Biology subjects

Gomez, N.

Publications and source records attributed to Gomez, N..

9 recordsLinked to original sources

bioRxiv2024

Molecular Xenomonitoring (MX) allows real-time surveillance of West Nile and Usutu virus in mosquito populations.

BackgroundWest Nile (WNV) and Usutu (USUV) virus are vector-borne flaviviruses causing neuroinvasive infections in both humans and animals. Entomological surveillance is a method of choice for identifying virus circulation ahead of the first human and animal cases, but performing molecular screening of vectors is expensive, and time-consuming. MethodsWe implemented the MX (Molecular Xenomonitoring) strategy for the detection of WNV and USUV circulation in mosquito populations in rural and urban areas in Nouvelle-Aquitaine region (France) between July and August 2023, using modified BG Sentinel traps. We first performed molecular screening and sequencing on excreta from trapped mosquitoes before confirming the results by detecting, sequencing and isolating viruses from individual mosquitoes. FindingsWe identified WNV and USUV-infected mosquitoes in 3 different areas, concurrently with the first human cases reported in the region. Trapped mosquito excreta revealed substantial virus co-circulation (75% of traps had PCR+ excreta for at least one of both viruses). Cx. pipiens was the most common species infected by both WNV and USUV. Genomic data from excreta and mosquitoes showed the circulation of WNV lineage 2 and USUV lineage Africa 3, both phylogenetically close to strains that circulated in Europe in recent years. Four WNV and 3 USUV strains were isolated from trapped mosquitoes. InterpretationMX strategy is easy and rapid to implement on the field, and has proven its effectiveness in detecting WNV and USUV circulation in local mosquito populations. FundingThis study received funding from the Direction Generale de lArmement (PDH 2 NBC-5-B-2212) and ARBOGEN (funded by MSDAVENIR). Research in contextO_ST_ABSEvidence before this studyC_ST_ABSWNV and USUV circulate through complex transmission cycles involving mosquitoes as vectors, birds as amplifying hosts and several mammal species as dead-end hosts. Transmission to humans primarily occurs through mosquito bites for both viruses. Notably, WNV can also be transmitted through blood donations and organ transplants. It is estimated that a significant proportion of both WNV and USUV infections in vertebrate hosts remain unreported due to their predominantly asymptomatic nature or nonspecific clinical presentation. Nevertheless, neuroinvasive and potentially fatal disease can occur, in particular among vulnerable populations such as elderly and immunocompromised patients. In France, after its first detection in 2015, USUV has been sporadically found in eastern and southern departments, with confirmed infections in birds, mosquitoes and mammals, and few human cases described. WNV has recently caused annual outbreaks of varying intensities involving humans, equids and avifauna in French departments mainly located in the Mediterranean area. Because of low viral loads and/or brief viremia, diagnosis of both pathogens is often based on serological evidence, and few genomic data are available on strains having circulated in France. Entomological surveillance can be used as an early warning method for WNV and USUV surveillance, but is costly to implement as it requires the collection of large numbers of mosquitoes to detect virus circulation when infection rates in mosquito populations are low. Therefore, viral surveillance in France still heavily relies on human and animal surveillance, i.e. late indicators of viral circulation. Added value of this studyThis study describes the implementation of the MX (Molecular Xenomonitoring) strategy for the effective surveillance of WNV and USUV circulation within mosquito populations. MX uses of modified BG Sentinels that allow (i) trapped mosquitoes to survive for several days and (ii) corresponding mosquito excreta to be collected and preserved on filter paper. MX has demonstrated many advantages over traditional entomological surveillance. Firstly, screening excreta collectively deposited by a community of trapped mosquitoes for the presence of viruses in the first instance is time and cost efficient, as one sample is tested for viral RNA, regardless of the number and species diversity in the trap. Second, filter papers with mosquito excreta can be transported from the field to the laboratory at room temperature by regular postal mail, bringing real-time detection within reach. WNV and USUV RNA have been detected and sequenced directly from the mosquito excreta shortly after collection. Thirdly, MX adapters increase the longevity of trapped mosquitoes, thereby allowing extension of the time between trap collections and increasing the likelihood of virus shedding by infected mosquitoes. Fourthly, this approach is easy to implement in the field and requires neither a strong entomological background nor specific technical skills. All these aspects make the MX strategy a powerful, non-invasive and cost-effective tool for real-time monitoring of enzootic WNV and USUV circulation. Authors should describe here how their findings add value to the existing evidence.WNV was never detected on the Atlantic seaboard of France until October 2022. Molecular evidence of WNV circulation was obtained in 3 symptomatic horses in the Nouvelle Aquitaine region in October 2022, concomitantly with an USUV human case with no recent travel history outside the region. This was a harbinger of an increase in cases over the next year. In 2023, MX succeeded in detecting the enzootic co-circulation of WNV and USUV in rural and urban areas of Nouvelle Aquitaine, simultaneously with the first cases of WNV detected by human and animal surveillance and the first human case of USUV diagnosed in the end of July 2023. Genomic and phylogenetic information was obtained directly from trapped mosquito excreta, before information derived from animal or human surveillance. Mosquitoes from traps with PCR-positive excreta were analysed individually, which allowed to calculate infection rates in mosquitoes. WNV and USUV were isolated from single Cx. pipiens mosquitoes. Cx. pipiens was the species most commonly found positive for either viruses although WNV was also detected in Ochlerotatus and Aedes mosquitoes, including one tiger mosquito (Ae. albopictus) in the urban environment. We argue that the MX approach is a major asset in the early warning detection of WNV and USUV circulation to alert health policy makers and take suitable control measures.

