Camille Jacqueline

Nathalie Bonnefoy

Guillaume M Charriere

Frederic Thomas

Benjamin Roche

Personal history of infections and immunotherapy: Unexpected links and possible therapeutic opportunities

Keywords: Immunotherapy, Infections, Personalized medicine, Cancer vaccine, Immune checkpoints, Adoptive cell transfe

The recent breakthroughs in the understanding of tumor immune biology have given rise to a new generation of immunotherapies, harnessing the immune system to eliminate tumors. As the typology and frequency of encountered infections are susceptible to shape the immune system, it could also impact the efficiency of immunotherapy. In this review, we report evidences for an indirect link between personal history of infection and different strategies of immunotherapy. In the current context of interest rise for personalized medicine, we discuss the potential medical applications of considering personal history of infection to design immunotherapeutic strategies.

Introduction

Genomic instability, common to all cancers, 1 is a challenge for current cytotoxic therapies and represents the main cause of therapy failure and cancer relapse. Indeed, treatments generally target the dominant clones within the tumor and result in massive cell death. Nevertheless, it may paradoxically provide a selective pressure for the expansion of variant cells resisting to the drug used. [START_REF] Greaves | Clonal evolution in cancer[END_REF] Combinations of chemotherapies have been tested, and have shown encouraging results, but only delay emergence of resistance and rarely avoid it. [START_REF] Bozic | Evolutionary dynamics of cancer in response to targeted combination therapy[END_REF] This is why the research on cancer treatment has evolved from relatively nonspecific cytotoxic agents to selective, mechanism-based therapeutics. These 'targeted therapies' have been designed to interfere with specific molecular targets that are known to have a critical role in tumor growth or progression. [START_REF] Sawyers | Targeted cancer therapy[END_REF] For instance, treatments based on VEGF have increased the time before the cancer progresses as well as improving survival for patient with renal cell carcinoma. [START_REF] Motzer | Lenvatinib, everolimus, and the combination in patients with metastatic renal cell carcinoma: a randomised, phase 2, open-label, multicentre trial[END_REF] Another key point in the recent evolution of cancer therapies is the observation that cancerous patients can be stabilized over metronomic treatments (chemo-, radio-or targeted therapies) through the induction of a long-term anti-cancer immune response or vaccine-like effect. [START_REF] Pasquier | Metronomic chemotherapy: new rationale for new directions[END_REF] To this extent, the recent breakthroughs in the understanding of tumor immune biology have given rise to a new generation of treatments which attempts to harness the exquisite power and specificity of the immune system for the treatment of malignancy. This cancer immunotherapy could allow overcoming immune tolerance to tumors and the production of adaptive anti-cancer responses able to deal with genomic instability. [START_REF] Yaghmour | Role of genomic instability in immunotherapy with checkpoint inhibitors[END_REF] Indeed, since the beginning of the 20th century, it has been hypothesized that immune system may play a role in cancer surveillance and prevention. The pioneering works of Dr William B. Coley have showed that stimulation of immune system by infections could be associated with tumor growth control. [START_REF] Coley | The treatment of inoperable sarcoma by bacterial toxins(the Mixed Toxins of the Streptococcus erysipelas and the Bacillus prodigiosus)[END_REF] However, direct evidence for immunosurveillance of human cancers has been hard to prove experimentally and immunotherapy has mainly consisted in passive transfer of anti-cancer antibodies and T cells from donor to induce an immediate immune reaction against cancer. [START_REF] Dougan | Immune therapy for cancer[END_REF] Recently, the clinical success of new immunotherapies, such as immune checkpoint inhibitors leading to complete tumor elimination in some patients with advanced cancers, [START_REF] Topalian | Survival, durable tumor remission, and long-term safety in patients with advanced melanoma receiving nivolumab[END_REF] supports the idea that immune system is a powerful weapon against cancer. Nevertheless, the successes of immunotherapy are extremely heterogeneous, sometimes associated with dramatic side-effects, 11 and resistance has already been reported. [START_REF] Restifo | Acquired resistance to immunotherapy and future challenges[END_REF] Thus, identification of predictive biomarkers allowing selection of patients predisposed to respond to immunotherapy remains a challenge for immunologists and oncologists. In addition to immunosurveillance of cancer, a prime function of immune system is the defense against infectious agents (i.e., viruses, bacteria, fungi, protozoans and metazoans that exploit other organisms, called hosts, to complete their life cycle). At an individual scale, humans are exposed to a high number of infections (through contact, ingestion or inhalation among other possibilities) which sometimes result in infraclinical symptoms. [START_REF] Barnes | Diseases and human evolution[END_REF] The community of organisms which have infected an individual during its life represents the personal history of infection. As cancer cells and infections interact with organisms' defense mechanisms, typology and frequency of encountered infections are susceptible to shape the functioning of immune system, and consequently the efficiency of immunotherapy. Here, we review the indirect evidences suggesting that personal history of infections may impact, in a non-negligible way, many mechanisms targeted by immunotherapeutic strategies and therefore may explain their contrasting successes as well as open new opportunities to improve them.

Strategies envisioned for immunotherapy

Today, several categories of immunotherapy exist which differ according to their target. First, we can distinguish immunotherapies designed to directly target tumor cells but which rarely result in a long-lasting anti-tumor immune response. It includes anti-tumor antibodies as well as cellular therapy, also known as adoptive cell transfer, aiming to select and/or to engineer tumor-specific immune cells that are thereafter administrated by local injection (i.e., intratumoral). [START_REF] Shi | Chimeric antigen receptor for adoptiveimmunotherapy of cancer: latest research and future prospects[END_REF] A second category includes immunotherapies that stimulate the patient own immune system to elicit cancer cells elimination and an immune memory protective against tumor-associated antigens (TAAs). They are generally administrated by intravenous injection to generate a systemic immune response. They include antibodies directed against immunemodulating molecules present on immune cells such as CTLA-4, PD-1 and PDL-1, PDL-1 being also expressed by tumor cells themselves. At the interface between these two strategies, oncolytic therapies are based on viruses that show a tropism for cancer cells. This cytotoxic therapy stimulates both cancer cell destruction as well as anti-tumor immune response. Finally, immunotherapy also includes strategies based on immune-mechanisms, such as vaccines that either treat existing cancer or prevent development of a cancer.

Cellular therapy

CAR T cells

Adoptive transfer of T cells engineered to express chimeric antigen receptor (CAR) has emerged as a powerful immunotherapeutic strategy and is now the focus of numerous clinical trials. Especially, CAR-modified T cells targeting the B cell specific antigen CD19 have shown great promise in the treatment of chronic lymphocytic leukemia (CLL) [START_REF] Kalos | Tcells with chimeric antigen receptors have potent antitumor effects and can establish memory in patients with advanced leukemia[END_REF] and acute lymphocytic leukemia (ALL). [START_REF] Grupp | Chimeric antigen receptor-modified T cells for acute lymphoid leukemia[END_REF] The most notable toxicity of this treatment is the cytokine release syndrome (CRS), which is a systemic inflammatory process resulting from attack by CAR T cells of nonmalignant cells that express targeted tumor antigen. [START_REF] Lee | How I Treat: Current concepts in the diagnosis and management of cytokine release syndrome[END_REF] In some patients, CD19-targeted T cells therapy has also been associated to neurological disturbances. A study has shown that CAR T cells are able to penetrate blood-brain barrier [START_REF] Hu | Predominant cerebral cytokine release syndrome in CD19directed chimeric antigen receptor-modified T cell therapy[END_REF] and they have been observed in the cerebrospinal fluid of the patient experiencing neurological toxicities. [START_REF] Davila | Efficacy and toxicity management of 19-28z CAR T Cell therapy in B Cell acute lymphoblastic leukemia[END_REF] We can expect that neurological complications may be more frequent in patients with a high level of B cell infiltration in the brain. Indeed, the immune response produced by CAR T cells recognizing CD19 B cells could be associated with cytotoxicity to the surrounding tissues. In addition to the natural variability between individuals, the level of cerebral infiltration by B cells could also be the result of past infections. Notably, acute malaria infection has been associated to a significant increase of CD19 B lymphocyte percentages in the brain of infected animals in a rodent malaria model. [START_REF] Lacerda-Queiroz | Plasmodium berghei NK65 induces cerebral leukocyte recruitment in vivo: An intravital microscopic study[END_REF] This increase is susceptible to protect against cerebral malaria, which is a frequent and deadly complication in children. [START_REF] Bao | CD19(C) B Cells confer protection against experimental cerebral malaria in semi-immune rodent model[END_REF] It may be worth mentioning that certain viral brain infections (i.e., cytomegalovirus (CMV), West Nile Virus) may also be associated with an infiltration of immune cells and especially immune-regulatory B cells in the brain, as suggested by an experimental study on mice. [START_REF] Mutnal | Infiltrating regulatory B cells control neuroinflammation following viral brain infection[END_REF] However, to our knowledge, the persistence of B cells within the brain after recovery of infection has not been investigated. If this was confirmed, neurological complications with CD19-targeted CAR T cells could be more common in patients with an history of malaria or such viral infections. Another approach to generate CAR T cells is to use natural virus-specific T cells that have been modified to express antitumor CAR. These techniques generally lay on the dual-specificity of CAR T cells for infected cells and for tumor cells. In this manner, clonal expansion resulting from response to infection may benefit to tumor cell control and elimination. For example, EBV-specific T cells have been transduced with tumor-specific chimeric receptors that allow the recognition of GD2, a TAA present on several tumors of neural crest origin. [START_REF] Mujoo | Functional properties and effect on growth suppression of human neuroblastoma tumors by isotype switch variants of monoclonal antiganglioside GD2 antibody 14.18[END_REF] The authors have observed that EBV-specific CAR T cells recognize EBV-infected targets through their conventional T-cell receptors and tumor targets through their chimeric receptors. [START_REF] Rossig | Epstein-Barr virusspecific human T lymphocytes expressing antitumor chimeric T-cell receptors: Potential for improved immunotherapy[END_REF] Consequently, they are maintained by the presence of EBVinfected B cells and induce a higher anti-tumor activity which has increased survival in two mice tumor models: neuroblastoma and Hodgkin diseases. Influenza and CMV have also been used to generate dual-specific CAR T cells [START_REF] Murphy | Antitumor activity of dual-specific T cells and influenza virus[END_REF][START_REF] Heemskerk | Reprogramming of virus-specific T cells into leukemiareactive T cells using T cell receptor gene transfer[END_REF] but their efficiencies in presence of the virus have not been tested. We would like to propose here that the establishment of personal history of infection (especially by viruses and P. falciparum), before treatment administration, may help to select patients in order to prevent the apparition of dramatic adverse-effect, such as those observed with CD19trageted CAR T cells. In addition, we can suspect that the use of virus-based CAR T cells in patients who have been already infected by the natural virus, could show a higher efficiency. For instance, EBV infects more than 80% of the adult population. [START_REF] Balfour | Age-specific prevalence of epstein-barr virus infection among individuals aged 6-19 Years in the united states and factors affecting its acquisition[END_REF] In these patients, the use of EBV-specific CAR T cells without simultaneous introduction of the virus, or with lower viral dose to avoid side-effects, could improve anti-tumor responses (Fig. 1). The generation of a cocktail of dual-specific CAR T cells, specific to persisting pathogens (EBV, CMV, herpesvirus) and engineered to target a panel of common TAAs, could improve immunotherapy efficiency and raises new perspectives for prevention.

