Vincent Guigoz

Stéphanie Bruyère

Sylvie Migot

Raphaël Schneider
email: raphael.schneider@univ-lorraine.fr

Thomas Gries
email: thomas.gries@univ-lorraine.fr

Enhanced photoelectrocatalytic hydrogen evolution using off-stoichiometry La 0.43 FeO y films

Keywords: LaxFeOy, off-stoichiometry, photoelectrochemical H2 production

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

As a respond to the global energy demand, hydrogen (H2) appears to be a promising energy carrier to replace fossil fuels, if produced renewably [START_REF] Andreas | Hydrogen: the future energy carrier[END_REF][START_REF] Mazloomi | Hydrogen as an energy carrier: Prospects and challenges[END_REF]. Photoelectrochemical (PEC) water splitting is one of the most promising process to generate H2 but the PEC efficiency is limited by the semiconductor photocatalyst performances [START_REF] Xu | A Perovskite Electrocatalyst for Efficient Hydrogen Evolution Reaction[END_REF][START_REF] Grätzel | Photoelectrochemical cells[END_REF][5][START_REF] Alfaifi | Photoelectrochemical solar water splitting: From basic principles to advanced devices[END_REF][START_REF] Piccinin | The band structure and optical absorption of hematite (α-Fe2O3): a first-principles GW-BSE study[END_REF]. Recently, perovskite materials with the general formula ABO3 (A and B being cations) have attracted great attention as photocatalysts [START_REF] Wang | Perovskite Oxide Based Electrodes for High-Performance Photoelectrochemical Water Splitting[END_REF][9][START_REF] Ismael | Perovskite-type LaFeO3: Photoelectrochemical Properties and Photocatalytic Degradation of Organic Pollutants Under Visible Light Irradiation[END_REF][START_REF] Tao | Strong Hollow Spherical La2 NiO4 Photocatalytic Microreactor for Round-the-Clock Environmental Remediation[END_REF]. Due to the abundance of its components, its low toxicity and its optical absorption in the visible light range, lanthanum ferrite perovskite (LaFeO3) was demonstrated to be a high value candidate for H2 photoproduction [START_REF] Ismael | Perovskite-type LaFeO3: Photoelectrochemical Properties and Photocatalytic Degradation of Organic Pollutants Under Visible Light Irradiation[END_REF][START_REF] Díez-García | Metal Doping to Enhance the Photoelectrochemical Behavior of LaFeO3 Photocathodes[END_REF][START_REF] Guigoz | Heterostructured thin LaFeO3/g-C3N4 films for efficient photoelectrochemical hydrogen evolution[END_REF][START_REF] Celorrio | Photoelectrochemical Properties of LaFeO3 Nanoparticles[END_REF][START_REF] Kanhere | Electronic Structure, Optical Properties, and Photocatalytic Activities of LaFeO3 -NaTaO3 Solid Solution[END_REF]. However, its performances are limited by a high charge recombination rate due to the modest electron transport [START_REF] Ismael | Perovskite-type LaFeO3: Photoelectrochemical Properties and Photocatalytic Degradation of Organic Pollutants Under Visible Light Irradiation[END_REF][START_REF] Wheeler | Photoelectrochemical Properties and Stability of Nanoporous p-Type LaFeO3 Photoelectrodes Prepared by Electrodeposition[END_REF][START_REF] Peng | Surface modification of LaFeO3 by Co-Pi electrochemical deposition as an efficient photoanode under visible light[END_REF][START_REF] Li | Photoinduced charge property of nanosized perovskite-type LaFeO3 and its relationships with photocatalytic activity under visible irradiation[END_REF][START_REF] Hu | Photocatalytic activities of LaFeO3 and La2FeTiO6 in p-chlorophenol degradation under visible light[END_REF][START_REF] Thirumalairajan | Shape evolution of perovskite LaFeO3 nanostructures: a systematic investigation of growth mechanism, properties and morphology dependent photocatalytic activities[END_REF]. In recent years, various methods have been developed to improve the light absorption and the charge transport in LaFeO3 photocatalysts such as the creation of heterojunctions, nanostructuration or doping [5,[START_REF] Díez-García | Metal Doping to Enhance the Photoelectrochemical Behavior of LaFeO3 Photocathodes[END_REF][START_REF] Low | Heterojunction Photocatalysts[END_REF][START_REF] Moniz | Visible-light driven heterojunction photocatalysts for water splitting -a critical review[END_REF][START_REF] Ge | A review of TiO2 nanostructured catalysts for sustainable H2 generation[END_REF][START_REF] Kumaravel | Photocatalytic hydrogen production using metal doped TiO2: A review of recent advances[END_REF][START_REF] Zhang | Noble-metal-free Ni3C as co-catalyst on LaNiO3 with enhanced photocatalytic activity[END_REF].

Another option is the development of non-stoichiometric materials responding to the general formula La1-xFeOy. Indeed, various studies have demonstrated the interest of modifying the stoichiometry (La/Fe ratio) of LaFeO3 for the catalytic conversion of gases such as NO or CO and of hydrocarbons [START_REF] Faye | Influence of lanthanum stoichiometry in La1-xFeO3-δ perovskites on their structure and catalytic performance in CH4 total oxidation[END_REF][START_REF] Schön | Non stoichiometric La1-yFeO3 perovskite-based catalysts as alternative to commercial three-way-catalysts? -Impact of Cu and Rh doping[END_REF][START_REF] Delmastro | Synthesis and characterization of non-stoichiometric LaFeO3 perovskite[END_REF]. The catalytic performances for the oxidation of CO, propene and CH4 increase using La1-xFeOy having an overstoichiometry in Fe [START_REF] Faye | Influence of lanthanum stoichiometry in La1-xFeO3-δ perovskites on their structure and catalytic performance in CH4 total oxidation[END_REF][START_REF]Noble Materials for Photoelectrochemical Water Splitting[END_REF]. This increase is linked to the smaller particle size and thus to the higher specific surface area, as particle growth is slowed down by the excess in Fe. X-ray photoelectron spectroscopy (XPS) measurements of La1-xFeOy particles also revealed the formation of a Fe2O3 phase on the surface of the particles [START_REF] Bhatt | Advancement of Platinum (Pt)-Free (Non-Pt Precious Metals) and/or Metal-Free (Non-Precious Metals) Electrocatalysts in Energy Applications: A Review and Perspectives[END_REF]. Fe2O3 was demonstrated to generate crystal defects that promote the adsorption of oxygen on the surface. The highest catalytic activity was observed for La0.9FeOy in the oxidation of CH4 and for Cu-doped La0.67Fe0.83Cu0.17O3 in the oxidation of CO and propene.

However, a significant excess in Fe can no longer be accommodated on the surface of La1-xFeOy and can generate surface defective layers containing stacking faults or displacive Fe leading to a decrease of catalytic activity [START_REF] Delmastro | Synthesis and characterization of non-stoichiometric LaFeO3 perovskite[END_REF]. Therefore, a fine tuning of the non-stoichiometric composition is of high importance to control the catalytic properties.

Herein, thin La1-xFeOy films were engineered to demonstrate the increase in photocatalytic activity compared to LaFeO3. The non-stoichiometric films were produced via the magnetron co-sputtering process and the deviation from the ideal ABO3 stoichiometry was finely tuned by adjusting the powers applied to the La and Fe targets. First, the influence of the composition on the photocatalytic properties is demonstrated. Next, the improvement of the photocatalytic properties by optimization of the oxidation treatment and of the layer thickness is discussed. Finally, H2 photoproduction by PEC water splitting under simulated solar illumination is presented to further support the enhanced photocatalytic activity of these La1-xFeOy nonstoichiometric layers.

Experimental Section

Elaboration of La1-xFeOy thin films

La1-xFe films were deposited on fluorine doped tin oxide coated glass (FTO, TEC 7 Ω.sq -1 ) and silicon substrates by direct current (DC) magnetron co-sputtering of two metal targets: Fe target (diameter: 50 mm, thickness: 0.25 mm, purity: 99.5%) and La target (diameter: 50 mm, thickness: 3 mm, purity: 99.9%). The targets were powered by two Advanced Energy MDX 500 DC generators. A base pressure of 5 × 10 -4 Pa in the sputtering chamber was provided by a mechanical and a turbo-molecular pump.

The experimental device was equipped with two independent magnetrons focused on the centre of the substrate-holder with an inclination of 30° with respect to the substrate normal, and the target to substrate distance was set equal to 10 cm. Thin films were synthesized at a working pressure of 0.58 Pa in a 10 vol.% H2 -90 vol.% Ar mixture.

The substrate-holder rotated at 28 rpm to ensure film homogeneity. Thin films were deposited without external heating and the self-established deposition temperature was below 50°C. At this temperature, films remained amorphous and a post-annealing in a tubular furnace at 500°C for 2 h in air was conducted to obtain the perovskite structure.

