Biology subjects

Kaptein, S.

Publications and source records attributed to Kaptein, S..

7 recordsLinked to original sources

bioRxiv2026

Rat hepatitis E virus infection has multiphasic viral replication kinetics in vivo

Background and AimsHepatitis E virus (HEV) infections are a growing threat to global public health. To obtain an in-depth understanding of HEV infections in untreated and ribavirin-treated rats, we characterized the early HEV viral kinetics using rat HEV (rHEV) as a surrogate model and using two routes of virus inoculation: intravenous (I.V.) or oral infection. Approach and ResultsWe frequently collected feces, serum, and tissue samples up to 60 days after infection in both infection models to characterize the rHEV viral kinetics. A ~2-week delay in quantifiable RNA levels in feces was observed in the oral versus the I.V. infection model. Early rHEV viral kinetics in feces were found to be multiphasic and showed good concordance with those in the various tissue compartments studied. Comparison of the viral kinetics in these samples also revealed that the liver may serve as the initial site of rHEV replication, followed by replication in the intestine and spleen. While a dosage of 60 mg/kg/day ribavirin was found optimal to maintain rHEV RNA levels at (nearly) undetectable in feces, levels were detectable in the liver and increased both in feces and liver after treatment discontinuation. ConclusionsWe found that the two rHEV infection models share similar multiphasic viral kinetics with the liver as the main site of viral replication. Additionally, the rHEV RNA load in feces could be used as a reliable proxy for that in the liver, spleen, and intestine. We also show that ribavirin at 60 mg/kg/day was partially effective in preventing viral rebound. These findings may aid in exploring the correlation between the infection phases and antiviral efficacy, ultimately guiding therapeutic decisions.

microbiology↗

bioRxiv2025

Oropouche virus results in severe congenital disease in embryonic mice

The recent outbreak of Oropouche virus (OROV) in Latin-America with unprecedented reports of vertical transmission resulting in congenital malformations and fetal death, redefines the threat posed by this arbovirus. We here explore the teratogenic potential of an epidemic strain of the virus in a mouse model, whereby a low inoculum of OROV is injected into the placenta. The virus replicates efficiently in multiple embryonic organs, with high titers in the brain, and causes intrauterine growth restriction. Infected embryos display multi-organ pathology, including pneumonitis, myocarditis, steatohepatitis and severe neuropathology. Embryonic brains exhibit microcephaly, ventriculomegaly and extensive neural loss, mirroring findings in human fetal cases. A dramatic alteration of the neural cell population correlates with massive cell death, demonstrating the extreme cytotoxicity of OROV for the developing brain. The infection also triggers neuroinflammation characterized by cytokine upregulation, microglial activation and neutrophil infiltration. Altogether, our findings establish a link between congenital OROV infection and neurodevelopmental disease, highlighting its teratogenic potential.

microbiology↗

bioRxiv2025

Discovery of orally bioavailable Zika and West Nile Virus antiviral compounds targeting the NS2B-NS3 protease

Flaviviruses are a class of pathogenic viruses with pandemic potential that are typically transmitted via infected arthropods. In addition, Zika virus is sexually transmissible and causes congenital malformations if infection occurs during pregnancy. Although over 1.5 million people were infected during the 2015-2016 Zika outbreak, to date, there are no clinical-stage vaccines or antivirals. Herein, we report the discovery of potent inhibitors of the Zika virus NS2B-NS3 protease that also show activity against the West Nile virus NS2B-NS3 protease. Starting from a crystallographic fragment screen, we employed a pharmacophore approach coupled with high-throughput library chemistry to elaborate fragments in the active site. Potent, metabolically stable, non-covalent, non-peptidomimetic inhibitors were identified with antiviral activity in vitro. The lead compound was progressed to a three-day Zika virus challenge study in AG129 mice, where it displayed significant viral RNA load reduction in both mouse plasma and spleen.

microbiology↗

bioRxiv2024

Hepatitis E virus replication is facilitated by epithelial cell turnover and targets enteroendocrine cells in human intestinal organoids

Hepatitis E virus (HEV) is a leading pathogen causing acute viral hepatitis globally. While HEV is primarily spread fecal-orally, the role of the gut in HEV pathogenesis remains largely unexplored, including how HEV disseminates from gut to liver, and whether the gut is an HEV reservoir. We here aimed to illuminate HEV biology in the gut using human intestinal enteroids (HIEs). Three strategies were explored to establish an HEV-HIE model: three-dimensional (3D) HIEs, two-dimensional (2D) HIEs in transwell, and HEV RNA-electroporated HIEs. HEV particles produced by electroporated HIEs were characterized by western blot and gradient centrifugation. The intestinal tropism of HEV was investigated through confocal fluorescent microscopy and gene expression analysis. HEV infection in 3D-HIEs and 2D-HIEs showed limited replication, whereas HIEs electroporation led to a sustained increase in the release of non-enveloped infectious virions. These virions could re-infect new 3D-HIEs, yielding a [~]2 log10 increase in HEV RNA. In electroporated HIEs, high expression of the infectious ORF2 capsid form was observed in the supernatant. Importantly, 70% of all HEV-infected cells were identified as proliferative cells. ORF2 staining was also observed in absorptive enterocytes, goblet, and enteroendocrine cells. Overall, we established a robust HEV-HIE model that yields high titers of infectious non-enveloped virions. Proliferative cells and the fast intestinal epithelial cell turnover are important features that facilitate efficient HEV replication, and likely also its dissemination. This study suggests that the gut is an HEV reservoir, capable of producing some of the non-enveloped HEV shed in the feces. SignificanceHepatitis E virus (HEV) causes [~]20 million enterically transmitted cases of viral hepatitis globally. To better understand the largely unexplored role of the gut during an HEV infection, we used human intestinal enteroids (HIEs). We show that HEV-electroporated HIEs produce highly infectious non-envelopedHEV particles, resulting in a more efficient re-infection compared with traditional gold-standard methods. Importantly, we detected HEV in multiple human intestinal epithelial cell types within the human intestinal epithelium, with the majority of proliferative cells infected. This model offers valuable insights into the intestinal tropism of HEV and can serve to further investigate chronic HEV infection in the gut. Moreover, this underscores the importance of using physiologically relevant models to unravel details of HEV infection in the gut.

microbiology↗

bioRxiv2022

A SCID mouse model to evaluate the efficacy of antivirals against SARS-CoV-2 infection

Ancestral SARS-CoV-2 lacks the intrinsic ability to bind to the mouse ACE2 receptor and therefore establishment of SARS-CoV-2 mouse models has been limited to the use of mouse-adapted viruses or genetically modified mice. Interestingly, some of the variants of concern, such as the beta B.1.351 variant, show an improved binding to the mouse receptor and hence better replication in different Wild type (WT) mice species. Here, we desribe the establishment of SARS-CoV-2 beta B.1.351 variant infection model in male SCID mice as a tool to assess the antiviral efficacy of potential SARS-CoV-2 small molecule inhibitors. Intranasal infection of male SCID mice with 105 TCID50 of the beta B.1.351 variant resulted in high viral loads in the lungs and moderate signs of lung pathology on day 3 post-infection (pi). Treatment of infected mice with the antiviral drugs Molnupiravir (200 mg/kg, BID) or Nirmatrelvir (300 mg/kg, BID) for 3 consecutive days significantly reduced the infectious virus titers in the lungs by 1.9 and 3.8 log10 TCID50/mg tissue, respectively and significantly improved lung pathology. Together, these data demonstrate the validity of this SCID mice/beta B.1.351 variant infection model as a convenient preclinical model for assessment of potential activity of antivirals against SARS-CoV-2. ImportanceUnlike the ancestral SARS-CoV-2 strain, the beta (B.1.351) VoC has been reported to replicate to some extent in WT mice (species C57BL/6 and BALB/c). We here demonstrate that infection of SCID mice with SARS-CoV-2 beta variant results in high viral loads in the lungs on day 3 post-infection (pi). Treatment of infected mice with the antiviral drugs Molnupiravir or Nirmatrelvir for 3 consecutive days markedly reduced the infectious virus titers in the lungs and improved lung pathology. The advantages of using this mouse model over the standard hamster infection models to assess the in vivo efficacy of small molecule antiviral drugs are (i) the use of a clinical isolate without the need to use mouse-adapted strains or genetically modified animals (ii) lower amount of the test drug is needed and (ii) more convenient housing conditions compared to bigger rodents such as hamsters.