TILs

Given the lack of well-defined TAAs for most human tumors, the use of naturally occurring tumor infiltrating lymphocytes (TILs) represents a good alternative to CAR T cells. In fact, TILs are able to recognize multiple TAAs that increase their ability to keep tumor growth in check. Nevertheless, insufficient migration [START_REF] Joyce | T cell exclusion, immune privilege, and thetumor microenvironment[END_REF] and decreased anti-tumor function in the immunosuppressive tumor microenvironment [START_REF] Gajewski | Innate and adaptive immune cellsin the tumor microenvironment[END_REF] are significant limitations to treat cancer patients with adoptive transfer of ex-vivo expanded TILs. However, the identification of cytotoxic tumorinfiltrating T cells that recognize clonal mutations, shared by all tumor cells, hold promise for adoptive cellular therapy strategies in combination with the appropriate immunomodulatory agent. [START_REF] Mcgranahan | Clonal neoantigens elicit T cell immunoreactivity and sensitivity to immune checkpoint blockade[END_REF] Recently, tumor-specific T cells from patients have been engineered to carry a second T cell receptor (TCR) recognizing antigens from the bacteria Listeria monocytogenes. [START_REF] Xin | Pathogen boosted adoptive cell transfer immunotherapy to treat solid tumors[END_REF] These polyclonal dual-specific T cells show a greater antitumor effect in presence of specific bacterial antigens, which are simultaneously injected at the tumor site. Indeed, the presence of bacterial antigens allows a greater clonal expansion of dual-specific T cells and decreases immunosuppression in tumor micro-environment. However, patients who have already been exposed to the bacteria, or a closely related one, could show a stronger immune response against these antigens. This may result in a quick elimination of the bacterial antigens, neutralizing the positive impact on clonal expression and thus decreasing therapy efficiency. But, the consequences of this pre-exposition in term of treatment efficacy have not been investigated. Thus, within the context of personalized medicine rise, the choice of an additional infectious agent in adoptive TILs transfer should rely on the personal history of infections of the patient in order to maximize therapy efficiency.

Immune checkpoint inhibitors

Anti-CTLA-4

Cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) is a negative regulator of T cell activation. CTLA-4 acts by competing with the co-stimulatory molecule CD28 for binding to shared ligands CD80 and CD86 expressed by antigen-presenting cells. Antibodies that block CTLA-4, such as the ipilimumab, are known to increase T-cell activation and proliferation. Therefore, blocking CTLA-4 has the potential to enhance antitumor immune responses. [START_REF] Leach | Enhancement of antitumorimmunity by CTLA-4 blockade[END_REF] Treatment with ipilimumab can cause significant tumor response in patients with metastatic melanoma. [START_REF] Hodi | Improved survival with ipilimumab in patients with metastatic melanoma[END_REF] However, complete response has rarely been observed [START_REF] Prieto | CTLA-4 blockade with ipilimumab: Long-term follow-up of 177 patients with metastatic melanoma[END_REF] and immune-related adverse events, related to tolerance break and consequent auto-immunity, happen at high frequency. [START_REF] Bertrand | Immune related adverse events associated with anti-CTLA-4 antibodies: Systematic review and meta-analysis[END_REF] Regarding the interactions between infections and CTLA4, a study has shown that the expression of CTLA-4 was depressed in peripheral blood mononuclear cells from tuberculosis patients, [START_REF] Gong | Interleukin-10 downregulates Mycobacterium tuberculosis-induced Th1 responses and CTLA-4 expression[END_REF] suggesting that people exposed to Mycobacterium tuberculosis could be more responsive to such immunotherapy. Nowadays, up to one-third of the world's population is infected with M. tuberculosis and the majority of infected persons have a clinically latent infection. [START_REF] Flynn | Tuberculosis : Latency and reactivation[END_REF] Even if the prevalence of tuberculosis is expected to be low in developed countries, where such immunotherapeutic strategy is available, this mechanism could be mimicked by TB vaccination (Bacillus Calmette-Guerin (BCG) is routinely administrated in many countries) or applied to infection with other mycobacteria. Interestingly, intravesical injection of BCG vaccine is approved for the treatment of noninvasive bladder cancer [START_REF] Gontero | The role of bacillus Calmette-Guerin in the treatment of non-muscle-invasive bladder cancer[END_REF] and its efficiency has been tested in combination with antiCTLA-4 antibodies in a murine model. It has been observed that mice treated with BCG and anti-CTLA-4 show an improved survival than mice receiving BCG only. [START_REF] Mangsbo | Enhanced tumor eradication by combining CTLA-4 or PD-1 Blockade with CpG Therapy[END_REF] Therefore, the use of ipilimumab in patients with a history of mycobacterial infection, where the target of ipilimumab is already under-expressed, could block the immunosuppressive action of remaining molecules and could therefore result in stronger immune activation (Fig. 2A). Thus, it would be intriguing to evaluate in clinical trials whether tuberculosis/vaccination status might have an impact on response to anti-CTLA-4 but also whether it would allow diminishing the dose of administration to avoid immuneregulation of immune system. Expression of PD-1, signed related adverse events (Fig. 2B).

Anti-PD-1 & anti-PD-L1

Programmed death 1 (PD-1) is a key immune-checkpoint receptor expressed by activated T cells that mediates downregulation of immune system. Expression of PD-1, signed the "exhausted phenotype" of effector T cells, has been first identified through studies of chronic viral infections in mice in which PD-1/PD-L1 was found to be an important negative regulatory feedback loop to ensure immune homeostasis.40 Cancer immunotherapies based on the administration of anti-PD-1 (such as nivolumab and pembrolizumab) or anti-PD-L1 (such as atezolizumab, avelumab or durmalumab) aim to inhibit the interaction between PD-L1, expressed by tumor cells but also by stromal cells, and PD1, expressed by T cells, and thus restore T-cell antitumor activity.41 Many investigators have addressed the question of whether the expression of PD-1 or PD-L1 might be a prognostic marker of treatments with immune checkpoint inhibitors.42,43 Although it is still a matter of debate for PD-L1, PD-1 expression within the tumor reflecting the presence of specific anti-tumor lymphocytes has been associated with a positive prognostic feature in many cancers. 44 Importantly, the basal level of PD-1 expression by T cells could be shaped by personal history of infection. [START_REF] Salazar M D'a | Evaluation of innate and adaptive immunity contributing to the antitumor effects of PD1 blockade in an orthotopic murine model of pancreatic cancer[END_REF] As mentioned above, the increase of PD-1 expression on antigen-experienced effector T cells during chronic infections, is a physiological mechanism to constrain the adaptive immune response and prevent auto-immunity. [START_REF] Sharpe | The function of programmed cell death 1 and its ligands in regulating autoimmunity and infection[END_REF] For instance, HIV infection has been associated with an increase of PD-1 expression on NK cells. [START_REF] Norris | PD-1 expression on natural killer cells and CD8 C T cells during chronic HIV-1 infection[END_REF] The alteration of PD-1 expression in the context of acute infections has also attracted increasing attention. It has been reported that acute infections with M. tuberculosis and influenza virus result in a significant increase of PD-1 on NK cells and CD8C T cells respectively. [START_REF] Alvarez | Role played by the programmed death1-Programmed death ligand pathway during innate immunity against mycobacterium tuberculosis[END_REF][START_REF] Kohlhapp | Nononcogenic acute viral infections disrupt anti-cancer responses and lead to accelerated cancer-specific host death[END_REF] Moreover, evidences suggest that PD-1 expression could also be modified at long-term through immunological memory. In fact, memory CD8 C T cells excessively up-regulate PD-1 expression following vaccinia virus reinfection. [START_REF] Fuse | Recall responses by helpless memory CD8C T cells are restricted by the up-regulation of PD-1[END_REF] Hence patients who have experienced a relatively high number of infections may also express high level of PD-1, in activated as well as memory immune cells, reflecting the immunosuppression of pre-existing T cell immunity. Contrarily to NK cells, CD8 C T cells are antigen-dependant and thus specific T cells could target cancer cells only if TAAs share similarities with infectious agents. This situation has already be suggested between influenza virus and lung cancer cells. [START_REF] Iheagwara | Influenza virus infection elicits protective antibodies and T cells specific for host cell antigens also expressed as tumor associated antigens : A new view of cancer immunosurveillance[END_REF] In this context, we could expect that checkpoint inhibitors will present a lower efficacy in patients with a high basal level of PD-1. We also suggest that this effect may not be limited by clonal selection of cancer cells. In fact, cancer cells which harbor the ligand PD-L1 may have been favored because it allows them to escape immune destruction by the high number of immune cells expressing PD-1. If so, new strategies, aiming to increase immunotherapy efficiency and avoid dangerous sideeffects, should consider adapting the dose of treatments with antagonists of the PD-1/PDL-1 interaction, according to the past history of infections of each patient.

