Work Description

Title: Raw microscopy data for Wolbachia infection of JW18 cells Open Access Deposited

https://creativecommons.org/licenses/by-nc/4.0/
Attribute Value
Abstract
  • Wolbachia pipientis is an incredibly widespread bacterial symbiont of insects, present in an estimated 25-52% of species worldwide. Wolbachia is faithfully maternally transmitted both in a laboratory setting and in the wild. In an established infection, Wolbachia is primarily intracellular, residing within host-derived vacuoles that are associated with the endoplasmic reticulum. However, Wolbachia also frequently transfer between host species, requiring an extracellular stage to their life cycle. Indeed, Wolbachia has been moved between insect species for the precise goal of controlling populations. The use of Wolbachia in this application requires we better understand how it initiates and establishes new infections. Here we designed a novel method for live-tracking Wolbachia during infection using a combination of stains and microscopy. We show that live Wolbachia are taken up by host cells at a much faster rate than dead Wolbachia cells, indicating that Wolbachia play a role in their own uptake and that Wolbachia colonization is not just a passive process. We also show that the host actin cytoskeleton must be intact for this to occur, and that drugs that disrupt the actin cytoskeleton effectively abrogate Wolbachia uptake. The development of this live infection assay will assist in future efforts to characterize Wolbachia factors used during host infection.
Methodology
  • Wolbachia isolated from infected JW18 cells were stained with a 1:1 mixture of SYTO9 (ThermoFisher) and Propidium Iodide (ThermoFisher) either immediately following isolation (live) or after storage at -20°C for at least 3 days prior to imaging (dead). Live Wolbachia were used straight away, without any long incubations and kept away from light prior to imaging using the same magnification, intensities, and exposure times. Microscopy and Analysis of Images All images were obtained using a Nikon Ti2 Eclipse inverted microscope at 100x magnification with a Nikon DS-Qi2 camera attachment. As noted above, cells were seeded at a density no greater than 2000 cells per well; this allowed us to more clearly delineate individual cells avoid cells clumps during the infection and made the image processing more smooth. Intensities and exposure times were as follows: green/FITC for 200ms with SOLA pad at 4%, red/Texas Red for 400ms with SOLA pad at 8%. All Z stacks were taken with a step size of 0.5μm with between 100-300 contiguous planes imaged per field, with identical settings applied across all fields, regardless of treatment and condition.
Depositor
Citations to related material
Resource type
Last modified
  • 06/20/2024
Language
Subject
License
Description sponsorship
  • NIH grant R01 AI144430
To Cite this Work:
Raw microscopy data for Wolbachia infection of JW18 cells [Data set]. Indiana University - DataCORE.

Relationships

Files (Count: 7; Size: 1.38 MB)

Thumbnail Title Original Upload Last Modified File Size Access Actions
Wolbachia_internalization.mp4 2025-05-19 2025-05-19 1.38 MB Open Access
Dead_Infection_A.zip 2025-05-19 2025-05-19 Open Access
Dead_Infection_B.zip 2025-05-19 2025-05-19 Open Access
Dead_Infection_C.zip 2025-05-19 2025-05-19 Open Access
Live_Infection_A.zip 2025-05-19 2025-05-19 Open Access
Live_Infection_B.zip 2025-05-19 2025-05-19 Open Access
Live_Infection_C.zip 2025-05-19 2025-05-19 Open Access

Download All Files

Some files in this dataset must be individually downloaded, and will not be included in the zip download:
Files can be downloaded individually in the "Files" panel above.

Best for data sets < 3 GB. Downloads all files plus metadata into a zip file.