microbiology↗

bioRxiv2021

Alphavirus and flavivirus infectivity is disrupted by components of the bacterial cell wall

The impact of the host microbiota on arbovirus infections is currently not well understood. Arboviruses are viruses transmitted through the bites of infected arthropods, predominantly mosquitoes or ticks. The first site of arbovirus inoculation is the biting site in the host skin, which is colonized by a complex microbial community that could possibly influence arbovirus infection. We demonstrated that pre-incubation of arboviruses with certain components of the bacterial cell wall, including lipopolysaccharides (LPS) of some Gram-negative bacteria and lipoteichoic acids or peptidoglycan of certain Gram-positive bacteria, significantly reduced arbovirus infectivity in vitro. This inhibitory effect was observed for arboviruses of different virus families, including chikungunya virus of the Alphavirus genus and Zika virus of the Flavivirus genus, showing that this is a broad phenomenon. A modest inhibitory effect was observed following incubation with a panel of heat-inactivated bacteria, including bacteria residing on the skin. No viral inhibition was observed after pre-incubation of cells with LPS. Furthermore, a virucidal effect of LPS on viral particles was noticed by electron microscopy. Therefore, the main inhibitory mechanism seems to be due to a direct effect on the virus particles. Together, these results suggest that bacteria are able to decrease the infectivity of alphaviruses and flaviviruses. ImportanceDuring the past decades the world has experienced a vast increase in epidemics of alphavirus and flavivirus infections. These viruses can cause severe diseases such as hemorrhagic fever, encephalitis and arthritis. Several alpha- and flaviviruses, such as chikungunya virus, Zika virus and dengue virus, are significant global health threats because of their high disease burden, their widespread (re-)emergence and the lack of (good) anti-arboviral strategies. Despite the clear health burden, alphavirus and flavivirus infection and disease are not fully understood. A knowledge gap in the interplay between the host and the arbovirus is the potential interaction with host skin bacteria. Therefore, we studied the effect of (skin) bacteria and bacterial cell wall components on alphavirus and flavivirus infectivity in cell culture. Our results show that certain bacterial cell wall components markedly reduced viral infectivity by directly interacting with the virus particle.

microbiology↗

bioRxiv2021

Drug development of an affinity enhanced, broadly neutralizing heavy chain only antibody that restricts SARS-CoV-2 in hamsters

We have identified camelid single-domain antibodies (VHHs) that cross-neutralize SARS-CoV-1 and -2, such as VHH72, which binds to a unique highly conserved epitope in the viral receptor-binding domain (RBD) that is difficult to access for human antibodies. Here, we establish a protein engineering path for how a stable, long-acting drug candidate can be generated out of such a VHH building block. When fused to human IgG1-Fc, the prototype VHH72 molecule prophylactically protects hamsters from SARS-CoV-2. In addition, we demonstrate that both systemic and intranasal application protects hACE-2-transgenic mice from SARS-CoV-2 induced lethal disease progression. To boost potency of the lead, we used structure-guided molecular modeling combined with rapid yeast-based Fc-fusion prototyping, resulting in the affinity-matured VHH72_S56A-Fc, with subnanomolar SARS-CoV-1 and -2 neutralizing potency. Upon humanization, VHH72_S56A was fused to a human IgG1 Fc with optimized manufacturing homogeneity and silenced effector functions for enhanced safety, and its stability as well as lack of off-target binding was extensively characterized. Therapeutic systemic administration of a low dose of VHH72_S56A-Fc antibodies strongly restricted replication of both original and D614G mutant variants of SARS-CoV-2 virus in hamsters, and minimized the development of lung damage. This work led to the selection of XVR011 for clinical development, a highly stable anti-COVID-19 biologic with excellent manufacturability. Additionally, we show that XVR011 is unaffected in its neutralizing capacity of currently rapidly spreading SARS-CoV-2 variants, and demonstrate its unique, wide scope of binding across the Sarbecovirus clades.

molecular biology↗