Tumor-targeting monoclonal antibodies

Although developed to directly target malignant cells, it is now recognized that the therapeutic activity of tumor antigentargeting monoclonal antibodies (mAb) is associated to their capacity to mobilize, through FcgR-dependent mechanisms, both innate and adaptive immune cells. However, tumor escape from mAb-induced tumor immune surveillance remains one of the main clinical issues. [START_REF] Michaud | Tumor antigen-targeting monoclonal antibody-based immunotherapy: Orchestrating combined strategies for the development of long-term antitumor immunity[END_REF][START_REF] They | PD-1 blockade at the time of tumor escape potentiates the immune-mediated antitumor effects of a melanoma-targeting monoclonal antibody[END_REF] As an example, anti-HER2/ErbB2 mAb (Trastuzumab) aims to reduce cell proliferation by blocking a receptor (ErbB2) of a growth factor highly expressed by many cancer cells. This therapy has substantially improved outcome of HER2-positive breast cancer [START_REF] Swain | Pertuzumab, trastuzumab, and docetaxel in HER2positive metastatic breast cancer[END_REF] and, given the importance of immune responses in trastuzumab therapy, [START_REF] Loi | Tumor infiltrating lymphocytes are prognostic in triple negative breast cancer and predictive for trastuzumab benefit in early breast cancer: Results from the FinHER trial[END_REF] therapy combined with immune stimulating agents have been used with success. [START_REF] Nava-Parada | Peptide vaccine given with aToll-like receptor agonist is effective for the treatment and prevention of spontaneous breast tumors[END_REF] Recently, co-injection of polyinosinic-polycytidylic acid (PolyI:C) and CpG oligodeoxynucleotides (CpG) with trastuzumab shows higher efficiency than anti-ErbB2 therapy used alone. [START_REF] Charlebois | PolyI:C and CpG synergize anti-ErbB2 mAb for treatment of breast tumors resistant to immune checkpoint inhibitors[END_REF] The authors suggest that these molecules may represent pertinent adjuvant stimulating innate responses including interferon expression which is required in trastuzumab therapy. [START_REF] Stagg | Anti-ErbB-2 mAb therapy requires type I and II interferons and synergizes with anti-PD-1 or anti-CD137 mAb therapy[END_REF] PolyI:C is a synthetic viral-like double-stranded RNA (dsRNA) which bind to the innate immunity receptors TLR3, RIG-I and MDA-5. It has been used to mimic activation of dendritic cells by viral infections as most of viruses produce dsRNA at some point in their replication. [START_REF] Jacobs | When two strands are better than one: themediators and modulators of the cellular responses to doublestranded RNA[END_REF] CpG, a ligand for TLR9, mimics the presence of bacterial DNA and is also considered as a pathogen-associated molecular pattern that activates DC and B cells in particular. [START_REF] Chaung | CpG oligodeoxynucleotides as DNA adjuvants in vertebrates and their applications in immunotherapy[END_REF] Thus, whether a history of viral and/or bacterial infections could mimic the action of these molecules, and especially the increase expression of interferons, remains an interesting hypothesis. In accordance with this idea, it has been shown that interferon (a and g) levels were significantly higher in patients with chronic hepatitis B infection but also in patients who cleared the virus. [START_REF] Chu | Serum levelsof interferon-alpha and -gamma in acute and chronic hepatitis B virus infection[END_REF] Thus, one can suggest that personal history of hepatitis B infection might naturally boost anti-tumor responses in patients who received anti-HER2 therapy. In addition, over the last few years, an increasing number of works uncovered that innate immune cells (including monocytes, macrophages, and natural killer cells) display prolonged changes in their functionalities after infections or vaccination therapies, a phenomenon called "trained immunity". [START_REF] Netea | Trained immunity: A program of innate immune memory in health and disease[END_REF] According to this idea, innate immune cells could harbor an increased production of cytokines and other inflammatory mediators, including interferons, after recent infections. The increase in innate immune cells responsiveness may potentiate the adjuvant effect and thus the efficiency of anti-tumor targeting therapies.

Oncolytic viruses

Oncolytic viruses have emerged as a novel class of immunebased cancer therapy through their natural preference for cancerous cells and their capacity to trigger lysis of tumor cells as well as activation of anti-tumor immunity. [START_REF] Russell | Oncolytic virotherapy[END_REF] Advances in molecular biology have also allowed the modification of other viruses to make them specific to neoplastic tissues and/or to combine them with immunogenic chemotherapy or immune checkpoint blockers to break tumor-induced immune tolerance. [START_REF] Bell | Viruses for tumor therapy[END_REF][START_REF] Fend | Immune checkpoint blockade, immunogenic chemotherapy or IFN-a Blockade boost the local and abscopal effects of oncolytic virotherapy[END_REF] For instance, recombinant measles viruses have been intravenously injected to treat human patients with bone-marrow cancer. [START_REF] Russell | Remission of disseminated cancer after systemic oncolytic virotherapy[END_REF] As for any non-oncolytic infections, the strength of the immune response to oncolytic viruses may be dependent of previous exposition. For instance, treatment based on the recombinant measles viruses led to a significant resolution of tumor only in the two patients who were measles-seronegative, [START_REF] Russell | Remission of disseminated cancer after systemic oncolytic virotherapy[END_REF] suggesting that absence of prior exposure to measles virus has allowed mounting an immune response strong enough to eliminate cancerous cells. This highlights that personal infection history could be crucial to identify which oncolytic agents can be envisioned in these strategies. In addition, a genetically engineered herpes simplex virus, called T-VEC, has been recently approved to treat advanced melanoma. [START_REF] Ledford | Cancer-fighting viruses near market[END_REF] The treatment is administrated by intra-tumoral injection to seronegative patients. [START_REF] Rehman | Into the clinic: Talimogene laherparepvec (T-VEC), first-in-class intratumoral oncolytic viral therapy[END_REF] However, 67% of the population (0-49 years) had current herpes simplex virus-1 infection in 2012 (WHO). Thus, the proportion of patients who could receive the treatment could be lower than excepted. Such considerations could be applied to other oncolytic therapy such as poxvirus vaccine in the treatment of prostate cancer [START_REF] Kaufman | Phase II randomized study of vaccine treatment of advanced prostate cancer (E7897): A trial of the eastern cooperative oncology group[END_REF] or BCG therapy for bladder cancer. [START_REF] Gontero | The role of bacillus Calmette-Guerin in the treatment of non-muscle-invasive bladder cancer[END_REF] Thus, it suggests the necessity to investigate the potential of less common virus to design new oncolytic therapies.

Prevention strategies

Prevention with infectious vaccines

Several evidences show that vaccination against specific infectious agents could be used to prevent cancer. Protection against melanoma, lymphoma or leukemia has been reported 10 years after BCG, vaccinia or yellow fever vaccination. [START_REF] Villumsen | Risk of lymphoma and leukaemia after bacille Calmette-Guerin and smallpox vaccination: a Danish casecohort study[END_REF][START_REF] Mastrangelo | Does yellow fever 17D vaccine protect against melanoma? Vaccine[END_REF] These findings might be explained by non-specific effects of these vaccines through the shifting of the immune response towards a Th1 profile or through cross reactivity. [START_REF] Grange | Vaccination strategiesto reduce the risk of leukemia and melanoma[END_REF] Indeed, vaccines may contain pathogen antigens with amino-acid sequences that are homologous with those of certain TAAs. [START_REF] Krone | Protection against melanoma by vaccination with Bacille Calmette-Guerin (BCG) and/or vaccinia: An epidemiology-based hypothesis on the nature of a melanoma risk factor and its immunological control[END_REF] By this crossreactive effect, vaccination allows eliminating malignant cells as soon as they expressed the shared-antigens. For instance, a prior immunization with BCG vaccine, which has antigenic similarity with human endogenous retroviruses (HERV-KMEL) expressed in 95% of malignant melanocytes, [START_REF] Schiavetti | A humanendogenous retroviral sequence encoding an antigen recognized on melanoma by cytolytic T lymphocytes[END_REF] has been associated with a better survival in patients with melanoma. [START_REF] K€olmel | Prior immunisation of patients with malignant melanoma with vaccinia or BCG is associated with better survival. An European organization for research and treatment of cancer cohort study on 542 patients[END_REF] Such considerations highlight the importance to consider vaccination as a component of personal history of infection and to study their potential for immunotherapeutic prevention. However, such studies may appear hardly conceivable because of methodological constrains such as cohort size and follow-up duration. Meta-analyses of several cohort studies and systematic reviews should nevertheless be considered to circumvent these limitations.

Preventive cancer vaccines

The potential of TAAs as targets for immunotherapy has already been largely investigated. Recently, it has been suggested that these antigens could be also candidates for preventive vaccines, which could hamper initiation of cancer cell proliferation. [START_REF] Finn | Cancer immunoprevention[END_REF] However, as it seems to be the case for therapy, the response to cancer vaccines could be conditioned by past history of infection. Here, we report examples of interaction between immune responses to infections and to cancer associated antigens.

Mumps and MUC1 vaccine

MUC1 is a normal epithelial cell antigen that was characterized as a TAA due to its abnormal expression in most epithelial adenocarcinomas, and the presence of anti-MUC1 antibodies in patients with cancer has already been associated with a more favorable prognosis. [START_REF] Hamanakai | Circulating anti-MUC1 IGG antibodies as a favorable prognostic factor for pancreatic cancer[END_REF] This suggests that pre-existing antiMUC1 humoral response could help to control tumor growth or even prevent cancer proliferation. Interestingly, it has been described that mumps parotitis causes abnormal MUC1 expression on salivary gland and a negative correlation has early been observed between ovarian cancer and past history of mumps infection. [START_REF] West | Epidemiologic study of malignancies of the ovaries[END_REF][START_REF] Menczer | Possible role of mumpsvirus in the etiology of ovarian cancer[END_REF] Taken together, it has suggested that generation of anti-MUC1 immunity during mumps infection might protect against cancer development. Reinforcing such hypothesis, it has been reported that women who have experienced childhood mumps show a higher proportion of antiMUC1 antibodies which could increase humoral responses and immune recognition of MUC1. [START_REF] Cramer | Mumps and ovarian cancer: moder interpretation of an historic association[END_REF] Accordingly, MUC1 has been studied as a target for antibody-based immunotherapies, some of which have been able to induce the elimination of ovarian cancer cells which express MUC1. [START_REF] Deng | The role of tumourassociated MUC1 in epithelial ovarian cancer metastasis and progression[END_REF] Multiple MUC1 vaccines for treatment of ovarian cancer are now in development and have given significant results. [START_REF] Gray | Progression-free and overall survival in ovarian cancer patients treated with CVac, a mucin 1 dendritic cell therapy in a randomized phase 2 trial[END_REF] The elaboration of prophylactic vaccines has also been proposed and tested in a murine model of colon adenoma where it elicits a long term memory. [START_REF] Kimura | MUC1 vaccine for individuals with advanced adenoma of the colon: A cancer immunoprevention feasibility study[END_REF] However, mumps is a very common childhood disease suggesting that a large proportion of the population could have been exposed. We proposed that the consequences of a pre-existing immune memory developed against the infectious antigens should be investigated in the case of vaccines based on similar antigens. Indeed, it may help to better adapt the number of vaccine boosts which are necessary to reestablish a potent anti-tumor immune environment. [START_REF] Finn | Vaccines for cancer prevention: a practical and feasibleapproach to the cancer epidemic[END_REF]