molecular biology↗

bioRxiv2024

Counter-regulation of RNA stability by UPF1 and TDP43

RNA quality control is crucial for proper regulation of gene expression. Disruption of nonsense mediated mRNA decay (NMD), the primary RNA decay pathway responsible for the degradation of transcripts containing premature termination codons (PTCs), can disrupt development and lead to multiple diseases in humans and other animals. Similarly, therapies targeting NMD may have applications in hematological, neoplastic and neurological disorders. As such, tools capable of accurately quantifying NMD status could be invaluable for investigations of disease pathogenesis and biomarker identification. Toward this end, we assemble, validate, and apply a next-generation sequencing approach (NMDq) for identifying and measuring the abundance of PTC-containing transcripts. After validating NMDq performance and confirming its utility for tracking RNA surveillance, we apply it to determine pathway activity in two neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) characterized by RNA misprocessing and abnormal RNA stability. Despite the genetic and pathologic evidence implicating dysfunctional RNA metabolism, and NMD in particular, in these conditions, we detected no significant differences in PTC-encoding transcripts in ALS models or disease. Contrary to expectations, overexpression of the master NMD regulator UPF1 had little effect on the clearance of transcripts with PTCs, but rather restored RNA homeostasis through differential use and decay of alternatively poly-adenylated isoforms. Together, these data suggest that canonical NMD is not a significant contributor to ALS/FTD pathogenesis, and that UPF1 promotes neuronal survival by regulating transcripts with abnormally long 3UTRs.

neuroscience↗

bioRxiv2023

Stress granule formation helps to mitigate neurodegeneration

Cellular stress pathways that inhibit translation initiation lead to transient formation of cytoplasmic RNA/protein complexes known as stress granules. Many of the proteins found within stress granules and the dynamics of stress granule formation and dissolution are implicated in neurodegenerative disease. Whether stress granule formation is protective or harmful in neurodegenerative conditions is not known. To address this, we took advantage of the alphavirus protein nsP3, which selectively binds dimers of the central stress granule nucleator protein G3BP (rin in Drosophila) and markedly reduces stress granule formation without directly impacting the protein translational inhibitory pathways that trigger stress granule formation. In Drosophila and rodent neurons, reducing stress granule formation with nsP3 had modest impacts on lifespan even in the setting of serial stress pathway induction. In contrast, reducing stress granule formation in models of ataxia, amyotrophic lateral sclerosis and frontotemporal dementia largely exacerbated disease phenotypes. These data support a model whereby stress granules mitigate, rather than promote, neurodegenerative cascades.

