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DNA fingerprinting of Mycobacterium tuberculosis has been shown to be a powerful epidemiologic tool. We propose a standardized technique which exploits variability in both the number and genomic position of IS6110 to generate strain-specific patterns. General use of this technique will permit comparison of results between different laboratories. Such comparisons will facilitate investigations into the international transmission of tuberculosis and may identify specific strains with unique properties such as high infectivity, virulence, or drug resistance.

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In this study we established the usefulness of DNA fingerprinting for the epidemiology of tuberculosis on the basis of the DNA polymorphism generated by the insertion sequence (IS) IS986. Although clinical isolates of Mycobacterium tuberculosis displayed a remarkably high degree of restriction fragment length polymorphism, we showed that transposition of this IS element is an extremely rare event in M. tuberculosis complex strains grown either in vitro or in vivo for long periods of time. The M. tuberculosis and Mycobacterium africanum strains tested in this study contained 6 to 17 IS copies. In the Mycobacterium bovis strains, the copy numbers ranged between 1 and 5, and all 27 M. bovis BCG strains investigated invariably contained a single IS copy. This copy was located at a unique chromosomal position, reinforcing the idea that the frequency of IS transposition is very low in M. tuberculosis complex strains. Various microepidemics are described in which each microepidemic corresponds to a particular fingerprint type. The extent of similarity between Dutch and African strains was quantitatively assessed by computer-assisted analysis of DNA fingerprints. The results indicate that M. tuberculosis strains from regions in central Africa, where tuberculosis is highly prevalent, are generally more related to each other than isolates from the Netherlands, where the transmission rate is low and where the majority of the tuberculosis cases are presumed to be the result of reactivation of previously contracted M. tuberculosis infections. MATERIALS AND METHODS Bacterial strains and plasmids. In this study, 222 M. tuberculosis strains, 5 Mycobacterium africanum strains, 24 M. bovis strains, and 27 M. bovis BCG strains were inves-2578 on September 28, 2020 by guest http://jcm.asm.org/ Downloaded from USE OF IS ELEMENTS IN TUBERCULOSIS EPIDEMIOLOGY 2579 TABLE 1. Bacterial strains used in this study Bacterial strain no. Species Origin Source or reference 5, 14, 15, 97, 164, 265, 266, 319-323 M. tuberculosis The Netherlands This laboratory 272-285 M. tuberculosis Czechoslovakia J. Ivanyib 34, 165, 302-316 M. tuberculosis The Netherlands P. G. H. Peerboomsc 324-333 M. tuberculosis Ruwanda F. Portaelsd 334-335 M. tuberculosis Central African Republic F. Portaelsd 336-343 M. tuberculosis Burundi F. Portaelsd 317-318 M. tuberculosis Belgium F. Portaelsd 116-126, 286-292 M. tuberculosis The Netherlands P. L. van Puttene 267-271, 108, 109, 111, 114 M. tuberculosis The Netherlands J. Steensmaf 295-301, 168-171 M. bovis The Netherlands This laboratory 149-152, 293, 294, 45, 106 M. bovis BCG5 The Netherlands This laboratory 44, 104 M. bovis BCG5 The Netherlands This laboratory 102 M. bovis BCG9 Organon Teknikah 103 M. bovis BCG5 Armand Frappier' a Strains used only for standard RFLP typing are not mentioned in this table.

An inverse correlation between colonization of the human nasopharynx by Streptococcus pneumoniae and Haemophilus influenzae, both common upper respiratory pathogens, has been reported. Studies were undertaken to determine if either of these organisms produces substances which inhibit growth of the other. Culture supernatants from S. pneumoniae inhibited growth of H. influenzae, whereas culture supernatants from H. influenzae had no effect on the growth of S. pneumoniae. Moreover, coculture of S. pneumoniae and H. influenzae led to a rapid decrease in viable counts of H. influenzae. The addition of purified catalase prevented killing of H. influenzae in coculture experiments, suggesting that hydrogen peroxide may be responsible for this bactericidal activity. H. influenzae was killed by concentrations of hydrogen peroxide similar to that produced by S. pneumoniae. Hydrogen peroxide is produced by the pneumococcus through the action of pyruvate oxidase (SpxB) under conditions of aerobic growth. Both an spxB mutant and a naturally occurring variant of S. pneumoniae, which is downregulated in SpxB expression, were unable to kill H. influenzae. A catalase-reversible inhibitory effect of S. pneumoniae on the growth of the respiratory tract pathogens Moraxella catarrhalis and Neisseria meningitidis was also observed. Elevated hydrogen peroxide production, therefore, may be a means by which S. pneumoniae is able to inhibit a variety of competing organisms in the aerobic environment of the upper respiratory tract.