DOI: 10.57745/ghnfyi
title: Structural biofilm characterisation of 40 Tenacibaculum spp. isolates

description: The establishment of comprehensive datasets on Tenacibaculum spp. biofilms is essential for understanding the environmental persistence and potential pathogenesis of these marine bacteria within aquaculture systems. To characterise the growth strategies of 40 clinical and environmental Tenacibaculum spp. isolates, we present a structured phenotypic dataset across three distinct growth models (planktonic, macro-colony biofilms, and submerged biofilms). Datasets from these three models are represented by the repository in three primary folders: "Tenaci_Planktonic data", "Tenaci_Macro-colony data", and "Tenaci_Confocal data". To study planktonic growth, kinetic OD600 data were acquired every 20 minutes over a 72-hour period using 96-well plates. These datasets, including both the initial readings and the annotated versions, are stored in the "Planktonic_raw data" and "Planktonic_annotated data" subfolders within the "Tenaci_Planktonic data" primary folder. Macro-colony biofilm model was used to study bacterial colony growth at the surface/air interface. Bacteria were cultivated on filtered marine agar within 12-well plates and incubated under static conditions; the Reshape Biotech imaging platform (https://www.reshapebiotech.com/) captured images hourly for 7 days. The platform documented physical expansion and morphological differentiation using colony area measurements generated via deep learning algorithms. These datasets, images and videos (timelapses) are stored into the "Macro-colony_images" and "Macro-colony_raw data" subfolders of the "Tenaci_Macro-colony data" primary folder. Submerged biofilms growing at the surface/liquid interface were studied using a confocal laser scanning microscope (CLSM) Leica SP8 AOBS (https://doi.org/10.15454/1.5572348210007727E12). Prior to acquisition, biofilms were stained with SYTO9 for nucleic acid visualisation and Congo red to contrast part of the extracellular polymeric substances (EPS). Acquisitions were performed with a 63x water-immersion objective and z-stacks at 1 µm intervals. Both dyes were excited at 488 nm. Emitted fluorescence was collected from 500 to 550 nm for SYTO9 and from 600 to 750 nm for Congo red (CR). Images have a resolution of 512 × 512 pixels, with a pixel size of 0.361 µm and support the quantification of bacterial biomass and CR-labelled EPS. These high-resolution 3D image acquisitions are provided in the "Tenaci_Confocal data" primary folder, organised into time-resolved subfolders for T0h, T24h, and T72h. Each of these subfolders contains individual strain folders where the raw .lif files are stored. To navigate the 40-strain collection, a "Table 1_Tenaci stock list" Excel file is included in every primary folder to map alphanumeric codes to metadata (species name, pathogenicity, and origin). Additional "Plate Plan" or "Plate Map" Excel files define the spatial orientation of high-throughput assays by identifying the specific well assignment for each strain or replicate within the 96-well and 12-well plate formats. For every experiment, at least three biological replicates were acquired; for the confocal imaging specifically, each biological replicate included four technical replicates, representing twelve technical values per condition. These biological replicates are identified by the suffixes x, y, and z (e.g., 20x, 20y, and 20z) within the file naming convention to ensure data traceability and robust comparative analysis.