DprA-Dependent Exit from the Competent State Regulates Multifaceted Streptococcus pneumoniae Virulence

Abstract

Streptococcus pneumoniae (pneumococcus) causes multiple infectious diseases. The pneumococcal competence system facilitates genetic transformation, spreads antibiotic resistance, and contributes to virulence. DNA-processing protein A (DprA) regulates the exit of pneumococcus from the competent state. Previously, we have shown that DprA is important in both bacteremia and pneumonia infections. Here, we examined the mechanisms of virulence attenuation in a ΔdprA mutant. Compared to the parental wild-type D39, the ΔdprA mutant enters the competent state when exposed to lower concentrations of the competence-stimulating peptide CSP1. The ΔdprA mutant overexpresses ComM, which delays cell separation after division. Additionally, the ΔdprA mutant overexpresses allolytic factors LytA, CbpD, and CibAB and is more susceptible to detergent-triggered lysis. Disabling of the competent-state-specific induction of ComM and allolytic factors compensated for the virulence loss in the ΔdprA mutant, suggesting that overexpression of these factors contributes to virulence attenuation. Finally, the ΔdprA mutant fails to downregulate the expression of multiple competence-regulated genes, leading to the excessive energy consumption. Collectively, these results indicate that an inability to properly exit the competent state disrupts multiple cellular processes that cause virulence attenuation in the ΔdprA mutant.

Streptococcus pneumoniae Elaborates Persistent and Prolonged Competent State during Pneumonia-Derived Sepsis

“…We used the serotype 2 strain D39 to construct the reporter strain AD2501 harboring a firefly luciferase transcriptionally fused to the ssbB gene (D39-ssbB-luc; Fig. 1A) (29). The ssbB gene encodes a single-stranded DNAbinding protein and is uniquely induced during competence induction and directly protects internalized single-stranded DNA (30).…”

“…For example, we found that D39 was not able to enter the competent state spontaneously when cultured in the nutrient-rich THY medium most widely used to cultivate pneumococci ( Fig. 1) (29). The competence-permissible CϩY medium, which contains defined amino acids, bovine serum albumin (BSA), and other nutrients, was formulated based on early findings that BSA or bodily fluids, including peritoneal washings and blood, provided better conditions for competence induction (6)(7)(8).…”

“…Under in vitro conditions, it has been reported that DprA is highly expressed and stable (34), rendering competent pneumococcal cells refractory to subsequent induction by CSP (35). Deletion of the dprA gene causes elevated and prolonged expression of "late" competence genes (16,17,29,34). Because of the prolonged and persistent competent state during pneumonia-derived sepsis, we examined whether the expression of DprA was compromised in vivo.…”

“…We predicted that such high levels of CSP should be able to stimulate a noncompetent recipient reporter strain. We generated a ssbB-luc reporter strain in the ΔdprA genetic background (ΔdprA-ssbB-luc, AD2506), which has heightened sensitivity and response to competence induction due to its inability to shut off the competent state (29). The pneumococcal cells that secreted these CSPs were removed from the blood samples by filtration to eliminate possible competence transfer by cell-cell direct contact mechanism.…”

A ComRS competence pathway in the oral pathogen Streptococcus sobrinus

“…sobrinus is one of the few species of oral streptococci that lack a natural competence pathway. Competence is central to pathogenicity in streptococci, and competence pathways are connected to metabolism [4], virulence [5,6], quorum sensing [7], and antibiotic tolerance [8]. The natural competence of streptococci has been known for over 75 years [9].…”