Varicella and vaccine against cyclin B1

Several tumors express high level of cyclin B1 such as lung, breast and prostate cancers, [START_REF] Finn | Cancer immunoprevention[END_REF] and it has been demonstrated that cyclin B1-specific immune responses may be important in cancer immunosurveillance. Indeed, a prophylactic vaccine against cyclin B1 can delay spontaneous cyclin B1C tumor growth in a murine lymphoma model and increases median survival. [START_REF] Vella | Cyclin B1 vaccine delays spontaneous tumors[END_REF] Interestingly, aberrant expression of cyclin B1-specific antibodies has also been reported in healthy individuals [START_REF] Vella | Healthy individuals have T-cell and antibody responses to the tumor antigen cyclin B1 that when elicited in mice protect from cancer[END_REF] and infections seem to be a reasonably good candidate to explain the presence of cyclin B1-specific responses in healthy individuals. Indeed, a study has shown that cyclins are highly induced in varicella-zoster virus (VZV) infected cells. [START_REF] Leisenfelder | Varicella-zoster virus infection of humanforeskin fibroblast cells results in atypical cyclin expression and cyclin-dependent kinase activity[END_REF] After the primary infection, Varicella-Zoster virus is able to establish a latent infection in the nervous system. [START_REF] Croen | Patterns of gene expression and sites of latency in human nerve ganglia are different for varicella-zoster and herpes simplex viruses[END_REF] The immune memory against the virus can persist for more than 20 years after the primoinfection [START_REF] Arvin | Humoral and cellular immunity to varicellazoster virus: An Overview[END_REF] and immune responses can be subsequently boosted either by endogenous reexposure (silent reactivation of latent virus) or exogenous re-exposure (environmental). [START_REF] Weinberg | Current topics in microbiology and immunology[END_REF] Thus, one can speculate that immune memory against VZV infection could boost immunosurveillance of cancer cells but could also interfere with the efficiency of cycline B1based vaccines. In this context, the number of vaccine boosts should be determined in accordance with past history of VZV infection. [START_REF] Finn | Vaccines for cancer prevention: a practical and feasibleapproach to the cancer epidemic[END_REF]

Personal history of infection and vaccine elaboration

There is a growing body of evidence showing that infections, both in childhood and adulthood, can provide a protection against the risk of developing malignancies. [START_REF] Kleef | Fever, cancer incidenceand spontaneous remissions[END_REF] For example, exposures to childhood diseases have been observed to reduce the risk of developing multiple cancers, including melanoma. [START_REF] Hoption Cann | Acute infections as ameans of cancer prevention: opposing effects to chronic infections?[END_REF] Among childhood diseases, a particular attention has been paid to measles virus. It has been observed that specific MHC class I self peptide (Hsp90b570-578) are expressed both on melanoma cells [START_REF] Jarmalavicius | High immunogenicity of thehuman leukocyte antigen peptidomes of melanoma tumor cells[END_REF] and on measles-infected cells. [START_REF] Herberts | Autoreactivity against induced or upregulated abundant selfpeptides in HLA-A0201 following measles virus infection[END_REF] Thus, the autoreactive CD8 C T cells generated after measles infection could persist and enhance immunosurveillance of cancer cells, giving another example of cross-reactivity. During adulthood, history of acute infections (i.e., common colds and influenza) has been associated with a decreased of cancer risk in controlcase study. [START_REF] Abel | Common infections in the history of cancer patients and controls[END_REF] This has been experimentally confirmed by a study showing that influenza infected mice harbor smaller lung tumor than na€ıve mice. They have also reported the overexpression of 5 TAAs (GAPDH, H4, MDH2, A2) both in infected cells and in tumor cells. [START_REF] Iheagwara | Influenza virus infection elicits protective antibodies and T cells specific for host cell antigens also expressed as tumor associated antigens : A new view of cancer immunosurveillance[END_REF] These evidences suggest that immune memory for antigens produced during influenza or measles infection has allowed the production of long-lasting antibodies against these specific TAAs which could be involved in cancer cell elimination. From these two examples, it seems that therapeutic potential of molecules, such as Hsp90b570-578 and GAPDH, should be investigated. However, studies should not under-estimate the effects of previous natural infections, especially in the case of recurrent infections of fast-evolving virus (i.e., influenza virus), on vaccine efficiency.

Conclusions

Even if accumulating evidence suggests that infections may play a role in cancer risk, [START_REF] Jacqueline | Interactions between immune challenges and cancer cells proliferation: timing does matter![END_REF][START_REF] Jacqueline | Infections and cancer: the "fifty shades of immunity" hypothesis[END_REF] their influence in the responses to immunotherapy has been rarely investigated. Here, we report the evidences for indirect links between personal history of infections and different immunotherapeutic strategies. First, infections could be associated with a modified expression of the therapeutic target, such as increase of PD-1 following acute infections. Second, preexposition to additional infectious agents (as latent EBV infection in the case of EBV-specific CAR T cells) could increase cancer therapy efficiency. Finally, nononcogenic pathogens may induce long-lasting immunity against antigens shared with cancerous cells (i.e., mumps and antigens associated with ovarian cancer) and thus modify immunotherapy efficiency. Some immunotherapeutic strategies already mimic or rely on the effect of infection on immune system. [START_REF] Lichty | Going viral with cancerimmunotherapy[END_REF] It confirms that approaches, linking immunotherapy and personal history of infection, should be developed in a greater extent to consider the huge diversity of infectious agents, chronic but also acute, that may play a role. While the potential interactions of infectious agents with immunotherapies cited here need to be experimentally tested and clarified, identifying the links between personal history of infection and immunotherapy could help clinicians to personalize their immunotherapeutic strategies, improving them or even envisioning new ones. First, it may help to adapt therapies through the determination of an optimal dose of administration. In addition, it may orient the clinicians' choice toward the most adapted strategy, in accordance with the patient's immune profile. Second, it could bring crucial insights to understand the diversity of responses to immunotherapy in cancer patients but also to forecast the apparition of adverse effects which could be crucial to design safe treatments. Third, considering personal history of infection could help to design new oncolytic therapies based on less common viruses as it is expected that they will generate stronger immune responses. Finally, it may allow the elaboration of new preventive measures based on the knowledge of immune mechanisms involved in response to infections. It is worth mentioning that the role of vaccines to induce some of the immune mechanisms described in the context natural infections should be investigated as they represent a good opportunity of prevention. Retrospective analyses of data coming from immunotherapy trials could already give crucial clues to figure out the contribution of personal history of infection and vaccination on the heterogeneity of outcomes observed after the treatments.

Figure 1

Influence of latent EBV infection on treatment with dual-specific CAR T cells. Latent and introduced virus could both be recognized by T-cell receptors and could increase CAR T cell clonal expansion. Thus, in patient with latent viral infection, the use of EBV-specific CAR T cells could result in stronger anti-tumor responses even without introduced EBV. TAA: tumor-associated antigens.

Figure 2

TB -CTLA-4 interaction hypothesis. (A) In patients with BCG vaccination or past history of tuberculosis (TB), CTLA-4 expression is down-regulated. For the same dose of ipilimumab (or other anti-CTLA-4 drugs), these patients will better respond to the treatment as more lymphocytes T (LT) will be targeted. (B) Proposition of clinical trial design to measure the impact of TB pre-exposure or BCG vaccination on treatment efficiency.

Acknowledgments

The authors thank CREEC sponsors CNRS, Labex MabImprove, and Andre HOFFAMNN (Fondation Mava).

Funding

Funding with Agence Nationale pour la Recherche, EVOCAN and STORY.

Competiting interest

The authors declare no competing interests.


Mathieu Lambotte

Stéphane De Cara

Catherine Brocas

Valentin Bellassen

Carbon footprint and economic performance of dairy farms: the case of protected designation of origin dairy farms in France

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

With livestock supply chains accounting for 14.5% of global anthropogenic greenhouse gas (GHG) emissions [START_REF] Gerber | Tackling climate change through livestock -A global assessment of emissions and mitigation opportunities[END_REF], the role of the animal sector is under increasing scrutiny in the climate change debate [START_REF] Herrero | Biomass use, production, feed efficiencies, and greenhouse gas emissions from global livestock systems[END_REF]. In France, meeting the ambitious GHG mitigation targets set by National Low Carbon Strategya reduction of its agricultural GHG emissions by 46% before 2050 (Ministère de la Transition Ecologique et Solidaire, 2018)will require mitigation strategies in the livestock sector.

One major difficulty in reducing livestock-related emissions is that it may severely affect farm income (e.g. Pellerin et al, 2017). This is particularly true in the EU dairy sector, since the abolishment of milk quotas in 2015 has driven milk prices down which threatens the least productive farming systems (Salou et al., 2017b). Moreover, most farmers will only adopt greener farming practices if they do not threaten their profitability [START_REF] Kiefer | The effect of feed demand on greenhouse gas emissions and farm profitability for organic and conventional dairy farms[END_REF]. While the classical economic response to this conundrum would be a tax on GHG emissions, the gilet jaune (Yellow Vests) uprising renders any new environmental tax very unlikely in the near future, which also pleads for addressing the environmental and economic performances simultaneously.

An interrogation that policy makers face and that we analyse is: can dairy farmers reduce GHG emissions while at the same time maintaining profits?

To approach this question in the case of extensive dairy farms, a rich and original dataset is mobilized (more than a thousand technical variables, used for life cycle inventories), with a relatively large sample size (n=95).

Moreover, the farms observed in the dataset are all producing under a Protected Designation of Origin (PDO) label, with specific production constraints, in mountainous areas in eastern France. Thus, they share a homogeneously extensive "production situation", where the bio-physical and socio-economical drivers of the environmental and economic performances are common to all farms [START_REF] Lechenet | Profiling farming management strategies with contrasting pesticide use in France[END_REF]. In a specific production situation, as the external setting of the farms is homogenous, an analysis of the drivers of the performances will isolate the managerial and agricultural practices that explain the difference in performances among the farms, limiting endogeneity issues. Moreover, in French dairy systems, the variability in GHG emissions within each production systemintensive or extensive -is greater than the variations between production systems [START_REF] Gac | Le potentiel d'atténuation des gaz à effet de serre en productions bovines[END_REF]. Thus, there exists a knowledge gap in explaining the variability of the performances of farms sharing the same production conditions. PDO farmers receive a "quality" premium (around 30%) on their milk selling price which enhances their profitability. To receive this premium, they must comply with specific requirements which limit both their production capacity and intensity, to enhance milk quality. These requirements are specifically related to extensive farming practices and could increase the environmental performance of PDO farming: low livestock density, lots of pastures, low use of concentrates, restricted use of fertilizers and so on [START_REF] Hocquette | Indicators of Milk and Beef Quality[END_REF][START_REF] Kop | Origin-based Products: Lessons for pro-poor market development[END_REF].

