Data on the bioluminescence, cell viability, and biofilm formation of Photobacterium leiognathi strains exposed to ground microplasticss

Description

ABSTRACT This dataset provides comprehensive measurements of bioluminescence, cell viability, and biofilm formation in Photobacterium leiognathi strains (LB01 and LB09) exposed to varying concentrations of ground microplastics (GMPs). The study aims to investigate the effects of GMPs exposure on these critical bacterial functions, offering insights into the environmental impact of MPs on marine luminous bacteria. DATA DESCRIPTION The bioluminescence data, which expressed as specific bioluminescence (SBF), includes measurements recorded in Log relative light units (Log RLUs) using Glomax Discover (Promega Instrument). These measurements were taken at multiple time intervals, such as every 5 h for 3.1 days (75 h) or 24 h for 7 days, to capture the temporal changes in bioluminescence activity during and after (post-exposure experiment) exposure to GMPs (Tables S1-S4). For cell viability, the data represented as optical density (ODs) at 560 nm. MTT assays were conducted to determine cell viability. The wells were added with MTT solution (5 mg/mL concentration), incubated for 8 h and dissolved with dimethyl sulfoxide. This analysis enabling an examination of the impact of GMPs on bacterial growth and survival (Tables S5-S6). Biofilm formation data was obtained by quantifying the biofilm biomass using 1% crystal violet solution. This involved measuring the ODs at 570 nm to determine the amount of biofilm produced by Phb. leiognathi under each condition. The dataset includes biofilm measurements for both the control (unexposed to GMPs) and experimental groups, corresponding to the same GMP concentrations used in the bioluminescence and cell viability experiments (Tables S7-S8). This dataset is valuable for researchers studying the environmental effects of microplastics on marine bacteria, particularly those interested in the functional responses of bioluminescent organisms to plastic pollution. The comprehensive data on bioluminescence, cell viability, and biofilm formation provides a robust foundation for understanding the ecophysiological effects of GMPs to Phb. leiognathi.

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1. GMP Exposure Experiment Preparation of Bacterial Cultures: Fresh cultures of Phb. leiognathi strains were grown to the exponential phase. Cells were harvested by centrifugation at 6000 × g for 10 min, washed twice with PBS (pH 7.4), and adjusted to an OD600 of 0.5–0.8 for immediate use. Experimental Setup: Bacteria were exposed to various GMP concentrations (0.25%, 0.50%, 1%, and 2% w/v) in ASW-YE-T broth. Control tubes contained broth without GMPs. A 10 µL aliquot of the bacterial solution was added to each tube, incubated at 22°C or 30°C while shaking at120 rpm. Bioluminescence Monitoring: Bioluminescence was measured to assess immediate effects every 5 h for 75 h and long-term effects every 24 h for 7 d, with double-strength ASW-YE-T broth added to prevent nutrient depletion. Measurements involved transferring 200 µL samples to black microtiter plates, covering them with foil, and allowing them to stand for 10 min. The experiment was repeated three times with triplicate wells for each condition. 2. Post-Exposure Recovery Experiment Procedure: Bacterial cultures were harvested, washed with PBS, resuspended in PBS with 20% glycerol, and stored at -20°C. For recovery, 10 µL of bacterial suspensions were transferred into ASW-YE-T broth and incubated for 35 h at 22°C or 30°C while shaking at 120 rpm. Bioluminescence Monitoring: Measurements were taken every 5 h during the 35-h recovery period by transferring 100 µL samples to black microtiter plates, covering with foil, and leaving undisturbed for 10 min. The experiment was conducted three times with triplicate wells for each condition. 3. MTT Assay for Cell Viability Preparation of MTT Solution: 5 mg/mL concentration of MTT powder was prepared and stored at -20°C. Experimental Procedure: Bacterial suspensions (3 µL) were inoculated into wells of a 96-well microtiter plate containing 100 µL of ASW broth, incubated at 22°C or 30°C for 8 h. After adding 10 µL of MTT solution and standing for 20 m, broth was removed, 100 µL of dimethyl sulfoxide added, and OD560 measured. OD values were corrected based on blanks, with twelve replicates for each condition. 4. Crystal Violet Biofilm Formation Assay Preparation of Microtiter Plates: Wells of a 96-well plate were coated with a 1% (w/v) squid’s ink solution, incubated overnight, and filled with double-strength ASW-YE-T broth. Biofilm Formation Procedure: Bacterial suspensions (3 µL) were inoculated into the wells and incubated at 30°C or 22°C while shaking at 150 rpm for 48 h. Broth was replenished with fresh ASW-YE-T and incubated again. Staining and Measurement: Wells were washed with PBS, stained with 1% (v/v) crystal violet solution, washed with distilled water, and solubilized with absolute ethanol. OD560 was measured to assess biofilm formation, with specific biofilm formation calculated by subtracting non-specific OD values and normalizing to bacterial growth in ASW-YE-T broth. The assay was performed in nine replicates for each condition.

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