Tracing amino acid exchange during host-pathogen interaction by combined stable-isotope time-resolved Raman spectral imaging

This study investigates the temporal and spatial interchange of the aromatic amino acid phenylalanine (Phe) between human retinal pigment epithelial cell line (ARPE-19) and tachyzoites of the apicomplexan protozoan parasite Toxoplasma gondii (T. gondii). Stable isotope labelling by amino acids in cell culture (SILAC) is combined with Raman micro-spectroscopy to selectively monitor the incorporation of deuterium-labelled Phe into proteins in individual live tachyzoites. Our results show a very rapid uptake of L-Phe(D8) by the intracellular growing parasite. T. gondii tachyzoites are capable of extracting L-Phe(D8) from host cells as soon as it invades the cell. L-Phe(D8) from the host cell completely replaces the L-Phe within T. gondii tachyzoites 7–9 hours after infection. A quantitative model based on Raman spectra allowed an estimation of the exchange rate of Phe as 0.5–1.6 × 104 molecules/s. On the other hand, extracellular tachyzoites were not able to consume L-Phe(D8) after 24 hours of infection. These findings further our understanding of the amino acid trafficking between host cells and this strictly intracellular parasite. In particular, this study highlights new aspects of the metabolism of amino acid Phe operative during the interaction between T. gondii and its host cell

CHEMICAL CARTOGRAPHY with APOGEE: METALLICITY DISTRIBUTION FUNCTIONS and the CHEMICAL STRUCTURE of the MILKY WAY DISK

Using a sample of 69,919 red giants from the SDSS-III/APOGEE Data Release 12, we measure the distribution of stars in the [/Fe] versus [Fe/H] plane and the metallicity distribution functions (MDFs) across an unprecedented volume of the Milky Way disk, with radius 3 < R < 15 kpc and height kpc. Stars in the inner disk (R < 5 kpc) lie along a single track in [/Fe] versus [Fe/H], starting with -enhanced, metal-poor stars and ending at [/Fe] ∼ 0 and [Fe/H] ∼ +0.4. At larger radii we find two distinct sequences in [/Fe] versus [Fe/H] space, with a roughly solar- sequence that spans a decade in metallicity and a high- sequence that merges with the low- sequence at super-solar [Fe/H]. The location of the high- sequence is nearly constant across the disk

Galactic archaeology with asteroseismology and spectroscopy: Red giants observed by CoRoT and APOGEE

With the advent of the space missions CoRoT and Kepler, it has recently become feasible to determine precise asteroseismic masses and relative ages for large samples of red giant stars. We present the CoRoGEE dataset, obtained from CoRoT light curves for 606 red giants in two fields of the Galactic disc that have been co-observed by the Apache Point Observatory Galactic Evolution Experiment (APOGEE). We used the Bayesian parameter estimation code PARAM to calculate distances, extinctions, masses, and ages for these stars in a homogeneous analysis, resulting in relative statistical uncertainties of ≲2% in distance, ~4% in radius, ~9% in mass and ~25% in age. We also assessed systematic age uncertainties stemming from different input physics and mass loss. We discuss the correlation between ages and chemical abundance patterns of field stars over a broad radial range of the Milky Way disc (5 kpc <RGal< 14 kpc), focussing on the [α/Fe]-[Fe/H]-age plane in five radial bins of the Galactic disc. We find an overall agreement with the expectations of pure chemical-evolution models computed before the present data were available, especially for the outer regions. However, our data also indicate that a significant fraction of stars now observed near and beyond the solar neighbourhood migrated from inner regions. Mock CoRoGEE observations of a chemodynamical Milky Way disc model indicate that the number of high-metallicity stars in the outer disc is too high to be accounted for even by the strong radial mixing present in the model. The mock observations also show that the age distribution of the [α/Fe]-enhanced sequence in the CoRoGEE inner-disc field is much broader than expected from a combination of radial mixing and observational errors. We suggest that a thick-disc/bulge component that formed stars for more than 3 Gyr may account for these discrepancies. Our results are subject to future improvements due to (a) the still low statistics, because our sample had to be sliced into bins of Galactocentric distances and ages; (b) large uncertainties in proper motions (and therefore guiding radii); and (c) corrections to the asteroseismic mass-scaling relation. The situation will improve not only upon the upcoming Gaia data releases, but also with the foreseen increase in the number of stars with both seismic and spectroscopic information