In this sense, the French government has pointed out the development of PDO farming as a way to achieve a low carbon agriculture while maintaining farmers' profitability (Ministère de la Transition Ecologique et Solidaire, 2018).

Despite large market share of the PDO quality sign in the dairy sector (e.g.

10% in the EU for cheese [START_REF] Chever | Value of production of agricultural products and foodstuffs, wines, aromatised wines and spirits protected by a geographical indication (GI)[END_REF] compared with around 3% for the organic sign in France (Augere-Granier, 2018)), the economic and environmental performances of the PDO dairy sector have never been studied jointly. In the European dairy sector, this joint performance has only been investigated, to the authors' knowledge, in extensive Irish systems [START_REF] O'brien | Relating the carbon footprint of milk from Irish dairy farms to economic performance[END_REF] and intensive Dutch systems [START_REF] Thomassen | Relating life cycle assessment indicators to gross value added for Dutch dairy farms[END_REF]. The question of the relationship between economic and environmental performances has also drawn a lot of interest with the assessment of the cost-effectiveness of mitigation measures, such as reducing stocking rates, nitrogen (N) fertilizers application or imported concentrates [START_REF] Beukes | Improving production efficiency as a strategy to mitigate greenhouse gas emissions on pastoral dairy farms in New Zealand[END_REF]Doole, 2014, for example). Moreover, whether extensive or intensive dairy systems pollute more is still debated and our study sheds some light on this issue, within PDO farms, which are mostly towards the "extensive" end of the spectrum [START_REF] Dollé | Combiner production et environnement, un défi pour la filière laitière Les enjeux environnementaux majeurs Les enjeux relatifs au changement 1-16[END_REF]. Thus, in this paper, we analyze the link between economic performancegross profit per liter of milk produced and per hectareand environmental performance -GHG emissionsincluding or not carbon sequestration, also per liter and per hectare. We go beyond existing literature by:  Quantifying the impact of farms' characteristics or practices on the environmental & economic performances of PDO farms simultaneously.

 Using a large sample size within a homogeneous production situation (PDO farms in mountainous Eastern France) which allows us to focus on the role of management practices.

 Designing and implementing a novel and simple approach to account for carbon sequestration related to land-use and land management changes in a net GHG emissions indicator for environmental performance.

 Outlining a lead that may reconcile the contradictory results on the relative merit of extensive and intensive systems with regards to climate mitigation: we confirm that more extensive systems perform better with a higher share of grass, possibly because grass is more expertly managed in extensive systems (e.g. through proper drying) than at the intensive end of the spectrum.

Methods and Data

Population characterization and notation

Our main data source is the field survey of 95 PDO farms in the Franche-Comté and Savoy regions, financed by the PDO consortia between 2013 and 2015 [START_REF] Michaud | Mise en place d'une démarche d'évaluation et d'amélioration du bilan carbone des exploitations agricoles[END_REF][START_REF] Perrard | Mise en place d'une démarche d'évaluation et d'amélioration du bilan carbone des exploitations agricoles[END_REF]. These surveys gather all the necessary technical and managerial information that is used to compute GHG emissions via the CAP'2ER Life Cycle Analysis (LCA) tool. These surveys also provide detailed information on farmers' practices and farms' characteristics, such as the farm's and herd's sizes, the amount of concentrate feed used, the cereals produced and used on farm, the fertilizer use or the labor use. The average farm of our sample has 125 (σ = 79) ha and 92 (σ = 51) cows, produces 348,158 (σ = 231,096) liters of milk per year, which amount to a productivity of 3,792 (σ = 758) liters per cow and 2,773 (σ = 1,205) liters per ha. The detailed descriptive statistics of our sample are provided in SM 2. In addition, PDO farms generally have Montbéliarde cows, fed mostly with grass and hay from set stocked pastures. The cows spend on average 208 days per year on pastures and otherwise are kept in barns with free stalls. The manure is usually not composted and stored in manure pit at least a week before being spread on the fields using a liquid manure tank. The farms are located in mountainous areas and do not use irrigation.

Consider a population of farms (indexed by

). Each farm is characterized by a matrix of outputs (e.g. liters of milk produced ( ), cereals and cows sold…) produced by combining two quasi-fixed inputs (land ( and herd size) and a matrix of variable inputs (e.g. fertilizer, concentrates, fuel…).

Denote by the gross profit, defined as where is a matrix of output prices and a matrix of input prices.

Moreover, each farm emits an amount of GHG as a negative externality of its production activity. As cropland and pastures can also sequester carbon in the soils, each farm sequesters an amount of carbon. Thus, each farm has a gross GHG emission amount and a net one, .

To measure the economic performance, we consider two indicators, the gross profit per liter of milk (fat-and-protein corrected, with 40g/kg and 33g/kg respectively) produced (variable output) and per hectare (fixed input), .

As indicators of the environmental performance we use the opposite of gross and net GHG emission per liter (fat-and-protein corrected) and per hectare, , , , respectively. We use both a productbased and an area-based indicator to account for two diverging hypotheses on the elasticities of demand. Indeed, if demand is infinitely elastic or if there is no substitute for PDO products, consumers will fully adjust to any change in the quantity produced and the product-based indicators are irrelevant. To the contrary, if demand is inelastic or if standard products are perfect substitutes for PDO products, a reduced production in the PDO area will be offset by an increase in production elsewhere, diminishing the relevance of area-based indicators.

In sum, the variables of interest are and the set of indicators .

Economic performance estimation

The gross margin is defined in this study as the difference between the farm's revenue and its costs, without accounting for taxes or subventions.

The former includes the revenues from the sale of the farm's outputs :

PDO milk, animals, cereals and roughage. Factor costs include the buying costs of the farm's inputs : forage, concentrates, fertilizer, electricity and fuel, contracted work and animals for the renewal of the herd. Family labor costs are valued at the average wage of paid labor (€20,965 per year).

To estimate the gross margin of each farm, these physical flows need to be multiplied by prices. The prices of most inputs and outputs are estimated using the FADN average for the corresponding year and the corresponding NUTS2 region, with the following exceptions:

 Since the FADN does not identify whether a farm is PDO certified, the price of PDO milk for each year and each PDO area comes from the PDO unions (Agreste Bourgogne-France Comté, 2015; Les fromages de Savoie, 2017).



The prices of fertilizers and concentrates, which cannot be derived directly from the FADN, are obtained from Eurostat (2018).



The buying and selling prices of dairy cows, cull cows and heifers is gathered from the Ministère de l'Agriculture et de l'Alimentation.

To test the robustness of this estimation, the average estimated profit is compared to the average reported profit for dairy farms in the Franche-Comté and Rhône-Alpes NUTS2 regions from FADN.

Estimation of the environmental performance

To assess the environmental performance, we focus on GHG emissions for two reasons: first because climate change is arguably one of the most pressing environmental challenge of the 21 st century and second because GHG emissions are correlated with environmental impacts such as eutrophication, acidification and energy use [START_REF] Guerci | Parameters affecting the environmental impact of a range of dairy farming systems in Denmark, Germany and Italy[END_REF]. Gross GHG emissions -without carbon emissions/sequestrations related to land use and management -are computed using CAP'2ER, a GHG emissions calculator developed by the Institut de l'Elevage and following Life Cycle Assessment (LCA) guidelines (Institut de L'Elevage, 2013). The system boundaries are therefore "cradle-to-farm gate", including enteric fermentation, manure management, fertilizers, fuel and energy use, but also the GHG emissions due to the production of concentrate feed and fertilizers.

Contrary to the default "energetic allocation" of CAP'2ER, these emissions are then allocated to the three products of farmsmilk, meat and cropsin proportion of the share of each product type in the farm revenue [START_REF] Baldini | A critical review of the recent evolution of Life Cycle Assessment applied to milk production[END_REF].

To estimate land-use related carbon sequestration (C i ), we also deviate from 2.09 t CO 2 eq ha -1 yr -1has been criticized as being much too large to be consistent with the current knowledge about carbon fluxes and stocks in grassland [START_REF] Smith | Do grasslands act as a perpetual sink for carbon?[END_REF].

In order to bridge this pitfall and provide a more robust estimate of landrelated GHG emissions, we develop an innovative methodology based on land use and land management changes. The land use and land management of each farm in our sample is compared to a reference, average farm. We then estimate the carbon fluxes which are being avoided by the choice of each farm to maintain its observed land use rather than transitioning towards the land use of the reference farm.

The share of land uses (cropland vs permanent grassland) in our reference farm is set to the sample average (82% permanent pasture, 18% temporary pasture and cropland). Note that the choice of the reference farm does not impact our results on the differences in environmental performance within the sample. Carbon fluxes (sequestration or emission) associated to each type of land-use changes include both the actual flux resulting from the change and the alteration of future carbon fluxes implied by the change. For example, a farm which has 100% of pasture on 100 ha of total land is estimated to sequester 3.72 t CO 2 eq. ha -1 .yr -1 on the 7 hectares which could have been converted to cropland to match the reference farm. The actual values and their sources are detailed in section 4.

Such an estimate is akin to direct LUC (dLUC) as defined by [START_REF] Herrero | Biomass use, production, feed efficiencies, and greenhouse gas emissions from global livestock systems[END_REF]. Indirect LUC (iLUC) is a more controversial topic and its estimates are laden with high uncertainties. Nevertheless, we attempt to provide an upper estimate of it in the context of French PDO farms. In our case, iLUC could occur because one hectare of cropland generally produces a higher nutritive capacity calorie-wise than grassland in our sample. Thus, assuming an inelastic demand, a farmer who converted some cropland into grassland would have to import feed to continue feeding the same herd. To produce this additional feed, either non-agricultural land is put into production (extensive margin) or the current production processes are intensified (intensive margin). We retain the extensive margin effect, and our study area being located in the Jura and Alps, non-agricultural land is most likely forest land. The combination of these two key hypothesizesinelastic demand and extensive marginyields an upper bound for the area estimate of iLUC. As such, they are not included in the indicator retained for "net environmental performance" and are only used as a robustness check (SM 10).

