Streptococcus pneumoniae Serine Protease HtrA, but Not SFP or PrtA, Is a Major Virulence Factor in Pneumonia

Abstract

Streptococcus (S.) pneumoniae is a common causative pathogen in pneumonia. Serine protease orthologs expressed by a variety of bacteria have been found of importance for virulence. Previous studies have identified two serine proteases in S. pneumoniae, HtrA (high-temperature requirement A) and PrtA (cell wall-associated serine protease A), that contributed to virulence in models of pneumonia and intraperitoneal infection respectively. We here sought to identify additional S. pneumoniae serine proteases and determine their role in virulence. The S. pneumoniae D39 genome contains five putative serine proteases, of which HtrA, Subtilase Family Protein (SFP) and PrtA were selected for insertional mutagenesis because they are predicted to be secreted and surface exposed. Mutant D39 strains lacking serine proteases were constructed by in-frame insertion deletion mutagenesis. Pneumonia was induced by intranasal infection of mice with wild-type or mutant D39. After high dose infection, only D39ΔhtrA showed reduced virulence, as reflected by strongly reduced bacterial loads, diminished dissemination and decreased lung inflammation. D39ΔprtA induced significantly less lung inflammation together with smaller infiltrated lung surface, but without influencing bacterial loads. After low dose infection, D39ΔhtrA again showed strongly reduced bacterial loads; notably, pneumococcal burdens were also modestly lower in lungs after infection with D39Δsfp. These data confirm the important role for HtrA in S. pneumoniae virulence. PrtA contributes to lung damage in high dose pneumonia; it does not however contribute to bacterial outgrowth in pneumococcal pneumonia. SFP may facilitate S. pneumoniae growth after low dose infection.

HtrA‐mediated selective degradation of DNA uptake apparatus accelerates termination of pneumococcal transformation

“…Previous studies show that pneumococcal HtrA is associated with inhibition of competence (Ibrahim et al , ; Sebert et al , ) and bacteriocin production (Dawid et al , ; Kochan and Dawid, ), tolerance to heat shock and oxidative stress (Ibrahim et al , ), and promotion of nasopharyngeal colonization (Sebert et al , ) and virulence (Ibrahim et al , ; de Stoppelaar et al , ). However, the protein targets of HtrA associated with these phenotypes have not been identified.…”

Stress Physiology of Lactic Acid Bacteria

“…Another highly conserved bacterial protein associated with the pathogenic potential of streptococcal species is HtrA. Deletion of htrA in S. pneumoniae significantly reduced nasopharyngeal colonization (486) and virulence in mouse models of pneumonia and bacteremia (487,488). The 50% lethal dose (LD 50 ) for an S. pyogenes htrA mutant in mice was 35-fold higher than that for the parent strain (489).…”

Non-Adhesive Surface Proteins of Streptococcus pneumoniae

“…Virulence of an htrA mutant was drastically reduced in animal models of pneumonia and bacteremia [51,55], which is not surprising as loss of HtrA will result in a fitness defect. Mutants of PrtA were attenuated in intraperitoneal infection [47], while in a pneumonia model a high-dose infection with D39ΔprtA induced significantly less lung inflammation without influencing bacterial loads [55], suggesting that PrtA plays a role in immunomodulation. A similar phenotype was observed with an SFP mutant, where colony-forming unit (CFU) count and inflammation were significantly lower in the lungs after a low-dose infection [55].…”

“…During cell division, HtrA was found to be predominantly located at the equators and septa of dividing pneumococcal cells, suggesting that it plays a general role in cell division by quality control of proteins exported by the Sec translocase in S. pneumoniae [54]. Virulence of an htrA mutant was drastically reduced in animal models of pneumonia and bacteremia [51,55], which is not surprising as loss of HtrA will result in a fitness defect. Mutants of PrtA were attenuated in intraperitoneal infection [47], while in a pneumonia model a high-dose infection with D39ΔprtA induced significantly less lung inflammation without influencing bacterial loads [55], suggesting that PrtA plays a role in immunomodulation.…”