Characterization

The La/Fe atomic ratio was estimated by energy dispersive spectrometry (EDS) coupled to a scanning electron microscope (SEM) FEI Quanta 650 FEG. Due to the uncertainty on the oxygen chemical contents measured by EDS, the La and Fe atomic concentrations were determined on metallic films, i.e. before annealing post-treatment.

Film thicknesses were measured by SEM cross-section observation. Transmission electron microscopy (TEM) investigations were performed using a cold FEG JEOL ARM200 microscope operating at 200 kV and equipped with spherical aberration (Cs) probe and image correctors (point resolution 0.12 nm in TEM mode). High-angle annular dark field image in scanning TEM (HAADF-STEM) and 2D elemental mapping using energy dispersive X-ray spectroscopy (EDX) are combined to obtain information about elemental distribution along film thickness. For these analyses, cross-section TEM samples of films deposited on silicon substrates were prepared using a focus ion beam (FIB) -SEM dual beam system FEI Helios NanoLab 600i. X-ray diffraction (XRD) was performed at room temperature using Bragg-Brentano θ/2θ measurements on a D8 Advance Bruker diffractometer (Cu Kα, λ = 1.54056 Å). The average crystallite size can be estimated from the full-width at half-maximum of diffraction peaks using the Scherrer's formula after correction from the experimental broadening (Eq. 1):

𝑑 = . ( 1
)
where dhkl is the average grain size along the hkl reticular distance, λ is the X-ray wavelength, θ is the diffraction angle and βhkl is the full-width at half-maximum of the diffraction peak. UV-visible absorption spectra were recorded on Cary 7000 UMS Agilent UV-visible spectrophotometer. Photoelectrochemical measurements of asprepared samples were performed by a SP 150 BioLogic potentiostat connected to a standard three electrode cell: Pt wire as the counter electrode, a saturated Ag/AgCl electrode as reference electrode and FTO glass coated with La1-xFeOy (1 cm × 1 cm) as the working electrode. A 0.1 M Na2SO4 (pH = 7) aqueous solution was used as electrolyte. All measurements were carried out under light illumination of 1 sun (100 mW•cm -2 ) provided from a Xenon lamp (Newport Xe 300 W ozone free) fitted with an air mass filter (AM 1.5G). Linear sweep voltammetry (LSV) measurements were obtained by scanning the potential from -0.8 V to 0.1 V vs Ag/AgCl at a scan rate of 5 mV.s -1 . The photocurrent density was evaluated from chronoamperometric measurements recorded at a constant potential of -0.30 V vs the reference electrode by cycling between dark and light illuminations of 30 s each for 10 min. Electrochemical impedance spectra (EIS) measurements were performed in the same workstation by applying a sinusoidal AC perturbation of 10 mV with a frequency range from 500 kHz to 0.01 Hz.

Hydrogen photoproduction

The photocatalytic hydrogen production experiments were performed in a sealed custom-made metallic PEC cell with an attached quartz viewport using the same experimental conditions as described for PEC measurements. The system was purged with Ar for 1 h before measurements. The amounts of produced H2 and O2 were determined during the water splitting reaction using an online quadrupole mass spectrometer (MS, Inficon Transpector MPS 100) operating with Ar as carrier gas.

Measurements were conducted at an interval of 1 h and averaged over 2 min.

Results

La1-xFeOy films synthesis and characterization

The La/Fe atomic ratio in the as-deposited films can be tuned by changing the discharge power applied to the Fe and La targets. The experimental parameters and the film compositions are summarized in Table 1. Seven samples with overstoichiometry in Fe were synthesized by progressively decreasing the power applied to the La target, the power applied to Fe target being fixed at 80 W: La0.11FeOy, La0.25FeOy, La0.33FeOy, La0.43FeOy, La0.54FeOy, La0.66FeOy and La0.82FeOy. Two samples were also elaborated with La over-stoichiometry: La1.26FeOy and La4FeOy. These samples will be compared to Fe2O3 and LaFeO3 films. The depositions durations were adjusted according to the sputtering rate of each target to keep the thickness constant after annealing, at about 500 nm. Fig. 1 shows the XRD patterns of the as-synthesized thin films with different compositions. The dotted peaks correspond to the FTO-coated glass substrate.

Depending on the film stoichiometry, the XRD patterns exhibit different crystal phases.

Samples can be divided into 4 groups.

-Group 1 corresponds to the films with a high Fe excess: Fe2O3 and La0.11FeOy.

Only peaks corresponding to Fe2O3 (rhombohedral hematite R-3c, PDF file 00-033-0664) could be observed.

-Group 2 corresponds to the films with an intermediate stoichiometry: La0.24FeOy, La0.33FeOy, La0.43FeOy and La0.54FeOy. Despite the oxidation treatment at 500°C for 2 h in air, the films were found to be amorphous as observed in the XRD patterns with a broad peak between 20 and 40°.

-Group 3 corresponds to films prepared with La/Fe atomic ratios close to 1:

La0.66FeOy and La1.26FeOy. The corresponding XRD patterns can be indexed to the orthorhombic LaFeO3 perovskite phase (Pnma, PDF file 04-011-7994).

-Group 4 corresponds to the deposit with a significant La over-stoichiometry:

La4FeOy. The XRD pattern can be assigned to the cubic structure of lanthanum oxide La2O3 (la-3, PDF file 03-065-3185). for an indirect allowed transition (Fig. S1b). A bandgap of ca. 2.28 eV was determined for LaFeO3 thin films deposited in the same experimental conditions, from a direct forbidden transition [START_REF] Guigoz | Heterostructured thin LaFeO3/g-C3N4 films for efficient photoelectrochemical hydrogen evolution[END_REF]. These values are in good agreement with those reported in the literature [START_REF] Piccinin | The band structure and optical absorption of hematite (α-Fe2O3): a first-principles GW-BSE study[END_REF][START_REF] Scafetta | Band structure and optical transitions in LaFeO3: theory and experiment[END_REF]. The UV-visible absorption spectra of the three non-stoichiometric films (groups 2 and 3) have an intermediate behaviour relative to Fe2O3 and LaFeO3 films. Furthermore, the nature of the transition does not seem to significantly influence the bandgap values of these intermediate non-stoichiometric films. For instance, for La0.43FeOy, an optical bandgap energy of 2.25 eV can be estimated from both indirect allowed and direct forbidden transitions (Fig. S1a andS1b). The absorption of the La4FeOy film markedly decreases at wavelengths above 350 nm, limiting its photocatalytic activity mostly to UV irradiation. This result can be correlated with the wide bandgap energy of La2O3 estimated to be 5.2 eV [START_REF] Pandey | Nonreducible, Basic La2O3 to Reducible, Acidic La2-xSbxO3 with Significant Oxygen Storage Capacity, Lower Band Gap, and Effect on the Catalytic Activity[END_REF][START_REF] Gedam | Electrical, dielectric and optical properties of La2O3 doped lithium borate glasses[END_REF][START_REF] Yadav | A simple chemical route for synthesis of microrods-like La2O3 thin films[END_REF][START_REF] Nohira | Atomic-scale depth profiling of composition, chemical structure and electronic band structure of La2O3/Si(100) interfacial transition layer[END_REF][START_REF] Liu | Band and Band Gap Characterization of La2O3 Films on Si Substrates Grown by Radio Frequency Magnetron Sputtering[END_REF].

Fig. Linear sweep voltammetry (J-V) curves of La1-xFeOy films were recorded under simulated sunlight illumination and a chopped beam was used to acquire both dark current and photocurrent in a single scan (Fig. 3a). The steady state photocurrent onset potential was around 0.6 V vs reversible hydrogen electrode (RHE) and the dark current remained stable until about 0.3 V vs RHE. All samples show an increase in negative current under illumination for potentials close to 0 V vs RHE, which corresponds to the behaviour of photocathodes. To determine the composition giving the highest PEC performance, chronoamperometric measurements under chopped illumination were conducted at a fixed potential of 0.3 V vs RHE (Fig. 3b). This potential value was chosen to obtain a dark current close to 0 µA. Fig. 3c shows the photocurrent density in absolute value as a function of the La stoichiometry of the films. These As previously indicated, XRD patterns showed that the group 2 samples (with a slight excess in Fe) remain amorphous after an oxidative treatment under air at 500°C for 2 h (Fig. 1). The XRD pattern of a film with a 500 nm thickness oxidized at 650°C for 2 h in air is shown in Fig. 4a. The presence of peaks corresponding to LaFeO3 can be observed, indicating the improved crystallinity after the thermal treatment at 650°C.

