Extraction and Quantification of Lipids from Plant or Algae.

In plants and algae, the glycerolipidome changes in response to environmental modifications. For instance, in phosphate starvation, phospholipids are degraded and replaced by non-phosphorus lipids, and in nitrogen starvation, storage lipids accumulate. In addition to the well-known applications of oil crops for food, algae lipids are becoming a model for potential applications in health, biofuel, and green chemistry and are used as a platform for genetic engineering. It is therefore important to measure accurately and quickly the glycerolipid content in plants and algae. Here we describe the methods to extract the lipid and quantify the fatty acid amount of the lipid extract and the different lipid classes that are present in these samples.

Quantitative Assessment of the Chloroplast Lipidome.

In plants and algae, photosynthetic membranes have a unique lipid composition. They differ from all other cellular membranes by their very low amount of phospholipids, besides some phosphatidylglycerol (PG), and high proportion of glycolipids. These glycolipids are the uncharged galactolipids, that is, mono- and digalactosyldiacylglycerol (MGDG and DGDG), and an anionic sulfolipid, that is, sulfoquinovosyldiacylglycerol (SQDG). In all photosynthetic membranes analyzed to date, from cyanobacteria to algae, protists, and plants, the lipid quartet constituted by MGDG, DGDG, SQDG, and PG has been highly conserved, but the composition in fatty acids of these lipids can vary a lot from an organism to another. To better understand the chloroplast biogenesis, it is therefore essential to know their lipid content. Establishing chloroplast lipidome requires first to purify chloroplast from plant or algae tissue. Here we describe the methods to extract the lipid, quantify the lipid amount of the chloroplast, and qualify and quantify the different lipid classes that might be present in these fractions.

A Gradient-Descent Optimization of CO2-CO-NOx Emissions over the Paris Megacity─The Case of the First SARS-CoV-2 Lockdown.

Urban greenhouse gas emissions monitoring is essential to assessing the impact of climate mitigation actions. Using atmospheric continuous measurements of air quality and carbon dioxide (CO2), we developed a gradient-descent optimization system to estimate emissions of the city of Paris. We evaluated our joint CO2-CO-NOx optimization over the first SARS-CoV-2 related lockdown period, resulting in a decrease in emissions by 40% for NOx and 30% for CO2, in agreement with preliminary estimates using bottom-up activity data yet lower than the decrease estimates from Bayesian atmospheric inversions (50%). Before evaluating the model, we first provide an in-depth analysis of three emission data sets. A general agreement in the totals is observed over the region surrounding Paris (known as Île-de-France) since all the data sets are constrained by the reported national and regional totals. However, the data sets show disagreements in their sector distributions as well as in the interspecies ratios. The seasonality also shows disagreements among emission products related to nonindustrial stationary combustion (residential and tertiary combustion). The results presented in this paper show that a multispecies approach has the potential to provide sectoral information to monitor CO2 emissions over urban areas enabled by the deployment of collocated atmospheric greenhouse gases and air quality monitoring stations.

NO2, BC and PM Exposure of Participants in the Polluscope Autumn 2019 Campaign in the Paris Region.

The Polluscope project aims to better understand the personal exposure to air pollutants in the Paris region. This article is based on one campaign from the project, which was conducted in the autumn of 2019 and involved 63 participants equipped with portable sensors (i.e., NO2, BC and PM) for one week. After a phase of data curation, analyses were performed on the results from all participants, as well as on individual participants' data for case studies. A machine learning algorithm was used to allocate the data to different environments (e.g., transportation, indoor, home, office, and outdoor). The results of the campaign showed that the participants' exposure to air pollutants depended very much on their lifestyle and the sources of pollution that may be present in the vicinity. Individuals' use of transportation was found to be associated with higher levels of pollutants, even when the time spent on transport was relatively short. In contrast, homes and offices were environments with the lowest concentrations of pollutants. However, some activities performed in indoor air (e.g., cooking) also showed a high levels of pollution over a relatively short period.

New particle formation at a peri-urban agricultural site.

