We examined the role of the neuraminidases NanA and NanB in colonization and infection in the upper and lower respiratory tract by Streptococcus pneumoniae, as well as the role of these neuraminidases in the onset and development of septicemia following both intranasal and intravenous infection. We demonstrated for the first time using outbred MF1 mouse models of infection that both NanA and NanB were essential for the successful colonization and infection of the upper and lower respiratory tract, respectively, as well as pneumococcal survival in nonmucosal sites, such as the blood. Our studies have shown that in vivo a neuraminidase A mutant is cleared from the nasopharynx, trachea, and lungs within 12 h postinfection, while a neuraminidase B mutant persists but does not increase in either the nasopharynx, trachea, or lungs. We also demonstrated both neuraminidase mutants were unable to cause sepsis following intranasal infections. When administered intravenously, however, both mutants survived initially but were unable to persist in the blood beyond 48 h postinfection and were progressively cleared. The work presented here demonstrates the importance of pneumococcal neuraminidase A and for the first time neuraminidase B in the development of upper and lower respiratory tract infection and sepsis.

Streptococcus pneumoniae colonizes the mucin-rich environment of the nasopharynx. As colonization may be the first stage of infection, investigation was carried out as to whether the pneumococcus could utilize mucin as a source of nutrient and whether its virulence gene expression is influenced by this glycoprotein. It was found that when grown in Sicard's defined medium supplemented with 1% w/v mucin, the organism grew at a rate similar to that in rich medium. The presence of sialate O-acetylesterase activity, an enzyme implicated in glycoprotein degradation, in pneumococcal cell extracts was also reported. The authors hypothesized that neuraminidase A, which is linked to pneumococcal virulence, plays an important role in mucin utilization. Growth in mucin resulted in an increase in nanA transcription and a DeltananA-deficient strain of pneumococcus could not grow when mucin was used as the sole carbon source.

In Streptococcus pneumoniae, the two-component signaling system MicAB was previously shown to contribute to repression of competence when oxygen is limited. In virulent strains expressing the serotype 2 and 6 capsule, mutation of the MicB kinase reduced the lag period of growth when cultures were switched from an aerobic to anaerobic atmosphere. After intranasal challenge of mice, the micB::km mutation decreased virulence, as shown by the absence of symptoms and by a lower level of recovery of CFU from lungs and blood. It is proposed that MicAB is involved in the adaptive response of the bacteria to changes in oxygen level during the course of infection.Streptococcus pneumoniae is an aerotolerant anaerobic human pathogen. During exponential growth in rich medium, central metabolism is essentially fermentative (4), despite NADH oxidase (1) and pyruvate oxidase (11), which reduce molecular substrate to H 2 O and H 2 O 2 , respectively. Implication of these oxidases in virulence (1, 11) indicates that oxidative metabolism is required for full virulence of S. pneumoniae. In vitro, oxidative metabolism also is involved in the control of genetic exchange via the regulation of competence for genetic transformation (1): competence is repressed under oxygen limitation (5).Integration of cellular redox status with competence occurs by phosphotransfer through the two-component systems (TCS) CiaRH and ComCDE (5-7). A third TCS, MicAB, contributes to repression of competence under oxygen limitation (7). Mutational analysis led to the proposal that the response regulator MicA is essential for bacterial growth, whereas the active form of the kinase, MicB, is not essential in vitro (7,9,12). MicB is the single protein in S. pneumoniae carrying the PAS signature (9).PAS domains are conserved motifs present in proteins that sense light or the redox and energetic status of the cell (2). Mutation of the PAS domain in MicB of S. pneumoniae abolishes the kinase activity of the recombinant protein and allows expression of competence under microaerobic conditions, suggesting that MicB phosphorylation both is dependent on PAS and constitutes one response to oxygen concentration (7).The requirement for this TCS in growth and in regulation of competence in vitro prompted us to investigate its role in the fate of pneumococci in vivo. The effect on in vivo growth and virulence of a mutation that annuls MicB kinase activity has been evaluated. Aerobic growth and anaerobic growth in vitro were studied for both serotype 2 and 6 pneumococcal micB::km mutants and their isogenic parents, and the virulence of these strains was determined with an in vivo model of pneumonia and bacteremia.The S. pneumoniae strains were a serotype 2 strain, D39 (NCTC 7466; National Collection of Type Cultures, London, United Kingdom), and a serotype 6B strain, S6 (10). Isogenic derivatives of these strains with the micB::km mutation were obtained by transformation of the wild-type strains with pPT12::Km, using 100 g of competence-stimulating peptide per ml (7). Trans...