The average grain size of the La0.43FeOy film was estimated by the Scherrer's formula (Eq. 1). A representative length of coherence domains of LaFeO3 crystallites of ca. 26 ± 2 nm was extracted using the peak corresponding to the (121) plane. A value of 39 ± 2 nm was obtained for a stoichiometric LaFeO3 film after a 2 h oxidative treatment at 500°C [START_REF] Guigoz | Heterostructured thin LaFeO3/g-C3N4 films for efficient photoelectrochemical hydrogen evolution[END_REF]. Such a decrease was also observed for La1-xFeOy nanoparticles synthesized via thermal decomposition of Fe and La nitrates and was attributed to the organization of excess Fe into FeO6 octahedra at the surface which limit the grain growth [START_REF] Delmastro | Synthesis and characterization of non-stoichiometric LaFeO3 perovskite[END_REF]. Measurements of the photocurrent density under constant applied potential (0.3V vs RHE) shows a significant improvement for La0.43FeOy films oxidized 650°C show an increase of 60% from -6.4 µA•cm -1 (500°C) to -10.2 µA•cm -1 (650°C) (Fig. 4b). This result can be attributed to a better electron transfer due to the improved crystallinity of the film. To gain additional information on the phase distribution in these non-stoichiometric films, cross-sectional TEM analyses were performed on a La0.43FeOy film (thickness of ca. 550 nm) deposited on a silicon substrate and oxidized at 650°C for 2 h. The bright field TEM image (Fig. 5a) and the corresponding EDS-STEM maps (Fig. S2) highlight an evolution of the composition and of the structure depending on the depth. The coating can be divided in 4 different zones (Fig. 5b). At the interface with the substrate, a thin layer (35-40 nm) with an excess of Fe (25 at.%) is observed (Zone 1).

EDS analyses show that the main part of the film (ca. 450 nm) contains Fe and La but with a higher concentration in La: 22 at.% Fe -12 at.% La (Zone 2). However, an evolution with the depth was noticed: close to the substrate, the composition is 13 at.% in La and 22 at.% in Fe, while on the other side, the composition is 11 at.% in La and 22 at.% in Fe. This evolution may originate from a diffusion of Fe atoms towards the interfaces during the oxidation treatment, which is consistent with the composition in zone 1 and zone 4. Noteworthy is that metal films synthesized by magnetron sputtering initially show a homogeneous composition in thickness (data not shown).

Close to the upper surface, a thin layer of about 60 nm containing a slightly higher atomic fraction in La (13% La and 21% Fe) was detected (Zone 3). Finally, the EDS-STEM maps (Fig. SI2) show the presence of a dense layer at the extreme surface (zone 4), composed only of Fe (30 at.%) and O (70 at.%). The high-resolution image and the corresponding Fast Fourier Transform (FFT) pattern recorded in zone 1 are characteristic of an amorphous phase (Fig. 6a and6b). The bright field micrograph in zone 2 shows differences in contrast, the brighter areas likely originate from less dense or even porous regions (Fig. 6c). The corresponding FFT pattern shows a diffuse circle related to an amorphous phase as well as spots characteristic of a polycrystalline material. The interplanar distances can be indexed to LaFeO3 planes (orthorhombic Pnma) and to Fe2O3 planes (rhombohedral hematite R-3c) (Fig. 6d). As observed for zone 2, the spots observed in the FFT pattern recorded for zone 3 (red square) can also be indexed to LaFeO3 and Fe2O3. However, the presence of a light diffuse circle suggests that an amorphous phase may also be present (Fig. 6e and6f). In zone 4, at the upper surface, the spots observed on the FFT pattern correspond only to Fe2O3 (Fig. 6g and6h). The organization in layers according to the depth of this non-stoichiometric film, summarized in Table 2, gives an average composition of 34 at.% La and 66 at.% Fe for the film, which is consistent with the composition estimated from EDS-SEM measurements. An increase of the Fe amount was also observed on the surface of non-stoichiometric LaFeO3 particles synthesized with a Fe excess [START_REF] Faye | Influence of lanthanum stoichiometry in La1-xFeO3-δ perovskites on their structure and catalytic performance in CH4 total oxidation[END_REF][START_REF] Schön | Non stoichiometric La1-yFeO3 perovskite-based catalysts as alternative to commercial three-way-catalysts? -Impact of Cu and Rh doping[END_REF]. This phenomenon seems to be similar to that described by Neagu et al. on non-stoichiometric perovskite particles of ABO3 formula in which the deficiency in A leads to the formation of metallic or oxidized nanoparticles of B on the surface [START_REF] Neagu | In situ growth of nanoparticles through control of non-stoichiometry[END_REF]. The presence of A deficiencies in the perovskite lattice decreases the stability of the crystal lattice and contributes to the spontaneous exsolution of B atoms to restore the stoichiometry in the lattice. The B atoms diffuse to the surface of the non-stoichiometric particles to form a shell [START_REF] Kwon | Exsolution trends and co-segregation aspects of self-grown catalyst nanoparticles in perovskites[END_REF]. This surface segregation phenomenon was also observed for La0.6Sr0.4CoO3-δ films prepared by pulsed laser deposition and in Ti-doped LaFeO3 particles [START_REF] Kubicek | Relationship between Cation Segregation and the Electrochemical Oxygen Reduction Kinetics of La0.6Sr0.4CoO3-δ Thin Film Electrodes[END_REF][START_REF] Chen | Segregated Chemistry and Structure on (001) and (100) Surfaces of (La1-xSrx)2CoO4 Override the Crystal Anisotropy in Oxygen Exchange Kinetics[END_REF][START_REF] Garcia-Muñoz | Synergy effect between photocatalysis and heterogeneous photo-Fenton catalysis on Ti-doped LaFeO3 perovskite for high efficiency light-assisted water treatment[END_REF]. This complex process of segregation leads to the formation of Fe2O3 layer at the surface of LaFeO3 and allows significant improvements in the catalytic properties, as demonstrated for perovskite electrodes used in fuel cells [START_REF] Chen | Nonstoichiometric Oxides as Low-Cost and Highly-Efficient Oxygen Reduction/Evolution Catalysts for Low-Temperature Electrochemical Devices[END_REF][START_REF] Li | Controlling cation segregation in perovskite-based electrodes for high electro-catalytic activity and durability[END_REF]. An increase of the temperature required to obtain the crystalline phase with modification of the perovskite structure was also observed by Porta et al. when doping LaFeO3 with Mg [START_REF] Porta | AFeO3 (A=La, Nd, Sm) and LaFe1-xMgxO3 perovskites: structural and redox properties[END_REF].

Influence of La0.43FeOy film thickness on the photoelectronic properties

To further optimize the photocatalytic activity, the influence of the thickness of the La0.43FeOy film was investigated. Indeed, the thickness of the photoelectrodes is a key parameter in PEC measurements as it affects light absorption and thus the formation of electron/hole pairs, but also the charge carrier recombination probability [START_REF] Pawar | Unbiased Spontaneous Solar Fuel Production using Stable LaFeO3 Photoelectrode[END_REF]. The magnetron sputtering duration was adjusted to obtain film thicknesses of 50, 100, 250 and 500 nm after oxidation at 650°C for 2 h, all other experimental parameters being kept constant. Photocurrent measurements were carried out on FTO electrodes coated La0.43FeOy films (Fig. 7a). The 50, 250 and 500 nm thick-films showed almost similar results. The slight decrease in photocurrent generation for the 250 nm film may originate from changes of the crystal size or from a modification of the composition gradient, affecting photocatalytic properties. For a 100 nm thick La0.43FeOy film, the photocurrent significantly increases by 57% compared to the 500 nm thick film synthesized in the same experimental conditions (-16 µA•cm -1 vs -10.2 µA•cm ). Thus, a thickness of 100 nm appears to be optimal for this film composition for a further increase of the photoelectronic properties. The corresponding EIS Nyquist plots show a significant decrease in semi-circle diameter for the 100 nm thick layer compared to the 500 nm one, which indicates a decrease of the charge transfer resistance at the electrode/electrolyte interface (Fig. 7b).

Photoelectrocatalytic H2 production

To confirm the improvement of the photocatalytic properties, measurements of H2 production in a PEC cell were conducted. The production of H2 from the photoelectrodes covered with a 100 nm thin layer of La0.43FeOy or LaFeO3 after oxidation was measured in a Na2SO4 solution (0.1 M) under irradiation of 1 sun (Xe lamp associated with an air mass filter AM 1.5G -100 mW.cm -2 ) (Fig. 8a). LaFeO3 coated photoelectrodes showed an average H2 production rate of 5.5 µmol•h -1 •cm -2 once the stationary state was reached, i.e. between 2 and 7 h. The average production rate for La0.43FeOy, between 2 and 7 h, reached 11.2 µmol-cm -2 , which corresponds to an increase in H2 production rate of 103%. These results further confirm the enhanced photo(electro)catalytic activity by simply tuning the stoichiometry of the photocatalytic film. The performance of the La0.43FeOy film is significantly improved compared to other photocathodes prepared with LaFeO3 films, including a photocell composed of a LaFeO3 photocathode and a Fe2O3 photoanode where H2 production is close to 20 μmol•h -1 using a Xe lamp without AM 1.5G filter [START_REF] Pawar | Unbiased Spontaneous Solar Fuel Production using Stable LaFeO3 Photoelectrode[END_REF][START_REF] Yu | A highly durable p-LaFeO3/n-Fe2O3 photocell for effective water splitting under visible light[END_REF][START_REF] Li | Dye-sensitized LaFeO3 photocathode for solar-driven H2 generation[END_REF].

