The influence of microbiomes on their hosts has emerged as one of the most promising factors able to modulate and improve livestock phenotypes, especially health and feed efficiency. Furthermore, increasing evidence is showing an involvement of microbiota into the gut-brain axis, which might contribute to individual welfare. Host-microbiota interactions are influenced by a myriad of external factors, from environment and diets to general management practices, but also by host genetics. The H2020 SFS-02-2020 project call targets these topics by asking for research on the interplay between the production animals and their microbial ecosystems in order to contribute to the improvement of sustainable livestock production. The aim of these ANR MRSEI project is to construct a network and provide a framework allowing the construction of a strong consortium for answering to the SFS-02-2020 project call. The project will contribute to the construction of the project consortium by funding three physical meetings of the whole consortium.

The severity of the global COVID-19 pandemic poses an urgent need for the development of efficient therapeutic strategies. To complete the available therapeutic arsenal, targeting the SARS-CoV-2 genome by antisense RNA therapy should be deeply investigated. We designed in silico antisense oligonucleotides (ASO) targeting viral genome to block the viral replication and transcription. The objective of the project is to validate the best ASO firstly by in vitro experiments on infected Vero E6 cultures, and secondly to test the best oligonucleotides antisense in vivo on infected animal model to perform a preclinical trial.

The release of metal nanoparticles (NPs) in ecosystems is constantly growing. NPs are massively manufactured for their useful properties such as UV-filter, anti-bacterial agents, remediation agent, fertilizers, or pesticides. Sampling from lakes, rivers, seas and even tap water indicated the presence of NPs, as well as in soils and in the air of densely urbanized and industrial areas. Whether animals living in these ecosystems are contaminated by NPs remains unknown.With iron oxide (Fe2O3) NPs, titanium dioxide (TiO2)-NPs is one of the most manufactured nanomaterial. Very emergent literature suggests an actual contamination in humans by TiO2-NPs in placental tissue, newborn stools, blood and colon tumor tissues. Whether animals, especially those raised for human consumption may be contaminated is unknown. Scarce studies in lactating rodents exposed to some NPs suggested the passage of NPs in milk and in offsprings whose development and survival were affected. In this context, NANOMILK proposes to evaluate the existence of an actual contamination by NPs in milk (WP1). Using combination of cutting-edge biophysical approaches, we will analyse the presence of NPs in milk from animals raised in urban farms with a known metal pollution in soils and vegetables. This will be compared to milk from animals raised in unpolluted arctic farms in Norway. To understand the mechanisms by which NPs may be released in milk, we will run a comprehensive analysis to characterize the molecular mechanisms underlying the secretion and intercellular transfer of NPs from mammary cells (WP2). To that end, we will investigate NP secretion in extracellular vesicles (EVs) which are an ensemble of membrane-limited carriers playing key roles in cell-to-cell communication and in physio-pathological processes such as immune response or cancer progression. Very recently, EVs have been detected in milk (milk-EVs). Secreted by mammary cells, they may transport information to offspring and influence their immunity and development. Whether milk-EVs secreted by mammary cells may also transport NPs is unknown but partners of NANOMILK have previously shown that Fe2O3-NPs and TiO2-NPs were transferred between non-mammary cells in a process involving EVs. Thorough analysis of the metal and protein content of EVs deriving from NPs-exposed mammary cells will be combined to transfer assays, indirect or direct in co-culture, coupled to silencing of genes involved in cell-cell communication. Furthermore the impact of NPs transferred from mammary cells to recipient cells will be evaluated on genome-wide expression profiles by transcriptomics in recipient cells. Finally, we propose to investigate in vivo the route of NPs within the entire mother-to-offpsring continuum (WP3). After a dose escalation study, we will analyse the behavior of NPs administrated by drinking water in lactating female rabbits. Biodistribution of NPs will be analysed in mother and offspring tissues and in milk-EVs collected at different lactation time. In addition to monitoring the effect of NPs on offspring growth and survival, we will also analyse their effect on milk-EV proteome to uncover potential effect on EVs biogenesis, origin and abundance. From on-field, to in vitro and to in vivo, NANOMILK investigates the spreading of contaminant NPs from mother-to-progeny. NANOMILK may reveal an actual contamination in milk samples from urban farms, influencing this agricultural trend of producing locally when location is polluted. Expected results will generate knowledge on mother-to-offspring communication by milk, and how this can be highjacked by pollutants. NANOMILK may provide new grounds for the use of NPs, for dairy industry, for nanoagriculture and breastfeeding in polluted areas.

