Spatiotemporal analysis of host cell modification during herpes simplex virus 1 replication

Scherer, Katharina M.; Manton, James D.; Soh, Timothy K.; Mascheroni, Luca; Connor, Viv; Crump, Colin M.; Kaminski, Clemens F.

doi:10.1101/2019.12.22.872002

Cited by 3 publications

(3 citation statements)

References 41 publications

(44 reference statements)

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“…The dose dependence on viral gene expression and productive replication aligns with other work that observed dosedependent HSV-1 replication in human foreskin fibroblasts (40). Similarly, the effects of lower inoculating dose leading to slower kinetics of expression and low rates of productive replication have been observed during HSV-1 infection of other non-neuronal cells (33,40). Notably, we were able to observe single-cell infection of neurons for longer observation periods.…”

Section: Discussionsupporting

confidence: 88%

“…Importantly, all RFP positive neurons were detectably YFP positive before 6 hpi, suggesting that early viral transcription is more likely to elicit productive viral replication. The dose dependence on viral gene expression and productive replication aligns with other work that observed dosedependent HSV-1 replication in human foreskin fibroblasts (40). Similarly, the effects of lower inoculating dose leading to slower kinetics of expression and low rates of productive replication have been observed during HSV-1 infection of other non-neuronal cells (33,40).…”

Section: Discussionsupporting

confidence: 85%

See 1 more Smart Citation

Single-cell Herpes Simplex Virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

Fredrikson,

Domanico,

Pratt

et al. 2023

Preprint

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Single-cell analyses of viral infections often reveal heterogeneity that is not detected by traditional population-level studies. This study applies drop-based microfluidics to investigate the dynamics of HSV-1 infection of neurons at the single-cell level. We used micron-scale Matrigel beads, termed microgels, to culture individual murine Superior Cervical ganglia (SCG) neurons or epithelial cells. Microgel-cultured cells are subsequently enclosed in individual media-in-oil droplets with a dual fluorescent-reporter HSV-1, enabling real-time observation of viral gene expression and replication. Infection within drops revealed that the kinetics of initial viral gene expression and replication were dependent on the inoculating dose. Notably, increasing inoculating doses led to earlier onset of viral gene expression and more frequent productive viral replication. These observations provide crucial insights into the complexity of HSV-1 infection in neurons and emphasize the importance of studying single-cell outcomes of viral infection. The innovative techniques presented here for cell culture and infection in drops provide a foundation for future virology and neurobiology investigations.

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Section: Discussionsupporting

confidence: 88%

Section: Discussionsupporting

confidence: 85%

Single-cell Herpes Simplex Virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

Fredrikson,

Domanico,

Pratt

et al. 2023

Preprint

View full text Add to dashboard Cite

show abstract

“…Expansion microscopy is a technique that has not yet been widely explored in virus research [ 40 ]. However, we show that it provides unprecedented detail on the interaction and localisation of virus particles with subcellular organelles.…”

Section: Discussionmentioning

confidence: 99%

Combining sample expansion and light sheet microscopy for the volumetric imaging of virus-infected cells with super-resolution

Mascheroni

Scherer

Manton

et al. 2020

Biomed. Opt. Express

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Expansion microscopy is a sample preparation technique that enables the optical imaging of biological specimens at super-resolution owing to their physical magnification, which is achieved through water-absorbing polymers. The technique uses readily available chemicals and does not require sophisticated equipment, thus offering super-resolution to laboratories that are not microscopy-specialised. Here we present a protocol combining sample expansion with light sheet microscopy to generate high-contrast, high-resolution 3D reconstructions of whole virus-infected cells. The results are superior to those achievable with comparable imaging modalities and reveal details of the infection cycle that are not discernible before expansion. An image resolution of approximately 95 nm could be achieved in samples labelled in 3 colours. We resolve that the viral nucleoprotein is accumulated at the membrane of vesicular structures within the cell cytoplasm and how these vesicles are positioned relative to cellular structures. We provide detailed guidance and a video protocol for the optimal application of the method and demonstrate its potential to study virus-host cell interactions.

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Combining sample expansion and light sheet microscopy for the volumetric imaging of virus-infected cells with optical super-resolution

Mascheroni

Scherer

Ward

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

Expansion microscopy is a sample preparation technique that enables the optical imaging of biological specimens at super-resolution owing to their physical magnification, which is achieved through water-absorbing polymers. The technique uses readily available chemicals and does not require sophisticated equipment, thus offering super-resolution to laboratories that are not microscopy-specialised. Here we present a protocol combining sample expansion with light sheet microscopy to generate high-contrast, high-resolution 3D reconstructions of whole virus-infected cells. The results are superior to those achievable with comparable imaging modalities and reveal details of the infection cycle that are not discernible before expansion. An image resolution of approximately 95 nm could be achieved in samples labelled in 3 colours. We clearly resolve the concentration of viral nucleoprotein on the surface of vesicular structures within the cell and their positioning relative to cellular organelles. We provide detailed guidance and a video protocol for the optimal application of the method and demonstrate its potential to study virus-host cell interactions.

show abstract

Spatiotemporal analysis of host cell modification during herpes simplex virus 1 replication

Cited by 3 publications

References 41 publications

Single-cell Herpes Simplex Virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

Single-cell Herpes Simplex Virus type-1 infection of neurons using drop-based microfluidics reveals heterogeneous replication kinetics

Combining sample expansion and light sheet microscopy for the volumetric imaging of virus-infected cells with super-resolution

Combining sample expansion and light sheet microscopy for the volumetric imaging of virus-infected cells with optical super-resolution

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