neuroscience↗

bioRxiv2023

Multi-omic characterization of antibody-producing CHO cell lines elucidates metabolic reprogramming and nutrient uptake bottlenecks

Characterizing the phenotypic diversity and metabolic capabilities of industrially relevant manufacturing cell lines is critical to bioprocess optimization and cell line development. Metabolic capabilities of the production hosts limit nutrient and resource channeling into desired cellular processes and can have a profound impact on productivity but cannot be directly inferred from measured data such as spent media concentrations or transcriptomics. Here, we present an integrated multi-omic characterization approach combining exo-metabolomics, transcriptomics, and genome-scale metabolic network analysis and apply it to three antibody-producing Chinese Hamster Ovary cell lines to reprogramming features associated with high-producer clones and metabolic bottlenecks limiting product production in an industrial bioprocess. Analysis of individual datatypes revealed a decreased nitrogenous byproduct secretion in high-producing clones and the topological changes in peripheral metabolic pathway expression associated with phase shifts. An integrated omics analysis in the context of the genome-scale metabolic model elucidated the differences in central metabolism and identified amino acid utilization bottlenecks limiting cell growth and antibody production that were not evident from exo-metabolomics or transcriptomics alone. Thus, we demonstrate the utility of a multi-omics characterization in providing an in-depth understanding of cellular metabolism, which is critical to efforts in cell engineering and bioprocess optimization.

systems biology↗

bioRxiv2023

Multiplexed amplicon sequencing reveals the heterogeneous spatial distribution of pyrethroid resistance mutations in Aedes albopictus mosquito populations in Southern France.

The risk of mosquito-borne diseases transmission is moving fast toward temperate climates with the colonization and proliferation of the Asian tiger mosquito vector Aedes albopictus and the rapid and mass transport of passengers returning from tropical regions where the viruses are endemic. The prevention of major Aedes-borne viruses heavily relies on the use of insecticides for vector control, mainly pyrethroids In Europe. High-throughput molecular assays can provide a cost-effective surrogate to phenotypic insecticide resistance assays when mutations have been previously linked to a resistance phenotype. Here, we screened for the spatial distribution of kdr mutations at a large scale using a two-step approach based on multiplexed amplicon sequencing and an unprecedented collection of field-derived mosquitoes in South of France. We identified the presence of the V1016G allele in 14 sites. The V1016G allele was predominantly found in South-East France close to the Italian border with two additional isolated sites close to Bordeaux and Marmande. All mosquitoes were heterozygous for this mutation and should not be phenotypically resistant to pyrethroid insecticide. Four other mutations were identified in our targeted genomic sequence: I1532T, M1006L, M1586L, M995L. Sequencing a section of maternally inherited mitochondrial genome confirmed that the spread of Ae. albopictus in France originated from founders with haplogroup A1. These findings contribute to the broader understanding of resistance dynamics in Europe and can inform targeted approaches to mitigate the impact of resistance on vector control.

genetics↗

bioRxiv2022

RNA methylation influences TDP43 binding and disease pathogenesis in models of amyotrophic lateral sclerosis and frontotemporal dementia

Methylation of RNA at the N6 position of adenosine (m6A) is one of the most common RNA modifications, impacting RNA stability as well as its transport and translation. Previous studies uncovered RNA destabilization in models of amyotrophic lateral sclerosis (ALS), in association with accumulation of the RNA-binding protein TDP43, which is itself mislocalized from the nucleus in >95% of those with ALS. Here, we show that TDP43 recognizes m6A-modified RNA, and that RNA methylation is critical for both TDP43 binding and autoregulation. We also observed extensive hypermethylation of coding and non-coding transcripts in ALS spinal cord, many of which overlap with methylated TDP43 target RNAs. Emphasizing the importance of m6A for TDP43 binding and function, we identified several m6A factors that enhance or suppress TDP43-mediated toxicity via a single-cell CRISPR/Cas9 candidate-based screen in primary neurons. The most promising genetic modifier among these--the canonical m6A reader YTHDF2-- accumulated within spinal motor neurons in ALS postmortem sections, and its knockdown prolonged the survival of human neurons carrying ALS-associated mutations. Collectively, these data show that m6A modifications modulate RNA binding by TDP43, and that m6A is pivotal for TDP43-related neurodegeneration in ALS.