ROBUSTNESS in farm animals was defined by Knap as ‘the ability to combine a high production potential with resilience to stressors, allowing for unproblematic expression of a high production potential in a wide variety of environmental conditions’. The importance of robustness-related traits in breeding objectives is progressively increasing towards the production of animals with a high production level in a wide range of climatic conditions and production systems, together with a high level of animal welfare. Current strategies to increase robustness include selection for ‘functional traits’, such as skeletal and cardiovascular integrity, disease resistance and mortality in various stages. It is also possible to use global evaluation of sensitivity to the environment (e.g. reaction norm analysis or canalization), but these techniques are difficult to implement in practice. STRESS is defined as the non-specific response of the organisms to any challenge. In vertebrate food-producing animal species, the hypothalamic–pituitary–adrenocortical (HPA) axis is the most important stress-responsive neuroendocrine system. Cortisol (or corticosterone) released by the adrenal cortices exerts a large range of effects on metabolism, the cardiovascular system, inflammatory processes and brain function, for example. Large individual variations have been described in the HPA axis activity with important physiopathological consequences. In terms of animal production, higher cortisol levels have negative effects on growth rate and feed efficiency and increase the fat/lean ratio of carcasses. On the contrary, cortisol has positive effects on traits related to robustness and adaptation. Intense selection for lean tissue growth during the last decades has concomitantly reduced cortisol production, which may be responsible for the negative effects of selection on robustness traits. The strategy that we propose is to change the balance between production and robustness by selecting animals with higher HPA axis activity (Mormède et al. Animal 5:651, 2011). The first aim of the present project SUSoSTRESS is to produce experimental evidence supporting this strategy. The proof of concept will be given by the study of production and robustness traits in two lines of pigs divergently selected for their cortisol response to ACTH stimulation. The critical advantage of this strategy for animal breeding is that it relies on a single, well-defined physiological system to increase robustness and adaptability, making genetic selection more easily reachable as well as the generalisation from the species under study, the pig, to other farm animal species. Numerous candidate genes and molecular polymorphisms have been described for genetic-based individual differences in HPA axis function, including hormone production and release by the adrenal cortices, bioavailability of hormones as well as receptor and post-receptor mechanisms (Mormède et al. ANYAS 1220:127, 2011). In order to use this molecular information for genetic selection, we need an integrated systems genetic approach. The second aim of the project is the elaboration of a model of genetic variation (genetic architecture) of the HPA axis and its physiological activity in relation to animal performance on both robustness and production traits, as the basis for genomic selection. The third aim of the project is to deliver practical information for genomic selection of more robust animals, by combination (integration) of the genetic model, the thorough bioinformatics analysis of the HPA axis components and targets, and the high-density genotyping of the bi-directionally selected animals. This research is therefore an exemplar of basic research with clear application for farm animal selection.

The PaterLeg project addresses the emerging issue of the inheritance of small non-coding RNAs from sperm and their potential to shape embryo development and to transmit possible paternally acquired phenotypes. Deciphering the "code of sperm RNA" and its consequences for offspring gene expression could move the field towards applications in human reproductive medicine and precision breeding. Here we aim to assess the extent to which sperm sncRNAs can influence the early development of the mammalian embryo. We will use the bovine embryo as a non-rodent embryo model. Indeed, the mouse presents particular characteristics of early development regulation, which do not always make it a relevant model for other mammals. Furthermore, the economic impact of sperm and embryo biotechnology in genetic selection is important for the cattle industry. We will first assess the paternal supply of sncRNAs to the early embryo, comparing it to the content of the oocyte and to the sncRNAs synthesised by the embryo itself. We will then explore the role of paternally inherited sncRNAs by overexpression or artificial inhibition experiments in the embryo of small groups of sncRNAs selected from scnRNAs differentially present in spermatozoa of fertile versus subfertile bulls, and/or sncRNAs acquired late by the spermatozoon during its passage through the epididymis. The resulting embryos will be phenotyped by morphokinetic analysis of their preimplantation development, integration of RNAseq and sncRNAseq data and by proteome analysis. Using novel mathematical approaches, gene networks will be inferred from the temporal and interventional datasets generated by PaterLeg to formulate and validate new hypotheses on the role of sperm sncRNAs. PaterLeg will thus improve our knowledge of the molecular mechanisms governing mammalian development. It will also open up the possibility of improving early embryonic development through new biotechnological approaches and contribute to the validation of candidate sncRNAs as markers of fertility in bulls.