The simultaneous production of O2 at the counter-electrode was also measured (Fig. 8b). An O2 production rate of about 5.1 µmol•h -1 •cm -2 was observed, corresponding to ca. the half of H2 production as expected for the water splitting reaction. This quasistoichiometric production of H2 and O2 by PEC with a photocathode composed of a La0.43FeOy film was confirmed by the pH stability of the electrolyte. Only a slight decrease was observed during the 7 h of irradiation (from 7.45 to 6.80), indicating that the concentration of H + and OH -ions present in the solution remained almost constant.

The stability of La0.43FeOy film was also confirmed by chronoamperometric measurements (at 0.3 V vs RHE) over 10 h under a continuous simulated sunlight illumination and the photocurrent density was found to remain almost constant (Fig. S3). The non-stoichiometric composition leading to exsolution and segregation mechanisms observed during the thermal oxidation process has no influence on the stability of photoelectrodes, which is a key parameter for their practical application.

PEC mechanism

The Fe excess in La1-xFeOy films was shown to produce Fe2O3 crystallites within the film by exsolution. Moreover, a Fe2O3 thin layer grows at the upper surface during thermal oxidation as observed by TEM. Therefore, the formation of a heterojunction between LaFeO3 and Fe2O3 can be assumed.

The edge potentials of the valence band (VB) and conduction band (CB) of Fe2O3 were estimated using the semi-empirical calculation from Mulliken's electronegativity at the point of zero-charge. Using the experimentally determined optical bandgap energy (Fig. S1b), ECB and EVB values of 0.3 and 2.48 V vs RHE were determined, respectively. The band edge potentials of LaFeO3 were experimentally determined by EIS and from the bandgap estimated from UV-visible absorption measurements [START_REF] Guigoz | Heterostructured thin LaFeO3/g-C3N4 films for efficient photoelectrochemical hydrogen evolution[END_REF].

The positions of the VB edges of Fe2O3 and LaFeO3 were used to estimate the band energy positions before contact (Fig. 9a). Within the film and at the upper surface, the heterojunction formed between the two semiconductors will favour a shift of band energies and at the junction, the Fermi levels of both semiconductors are equal and constant under equilibrium conditions. As a consequence, an electric field can be generated at the interface between Fe2O3 and LaFeO3 leading to a better separation of the electron/hole pairs and thus to a reduction of their recombination probability. to the VB of LaFeO3 (Fig. 9b). The electrons photogenerated in Fe2O3 combined to those transferred from the CB of LaFeO3 likely reach an excited level in the CB of Fe2O3 with a potential sufficiently negative to allow the reduction of water into hydrogen (Fig. 9a). Similar results were previously described for heterojunctions composed of NiO or CuOx underneath a thin WO3 layer [START_REF] Wu | Flake-like NiO/WO3 p-n heterojunction photocathode for photoelectrochemical water splitting[END_REF][START_REF] Xie | A promising CuOx/WO3 p-n heterojunction thin-film photocathode fabricated by magnetron reactive sputtering[END_REF]. Furthermore, the presence of Fe2O3 crystals within the film, as demonstrated by TEM measurements, could also improve the bulk charge transfer, reducing their recombination probability before reaching the surface.

Conclusion

The influence of the stoichiometry in La1-xFeOy films on their photocatalytic activity for H2 production was investigated. The modification of the power applied to the metallic targets during magnetron co-sputtering process allows a fine control of the film composition. The highest photoactivity was observed for a 100 nm-thick La0.43FeOy film oxidized at 650°C for 2 h.

Fig. 1 .

 1 Fig. 1. Evolution of XRD patterns as a function of the sample composition after oxidation at 500°C for 2 h.

Fig. 2

 2 Fig.2shows the UV-visible absorption of the reference films (LaFeO3 and Fe2O3), of the films with intermediate compositions (La0.33FeOy and La0.43FeOy), of the film with near stoichiometric composition (La0.82FeOy) and of the film mainly composed of La4FeOy. The absorption of Fe2O3 shows a transition towards 550 nm. The optical bandgap energy was determined using Tauc plot and a value of 2.18 eV was obtained

  values have been determined at t = 525 s, i.e. in the middle of a light exposure period, when the spike corresponding to the formation of a double layer has passed, and after around 9 min of measurement, confirming the high stability of non-stoichiometric films.The photocurrent measured for group 1 films (high Fe excess) and for group 3 films (compositions close to LaFeO3) is almost similar with values close to -1.8 µA•cm -2 . The group 2 samples (intermediate compositions) exhibit much higher photocurrents, with a maximum value of -6.40 µA•cm -2 reached for the La0.43FeOy film. The photocurrent density is ca. 3.5 higher for the non-stoichiometric La0.43FeOy film compared to the stoichiometric LaFeO3 film synthesized under the same experimental conditions. As the photogenerated current density shows a markedly enhanced value for the La0.43FeOy film, a specific study will be focused on this film to further optimise its PEC performances by adjusting thermal annealing step and film thickness (vide infra).

Fig. 3 .

 3 Fig. 3. (a) Current-potential curves and (b) current-time curves at a constant potential of 0.3 V vs RHE for the different La1-xFeOy thin films under chopped illumination. (c) Photocurrent densities (absolute value) as a function of x' in Lax'FeOy films (0.1 M Na2SO4, pH 7, 1 sun).

Fig. 4 .

 4 Fig. 4. (a) XRD pattern of the La0.43FeOy sample after the ex situ oxidative treatment at 650°C for 2 h and (b) photocurrent density measurements of La0,43FeOy films oxidized at 500 and 650°C (0.3 V vs RHE, 0,1M Na2SO4 pH 7, 1 sun).

Fig. 5 .

 5 Fig. 5. TEM cross sectional micrograph of a La0.43FeOy film deposited on a Si substrate, after oxidation at 650°C for 2 h and (b) HAADF-STEM image showing the different zones and their composition.

Fig. 6 .

 6 Fig. 6. High resolution TEM micrographs recorded in the 4 zones and their corresponding FFT patterns: (a) and (b) = zone 1, (c) and (d) = zone 2, (e) and (f) = zone 3, (g) and (h) = zone 4.

Fig. 7 .

 7 Fig. 7. (a) Current-time curves of La0.43FeOy coated photoelectrodes at 0.3 V vs RHE under chopped illumination as a function of the film thickness and (b) EIS Nyquist plots of 500 and 100 nm thick La0.43FeOy films (0.1M Na2SO4, pH 7, 1 sun).

Fig. 8 .

 8 Fig. 8. (a) H2 production rates of 100 nm LaFeO3 and La0.43FeOy oxidized films coated photoelectrodes, (b) H2 and O2 production rate evolutions of La0.43FeOy photoelectrode (0.3 V vs RHE, 0.1 M Na2SO4, pH 7, 1 sun).

Fig. 9 .

 9 Fig. 9. (a) Diagram of band energy positions of Fe2O3 and LaFeO3 before contact and (b) schematic representation of the separation and transfer pathway of photogenerated charge carriers for PEC H2 production using La0.43FeOy films under solar illumination.

  TEM analyses show the presence of LaFeO3 within the film, underneath a thin(15 nm) and continuous polycrystalline Fe2O3 layer at the upper surface. The presence of Fe2O3 originates from an exsolution mechanism typically observed for nonstoichiometric perovskites. The improvement of the photocatalytic properties by optimizing the stoichiometry, the oxidation temperature and the film thickness was confirmed by a large increase of H2 production rates by comparison with a stoichiometric LaFeO3 film prepared under the same experimental conditions: from 5.5 to 11.2 µmol•cm -2 , i.e. 103%. This increase in catalytic performances associated to the high stability, visible light photoactivation and facile, low cost and scalable synthesis process make La0.43FeOy films of high interest as photocathode for PEC water splitting systems.CRediT authorship contribution statementV. Guigoz: Investigation, Writing -original draft. S. Bruyère: Investigation, Writingreview & editing. S. Migot: Investigation, Writing -review & editing. R. Schneider: Supervision, Funding acquisition, Writing -review & editing. T. Gries: Supervision, Funding acquisition, Writing -review & editing.

Table 1 :

 1 Deposition conditions, thickness and composition of La1-xFeOy thin films synthesized by magnetron co-sputtering of La and Fe targets and annealed at 500°C for 2 h in air.

	Power Fe	Power La	Deposition time	Sample composition
	(W)	(W)	(min)
	80	0	19.5	Fe2O3
	80	15	18	La0.11FeOy
	80	30	16	La0.24FeOy
	80	40	15	La0.33FeOy
	80	45	14	La0.43FeOy
	80	50	13	La0.54FeOy
	80	55	12	La0.66FeOy
	80	60	11	La0.82FeOy
	80	65	10	LaFeO3
	60	65	9	La1.26FeOy
	20	65	5	La4FeOy

Table 2 .