New Particle Formation (NPF) is a major source of ultrafine particles that affect both air quality and climate. Despite emissions from agricultural activities having a strong potential to lead to NPF, little is known about NPF within agricultural environments. The aim of the present study was to investigate the occurrence of NPF events at an agricultural site, and any potential relationship between agricultural emissions and NPF events. A field campaign was conducted for 3 months at the FR-Gri-ICOS site (France), at an experimental farm 25 km west of Paris city centre. 16 NPF events have been identified from the analysis of particle number size distributions; 8 during the daytime, and 8 during the night-time. High solar radiation and ozone mixing ratios were observed during the days NPF occurred, suggesting photochemistry plays a key role in daytime NPF. These events were also associated with higher levels of VOCs such as isoprene, methanol, or toluene compared to non-event days. However, ammonia levels were lower during daytime NPF events, contributing to the hypothesis that daytime NPF events were not related to agricultural activities. On the other hand, temperature and ozone were lower during the nights when NPF events were observed, whereas relative humidity was higher. During these nights, higher concentrations of NO2 and ammonia were observed. As a result, agricultural activities, in particular the spreading of fertiliser on surrounding crops, are suspected to contribute to night-time NPF events. Finally, all the identified NPF events were also observed at SIRTA monitoring station 20 km from the FR-Gri ICOS site, showing that both night-time and daytime NPF events were regional processes. We hypothesise that night-time NPF may be related to fertiliser spreading over a regional scale, as opposed to the local activities at the farm. To our knowledge, this is the first time night-time NPF has been observed in the agricultural context.

Personal Exposure to Black Carbon, Particulate Matter and Nitrogen Dioxide in the Paris Region Measured by Portable Sensors Worn by Volunteers.

Portable sensors have emerged as a promising solution for personal exposure (PE) measurement. For the first time in Île-de-France, PE to black carbon (BC), particulate matter (PM), and nitrogen dioxide (NO2) was quantified based on three field campaigns involving 37 volunteers from the general public wearing the sensors all day long for a week. This successful deployment demonstrated its ability to quantify PE on a large scale, in various environments (from dense urban to suburban, indoor and outdoor) and in all seasons. The impact of the visited environments was investigated. The proximity to road traffic (for BC and NO2), as well as cooking activities and tobacco smoke (for PM), made significant contributions to total exposure (up to 34%, 26%, and 44%, respectively), even though the time spent in these environments was short. Finally, even if ambient outdoor levels played a role in PE, the prominent impact of the different environments suggests that traditional ambient monitoring stations is not a proper surrogate for PE quantification.

Variability and sources of non-methane hydrocarbons at a Mediterranean urban atmosphere: The role of biomass burning and traffic emissions.

Levels and sources of non-Methane Hydrocarbons (NMHCs) were investigated at the urban background Thissio station, close to the historical center of Athens (Greece) from March 2016 to February 2017 (12 months), by means of an automated GC-FID. Alkanes dominated over aromatics and alkenes, with hourly mean levels ranging from detection limit up to 60 µg m-3 for i-pentane and 90 µg m-3 for toluene. Higher levels were recorded in the cold period relative to the warmer one. In addition, NMHCs seasonal diurnal cycles were characterized by a bimodal pattern, following the trend of tracers of anthropogenic sources. The Positive Matrix Factorization (PMF) was used for the allocation of NMHC to their sources. Five factors were identified and quantified, with traffic-related sources being the main one contributing up to 60% to total NMHCs, while biomass burning contributes up to 19%. A supplementary PMF assimilation was applied on a seasonal basis further including α-pinene, C6-C16 alkanes and aromatics. This PMF resulted to a seven-factor solution that allowed the examination of two additional sources, in addition to five already identified, highlighting the main contribution of anthropogenic sources (70%) to α-pinene.

Consequences of Mixotrophy on Cell Energetic Metabolism in Microchloropsis gaditana Revealed by Genetic Engineering and Metabolic Approaches.

