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Bacterial biofilms formation is one of the major reasons for treatment failure in chronic wound infections. Therefore, diagnostic biomarkers remain the best option for prevention and treatment of chronic wound infections by biofilms. Herein,  Pseudomonas aeruginosa  PAO1 was used to mimic biofilm development in porcine skin explants wells as ex vivo wound model. The microscopic imaging showed that PAO1 in porcine skin explants wells formed micro-colonies at 24 h, developed mushroom-like structure at 48 h, and at 72 h mushroom-like structure disappeared, remaining a thin bacterial lawn. RNA-seq data analysis revealed that the expression levels of genes involved in the type II  hxc  secretion system were significantly higher in biofilms than in planktonic cells, especially the expression of  lap A encoding alkaline phosphatase. However, the expression levels of genes associated with denitrification pathway were markedly decreased in biofilms, especially the transcription of  nir S encoding nitrite reductase to produce nitric oxide (NO). Therefore, their expressions and products were further detected using RT-qPCR and biochemical assays, respectively. The results found that the expression of  lap A and alkaline phosphatase activity were induced, but the expression of  nir S and intracellular NO were reduced at the whole biofilms cycle. The study indicates that LapA and NO would play an important role for  P. aeruginosa  biofilm formation in chronic wound infections. LapA would serve as potential target to monitor chronic wound infections by  P. aeruginosa  biofilms. Inducing NO would be used to treat chronic wound infections due to  P. aeruginosa  biofilms.

Transcriptional analysis and target genes discovery of Pseudomonas aeruginosa biofilm developed ex vivo chronic wound model