 2 Composition, thickness, and structure of the different layers present in the La0.43FeOy film.

	Zone	Composition (at.%)	Thickness	Structure
	1	O 66%, Fe 25%, La 9%	35-40 nm	Amorphous
	2	O 66%, Fe 22%, La 12%	450 nm	Polycrystalline (LaFeO3 and
				Fe2O3) + amorphous phase
	3	O 66%, Fe 21%, La 13%	60 nm	Polycrystalline (LaFeO3 and
				Fe2O3) + amorphous phase
	4	O 70%, Fe30%, La 0%	15 nm	Fe2O3

Acknowledgements

This work was supported partly by the French PIA project « Lorraine Université d'Excellence », reference ANR-15-IDEX-04-LUE.

Appendix A. Supplementary data

Supplementary data to this article can be found at http://doi.org/

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.


Gaspard Kerner

Etienne Patin

Lluis Quintana-Murci

Quintana-Murci Lluis

New insights into human immunity from ancient genomics

Population genetic studies have clearly indicated that immunity and host defense are among the functions most frequently subject to natural selection, and increased our understanding of the biological relevance of the corresponding genes and their contribution to variable immune traits and diseases. Herein, we will focus on some recently studied forms of human adaptation to infectious agents, including hybridization with now-extinct hominins, such as Neanderthals and Denisovans, and admixture between modern human populations. These studies, which are partly enabled by the technological advances in the sequencing of DNA from ancient remains, provide new insight into the sources of immune response variation in contemporary humans, such as the recently reported link between Neanderthal heritage and susceptibility to severe COVID-19 disease. Furthermore, ancient DNA analyses, in both humans and pathogens, allow to measure the action of natural selection on immune genes across time and to reconstruct the impact of past epidemics on the evolution of human immunity.

Introduction

The relationship between humans and microbes is double-edged; they can complement each other to maintain homeostasis, like in the case of microbiota, or risk fatal conflict, like when microbes cause infectious disease. Microbes have accompanied humans since their origins in Africa ~200,000-300,000 years ago and through their subsequent dispersals around the world over the last ~60,000 years [START_REF] Karlsson | Natural selection and infectious disease in human populations[END_REF]. During their spread, humans have also encountered a highly diverse set of new climatic, nutritional, and pathogenic conditions, to which they had to adapt [START_REF] Fan | Going global by adapting local: A review of recent human adaptation[END_REF]. Given the strong mortality burden imposed by pathogens, it is not surprising that human genes involved in immune functions have evolved under the strongest selection pressures, whether purifying, positive or balancing [START_REF] Quintana-Murci | Human Immunology through the Lens of Evolutionary Genetics[END_REF][START_REF] Barreiro | Evolutionary and population (epi)genetics of immunity to infection[END_REF]. By exploring how natural selection has shaped the variability of immune genes, population genetic studies have informed their biological relevance-essentiality, redundancy or adaptability-and provided a valuable complement to clinical, epidemiological and immunological approaches [START_REF] Quintana-Murci | Immunology in natura: clinical, epidemiological and evolutionary genetics of infectious diseases[END_REF]. Furthermore, there is increasing evidence to suggest that natural selection has contributed to the observed population differences in immune responses and susceptibility to infectious, inflammatory and autoimmune diseases [START_REF] Brinkworth | The contribution of natural selection to present-day susceptibility to chronic inflammatory and autoimmune disease[END_REF][START_REF] Sanz | Genetic and evolutionary determinants of human population variation in immune responses[END_REF].

In this review, we will not focus on the insights provided by population genetic studies of the immune system, as they have been extensively reviewed elsewhere [START_REF] Karlsson | Natural selection and infectious disease in human populations[END_REF][START_REF] Quintana-Murci | Human Immunology through the Lens of Evolutionary Genetics[END_REF][START_REF] Barreiro | Evolutionary and population (epi)genetics of immunity to infection[END_REF][START_REF] Quintana-Murci | Immunology in natura: clinical, epidemiological and evolutionary genetics of infectious diseases[END_REF][START_REF] Brinkworth | The contribution of natural selection to present-day susceptibility to chronic inflammatory and autoimmune disease[END_REF][START_REF] Sanz | Genetic and evolutionary determinants of human population variation in immune responses[END_REF][START_REF] Quintana-Murci | Population genetic tools for dissecting innate immunity in humans[END_REF][START_REF] Quach | Living in an adaptive world: Genomic dissection of the genus Homo and its immune response[END_REF]. Instead, we will discuss how hybridization with archaic hominins and modern admixture between populations (Figure 1) can facilitate human adaptation to pathogens and how such events have affected present-day individual and population differences in immune responses.

Finally, we will highlight how the use of ancient DNA, from humans and pathogens, can inform the dynamics of past epidemics and pinpoint genes and functions of key importance in immunity to infection.

The legacy of archaic hominins on human immunity

When modern humans left Africa to settle the rest of the world, they encountered 'archaic' hominins such as Neanderthals in Eurasia and Denisovans in the Asia-Pacific region [START_REF] Dannemann | Something old, something borrowed: admixture and adaptation in human evolution[END_REF].

The sequencing of the genomes of archaic hominins, including high-coverage genomes from three Neanderthals [START_REF] Mafessoni | A high-coverage Neandertal genome from Chagyrskaya Cave[END_REF][START_REF] Prufer | A high-coverage Neandertal genome from Vindija Cave in Croatia[END_REF][START_REF] Prufer | The complete genome sequence of a Neanderthal from the Altai Mountains[END_REF] and one Denisovan [START_REF] Meyer | A high-coverage genome sequence from an archaic Denisovan individual[END_REF], as well as several low-coverage Neanderthal genomes [START_REF] Hajdinjak | Reconstructing the genetic history of late Neanderthals[END_REF], has generated unprecedented knowledge on the extent of admixture between modern and archaic humans. We have learned that most of the Neanderthal ancestry present in the genomes of modern humans results from a single admixture event occurred after the out-of-Africa, while there is increasing evidence suggesting that several admixture events with Denisovans took place in different regions, and at different times [START_REF] Jacobs | Multiple Deeply Divergent Denisovan Ancestries in Papuans[END_REF][START_REF] Browning | Analysis of Human Sequence Data Reveals Two Pulses of Archaic Denisovan Admixture[END_REF][START_REF] Choin | Genomic insights into population history and biological adaptation in Oceania[END_REF]. This has resulted in ~2% Neanderthal ancestry in the genomes of all non-Africans, <1% Denisovan ancestry in East and South East Asians, and up to 5% Denisovan ancestry in some Pacific groups [START_REF] Choin | Genomic insights into population history and biological adaptation in Oceania[END_REF][START_REF] Qin | Denisovan Ancestry in East Eurasian and Native American Populations[END_REF][START_REF] Vernot | Excavating Neandertal and Denisovan DNA from the genomes of Melanesian individuals[END_REF][START_REF] Sankararaman | The Combined Landscape of Denisovan and Neanderthal Ancestry in Present-Day Humans[END_REF].

Recent studies have shown that hybridization with closely related species can be a source of advantageous variants facilitating the acquisition of beneficial traits (i.e., adaptive introgression) [START_REF] Taylor | Insights from genomes into the evolutionary importance and prevalence of hybridization in nature[END_REF]. Although the introgression of archaic material was generally selected against in humans [START_REF] Petr | Limits of long-term selection against Neandertal introgression[END_REF], high levels of Neanderthal or Denisovan ancestry at specific loci can be indicative of adaptive introgression [START_REF] Dannemann | Something old, something borrowed: admixture and adaptation in human evolution[END_REF]. Over the past few years, several studies have shown that archaic introgression has affected human immune functions, supporting the notion that ancient admixture facilitated genetic adaptation of modern humans to the new pathogens they encountered around the world. An influential study showed that innate immunity genes are enriched in Neanderthal ancestry, highlighting the beneficial nature of Neanderthal introgression [START_REF] Deschamps | Genomic Signatures of Selective Pressures and Introgression from Archaic Hominins at Human Innate Immunity Genes[END_REF]. Since then, other reports have detected high levels of Neanderthal or Denisovan ancestry at some specific genes, including the antiviral OAS genes, the TLR1-6-10 gene cluster or the inflammation-related TNFAIP3 gene [START_REF] Deschamps | Genomic Signatures of Selective Pressures and Introgression from Archaic Hominins at Human Innate Immunity Genes[END_REF][START_REF] Gittelman | Archaic Hominin Admixture Facilitated Adaptation to Out-of-Africa Environments[END_REF][START_REF] Racimo | Signatures of Archaic Adaptive Introgression in Present-Day Human Populations[END_REF][START_REF] Sams | Adaptively introgressed Neandertal haplotype at the OAS locus functionally impacts innate immune responses in humans[END_REF][START_REF] Dannemann | Introgression of Neandertal-and Denisovan-like Haplotypes Contributes to Adaptive Variation in Human Toll-like Receptors[END_REF][START_REF] Zammit | Denisovan, modern human and mouse TNFAIP3 alleles tune A20 phosphorylation and immunity[END_REF]. More recently, a genomic study of Pacific populations, who present the highest levels of archaic ancestry worldwide [START_REF] Qin | Denisovan Ancestry in East Eurasian and Native American Populations[END_REF][START_REF] Vernot | Excavating Neandertal and Denisovan DNA from the genomes of Melanesian individuals[END_REF][START_REF] Sankararaman | The Combined Landscape of Denisovan and Neanderthal Ancestry in Present-Day Humans[END_REF], has shown that while Neanderthals facilitated human adaptation related to phenotypes as diverse as immunity, neuronal development, metabolism and dermatological/pigmentation phenotypes, the beneficial nature of Denisovan introgression is primarily restricted to immune functions [START_REF] Choin | Genomic insights into population history and biological adaptation in Oceania[END_REF].

