Biology subjects

Halle, S.

Publications and source records attributed to Halle, S..

5 recordsLinked to original sources

bioRxiv2026

Single cell in vivo analysis of type I IFN and NK cell-mediated control of B cell infection densities during acute gammaherpesvirus infection

Immunodeficient patients are at risk of severe complications following infection with the human gammaherpesviruses Epstein-Barr virus (EBV) and Kaposis sarcoma-associated herpesvirus (KSHV). Due to the strict host specificity of human gammaherpesviruses, murine gammaherpesvirus 68 (MHV68) is widely used as an in vivo model to study gammaherpesvirus pathogenesis. While type I interferons (IFNs) and natural killer (NK) cells are known to contribute to antiviral defense during MHV68 infection, how these innate immune mechanisms control infection within lymphoid tissues at the level of individual infected cells remains incompletely understood. Here, we used an MHV68-DsRed reporter virus to visualize infected cells in the lymph nodes of wildtype and type I IFN receptor-deficient (Ifnar1-/-) mice by flow cytometry, immunohistochemistry, and two-photon microscopy. We found that type I IFN signaling plays a major role in limiting both local viral infection in draining lymph nodes and systemic dissemination to the spleen during acute infection. In the absence of IFNAR signaling, infected B cells accumulated to higher numbers, including an increased proportion of infected germinal center phenotype B cells, and this was accompanied by enhanced activation of T and B lymphocytes. Using two-photon microscopy, we further examined NK cell behavior within infected lymph nodes. NK cells were rapidly recruited to sites of infection but did not form stable clusters or prolonged contacts with infected cells. Nevertheless, NK cell depletion resulted in increased numbers of infected cells, indicating that NK cells contribute to the control of acute MHV68 infection despite the absence of detectable long-lasting interactions with infected cells. Together, our findings provide a single-cell view of acute gammaherpesvirus infection and innate immune control within lymph nodes in vivo. These data refine our understanding of how type I IFN responses and NK cells restrict early gammaherpesvirus spread and shape infection dynamics within lymphoid tissues. Author SummaryCertain viruses can remain in the body for life and cause little harm in healthy individuals. However, when the immune system is weakened, these infections can lead to cancer or other life-threatening diseases. To study human gammaherpesviruses within the living body, scientists often use a closely related mouse virus to understand how the immune system controls these infections. In this study, we tracked virus-infected cells within lymph nodes, important sites of the immune responses. Using a fluorescent virus together with advanced imaging techniques, we visualized infected cells directly in intact tissues. We found that type I interferons, a key component of the early antiviral defense, strongly limited the number of infected cells. Mice that could not respond to type I interferons developed substantially larger infections. We also investigated the role of natural killer cells, immune cells that provide rapid protection against viral infections. Although these cells quickly accumulated at sites of infection, they rarely formed prolonged contacts with infected cells. Together, our findings provide a detailed view of how innate immune defenses restrict gammaherpesvirus infection within lymph nodes. By revealing how early immune responses limit viral spread, this work improves our understanding of mechanisms that protect against severe herpesvirus-associated disease.

immunology↗

bioRxiv2025

Counterproductive: coinfection of a water flea by a fungus and a microsporidium reduces the reproductive outputs of all parties.

Organisms are often coinfected by more than one parasite species. These coinfections can alter the fitness of both host and its coinfecting parasites. Changes in host fitness are usually estimated by comparing parasite virulence under coinfection to the virulence of each parasite under single infection. While many studies focus on host survival as proxies of virulence, virulence can be expressed via reduced host fecundity. Here, we studied the outcome of coinfection of water fleas (Daphnia magna) by two microparasitic species, the fungus Metschnikowia bicuspidata and the microsporidium Hamiltosporidium tvaerminnensis. We found that Metschnikowia expressed its virulence mainly via host mortality and Hamiltosporidium expressed it mainly via host reproduction. Despite their competitive interaction, indicated by their reduced spore production, both parasites were able to fully express their virulence during coinfection. Host survival was dominated by Metschnikowia while clutch-size was dominated by Hamiltosporidium. Furthermore, coinfected hosts experienced increased virulence that was manifested only by additional reduction in reproduction (less clutches were released). Our results demonstrate that focusing only on the survival component of host fitness can miss important outcomes of coinfections. Since reduced host fecundity is a common outcome of parasitism, the influence of coinfection on host reproduction deserves more attention.

evolutionary biology↗

bioRxiv2024

Herpes Simplex Virus Assembly and Spread in murine skin after infection from the outside

