Abstract:

This dataset includes microscopy videos showing how the motility of hydrocarbon-degrading bacteria is influenced by increased hydrostatic pressure. To obtain this data, a high-pressure microscope chamber (described in Nishiyama and Sowa, 2012, Biophysical Journal) was used. Bacteria were grown in Marine Broth 2216 at 23C and 5C until late log phase, then transferred to the microscope chamber (set to 23C and 7C, respectively). 7C was used (instead of 5C) as it is as cold as we can get the system without compromising data quality. The closed high-pressure chamber was hooked up to a phase-contrast microscope with video capabilities. Two types of experiments were performed: 1) Pressure was incrementally increased by 20 MPa, and 20-second videos of motility were taken at each pressure. Once motility appeared to cease, the system was depressurized incrementally by 20 MPa, and videos were again taken; 2) The samples were pressurized to high pressure and left there for 2 minutes. Then, the pressure was released to atmospheric pressure (0.1 MPa) and the recovery of motility after pressure exposure was recorded.

Suggested Citation:

Kelli Mullane, Douglas Bartlett, Masayoshi Nishiyama. 2020. Microscopy videos of microbial motility at high hydrostatic pressure. Distributed by: GRIIDC, Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/SBJYTWB6

Purpose:

The goal of this work was to assess pressure effects on the motility of hydrocarbon-degrading bacteria.

Data Parameters and Units:

The dataset contains 2 Excel files and several sub-folders of video files with microscopy videos showing how the motility of hydrocarbon-degrading bacteria is influenced by increased hydrostatic pressure. The file "XY Microscope Calibration.xlsx" contains the calibration information organized under different worksheets named as per the date. The file “High-Pressure Microscopy Notes.xlsx” include the information about the experiment as named on the individual spreadsheets - OD (optical density) Measurements, Motility Checks, High-Pressure Microscopy, Number of HP Experiments, and 7C High-Pressure Microscopy. The worksheets “High-Pressure Microscopy and 7C High-Pressure Microscopy” describe the experimental conditions under which videos were taken. Experimental conditions such as pressure and room temperature are linked with the folders of videos by the filename which has the format-YY-MM-DD HH-MM-SS. These worksheet includes - Strain, Culture Start Date (MM/DD/YYYY), Date of Microscopy (MM/DD/YYYY), Experimental Temperature (Celsius), File Name (YY-MM-DD HH-MM-SS), Pressure (MPa), NOTES (experimental details), HP Chamber, and HP Cap, and Light Filter Wavelength (nm). The worksheet "OD Measurements" contains: Strain, OD660 (measured), OD600 (calculated), Calibration Medium (aka Growth Medium), Date of Inoculation (MM/DD/YYYY), Date of Measurement (MM/DD/YYYY), and NOTES (experimental details). The worksheet "Motility Checks" contains: Strain, File Name (YY-MM-DD HH-MM-SS), Room Temp. (Celsius), Culture Start Date (MM/DD/YYYY), Date of Microscopy (MM/DD/YYYY), and NOTES (experimental details). The worksheet "Number of HP Experiments" contains: Strain, #1 Date (MM/DD/YYYY), #2 Date (MM/DD/YYYY), #3 Date (MM/DD/YYYY), #4 Date (MM/DD/YYYY), #5 Date (MM/DD/YYYY), and #6 Date (MM/DD/YYYY). Microscope Settings: 40x objective lens, 1x eye-piece, Ph2 phase contrast. Camera visual rate = 30 frames/second. Camera calibration = 0.24 um/pixel.

Methods:

Bacteria were grown in Marine Broth 2216 at 23C and 4C until late log phase, then transferred to the microscope chamber (set to 23C and 7C, respectively). 7C was used as it is as cold as we can get the system without compromising data quality. The closed high-pressure chamber was hooked up to a phase-contrast microscope with video capabilities. Two types of experiments were performed - both described below. 1) For the first type of experiment, the pressure was incrementally increased by 20 MPa, and 20-second videos of motility were taken at each pressure. Once motility appeared to cease, the system was depressurized incrementally by 20 MPa, and videos were again taken. 2) For the second type of experiment, the samples were pressurized to high pressure and left there for 2 minutes. Then, the pressure was released to atmospheric pressure (0.1 MPa) and the recovery of motility after pressure exposure was recorded.

Instruments:

Microscope used: Nikon Eclipse Ti Camera used: WAT-120N+

Provenance and Historical References:

Nishiyama, M., & Sowa, Y. (2012). Microscopic Analysis of Bacterial Motility at High Pressure. Biophysical Journal, 102(8), 1872–1880. doi:10.1016/j.bpj.2012.03.033

Individual Files: