Introduction

Streptococcus pneumoniae (the pneumococcus) is a major human pathogen that

colonizes the upper respiratory tract and causes both life-threatening diseases such as
pneumonia, sepsis, and meningitis and milder but common diseases, like sinusitis and otitis
media . Colonization of the nasopharynx by S. pneumoniae begins early on in life and peaks
(up to 55%) at the age of 3 years .Colonization not only provides the basis for pneumococcal
horizontal spread but is also necessary for invasive disease. The pneumococcus produces a
plethora of virulence factors that allow bacteria to spread from the upper to the lower
respiratory tract, leading to pneumonia and invasive disease. The latest report from the
Centers for Disease Control and Prevention in the United States estimated that in 2006
pneumococci caused 41,400 invasive infections and 5,000 deaths, most of which were due
to bacteremic pneumonia. However, each year at least 1 million children younger than 5
years of age die of pneumonia and invasive diseases in developing countries . Mortality
rates for pneumonia and meningitis are especially high for young children, the elderly, and
immunocompromised individuals , including patients with human immunodeficiency virus .
Pneumococcal diseases are treated with -lactams (penicillin G, amoxicillin, cephalosporins),
respiratory fluoroquinolones, macrolides, and vancomycin. However, efforts to treat
pneumococcal diseases have been complicated by increasing resistance to antibiotics. A
recent work reported high rates of resistance for some antimicrobials such as penicillin
(34.2%), trimethoprim-sulfamethoxazole (31.9%), and erythromycin (29.5%). For the
prevention of pneumococcal disease, there is a licensed 23-valent polysaccharide vaccine
which is effective against 23 out of 90 pneumococcal serotypes but does not confer
protection in children younger than 2 years of age or in the elderly. A 7-valent conjugate
vaccine has also been available since 2000 and is effective in children, although its cost is
high and protection is induced only against serotypes included in the vaccine formulation .
Both 9- and 11-valent conjugate vaccines will be licensed in the near future. Protein-based
or protein subunit vaccines common to different capsular serotypes represent attractive
alternatives to improve protection and to address the current limitations of polysaccharide
vaccines.

Methodology
Bacterial isolates and growth conditions. S. pneumoniae serotype 6A strains that were
isolated from patients with either invasive pneumococcal disease (n = 5) or asymptomatic
NP carriage (n 5) were used for C3 binding studies.
Sera and complement reagents. Immune serum was obtained from two healthy volenteers
4 weeks after vaccination woth the 23 valent pneumococcal polysaccharides vaccine.
Antibodies. MAbs directed against serotype 6A pneumococcal polysaccgarides were
obtained as a courtesy.
Multilocus sequences typing.The genotypes of the pneumococcal strains used in this
study were analyzed by multilocus sequence typing (MLST).
Colony morphology. Bacteria grown overnight in BHI broth supplemented with 10
g/ml

hemin

and

g/ml NAD at 31C were reinoculated into fresh media the next day and grown to the mid-log
phase. Bacteria were inoculated onto tryptic soy agar plates (Remel), each containing 100
l (3,000 U) of catalase.
Evaluation of complement and antibody deposition and surface of S. pneumoniae.
EOM model. An experimental chinchilla model of acute otitis media was used
Capsule qualification. Capsule qualification both that on intact becteria and that released
into culture supernatants during growth in liquid media.

Results and discussion

Innate immunity and interaction between the pneumococcus and other microbes
For ease of study, most work on the interaction of the pneumococcus with the innate
immune system has employed pure cultures. However, the mucosa of the upper respiratory
tract is colonized by a diverse array of microbial species. Indeed, analysis of DNA from
human airway surface fluid suggests the presence of more than 500 bacterial species
(Paster et al., 2001). Concurrent stimulation of the innate immune system by multiple
species appears to have effects distinct from those of single-species interactions (Neish et
al., 2000; Tong et al., 2003), and this has recently been shown to have relevance to the

pneumococcus (Ratner et al., 2005). Co-stimulation of human respiratory epithelia cells in

vitro by S. pneumoniae and Haemophilus influenzae, also an inhabitant of the upper
respiratory tract, results in synergistic production of IL-8 (Ratner et al., 2005). This has been
extended to a mouse-colonization model, with a synergistic effect on MIP-2 production and
inflammatory influx into the upper airways. This synergy is independent of TLR2 and TLR4,
but involves NF-k B translocation to the nucleus and phosphorylation of p38 MAPK. With
regard to the microbial products involved, pneumolysin can substitute for the pneumococcus,
but the PdB pneumolysin toxoid, lacking cytolytic activity, is inactive. It is therefore
speculated that the pore-forming activity of pneumolysin leads to enhanced delivery of
microbial products, such as the soluble inflammatory protein SCF from H. influenzae, into
the host cell, where recognition by Nod1 and Nod2 would result in increased stimulation
(Ratner et al., 2005). This proinflammatory activity of pneumolysin is therefore distinct from
its effects on macrophages that are mediated through TLR4 and are independent of poreforming activity.
Another important microbial interaction is that of the pneumococcus and the influenza A
virus. Subsequent to influenza A outbreaks, secondary pneumococcal infectionis an
important cause of morbidity and mortality. This heightened susceptibility to pneumococcal
disease can be reproduced in animal models, allowing investigation of the mechanisms
involved. While viral neuraminidase contributes to this phenomenon by exposing
pneumococcal receptors (McCullers & Bartmess, 2003; Tong et al., 2001), alterations in the
immune response also seem to contribute. Prior influenza A infection in mice primes for an
exaggerated inflammatory response to subsequent pneumococcal infection (van der Sluijs et
al., 2004). Increased levels of IL-10 in this response likely contribute to increased
susceptibility, as neutralization of this cytokine improves disease outcome (van der Sluijs et
al., 2004). In vitro exposure to both influenza A and the pneumococcus results in a
synergistic inflammatory response from human middle-ear epithelial cells (Tong et al., 2003).
Microarray gene expression analysis of these cells following influenza A infection provides
insight into the possible mechanisms behind this synergy (Tong et al., 2004). For example, it
is found that tlr2 expression is upregulated by influenza A infection. This may make the cell
more responsive to stimulation by pneumococcal peptidoglycan and LTA (Tong et al., 2004).
Global analysis of host responses
The advent of microarray technology allows greater insight into the host-cell response to the
pneumococcus. The response of the human monocytic cell line THP-1 has been assessed
by microarray analysis following exposure to S. pneumoniae and an isogenic mutant lacking
pneumolysin (Rogers et al., 2003). After a three-hour exposure to the pneumococcus,

expression differences were revealed in 182 host genes from the 4133 examined, illustrating
the potential for large-scale expression changes induced by the pneumococcus (Rogers et
al., 2003). Of these 182 genes, 142 were responsive to pneumolysin, showing the dominant
nature of this virulence factor in host responses. While this study will not be comprehensive
in fully documenting the host response, it illustrates the complexity of the interaction between
host cells and the pneumococcus. An important future challenge will be to understand the
significance of these expression changes in the disease process.
In addition to the host response, global analysis of bacterial gene expression in vivo has also
been studied using models of meningitis and bacteraemia (Orihuela et al., 2004). Together,
these studies of host and bacterial response in vivo provide a platform to understand in
molecular detail pneumococcalhost interactions from both perspectives.

Conculusion
This studies using MARCO deficient mice show and important function for this AM class A
scavenger receptor in host defense against pnemococcal pneumonia and against pulmonary
inflammation from otherwise inert environmental dust.

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