The introgression of archaic material in humans has been also found to affect molecular phenotypes, such as gene expression [START_REF] Dannemann | The Contribution of Neanderthals to Phenotypic Variation in Modern Humans[END_REF][START_REF] Mccoy | Impacts of Neanderthal-Introgressed Sequences on the Landscape of Human Gene Expression[END_REF]. For example, two studies found that mutations associated with gene expression variation (expression quantitative trait loci, or eQTLs) in monocytes and macrophages are enriched in Neanderthal ancestry in Europeans, particularly eQTLs associated with antiviral responses [START_REF] Nédelec | Genetic Ancestry and Natural Selection Drive Population Differences in Immune Responses to Pathogens[END_REF][START_REF] Quach | Genetic Adaptation and Neandertal Admixture Shaped the Immune System of Human Populations[END_REF]. Supporting further the notion that Neanderthals facilitated genetic adaptation of early Eurasians to viral challenges, viralinteracting proteins (VIPs) also are enriched in Neanderthal ancestry in modern humans, specially proteins interacting with RNA viruses [START_REF] Enard | Evidence that RNA Viruses Drove Adaptive Introgression between Neanderthals and Modern Humans[END_REF]. A recent study has explored how Neanderthal ancestry has affected other layers of gene regulation, including promoters, enhancers and miRNA-mediated regulation, and found that Neanderthal ancestry has mostly affected enhancers that are active in adipose-related tissues and various types of primary T cells [START_REF] Silvert | Impact and Evolutionary Determinants of Neanderthal Introgression on Transcriptional and Post-Transcriptional Regulation[END_REF]. Collectively, these studies highlight the importance of archaic introgression as a vehicle for modern human adaptation, by serving as a possible reservoir of resistance alleles against newly-encountered pathogens.

Neanderthal heritage and the recent COVID-19 pandemic

Recent works have explored the links between admixture with archaic hominins and human immunity against SARS-CoV-2 infection [START_REF] Zeberg | The major genetic risk factor for severe COVID-19 is inherited from Neanderthals[END_REF][START_REF] Zeberg | A genomic region associated with protection against severe COVID-19 is inherited from Neandertals[END_REF]. Several genomic regions have been associated with increased susceptibility of developing severe forms of COVID-19, including a region in chromosome 3 that spans 50 kb and contains 6 genes [START_REF] Covid | Genomewide Association Study of Severe Covid-19 with Respiratory Failure[END_REF]. The risk haplotype, which increases by 60% the odds of being hospitalized because of COVID-19, is of Neanderthal origin and is present in 16% of individuals from Europe, 50% of people from India and up to 63% of individuals from Bangladesh (Figure 2) [START_REF] Zeberg | The major genetic risk factor for severe COVID-19 is inherited from Neanderthals[END_REF]. Interestingly, an independent study has shown that individuals of Bangladeshi origin living in the UK has twice more chances to die from COVID-19 than the general population, supporting the deleterious nature of the genetic risk factor [START_REF] England | COVID-19: Review of Disparities in Risks and Outcomes[END_REF]. Yet, the high frequency of the risk haplotype among some contemporary populations suggests a different, even opposite adaptive value for it in past populations, when challenged by other pathogens.

Of note, the Neanderthal risk haplotype on chromosome 3 is absent among East and South-East Asian populations, an observation that well fits the extraordinarily low mortality observed in this region with respect to the number of cases. Although other factors may explain such low mortality rates, including socio-cultural and lifestyle factors, and a different management of the pandemic, a recent study argues that East Asians present some form of biological adaptation to coronaviruses. By analyzing selection signals in 420 genes encoding proteins that interact with coronaviruses in 26 worldwide populations, researchers have shown that the signals of positive selection, dating back to 20 000 years ago, are only present in populations of East-Asian ancestry [START_REF] Souilmi | An ancient viral epidemic involving host coronavirus interacting genes more than 20,000 years ago in East Asia[END_REF]. This suggests that an ancient coronavirus-like epidemic occurred in the ancestors of East Asians, the genetic legacy of which might confer, at least to some extent, higher levels of protection against severe COVID-19 among contemporary populations living in this region.

Finally, another report shows that the legacy of Neanderthals has not only been detrimental for modern humans in the context of the COVID-19 pandemic. A Neanderthal haplotype on chromosome 12, overlapping the antiviral OAS1, OAS2, and OAS3 and present at frequencies of ~30% in all non-African groups (Figure 2), has been found to reduce the odds to develop severe COVID-19 by 22% [START_REF] Zeberg | A genomic region associated with protection against severe COVID-19 is inherited from Neandertals[END_REF]. This observation, together with previouslyreported signals of positive selection targeting the Neanderthal OAS haplotype in humans [START_REF] Sams | Adaptively introgressed Neandertal haplotype at the OAS locus functionally impacts innate immune responses in humans[END_REF], lend support to the notion that Neanderthal introgression has been mostly beneficial for human adaptation to viral challenges [START_REF] Quach | Genetic Adaptation and Neandertal Admixture Shaped the Immune System of Human Populations[END_REF][START_REF] Enard | Evidence that RNA Viruses Drove Adaptive Introgression between Neanderthals and Modern Humans[END_REF].

Modern human admixture as a source of beneficial immune variation

Population genetic studies have distinguished two important periods in human evolution: the Late Pleistocene, when populations of hunter-gatherers dispersed over the globe, encountered archaic hominins and diversified, and the Holocene, a more recent period when large-scale population dispersals, facilitated by technological and/or demographic changes, resulted in admixture between differentiated populations. Ancient DNA studies have shown, for example, that admixture has occurred from ~7,500 years ago onward in Western Europe, between autochthonous hunter-gatherers and farmers originating from Anatolia [START_REF] Lazaridis | Ancient human genomes suggest three ancestral populations for present-day Europeans[END_REF][START_REF] Lipson | Parallel palaeogenomic transects reveal complex genetic history of early European farmers[END_REF].

Likewise, following their dispersals from western Central Africa ~5,000-3,000 years ago, Bantu-speaking populations admixed with hunter-gatherer and pastoralist groups from the rainforest, the East African highlands and the Kalahari Desert [START_REF] Busby | Admixture into and within sub-Saharan Africa[END_REF][START_REF] Patin | Dispersals and genetic adaptation of Bantuspeaking populations in Africa and North America[END_REF]. In the Pacific, population dispersals starting from Taiwan, known as the Austronesian expansion, resulted in extensive admixture with populations from Indonesia, the Philippines and Near Oceania within the last ~3,000 years [START_REF] Choin | Genomic insights into population history and biological adaptation in Oceania[END_REF][START_REF] Lipson | Reconstructing Austronesian population history in Island Southeast Asia[END_REF][START_REF] Pugach | The Gateway from Near into Remote Oceania: New Insights from Genome-Wide Data[END_REF]. Together, these studies indicate that recent admixture has been pervasive and has repeatedly shaped the genetic diversity of modern humans [START_REF] Hellenthal | A genetic atlas of human admixture history[END_REF].

Similarly to adaptive archaic introgression, theoretical and empirical evidence suggests that modern admixture has also facilitated human genetic adaptation [START_REF] Jeong | Admixture facilitates genetic adaptations to high altitude in Tibet[END_REF][START_REF] Pagani | Ethiopian genetic diversity reveals linguistic stratification and complex influences on the Ethiopian gene pool[END_REF][START_REF] Slatkin | Gene flow and the geographic structure of natural populations[END_REF] (Figure 1).

Although the role of this process, known as 'adaptive admixture' (or adaptive gene flow), in human evolution is yet to be fully determined, there is a growing number of cases of adaptive traits that have been acquired via intraspecies admixture, including resistance to pathogens.

