Diversification of After Inhaled Tobramycin Therapy of Cystic Fibrosis Patients: Genotypic and Phenotypic Characteristics of Paired Pre- and Post-Treatment Isolates.

This study examines the impact of inhaled tobramycin therapy on the within-host changes in strains isolated from Bulgarian patients with CF prior to and post treatment. Genotypic comparison by RAPD-PCR indicated that most of the pre-treatment isolates had a high similarity and were genetically comparatively close to strains from other countries with known increased morbidity or treatment requirements. Most of the post-treatment isolates were, however, genetically distant from their pre-treatment counterparts, showing genotypic diversification after the treatment. Phenotypic comparisons showed a lower ODmax reached during groswth and an increased lag-time in the post-treatment isolates. All strains were capable of invasion and intracellular reproduction within A549 cultured cells. The addition of sub-inhibitory amounts (1/4 or 1/2 MIC) of tobramycin during growth showed the higher relative fitness (as a percentage of the untreated control) of the post-treatment strains. The effects of sub-MICs on biofilm growth did not show such a pronounced trend. However, when a resazurin-based viability test was applied, the advantage of the post-treatment strains was confirmed for both broth and biofilm cultures. In spite of that, according to the determined MIC values, all isolates were tobramycin-sensitive, and the data from this study imply the development of tolerance to the antibiotic in the strains that survived the treatment.