Pneumococcal Virulence Gene Expression and Host Cytokine Profiles during Pathogenesis of Invasive Disease

Abstract

Pneumococcal disease continues to account for significant morbidity and mortality worldwide. For the development of novel prophylactic and therapeutic strategies against the disease spectrum, a complete understanding of pneumococcal behavior in vivo is necessary. We evaluated the expression patterns of the proven and putative virulence factor genes adcR, cbpA, cbpD, cbpG, cpsA, nanA, pcpA, piaA, ply, psaA, pspA, and spxB after intranasal infection of CD1 mice with serotype 2, 4, and 6A pneumococci by real-time reverse transcription-PCR. Simultaneous gene expression patterns of selected host immunomodulatory molecules, CCL2, CCL5, CD54, CXCL2, interleukin-6, and tomor necrosis factor alpha, were also investigated. We show that pneumococcal virulence genes are differentially expressed in vivo, with some genes demonstrating niche-and serotype-specific differential expression. The in vivo expression patterns could not be attributed to in vitro differences in expression of the genes in transparent and opaque variants of the three strains. The host molecules were significantly upregulated, especially in the lungs, blood, and brains of mice. The pneumococcalgene expression patterns support their ascribed roles in pathogenesis, providing insight into which protein combinations might be more appropriate as vaccine antigens against invasive disease. This is the first simultaneous comparison of bacterial-and host gene expression in the same animal during pathogenesis. The strategy provides a platform for prospective evaluation of interaction kinetics between invading pneumococci and human patients in culture-positive cases and should be feasible in other infection models.

Important Role for CC Chemokine Ligand 2-Dependent Lung Mononuclear Phagocyte Recruitment to Inhibit Sepsis in Mice Infected with Streptococcus pneumoniae

“…The pathogenicity profile of S. pneumoniae not only differs between, but also among serotypes (10). In addition, both capsular and noncapsular virulence factors contribute to invasiveness of a given strain.…”

“…These data uncovered the CCL2-CCR2 axis as an integral component of lung innate immunity to inhaled S. pneumoniae (3,(7)(8)(9). However, since S. pneumoniae differs in terms of pathogenicity profiles not only between, but also among serotypes (10), the relevance of host-derived mechanisms to contribute to lung-protective immunity against S. pneumoniae, such as chemokine-induced lung phagocyte trafficking, can only be fully evaluated in comparative analyses using both highly virulent and less virulent pneumococcal strains. Such comparative analyses are of major clinical importance, because a patient's outcome in infectious disease not only depends on the pathogen's virulence, but also on the patient's resistance to a given pathogen.…”

Identification of a novel pneumococcal vaccine antigen preferentially expressed during meningitis in mice

“…RNA for microarray experiments was isolated and purified from either blood-borne bacteria or a mixture of bacteria that had either crossed over to the brain or were still in transition (attached to the microvascular endothelium) with acid phenol:chloroform:isoamyl alcohol (125:24:1; pH 4.5; Ambion). Extracted RNA samples were checked for purity and integrity as described previously (26,27). Bacterial RNA samples from a minimum of 5 mice that satisfied these criteria from a specific niche were pooled.…”

“…To test this, we conducted microarray transcriptomic analysis of bacteria harvested from the blood and brain of mice at various stages of the pathogenic process, examined the role of upregulated genes in pathogenesis, and tested the protective immunogenicity of one of the gene products. (26,27). In one of those studies, we observed that 2 clinical strains, WCH16 (serotype 6A) and WCH43 (serotype 4), had a high propensity to progress to the brain of mice and proliferate to cause meningitis (27).…”

“…(26,27). In one of those studies, we observed that 2 clinical strains, WCH16 (serotype 6A) and WCH43 (serotype 4), had a high propensity to progress to the brain of mice and proliferate to cause meningitis (27). Therefore, the pathogenesis of the 2 strains was followed at 48, 72, and 96 hours after i.n.…”