Nasal MRSA colonization in healthy children in Nashville has increased significantly in the past 3 years. As colonization typically precedes infection, this increase may be a major factor in the emergence of community-associated MRSA as a pathogen of healthy children.

Superoxide dismutase (SOD) is a ubiquitous metalloenzyme in aerobic organisms that catalyzes the conversion of superoxide anion to hydrogen peroxide. Mycobacterium tuberculosis is unusual in that it secretes large quantities of iron-cofactored SOD. To determine the role of SOD in pathogenesis, we constructed mutants of M. tuberculosis H37Rv with reduced SOD production. Compared with controls, SOD-diminished isolates were more susceptible to killing by hydrogen peroxide. The isolates were markedly attenuated, exhibiting nearly 100,000-fold fewer bacilli than virulent control strains in the lungs and spleens of C57BL/6 mice 4 wk after intravenous inoculation. In the lung, SOD-attenuated M. tuberculosis induced robust interstitial mononuclear cell infiltration within 24 h and many cells were apoptotic by TUNEL staining, whereas virulent H37Rv exhibited minimal early inflammatory response and only rare interstitial mononuclear cell apoptosis. During prolonged infections, C57BL/6 mice tolerated SOD-attenuated M. tuberculosis better than BCG, exhibiting 68% greater weight gain, quicker eradication of bacilli from the spleen, and less alveolar lung infiltration. These results establish the importance of SOD in the pathogenesis of tuberculosis. Its effect appears to be mediated in part by inhibiting innate host immune responses, including early mononuclear cell infiltration of infected tissues and apoptosis.

We examined the role of redox signaling generated by NADPH oxidase in activation of NF-κB and host defense against Pseudomonas aeruginosa pneumonia. Using mice with an NF-κB-driven luciferase reporter construct (HIV-LTR/luciferase (HLL)), we found that intratracheal administration of P. aeruginosa resulted in a dose-dependent neutrophilic influx and activation of NF-κB. To determine the effects of reactive oxygen species generated by the NADPH oxidase system on activation of NF-κB, we crossbred mice deficient in p47phox with NF-κB reporter mice (p47phox−/−HLL). These p47phox−/−HLL mice were unable to activate NF-κB to the same degree as HLL mice with intact NADPH oxidase following P. aeruginosa infection. In addition, lung TNF-α levels were significantly lower in p47phox−/−HLL mice compared with HLL mice. Bacterial clearance was impaired in p47phox−/−HLL mice. In vitro studies using bone marrow-derived macrophages showed that Toll-like receptor 4 was necessary for NF-κB activation following treatment with P. aeruginosa. Additional studies with macrophages from p47phox−/− mice confirmed that redox signaling was necessary for maximal Toll-like receptor 4-dependent NF-κB activation in this model. These data indicate that the NADPH oxidase-dependent respiratory burst stimulated by Pseudomonas infection contributes to host defense by modulating redox-dependent signaling through the NF-κB pathway.

Staphylococcus aureus produces four types of ,-lactamase (A, B, C, and D). To (13,14,16,20,21,24,25,27

We previously have shown that extracts of S. aureus isolates which produce the recognized serotypes of staphylococcal I-lactamase (A, B, C, D) differ in the rates at which they hydrolyze selected cephalosporins, exhibiting substrate profiles which are distinctive for each serotype. In an effort to simplify the methods employed in identifying the different staphylococcal ,-lactamases, we evaluated whether distinctive substrate profiles could be obtained by using whole-cell suspensions of 115 ,I-lactamase-producing isolates of S. aureus. Compared with extracts from the same strains, the whole-cell bacterial suspensions not only were simpler to prepare but enabled P-lactamase typing of a higher proportion of the evaluated strains (86 versus 97%, respectively). Furthermore, the use of whole-cell bacterial suspensions enabled the simultaneous quantitation of the P-lactamase activity exhibited by each strain. Additionally, by comparing the quantitative activity of I8-lactamase-induced and -uninduced preparations of the same strain, induction ratios (i.e., induced/uninduced activity) could be derived, yielding information regarding the regulation of ,-lactamase production by each strain. We believe that the utilization of whole-cell methods, such as those employed in this study, will facilitate the investigation of qualitative and quantitative differences in ,B-lactamase production among clinical and reference isolates of S. aureus.There are four known serotypes of ,-lactamase produced by Staphylococcus aureus (11,12,14). We recently have reported that extracts of strains known to produce these ,3-lactamase variants exhibit differences in the relative rates of hydrolysis of selected cephalosporin antimicrobial agents, permitting the identification of the different staphylococcal ,-lactamases (4). In the present investigation, we explored the utility and limitations of these techniques in determining the type of P-lactamase produced by 115 strains of S. aureus.Alternative methods which involve the preparation of wholecell suspensions of S. aureus suitable for hydrolysis assays are described. MATERIALS AND METHODSAcquisition of S. aureus isolates. A total of 115 3-lactamase-producing strains of S. aureus were evaluated. A total of 102 isolates were recovered from clinical and surveillance cultures performed between January 1986 and June 1988 at Saint Thomas Hospital, Nashville, Tenn., and LDS Hospital, Salt Lake City, Utah (LDS isolates provided courtesy of D. C. Classen and J. P. Burke). Thirteen reference strains, nine of which had been reported to produce different variants of staphylococcal P-lactamase with Richmond's typing antisera (11), also were evaluated. The source and pedigree of six reference strains have been previously described (4); these strains include PC1 and NCTC 9789 (type A ,Blactamase), V137 (type C ,-lactamase), and FAR8, FAR10, and FAR19 (type D ,-lactamase). Strains RN11(pI258) and RN98(pII147) produce the A and C variants of staphylococ- 10972 (propagating strain for bacteriophage 96), ST79/741,...