COVID-19 represents an unprecedented world-wide challenge due to the transmissibility of the virus, its impact on morbidity and mortality, the uncertainty regarding the development of long-term immunity, and the current paucity of efficient therapeutic options. Understanding the kinetics of shedding of Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2), and the duration of viral infectivity is of critical importance for controlling the pandemic. Such understanding is most crucial for the patients experiencing an asymptomatic or pauci-symptomatic infection as a large proportion of transmissions appears to occur from these subjects. Very few studies investigated the duration of detection of viral RNA in asymptomatic or pre-symptomatic subjects, and more specifically in children. They suggest that the presence of infectious virus is correlated to the overall viral load and inversely correlated with the delay from symptoms onset and the appearance of a neutralizing antibody response. Other factors that could have an impact on infectivity duration are still uncertain. This includes the innate and the adaptive immune response to SARS-CoV-2 as well as the concomitant presence of other viruses which can elicit cross-immunity. In particular, the impact of prior immunity to SARS-CoV-2 or to seasonal coronaviruses on the duration of SARS-CoV-2 infectivity warrants to be explored further. The objective of the PED-COVID_Infect project is to measure SARS-CoV-2 infectivity in a cohort of pauci-symptomatic children and their parents to evaluate how the duration and intensity of infectious virus shedding are affected by viral and immune interference, in comparison to adults. The project will benefit from the unique longitudinal collection of nasopharyngeal, saliva and blood samples from the PED-COVID cohort currently implemented in children and adults undergoing asymptomatic or pauci-symptomatic SARS-CoV-2 infection, already funded by a PHRC. We will determine the kinetics of viral clearance by assessing SARS-CoV-2 virus load and viability in the upper respiratory airways and the saliva, and its modulation by 4 main factors: (i) anti-SARS-CoV-2 antibody response at the local and general level; (ii) pre-existing antibodies towards seasonal coronaviruses; (iii) co-infections with specific respiratory pathogenic viruses; (iv) composition of the whole local eucaryotic virome. Virological characterization will be based on RT-PCR for quantification, RNA sequencing and virus infectivity on VeroE6 cells. Anti-SARS-CoV-2 and seasonal coronavirus virus antibodies will be assessed in serum by a liquid-phase Luciferase Immunoprecipitation System (LIPS)-based assay and in the saliva and the nasopharyngeal mucosa by ELISA. NGS-based agnostic metagenomics will focus on the whole viral population of the nasopharyngeal mucosa. The outcomes of our project, which combines virological, serological and metagenomic data, will provide a better understanding of the duration of SARS-CoV-2 infectivity in pauci-symptomatic children and adults, as well as new insights as to factors that could modulate virus infectivity. Such knowledge is key to inform public health policy, including quarantine, isolation and contact tracing.

Aneuploidy, the state of cells containing an incorrect number of chromosomes, is associated with defective cellular functions. Aneuploidy occurs frequently during human embryonic development and often causes miscarriages or genetic diseases. However, all chromosomes are not equally likely to become aneuploid in human embryos. It has been proposed that chromosome-specific aneuploidy could arise from different propensities to be mis-segregated during mitosis as a result from the interaction of the mitotic spindle with the chromosome-specific centromeres. Also, during embryonic development, repair mechanisms could eliminate aneuploid cells into extra-embryonic tissues. However, this could not be studied systematically due to lack of tools to generate chromosome-specific aneuploidy. Our project CaCAn proposes to use newly developed biophysical and biochemical approaches to identify the causes and consequences of chromosome-specific aneuploidy in cellulo and in vivo. First, we will probe the topological and mechanical properties of human centromeres using our 2 newly developed methods for centromere enrichment and magnetic manipulation. Second, we will force chromosome-specific aneuploidy by directly pulling on chromosomes to reveal connections between chromosome positions and aneuploidy. Finally, we will study the behavior of aneuploid cells in both mouse and human preimplantation embryos and relate this to the unbalance of specific chromosomes. By providing molecular, physical and cellular insights into the causes and consequences of chromosome-specific aneuploidy, CaCAn will reveal the mechanisms of a fundamental cellular defect that is frequently at the origin of many pathologies.

Cerebral folate deficiency (CFD), defined by a low folate cerebrospinal fluid (CSF) concentration, can be linked to genetic defects of folate metabolism or be secondary to various diseases without clear causal link, and be associated to potentially very disabling cognitive and motor symptoms. We identified a neurological syndrome (named LHIPFOLD for Leukoencephalopathy with High CSF Protein and FOLate Deficiency) characterized by deep CFD with normal blood folate, high protein level in CSF (>1g/L) and a specific leukoencephalopathy. A folate supplementation may stabilize or even improve symptoms in some patients, but with limited impact. In some patients, this syndrome is associated to genetic mutations leading to a mitochondrial malfunction, without connection to folate metabolism, whereas LHIPFOLD syndrome remains unexplained in others even after genome sequencing. We hypothesize that CFD in LHIPFOLD is due to generalized Choroid Plexus (CP) dysfunction, a brain organ that expresses transporters regulating flux between blood and CSF of numerous metabolites (including folate), and secretes CSF and specific proteins in CSF with various functions (trophic function, adult neurogenesis, etc). Consequently, other potentially treatable biochemical abnormalities due to PC dysfunction may exist in LHIPFOLD, beyond CFD. Currently, there are no available clinical explorations to evaluate CP functions, whereas we consider LHIPFOLD a very useful model to validate the capacity of some relevant diagnostic tools to do so. Therefore, our objectives are to identify a CP-related MRI and biochemical signature in LHIPFOLD patients, using morphological and functional imaging (CP capillary permeability and CP macrovascular perfusion), and blood/CSF metabolomics/proteomics approaches (untargeted then targeted validation of candidate biomarkers related to CP physiology); to identify CP-related biochemical therapeutic targets in LHIPFOLD patients that could improve patients condition combined to folate supplementation; and to set-up imaging and biochemical diagnostic tests investigating CP function for clinical practice. For this, brain MRI data and blood/CSF samples will be collected during 2 years from LHIPFOLD patients and controls (healthy volunteers and neurological patients). Some experimental data indicate that the innovative concept of generalized PC dysfunction as part of a more global pathophysiology has the potential to be applied to other neurological diseases like Alzheimer’s disease or multiple sclerosis. Therefore, efficient diagnostic tools exploring CP function will be of great utility not only in suspected LHIPFOLD patients but also in more common neurological diseases, potentially leading to original therapeutic approaches.