Algae belonging to the Microchloropsis genus are promising organisms for biotech purposes, being able to accumulate large amounts of lipid reserves. These organisms adapt to different trophic conditions, thriving in strict photoautotrophic conditions, as well as in the concomitant presence of light plus reduced external carbon as energy sources (mixotrophy). In this work, we investigated the mixotrophic responses of Microchloropsis gaditana (formerly Nannochloropsis gaditana). Using the Biolog growth test, in which cells are loaded into multiwell plates coated with different organic compounds, we could not find a suitable substrate for Microchloropsis mixotrophy. By contrast, addition of the Lysogeny broth (LB) to the inorganic growth medium had a benefit on growth, enhancing respiratory activity at the expense of photosynthetic performances. To further dissect the role of respiration in Microchloropsis mixotrophy, we focused on the mitochondrial alternative oxidase (AOX), a protein involved in energy management in other algae prospering in mixotrophy. Knocking-out the AOX1 gene by transcription activator-like effector nuclease (TALE-N) led to the loss of capacity to implement growth upon addition of LB supporting the hypothesis that the effect of this medium was related to a provision of reduced carbon. We conclude that mixotrophic growth in Microchloropsis is dominated by respiratory rather than by photosynthetic energetic metabolism and discuss the possible reasons for this behavior in relationship with fatty acid breakdown via ß-oxidation in this oleaginous alga.

Atmospheric reactivity of biogenic volatile organic compounds in a maritime pine forest during the LANDEX episode 1 field campaign.

Trace gas measurements were performed during the LANDEX (the LANDes EXperiment) Episode 1 field campaign in the summer 2017, in one of the largest European maritime pine forests (> 95% Pinus pinaster) located in southwestern France. Efforts have been focused on obtaining a good speciation of 20 major biogenic volatile organic compounds (BVOCs, including pinenes, carenes, terpinenes, linalool, camphene, etc.). This was made possible by the development of a new and specific chromatographic method. In order to assess the role of BVOCs in the local gas phase chemistry budget, their reactivity with the main atmospheric oxidants (hydroxyl radicals (OH), ozone (O3) and nitrate radicals (NO3)) and the corresponding consumption rates were determined. When considering the OH reactivity with BVOCs, isoprene and linalool accounted for 10-47% of the OH depletion during daytime, and monoterpenes for 50-65%, whereas monoterpenes were the main contributors during the night (70-85%). Sesquiterpenes and monoterpenes were the main contributors to the ozone reactivity, especially ß-caryophyllene (30-70%), with a maximum contribution during nighttime. Nighttime nitrate reactivity was predominantly due to monoterpenes (i.e. 90-95%). Five specific groups have been proposed to classify the 19 BVOCs measured in the forest, according to their reactivity with atmospheric oxidants and their concentrations. The total amount of BVOCs consumed under and above the forest canopy was evaluated for 7 BVOCs (i.e. isoprene, α-pinene, ß-pinene, myrcene, limonene + cis-ocimene and Δ3-carene). The reactivity of atmospheric oxidants and BVOCs at a local level are discussed in order to highlight the compounds (BVOCs, other VOCs), the atmospheric oxidants and the main associated reactive processes observed under the canopy of a maritime pine forest.

Atmospheric Biodetection Part I: Study of Airborne Bacterial Concentrations from January 2018 to May 2020 at Saclay, France.

Background: The monitoring of bioaerosol concentrations in the air is a relevant endeavor due to potential health risks associated with exposure to such particles and in the understanding of their role in climate. In this context, the atmospheric concentrations of bacteria were measured from January 2018 to May 2020 at Saclay, France. The aim of the study was to understand the seasonality, the daily variability, and to identify the geographical origin of airborne bacteria. Methods: 880 samples were collected daily on polycarbonate filters, extracted with purified water, and analyzed using the cultivable method and flow cytometry. A source receptor model was used to identify the origin of bacteria. Results: A tri-modal seasonality was identified with the highest concentrations early in spring and over the summer season with the lowest during the winter season. Extreme changes occurred daily due to rapid changes in meteorological conditions and shifts from clean air masses to polluted ones. Conclusion: Our work points toward bacterial concentrations originating from specific seasonal-geographical ecosystems. During pollution events, bacteria appear to rise from dense urban areas or are transported long distances from their sources. This key finding should drive future actions to better control the dispersion of potential pathogens in the air, like persistent microorganisms originating from contaminated areas.

