Population transcript accumulation of Pseudomonas aeruginosa exotoxin A and elastase in sputa from patients with cystic fibrosis

Abstract

The in vivo regulation of Pseudomonas aeruginosa virulence factors during the chronic lung infections associated with cystic fibrosis is poorly understood. We have developed an approach for the analysis of transcript accumulation of individual virulence factors from the P. aeruginosa populations found in the sputa of patients with cystic fibrosis. This method has been named population transcript accumulation, since we examine the transcript accumulation patterns in RNA extracted from the total bacterial population found in the sputum samples. DNA probes specific for P. aeruginosa elastase (lasB) and exotoxin A (toxA) were used to examine the population transcript accumulation of 21 sputum samples taken from 10 patients. We detected three patterns of population transcript accumulation: lasB and toxA, lasB alone, and neither lasB nor toxA. We also measured the relative levels of elastase and exotoxin A transcript accumulation in 19 of these samples. In the six samples containing both toxA and lasB transcripts, we found that the levels of lasB transcripts were consistently higher than those of toxA. Differences in the stability of the two mRNA species could not completely account for the higher level of lasB population transcript accumulation, since we showed that the mRNA half-life of lasB (11 min) was similar to that of toxA (10 min). Finally, we showed that elastase transcripts could be detected in some samples which contained only mucoid isolates. This finding suggests that both mucoid and nonmucoid populations may be transcribing lasB in the lungs of patients with cystic fibrosis.