neuroscience↗

bioRxiv2021

Mosquito excreta reveals circulation of West Nile virus and its underlying ecosystem

Emerging and endemic mosquito-borne viruses can be difficult to detect and monitor because they often cause asymptomatic infections in human or vertebrate animals or cause nonspecific febrile illness with a short recovery waiting period. Cases detection in vertebrate hosts can be complemented by entomological surveillance, but this method is not adapted to low infection rates in mosquito populations that typically occur in low or non-endemic areas. We identified West Nile Virus circulation in Camargue, a wetland area in South of France, using a cost effective innovative xenomonitoring method based on the molecular detection of virus in excreta from trapped mosquitoes. We also succeeded at identifying the mosquito community diversity dynamic on several sampling sites, together with the vertebrate hosts on which they fed prior to be captured using amplicon-based metagenomic on mosquito excreta without processing any mosquito. Mosquito excreta-based virus surveillance can be considered as a cost-effective and non-invasive strategy that offers the additional asset to reveal the ecological network underlying arbovirus circulation.

ecology↗

bioRxiv2020

TDP-43 stabilizes transcripts encoding stress granule protein G3BP1: potential relevance to ALS/FTD

TDP-43 nuclear depletion and concurrent cytoplasmic accumulation in vulnerable neurons is a hallmark feature of progressive neurodegenerative proteinopathies such as amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Cellular stress signalling and stress granule dynamics are now recognized to play a role in ALS/FTD pathogenesis. Defective stress granule assembly is associated with increased cellular vulnerability and death. G3BP1 (Ras-GAP SH3-domain-binding protein 1) is a critical stress granule assembly factor. Here, we define that TDP-43 stabilizes G3BP1 transcripts via direct binding of a highly conserved cis regulatory element within the 3UTR. Moreover, we show in vitro and in vivo that nuclear TDP-43 depletion is sufficient to reduce G3BP1 protein levels. Finally, we establish that G3BP1 transcripts are reduced in ALS/FTD patient neurons bearing TDP-43 cytoplasmic inclusions/nuclear depletion. Thus, our data suggest that, in ALS/FTD, there is a compromised stress granule response in disease-affected neurons due to impaired G3BP1 mRNA stability caused by TDP-43 nuclear depletion. These data implicate TDP-43 and G3BP1 loss of function as contributors to disease.

neuroscience↗

bioRxiv2020

Astrocyte-neuron crosstalk through Hedgehog signaling mediates cortical circuit assembly

Neuron-glia relationships play a critical role in the regulation of synapse formation and neuronal specification. The cellular and molecular mechanisms by which neurons and astrocytes communicate and coordinate are not well understood. Here we demonstrate that the canonical Sonic hedgehog (Shh) pathway is active in cortical astrocytes, where it acts to coordinate layer-specific synaptic connectivity and functional circuit development. We show that Ptch1 is a Shh receptor that is expressed by cortical astrocytes during development and that Shh signaling is necessary and sufficient to promote the expression of layer-specific astrocyte genes involved in regulating synapse formation and function. Loss of Shh in layer V neurons reduces astrocyte complexity and coverage by astrocytic processes in tripartite synapses, moreover, cell-autonomous activation of Shh signaling in astrocytes promotes cortical excitatory synapse formation. Together, these results suggest that Shh secreted from deep layer cortical neurons acts to specialize the molecular and functional features of astrocytes during development to shape circuit assembly and function.

neuroscience↗