TacL-mediated lipoteichoic acid biosynthesis regulates pneumolysin release via modulating bacterial surface hydrophobicity and autolysis in Streptococcus pneumoniae D39 grown in C+Y medium

Streptococcus pneumoniae modulates its virulence by regulating the release of pneumolysin (Ply), a key pore-forming toxin whose secretion is tightly governed by the bacterial cell wall. Lipoteichoic acids (LTAs), a major structural component of the Gram-positive bacterial cell wall, have an uncharacterized role in the regulation of Ply release. Here, we demonstrate that TacL-mediated (LTAs) biosynthesis controls Ply release in S. pneumoniae by altering cell wall hydrophobicity during the early logarithmic phase and bacterial autolysis during the middle-late logarithmic phase. Deletion of the tacL gene disrupted LTAs biosynthesis in S. pneumoniae strain D39 cultured in C+Y medium, resulting in a significant decrease in Ply levels in the culture supernatant and a concurrent increase in bacterial cell‑associated Ply during the early logarithmic phase. Notably, neither ply gene transcription nor the total Ply abundance in bacterial suspensions was significantly altered by tacL deletion, indicating that TacL regulates Ply release rather than its synthesis. The tacL mutant exhibited significantly increased surface hydrophobicity during the early logarithmic phase; importantly, mitigation of this enhanced hydrophobicity by treatment with Tween-80 restored Ply secretion into the supernatant and reduced intracellular Ply retention. Collectively, these findings establish a novel link between LTAs biosynthesis and Ply release in S. pneumoniae, mediated by the regulation of cell wall hydrophobicity and bacterial autolysis. This study advances our understanding of the molecular mechanisms underlying Ply release in S. pneumoniae D39 and provides new insights into the role of LTAs in modulating the virulence of this opportunistic pathogen.