Analysis of Pseudomonas aeruginosa clinical isolates for possible variations within the virulence genes exotoxin A and exoenzyme S

We have previously characterized several Pseudomonas aeruginosa isolates th at were obtained from patients with tracheal, urinary tract, or wound infec tions (A. H. Hamood, J. A. Griswold, and C. M. Duhan, 1996, J. Surg. Res. 6 1: 425). Analysis of additional isolates showed that regardless of the isol ation site, some isolates produced significantly higher or significantly lo wer levels of either exotoxin A or exoenzyme S proteins. These variations d id not correlate with the mucoid phenotype of the isolates. One aim of this study was to determine if the variations in the level of exotoxin A or exo enzyme S are due to DNA rearrangements within either the toxA or the exoS g ene. This was accomplished by Southern blot hybridization experiments using a toxA internal probe, a toxA upstream probe, or an exoS internal probe. H ybridization with the toxA internal probe produced a 0.8-kb hybridizing fra gment, whereas hybridization with the exoS internal probe produced either a 2.0- or a 2.3-kb hybridizing fragment. Hybridization with the toxA upstrea m probe, however, produced hybridizing fragments of varying sizes, regardle ss of their isolation site. Isolates that showed a similar hybridization fr agment with either the toxA upstream probe or the exoS internal probe produ ced variable levels of exotoxin A or exoenzyme S. These results suggest tha t: [1] specific location within the host has no effect on either the mucoid phenotype of the isolate or the level of exotoxin A or exoenzyme S produce d by the isolates; [2] although restriction polymorphism exists within the toxA upstream region, both the toxA and the exoS structural genes are relat ively conserved; and [3] variations in the level of exoenzyme S and exotoxi n A produced by different isolates do not correlate with either the observe d heterogeneity within the toxA upstream region or the mucoid phenotype of the isolates. (C) 1999 Academic Press.