The most striking example is the Duffy-null FY*O allele, which is thought to confer protection against Plasmodium vivax malaria (Figure 3). Indeed FY*O carriers do not express the Duffy antigen/chemokine receptor, which is required for the invasion of human erythrocytes by vivax merozoites [START_REF] Miller | The resistance factor to Plasmodium vivax in blacks. The Duffy-blood-group genotype, FyFy[END_REF][START_REF] Tournamille | Disruption of a GATA motif in the Duffy gene promoter abolishes erythroid gene expression in Duffy-negative individuals[END_REF]. FY*O occurred ~42,000 years ago in sub-Saharan Africa, where it has reached near fixation, possibly because vivax malaria was once endemic in the region [START_REF] Mcmanus | Population genetic analysis of the DARC locus (Duffy) reveals adaptation from standing variation associated with malaria resistance in humans[END_REF]. Interestingly, FY*O has been shown to be under strong, recent positive selection in African-descent admixed populations from various regions where vivax malaria is nowadays endemic, including Madagascar [START_REF] Hodgson | Natural selection for the Duffy-null allele in the recently admixed people of Madagascar[END_REF][START_REF] Pierron | Strong selection during the last millennium for African ancestry in the admixed population of Madagascar[END_REF], Cabo Verde [START_REF] Hamid | Rapid adaptation to malaria facilitated by admixture in the human population of Cabo Verde[END_REF], Sudan [START_REF] Triska | Extensive Admixture and Selective Pressure Across the Sahel Belt[END_REF] and Pakistan [START_REF] Laso-Jadart | The Genetic Legacy of the Indian Ocean Slave Trade: Recent Admixture and Post-admixture Selection in the Makranis of Pakistan[END_REF]. In the context of malaria, a recent study has shown that the hemoglobin β S sickle mutation, a well-known resistance factor against falciparum malaria, occurred ~22,000 years ago in the ancestors of present-day Bantu-speaking farmers from Central Africa, and was acquired by rainforest hunter-gatherers through adaptive admixture over the last 6,000 years [START_REF] Laval | Recent Adaptive Acquisition by African Rainforest Hunter-Gatherers of the Late Pleistocene Sickle-Cell Mutation Suggests Past Differences in Malaria Exposure[END_REF]. These findings collectively reinforce the view that Plasmodium parasites have imposed a strong burden on human populations for more than 20,000 years [START_REF] Mcmanus | Population genetic analysis of the DARC locus (Duffy) reveals adaptation from standing variation associated with malaria resistance in humans[END_REF][START_REF] Laval | Recent Adaptive Acquisition by African Rainforest Hunter-Gatherers of the Late Pleistocene Sickle-Cell Mutation Suggests Past Differences in Malaria Exposure[END_REF].

One of the earliest accounts of adaptive admixture in humans was described for the human leukocyte antigen (HLA) loci, which show an excess of African ancestry in admixed Puerto Ricans [60]. Although it has been advocated that different methodological artifacts can confound the analyses [START_REF] Price | Long-range LD can confound genome scans in admixed populations[END_REF], the signal has been consistently detected in populations from Mexico, Colombia, Costa Rica and Argentina [START_REF] Deng | Ancestry variation and footprints of natural selection along the genome in Latin American populations[END_REF][START_REF] Zhou | Strong Selection at MHC in Mexicans since Admixture[END_REF][START_REF] Norris | Admixture-enabled selection for rapid adaptive evolution in the Americas[END_REF]. Given the role of HLA in resistance to infections [START_REF] Tian | Genomewide association and HLA region fine-mapping studies identify susceptibility loci for multiple common infections[END_REF][START_REF] Sveinbjornsson | HLA class II sequence variants influence tuberculosis risk in populations of European ancestry[END_REF], it has been suggested that HLA alleles of African-descent have been selected to protect Native Americans from diseases introduced during the European colonization, such as smallpox, measles, and typhus. However, it is unclear which pathogen(s) was (were) responsible for the demographic collapse of Native Americans, a question that ancient DNA studies may soon resolve [START_REF] Vagene | Salmonella enterica genomes from victims of a major sixteenth-century epidemic in Mexico[END_REF]. Adaptive admixture at HLA genes has also been described in Central Africa [START_REF] Patin | Dispersals and genetic adaptation of Bantuspeaking populations in Africa and North America[END_REF] and Near Oceania [START_REF] Isshiki | Admixture and natural selection shaped genomes of an Austronesian-speaking population in the Solomon Islands[END_REF], highlighting HLA as a hotspot of recent selection. Overall, these findings suggest that, in addition to adaptive introgression, intraspecies admixture has facilitated genetic adaptation to the ever-changing world of pathogens that threatens our species.

Understanding recent immune evolution using ancient DNA data

Our understanding of the evolution of immune response genes has long been limited to statistical inferences from patterns of genetic variation in contemporary populations, including those harboring genetic material from archaic hominins. However, ancient DNA techniques have facilitated studies of human adaptation to pathogen exposure; by analyzing the genomes of populations from different epochs, one can directly measure the effects of natural selection (Figure 4). In doing so, recent studies have increased our understanding of the last 10,000 years of human evolution, a time transect that benefits from more than 5,000 available ancient human genomes, covering mainly Europe but also other regions of the world.

A pioneering study, based on 230 Eurasian samples across the Holocene period, revealed 12 loci under strong positive selection, including three that are directly linked to immune functions: the TLR10-TLR1-TLR6 gene cluster, genes regulating autophagy and the HLA region [START_REF] Mathieson | Genome-wide patterns of selection in 230 ancient Eurasians[END_REF]. Another study sought to explore how the arrival of Europeans to the Americas may have altered the exposure of Native Americans to new pathogen cues. Based on exome sequences from a population of First Nations of the northwest coast of Canada, dating from before and after the first contact with Europeans, the authors observed a recent decrease in frequency of formerly beneficial HLA-DQA1 alleles in the indigenous population, which they attribute to the environmental changes triggered by the European arrival [START_REF] Lindo | A time transect of exomes from a Native American population before and after European contact[END_REF].

However, which precise HLA alleles or haplotypes have played a major role in human biological adaptation remains an open question [START_REF] Barquera | An ancient view on host pathogen interaction across time and space[END_REF]. In this context, the reconstruction of HLA class I and II alleles for a large number of Europeans belonging to the Wartberg culture (5,500-4,800 years ago) indicates that immune response was probably more adapted to viral than bacterial infections during the Neolithic [START_REF] Immel | Genome-wide study of a Neolithic Wartberg grave community reveals distinct HLA variation and hunter-gatherer ancestry[END_REF].

Interestingly, no biological or archeological evidence for diseases of epidemic proportions have been noted earlier than the Bronze Age in Europe [START_REF] Fuchs | Infectious diseases and Neolithic transformations: Evaluating biological and archaeological proxies in the German loess zone between 5500 and 2500 BCE[END_REF]. The notion that the Bronze Age has been an important period of biological adaptation to pathogens, and particularly to bacteria, has been recently reinforced by a study that focused on human genetic susceptibility to Mycobacterium tuberculosis (M. tuberculosis) [START_REF] Kerner | Human ancient DNA analyses reveal the high burden of tuberculosis in Europeans over the last 2,000 years[END_REF]. Using a database composed of more than 1,000 ancient European genomes, dating from the Mesolithic to the Middle Ages, we have analyzed the frequency trajectory of the only mutation so far identified as underlying, in homozygosity, a common monogenic form of human tuberculosis (TB), the TYK2 P1104A variant, which is present in 2-4% of contemporary individuals of European ancestry [START_REF] Boisson-Dupuis | Tuberculosis and impaired IL-23dependent IFN-gamma immunity in humans homozygous for a common TYK2 missense variant[END_REF][START_REF] Kerner | Homozygosity for TYK2 P1104A underlies tuberculosis in about 1% of patients in a cohort of European ancestry[END_REF]. The study has shown that the TB-risk mutation appeared in western Eurasians ~30,000 years ago, but has evolved under strong purifying selection only for the last 2,000 years [START_REF] Kerner | Human ancient DNA analyses reveal the high burden of tuberculosis in Europeans over the last 2,000 years[END_REF]. These analyses indicate that TB has imposed a heavy burden on European health over the last two millennia, a date that coincides with the emergence of some specific M. tuberculosis strains in the region [START_REF] O'neill | Lineage specific histories of Mycobacterium tuberculosis dispersal in Africa and Eurasia[END_REF][START_REF] Sabin | A seventeenth-century Mycobacterium tuberculosis genome supports a Neolithic emergence of the Mycobacterium tuberculosis complex[END_REF][START_REF] Kay | Eighteenth-century genomes show that mixed infections were common at time of peak tuberculosis in Europe[END_REF].

