Culture-Negative Childhood Empyema Is Usually Due to Penicillin-Sensitive Streptococcus pneumoniae Capsular Serotype 1

Abstract

Two studies in the United States have shown that isolates of Streptococcus pneumoniae from children with empyema are more likely to be penicillin sensitive and of capsular serotype 1 than those from uncomplicated pneumococcal pneumonia (1, 4). In the United Kingdom, childhood empyema is becoming more common (S. D. Playfor, A. R. Smyth, and R. J. Stewart, Letter, Thorax 52:932, 1977; J. H. M. Rees, D. A. Spencer, D. Parikh, and P. Weller, Letter, Lancet 349:402, 1997). Many cases have no definitive etiology, despite attempts at noncultural diagnosis by antigen detection, and are most likely due to previous antibiotic therapy. It is uncertain, therefore, if these culture-negative cases of childhood empyema are from the same population as the S. pneumoniae culture-positive ones or whether they are a separate and different problem. We have examined pleural aspirates from 43 children (aged 6 months to 17 years; mean age, 6.2 years; median age, 6 years; 30 of whom were male) presenting with empyema thoracis over a 4-year period to the Freeman Hospital in Newcastle upon Tyne, a tertiary referral center for the North East of England. Routine culture and a latex agglutination test for pneumococcal antigen (SLIDEX; BioMerieux, Marcy l'Etoile, France) were performed with these specimens, after which the remaining material was digested with dithiothreitol. A 1-ml aliquot of the digest was centrifuged at 10,000 × g for 10 min, and the pellet was washed twice in phosphate-buffered saline (PBS). The pellet was resuspended in 12.5% Chelex-100 resin (Bio-Rad Laboratories Ltd.) and heated at 56°C for 30 min and then at 100°C for 15 min. The supernatant was then tested for the presence of S. pneumoniae DNA by a previously described real-time PCR assay targeting the pneumolysin gene (ply) (A. M. Kearns, R. Freeman, O. M. Murphy, P. R. Seiders, M. Steward, and J. Wheeler, Letter, J. Clin. Microbiol. 37:3434, 1999). Positive samples underwent a second real-time PCR assay targeting a conserved sequence of the PBP2B gene (pbp2b) (3) found in all strains of penicillin-susceptible S. pneumoniae (PSSP) for which the MIC of penicillin was <0.06 mg/liter. Finally, whenever sample volumes permitted, 250 μl of the digested sample was mixed with 750 μl of PBS and heated to 100°C for 10 min and then centrifuged for 2 min (21,000 × g). The supernatant was used in solid-phase indirect sandwich enzyme-linked immunosorbent assays (ELISAs) for the detection of type-specific pneumococcal capsular polysaccharides, with monoclonal antibodies to types 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, and 23F donated by Wyeth Vaccines Research, West Henrietta, N.Y. Two samples were culture positive. One of them, for which all tests for S. pneumoniae were negative, yielded Staphylococcus aureus. The other grew scanty Staphylococcus epidermidis, which was regarded as a skin contaminant. All pbp2b assays indicated “penicillin sensitivity.” Other results are found in Table ​Table11. TABLE 1. Results of real-time PCR assays for pneumococcal DNA and ELISAs for 13 pneumococcal capsular serotype antigens on 43 pleural aspirates from children with empyema We found evidence of S. pneumoniae infection in over 70% of culture-negative empyema fluids. The infecting strains were likely to have been highly penicillin sensitive, and at least 60% were of capsular serotype 1, which, in the United Kingdom, as in the United States, is not the commonest serotype in invasive pneumococcal disease (2). We conclude that cases of culture-negative childhood empyema presenting at tertiary referral centers are not an etiologically distinct group, that childhood pneumococcal vaccines must include capsular serotype 1 antigen, and that early detection of S. pneumoniae capsular serotype 1 antigen in cases of childhood pneumonia may aid management and predict complications, especially empyema.