Designed Reduction of Streptococcus pneumoniae Pathogenicity via Synthetic Changes in Virulence Factor Codon-pair Bias

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.

A Serotype 3 Pneumococcal Capsular Polysaccharide-Specific Monoclonal Antibody Requires Fcγ Receptor III and Macrophages To Mediate Protection against Pneumococcal Pneumonia in Mice

“…Nonetheless, and corresponding to the lack of early bacterial clearance in the lungs of 1E2-treated mice, 1E2-opsonized ST3 induced significantly less intracellular bacterial killing in vitro after 1 h of incubation. Although ample data establish that mouse survival in pneumococcal infection requires bacterial clearance and control of bloodstream invasion (8,13,27,44,54), this study did not directly demonstrate how 1E2 mediates bacterial clearance. Given that pneumococcal pathogenesis stems from bacterial ligands binding to host receptors (3, 34) 1E2-induced macrophage internalization of ST3 could temporarily limit ST3 access to host receptors.…”

“…ELISA Duoset kits for macrophage inflammatory protein-2 (MIP-2), interleukin-6 (IL-6), and tumor necrosis factor-␣ (TNF-␣) (R&D Systems) were used for cytokine determinations according to the manufacturer's protocols. These mediators were chosen because they are increased by Fc␥RIII-mediated phagocytosis (21), in models of pneumococcal pneumonia and sepsis (13,14,24,27,54) and pneumococcal phagocytosis in S. pneumoniae-infected inhibitory Fc␥R (Fc␥RIIB)-deficient mice (12).…”

Attenuation of human respiratory syncytial virus by genome-scale codon-pair deoptimization

“…Because amino acid coding is unaffected, CPD strains provide the same repertoire of epitopes for inducing cellular and humoral immunity as the WT pathogen. Recently, the CPD approach has been used successfully to attenuate poliovirus, influenza A virus, Streptococcus pneumonia, and HIV type 1 (5,(10)(11)(12)(13).…”

HP-PRRSV is attenuated by de-optimization of codon pair bias in its RNA-dependent RNA polymerase nsp9 gene

“…Although it is not clear how codon pair preference is formed (Buchan et al, 2006), the codon pair usage in an open reading frame can influence the gene expression (Irwin et al, 1995). It has been shown that synthesizing novel poliovirus and influenza virus to contain underrepresented codon pairs while exactly preserve the codon usage and amino acid sequence, can dramatically attenuate the virus (Coleman et al, 2008(Coleman et al, , 2011Mueller et al, 2010;Yang et al, 2013). Recently, a report showed that deoptimization of the major envelope GP5 gene attenuated PRRSV (Ni et al, 2014).…”