Alkaline phosphatase LapA regulates quorum sensing–mediated virulence and biofilm formation in
Pseudomonas aeruginosa
PAO1 under phosphate depletion stress
Authors: Tan Xiaojuan , Cheng Xi , Xiao Jingjing , Liu Qianqian , Du Dongsheng , Li Minghui , Sun Yang , Zhou Jinwei , Zhu Guoping 
Published: October 2023

Among various environmental stresses, the depletion of inorganic phosphate, which is an important signaling molecule that modulates virulence production, is notable. In our previous study, we demonstrated that
lap
A, encoding alkaline phosphatase, plays a key role in
Pseudomonas aeruginosa
biofilm formation in porcine skin explants, an
ex vivo
chronic wound model. However, the mechanism by which
lap
A participates in the virulence and biofilm formation of
P. aeruginosa
remains unclear. In the present study, the
lap
A deletion mutant was constructed in
P. aeruginosa
PAO1, and the phenotypic characterization was assessed. Our data demonstrated that phosphate depletion stress significantly enhanced elastase activity, hemolysis, rhamnolipid production, swarming and swimming motilities, and 3-oxo-C12-homoserine lactone (HSL) production in
P. aeruginosa
. Moreover, phosphate starvation evidently induced the virulence of
P. aeruginosa
in
Caenorhabditis elegans
(
C. elegans
) through fast-kill and slow-kill infections. Deletion of the
lap
A gene led to broad phenotypic changes, including reduced elastase activity, swimming motility, C4-HSL, and 3-oxo-C12-HSL production, as well as increased chitinase activity and rhamnolipid production under phosphate depletion stress, which was unrelated to C4-HSL. Deletion of the
lap
A gene also significantly inhibited PAO1 biofilm formation in porcine skin explants by reducing the expression levels of
las
and
rhl
quorum sensing systems and extracellular polymeric substance synthesis. Finally,
lap
A deletion also reduced the virulence of PAO1 in
C. elegans
in fast-kill and slow-kill infection assays. Thus, the findings suggest that targeting
lap
A can help control biofilm formation and reduce virulence in nonhealing-infected wounds, where
P. aeruginosa
is a persistent problem.

IMPORTANCE

Our previous study demonstrated that the expression of lapA was induced under phosphate depletion conditions, but its roles in virulence and biofilm formation by
Pseudomonas aeruginosa
remain largely unknown. This study presents a systematic investigation of the roles of lapA in virulence induction and biofilm formation by constructing a lapA-deficient strain with
P. aeruginosa
PAO1. The results showed that deletion of the lapA gene evidently reduced elastase activity, swimming motility, C4-HSL, and 3-oxo-C12-HSL production, and increased rhamnolipid production under phosphate depletion stress. Moreover, lapA gene deletion inhibited PAO1 biofilm formation in porcine skin explants by reducing the expression levels of las and rhl quorum sensing systems and extracellular polymeric substance synthesis. Finally, lapA gene deletion also reduced the virulence of PAO1 in
Caenorhabditis elegans
in fast-kill and slow-kill infection assays. This study provides insights into the roles of lapA in modulating
P. aeruginosa
virulence and biofilm formation under phosphate depletion stress.