Upon invasion into the host cell, a subset of bacterial pathogens resides exclusively in the cytosol. While previous research revealed how they reshape the plasma membrane during invasion, subvert the immune response, and hijack cytoskeletal dynamics to promote their motility, it was unclear if these pathogens also interacted with the organelles in this crowded intracellular space. Here, we examined if the obligate intracellular pathogen Rickettsia parkeri interacts with the endoplasmic reticulum (ER), a large and dynamic organelle spread throughout the cell. Using live-cell microscopy and transmission and focused-ion-beam scanning electron microscopy, we show that R. parkeri forms extensive contacts with the rough ER that are ∼55 nm apart and cover more than half the bacterial surface. Depletion of the ER-specific tethers VAPA and VAPB reduced rickettsia–ER contacts, and VAPA and VAPB were localized around intracellular rickettsiae. Overall, our findings illuminate an interkingdom ER contact uniquely mediated by rickettsiae that mimics some characteristics of traditional host membrane contact sites.

Interorganelle communication regulates cellular homeostasis through the formation of tightly-associated membrane contact sites 1-3. Prior work has identified several ways that intracellular pathogens alter contacts between eukaryotic membranes 4-6, but there is no existing evidence for contact sites spanning eukaryotic and prokaryotic membranes. Here, using a combination of live-cell microscopy and transmission and focused-ion-beam scanning electron microscopy, we demonstrate that the intracellular bacterial pathogen Rickettsia parkeri forms a direct membrane contact site between its bacterial outer membrane and the rough endoplasmic reticulum (ER), with tethers that are approximately 55 nm apart. Depletion of the ER-specific tethers VAPA and VAPB reduced the frequency of rickettsia-ER contacts, suggesting these interactions mimic organelle-ER contacts. Overall, our findings illuminate a direct, interkingdom membrane contact site uniquely mediated by rickettsia that seems to mimic traditional host MCSs.

Acoustic overexposure can eliminate synapses between inner hair cells (IHCs) and auditory nerve fibers (ANFs), even if hair-cell function recovers. This synaptopathy has been extensively studied by confocal microscopy, however, understanding the nature and sequence of damage requires ultrastructural analysis. Here, we used focused ion-beam scanning electron microscopy to mill, image, segment and reconstruct ANF terminals in mice, 1 day and 1 week after synaptopathic exposure (8–16 kHz, 98 dB SPL). At both survivals, ANF terminals were normal in number, but 62% and 53%, respectively, lacked normal synaptic specializations. Most non-synapsing fibers (57% and 48% at 1 day and 1 week) remained in contact with an IHC and contained healthy-looking organelles. ANFs showed a transient increase in mitochondrial content (51%) and efferent innervation (34%) at 1 day. Fibers maintaining synaptic connections showed hypertrophy of pre-synaptic ribbons at both 1 day and 1 week. Non-synaptic fibers were lower in mitochondrial content and typically on the modiolar side of the IHC, where ANFs with high-thresholds and low spontaneous rates are normally found. Even 1 week post-exposure, many ANF terminals remained in IHC contact despite loss of synaptic specializations, thus, regeneration efforts at early post-exposure times should concentrate on synaptogenesis rather than neurite extension.

Journal Article From Archeology to the Malaria Parasite, the Exciting Quests of Microscopy Get access David C Bell, David C Bell Harvard John A. Paulson School of Engineering and Applied Sciences, Harvard University, Cambridge, MA, USA Corresponding author: dcb@seas.harvard.edu Search for other works by this author on: Oxford Academic Google Scholar Hao-Yu Greg Lin, Hao-Yu Greg Lin Center for Nanoscale Systems, Harvard University, Cambridge, MA, USA Search for other works by this author on: Oxford Academic Google Scholar Austin Akey, Austin Akey Center for Nanoscale Systems, Harvard University, Cambridge, MA, USA Search for other works by this author on: Oxford Academic Google Scholar Stephan Kraemer, Stephan Kraemer Center for Nanoscale Systems, Harvard University, Cambridge, MA, USA Search for other works by this author on: Oxford Academic Google Scholar Jeffrey T Borenstein, Jeffrey T Borenstein Draper Laboratory, Cambridge, MA, USAResearch groups thereof Search for other works by this author on: Oxford Academic Google Scholar Jeffrey D Dvorin, Jeffrey D Dvorin Boston Children’s Hospital, Boston, MA, USAResearch groups thereof Search for other works by this author on: Oxford Academic Google Scholar Angela Chang Angela Chang Harvard Art Museums, Harvard University, Cambridge, MA, USAResearch groups thereof Search for other works by this author on: Oxford Academic Google Scholar Microscopy and Microanalysis, Volume 29, Issue Supplement_1, 1 August 2023, Pages 86–87, https://doi.org/10.1093/micmic/ozad067.035 Published: 22 July 2023

New techniques for obtaining electron microscopy data through the cell volume are being increasingly utilized to answer cell biologic questions. Here, we present a three-dimensional atlas of Plasmodium falciparum ultrastructure throughout parasite cell division. Multiple wild type schizonts at different stages of segmentation, or budding, were imaged and rendered, and the 3D structure of their organelles and daughter cells are shown. Our high-resolution volume electron microscopy both confirms previously described features in 3D and adds new layers to our understanding of Plasmodium nuclear division. Interestingly, we demonstrate asynchrony of the final nuclear division, a process that had previously been reported as synchronous. Use of volume electron microscopy techniques for biological imaging is gaining prominence, and there is much we can learn from applying them to answer questions about Plasmodium cell biology. We provide this resource to encourage readers to consider adding these techniques to their cell biology toolbox.

During the blood stage of human malaria, Plasmodium falciparum parasites divide by schizogony-a process wherein components for several daughter cells are produced within a common cytoplasm and then segmentation, a synchronized cytokinesis, produces individual invasive daughters. The basal complex is hypothesized to be required for segmentation, acting as a contractile ring to establish daughter cell boundaries. Here we identify an essential component of the basal complex which we name PfCINCH. Using three-dimensional reconstructions of parasites at electron microscopy resolution, we show that while parasite organelles form and divide normally, PfCINCH-deficient parasites develop inviable conjoined daughters that contain components for multiple cells. Through biochemical evaluation of the PfCINCH-containing complex, we discover multiple previously undescribed basal complex proteins. Therefore, this work provides genetic evidence that the basal complex is required for precise segmentation and lays the groundwork for a mechanistic understanding of how the parasite contractile ring drives cell division.

Recent studies predict that LaPtBi is a half-Heusler has multi-functionalities: the superconductivity and topological edge states, a namely topological superconductor under certain conditions. It can be stimulated under substantial uniaxial strain conditions. The superconductivity in LaPtBi has been shown in bulk. However, the topological superconductive surface state can not be observed yet. In this study, we report the transmission electron microscopy work for observation of superconductivity and its relationship with a uniaxial strain in MBE grown epitaxial non-centrosymmetric LaPtBi film on MgO (001).

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