RESUMEN
While agricultural activities, such as the application of manure on arable land and animal grazing on pastures, provide economic and environmental benefits, they may also pose microbial risks to water sources. The aim of this paper was to study the microbial fate and transport in an agricultural catchment and recipient water source through further development of the hydrological model HYPE. Hydrological modelling was combined with hydrodynamic modelling to simulate the fate and transport of Salmonella spp., verotoxin-producing Escherichia coli O157:H7 (VTEC) and Cryptosporidium parvum in an agricultural catchment of a drinking water source, Lake Vombsjön, in Sweden. This approach was useful to study the influence of different processes on the pathogen fate and transport, and to interpret the relative changes in the simulated concentrations. Sensitivity analysis indicated that the largest uncertainties in the model were associated with the estimation of pathogen loads, parameterisation of the pathogen processes, and simulation of partitioning between surface runoff and infiltration. The proposed modelling approach is valuable for assessing the relative effect of different risk-reducing interventions.
Asunto(s)
Bacterias/clasificación , Simulación por Computador , Heces/microbiología , Modelos Teóricos , Calidad del Agua , Agricultura , Animales , Bovinos , Monitoreo del Ambiente , Humanos , Lagos , Suecia , Microbiología del Agua , Contaminantes del Agua , Contaminación del Agua/prevención & controlRESUMEN
Conceptual hydrological models can move towards process-oriented modelling when addressing broader issues than discharge modelling alone. For instance, water quality modelling generally requires understanding of both pathways and travel times which might not be easily identified because observations at the outlet aggregate all processes at the catchment scale. In this study we tested if adding a second kind of observation, specifically sediment data, can help distinguish overland flow from total discharge. We applied a multi-objective calibration on both discharge and suspended sediment concentration simulation performance to the World-Wide Hydrological Predictions for the Environment (HYPE) model for 111 catchments spread over the USA. Results show that in comparison to two calibrations made one after the other, the multi-objective calibration leads to a significant improvement on the simulation performance of suspended sediments without a significant impact on the performance of discharge. New modelling hypotheses for overland flow calculations are proposed and resulted in similar discharge performances as the original one but with fewer parameters, which reduces equifinality and can prevent unwarranted model complexity in data-poor areas.
RESUMEN
The activity of natural killer (NK) cells is regulated by a fine-tuned balance between activating and inhibitory receptors. Dual-color fluorescence cross-correlation spectroscopy (FCCS) was used to directly demonstrate a so-called cis-interaction between a member of the inhibitory NK cell receptor family Ly49 (Ly49A), and its ligand, the major histocompatibility complex (MHC) class I, within the plasma membrane of the same cell. By a refined FCCS model, calibrated by positive and negative control experiments on cells from the same lymphoid cell line, concentrations and diffusion coefficients of free and interacting proteins could be determined on a collection of cells. Using the intrinsic intercellular variation of their expression levels for titration, it was found that the fraction of Ly49A receptors bound in cis increase with increasing amounts of MHC class I ligand. This increase shows a tendency to be more abrupt than for a diffusion limited - three dimensional bimolecular reaction, which most likely reflects the two-dimensional confinement of the reaction. For the Ly49A- MHC class I interaction it indicates that within a critical concentration range the local concentration level of MHC class I can provide a distinct regulation mechanism of the NK cell activity.
Asunto(s)
Membrana Celular/metabolismo , Antígenos HLA/metabolismo , Subfamilia A de Receptores Similares a Lectina de Células NK/metabolismo , Espectrometría de Fluorescencia/métodos , Alelos , Animales , Línea Celular Tumoral , Supervivencia Celular , Difusión , Proteínas Fluorescentes Verdes/metabolismo , Antígenos HLA/genética , Ratones , Unión ProteicaRESUMEN
We present a fluorescence recovery after photobleaching-based method for monitoring the progression of septal Z-ring contraction in dividing Escherichia coli cells. In a large number of cells undergoing division, we irreversibly bleached cytosolically expressed Enhanced Green Fluorescent Protein on one side of the septal invagination and followed the fluorescence relaxation on both sides of the septum. Since the relaxation time depends on the cross-sectional area of the septum, it can be used to determine the septal radius r. Assuming that the fraction of the observed cells with r-values in a given interval reflects the duration of that interval in the division process we could derive an approximate time-course for the contraction event, as a population average. By applying the method repeatedly on individual cells, the contraction process was also followed in real time. On a population average level, our data are best described by a linear contraction process in time. However, on the single cell level the contraction processes display a complex behaviour, with varying levels of activity. The proposed approach provides a simple yet versatile method for studying Z-ring contraction in vivo, and will help to elucidate its underlying mechanisms.
Asunto(s)
Proteínas Bacterianas/metabolismo , División Celular , Proteínas del Citoesqueleto/metabolismo , Escherichia coli/fisiología , Sustancias Macromoleculares/metabolismo , Multimerización de Proteína , Técnicas Bacteriológicas/métodos , Escherichia coli/metabolismo , Fluorescencia , Genes Reporteros , Proteínas Fluorescentes Verdes/metabolismo , Fotoblanqueo , Factores de TiempoRESUMEN
An approach to study bimolecular interactions in model lipid bilayers and biological membranes is introduced, exploiting the influence of membrane-associated electron spin resonance labels on the triplet state kinetics of membrane-bound fluorophores. Singlet-triplet state transitions within the dye Lissamine Rhodamine B (LRB) were studied, when free in aqueous solutions, with LRB bound to a lipid in a liposome, and in the presence of different local concentrations of the electron spin resonance label TEMPO. By monitoring the triplet state kinetics via variations in the fluorescence signal, in this study using fluorescence correlation spectroscopy, a strong fluorescence signal can be combined with the ability to monitor low-frequency molecular interactions, at timescales much longer than the fluorescence lifetimes. Both in solution and in membranes, the measured relative changes in the singlet-triplet transitions rates were found to well reflect the expected collisional frequencies between the LRB and TEMPO molecules. These collisional rates could also be monitored at local TEMPO concentrations where practically no quenching of the excited state of the fluorophores can be detected. The proposed strategy is broadly applicable, in terms of possible read-out means, types of molecular interactions that can be followed, and in what environments these interactions can be measured.
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Colorantes Fluorescentes/química , Membrana Dobles de Lípidos/química , Rodaminas/química , Espectrometría de Fluorescencia/métodos , Óxidos N-Cíclicos , Difusión , Cinética , Liposomas/química , SolucionesRESUMEN
G protein-coupled receptors (GPCRs) constitute the largest family of cell surface receptors. They can exist and act as dimers, but the requirement of dimers for agonist-induced signal initiation and structural dynamics remains largely unknown. Frizzled 6 (FZD6) is a member of Class F GPCRs, which bind WNT proteins to initiate signaling. Here, we show that FZD6 dimerizes and that the dimer interface of FZD6 is formed by the transmembrane α-helices four and five. Most importantly, we present the agonist-induced dissociation/re-association of a GPCR dimer through the use of live cell imaging techniques. Further analysis of a dimerization-impaired FZD6 mutant indicates that dimer dissociation is an integral part of FZD6 signaling to extracellular signal-regulated kinases1/2. The discovery of agonist-dependent dynamics of dimers as an intrinsic process of receptor activation extends our understanding of Class F and other dimerizing GPCRs, offering novel targets for dimer-interfering small molecules.Frizzled 6 (FZD6) is a G protein-coupled receptor (GPCR) involved in several cellular processes. Here, the authors use live cell imaging and spectroscopy to show that FZD6 forms dimers, whose association is regulated by WNT proteins and that dimer dissociation is crucial for FZD6 signaling.
Asunto(s)
Receptores Frizzled/metabolismo , Dimerización , Células HEK293 , Humanos , Proteína Wnt-5a/metabolismoRESUMEN
In cold climates, fungicides are used on golf greens to prevent snow mould causing serious damage to the turf. However, fungicide residues have been detected in runoff from golf courses, which may lead to restrictions on use. There is therefore an urgent need to improve understanding of the processes affecting leaching of fungicides from turfgrass systems to allow identification of green construction and management practices that minimize environmental impacts. In this study we monitored the leaching of the fungicide iprodione in a putting green. Sorption and degradation of iprodione was measured in batch and incubation experiments, and the simulation model MACRO was used as a risk assessment tool. Degradation of iprodione was bi-phasic, with a rapid initial phase (half-life 17 h) caused by enhanced biodegradation. Degradation rates slowed considerably after 5 days, with half-lives of up to 38 days. Sorption of iprodione was linear, with a K(oc) value of ca 400 cm(3) g(-1). MACRO reasonably accurately matched measured drainflows and concentrations of iprodione in soil and drainflow. However, peak concentrations in drainage were underestimated, which was attributed to preferential finger flow due to water repellency. The results also showed the importance of the organic matter content in the green root zone in reducing leaching. It was concluded that, with 'reasonable worst-case' use, losses of iprodione from greens can occur at concentrations exceeding water quality limits for aquatic ecosystems. Snow mould problems should be tackled by adopting green root zone mixes that minimize leaching and 'best management practices' that would avoid the need for intensive prophylactic use of fungicides.