[Show abstract][Hide abstract] ABSTRACT: Bacteremic pneumonia with some pneumococcal capsular serotypes, including serotype 3 (ST3), has been associated with a higher risk of death, whereas others, such as ST8, are associated with a lower risk. To provide a molecular basis for understanding such differences, we used oligo cDNA microarrays to analyze and compare the gene expression profiles of the lungs of Balb/c mice infected intranasally with either ST3, strain A66.1, or ST8, strain ATCC 6308 (6308). Compared to uninfected controls, infection with either A66.1 or 6308 led to inoculum-dependent expression of IFN-γ inducible CXC chemokines among other pro-inflammatory genes. To investigate the role that IFN-γ inducible chemokines CXCL9, CXCL10 and CXCL11 play in A66.1- and 6308-induced pneumonia, we examined the effect of the absence of their common receptor, CXCR3, on intranasal infection in CXCR3(-/-) (Balb/c) mice. Compared to wild type (WT) mice, virulence of A66.1 but not 6308 was attenuated in CXCR3(-/-) mice. A66.1-infected CXCR3(-/-) mice had fewer lung neutrophils and more alveolar macrophages 48 h after infection and fewer blood CFU 72 h after infection. Histopathological examination of lung sections revealed less inflammation among A66.1-infected CXCR3(-/-) than WT mice. The reduced virulence of A66.1 in CXCR3(-/-) mice suggests that inhibition of the functional activity of IFN-γ inducible chemokines modulates the host response to A66.1, in turn suggesting a novel approach to improve vaccine-mediated protection against ST3 pneumonia.
[Show abstract][Hide abstract] ABSTRACT: In this study, we used a previously described method of controlling gene expression with computer-based gene design and de novo DNA synthesis to attenuate the virulence of Streptococcus pneumoniae. We produced 2 S. pneumoniae serotype 3 (SP3) strains in which the pneumolysin gene (ply) was recoded with underrepresented codon pairs while retaining its amino acid sequence and determined their ply expression and pneumolysin production in vitro and their virulence in a mouse pulmonary infection model. Expression of ply and production of pneumolysin of the recoded SP3 strains were decreased, and the recoded SP3 strains were less virulent in mice than the wild-type SP3 strain or a Δply SP3 strain. Further studies showed that the least virulent recoded strain induced a markedly reduced inflammatory response in the lungs compared with the wild-type or Δply strain. These findings suggest that reducing pneumococcal virulence gene expression by altering codon-pair bias could hold promise for rational design of live-attenuated pneumococcal vaccines.
The Journal of Infectious Diseases 02/2011; 203(9):1264-73. · 5.85 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The use of pneumococcal capsular polysaccharide (PPS)-based vaccines has resulted in a substantial reduction in invasive pneumococcal disease. However, much remains to be learned about vaccine-mediated immunity, as seven-valent PPS-protein conjugate vaccine use in children has been associated with nonvaccine serotype replacement and 23-valent vaccine use in adults has not prevented pneumococcal pneumonia. In this report, we demonstrate that certain PPS-specific monoclonal antibodies (MAbs) enhance the transformation frequency of two different Streptococcus pneumoniae serotypes. This phenomenon was mediated by PPS-specific MAbs that agglutinate but do not promote opsonic effector cell killing of the homologous serotype in vitro. Compared to the autoinducer, competence-stimulating peptide (CSP) alone, transcriptional profiling of pneumococcal gene expression after incubation with CSP and one such MAb to the PPS of serotype 3 revealed changes in the expression of competence (com)-related and bacteriocin-like peptide (blp) genes involved in pneumococcal quorum sensing. This MAb was also found to induce a nearly 2-fold increase in CSP2-mediated bacterial killing or fratricide. These observations reveal a novel, direct effect of PPS-binding MAbs on pneumococcal biology that has important implications for antibody immunity to pneumococcus in the pneumococcal vaccine era. Taken together, our data suggest heretofore unsuspected mechanisms by which PPS-specific antibodies could affect genetic exchange and bacterial viability in the absence of host cells. IMPORTANCE: Current thought holds that pneumococcal capsular polysaccharide (PPS)-binding antibodies protect against pneumococcus by inducing effector cell opsonic killing of the homologous serotype. While such antibodies are an important part of how pneumococcal vaccines protect against pneumococcal disease, PPS-specific antibodies that do not exhibit this activity but are highly protective against pneumococcus in mice have been identified. This article examines the effect of nonopsonic PPS-specific monoclonal antibodies (MAbs) on the biology of Streptococcus pneumoniae. The results showed that in the presence of a competence-stimulating peptide (CSP), such MAbs increase the frequency of pneumococcal transformation. Further studies with one such MAb showed that it altered the expression of genes involved in quorum sensing and increased competence-induced killing or fratricide. These findings reveal a novel, previously unsuspected mechanism by which certain PPS-specific antibodies exert a direct effect on pneumococcal biology that has broad implications for bacterial clearance, genetic exchange, and antibody immunity to pneumococcus.
[Show abstract][Hide abstract] ABSTRACT: Streptococcus pneumoniae is the most common cause of community-acquired pneumonia in the United States and globally. Despite the availability of pneumococcal capsular polysaccharide (PPS) and protein conjugate-based vaccines, the prevalence of antibiotic-resistant pneumococcal strains, serotype (ST) replacement in nonconjugate vaccine strains, and uncertainty as to whether the PPS vaccine that is used in adults protects against pneumonia emphasize the need for continued efforts to understand the nature of protective PPS antibody responses. In this study, we generated mouse monoclonal antibodies (MAbs) to a conjugate consisting of the PPS of serotype 8 (PPS8) S. pneumoniae and tetanus toxoid. Thirteen MAbs, including four IgMs that bound to PPS8 and phosphorylcholine (PC) and five IgMs and four IgG1s that bound to PPS8 but not PC, were produced, and their nucleotide sequences, epitope and fine specificity, and efficacy against lethal challenge with ST8 S. pneumoniae were determined. MAbs that bound to PPS8 exhibited gene use that was distinct from that exhibited by MAbs that bound to PC. Only PPS8-binding MAbs that did not bind PC were protective in mice. All 13 MAbs used germ line variable-region heavy (V(H)) and light (V(L)) chain genes, with no evidence of somatic hypermutation. Our data reveal a relationship between PPS specificity and V(H) gene use and MAb efficacy in mice. These findings provide insight into the relationship between antibody molecular structure and function and hold promise for the development of novel surrogates for pneumococcal vaccine efficacy.