Thalia I. Nicas

Role of exoenzyme S in chronic Pseudomonas aeruginosa lung infections.

Exoenzyme S is an extracellular ADP-ribosyltransferase enzyme produced byPseudomonas aeruginosa. Mutants ofPseudomonas aeruginosa deficient in this enzyme have been shown to have reduced virulence in infections of burned mice. The contribution of exoenzyme S to the pathogenesis of chronic lung infections with this organism was evaluated by examining the incidence of exoenzyme S production byPseudomonas aeruginosa strains isolated from cystic fibrosis patients and comparing an exoenzyme S deficient mutant and its exoenzyme S producing parent in a rat chronic lung infection model. Of 51 isolates examined, 43 % produced detectable levels of exoenzyme S. While both the exoenzyme S deficient mutant and its parent strain were equally capable of colonizing and persisting in rat lungs, the exoenzyme S producing parent elicited a greater degree of lung damage. These data suggest that exoenzyme S contributes to the pathogenesis of chronic lung infections.

CONTRIBUTION OF EXOENZYME S TO THE VIRULENCE OF PSEUDOMONAS AERUGINOSA

Exoenzyme S is an extracellular ADPR transferase produced by P. aeruginosa. Forms of this enzyme that have thus far been purified are not toxic, however, exoenzyme S clearly contributes to the virulence of strain 388. Thus, a Tn1 mutant deficient in exoenzyme S was found to be markedly less virulent than its exoenzyme S-producing parental strain in both a burned mouse infection model and a rat chronic lung infection model. Exoenzyme S does not appear to contribute to initial colonization of the rat lung or the burned mouse skin. Exoenzyme S does, however, appear to contribute to local tissue damage in the rat lung, and to dissemination of P. aeruginosa from the skin into the blood and distant organs of the burned mouse. Perhaps our most important observation is that specific antibody against exoenzyme S confers a high level of protection against subsequent infection of burned mice. While these results must be extended to include additional strains they are encouraging, and they underscore the relative importance of exoenzyme S in the pathogenesis of P. aeruginosa infections.

Genetic approaches to study Pseudomonas aeruginosa protein antigens.

Pseudomonas aeruginosa produces a large number of extracellular products which may play a role in pathogenesis. We have used genetic techniques to elucidate the relative contribution of these proteins to virulence, and as a method of producing safe toxoids. A mutant has been isolated which produces an immunologically reactive nontoxic form of toxin A, the most toxic extracellular protein produced by P. aeruginosa. Although there are difficulties in production of sufficient quantities of this CRM toxoid, these are likely to be solved by further genetic manipulation. Protection studies with toxin A antibody and studies of mutants deficient in toxin A have confirmed that toxin A plays a role in pathogenesis while clearly showing that toxin A alone cannot totally account for the virulence of P. aeruginosa. Studies of mutants specifically altered in three other products, exoenzyme S, and the two major proteases of P. aeruginosa, elastase and alkaline protease, have clarified the contribution of these products to virulence. Demonstration by genetic studies that exoenzyme S was a major factor in the virulence for one P. aeruginosa strain allowed us to correctly predict that antibody to this product would be protective against infection with that strain.

The Contribution of Exoproducts to Virulence of Pseudomonaa Aeruginosa

Pseudomonas aeruginosa produces a large number of extracellular products which may contribute to its virulence. We have employed a genetic approach to determine the contribution of toxin A, exoenzyme S, elastase and alkaline protease to the pathogenesis of P. aeruginosa. Mutations have been introduced with chemicals or transposons. Mutants have been identified using immunological, chemical, or toxicity assays. Mutants were extensively characterized in vitro to ascertain that they were identical to their parent strain except for the production of the desired product. Appropriate mutants were compared with their parent strains in several animal models: the burned mouse model, the mouse corneal infection model, and a rat model of chronic lung infection. The data indicate that virulence of P. aeruginosa is multifactorial. Further, the relative contribution of a given P. aeruginosa product may vary with the type of infection.