Figure 3
Genetic and phenotypic analysis of biofilm and quorum sensing coding genes. (A) Heatmap outlining the differential expression of genes associated with biofilm formation and quorum sensing in A. baumannii AB5075 Δhns vs. parental strain. The asterisks represent the DEGs (adjusted P-value < 0.05 with log2fold change > 1). Heat map was performed using GraphPad Prism version number 9 (GraphPad software, San Diego, CA, USA, https://www.graphpad.com/) (B) qRT-PCR of AB5075, AB5075 Δhns and AB5075 Δhns pMBLe-hns strains genes associated with biofilm and quorum sensing, the expression levels and the fold changes were determinate using the double ΔCt analysis. At least three independent samples were used, and three technical replicates were performed from each sample. Statistical significance (P-value < 0.05) was determined by ANOVA followed by Tukey’s multiple comparison test; one asterisks: P-value < 0.05; two asterisks: P-value < 0.01 and three asterisks: P-value < 0.001. (C) Biofilm assays performed in A. baumannii AB5075, AB5075 Δhns and AB5075 Δhns pMBLe-hns represented by OD580/OD600. Statistical analysis (t test) was performed and a P-value < 0.05 was considered significant. (D) Agar plate assay for the detection of AHL using A. tumefaciens. The presence of AHL was determined by the development of the blue color. Quantification of 5,5′-dibromo-4,4′-dichloro-indigo were estimated as the percentage relative to C10-AHL standard, measured with ImageJ (NIH). The mean ± SD is informed. Statistical significance (P-value < 0.05) was determined by ANOVA followed by Tukey’s multiple-comparison test. Experiments were performed in triplicate, with at least three technical replicates per biological replicate. This figure was performed using GraphPad Prism version number 9 (GraphPad software, San Diego, CA, USA, https://www.graphpad.com/).
