Biofilm Formation by Clostridium ljungdahlii Is Induced by Sodium Chloride Stress: Experimental Evaluation and Transcriptome Analysis
A) Effect of enzyme treatment. C. ljungdahlii biofilms were grown in well plates by adding 200 mM NaCl to the medium. After 2 days of incubation, the supernatant of all wells was removed and the biofilms were washed twice with PBS buffer. PBS buffer without (control 1) or with 0.2 mg·mL-1 proteinaseK or reaction buffer (10 mM Tris HCl pH 7.5, 2.5 mM MgCl2, 0.5 mM CaCl2) without (control 2) or with 2 U·mL-1 DNaseI was added to the wells (n = 3). After one hour of enzyme treatment at 37°C, the crystal violet assay (described in text) was performed. B) Confocal laser scanning microscopy images of component-specific stained C. ljungdahlii biofilms. C. ljungdahlii was grown in chamber slides with the addition of 200 mM NaCl to the medium and, after 2 days of incubation, the biofilms were stained as described in the text with SYPRO ruby red biofilm matrix stain (left) or calcofluor white (right). The scale bars are 50 μm long.