Fluorescence of nitrobenzoxadiazole (NBD)-labeled lipids in model membranes is connected not to lipid mobility but to probe location

Nitrobenzoxadiazole (NBD)-labeled lipids are popular fluorescent membrane probes. However, the understanding of important aspects of the photophysics of NBD remains incomplete, including the observed shift in the emission spectrum of NBD-lipids to longer wavelengths following excitation at the red edge of the absorption spectrum (red-edge excitation shift or REES). REES of NBD-lipids in membrane environments has been previously interpreted as reflecting restricted mobility of solvent surrounding the fluorophore. However, this requires a large change in the dipole moment (Dm) of NBD upon excitation. Previous calculations of the value of Dm of NBD in the literature have been carried out using outdated semi-empirical methods, leading to conflicting values. Using up-to-date density functional theory methods, we recalculated the value of Dm and verified that it is rather small (B2 D). Fluorescence measurements confirmed that the value of REES is B16 nm for 1,2-dioleoyl-sn-glycero-3- phospho-L-serine-N-(NBD) (NBD-PS) in dioleoylphosphatidylcholine vesicles. However, the observed shift is independent of both the temperature and the presence of cholesterol and is therefore insensitive to the mobility and hydration of the membrane. Moreover, red-edge excitation leads to an increased contribution of the decay component with a shorter lifetime, whereas time-resolved emission spectra of NBD-PS displayed an atypical blue shift following excitation. This excludes restrictions to solvent relaxation as the cause of the measured REES and TRES of NBD, pointing instead to the heterogeneous transverse location of probes as the origin of these effects. The latter hypothesis was confirmed by molecular dynamics simulations, from which the calculated heterogeneity of the hydration and location of NBD correlated with the measured fluorescence lifetimes/REES. Globally, our combination of theoretical and experiment-based techniques has led to a considerably improved understanding of the photophysics of NBD and a reinterpretation of its REES in particular.

Assessing the conservation status of Mediterranean temporary ponds using biodiversity: a new tool for practitioners

The assessment of the habitat condition is the first step of conservation actions and several tools are available to assess wetlands. However, only a few tools are adapted to the priority habitat Mediterranean temporary ponds. Thus, our objectives were (i) to identify biological indicators associated with the different conservation status of Mediterranean tem- porary ponds and (ii) to create an efficient evaluation tool for non-experts using indicators of conservation status. A total of 87 ponds were sampled in southwest Portugal to assess the presence of plants, large branchiopods, amphibians, threatened voles and bats. Ponds with favourable conservation status showed higher species richness of plants, large branchiopods and amphibians. We identified eighteen indicators for favourable ponds: 15 plants, one large branchiopod and two amphibian taxa. We propose a new tool to assess the conservation status of Mediterranean tem- porary ponds based on the presence of these indicators. This tool is an alternative to other common, but time- consuming, methods and can be readily used by trained practitioners. The replication and adaptation of this tool to other regions and habitats enables the collection of comparable data and the geographical scaling-up of the assessments.

Assessing the conservation status of Mediterranean temporary ponds using biodiversity: a new tool for practitioners

The assessment of the habitat condition is the first step of conservation actions and several tools are available to assess wetlands. However, only a few tools are adapted to the priority habitat Mediterranean temporary ponds. Thus, our objectives were (i) to identify biological indicators associated with the different conservation status of Mediterranean temporary ponds and (ii) to create an efficient evaluation tool for non-experts using indicators of conservation status. A total of 87 ponds were sampled in southwest Portugal to assess the presence of plants, large branchiopods, amphibians, threatened voles and bats. Ponds with favourable conservation status showed higher species richness of plants, large branchiopods and amphibians. We identified eighteen indicators for favourable ponds: 15 plants, one large branchiopod and two amphibian taxa. We propose a new tool to assess the conservation status of Mediterranean temporary ponds based on the presence of these indicators. This tool is an alternative to other common, but time consuming, methods and can be readily used by trained practitioners. The replication and adaptation of this tool to other regions and habitats enables the collection of comparable data and the geographical scaling-up of the assessments.