Tailored approach to study Legionella infection using a lattice light sheet microscope (LLSM)
Xiyu Yi1,2, Haichao Miao3, Jacky Kai-Yin Lo1
1Physical and Life Science Directorate, Lawrence Livermore National Laboratory, Livermore, CA, 94550, USA.
Abstract:
Legionella is a genus of ubiquitous environmental pathogens found in freshwater systems, moist soil, and composted materials. More than four decades of Legionella research has provided important insights into Legionella pathogenesis. Although standard commercial microscopes have led to significant advances in understanding Legionella pathogenesis, great potential exists in the deployment of more advanced imaging techniques to provide additional insights. The lattice light sheet microscope (LLSM) is a recently developed microscope for 4D live cell imaging with high resolution and minimum photo-damage. We built a LLSM with an improved version for the optical layout with two path-stretching mirror sets and a novel reconfigurable galvanometer scanner (RGS) module to improve the reproducibility and reliability of the alignment and maintenance of the LLSM. We commissioned this LLSM to study Legionella pneumophila infection with a tailored workflow designed over instrumentation, experiments, and data processing methods. Our results indicate that Legionella pneumophila infection is correlated with a series of morphological signatures such as smoothness, migration pattern and polarity both statistically and dynamically. Our work demonstrates the benefits of using LLSM for studying long-term questions in bacterial infection. Our free-for-use modifications and workflow designs on the use of LLSM system contributes to the adoption and promotion of the state-of-the-art LLSM technology for both academic and commercial applications.
Insights
Lattice light sheet microscopy (LLSM) offers new insights into Legionella pathogenesis. This advanced imaging reveals distinct bacterial morphological signatures during infection, advancing our understanding of this pathogen.
Area of Science:
- Microbiology
- Cell Biology
- Biophysics
Background:
- Legionella is a significant environmental pathogen with complex pathogenesis.
- Traditional microscopy has limitations in studying dynamic bacterial infections.
- Advanced imaging techniques like LLSM hold potential for deeper insights.
Purpose of the Study:
- To develop and apply a customized Lattice Light Sheet Microscope (LLSM) for studying Legionella pneumophila infection.
- To investigate the dynamic morphological changes of Legionella during host cell infection using LLSM.
- To establish a workflow for LLSM-based bacterial infection studies.
Main Methods:
- Construction of an improved LLSM with enhanced optical layout and a novel reconfigurable galvanometer scanner (RGS) module.
- Development of a tailored workflow encompassing instrumentation, experimental procedures, and data processing for Legionella infection studies.
- Utilizing 4D live cell imaging capabilities of LLSM for high-resolution, low-photodamage observation.
Main Results:
- Legionella pneumophila infection is characterized by specific, identifiable morphological signatures.
- These signatures include changes in bacterial smoothness, migration patterns, and polarity.
- Statistical and dynamic analyses confirmed the correlation between morphological changes and infection progression.
Conclusions:
- LLSM is a powerful tool for investigating long-term bacterial infection dynamics.
- The developed LLSM modifications and workflow facilitate the study of bacterial pathogenesis.
- This approach promotes the adoption of advanced imaging technologies in microbiology research.