PUB11-Dependent Ubiquitination of the Phospholipid Flippase ALA10 Modifies ALA10 Localization and Affects the Pool of Linolenic Phosphatidylcholine.

Biogenesis of photosynthetic membranes depends on galactolipid synthesis, which relies on several cell compartments, notably the endoplasmic reticulum (ER) and the chloroplast envelope. Galactolipid synthesis involves lipid trafficking between both membrane compartments. In Arabidopsis, ALA10, a phospholipid flippase of the P4 type-ATPase family, counteracts the limitation of monogalactosyldiacylglycerol (MGDG) production and has a positive effect on leaf development. ALA10 locates in distinct domains of the ER depending on the ALIS (ALA interacting subunit) subunit it interacts with: close to the plasma membrane with ALIS1, or next to chloroplasts with ALIS5. It interacts with FAD2 (Fatty acid desaturase 2) and prevents accumulation of linolenic (18:3) containing phosphatidylcholine (PC) stimulating an increase of MGDG synthesis. Here we report that ALA10 interacts with PUB11 (plant U-box type 11), an E3 protein ubiquitin ligase, in vitro and in vivo. ALA10 is however ubiquitinated and degraded by the 26S proteasome in a PUB11-independent process. In pub11 null mutant, the proteasome-dependent degradation of ALA10 is retained and ALA10 is still subject to ubiquitination although its ubiquitination profile appears different. In the absence of PUB11, ALA10 is constrained to the ER close to chloroplasts, which is the usual location when ALA10 is overexpressed. Additionally, in this condition, the decrease of 18:3 containing PC is no longer observed. Taken together these results suggest, that ALA10 contributes in chloroplast-distal ER interacting domains, to reduce the 18:3 desaturation of PC and that PUB11 is involved in reconditioning of ALA10 from chloroplast-proximal to chloroplast-distal ER interacting domains.

Substantial brown carbon emissions from wintertime residential wood burning over France.

Brown carbon (BrC) is known to absorb light at subvisible wavelengths but its optical properties and sources are still poorly documented, leading to large uncertainties in climate studies. Here, we show its major wintertime contribution to total aerosol absorption at 370 nm (18-42%) at 9 different French sites. Moreover, an excellent correlation with levoglucosan (r2 = 0.9 and slope = 22.2 at 370 nm), suggesting important contribution of wood burning emissions to ambient BrC aerosols in France. At all sites, BrC peaks were mainly observed during late evening, linking to local intense residential wood burning during this time period. Furthermore, the geographic origin analysis also highlighted the high potential contribution of local and/or small-regional emissions to BrC. Focusing on the Paris region, twice higher BrC mass absorption efficiency value was obtained for less oxidized biomass burning organic aerosols (BBOA) compared to more oxidized BBOA (e.g., about 4.9 ± 0.2 vs. 2.0 ± 0.1 m2 g-1, respectively, at 370 nm). Finally, the BBOA direct radiative effect was found to be 40% higher when these two BBOA fractions are treated as light-absorbing species, compared to the non-absorbing BBOA scenario.

Unveiling membrane thermoregulation strategies in marine picocyanobacteria.

The wide latitudinal distribution of marine Synechococcus cyanobacteria partly relies on the differentiation of lineages adapted to distinct thermal environments. Membranes are highly thermosensitive cell components, and the ability to modulate their fluidity can be critical for the fitness of an ecotype in a particular thermal niche. We compared the thermophysiology of Synechococcus strains representative of major temperature ecotypes in the field. We measured growth, photosynthetic capacities and membrane lipidome variations. We carried out a metagenomic analysis of stations of the Tara Oceans expedition to describe the latitudinal distribution of the lipid desaturase genes in the oceans. All strains maintained efficient photosynthetic capacities over their different temperature growth ranges. Subpolar and cold temperate strains showed enhanced capacities for lipid monodesaturation at low temperature thanks to an additional, poorly regiospecific Δ9-desaturase. By contrast, tropical and warm temperate strains displayed moderate monodesaturation capacities but high proportions of double unsaturations in response to cold, thanks to regiospecific Δ12-desaturases. The desaturase genes displayed specific distributions directly related to latitudinal variations in ocean surface temperature. This study highlights the critical importance of membrane fluidity modulation by desaturases in the adaptive strategies of Synechococcus cyanobacteria during the colonization of novel thermal niches.

