and lipofuscin-mediated macular degeneration. The retinoic acid derivative fenretinide (N-(4-hydroxyphenyl) retinamide; HPR) exerts therapeutic effects in mouse models of obesity, diabetes, and Stargardt’s disease by targeting RBP4. Fenretinide competes with retinoids for RBP4 binding, disrupts RBP4-transthyretin (TTR) complexes, and results in urinary secretion of RBP4 and systemic depletion of retinol. To enable the search for nonretinoid molecules with fenretinide-like activities we developed a HTS-compatible homogeneous TR-FRET assay monitoring the displacement of retinoic acid derivatives from RBP4 in high-density 384-well and 1536-well microtiter plate formats. The retinoid displacement assay proved to be highly sensitive and robust after miniaturization with IC₅₀s for fenretinide and retinol ranging around 50 and 100 nM, respectively, and Z′-factors around 0.7. In addition, a surface plasmon resonance (SPR)-based secondary assay was developed to interrogate small molecule RBP4 binders for their ability to modulate the RBP4-TTR interaction. Finally, a 1.6 × 10⁶ compound library was screened against the retinoid displacement assay. Several potent retinoid competitors were identified that also appeared to disrupt RBP4-TTR complexes. Some of these compounds could potentially serve as valuable tools to further probe RBP4 biology in the future.     

Activity of the Enterococcus faecalis EIIA PTS component and its strong interaction with EIIB

Eubacteria can import and simultaneously phosphorylate a range of different carbohydrates by means of sugar specific phosphoenolpyruvate (PEP) dependent sugar phosphotransferase systems (PTSs). Here, we report the biochemical characterization of the gluconate specific PTS component EIIA^gnt from Enterococcus faecalis and its unexpectedly strong complex with EIIB^gnt. We analyze the activity of the complex regarding phosphoryl transfer using kinetic measurements and demonstrate by mutagenesis that His-9 of EIIA^gnt is essential for this process and represents most likely the phosphoryl group carrier of EIIA^gnt. With a combination of isothermal titration calorimetry (ITC), analytical ultracentrifugation (AUC), native gel electrophoresis and chemical crosslinking experiments we show that EIIA^gnt and EIIB^gnt form a strong 2:2 heterotetrameric complex, which seems to be destabilized upon phosphorylation of EIIB^gnt.