Herpes simplex viruses (HSV) cause many skin diseases, particularly in immunocompromised patients. HSV-1 infection of murine skin recapitulates many aspects of human pathology. However, many protocols rely on mechanical or enzymatic skin disruption to induce lesions, although this can alter skin homeostasis and prime antiviral inflammation before inoculation. To investigate the initial events following HSV-1 primary skin infection before the onset of symptoms, we developed a novel murine ex vivo explant model using gentle depilation but no further scarification and infected keratinocytes from the outside with minimal tissue damage. Two-photon microscopy studies showed that HSV-1 spread exclusively in the epidermis. The infection centers increased in number and size over time and contained hundreds of infected keratinocytes. We investigated the HSV-1 spread at the cellular level, using reporter strains with fluorescently-tagged capsid protein VP26, and monitored the formation of nuclear capsid assembly sites, nuclear capsid egress, and the recruitment of the inner tegument protein pUL37GFP, the outer tegument protein VP11/12GFP, and the envelope protein gDGFP to cytoplasmic capsids. By electron microscopy, the skin appeared intact, and keratinocytes contained many nuclear capsids, primary virions in the nuclear envelope, cytosolic membrane-associated capsids, and enveloped virions. Our protocol provides a robust and reproducible approach to investigate the very early events of HSV-1 spread in the skin, to characterize the phenotypes of HSV-1 mutants in terminally differentiated skin tissues, and to evaluate potentially antiviral small molecules in a preclinical ex vivo infection model. IMPORTANCEThis study describes a novel murine ex vivo skin explant model to investigate early events in HSV-1 infection without causing significant tissue damage. To infect from the outside, via the apical keratinocytes, this method relies on gentle depilation, which maintains skin integrity. HSV-1 spread exclusively within the epidermis, with infection centers growing over time and involving hundreds of keratinocytes. Using advanced microscopy techniques, we tracked HSV-1 spread at the cellular level and intracellular assembly of all intermediate virus structures. This model offers a valuable tool for studying the initial stages of HSV-1 infection, assessing viral mutant phenotypes, and testing antiviral compounds in a more physiological context to provide critical insights into HSV-1 pathogenesis and therapeutic strategies.

microbiology↗

bioRxiv2024

Protective function of ex vivo expanded CD8 T cells in a mouse model of adoptive therapy for cytomegalovirus infection depends on integrin beta 1 but not CXCR3, CTLA4, or PD-1 expression

The adoptive transfer of virus-specific T cells (VSTs) represents a therapeutic option for viral infection treatment in immunocompromised patients. Before administration, ex vivo culture enables VST expansion. However, it is unclear how ex vivo expansion affects the circulation, homing, and intra-tissue migration of administered VSTs. We established a model of VST immunotherapy of acute cytomegalovirus infection using adoptive transfer of ex vivo expanded OT-I CD8 T cells (recognizing SIINFEKL peptide) into Rag2-/- mice infected with murine cytomegalovirus (MCMV) encoding for the SIINFEKL peptide. Ex vivo expansion induced an effector T cell phenotype and affected the expression of integrins and chemokine receptors. CRISPR/Cas9-mediated gene deletions enabled us to address the role of selected genes in the homing of VSTs following intravenous administration. We found that deletion of Itgb1, encoding for integrin beta 1, prevented OT-I cells from entering infected organs and drastically reduced their number in blood, suggesting that adoptively transferred VSTs primarily expand in the infected tissues. In contrast, Cxcr3-/- OT-I cells provided equal protection as their Cxcr3+/+ counterparts, indicating that this chemokine receptor does not contribute to VST entry into infected organs. Further, Pdcd1 and Ctla4 deletion did not impair the transferred OT-I cells ability to protect mice from MCMV, arguing against quick exhaustion of VSTs with an effector T cell phenotype. Together, these data indicate that ex vivo expansion affects migration and activation properties of VSTs and suggest that future clinical evaluation of adoptive T cell therapy efficacy should include homing molecule expression assessment.

immunology↗

bioRxiv2019

Evaluation of CD8 T cell killing models with computer simulations of 2-photon imaging experiments

In vivo imaging of cytotoxic T lymphocyte (CTL) killing activity revealed that infected cells have a higher observed probability of dying after multiple contacts with CTLs, suggesting memory effect in CTLs or infected cells. We developed a three-dimensional agent-based model of CTL killing activity to discriminate different hypotheses about how infected cells get killed based on quantitative 2-photon in vivo observations. We compared a constant CTL killing probability with mechanisms of signal integration in CTL or infected cells. The most likely scenario implied increased susceptibility of infected cells with increasing number of CTL contacts where the total number of contacts was a critical factor as opposed to signal integration over many contacts. However, when allowing in silico T cells to interact with apoptotic target cells (zombie contacts), a contact history independent killing mechanism was also in agreement with the experimental datasets. We showed that contacts that take place between CTLs and dying infected cells impact the observed killing dynamics because even in absence of modulation of cell properties, we saw an increase of the observed probability of killing infected cells with more interactions. The duration taken by an infected cell to die and the per capita killing rate (PCKR) of CTLs, parameters hard to measure directly, were determined from the model and turned out predictive to distinguish the different CTL killing models in future experiments. The comparison of observed datasets to simulation results, revealed limitations in interpreting 2-photon data, and provided prediction for additional measurements to distinguish CTL killing models. HighlightsO_LIKilling of infected cells by cytotoxic T cells typically involves more than a single contact. C_LIO_LICytotoxic T cells or infected cells integrate signals from multiple interactions. C_LIO_LIT cell contacts with dying infected cells have a major impact on in vivo data interpretation. C_LI Significance StatementDespite having a clear understanding of cytotoxic T lymphocyte (CTL) mediated cytotoxicity mechanisms, the quantitative dynamics remain unexplored at a cellular level. We developed an agent-based model to compare different hypotheses for mechanisms of CTL mediated cytotoxicity that could lead to an increase in observed probability of killing infected cells at higher interactions with CTLs as seen in vivo. We showed that this behaviour can be explained by modulation of properties by infected cells or CTLs with increasing number of contacts. For the modulation, we compared two modes of signal integration and showed that time is not a relevant parameter in signal integration. We also studied the impact of contacts between CTLs and apoptotic infected cells on observed killing properties.

systems biology↗