Origin of the Metallicity Distribution in the Thick Disc

Aims. Using a suite of cosmological chemodynamical disc galaxy simulations, we assess how (a) radial metallicity gradients evolve with scaleheight; (b) the vertical metallicity gradients change through the thick disc; and (c) the vertical gradient of the stellar rotation velocity varies through the disc. We compare with the Milky Way to search for analogous trends. Methods. We analyse five simulated spiral galaxies with masses comparable to the Milky Way. The simulations span a range of star formation and energy feedback strengths and prescriptions, particle- and grid-based hydrodynamical implementations, as well as initial conditions/assembly history. Disc stars are identified initially via kinematic decomposition, with a posteriori spatial cuts providing the final sample from which radial and vertical gradients are inferred. Results. Consistently, we find that the steeper, negative, radial metallicity gradients seen in the mid-plane flatten with increasing height away from the plane. In simulations with stronger (and/or more spatially-extended) feedback, the negative radial gradients invert, becoming positive for heights in excess of !1 kpc. Such behaviour is consistent with that inferred from recent observations. Our measurements of the vertical metallicity gradients show no clear correlation with galactocentric radius, and are in good agreement with those observed in the Milky Way’s thick disc (locally). Each of the simulations presents a decline in rotational velocity with increasing height from the mid-plane, albeit the majority have shallower kinematic gradients than that of the Milky Way. Conclusions. Simulations employing stronger/more extended feedback prescriptions possess radial and vertical metallicity and kinematic gradients more in line with recent observations. The inverted, positive, radial metallicity gradients seen in the simulated thick stellar discs originate from a population of younger, more metal-rich, stars formed in-situ, superimposed upon a background population of older migrators from the inner disc; the contrast provided by the former increases radially, due to the inside-out growth of the disc. A similar behaviour may be responsible for the same flattening seen in the radial gradients with scaleheight in the Milky Way

The Gaia-ESO Survey::the present-day radial metallicity distribution of the Galactic disc probed by pre-main-sequence clusters

Context. The radial metallicity distribution in the Galactic thin disc represents a crucial constraint for modelling disc formation and evolution. Open star clusters allow us to derive both the radial metallicity distribution and its evolution over time. Aims. In this paper we perform the first investigation of the present-day radial metallicity distribution based on [Fe/H] determinations in late type members of pre-main-sequence clusters. Because of their youth, these clusters are therefore essential for tracing the current interstellar medium metallicity. Methods. We used the products of the Gaia-ESO Survey analysis of 12 young regions (age < 100 Myr), covering Galactocentric distances from 6.67 to 8.70 kpc. For the first time, we derived the metal content of star forming regions farther than 500 pc from the Sun. Median metallicities were determined through samples of reliable cluster members. For ten clusters the membership analysis is discussed in the present paper, while for other two clusters (i.e. Chamaeleon I and Gamma Velorum) we adopted the members identified in our previous works. Results. All the pre-main-sequence clusters considered in this paper have close-to-solar or slightly sub-solar metallicities. The radial metallicity distribution traced by these clusters is almost flat, with the innermost star forming regions having [Fe/H] values that are 0.10−0.15 dex lower than the majority of the older clusters located at similar Galactocentric radii. Conclusions. This homogeneous study of the present-day radial metallicity distribution in the Galactic thin disc favours models that predict a flattening of the radial gradient over time. On the other hand, the decrease of the average [Fe/H] at young ages is not easily explained by the models. Our results reveal a complex interplay of several processes (e.g. star formation activity, initial mass function, supernova yields, gas flows) that controlled the recent evolution of the Milky Way

Exploring the potential of stable isotope (resonance) raman microspectroscopy and SERS for the analysis of microorganisms at single cell level.