Reconstructing past epidemics through ancient pathogen genomics

To understand how past epidemics have forged the evolution of the immune system, one can also leverage the field of ancient pathogen genomics, which has proven to be informative for identifying causative agents of past pandemics and reconstructing the history of human exposure to pathogens [START_REF] Valtueña | The Stone Age Plague and Its Persistence in Eurasia[END_REF][START_REF] Duchêne | The Recovery, Interpretation and Use of Ancient Pathogen Genomes[END_REF]. The most remarkable example relates to Yersinia pestis (Y. pestis), the causative agent of plague. By reconstructing the genome of Y. pestis preserved in Neolithic farmers of ancient Sweden, found to be basal to all modern and ancient known strains, a recent study envisioned the possibility of a prehistoric plague pandemic predating all other known human pandemics [START_REF] Rascovan | Emergence and Spread of Basal Lineages of Yersinia pestis during the Neolithic Decline[END_REF]. Yet, flea adaptation, known to enhance plague dispersal, was not acquired by Y. pestis until the end of the Neolithic, suggesting the rise of plague epidemics during the Bronze Age [START_REF] Spyrou | Analysis of 3800-year-old Yersinia pestis genomes suggests Bronze Age origin for bubonic plague[END_REF].

The field of ancient pathogen genomics can also inform the history of infectious agents that remain highly relevant to public health today, as is the case of M. tuberculosis. Whereas modern genetic data had suggested an African origin and out-of-Africa dispersal 70,000 years ago for the causative agent of TB [START_REF] Comas | Out-of-Africa migration and Neolithic coexpansion of Mycobacterium tuberculosis with modern humans[END_REF], studies based on a 1,000-year-old mycobacterial genome from Peru had re-estimated its emergence to a maximum of 6,000 years ago [START_REF] Bos | Pre-Columbian mycobacterial genomes reveal seals as a source of New World human tuberculosis[END_REF]. Other works, based on high-quality ancient mycobacteria of the 18 th and 17 th centuries, have also supported a recent origin of TB, and the appearance of one of the most prevalent lineages of M. tuberculosis less than 2,500 years ago [START_REF] Sabin | A seventeenth-century Mycobacterium tuberculosis genome supports a Neolithic emergence of the Mycobacterium tuberculosis complex[END_REF][START_REF] Kay | Eighteenth-century genomes show that mixed infections were common at time of peak tuberculosis in Europe[END_REF]. Altogether, the integration of both ancient human and pathogen genomes into disease modelling, combined with appropriate immune functional validation, will help delineate the most significant hostpathogen interactions during human evolution.

Conclusions

Understanding how immune gene variation has contributed to our species' survival and success through archaic or modern admixture has been a topic of active research over the past few years. The sequencing of the genomes of archaic hominins and their contribution to modern non-African ancestry [START_REF] Dannemann | Something old, something borrowed: admixture and adaptation in human evolution[END_REF] has allowed to unravel some key immune players that facilitated the adaptation of early non-Africans to the newly encountered environments.

Likewise, admixture between modern human groups during the Holocene has enabled genetic adaptation to pre-existing pathogens, supporting the poison/antidote model whereby a population transmits a new disease to another population (the poison) but also the means to fight it (the antidote) [START_REF] Enard | Evidence that RNA Viruses Drove Adaptive Introgression between Neanderthals and Modern Humans[END_REF]. Possible cases include that of rainforest hunter-gatherers from Central Africa who recently acquired the sickle-cell mutation from neighboring farmers to prevent malaria [START_REF] Laval | Recent Adaptive Acquisition by African Rainforest Hunter-Gatherers of the Late Pleistocene Sickle-Cell Mutation Suggests Past Differences in Malaria Exposure[END_REF], and that of Latin Americans who have most likely acquired beneficial HLA alleles through admixture with Africans, following the European colonization and the respective introduction of new pathogens [60, [START_REF] Deng | Ancestry variation and footprints of natural selection along the genome in Latin American populations[END_REF][START_REF] Zhou | Strong Selection at MHC in Mexicans since Admixture[END_REF][START_REF] Norris | Admixture-enabled selection for rapid adaptive evolution in the Americas[END_REF]. Despite these new findings, the study of how humans from different regions of the world have historically adapted to the diverse set of pathogens they have encountered remains in its infancy. Yet, the study of human genetic diversity is at the basis of understanding the observed population differences in immune responses, which is essential for the development of therapeutic treatments at a worldwide scale. Exploring, for example, immune gene diversity in Pacific populations, with the highest amount of combined archaic ancestry, and in African populations, with the lowest, might reveal key genetic components underlying population disparities in susceptibility to infectious, inflammatory, and autoimmune disorders.

Ancient DNA studies are also expected to shed light on the role of yet unknown immunerelated loci, especially on those having undergone purifying selection, that is, the selective removal of deleterious alleles from the population. The use of modern DNA datasets to study negatively-selected variants has been a daunting task because, in light of their deleterious nature, such variants are underrepresented, if not extinct, in contemporary populations. In this context, ancient genomic studies based on allele frequency fluctuations over time, as that on the TB-risk TYK2 P1104A allele [START_REF] Kerner | Human ancient DNA analyses reveal the high burden of tuberculosis in Europeans over the last 2,000 years[END_REF], can be used to delineate human genetic variants that have been selected due to microbial pressure. Studies of ancient populations dating from before, during and after massive death episodes, such as past epidemics, might reveal strong signatures of selection owing to specific infectious agents. For example, ancient DNA data should help reconstructing the impact of Black Death in European populations as well as the burden caused by the European arrival (and their pathogens) in Native American populations at the immune level. Furthermore, the reconstruction of how humans have adapted to major pandemics in the past will help understanding the genetic factors involved in our current predisposition to infectious disease. In this context, additional studies with denser ancient DNA datasets across time and space, combined with modern genomes from additional populations and innovative methodological tools to detect more complex patterns of selection, will improve our understanding of the determinants of human immune evolution and its current diversity. illustrated how population genetics can help reconstructing historical exposure to pathogens.

Figure 1 .Figure 2 .

 12 Figure 1. Graphical representation of the beneficial nature of ancient or modern admixture.On the left panel, the contribution of genetic material from archaic humans, such as Neanderthals or Denisovans, to non-Africans is represented. Neanderthals contributed genetic material to both Europeans and Asians, while the contribution of Denisovan ancestry is restricted to the Asia-Pacific region. In most cases, the introgression of archaic material was selected against in the genomes of modern humans, yet, in some cases, such archaic segments were beneficial for modern human adaptation a process known as 'adaptive introgression'. On the right panel, admixture, or gene flow, between two modern human populations is represented. Population 2 (the donor) has sent gene flow to Population 1 (the recipient); in some cases, the genetic material of the donor population can be beneficial for the recipient population, and such material can increase in frequency in the latter population by positive Darwinian selection, a phenomenon known as 'adaptive admixture' or 'adaptive gene flow'.

Figure 3 .

 3 Figure 3. Adaptive admixture at the Duffy-null FY*O allele in modern humans. Gray, light and dark orange areas indicate P. vivax-free regions and regions of unstable and stable P.vivax transmission, respectively (adapted from ref.[START_REF] Gething | A long neglected world malaria map: Plasmodium vivax endemicity in 2010[END_REF]). The dashed areas indicate regions where the Duffy-null FY*O allele is higher than 90%. In case 1, the region is free from P. vivax and most resident populations are resistant to vivax malaria. In case 2, P. vivax transmission is stable, and most resident populations are susceptible to vivax malaria. In case 3, populations have recently acquired P. vivax resistance through admixture with sub-Saharan Africans, and resistance is rapidly increasing in frequency under strong positive selection.

Figure 4 .

 4 Figure 4.A general sketch for the rational of studying past epidemics using aDNA data. Human remains of individuals that have been exposed to ancient pathogens at different epochs, sharing some degree of genetic homogeneity (genetic continuity), are collected for DNA extraction. Human DNA but also microbe DNA can be obtained from the collected bones. Genomics of ancient individuals are then compared, on the basis of prior hypothesis, using statistical tools. Genome-wide assessment of selection at the variant level (Manhattan plot), intensity for the selection coefficient of a significantly selected variant (blue distribution), age of the mutation and onset of selection for the selected variant (demographic model) and a phylogeny for the involved pathogen can all be investigated with the extracted aDNA data.

Acknowledgements

The laboratory of Human Evolutionary Genetics is supported by the Institut Pasteur, the Collège de France, the Centre Nationale de la Recherche Scientifique (CNRS), the Agence Nationale de la Recherche (ANR) grants LIFECHANGE (ANR-17-CE12-0018-02), MORTUI (ANR-19-CE35-0005) and CNSVIRGEN (ANR-19-CE15-0009-02), the French Government's Investissement d'Avenir program, Laboratoires d'Excellence "Integrative Biology of Emerging Infectious Diseases" (ANR-10-LABX-62-IBEID) and "Milieu Intérieur" (ANR-10-LABX-69-01), the Fondation pour la Recherche Médicale (Equipe FRM DEQ20180339214), the Fondation Allianz-Institut de France, and the Fondation de France (n°00106080).

Conflict of interest statement

Nothing declared