Biology subjects

Barcena, J.

Publications and source records attributed to Barcena, J..

5 recordsLinked to original sources

bioRxiv2026

Feline calicivirus encoding NanoLuc luciferase as a tool for assessing antibody neutralisation and antivirals

Feline calicivirus (FCV) is among the most common viruses to infect cats worldwide, with prevalence estimated to range from 10-90% depending on the population sampled. Typical FCV infection presents with oral ulcerations, fever and in some cases can also lead to clinical signs such as pneumonia or "limping syndrome". However, some FCV strains have been isolated from cats exhibiting virulent systemic (VS) disease, which is associated with high morbidity and mortality. Breakthrough VS-FCV infections have been recorded in vaccinated cats and, therefore, there is considerable interest in developing novel therapeutics for use in the face of VS-FCV outbreaks. However, to design effective therapeutics, a tractable system to systematically assess the efficacy of novel vaccine candidates or antivirals is required. Here, we used reverse genetics to develop an FCV reporter virus, inserting NanoLuc luciferase into the LC protein of FCV-Urbana (FCV-UrbanaNL). We characterised the replication kinetics of FCV-UrbanaNL in comparison to its parent virus and assessed the stability of the reporter over multiple passages. Subsequently, we developed virus neutralisation assays to assess a range of monoclonal antibodies that recognise FCV Urbana. We then assessed the breadth of neutralisation by exchanging the major capsid protein, VP1, of FCV Urbana with VP1 from the vaccine strain F9 and the VS-FCV strain NSW-E1. Finally, we evaluated the utility of the FCVNL reporter system to screen candidate antiviral compounds, identifying GS-441524 (the active metabolite of the parent nucleoside remdesivir) as having therapeutic potential against FCV. These findings highlight the potential of this reporter virus as a powerful molecular tool to accelerate the discovery and development of novel therapeutics.

microbiology↗

bioRxiv2025

Development of an ex vivo model to study the zoonotic transmission of noroviruses

Human noroviruses are the most common cause of viral gastroenteritis worldwide, causing sporadic cases and outbreaks. Genetically related animal noroviruses infect wild and domestic animals including pigs and dogs. To investigate the potential of noroviruses to jump the species barrier, we used porcine, canine, and avian precision-cut intestinal slices. We show that human norovirus can bind to pig and dog intestinal tissue and be taken up with a similar efficiency as the porcine and canine noroviruses, respectively. In contrast, no binding or internalization of human, porcine, or canine noroviruses was detected in chicken tissue. We further showed that human norovirus can replicate to some degree in pig intestinal tissue, infecting single cells. In contrast, while animal noroviruses attached to human intestinal cells, intracellular uptake was limited. This suggests that human-to-animal transmission is more likely than animal-to-human transmission and that viral uptake likely presents a species barrier.

microbiology↗

bioRxiv2025

Calicivirus assembly and stability are mediated by the N-terminal domain of the capsid protein with the involvement of the viral genome

Caliciviruses are important human and animal pathogens that cause varying clinical signs including gastroenteritis, respiratory illness, and hepatitis. Despite the availability of numerous calicivirus structures, relatively little is known about the mechanisms of capsid assembly and stability, or about genome packaging. Here we present the atomic structure of the RHDV virion and several related non-infectious virus-like particles, determined using cryo-EM at 2.5-3.3 [A] resolution. The inherent molecular switch, responsible for the conformational flexibility of the capsid protein VP1, is located in its N-terminal arm (NTA). The NTA establishes an extensive network of interactions on the inner capsid surface that stabilizes the hexamers and pentamers. For this structural polymorphism, we show that the NTA must interact with the RNA viral genome, that is, the genomic RNA acts concomitantly with the NTA as a molecular switch. The NTA-RNA interaction leads to specific conformational states that result in two types of VP1 dimers (the basic building blocks) necessary for T=3 capsid assembly. In addition, we used atomic force microscopy (AFM) to assess whether differences in genomic RNA content influence viral properties such as capsid stiffness in physiological conditions. These analyses highlight the mechanical role of packed RNA genome in RHDV virions, as the virion capsid pentamers are strengthened by interactions of the NTA star-like structure promoted by the viral genome. These results indicate that the interactions between the NTA and the viral genome guide the conformational states of VP1 dimers, directing capsid assembly and modulating its mechanical properties. Through interference with intermediate assemblies, the NTA network promoted by the genome could be an attractive target in future antiviral strategies. Author SummaryRabbit hemorrhagic disease virus (RHDV) is a highly contagious, often fatal calicivirus that poses a significant threat to rabbit populations worldwide. Although considerable progress has been made in elucidating the structural organization of RHDV capsids at near-atomic resolution, information regarding the nucleic acid organization within these virions and its interactions with the viral capsid remains limited. Understanding these interactions is essential for identifying mechanisms of structural polymorphism of the capsid protein, as well as virion assembly and stability. We report the cryo-electron microscopy structure, at near-atomic resolution, of RHDV virions and distinct virus-like particles, together with characterization of their biophysical properties using atomic force microscopy. The N-terminal region of the capsid protein, essential for its structural polymorphism, requires its interaction with genomic RNA. The interaction between N-terminal arms and the viral genome governs the conformational states of capsid protein dimers, thereby orchestrating capsid assembly and influencing its biophysical properties. By interfering with intermediate assemblies, the NTA network promoted by the genome could constitute an attractive target in antiviral strategies for several major human calicivirus pathogens that cause acute gastroenteritis and respiratory illness, including norovirus and sapovirus.

biophysics↗

bioRxiv2021

Comparison of biological features of wild European rabbit mesenchymal stem cells derived from different tissues.

Although the European rabbit is an "endangered" species and a notorious biological model, the analysis and comparative characterization of new tissue sources of rabbit mesenchymal stem cells (rMSCs) has not been well studied. Here we report for the first time the isolation and characterization of rMSCs derived from an animal belonging to a natural rabbit population within the species native region. New rMSC lines were isolated from different tissues: oral mucosa (rOM-MSC), dermal skin (rDS-MSC), subcutaneous adipose tissue (rSCA-MSC), ovarian adipose tissue (rOA-MSC), oviduct (rO-MSC), and mammary gland (rMG-MSC). The six rMSC lines showed plastic adhesion with fibroblast-like morphology and were all shown to be positive for CD44 and CD29 expression (characteristic markers of MSCs), and negative for CD34 or CD45 expression. In terms of pluripotency features, all rMSC lines expressed NANOG, OCT4, and SOX2. Furthermore, all rMSC lines cultured under osteogenic, chondrogenic, and adipogenic conditions showed differentiation capacity. In conclusion, this study describes the isolation and characterization of new rabbit cell lines from different tissue origins, with a clear mesenchymal pattern. We show that rMSC do not exhibit differences in terms of morphological features, expression of the cell surface, and intracellular markers of pluripotency and in vitro differentiation capacities, attributable to their tissue of origin.

cell biology↗

bioRxiv2021

Eukaryotic CRFK cells motion characterized with atomic force microscopy

We performed a time-lapse imaging with Atomic Force Microscopy (AFM) of the motion of eukaryotic CRFK (Crandell-Rees Feline Kidney) cells adhered onto a glass surface and anchored to other cells in culture medium at 37{degrees}C. The main finding is a gradient in the spring constant of the actomyosin cortex along the cells axis. The rigidity increases at the rear of the cells during motion. This observation as well as a dramatic decrease of the volume suggests that cells may organize a dissymmetry in the skeleton network to expulse water and drive actively the rear edge.

biophysics↗