Biology subjects

Bordes, L.

Publications and source records attributed to Bordes, L..

4 recordsLinked to original sources

bioRxiv2025

Multiplexed imaging of G-proteins and ERK activity upon activation of CaSR

We have previously shown that the calcium sensing receptor (CaSR) activates different G proteins and second messengers in single cells, and that GPCR and G protein-mediated ERK activity can be highly dynamic and heterogeneous at the single cell level. Here, we attempted to investigate the CaSR-Gi-ERK signaling pathway in single cells with previously characterized biosensors. We developed a strategy to simultaneously track G-protein and ERK activities in a single cell, using FRET-based and translocation sensors, a novel large-Stokes-shift green fluorescent protein, and an imaging setup with a single detector and a single wavelength for excitation. Extensive characterization and optimization to unmix the signals showed that the differences in dynamic range and type of read-out obtained from these biosensors made it very challenging to obtain robust FRET-measurements in the developed setup. When focusing solely on ERK, we found signs of possible ERK activity in response to calcium stimulation via CaSR, as well as milder changes in the absence of calcium treatment. Further optimization of the model, probes, and image processing will be necessary to develop a setup to robustly measure FRET and translocation simultaneously in single cells. We present the challenges we encountered and discuss future developments that can overcome them.

cell biology↗

bioRxiv2025

Mathematical modelling of in vitro replication dynamics for multiple highly pathogenic avian influenza clade 2.3.4.4 viruses in chicken and duck cells

The introduction and subsequent detection of highly pathogenic avian influenza (HPAI) in poultry is influenced by the virus replication fitness, transmission fitness, and virulence in poultry. These viral fitness parameters are important for implementing surveillance and control measures for poultry. This study investigates the potential application of an avian in vitro model using primary chicken embryo (CEF) and duck embryo fibroblasts (DEF) to identify the viral fitness for a reference panel of eight dominant HPAI clade 2.3.4.4 virus genotypes: four H5N1 viruses isolated between 2021 and 2024, as well as three H5N8 and one H5N6 virus isolated between 2014 and 2020. Infectious virus titre and cytopathogenicity were measured in the primary cell cultures over time and these data were analysed using a mathematical model which delineates cell populations into susceptible, latent, infectious, and dead compartments. In addition to obtaining "traditional" virological parameters such as peak virus replication and the time to 50% cell death, eight new parameters, key among those, the infecting time (tinf), generation time (tgen) and basic reproduction number (R0), were estimated using the mathematical model. Collectively, these parameters contribute to virus characterization, enhancing the resolution for comparing genetically similar viruses. This approach can allow for the evaluation of virus virulence, replication fitness, and, ideally, transmissibility fitness across different hosts. This study underscores the potential of integrating avian in vitro models with mathematical modeling and builds towards rapid risk assessments of novel HPAI viruses.

microbiology↗

bioRxiv2023

Cross-transmission of resistant gastrointestinal nematodes between wildlife and transhumant sheep

Wild and domestic ungulates can be infected with the same species of gastrointestinal parasitic nematodes. These parasites have free-living stages in the environment that contribute to the ease of transmission among different host species. In addition, gastrointestinal nematodes have developed resistance to anthelmintics which is now considered a major problem for the livestock sector. In a context where wild and domestic ungulates share the same pastures, the maintenance and circulation of resistant gastrointestinal nematodes between species have rarely been explored. In the European Alps, domestic sheep are driven to high-altitude summer pastures and live in sympatry with wild ungulates for several months each year. In this study, we investigated the nemabiome of domestic sheep and Alpine ibex, Capra ibex, in three different areas of the French Alps to evaluate parasite circulation between the two host species. The Alpine ibex is a protected mountain ungulate that is phylogenetically related to sheep and hosts nematode species common to sheep. Using internal transcribed spacer 2 (ITS-2) nemabiome metabarcoding, we found sheep and ibex share similar gastrointestinal nematodes, except for a few species such as Marshallagia marshalli and Trichostrongylus axei. This suggests that the long-term co-occurrence of sheep and ibex on mountain pastures has promoted the exchange of gastrointestinal nematodes between the two hosts. Based on the sequencing of the isotype 1 of the beta tubulin gene, associated with benzimidazole resistance, we found resistant nematodes in all sheep flocks and in all ibex populations. Our results demonstrated that ibex can host and shed resistant strains before transhumant sheep arrive on pastures, and thus could act as a refuge or even contribute to maintaining resistant gastrointestinal nematodes. The relative role of ibex in the maintenance and circulation of resistant strains in sheep remain to be determined.

ecology↗

bioRxiv2022

Highly pathogenic avian influenza H5N1 virus infections in wild red foxes (Vulpes vulpes) show neurotropism and adaptive virus mutations

During the 2020-2022 epizootic of highly pathogenic avian influenza virus (HPAI) several infections of mammalian species were reported in Europe. In the Netherlands, HPAI H5N1 virus infections were detected in three wild red foxes (Vulpes vulpes) that were submitted with neurological symptoms between December 2021 and February 2022. Histopathological analysis demonstrated the virus was mainly present in the brain, with limited or no detection in the respiratory tract and other organs. Phylogenetic analysis showed the three fox viruses were not closely related, but were related to HPAI H5N1 clade 2.3.4.4b viruses found in wild birds. In addition, limited virus shedding was detected suggesting the virus was not transmitted between the foxes. Genetic analysis demonstrated the presence of mammalian adaptation E627K in the polymerase basic two (PB2) protein of the two fox viruses. In both foxes the avian (PB2-627E) and the mammalian (PB2-627K) variant were present as a mixture in the virus population, which suggests the mutation emerged in these specific animals. The two variant viruses were isolated and virus replication and passaging experiments were performed. These experiments showed mutation PB2-627K increases replication of the virus in mammalian cell lines compared to the chicken cell line, and at the lower temperatures of the mammalian upper respiratory tract. This study showed the HPAI H5N1 virus is capable of adaptation to mammals, however more adaptive mutations are required to allow efficient transmission between mammals. Therefore, surveillance in mammals should be expanded to closely monitor the emergence of zoonotic mutations for pandemic preparedness. IMPORTANCEHighly pathogenic avian influenza (HPAI) viruses caused high mortality amongst wild birds in 2021-2022 in the Netherlands. Recently three wild foxes were found to be infected with HPAI H5N1 viruses, likely by feeding on infected birds. Although HPAI is a respiratory virus, in these foxes the viruses were mostly detected in the brain. Two viruses isolated from the foxes contained a mutation that is associated with adaptation to mammals. We show the mutant virus replicates better in mammalian cells than in avian cells, and at the lower body temperature of mammals. More mutations are required before viruses can transmit between mammals, or can be transmitted to humans. However, the infections in mammalian species should be closely monitored to swiftly detect mutations that may increase the zoonotic potential of the HPAI H5N1 viruses as these may threaten public health.

microbiology↗