Raman microspectroscopy is a prime tool to characterize the molecular and isotopic composition of microbial cells. However, low sensitivity and long acquisition times limit a broad applicability of the method in environmental analysis. In this study, we explore the potential, the applicability and the limitations of stable isotope Raman microspectroscopy (SIRM), resonance SIRM, and SIRM in combination with surface-enhanced Raman scattering (SERS) for the characterization of single bacterial cells. The latter two techniques have the potential to significantly increase sensitivity and decrease measurement times in SIRM, but to date there are no (SERS-SIRM) or only a limited number (resonance SIRM) of studies in environmental microbiology. The analyzed microorganisms were grown with substrates fully labeled with the stable isotopes 13C or 2H and compounds with natural abundance of atomic isotopes (12C 98.89% or 1H 99.9844%, designated as 12C- or 1H-, respectively). Raman bands of bacterial cell compounds in stable isotope-labeled microorganisms exhibited a characteristic red-shift in the spectra. In particular the sharp phenylalanine band was found to be an applicable marker band for SIRM analysis of the Deltaproteobacterium strain N47 growing anaerobically on 13C-naphthalene. The study of G. metallireducens grown with 13C- and 2H-acetate showed that the information on the chromophore cytochrome c obtained by resonance SIRM at 532 nm excitation wavelength can be successfully complemented by whole-organism fingerprints of bacteria cells achieved by regular SIRM after photobleaching. Furthermore, we present here for the first time the reproducible SERS analysis of microbial cells labeled with stable isotopes. Escherichia coli strain DSM 1116 cultivated with 12C- or 13C-glucose was used as a model organism. Silver nanoparticles synthesized in situ were applied as SERS media. We observed a reproducible red-shift of an adenine-related marker band from 733 cm 1 to 720 cm 1 in SERS spectra for 13C-labbeled cells. Additionally, Raman measurements of 12C/13C-glucose and -phenylalanine mixtures were performed to elucidate the feasibility of SIRM for nondestructive quantitative and spatially-resolved analysis. The performed analysis of isotopically labeled microbial cells with SERS- and resonance SIRM paves the way towards novel approaches to apply Raman microspectroscopy in environmental process studies

Exploring the Potential of Stable Isotope (Resonance) Raman Microspectroscopy and Surface-Enhanced Raman Scattering for the Analysis of Microorganisms at Single Cell Level

Raman microspectroscopy is a prime tool to characterize the molecular and isotopic composition of microbial cells. However, low sensitivity and long acquisition times limit a broad applicability of the method in environmental analysis. In this study, we explore the potential, the applicability, and the limitations of stable isotope Raman microspectroscopy (SIRM), resonance SIRM, and SIRM in combination with surface-enhanced Raman scattering (SERS) for the characterization of single bacterial cells. The latter two techniques have the potential to significantly increase sensitivity and decrease measurement times in SIRM, but to date, there are no (SERS-SIRM) or only a limited number (resonance SIRM) of studies in environmental microbiology. The analyzed microorganisms were grown with substrates fully labeled with the stable isotopes ¹³C or ²H and compounds with natural abundance of atomic isotopes (¹²C 98.89% or ¹H 99.9844%, designated as ¹²C or ¹H, respectively). Raman bands of bacterial cell compounds in stable isotope-labeled microorganisms exhibited a characteristic red-shift in the spectra. In particular, the sharp phenylalanine band was found to be an applicable marker band for SIRM analysis of the <i>Deltaproteobacterium</i> strain N47 growing anaerobically on ¹³C-naphthalene. The study of <i>G. metallireducens</i> grown with ¹³C- and ²H-acetate showed that the information on the chromophore cytochrome <i>c</i> obtained by resonance SIRM at 532 nm excitation wavelength can be successfully complemented by whole-organism fingerprints of bacteria cells achieved by regular SIRM after photobleaching. Furthermore, we present here for the first time the reproducible SERS analysis of microbial cells labeled with stable isotopes. <i>Escherichia coli</i> strain DSM 1116 cultivated with ¹²C- or ¹³C-glucose was used as a model organism. Silver nanoparticles synthesized <i>in situ</i> were applied as SERS media. We observed a reproducible red-shift of an adenine-related marker band from 733 to 720 cm^–1 in SERS spectra for ¹³C-labeled cells. Additionally, Raman measurements of ¹²C/¹³C-glucose and -phenylalanine mixtures were performed to elucidate the feasibility of SIRM for nondestructive quantitative and spatially resolved analysis. The performed analysis of isotopically labeled microbial cells with SERS-SIRM and resonance SIRM paves the way toward novel approaches to apply Raman microspectroscopy in environmental process studies