Using the formalization of [START_REF] Plevin | Greenhouse gas emissions from biofuels' indirect land use change are uncertain but may be much greater than previously estimated[END_REF], our reduced-form models of carbon sequestration for dLUC and iLUC are therefore expressed in equations 1 and 2.

(

(2)

Where (see SM 1 for demonstration).

To compute the land-use related emissions ( ), we use the parameters presented in Table 1.

Table 1. Specification of the carbon sequestration methods

Emission factor cropland to grassland (dEmissionFactor) -74.3 t CO2eq.ha -1 Source: [START_REF] Efese | La séquestration de carbon par les écosystèmes en France[END_REF].

Emission factor forest to cropland (iEmissionFactor)

749.4 t CO2eq.ha -1 Source: [START_REF] Efese | La séquestration de carbon par les écosystèmes en France[END_REF].

Nutritious content (Nutri)

Nutri C = 3840 kcal.

Nutri G = 4010 kcal. In addition to the estimation of carbon sequestration from dLUC and iLUC, our method allows the estimation of the impacts of some management practices on biomass and soil carbon. Based on a recent review in France [START_REF] Pellerin | Stocker du carbone dans les sols français , Quel potentiel au regard de l'objectif 4 pour 1000 et à quel coût? Synthèse du Rapp[END_REF], we identify three practices that are relevant in PDO dairy farming and that change biomass and soil carbon stocks: the share of temporary grasslands in crops rotation, the amount of nitrogen (mineral or organic) fertilization in pastures and the amount of hedges. The carbon impact of these practices follows a temporal pattern similar to the carbon impact of LUC: a change in practice leads to carbon sequestration or emissions which saturate over time as soil and biomass carbon reach a new steady-state equilibrium. Similar to our LUC model, only the differences from the reference farm are therefore considered. [START_REF] Pellerin | Stocker du carbone dans les sols français , Quel potentiel au regard de l'objectif 4 pour 1000 et à quel coût? Synthèse du Rapp[END_REF] estimates that on average 63.7 linear meters of hedges sequesters 259 kg C.ha -1 .yr -1 in the soil and biomass on cropland and 242 kg C.ha -1 .yr -1 on pasture. Here, a linear meters of hedge is associated to 2 square meters of hedge and 1.5 square meters of uncultivated area both side of the hedge. As our dataset only contains the cumulative length of hedges for each farm, we allocate these hedges proportionally to grassland and cropland, based on the land-use of each farm. Emissions or sequestration are then added to the carbon budget of each farm based on the difference with the reference farm for both the amount of hedges in grassland and the amount of hedges in cropland.

Nitrogen fertilisation on pasture stimulates the biomass growth and thus soil carbon sequestration. Several reviews conclude an almost linear relationship between nitrogen and carbon sequestration in grasslands, with an average ratio of 1.2 kg C per kg N [START_REF] Eze | Soil organic carbon stock in grasslands: Effects of inorganic fertilizers, liming and grazing in different climate settings[END_REF][START_REF] Fornara | Soil carbon sequestration in prairie grasslands increased by chronic nitrogen addition[END_REF][START_REF] Pellerin | Stocker du carbone dans les sols français , Quel potentiel au regard de l'objectif 4 pour 1000 et à quel coût? Synthèse du Rapp[END_REF]. Here again, differences in nitrogen fertilizationboth mineral and organicwith the reference farm are translated into carbon emissions or sequestration, using the average ratio above.

The share of temporary pasture in rotation with crops also improves carbon sequestration in soil. For France, [START_REF] Pellerin | Stocker du carbone dans les sols français , Quel potentiel au regard de l'objectif 4 pour 1000 et à quel coût? Synthèse du Rapp[END_REF] estimate that including 50% of temporary pasture in rotation with crops, compared to crops only, sequesters an additional 466 kgC.ha -1 .yr -1 . More generally, the relationship between the annual increase of SOC and the share of temporary pasture in the rotation follow a linear pattern from rotations dominated by crop (0% of grass) to rotation dominated by grassland (100% of grass) [START_REF] Vertès | Modelling the long term SOM dynamics in fodder rotations with a variable part of grassland[END_REF]. Accordingly, we assume that soil carbon sequestration and the share of temporary pasture in the rotation are positively and linearly correlated. To be consistent with our LUC estimates, temporary grassland is therefore assumed to increase carbon sequestration by 37.15 kgCO2e/% of temporary grassland/year. For example, as temporary grasslands represent 71% of the UAA (excluding permanent grassland) in the reference farm, a farm with no temporary grassland would be estimated to be emitting 2.6 tCO2e.yr -1 .ha -1 of UUA excluding permanent pasture.

The results based on this estimation of GHGE including impacts of management practices on carbon sequestration are however only used as robustness check because of multicollinearity issues (SM 9).

Econometric analysis on the whole sample

We aim at identifying the practices which create synergies between economic and environmental performances i.e. that influence in the same direction both performances. The annual variation of weather, production or prices, that can impact the environmental or economic performances, is accounted for by using year dummies and pedo-climatic variables (slope, temperature, rainfall, type of soil).

We use six separate Ordinary Least Squares (OLS) regression models (Model 1 to 6), with each of the indicators in the set being the dependent variable of a model. As independent variables, we use farms' characteristics and practices, described in SM 2. The 6 separate regression equations following the classical linear form: The regression coefficients are compared to detect the explanatory variables that affect in the same direction both the environmental and economic performances (synergies). To identify practices which have an important effect on the performances, we calculate the effect size as the product of the difference between the first and third quartiles in Xto capture the actual variability in the samplewith the associated regression coefficients. Then, we divide these effect sizes by the average performance in the sample to obtain a relative effect size. For a given practice, this quantifies by how much the environmental or economic performanceper liter or hectare -could be increased if the median farm in the worst half of the sample would adopt the same practice as the median farm in the best half. All the statistical analysis is performed using R language (R Core Team, 2020) and the data visualization is done with the ggplot2 [START_REF] Wickham | ggplot2: Elegant Graphics for Data Analysis[END_REF] and corrplot [START_REF] Wei | R package "corrplot": Visualization of a Correlation Matrix[END_REF] packages. The whisker boxes represent the average, first and third quartiles, and minimum and maximum

Results

3

Gross profit per liter averages at €0.34 per liter and is higher than the FADN average for PDO, primarily because of lower costs. Indeed, concentrates costs may be underestimated in our estimation: it is one of the few cost categories for which we use prices from Eurostats (2018), as the FADN does not provide detailed prices for the concentrates purchased. These national prices underestimate this type of costs for PDO farms which are subject to specific constraints (many feed types are forbidden, local production of concentrates is mandatory, …). Otherwise, the higher revenues and profits of PDO farms is confirmed.

Note that the standard deviation of our two economic indicators, profit per liter and per hectare, is large: 32% and 49% respectively (SM 2). This important variability is promising for the econometrical analysis. The results are similar when the environmental performance is measured per liter, except that gross GHGE does not vary strongly with farming intensity (Figure 3). 3.2.2. No synergetic practice but many levers on either the economic or environmental performance.

No synergetic farming practice could be identified for the economic nor environmental performances: no variable with a significant regression coefficient for economic performance has a significant effect of the same sign on environmental performance and vice-versa (Figure 5, Table 2).

Trade-offs are also scarce: only the organic and mineral N spread on pastures improve environmental performance per hectare at the expense of economic performance per hectare. Several levers are however identified, which may improve either the environmental or economic performances by 7 to 21% without deteriorating the other. Although these alternative specifications are sometimes useful in interpreting the results, none of them trigger major changes in the estimators or their significance. A notable exception is the inclusion of our higher-end estimate of iLUC which turns the amount of organic N on pasture into a significant positive lever on net environmental performance per liter.

Another exception is the inclusion of the impacts of management practices on carbon sequestration, which turns the labor use per cow into a significant and negative lever of the net environmental performance per liter and the age of first calving into a significant and negative lever of the net environmental performance per hectare.

The colored bars represent the relative effect sizes on the performance and the black lines the relative confidence intervals of the coefficients.

Impacts of levers on the environmental performance per liter

If the farmers of the quartile using the least electricity per cow could upgrade their hay drying equipment and reach the environmental performance of the upper quartile on electricity consumption, they would decrease their net GHG emissions by 0.08±0.07kg CO 2 eq.L -1 (net environmental performance increased by 7.1% of total sample average) without any significant profitability change (Figure 6).

Similarly, farmers at the upper quartile of organic N on pasture would decrease their gross GHG emissions by 0.14±0.12 kg CO 2 eq.L -1 (12.3% of Figure 7. Relative effect sizes of selected practices on the net environmental and economic performances per hectare total sample average) if they could move down to the lower quartile (Figure 6).

If farmers would stop the practice manure composting, they would decrease their net GHG emissions by 0.14±0.13 kg CO 2 eq.L -1 (13% of total sample average) (Figure 6).

Impacts of levers on the economic performance per liter

Reducing the labor use per cow, the share of protein in cows' diet and the ecological focus area could increase the economic performance of the highest half of the sample by €0.07±€0.02. L -1 , €0.03±€0.02. L -1 and €0.02±€0.02. L -1 respectively (21%, 9.2% and 7.2% of total sample average), all without any significant environmental damage (Figure 6).

Impacts of levers on the environmental performance per hectare

As expected, reducing fuel use per hectare could increase the gross environmental performance of the highest half of the sample by 208±153 kg CO 2 eq.ha -1 (5.9% of total sample average) (Figure 7).

More interestingly, reducing the share of manure in organic fertilizers could increase the gross environmental performance of the highest half of the sample by 262±248 kg CO 2 eq.ha -1 (7.4% of total sample average) (Figure 7).

Impacts of levers on the economic performance per hectare

Reducing labor use per cow, the share of protein in cows' diet and the Ecological Focus Area of the highest half of the sample could increase their economic performance by €174±€65.ha -1 , €86±€68.ha -1 and €120±€95.ha -1 respectively (17.5%, 8.6% and 12% of total sample average) (Figure 7).

Impacts of trade-offs on the performances per hectare

Reducing mineral and organic N spread on pastures in the highest half of the sample would increase their gross environmental performance by 419±182 kg CO 2 eq.ha -1 and 1178±198 kg CO 2 eq.ha -1 respectively (11.8% and 33.3% of total sample average) but decrease their economic performance by €143±€96.ha -1 and €277±104.ha -1 respectively (14.4% and 27.8% of total sample average) (Figure 7).

