Journal: bioRxiv

Article Title: Discovery of a new fibronectin-binding surface protein of Streptococcus canis with serum opacification activity through Transposon Directed Insertion-Site Sequencing

doi: 10.64898/2025.12.05.692515

Figure Lengend Snippet: A) Serum opacification activity of S. canis strain IMT49926 and 49926Δ sof , S. suis M10 and a negative serum control. Showing (left to right): overnight incubation with heat-inactivated bacterial supernatant, heatinactivated bacterial sediment suspension from logarithmic growing bacteria and 1% SDS extracts of overnight cultures. B) Radius of the haemolytic zone measured with ImageJ based on pictures of blood agar plates of 3 biological replicates comparing IMT49926 and 49926Δ sof every time on the same plate. Statistical significance was calculated by Two-way ANOVA. *** indicates p < 0.001 and **** indicates p < 0.0001. C) Representative picture of a blood agar plate with colonies of S. canis IMT49926 (left) and 49926Δ sof (right).

Article Snippet: Primary HUVECs (PromoCell, Germany) were cultured in endothelial cell growth medium (C-22010, PromoCell, Germany) and infected with mid-log phase S. canis TraDIS library, S. canis WT or ScSOF knockout strains at multiplicity of infection (MOI) 10.

Techniques: Activity Assay, Control, Incubation, Suspension, Bacteria

Journal: bioRxiv

Article Title: Discovery of a new fibronectin-binding surface protein of Streptococcus canis with serum opacification activity through Transposon Directed Insertion-Site Sequencing

doi: 10.64898/2025.12.05.692515

Figure Lengend Snippet: A) S. canis IMT49926 and 49926Δ sof stained with specific antibodies (green) on a fibronectin-coated coverslip fixed at different timepoints. Images are acquired with a DMI6000B fluorescence microscope. Scale bars indicate 10 µm. B) Area covered with bacteria (determined by ImageJ analysis of images of three independent experiments. C) FACS analysis of the direct fibronectin-binding ability of IMT49926 and 49926Δ sof using FITC-conjugated fibronectin. Data represents geometric mean fluorescence intensity (MFI) ± SD of three independent experiments.

Article Snippet: Primary HUVECs (PromoCell, Germany) were cultured in endothelial cell growth medium (C-22010, PromoCell, Germany) and infected with mid-log phase S. canis TraDIS library, S. canis WT or ScSOF knockout strains at multiplicity of infection (MOI) 10.

Techniques: Staining, Fluorescence, Microscopy, Bacteria, Binding Assay

Journal: bioRxiv

Article Title: Discovery of a new fibronectin-binding surface protein of Streptococcus canis with serum opacification activity through Transposon Directed Insertion-Site Sequencing

doi: 10.64898/2025.12.05.692515

Figure Lengend Snippet: A) Endothelial cell morphology was visualized by the detection of actin cytoskeleton with Alexa Fluor 488-conjugated phalloidin in combination with DAPI stain of nuclei at time points 6 h, 24 h and 48 h. Representative overlay images are shown for each time point of the CSMA without bacteria (ctrl), and in the presence of WT S. canis or 49926Δ sof . Images were generated using the CLSM (Leica Sp8) at 200-fold magnification and a zoom factor of 0.75. White arrows mark the direction of flow. Scale bar represents 100 μm. B) Representative microscopic images of CSMA results at 630-fold magnification. Blue arrows point to cell evaginations at the growing cell borders (ctrl). Yellow arrows point to adherent S. canis , and red arrows point to internalized bacteria. White arrows mark the direction of flow. Images generated using CLSM (Leica Sp8) at 630-fold magnification and a zoom factor of 0.75. Scale bar represents 10 μm. C) At indicated timepoints, progress of gap closure was calculated in percent in relation to the starting point, which is defined as 0% gap closure. Values are expressed as mean values with standard deviation. * Indicates p < 0.05, ** indicates p < 0.01, and *** indicates p < 0.001, according to one-way ANOVA and Tukey HSD post hoc tests. For the 24 h and 48 h samples homoscedasticity was not met (Levene p < 0.05); therefore, Welch’s ANOVA and Games-Howell post hoc testing were applied.

Article Snippet: Primary HUVECs (PromoCell, Germany) were cultured in endothelial cell growth medium (C-22010, PromoCell, Germany) and infected with mid-log phase S. canis TraDIS library, S. canis WT or ScSOF knockout strains at multiplicity of infection (MOI) 10.

Techniques: Staining, Bacteria, Generated, Standard Deviation