A methodology for the characterization of portable sensors for air quality measure with the goal of deployment in citizen science.

The field of small air quality sensors is of growing interest within the scientific community, especially because this new technology is liable to improve air pollutant monitoring as well as be used for personal exposure quantification. Amongst the myriad existing devices, the performances are highly variable; this is why the sensors must be rigorously assessed before deployment, according to the intended use. This study is included in the Polluscope project; its purpose is to quantify personal exposure to air pollutants by using portable sensors. This paper designs and applies a methodology for the evaluation of portable air quality sensors to eight devices measuring PM, BC, NO2 and O3. The dedicated testing protocol includes static ambient air measurements compared with reference instruments, controlled chamber and mobility tests, as well as reproducibility evaluation. Three sensors (AE51, Cairclip and Canarin) were retained to be used for the field campaigns. The reliability of their performances were robustly quantified by using several metrics. These three devices (for a total of 36 units) were deployed to be worn by volunteers for a week. The results show the ability of sensors to discriminate between different environments (i.e., cooking, commuting or in an office). This work demonstrates, first, the ability of the three selected sensors to deliver data reliable enough to enable personal exposure estimations, and second, the robustness of this testing methodology.

Characterization of particulate and gaseous pollutants from a French dairy and sheep farm.

Agricultural activities highly contribute to atmospheric pollution, but the diversity and the magnitude of their emissions are still subject to large uncertainties. A field measurement campaign was conducted to characterize gaseous and particulate emissions from an experimental farm in France containing a sheep pen and a dairy stable. During the campaign, more than four hundred volatile organic compounds (VOCs) were characterized using an original combination of online and off-line measurements. Carbon dioxide (CO2) and ammonia (NH3) were the most concentrated compounds inside the buildings, followed by methanol, acetic acid and acetaldehyde. A CO2 mass balance model was used to estimate NH3 and VOC emission rates. To our knowledge, this study constitutes the first evaluation of emission rates for most of the identified VOCs. The measurements show that the dairy stable emitted more VOCs than the sheep pen. Despite strong VOC and NH3 emissions, the chemical composition of particles indicates that gaseous farm emissions do not affect the loading of fine particles inside the farm and is mainly explained by the low residence time inside the buildings. The experimental dataset obtained in this work will help to improve emissions inventories for agricultural activities.

Wood burning: A major source of Volatile Organic Compounds during wintertime in the Paris region.

Wood burning is widely used for domestic heating and has been identified as a ubiquitous pollution source in urban areas, especially during cold months. The present study is based on a three and a half winter months field campaign in the Paris region measuring Volatile Organic Compounds (VOCs) by Proton Transfer Reaction Mass Spectrometry (PTR-MS) in addition to Black Carbon (BC). Several VOCs were identified as strongly wood burning-influenced (e.g., acetic acid, furfural), or traffic-influenced (e.g., toluene, C8-aromatics). Methylbutenone, benzenediol and butandione were identified for the first time as wood burning-related in ambient air. A Positive Matrix Factorization (PMF) analysis highlighted that wood burning is the most important source of VOCs during the winter season. (47%). Traffic was found to account for about 22% of the measured VOCs during the same period, whereas solvent use plus background accounted altogether for the remaining fraction. The comparison with the regional emission inventory showed good consistency for benzene and xylenes but revisions of the inventory should be considered for several VOCs such as acetic acid, C9-aromatics and methanol. Finally, complementary measurements acquired simultaneously at other sites in Île-de-France (the Paris region) enabled evaluation of spatial variabilities. The influence of traffic emissions on investigated pollutants displayed a clear negative gradient from roadside to suburban stations, whereas wood burning pollution was found to be fairly homogeneous over the region.

Measurement of Lipid Transport in Mitochondria by the MTL Complex.

Membrane biogenesis requires an extensive traffic of lipids between different cell compartments. Two main pathways, the vesicular and non-vesicular pathways, are involved in such a process. Whereas the mechanisms involved in vesicular trafficking are well understood, fewer is known about non-vesicular lipid trafficking, particularly in plants. This pathway involves the direct exchange of lipids at membrane contact sites (MCSs) between organelles. In plants, an extensive traffic of the chloroplast-synthesized digalactosyldiacylglycerol (DGDG) to mitochondria occurs during phosphate starvation. This lipid exchange occurs by non-vesicular trafficking pathways at MCSs between mitochondria and plastids. By a biochemical approach, a mitochondrial lipoprotein super-complex called MTL (Mitochondrial Transmembrane Lipoprotein complex) involved in mitochondria lipid trafficking has been identified in Arabidopsis thaliana. This protocol describes the method to isolate the MTL complex and to study the implication of a component of this complex (AtMic60) in mitochondria lipid trafficking.

New application of direct analysis in real time high-resolution mass spectrometry for the untargeted analysis of fresh and aged secondary organic aerosols generated from monoterpenes.

RATIONALE: Secondary organic aerosols (SOAs) represent a significant portion of total atmospheric aerosols. They are generated by the oxidation of volatile organic compounds (VOCs), and particularly biogenic VOCs (BVOCs). The analysis of such samples is usually performed by targeted methods that often require time-consuming preparation steps that can induce loss of compounds and/or sample contaminations. METHODS: Recently, untargeted methods using high-resolution mass spectrometry (HRMS) have been successfully employed for a broad characterization of chemicals in SOAs. Herein we propose a new application of the direct analysis in real time (DART) ionization method combined with HRMS to quickly detect several hundred chemicals in SOAs collected on a quartz filter without sample preparation or separation techniques. RESULTS: The reproducibility of measurements was good, with several hundred elemental compositions common to three different replicates. The relative standard deviations of the intensities of the chemical families ranged from 6% to 35%, with sufficient sensitivity to allow the unambiguous detection of 4 ng/mm2 of pinic acid. The presence of oligomers and specific tracers was highlighted by MSn (n ≤ 4) experiments, an achievement that is difficult to attain with other ultrahigh-resolution mass spectrometers. Contributions of this untargeted DART-HRMS method were illustrated by the analysis of fresh and aged SOAs from different gaseous precursors such as limonene, a ß-pinene/limonene mixture or scots pines emissions. CONCLUSIONS: The results show that it is possible to use DART-HRMS for the identification of tracers of specific aging reactions, or for the identification of aerosols from specific biogenic precursors.

Extraction and Quantification of Lipids from Plant or Algae.

In plants and algae, the glycerolipidome changes in response to environmental modifications. For instance, in phosphate starvation, phospholipids are degraded and replaced by nonphosphorus lipids and in nitrogen starvation, storage lipids accumulate. In addition to the well-known applications of oil crops for food, algae lipids are becoming a model for potential applications in health, biofuel, and green chemistry and are used as a platform for genetic engineering. It is therefore important to measure accurately and quickly the glycerolipid content in plants and algae. Here we describe the methods to extract the lipid, quantify the fatty acid amount of the lipid extract and to quantify the different lipid classes that are present in these samples.

Quantitative Assessment of the Chloroplast Lipidome.

In plants and algae, photosynthetic membranes have a unique lipid composition. They differ from all other cellular membranes by their very low amount of phospholipids, besides some phosphatidylglycerol (PG), and high proportion of glycolipids. These glycolipids are the uncharged galactolipids, i.e., monogalactosyldiacylglycerol and digalactosyldiacylglycerol (MGDG and DGDG), and an anionic sulfolipid, i.e., sulfoquinovosyldiacylglycerol (SQDG). In all photosynthetic membranes analyzed to date, from cyanobacteria to algae, protists, and plants, the lipid quartet constituted by MGDG, DGDG, SQDG, and PG has been highly conserved but the composition in fatty acids of these lipids can vary a lot from an organism to another. To better understand chloroplast biogenesis, it is therefore essential to know their lipid content. Establishing chloroplast lipidome requires first to purify chloroplast from plant or algae tissue. Here we describe the methods to extract lipids, quantify the lipids of the chloroplast, and qualify and quantify the different lipid classes that might be present in these fractions.