Discussion

4.1. Possible levers for performance improvement: tillage, logistics, milking equipment and labor efficiency

The econometric analysis shows that 6% can be gained on the environmental side by reducing fuel use without impairing economic performance. This possibility likely emerges from the potential to increase the share of grazed pasture (rather than mowed pasture): the share of grazing in the diet is negatively correlated with fuel use and when an interaction between the two is added in the regression model, its estimator is negative (although not significant, see SM 11). Another possible practice allowing the reduction of fuel use is the optimization of logistics, although this possibility may be constrained by the spatial distribution of fields and their distance from stables.

Conversely, a higher electricity use per cow increases the environmental performance per liter by 7% without decreasing the economic one. Indeed, electricity production results in little emissions in France (nuclear energy).

Moreover, electricity use is mainly linked to milking equipment and the drying of hay, both of which increase milk production. Indeed, the share of hay is positively correlated to the electricity use and the estimator of the interaction is positive although not significant (SM 11).

Manure composting deteriorates the environmental performances per liter by 13%: most farm composters are not equipped to capture or flare methane, thus releasing considerable amounts of it during composting [START_REF] Hao | Carbon, Nitrogen Balances and Greenhouse Gas Emission during Cattle Feedlot Manure Composting[END_REF].

The share of protein in the diet largely reduces the economic performance, both per liter (21%) and per hectare (9%). Soy-based concentrates are indeed costlier and therefore do not seem to proportionally increase cow productivity. This practice is positively correlated with the amount of concentrates fed to the cows and thus decreases the environmental performance per ha, as the GHG emissions from the production and transportation costs of the concentrates are accounted for in our analysis.

Labor efficiency is also an avenue worth exploring to substantially improve economic performances per liter and per hectare without impairing environmental performance. Indeed, labor costs weight 53% of total costs, as PDO dairy farming is a labor-intensive technology (Bouamra-Mechemache and Chaaban, 2010). This lever seems partly related to economies of scale as our alternative models with variable selection mostly remove farm size from the set of dependent variables (SM 12). Natural constraints also play a role: labor intensity is correlated with steeper slope, scarcer rainfall and lower temperature.

Lastly, the positive influence of nitrogen on profit shows that PDO farms are not wasting nitrogen on pasture. However, the use of mineral and organic nitrogen on pasture is detrimental to environmental performance.

Moreover, testing for an interaction between the amount of mineral and organic N spread on pasture reveals a negative and significant interaction effect on the environmental and economic performances. This indicates a potential synergy: where organic N fertilization is already high, reducing mineral fertilization would simultaneously increase both performances.

The share of grass in the diet, and in particular of grazing, is paramount in the technical specifications of these PDOs, but also in other quality signs such as organic farming. Here we do not identify these as important levers, neither for economic performance nor for environmental performance. This may be due to a rather small variance in these variables because all our farms follow the PDO specifications or to their correlation with fuel and electricity uses: the share of grass and hay in the diet are mostly excluded from the variables selection procedure, while the fuel and electricity uses are kept (SM 11).

Correlation between environmental and economic performances

The negative correlation between the environmental and economic performances per hectare is partly due to the intensification of farming practices: when production is intensified per unit of land, more feed, enteric fermentation and manure are taking place in the same area.

When the performances are measured per liter, we find a weak positive correlation between gross environmental and economic performances (ρ = In the case of intensive Dutch farms, [START_REF] Thomassen | Relating life cycle assessment indicators to gross value added for Dutch dairy farms[END_REF] show that a high share of concentrate feed in cows' diet results in lower GHG emissions per liter thanks to higher milk productivity and lower emissions per unit of feed [START_REF] Liang | Optimizing productivity, herd structure, environmental performance, and profitability of dairy cattle herds[END_REF][START_REF] Lovett | A systems approach to quantify greenhouse gas fluxes from pastoral dairy production as affected by management regime[END_REF]. However, gross margin per liter is also reduced because of feed costs. Hence its conclusion is that environmental performance cannot be enhanced without decreasing farms' profitability.

Our results lie somewhat in between: similarly to O'Brien (2015), we find that concentrates may be overused in the sense that their reduction improves the economic performance in our sample of extensive farms. However, the environmental benefit is not sufficient to suggest a synergy when economic and environmental performances are expressed per liter.

We think that farmer know how in the grass management may provide the key to reconcile these contradictory results. Indeed, mowed grass tends to lose rapidly its nutritious content. The antagonism identified in Thomassen et al. ( 2009) may be explained by the limited presence of grazing in their sample farms, associated with a limited farmer know-how on grass management. In this context, a higher use of concentrates can be an effective way to reduce GHG emissions by lowering enteric fermentation [START_REF] Lovett | Greenhouse gas emissions from pastoral based dairying systems: The effect of uncertainty and management change under two contrasting production systems[END_REF] and to increase profitability by rising the cows' productivity [START_REF] Thomassen | Relating life cycle assessment indicators to gross value added for Dutch dairy farms[END_REF]. But, as our study and the Irish case demonstrate, farms with high shares of pastures tend to create a synergy between environmental and economic performances as increasing the grass in the cows' diet can improve the digestibility of the forage and thus reduce the enteric fermentation and the CH4 emission [START_REF] Dillon | The effect of forage type and level of concentrate supplementation on the performance of spring-calving dairy cows in early lactation[END_REF], especially if the cut grass is harvested in an early maturity stage [START_REF] Van Middelaar | Cost-effectiveness of feeding strategies to reduce greenhouse gas emissions from dairy farming[END_REF]. The positive influence of hay drying equipment and positivealthough not significanteffect of the square of the share of grass on the gross environmental performance per liter are consistent with this interpretation (SM 10): the grass management know-hows of extensive farmers allow them to increase their environmental performance with a higher share of grassland while intensive farmers would suffer from a degraded digestibility of grass when their share of grassland increases.

Ultimately however, all these results rely on parameters choices for the digestibility of feed which are known to be very uncertain (IPCC, 2019). Producing electricity with nuclear energy does not emit GHG whereas coal does, even if nuclear energy creates wastes that impact the environment but not through global warming.

In this debate, the originality of our study is to propose another statistical approach to this question and another method for the carbon sequestration, as well as using both product-based and area-based indicators. We find that the amount of concentrate only has a significant negative influence on the Moreover, proposing several indicator of the environmental performances (gross GHGE, net GHGE and iLUC GHGE) increases the validity of the results, as including carbon sequestration and how management practices impact it as well as indirect land-use changes can by itself give the advantage to either intensive or extensive dairy farming as the most environmentally performant system [START_REF] Meier | Environmental impacts of organic and conventional agricultural products -Are the differences captured by life cycle assessment?[END_REF].

Omitted variables bias

The main methodological limit in this study is related to the econometric models. Some important variables are likely to have been omitted, at least in the models with low adjusted r-square. Classical omitted variables, such as farmer's dynamism or competence, could be correlated with both the dependent variables and the practices, biasing the estimators (endogeneity).

However, such omitted variable bias is limited: the heterogeneity of these classical omitted variables is likely to be limited in our sample (same production situation, same region, all PDO farms included in the same farmer association, …). However, we cannot fully rule out and the causality of the relationships we identify must be carefully pondered. Other methods, such as farm system modelling or Data Envelopment Analysis can also successfully identify mitigation practices that increase the economic or environmental performances of dairy farms and that are similar to the levers discussed above [START_REF] Beukes | Improving production efficiency as a strategy to mitigate greenhouse gas emissions on pastoral dairy farms in New Zealand[END_REF][START_REF] Doole | Least-cost greenhouse gas mitigation on New Zealand dairy farms[END_REF][START_REF] Iribarren | Benchmarking environmental and operational parameters through eco-efficiency criteria for dairy farms[END_REF].

Conclusion

Our regression models question the possibility of synergies between drivers of economic and environmental performance, but also the existence of necessary trade-offs. We identify however several levers: investing in milking equipment and hay drying equipment, reducing the livestock density, abandoning manure composting or optimizing fuel use increase the environmental performance by 5 to 13% without impairing gross margins, while increasing labor productivity and reducing the share of protein in the diet enhance the economic performance by 7 to 21% without increasing GHG emissions.

Our results also bring new insights on the debated merits of extensive milk farming, suggesting that concentrate use is detrimental to both economic and environmental performance as long as grass retains its nutritious content, for example via grazing. This would be worth confirming with a similar analysis on a sample containing both extensive and intensive dairy farms.

We also develop a novel and simple methodology for the estimation of landuse related emissions and sequestration based on potential land-use changes compared with a reference farm. By doing so, we provide new information on the sustainability of specific practices and a complete methodology that could be used in further studies on environmental and economic performances.

Beyond the methodological limit posed by a possible, although likely moderate, omitted variable bias, the main limit of this paper comes from the restricted study region and the possible sample selection. Indeed, we study the performances of the PDO farms among the same region and using only PDO farms in our statistical population. While this can be beneficial to limit endogeneity, as we can compare farms that share a similar production situation, it limits the validity of any comparison with conventional dairy farming or PDO farming in other areas. Thus, the research on PDO farming and sustainable practices in agriculture could be improved by an analysis that would compare PDO farming in different countries or production situations. Reproducing our analysis for the conventional dairy sector in

France and comparing the results could help determine if PDO dairy farming is more economically and environmentally performant, so more sustainable, than the conventional one. Furthermore, the levers of the performances that we uncover in this paper could be compared to the ones in the conventional dairy sector.

  CAP'2ER for two main reasons. Firstly, CAP'2ER attributes carbon sequestration to static land managementsuch as permanent pasturewhereas the only stabilized results for cropland and grassland related carbon fluxes in the literature concern land-use changes (LUC). Indeed, the latest IPCC guidelines (IPCC, 2019) estimate carbon fluxes to be null for croplands and grasslands which did not undergo recent land use or management changes. Secondly because the sequestration factor used by CAP'2ER for permanent grassland derived from Soussana et al. (2010) -

  one of the above 6 measures of performance for each farm, is a [ ] matrix of regression coefficients, different for each of the 6 models, is a [ ] matrix, similar for each equation (SM 2) and is a [ ] matrix of error terms, with being the sample size and the number of parameters.

. 1 .

 1 Economic and environmental performances of PDO farms 3.1.1. Economic performance of PDO farmsThe average estimated farm revenue in our sample is €210,813 and the average total factor cost amounts to €83,538. The average gross margin is thus €127,274. The averaged reconstituted revenue, cost and profit per liter are comparable to FADN averages for PDO farms in the same regions (Figure1).

Figure 1 .

 1 Figure 1. Comparison of the distribution of the estimated economic performance per liter and theFADN's value (2013FADN's value ( -2015) )

3. 1 . 2 .

 12 Impacts of stocking rate and system boundaries on GHGETo illustrate the results of the theoretical LUC model, the GHGE are computed for each indicator, both harmonized per liter and hectare, and presented depending of the quartile of stocking rates, to represent the variation of farming intensity in the sample. When the GHGE are measured per hectare, the most extensive farms emit less, except when iLUC are accounted for. The difference of GHGE between the most extensive and intensive farms becomes larger with the increasing comprehensiveness of the LCA perimeter, until iLUC are included (Figure2). Indeed, when iLUC are accounted for in the LCA, the GHGE of extensive farms is higher than intensive farms' ones, because the difference in nutritive capacity between maize and grass is high in PDO farms and thus the iLUC effects attributed to extensive farms are large.

Figure 2 .

 2 Figure 2. Carbon footprint of indicators per hectare-with different LCA perimetersper stocking rate quartile

Figure 3 . 1 .

 31 Figure 3. Carbon footprint of indicators per liter-with different LCA perimetersper stocking rate quartile

Figure 4 .

 4 Figure 4. Correlation of environmental and economic performances

Figure

  Figure 5. Synergies, levers and antagonisms in economic and environmental

  distribution of the residuals. To further asses the linearity of the relationships between the indicators of performances and the farms' inputs, a general additive model specification was tested but produced lower R² (coefficient of determination). With the exception of organic N spread on pastures, whose positive coefficients for models 1,2 and 6 saturate after 120 N unit per ha, no other input shows nonlinear effects (labor per cow, concentrates, N, P and K spread on cereals or pastures). Several alternative indicators have been attempted to test the robustness of these results such as allocating all GHG emissions to milk production or restricting the perimeter of GHG emissions to the farms by ignoring emissions from the production and transportation of concentrates and fertilizers (SM 7 & SM 8). Indicators including the impacts of several management practices on carbon sequestration in the farms' GHGE have been estimated (SM9). Similarly, indicators including an upper bound estimate of indirect land-use changes are summarized in SM 10. Alternative specifications, with interaction effects (SM 11) or variable selection (SM 12) have also been tested.

Figure 6 .

 6 Figure 6. Relative effect sizes of selected practices on the net environmental and economic performances per liter

0

  .18) but a strong and negative correlation between the net environmental and economic performances (ρ = -0.33).[START_REF] O'brien | Relating the carbon footprint of milk from Irish dairy farms to economic performance[END_REF], who only use per liter indicators, finds a positive correlation between economic and gross/net environmental performances (ρ = 0.3 to 0.5). This may be explained by the difference in carbon sequestration estimation method. Indeed, O'Brien et al. (2015) uses a sequestration factor of 1.36 t of CO 2 eq per ha of grassland and per year based on[START_REF] Soussana | Mitigating the greenhouse gas balance of ruminant production systems through carbon sequestration in grasslands[END_REF], which overestimates carbon sequestration as discussed in section 2.3.[START_REF] Thomassen | Relating life cycle assessment indicators to gross value added for Dutch dairy farms[END_REF] however find a negative correlation between the gross environmental and economic performances per liter (ρ = -0.31), in the case of intensive farms.In Italy,[START_REF] Fiore | GHG and cattle farming: CO-assessing the emissions and economic performances in Italy[END_REF] choose to cluster farms by their environmental performance (GHG emissions) and finds 3 clusters, with an antagonism between environmental and economic performances in each cluster. 4.3. Diverging results on the effects of farms' characteristics and practices on the performances In the case of extensive Irish farms (O'Brien et al. (2015)), the length of the grazing season is the most important lever on both the environmental and economic performances, i.e. creates a synergy. The conclusions drawn are that extensive livestock farming, limiting concentrate feed (which has a negative influence on both performances in their study) and better valorizing pastures and meadows can outperform more intensive systems[START_REF] Ledgard | Temporal, spatial, and management variability in the carbon footprint of New Zealand milk[END_REF], mainly because pastures imply carbon sequestration in soils. We verify these results, even when carbon sequestration is not accounted for (gross vs net GHG emissions), or integrates indirect land use changes (SM 9). Moreover, because both the length of the grazing season and the yield of milk per hectare or per cow are negatively correlated with GHG emissions per liter,[START_REF] O'brien | Relating the carbon footprint of milk from Irish dairy farms to economic performance[END_REF] show that extensive diets can also result in low carbon footprints. At the same time, by reducing feed costs, extensive grazing can reduce the farms' costs and thus extend their margins.

Kiefer,

  [START_REF] Kiefer | The effect of feed demand on greenhouse gas emissions and farm profitability for organic and conventional dairy farms[END_REF] compare organic and conventional dairy farms in Germany and also find that limiting concentrates use reduces GHG emissions and increases profitability. Similarly, Thomassen, van Calker, Smits, Iepema and de Boer (2008) recommend to decrease concentrate use per kilogram of milk, especially concentrates with a high environmental impacts (soy). Moreover, Arsenault, Tyedmers and Fredeen (2009) find that the high concentrates use, fuel use and N fertilizers are the main drivers of environmental impacts in Canadian dairy farms. In their study, electricity is also an important contributor to GHG emissions, but our diverging results are straightforwardly explained by the sources of electricity: mainly nuclear energy in our French, context versus 75% of coal in Nova Scotia (Canada).

  Note: *** p < 0.001, ** p < 0.01, * p < 0.05, standard deviations of the coefficients are included between brackets. Only variables which have a significant and large impact on indicators are represented (p.value <5% and relative effect size > 5%) The full table, including pedo-climatic variables and year dummies is provided in SM 4.

	400		A greenup arrow indicates an improvement of the indicator whereas
	401		a reddown arrow indicates a deterioration of the indicator. Only
	402		variables which have a significant and large impact on indicators
	403		are represented (p.value <5% and relative effect size > 5%). In the
	404		case of trade-offs, the first arrow always represent the impact on the
	405	environmental performances. The independent variables retained in our models explain most of the
	406	variance of the indicators expressed on a per hectare basis, but a smaller
		share of per liter indicators (Table 2, SM 4). This could be expected as more
	407	Table 2. Selected results of the OLS models independent variables are expressed on a per hectare basis. Residuals vs
		fitted values plots do not indicate heteroscedasticity of the residuals or non-
		Net Environmental performance per L (1) linear relationships between the variables, even if some outliers can be Gross Environmental performance per L (2) Economic performance per L (3) Net Environmental performance per Ha (4) Gross Environmental performance per Ha (5) detected (SM 6). Shapiro-Wilk tests successfully assert the normality of the	Economic performance per Ha (6)
	Labor Use per cow	5. Synergies, levers and antagonisms in economic and environmental -5.00 -1.04 -6.08 *** 1,745.81 6,819.09 (3.38) (2.24) (0.70) (11,773.86) (5,484.13)	-15,319.7 *** (2,878.36)
	Fuel per Ha	performance -0.002 * (0.001)	-0.001 (0.001)	0.0002 (0.0002)		-4.31 (4.15)	-5.22 *** (1.93)	0.61 (1.01)
	Electricity per cow Concentrate per	0.0004 ** (0.0002) -0.0000	0.0003 ** (0.0001) 0.0000	Indicators per liter -0.0000 (0.0000) -0.0000		-0.03 (0.67) -0.14	-0.54 * (0.31) Indicators per Ha -0.42 *	0.17 (0.16) 0.02
	cow		(0.0001)	(0.0001)	(0.0000)		(0.46)	(0.21)	(0.11)
	Synergy Share protein in	3.50	-1.91	-2.91 ***		2,058.35	-5,106.24	-8,415.21 **
	the diet		(3.99)	(2.64)	(0.83)	(13,888.51)	(6,469.11)	(3,395.33)
	Lever on the environmental performance Ecological Focus -0.0001 Area (0.0002) Mineral N spread 0.003	Electricity per cow -0.0002 -0.0001 ** (0.0001) (0.0000) Organic N on pasture -0.0002 0.0001		0.69 (0.69) -14.56 Fuel per ha (0.32) -0.17 Share manure in organic -19.47 ***	-0.43 ** (0.17) 6.68 ***
	on pasture Organic N on		(0.003) -0.004 **	(0.002) Manure composting (0.001) 0.0005 0.0000		fertilizers -38.55 *** (9.10)	(4.24) -32.19 ***	(2.23) 7.57 ***
	Lever on the economic pasture (0.002) -0.14 ** Manure composting (0.07) performance Share of manure in -0.69 organic fertilisers (0.49)	Labor use per cow Share protein in the diet (0.001) (0.0003) -0.09 ** -0.01 (0.04) (0.01) Ecological Focus Area -0.08 0.01 (0.32) (0.10)	(5.81) -306.13 Labor use per cow (2.71) -84.75 Share protein in the diet (235.60) (109.74) -3,593.83 ** -305.90 Ecological Focus Area (1,705.98) (794.63)	(1.42) -97.04 * (57.60) 222.04 (417.06)
	Constant Trade-off Observations	1.01 (1.23) 95	-0.53 (0.81) 95	0.81 *** (0.25) 95		Mineral N on pasture 2,645.66 -43.49 (4,267.82) (1,987.90) Organic N on pasture 95 95	2,579.30 ** (1,043.36) 95
	R 2		0.75	0.35	0.82		0.87	0.94	0.86
	Adjusted R 2		0.65	0.09	0.75		0.81	0.91	0.80
					22

gross environmental performance per hectare. It also decreases net environmental performance per liter, but not significantly. As explained above, product-based indicators strongly respond to practices influencing cows' productivity. Thus, the non-significant effect of the concentrate use on the environmental performance may be explained by its limited effect on cows' productivity in our sample. Indeed, in our PDO sample, the capacity of the farmers to buy fodder crops and feed from the outside the PDO area is limited by the label's constraints, which forces them to develop other feeding practices, such as grazing and mowing.

Methodological advantages of the study

We find that using two indicators for performances, per liter and per hectare, is helpful in providing meaningful interpretations. Indeed, reasoning with product-based indicators presents the risk of underestimating the environmental impact of intensive practices (Salou et al., 2017a). As the per liter measure of the environmental performance is defined as the ratio between GHG emissions and milk production, if a practice increases the cows' productivity more than the GHG emissions, it will rise the environmental performance per liter. However, such practices would increase the absolute farm's GHG emissions, as well as GHG emissions per cow or per hectare. For example, in our study, only half of the significant practices impact both performances per liter and hectare (fuel per ha, share of protein in the diet and labor use per cow). The other identified levers are less robust and the recommendations to the farmers thus depend on the choice of the indicator. Note that the indicator selected for economic performance are correlated with other possible choices such as gross margin per labor unit.