Serial isolates of Pseudomonas aeruginosa from a cystic fibrosis patient have identical pilin sequences

Abstract

Five isolates of Pseudomonas aeruginosa (CD2, CD3, CD4, CD5, and CD10) from a patient with cystic fibrosis were examined with regard to several genotypic and phenotypic characteristics to determine whether the patient was colonized with one or several distinct strains. Isolates CD2, CD3, and CD4 were obtained from a single sputum sample, and CD5 and CD10 were obtained 1 and 2 years later, respectively. On the basis of colonial morphology, serotyping, and antibiograms, the five isolates appeared to be different strains. However, Southern blot analysis with a 1.2-kilobase DNA probe containing the P. aeruginosa PAK pilin gene indicated that all five strains were identical at that genetic locus. The pilin genes of the five isolates were cloned and sequenced at the nucleotide level and found to be identical. Southern blot analysis with a probe from a separate region of the P. aeruginosa chromosome, a 741-base-pair PstI-NruI DNA fragment adjacent to the exotoxin A gene, also revealed genetic identity among these five clinical isolates. On this basis, it was concluded that this patient was colonized with a single strain of P. aeruginosa and that the strain had remained genetically stable over a period of 2 years. The predicted pilin sequence of the CD isolates was almost identical to that of strain PA103 (97% homology) and serologically related to PAO pilin, with which it shared 80% homology. No immunological cross-reactivity was detected between the CD and PAK pilins, which shared the least homology (62%) among the four pilins considered in this study. Although all five CD isolates contained identical pilin genes, three had acquired mutations which prevented normal expression of the pilus system. CD3 was a putative regulatory mutant which was unable to produce normal amounts of pilin, and CD4 and CD10 were putative assembly mutants which produced normal amounts of pilin but were unable to assemble the pilin subunit into intact pili.

Heterogeneity, persistence, and distribution of Pseudomonas aeruginosa genotypes in cystic fibrosis patients

“…Most recent research suggests that, once one strain colonizes the lung of the CF patient, this strain remains the dominant strain within the lung, and changes in the strain type are very rare (8,12,14,23,28). The results we obtained support this.…”

Use of a pilin gene probe to study molecular epidemiology of Pseudomonas aeruginosa

“…Most of such strains are sensitive to the bactericidal effect of normal human serum and are nontypeable, autoagglutinable, or agglutinable by multiple typing sera (9,10,26,27). Moreover, we and others (19, 21) have demonstrated that isolates with different LPS serotypes often prove to be clonally related when examined by gene probing and restriction fragment length polymorphism (RFLP) techniques. Therefore, conventional serotyping methods do not offer the discriminatory power necessary to distinguish among isolates and provide answers to basic epidemiological questions.…”

T-cell modulation of the antibody response to bacterial polysaccharide antigens

“…The polysaccharides used are all clinically relevant antigens. P. aeruginosa is an important cause of infection in cystic fibrosis and burn patients (24,26), S. mutans is strongly implicated in the development of dental caries (20), Neisseria meningitidis causes meningitis primarily in children (27), and S. pneumoniae causes middle ear infection in children and pneumonia in the elderly (23). Our present studies show, for instance, that the antibody response to SM can be significantly enhanced by increasing Ta-cell activity.…”